Document ymgyNYZmKRm5DwvNgZxLqNbaX

3M Katherine E. Reed, Ph.D. Start' Vice President CERTIFIED MAIL 3M Environmental, Health and Safety Operations RECEIVED ADDT DD'P 06JUM-7 AH 6:21 900 Bush Avenue. Building 42-2F.-26 PO Box 33331 St. Paul. MN 55133-3331 651 77S 4331 May 30, 2006 Document Processing Center EPA East - Room 6428 Attn: Section 8(e) Office of Pollution Prevention and Toxics U.S. EPA 1200 Pennsylvania Avenue NW Washington, DC 20460-0001 Re: TSCA 8(e) Substantial Risk Notice: Sulfonate-based and Carboxylic-based Fluorochemicals, Docket Nos. 8EHQ-1180-374; 8EHQ-0381-0394; 8EHQ-0598-373 [check these numbers] - Results from Analysis of Groundwater, Sediments, Surface Water, Fish and Clams in the Decatur, Alabama Area To whom it may concern: 3M is submitting this notice to supplement its previous submissions on sulfonyl and carboxylicbased fluorochemicals. More specifically, the data contained in this submittal have been generated as part of a site-related environmental assessment for fluorochemicals that 3M is performing for its 3M Decatur, Alabama manufacturing facility. As part of this effort, 3M has recently received the enclosed final analytical reports for various sampling activities conducted in the vicinity of our Decatur manufacturing facility: Property Adjacent to 3M Decatur Facility 3M Environmental Laboratory Report No. E06-0199 - Analysis o f PFOA, PFOS, PFHS, and PFBS in Aqueous Samples: 3M Decatur Offsite Sampling Event (BPP Sample Sites) Exygen Study No. P0000760, Interim Report #23, E05-0209, Analysis of Sediment Samples Exygen Study No. P0001131, Interim Report #23, E05-0210, Analysis o f Sediment Samples Tennessee River Biota Sampling Exygen Study No. P0001131, Interim Report #10, E05-0210, Analysis of Fish and Clam Samples The first three reports contain data for samples collected from property adjacent and immediately to the west of the 3M Decatur site. More specifically, three locations within an open water marsh area were sampled for surface water and sediments. The identification numbers for these locations are: 89060000336 S 9 0 6 0 0 0 O V S6 JZ <j (j $ 1 U.S. EPA Document Processing Center May 30, 2006 Page 2 Surface Water DAL-S WS-BPPO1-0-060413 DAL-S WS-BPP02-0-060413 DAL-S WS-BPP03-0-060413 Sediments DAL-SD-BPPO1-0-060421 DAL-SD-BPP02-0-060421 DAL-SD-BPP03-0-060421 In general, the BPPOI location represented the eastern end of the marsh, closest to inlet water being supplied by a nearby well. The BPP02 location was towards the northern edge of the marsh and the BPP03 location towards the south and eastern portion of the marsh, near the outlet to adjacent wetlands. In addition to the samples referenced above, groundwater from a well located on the western adjacent property and used to supply the marsh with water was also sampled. This location is identified as DAL-GWS-BPWELL-0-060413 in the first report listed above. There are a two additional items pertaining to Report No. E06-0199 that merit discussion. First, the report represents a subreport for a series of samples collected adjacent to, but off of, the 3M Decatur facility. Analytical results for additional off-site samples are pending but not final at this time. Second, there are several results that have been presented as estimated minimum values based on associated quality control samples that did not demonstrate adequate method performance. We are working to better quantify these results and may have revised values when the entire laboratory report is finalized. The fourth report listed above contains analytical data for fish and clams collected in late 2004 from the Tennessee River near the Decatur, AL area. Again, these laboratory results were just recently finalized. The enclosed figure provides sample locations corresponding to the results tabulated in the analytical report. While 3M does not believe that any of these data taken alone or cumulatively meet the "substantial risk" reporting threshold, we nevertheless recognize the ongoing work by U.S. EPA to assess fluorochemica! exposure pathways. Therefore, we are placing these results in the 8(e) docket as a supplement to previous submissions. If you have any questions, please do not hesitate to contact Gary Hohenstein at (651)778-5150. Sincerely, yuL Katherine E. Reed Staff Vice President Environmental, Health and Safety Operations 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 Subreport Analysis of PFOA, PFOS, PFHS, and PFBS in Aqueous Samples: 3M Decatur Offsite Sampling Event (BPP Sample Sites) Laboratory Request Number: 06-0199 Method Requirement: 3M Method ETS-8-154.1 (modified) Testing Laboratory 3M EHS Operations 3M Environmental Laboratory Building 2-3E-09 935 Bush Avenue St. Paul, MN 55106 Requester Gary Hohenstein 3M Building 42-02-E-27 PO Box 3331 Saint Paul, MN 55133-3331 Phone: (651)778-5150 Fax: (651) 778-7203 The testing reported herein meet the requirements of ISO/IEC 17025-1999 "General Requirements for the Competence of Testing and Calibration laboratories", in accordance with the A2LA Certificate #2052-01. Testing that complies with this International Standard also operate in accordance with ISO 9001/ISO 9002 (1994). Certificate #2052-01 PAGE 1 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO E06-0199 3M Environm ental Laboratory 3M Environmental Laboratory Manager: William K. Reagen, Ph.D. 3M Technical Lead: Michelle Malinsky, Ph.D. Report Author: Micheile Malinsky, Ph.D. Analytical Subreport E06-0199 Surface Water Samples BPP01, BPP02, and BPP03; Ground Water Sampile BPWell Decatur Offsite Sampling Event Report Date: May 1,2006 1 Introduction/Summary The 3M Environmental Laboratory extracted and analyzed ground water and surface water samples collected by Weston Solutions personnel on April 12 and 13, 2036 near the 3M Decatur facility. Samples were returned to the 3M Environmental Laboratory for analysis of perfluorooctanoate (PFOA), perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHS) and perfluorobutane sulfonate (PFBS) under laboratory project number E06-0199 using modified 3M Environmental Laboratory Method ETS 8-154.1 "Determination of Perfluorinated Adds, Alcohols, Amides, and Sulfonates in Water by Solid Phase Extraction and High Performance Liquid Chromatography/Mass Spectrometry". Per the dient's request, an initial subreport is being issued for the following four sample locations only: DAL SWS BPP01 DAL SWS BPP02 DAI. SWS BPP03 DAL. GWS BPWELL This report will contain information relevant to these four sample collection points only. These results will be reissued in a comprehensive final project report that indudes results for all samples collected under E06-0199. The 3M Environmental Laboratory prepared sets of sample containers for thirty-two separate sampling locations. At a minimum, each sample set consisted of a field sample, feld sample duplicate, low field spike and high field spike. For some locations, s third mid-level field spike was prepared. Each empty container was marked with a "fill to here" line which corresponded to a final volume of 450 mL. Containers reserved for field matrix spikes were fortified with an appropriate matrix spike solution containing the four target analytes prior to being sent *o the field for sample collection. The spike amounts prepared for each, of the four locations presented here wili be provided later in this report. Certificate #2052-01 The testing reported herein meet the requirements of ISO/IEC 17025-1999 "General Requirements for the Competence of Testing and Calibration Laboratories", in accordance with the A2LA Certificate #2052-01. Testing that complies with this International Standard also operate in accordance with ISO PAGE 2 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 Samples were initially extracted and analyzed on April 19, 2006. Samples were diluted,, reextracted, and analyzed on April 21, 2006. Laboratory matrix spikes were prepared, diluted, extracted, and analyzed on April 25, 2006 in the analytical batch initiated on April 24, 2006., Table 1 below summarizes the sample results. All results for quality control samples prepared and analyzed with the samples will be reported and discussed elsewhere in this report. All samples were diluted 1:50 in ASTM Type I water prior to extraction. Table 1. Sample Results Summary. 3M UNIS ID E06-O199-055 E06-0199-056 E06-0199-060 E06-0199-061 E06-O199-065 E06-0199-066 E06-0199-070 E06-O199-071 Sample Description . mPFOA Concentration (ng/mL) PFOS Concentration (ng/mL) DAL SWS BPP01 0 060413 DAL SWS BPP01 DB 060413 Average 293 307 300 >423 >380 >402 %RPD Sample/Sample Dup . . DAL SWS BPP02 0 060413 DAL SWS BPP02 DB 060413 Average %RPD Sample/Sample Dup DAL SWS BPP03 0 060413 DAL SWS BPP03 DB 060413 : Average %RPD Sample/Sample Dup DAL GWS BPWELL 0 060413 DAL GWS BPWELL DB 060413 Average %RPD Sample/Sample Dup 206 J 219 ~ 212 6-1 , 219 214 216 2.3 (2|>774 >779 >776 N NA >295 >314 >304 NA >272 >256 >264 NA >1460 >1170 >1320 NA mPFHS Concentration (ng/mL) 53.3 . 54.3 53,8 1.8 33.2 39.8 39.0 ' 4.1 39.1 40.1 39.6 2.5 >145 >145 >145 NA n PFBS Concentration (ng/mL) 6.98 7.17 7.08 Z7 4.30 4.72 4.66 2.6 4.69 5.05 4.87 7.4 17.2 17.6 17.4 2.3 (1) The analytical method uncertainty for PFOA, PFHS, and PFBS was 10011%, 10Q5.8%, and 1008.8%, respectively based on method accuracy and precision. Sed Section 3.7 for additional explanation. The limit of quantitation (LOQ) for these sampbs only was 2.50 ng/mL (PFOA) and 1.25 ng/mL (PFHS and PFBS). Results and average values are rounded to three significant figures according to EPA rounding rules. Percent relative difference (%RPD) values are rounded to two significant figures. Because of rounding, values may vary slightly from those listed in the raw dbta. (2) All PFOS sample results presented here should be considered estimated minimum values as quality control samples (field matrix spikes and lab matrix spikes) associated with these four samples did not demonstrate adequate method performance. The accuracy of the PFOS results could be within an order o f magnitude. %RPD values not calculated. BPWell results for PFOA and PFHS should also be considered estimated minimums as field matrix spike recoveries did not meet method acceptance criteria o f 10030%. The analytical accuracy of the PFOA and PFHS estimated values for this sample only could be within an order of magnitude. %RPD values not calculated. 2 Methods - Analytical and PreparatoryJ ,f! \ 2.1 Sample Collection Samples were collected in NalgeneTM (low-density polyethylene) bottles prepared at the 3M Environmental Laboratory. Prior to sample collection, bottles designated for field matrix spikes were spiked in the laboratory with a known volume of an appropriate matrix spiking solution containing the PAGE 3 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO E06-0199 four analytes cf interest (PFOA, PFOS, P^HS and PFBS). Collected sample bottles were received at the laboratory at ambient conditions on April 19, 2006. Table 2 below details the samples collected and spikes added to each bottle Table 2. Sample Collection and Spike information. 3 M U M S ID Weston Sample Description E06-019S-055 E06-C199-056 E0S-0199-057 E06-0193-058 E06-0199-059 DAL SWS BPP01 0 060413 DAL SWS BPP01 DB '060413 DAL SWS BPF01 LS 9C412 DAL SWS BPP01 MS 960413 DAL SWS BPP01 HS 060413 E06-0199-060 E06-0199-061 E06-0199-062 E06-0199-063 E06-0199-064 DAL SWS BPP02 0 060413 DAL SWS BPP02 DB 060413 DAL SWS BPP02 LS 360413 DAL SWS BPP02 MS 060413 DAL SWS BPP02 HS 060413 E06-0199-065 E06-0199-066 E06-0199-067 E06-0199-068 E06-0199-069 DAL SWS BPP03 0 060413 DAL SWS BPP03 DB 060413 DAL SWS BPP03 LS 060413 DAL SWS BPP03 MS 060413 DAL SWS BPP03 HS 060413 E06-0199-070 DAL GWS BPWELL 0 060413 E06-0199-071 DAL GWS BPWELL DB 060413 E06-0199-072 DAL GWS BPWELL LS 060413 E06-0199-073 DAL GWS BPWELL MS 060413 E06-0199-074 DAL GWS BPWELL HS 060413 DAL = Decatur, AL SWS = Surface Water Sample GWS = Ground Water Sample 0 = Sample DB = Duplicate LS = Low Spike MS = Mid Spike HS = High Spike 3M Sollte Number 109 110 4.414i *112 113 119 120 121 122 123 114 115 116 117 113 72 73 74 75 75 BoWe Designation Sample Sample Dup FMS Low FMS Mid FMS High Sample Sample Dup FMS Low FMS Mid FMS High Sample Sample Dup FMS Low FMS Mid FMS High Sample Sample Dup FMS Low FMS Mid FMS High Nominal Spike Level (ng/mL) NA NA 1.0 10 100 NA NA 1.0 10 100 NA NA 1.0 10 100 NA NA 10 100 1000 GPO Location Comment FSIA Drainage FSIA Drainage FSIA Drainage FSiA Drainage FSIA Drainaqe FSIA Drainage FSIA Drainage FSIA Drainage FSIA Drainage FSiA Drainaqe FSIA Drainage FSIA Drainage FSIA Drainage FSIA Drainage FSIA Drainaqe BP Property BP Property BP Properly BP Property BP Property 2.2 Extraction All samples, calibration standards, and associated quality control samples were extracted using a modified procedure of ETS-8-154.1 "Determination of Perfluorinated Acids, Alcohols, Amides, and Sulfonates in Water by Solid Phase Extraction and High Performance Liquid Chromatography/Mass Spectrometry". Briefly, 40 mL of sample were loaded onto a pre-conditioned Waters C18 solid phase extraction (SPE) cartridge (Sep-Pak, 1.0 g, 6 cc) using a vacuum manifold. The loaded SPE cartridges were then eluted with 5 mL of methanol. This extraction procedure concentrates the samples by a factor of eight. (Initial volume = 40 mL, final volume = 5 mL). Lab control spikes extracted in the same manner cross-validate all the method modifications/deviations from ETS-8-154.1. See Section 3.6 for additional information. PAGE 4 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 The samples and sample duplicates for BPP01, BPP02, and BPP03 were initially extracted undiluted on April 19, 2006. Initial analysis of the undiluted samples exceeded the upper limit of quantiation (25 ng/mL nominal) for PFOA and PFOS. Likewise, the sample and sample duplicate for BPWELL were initially extracted on April 19, 2006. The final extracts were diluted 1:10 prior to analysis. Analysis of these diluted extracts also exceeded the upper limit of quantitation. ,A|I the samples, sample duplicates, and field matrix spikes were reextracted on April 21,2006. For BPP01, BPP02, and BPP03, the samples and associated QC were diluted 1:50 using ASTM Type I water prior to extraction. For BPWELL, the sample, sample duplicate,,low spike and mid spike were diluted 1:50, while the high spike was diluted 1:500. ' * ? . ? .. w 2.3 Analysis All sample and quality control extracts were analyzed for PFOA, PFOS, PFHS, and PFBS using high performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). Pertinent instrument parameters, the liquid chromatography gradient program, and the specific mass transitions analyzed are described in the tables below. Table 3. Instrument Parameters. Instrument Name Liquid Chromatograph Guard column Analytical column Injection Volume Mass Spectrometer Ion Source Electrode Polarity Software ETSStan Agilent 1100 Betasil C18 (2.1 mm.XlOO mm), 5 urn Betasil C18 (2.1 mm X 100 mm), 5 urn 5 gL Applied Biosystems API 4000 Turbo Spray Z-spray Negative_________ _ Analyst 1.4.1 Table 4. Liquid Chromatography Gradient Program. Step Number 0 1 2 3 4 5 Total Time (min) 0 1.0 14.5 15.5 16.5 20.0 Flow Rate (pL/min) 300 300 300 300 300 300 Percent A (2 mM ammonium acetate) 80.0 80.0 10.0 10.0 80.0 80.0 Percent B (Methanol) 20.0 20.0 90.0 90.0 20.0 20.0 PAGE 5 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 Table 5. Mass Transitions. . Analtfe ' ' 'PFBS " 'PFHS " 'PFOS " 'PFOA Mass Transition Q1/Q3 2S9/80 299/99 399/80 , 399/99 499/130 499/99 499/80 413/369 413/219 413/169 Dive// Time (msec) 125 125 125 125 125 125 125 125 125 125 (1) The individual transitions were summed to produce a "total ion chromatogram" (T IC). Tire TICs were used for quantitation. 3 Data Analysis 3.1 Calibration Calibration standards were prepared by spiking known amounts of stock solutions containing the target analytes into 40 mL of ASTM type I water, Each spiked water standard was then extracted in the same manner as the collected samples. A total of twelve spiked standards ranging from 0.025 ng/mL to 25 ng/mL (nominal) were prepared. A quadratic, 1/x weighted, calibration curve was used to fit the data for each analyte. The data were not forced th-cugh zero during the fitting process. Calculating the standard concentration using the peak area counts and the resultant calibration curve confirmed accuracy o f each curve point. Each extracted calibration standard used to generate the final calibration curve met the method calibration accuracy requirement of 10025%. The correlation coefficients (r) were greater than 0.999 for all analytes. 3.2 Limit of Quantitation. (LOQ) The LOQ for this analysis, as defined In ETS-8-154.1, is the 'owest non -zero calibration standard in the curve in which the area counts are at least twice those of the method and solvent blank(s). The LOQs for PFOA, PFOS, PFHS, and PFBS were 0.050C ng/mL, 0.C999 ng/mL, 0.0250 ng/mL, and 0.0250 ng/mL, respectively. Note: these LOQ values have not been corrected for sample specific dilution factors. Area count comparison for the method blanks and the lowest calibration standard will be provided in Section 3.5.1. 3.3 System Suitability Replicate injections of the 1 ng/mL extracted-calibration standard were analyzed at the beginning and end of the analytical sequence to demonstra*e overall system suitability. All analytes met method acceptance criteria of less than 5% relative standard deviation (RSD) for peak area and less than 2% RSD for retention time for both opening and closing system suitability injections. 3.4 Continuing Calibration During the course of the analytical sequence, several continuing calibration verification samples (CCVs) were analyzed to confirm that the instrument response and the initial calibration curve were still in control. One PFOS CCV (139%) and one PFHS CCV (153%) exceeded method acceptance criteria of 10025% for accuracy. Both of these non-ccmpliant CCVs followed a contaminated solvent blank and the high recovery is most likely carryover from the previous injection. A method deviation has been PAGE GOF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 issued. The overall impact to the data quality objectives for the sample results is considered minimal as all sample injections and sample related QC injections were bracketed by compliant CCVs. 3.5 Blanks Three types of blanks were prepared and analyzed with the samples: method blanks, solvent blanks, and field/trip blanks. Each blank type is described below. 3.5.1 Method Blanks Several method blanks were prepared by loading 40 mL of ASTM Type f water onto a C18 SPE cartridge and eluting with 5 mL of methanol using the same extraction procedure as the samples. Method blanks were prepared to evaluate the levels of background contamination in the overall extraction process (reagent water, glassware, SPE cartridges, etc.) The calibration curve was prepared on a separate day as the samples. (Calibration standards: April 10, 2006; diluted samples: April 21, 2006). The method blanks prepared with the calibration standards as well as with the samples were both analyzed during the April 21, 2006 analysis. Additional lab matrix spikes were prepared and analyzed in a separate analytical run initiated on April 24, 2006. Again, the method blanks prepared with the calibration curve and the method blanks prepared with the spikes were analyzed. Method blank area count comparisons to the LOQ standard are provided in the Attachments section at the end ofthis report. 3.5.2 Solvent Blanks Several methanol solvent blanks were analyzed to assess system contamination and/or instrument carryover. For the April 21, 2006 analysis, three separate vials of methanol solvent blanks were contaminated and produced area counts of the target anaiytes greater than one-half those of the LOQ standard. A method deviation has been issued. The solvent blank area count comparison for the April 21, 2006 is provided in the Attachments section at the end of this report. For the lab matrix spikes analyzed on April 24, 2006, only the initial solvent blanks injected before the opening system suitabilities ana the solvent blanks analyzed after the highest calibration standard exhibited area counts greater than one-half the LOQ standard. 3.5.3 Field/Trip Blanks Prior to sample collection, seven sets of trip/field blanks were prepared. A trip blank consists of a sample container filled with 450 mL of ASTM Type I water, seafed, and shipped to the sample collection site along with the empty containers. The trip blank serves as an additional method blank that accounts for any storage conditions and/or holding time issues that the samples may experience. Results of the trip blank analyses will be provided in the final comprehensive report. 3.6 Lab Control Spikes (LCSs) Lab control spikes at nominal concentrations of 0.050 ng/mL, 5.0 ng/mL, 100 ng/mL, and 1000 ng/mL spikes were prepared and analyzed each day samples were extracted and analyzed. The 0.050 ng/mL and 5.0 ng/mL LCSs were prepared by spiking known mounts of the analytes into 40 mL of ASTM Type I water to produce the desired concentration. The spiked water samples were then extracted and analyzed in the same manner as the samples. For the 100 ng/mL and 1000 ng/mL LCSs, target analytes were added to 450 mL o f ASTM Type I water to produce the final desired concentration. An aliquot of the LCS was then diluted 1:50 or 1:500 with ASTM Type I water prion to extraction, in the same manner as the diluted samples. This technique was done to demonstrate that the dilution procedure used for the samples was able to produce accurate results. Analysis of replicate LCSs at the various levels cross-validates the analytical method, as used here, for any modifications/deviations from ETS 8-154.1. Additionally, LCS results will be used to determine overall method uncertainty in Section 3.7. Table 6 summarizes the LCS recovery results. PAGE 7 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 LrC'CS' nPercen.t nRecovery = -C--a--l-c-u--l-a-t-e--d---C--o--n--oe---n--t-r-a-t-i-o--n- *. 1.00% Spike Concentration LCS% RSD = standard deviation LCS replicates . 100% average LCS recovery PAGE 8 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO E06-0199 Table 6. *\ab `Control Spike Results. Sample Description Lab ID 0.05 ppb LCS LCS-060410-1 0,05 ppb LCS LCS-060410-2 0.05 ppb LCS LCS-060410-3 5.0 ppb LCS LCS-060410-4 5.0 ppb LCS LCS-060410-5 5 0 ppb LCS LCS-060410-6 0.05 ppb LCS LCS-060421-1 0 05 ppb LCS LCS-060421-2 0.05 ppb LCS LCS-060421-3 5.0 ppb LCS LCS-060421-4 5.0 ppb LCS LCS-060421-5 5.0 ppb LCS LCS-060421-6 100 ppb LCS LCS-060421-7 100 ppb LCS LCS-060421-8 100 ppb LCS LCS-060421-9 1000 ppb LCS LCS-060421 -10 1000 ppb LCS LCS-060421-11 1000 ppb LCS LCS-060421-12 0.05 ppb LCS LCS-060425-1 0.05 ppb LCS LCS-060425-2 5.0 ppb LCS LCS-060425-3 5.0 ppb LCS LCS-060425-4 100 ppb LCS LCS-060425-5 100 ppb LCS LCS-060425-6 1000 ppb LCS LCS-060425-7 1000 ppb LCS LCS-060425-fl Average %Recovery J%RSD PFOA Spiked Cone. (ng/mL) Calc. Cone. (ng/mL) % Recovery 0.0500 0.0500 0.0500 5.00 5.00 5.00 0.0500 0.0500 0.0500 5.00 5.00 5.00 100.0 100.0 100.0 1000 1000 1000 0.0500 0.0500 5.00 5.00 100.0 100.0 1000 1000 <0.0500 <0,0500 0.0517 4.93 4.86 5.30 <0.05 <0.05 0.0514 5.02 4.81 4.82 95.0 99.7 96.9 928 977 1030 0.0689 0.0600 4.91 4.87 95.3 98.6 977 972 101%9.7% |2'n a (2,n a 104 98.6 97.2 106 (2|n a ,2,n a 103 100 96.2 96.4 95.0 99.7 96.9 92 8 97.7 103 |4,138 120 98.2 97.3 95.3 98.6 97.7 97.2 PFOS Spiked Cone. (ng/mL) Calc. Cone. (ng/mL) % Recovery 0.0500 <0 0999 0.0500 <0.0999 0.0500 <0 0999 5.00 493 5.00 5.01 5.00 5.05 0.0500 <0.0999 0.0500 <0.0999 0.0500 <0 0999 5.00 4.53 5.00 4.34 5.00 4.49 99.9 87.3 99.9 89.2 99.9 88,5 999 855 999 899 999 929 0.0500 0.0631 0.0500 0.152 5.00 4.79 5.00 4.94 99.9 84.5 99.9 89.8 999 878 999 849 93.1%10% |2,n a |2in a (2,n a 98.7 100 101 (2>n a k na (2'n a 90.7 86.9 89.9 87.4 89.3 88.6 85.6 90.0 93.0 |4,126 |5|304 95.9 98.9 84.6 89.9 87.9 85.0 PFHS PFBS Spiked Cone. (ng/mL) Calc. Cone. (ng/mL) % Recovery 0.0500 00500 00500 5.00 5.00 5.00 0.0500 00500 0.0500 5.00 5.00 5.00 100 100 100 1000 1000 1000 0.0500 0.0500 5.00 5.00 100 100 1000 1000 0.0504 0.0480 0.0483 4.98 4 94 4.98 0.0501 00485 0.145 4.88 479 4,90 98.1 100.0 994 940 988 1020 0.0578 0.0592 4.98 5.08 95.9 96.8 978 966 99.8%S.6% 05 00 CM 101 96,0 96.6 99.6 98.8 99.6 100 97.0 97.6 95.8 98.0 98.1 100.0 994 94.0 98.8 102 116 118 99.6 102 95.9 96.8 97.8 96.6 Spiked Cone. (ng/mL) Calc. Cone. (ng/mL) % Recovery 0.0499 0.0499 0.0499 4.99 4.99 4.99 0.0499 0.0499 0.0499 4.99 4.99 4.99 99.9 99.9 99.9 999 999 999 0.0499 0.0499 4.99 4.99 99.9 99.9 999 999 0.0505 0.0456 0.0480 4.94 4.88 4.94 0.0426 0.0427. 0.0441 4.80 4.71 4.76 . 98.5100.0 : 98.0 909 973 1040 0.0563 0.0507 4.89 4.86 95.2 96.6 988 975 96.8%S.8% 101 91.3 96.1 98.9 97.7 98.9 85.3 85.5 88.3 96.1 94.3 95.3 98.6 100 98.1 91.0 97.4 104 113 102 979 97.3 95.3 96.7 989 976 PAGE 9 OF 21 3MENVIRONMENTAL LABORATORY REPORT NO. E06-0199 (1) All results and average values listed to three significant figures according to EPA rounding rules. %RSD values given to two significant figures. Due to rounding, values may vary slightly from those in the raw data. (2) Low end of the calibration curve disabled for PFOS and PFOA due to solvent blank contamination. Recovered LCS concentrations were below the established LOQ and are not reported. (3) High recovery due to instrument carryover from the previous injection of a contaminated methanol blank. Recovery excluded fi um average and %RSD calculations. (4) Recovery was outside the method acceptance criteria of 10025%; however, the recovery was included in the calculation of the overall average and %RSD. (5) Recovery was outside the method acceptance criteria of 10025%; the recovery value was not included In the calculation of the overall average and %RSD as it was clearly a statistical outlier. The 5 ppb, 100 ppb, and 1000 ppb LCSs are more representative of the sample concentrations for the results reported in this subreport. Impact to data quality objectives is minimal. PAGE 10 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. 06-0)99 3.7 Analytical Method Uncertainty Both the accuracy (percent recovery) and precision (%RSD) of the lab control spikes were used to estimate the overall method's analytical uncertainty for a given analyte. For example, the overall accuracy and precision for PFOA based on LCS results was 101%9.7%. The measured precision (%RSD) is then used to determine the range of the accuracy. Example: 101 *(0.097) = 9.84 101+9.84=111.1; 1 0 1 -9 .8 4 = 91.45 Thus, LCS accuracy results range from 91.45% to 111.1 %. The absolute difference of the low and high ends of this range, when compared to 100%, are then calculated. 111.1 %-100 = 11.1% 100%-91.45 = 8.54%. The most conservative (largest) absolute difference is then used as the analytical uncertainty for the given analyte. Therefore, the analytical uncertainty for PFOA is given as 10011% (two significant figures) for these results. For PFOS, PFHS, and PFBS, the analytical method uncertainty is 10016%, 1005.8%, and 1008.8%, respectively. It should be emphasized that the analytical method uncertainty described above only applies to the method and the procedures performed within. Additional quality control samples specific to the sample matrix, in the form of field matrix spikes and lab matrix spikes, demonstrate that the individual sample results are accurate to within 10030%, the targeted data quality objectives. 3.8 Field Matrix Spikes (FMS) At a minimum, low and high field matrix spikes were collected at each sampling point to verify that the analytical method is applicable to the collected matrix. For some locations, a mid-level spike was also collected. Field matrix spike recoveries within method acceptance criteria of 10030% confirm that "unknown" components in the sample matrix do not interfere with the extraction and analysis of the analytes of interest. Field matrix spikes will be presented in the next section with the sample data. The field matrix spike should be at least 50% of the endogenous sample concentration to be considered an appropriate spike level for the given sample matrix. FMS Recovery - (SamP|e Concentration of FMS - Average Concentration: Field Sample & Field Sample Pup.) , 1QQ0/ Spike Concentrator! 3.9 Lab Matrix Spikes (LMS) If the sample and sample duplicate produced concentrations where none of the associated field matrix spikes were at least 50% of the endogenous concentration, then laboratory matrix spikes (LMS) were prepared. LMSs were prepared by spiking a known amount of target analytes into a measured aliquot of the sample matrix. The spiked matrix was then diluted, if necessary, and extracted in the same manner as the samples. LMS recoveries within 10030% demonstrated that the analytical method was appropriate for the given sample matrix only in the absence of an appropriate field matrix spike. 4 Data Summary The data tables below summarize the sample results, field matrix spike recoveries, and laboratory matrix spike recoveries for the four locations (BPP01, BPP02, BPP03, and BPWell). Each table provides the average concentration and the relative percent difference (RPD) of the sample and sample duplicate. PAGE 11 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO E06-0199 BPP01 The endogenous concentration for BPP01 was at least twice the spike concentration for the low spike (1 ng/mL), mid spike (10 ng/mL) and high spike (100 ng/mL) for both PFOA and PFOS. A laboratory matrix spike at 500 ng/mL was prepared and analyzed. The LMS recovery met method acceptance criteria of 10030% for PFOA (79.6%), but not for PFOS (60.6%). Therefore, the sample results for PFOS should be considered estimated minimums with a sample analytical accuracy within an order of magnitude as the analytical method has not been demonstrated to be suitable for the given sample matrix. FMS and/or LMS recoveries within 10030% for PFOA, PFHS, and PFBS demonstrate that the analytical method is appropriate for the given sample matrix for these analytes. BPP02 The endogenous concentration for BPP02 was at least twice the spike concentration for the low spike (1 ng/mL) and mid spike (10 ng/mL) for both PFOA and PFOS as well as the high spike (100 ng/mL) for PFOS only. A laboratory matrix spike at 250 ng/mL was prepared and analyzed. The LMS recovery met method acceptance criteria of 10030% for PFOA (75 0%), but not PFOS (59.8%). Therefore, the sample results for PFOS should be considered estimated minimums with a sample analytical accuracy within an order of magnitude as the analytical method has not been demonstrated to be suitable for the given sample matrix. FMS and/or LMS recoveries within 10030% for PFOA, PFHS, and PFBS demonstrate that the analytical method is appropriate for the given sample matrix for these analytes. BPP03 The endogenous concentration for BPP03 was at east twice tne spike concentration for the low spike (1 ng/mL) and mid spike (10 ng/mL) for both PFOA and PFOS as well as the high spike (100 ng/mL) for PFOS only. A laboratory matrix spike at 250 ng/mL was prepared and analyzed. The LMS recovery met method acceptance criteria of 100+30% for PFOA (73.8%), but not PFOS (60.8%). Therefore, the sample results for PFOS should be considered estimated minimums with a sample analytical accuracy within an order of magnitude as the analytical method has not been demonstrated to be suitable for the given sample matrix. The high level PFBS FMS exceeded method acceptance criteria of 10030% (134%). However, the PFBS results for this sample are still considered accurate within 10030% because the criteria exceedence was minimal (within 4%) and mid level FMS and LMS recoveries were excellent. 101% and 90.2%, respectively. High level FMS and LMS recoveries within 10030% for PFOA and PFHS also demonstrate that the analytical method is appropriate for the given sample matrix for these two analytes as well. BPWell The endogenous concentration for BPWell was at least twice the spike concentration for the low spike (10 ng/mL) for PFOA, PFOS, PHFS, and PFBS as well as the mid spike for PFOA, PFOS, and PFHS. Although the high level spike (1000 ng/mL) was an appropriate level for all four analytes, the recoveries were consistently less than 30% for all analytes, with PFOS showing no measuring recovery. The high FMS was reextracted to rule out a potential preparatory error during the extraction process. The second extraction produced similar recoveries as the first. A laboratory matrix spike at 1000 ng/mL was also prepared. The LMS recovery met method acceptance criteria of 10030% for PFOA (85.4%), PFHS(93.5%), and PFBS(97.2%) but not for PFOS (13.5%). Therefore, the sample results for PFOS should be considered estimated minimums with a sample analytical accuracy within an order o f magnitude as the analytical method has not been demonstrated to be suitable for the given sample matrix. Although the LMS produces acceptable recoveries for PFOA and PFHS, the sample results should be considered estimated minimums as no appropriate supplementary FMS recovery was available. Low and mid level FMS and LMS recoveries within 10030% for PFBS demonstrate that the analytical method is appropriate for the given sample matrix for this analyte only. Further method development for the BP Well sample matrix will be performed to see if improved PFOA, PFOS, and PFHS recoveries can be achieved. PAGE 12 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO E06-0199 Table 7. ,1,BPP01 Results. PFOA PFOS PF HS PFBS 3M U M S ID Description E06-0199-055 DAL SWS BPP01 0 060413 E06-0199-056 DAL SWS BPP01 DB 060413 E06-0199-057 DAL SWS BPP01 LS 060413 E06-0199-058 DAL SWS BPP01 MS 060413 E06-0199-059 DAL SWS BPP01 H3 060413 ... LMS-060425-1 DAL SWS BPP01 0 060413 500ppb LMS Average Concentration (ng/mL) %RPD Cone. (ng/mL) % Recovery 293 NA 307 NA 292 " NR 328 " NR. 361 ('"NFL 698 79.6 3004.7% Cone. (ng/mL) % Recovery 423 NA 380 NA 338 ,2,n r 484 (2In r 365 ,2,n r 704 ,3160.6 ' ,4I>402 Cone. (ng/mL) % Recovery 53.3 NA 54.3 NA 53.4 i2,NR 66.6 (2,n r 136 82.2 485 86.2 53.81.9% Cone. (ngAnL) % Recovery 6.98 NA 7.17 NA 7.44 |2|n r 188 117 93.8 86.8 444 87.5 7.08zr/. NA= Not Applicable * (1) Table displays rounded values for all concentrations and percent recovery values (3 significant figures). %RPD values (2 significant figures). Recovery and RPD values may vary slightly from the values in the raw data. All samples and associated FMSs were diluted 1:50 in ASTM Type I water prior to extraction: LMS was diluted 1:100 prior to extraction. (2) NR=Not reportable. FMS level not appropriate for the given matrix. The endogenous sample concentration was at least twice spike level. " (3) LMS recovery outside of method acceptance criteria of 10030%. , _. (4) PFOS sample results should be considered estimated minimums only. Sample matrix specific QC did not meet method criteria. The analytical sample accuracy is arbitrarily estimated to within an order of magnitude. %RPD values not reported. ,- PAGE 13 OF 21 3M ENVIRONMENTAL LABORA TORY REPORT NO. E06-0199 Table 8. ,1|BPP02 Results. PFOA PFOS PFHS PFBS 3MLIMSID Description E06-O199-060 DAL SWS BPP02 0 060413 E06-0199-061 DAL SWS BPP02 DB 060413 E06-0199-062 DAL SWS BPP02 LS 060413 E06-0199-063 DAL SWS BPP02 MS 060413 E06-0199-064 DAL SWS BPP02 HS 060413 LMS-060425-2 DAL SWS BPP02 0 060413 250ppb Average Concentration (ng/mL) %RPD Cone. (ng/mL) % Recovery 206 NA 219 NA 234 |2|n r 220 ,2In r 314 102 400 75.0 2126.1% Cone. (ng/mL) % Recovery 295 NA 314 NA 388 ,2|n r 346 ,2,n r 437 *N R 454 |3|59.8 <4>>304 Cone. (ng/mL) % Recovery 38.2 NA 39.8 NA 42.3 |2,n r 48.0 b,NR 134 95.0 256 86.8 39.04.1% Cone. (ng/mL) % Recovery 4.6 NA 4.72 NA 5.9 (2,nr 14.4 97.5 102 97.5 223 87.4 4.662.6% NA=Not Applicable. (1) Table displays rounded values `or all concentrations and percent recovery values (3 significant figures), %RPD values (2 significant figures). Recovery and RPD values may vary slightly from the values in the raw date All camples end associated QC were diluted 1.50 in ASTM Ty.Xi I water prior to extraction. (2) NR=Not reportable. FMS level not appropriate for the given matrix. Endogenous concentration was at least twice spike level. (3) LMS recovery outside of method acceptance criteria of 10Qt30% (4) PFOS sample results should be considered estimated minimums only. Sample matrix specific QC did not meet method criteria. The analytical sample accuracy is arbitrarily estimated to within an order of magnitude. %RPD values not reported. PAGE 14 OF 21 3MENVIRONMENTAL LABORATORY REPORT NO. E06-0199 Table 9. BPP03 Results. PFOA PFOS PFHS PFBS 3 M U M S ID Description Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery E06-0199-065 DAL SWS BPP03 0 060413 219 NA 272 NA 39.1 NA 4.69 NA E06-0199-066 DAL SWS BPP03 DB 060413 214 NA 256 NA 40.1 NA 5.05 NA E06-0199-067 E06-0199-068 DAL SWS BPP03 LS 060413 DAL SWS BPP03 MS 060413 211 |2)n r 258 |2|n r 40.2 " NR 5.76 |2>n r 228 " NR 255 (2,n r 50.0 |2>n r 15.0 101 E06-0199-069 DAL SWS BPP03 HS 060413 324 108 315 (2,n r 143 103 139 (31134 LMS-060425-3 DAL SWS BPP03 0 060413 250ppb LMS 401 73.8 416 ,4,60.8 262 89.0 230 90.2 Average Concentration (ng/mL) %RPD 216Z3% ffl>264 39.62.5% 4.877.4% NA=Not Applicable. (1) Table displays rounded values for all concentrations and percent recovery values (3 significant figures), %RPD values (2 significant figures). Recovery and RPD values may vary slightly from the values in the raw data. All samples and associated QC were diluted 1:50 in ASTM Type I water prior to extraction. (2) NR=Not reportable. FMS level not appropriate for the given matrix. Endogenous concentration was at least twice spike level. (3) FMS recoveries outside method acceptance criteria of 10030%. (4) LMS recovery outside of method acceptance criteria of 10030%. . (5) PFOS sample results should be considered estimated minimums only. Sample matrix specific QC did not meet method criteria. The analytical sample accuracy is arbitrarily estimated to within an order of magnitude. %RPD values not reported. PAGE 15 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 Table 10. |1|BPWell results. PFOA PFOS PFHS PFBS 3M U M S ID Description Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery Cone. (ng/mL) % Recovery E06-0199-070 DAL GWS BPWELL 0 060413 774 NA 1460 NA 145 NA 17.2 NA E06-0199-071 DAL GWS BPWELL DB 060413 779 NA 1170 NA 145 NA i 7.o NA E06-0199-072 DAL GWS BPWELL LS 060413 723 l2,NR 960 (2|n r 152 |2,n r 26.9 95.1 E06-0199-073 DAL GWS BPWELL MS 060413 821 |2|n r 1020 I2|n r 244 ,2|n r 122 105 ,3,E06-0199-074 |4,E06-0199-074 DAL GWS BPWELL HS 060413 DAL GWS BPWELL HS 060413 1020 |S|24.4 1200 |6,n m r 375 ,5|23.0 280 ,S|26.9 987 |5|21.0 1320 lf|NMR 348 (51203 254 |5)23.7 LMS-O60425-4 DAL GWS BPWELL 0 060413 1000ppb LMS 1630 85.4 1450 <7)13.5 1080 93.5 988 97.2 Average Concentration (ng/mL) %RPD NA=Not Applicable. '' >776 m>1T2u ">145 17.412.3% (1) Table displays rounded values for all concentrations and percent recovery values (3 significant figures), %RPD values (2 significant fiqures). Recovery and RPD values may vary slightly from the values in the raw data. The sample, sample duplicate, low CMS, and mid FMS were diluted 1:50 in AS7M Type I water prior to extraction. The high FMS and 1000 ppb LMS were diluted 1:500 prior to extraction. (2) NR=Not reportable. FMS level not appropriate for the given matrix. Endogenous concentration was at least twice spike level. (3) FMS extracted on 4-21-2006 with the samples. (4) FMS re-extracted on 4-25-0006. (5) FMS recovery outside of method acceptance criteria of 10030%. (6) NMR = No measurable recovery. (7) LMS recovery outside method acceptance criteria of 1U030%. (8) PFOA, PFOS, and PFHS sample results should be considered estimated minimums only. Field matrix spikes did not meet method criteria. The analytical sample accuracy is arbitrarily estimated to within an order of magnitude. %RPD values not reported. PAGE 16 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. EO&-0199 5 Conclusion Sample matrix QC samples for the four samples reported herein did not meet method acceptance criteria of 10030% for PFOS. Therefore, the values listed in Table 1 and Tables 7-10 forPFOS should be considered estimated minimums. The sample analytical accuracy is arbitrarily estimated to be within an order of magnitude. Further method development is needed for this matrix as the QC results demonstrated that unknown matrix components preclude accurate analysis with the method as performed here. Field matrix spikes yielding recoveries within 10030% demonstrated that the analytical method was suitable for the given sample matrices for PFOA, PFHS, and PFBS for BPP01, BPP02, and BPP03. The analytical method uncertainty for these analytes was presented in T a b le l. PFOA and PFHS results for BPWeH should also be considered estimated minimums with a sample analytical accuracy within an order o f magnitude as no acceptable held matrix spikes recoveries were produced. g Data / Sample Retention All remaining sample and associated project data (hardcopy and electronic) will be archived according to 3M Environmental Laboratory standard operating procedures. PAGE 17 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. E06-0199 7 Signatures \ Michelle D. Malinsky, Ph.D.ySM Technical llead ^ / f/ o L Date Kent R. Lindstrom,` 3M Technical Reviewer r < / j / c - ' Date ___________ William K. Reagen, Ph.D., Environmental Laboratory Management S / C i/ / {}^ Date The 3M Environmental Laboratory's Quality Assurance Unit has audited the data and report for this project. Quality A s k a n c e Representative Date 3M CONFIDENTIAL PAGE 18 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO. 06-0 J99 8 Attachments 8.1 Method Blank/LOQ Area Count Comparison. Table 11. Method Blank/LOQ Area Count Comparison: April 21, 2006. Analytical Run: April 21, 2006 Datafile Sample Description Sample Name PFOA Area Counts PFOS PFHS PFBS Method Blanks Prepared with the Extracted Curve s060421a025 Method Blank-1 MB-060410-1 55982 3413 1591 2760 s060421a026 s060421a027 s060421a028 s060421a029 s060421a030 s060421a031 s060421a032 Method Blank-2 Method Blank-3 Method Blank-4 Method Blank-5 Method Blank-6 Method Blank-7 Method Blank-8 MB-060410-2 MB-060410-3 MB-060410-4 MB-060410-5 MB-060410-6 MB-060410-7 MB-060410-8 18376 16127 22714 21587 25928 21060 16383 1082 1718 1116 792 4338 2143 1776 668 1970 639 1686 2077 2436 1660 2001 2336 1305 2109 5711 3561 4235 Method Blanks Prepared with the Extracted Samples s060421a045 s060421a046 s060421a047 s060421a048 s060421a049 s060421a050 s060421a051 s060421a052 Method Blank-1 Method Blank-2 Method Blank-3 Method Blank-4 Method Blank-5 Method Blank-6 Method Blank-7 Method Blank-8 MB-060421-1 MB-060421 -2 MB-060421-3 MB-060421-4 MB-060421-5 MB-060421-6 MB-060421-7 MB-060421-8 12933 12809 13875 22242 19675 17774 13843 14073 1437 1446 2228 2053 1762 1568 1084 1492 '" 121128 '"129379 |1)66834 |1)42902 '"33923 '"13105 6059 5570 277 715 595 1052 826 2071 m19419 450 Area Counts of the LOQ standard 163373 14600 13176 25766 Area Counts of the LOQ standard*0.5 81686.5 7300 6588 12883 Concentration of LOQ standard (ng/mL) 0.0500 0.0999 0.025 0.025 (1) Method blanks Injected immediately after a highly contaminated solvent blank. Carryover present. Values excluded when establishing the LOQ for PFHS. (2) Method blank determined to be a statistical outlier at the 99% confidence level using Dixon's Q-test. Value exduded when establishing the LOQ for PFBS. PAGE 19 OF21 3MENVIRONMENTAL LABORATORY REPORT NO E06-0199 Table 12. Method Blank/LOQ Area Count Comparison:' . April 24, 2006. Analytical Run: April 24, 2006 Datafile Sample IDescription Sample Name PFOA Method Blanks Prepared with the Extracted Curve s080424a025 Method Blank-1 MB-0604-10-1 (1*36625 s060424a026 Method Blank-2 MB-06410-2 '" 42239 s060424a027 Method Blank-3 MB-060410-3 25302 s060424a028 Method Blank-4 MB-060410-4 22151 s060424a029 Method Blank-5 MB-060410-5 30401 s060424a030 Method Biank-6 MB-06041 -6 17458 s060424a031 Method Biank-7 MB-060410-7 230/4 s060424a32 Method Blank-8 MB-060410-8 22049 Method Blanks Prepared with the Extracted Laboratory Matrix Spikes s060424a090 Method Biank-1 MB-060425-1 30448 s060424a091 Method Blank-2 MB-060425-2 15006 s060424a092 Method Blank-3 MB-00425-3 11262 s060424a093 Method Blank-4 MB-060425-4 11867 s060424a094 Method Blank-5 MB-060425-5 16236 s060424a095 Method Blank-6 MB-060425-6 15924 s060424a026 Method Blank-7 s060424a097 Method Blank-8 Area Counts of the LOQ standard Area Counts of the LOQ standard*0.5 Concentration of LOQ standard (ng/mL) MB-030475-7 MB-060425-8 22750 ! 1928-! ! 36060 I 18030 ! 0.025 Area Counts PFOS PFHS ! PFBS 3514 2274 2950 804 2565 827 1195 1192 2301 1266 991 836 1024 1107 1530 1071 6724 3362 0.04 2029 2890 1147 1197 1955 1275 1974 1512 1778 1311 1039 707 717 589 1038 1135 12166 6083 0.025 3078 3938 2248 1638 2593 5242 3516 4294 1906 2990 1703 6142 2723 5446 435S 1677 27032 13516 0.025 (1) Value excluded for the LOQ determination for the given set. Area counts are very close to one-half the LOG value. Prior analysis of this extract demonstrated area counts below one-half the 25 ppt standard. LMSs analyzed on this day are well above the LOQ. PAGE 20 OF 21 3M ENVIRONMENTAL LABORATORY REPORT NO EO6-0199 8.2 Solvent Blank/LOQ Comparison. Table 13. Area Counts of Solvent Blank. Area Counts File Vial Position PFOA PFOS PFHS s060421a001 91 70233 3067 1853 s060421a002 91 16249 2196 1851 s060421a003 91 18222 .1642 . 56? . s060421a009 91 48287 4524 1505 s060421a010 92 |1|84762 5006 2424 s060421a023 92 48308 . ,4192 4524 s060421a024 92 '"84666 ., -.5317, 4738 s060421a035 93 '" 162166 '"283547 . '" 52950 S060421a037 93 23806 . '"8904 2706 s060421a042 93 15939 2135 1693 s060421a044 s060421a055 94 49169 . 4?W , |1|274j^54 94 1268) ,1141 '"328841 s060421a057 94 47071 338? '"257946 s060421a068 95 53698 2525 5237 s060421a070 95 74125 , 5907 6588 s060421a081 95 62139 2124 3945 s060421a083 95 68329 3139 3951 s060421a094 96 76799 4192 3512 s060421a096 96 77269, 3358 3905 s060421a107 97 14824 1166 852 s060421a109 97 19120 1763 1499 s060421a120 97 9918 1028 , ,1024 s060421a122 98 24599 1589 2321 s060421a127 98 37573 1305 6127 s060421a129 99 18673 110 1909 s060421a135 99 31743 1944 1472 LOQ Standard 163373 14600 13176 Vi'LOQ Standard 81687 7300 6588 (1) Area counts of the solvent blank were greater than '/2*the LOQ standard area counts. PFBS 192 563 471 447 889 6609 7007 '"41582 2357 , 1823 1118, 547 946 1137 975 1225 1149 1054 v 1173 335 ,, 1289 698 3275 2190 937 632 25766 12883 PAGE 21 OF 21 INTERIM REPORT # 1 0 - Analysis of Fish and Clam Samples STUDY TITLE Analysis o f Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program DATA REQUIREMENTS EPA TSCA Good Laboratory Practice Standards 40 CFR 792 STUDY DIRECTOR Jaisimha Kcsari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 Phone: 610-701-3761 INTERIM REPORT COMPLETION DATE May 09, 2006 PERFORMING LABORATORY Exygen Research 3058 Research Drive State College, PA 16801 P h o n e :814-272-1039 STUDY SPONSOR 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 Phone: 651-733-6374 PROJECT Protocol Number P0001131 Exygen Study Number: P0001131 Total Pages: 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: POOOl 131 GOOD LABORATORY PRACTICE COM PLIANCE STATEM ENT Exygen Study Number POOOl 131, entitled "Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program," conducted for 3M Company, is being performed in compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792 by Exygen Research. 'Principal Investigator Exygen Research Jaisimha , ___ Study Director Weston Solutions, Inc. W ^ckr-t Michael A. S Sponsor Repi 3M Company Exygen Research Date Page 2 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Q UALITY ASSURANCE STATEM ENT Exygen Research's Quality Assurance Unit reviewed Exygen Study Number P0001131, entitled, "Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program". All reviewed phases1 were inspected for conduct according to Exygen Research's Standard Operating Procedures, the Study Protocol, and all applicable Good Laboratory Practice Standards. All findings were reported to the Exygen Principal Investigator and Management and to the Study Director. Phase 18. Raw Data Review and Interim Analytical Report Review 26. Raw Data Review and Interim Analytical Report Review Date Inspected Date Reported to Date Reported to Principal Exygen Investigator Management Date Reported to Studv Director 08/01-05/05 09/20/05 09/21/05 09/21/05 05/03/06 05/08/06 05/09/06 05/09/06 'Note: All in-lab inspections will be documented in the QA statement for the final analytical report at the conclusion of the study. This QA statement involves only the review of the interim report and associated raw data. Exygen Research Page 3 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: POOOi 131 CERTIFICATION O F A UTH ENTIC ITY This interim report, for Exygen Study Number P0001131, is a true and complete representation of the raw data. Submitted by: Exygen Research 3058 Research Drive State College, PA 16801 (814)272-1039 Principal Investigator, Exygen: Date Exygen Research Facility Management: Date Study Director, Weston Solutions, Inc Jaisimha Kesari P.E., DEE Weston Solutions, Inc. Sponsor Representative, 3M Company: Michael A. Santoro Director o f Regulatory Affairs Exygen Research Page4ofl32 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : P0001131 STUDY IDENTIFICATION Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program PROTOCOL NUMBER: P0001131 EXYGEN STUDY NUMBER: P0001131 TYPE OF STUDY: Residue SAMPLE MATRIX: Fish and Clams TEST SUBSTANCE: Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) SPONSOR: 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 STUDY DIRECTOR: Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 STUDY MONITOR: Michael A. Santoro 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 PERFORMING LABORATORY: Exygen Research 3058 Research Drive State College, PA 16801 ANALYTICAL PHASE TIMETABLE: Study Initiation Date: 11/05/04 Interim Analytical Start Date: 12/27/04 Interim Analytical Termination Date: 04/17/06 Interim Report Completion Date: 05/09/06 Exygen Research Page 5 of 132 Interim P,,eport #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 PROJECT PERSONNEL The Study Director for this project is Jaisimha Kesari at Weston Solutions, Inc. The following personnel from Exygen Research were associated with various phases of this interim portion of the study: Name John Flaherty Karen Risha Paul Connolly Amy Sheehan Christine Edwards Brittany Kravets Mindy Cressley Krista Gallant Scott Crain Eric Hoover Carlyle Horrell Shawn Robb Mark Ammerman Eric Edwards Edward Cams Title Vice President Laboratory Supervisor Technical Lead-LC/MS Associate Scientist Technician Technician Technician Technician Technician Technician Intern Technician Technical Lead - Facilities Sample Custodian Sample Custodian Sample Custodian Exygen Research Page 6 of 132 Interim Report #1O-Analysis o f Fish and Clam Samples - Exygen Study No. : P0001131 TABLE OF CONTENTS Page TITLE PAGE..........................................................................................................................1 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT.............................. 2 QUALITY ASSURANCE STATEMENT........................................................................... 3 CERTIFICATION OF AUTHENTICITY..............................,.............................................4 STUDY IDENTIFICATION..................................................................................................5 PROJECT PERSONNEL....................................................................................................... 6 TABLE OF CONTENTS....................................................................................................... 7 LIST OF TABLES................................................................................................................. 8 LIST OF FIGURES........................................................ 9 LIST OF APPENDICES...................................................................................................... 10 1.0 SUMMARY.......... ........................................................................................................11 2.0 OBJECTIVE..................................................................................................................12 3.0 INTRODUCTION............................................. 13 4.0 ANALYTICAL TEST SAMPLES.............................................................................13 5.0 REFERENCE MATERIAL................................................ 13 6.0 DESCRIPTION OF ANALYTICAL METHOD......................................................... 15 6.1 Extraction Procedure.................................................................................................. 15 6.1.1 Sample Preparation......................................................................................15 6.1.2 Glassware Preparation.............................. 15 6.1.3 Column Preparation.....................................................................................15 6.1.4 Sample Extraction........................................................................................15 6.2 Preparation of Standards and Fortification Solutions................................................ 16 6.3 Chromatography......................................................................................................... 17 6.4 Instrument Sensitivity................................................................................................. 17 6.5 Description of LC/MS/MS Instrument and Operating Conditions........................... 17 6.6 Quantitation and Example Calculation.......................................................................18 7.0 EXPERIMENTAL DESIGN........................................................................................20 8.0 RESULTS..................................................................................................................... 20 9.0 CONCLUSIONS...........................................................................................................21 10.0 RETENTION OF DATA AND SAMPLES.............................................................. 21 Exygen Research Page 7 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 LIST OF TABLES Eage Table L Summary of PFBS, PFHS and PFOS in Fish Samples........................................23 Table II. Summary of PFOS in Re-extracted Fish Samples...............................................30 Table HI. Summary o f PFBS, PFHS and PFOS in Clam Samples....................................32 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples................... 33 Table V. Matrix Spike Recovery of PFOS in Re-extracted Fish Samples........................ 46 Table VI. Matrix Spike Recovery PFBS, PFHS and PFOS in Clam Samples..................49 Exygen Research Page 8 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Figure 1. LIST OF FIGURES Page Typical Calibration Curve for PFBS in Methanol.......................................... 5 1 Figure 2. Non-Extracted Standards of PFBS in Methanol, 0.0005 pg/mL and 0.001 pg/mL, Respectively.......................................................... ...................52 Figure 3. PFBS in Control Fish Blank, 0.00625 pg/mL Fortified Control Fish Spk A, and 0.025 pg/mL Fortified Control Fish Spk B, Respectively............................. 53 Figure 4. Chromatogram Representing a Fish Sample Analyzed for PFBS (Exygen ED: C0049478, Data Set:061605E)...................................................54 Figure 5. Typical Calibration Curve for PFHS in Methanol.......................................... 55 Figure 6. Non-Extracted Standards of PFHS in Methanol, 0.0005 pg/mL and 0.001 pg/mL, Respectively.............................................................................. 56 Figure 7. PFHS in Control Fish Blank, 0.00625 pg/mL Fortified Control Fish Spk A, and 0.025 pg/mL Fortified Control Fish Spk B, Respectively.............................. 57 Figure 8. Chromatogram Representing a Fish Sample Analyzed for PFHS (Exygen ID: C0049478, Data Set: 061605E)..................................................58 Figure 9. Typical Calibration Curve for PFOS in Methanol.......................................... 59 Figure 10. Non-Extracted Standards o f PFOS in Methanol, 0,0005 pg/mL and 0.001 pg/mL, Respectively..................................... 60 Figure 11. PFOS in Control Fish Blank, 0.00625 pg/mL Fortified Control Fish Spk A, and 0.025 pg/mL Fortified Control Fish Spk B, Respectively............................. 61 Figure 12. Chromatogram Representing a Fish Sample Analyzed for PFOS (Exygen ID: C0049478, Data Set:061605E)...................................................62 Exygen Research Page 9 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 LIST OF APPENDICES Page Appendix A Study Protocol P0001131 (Exygen Study No. P0001131) with Analytical Method V0001783: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Fish and Clams by LCMS/MS", Protocol Amendments 1 and 2, and Deviations 2 and 3................................................................................................................... 63 Exygen Research Page 10 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : P0001131 1.0 SU M M A R Y Exygen Research extracted and analyzed fish and clam samples for the determination of perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS), and perfluorooctanesulfonate (PFOS) according to Exygen Method V0001783 (Appendix A). The limit o f quantitation (LOQ) for PFBS, PFHS, and PFOS in fish and clams was 0.2 ng/g. For one sample (C0048133) only one gram of fish was used for extraction, so the limit of quantitation for PFBS, PFHS, and PFOS in this fish sample was 1.0 ng/g. The initial analytical results associated with fifty-one fish samples are not reported (NR) in Table I because the endogenous levels of PFOS were too high to calculate accurate matrix spike recoveries. These samples were re-extracted and the results of the re analyses are summarized in Tables II and V. Analytical results and assessed accuracies for the analysis of PFBS, PFHS and PFOS found in fish samples are summarized in Table I. Analytical results and assessed accuracies for the analysis of PFOS found in the re-extracted fish samples are summarized in Table II. Analytical results and assessed accuracies for the analysis of PFBS, PFHS and PFOS found in clam samples are summarized in Table III. Laboratory matrix spikes that were in the appropriate concentration range (primary concentration less than three times the spiking level) were used to calculate the assessed accuracies. The average percent recoveries standard deviations for PFBS, PFHS, PFOS in fish samples were 79% 13%, 94% 12%, and 85% 20%, respectively. Fortification recoveries for PFBS, PFHS, and PFOS in fish samples are summarized in Table IV. The average percent recovery standard deviation for PFOS in the re-extracted fish samples was 109% 20%. Fortification recoveries for PFOS in the re-extracted fish samples are summarized in Table V. The average percent recoveries standard deviations for PFBS, PFHS, PFOS in clam samples were 84% 10%, 93% 13%, and 73% 16%, respectively. Fortification recoveries for PFBS, PFHS, and PFOS in clam samples arc summarized in Table VI. The sample IDs assigned by the client follow this formula: Dxx-x01-xxxxxx-x-04xxxx. The first string begins with D for Decatur, Alabama and the xx defines the sampling area within the Wheeler Reservoir of the Tennessee River where: DL3 = Up river Letter of Intent (LOI) sampling location LOC-3 DL2 - Cross river LOI sampling location LOC-2 DL1 - Downstream Fox Creek mouth LOI sampling location LOC-1 DBC = Bakers Creek mouth DOU = Cove at 3M WWTP outfall DMC = Downstream Mallard Creek mouth Exygen Research Page 11 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 The second string defines the general category of the biota sample and the sampling round where: F01 = Fish sample first round 101 - Invertebrate sample first round The third string defines the species (first two characters), the sample type (third character), and the sample number (last three characters) within that sampling area and type of sample where: IP = channel catfish (Ictalurus pu n ctatu s) MS = largemouth bass (M icro p ieru s sa lm o id es) CF = Asiatic clam ( C orbicu la flu m in ea) F =*Filet tissue sample W = Whole body sample 001 through 005 = sample number within the sampling area and sample type The fourth string (single character) additionally defines the sample type where: 0 " Primary field sample 1 = Field duplicate sample 2 = Equipment rinseate blank sample The fifth string defines the sample preparation date in YYMMDD format. Examples of this sample ID system are: Sample DBC-F01-IPF004-0-041021 is the filet tissue sample prepared on October 21, 2004 from the fourth channel catfish collected at the Bakers Creek mouth sampling location. Sample DLI-F01-MSW002-0-041104 is the whole body sample prepared on November 4,2004 from the second largemouth bass collected at the Fox Creek mouth sampling location. 2.0 OBJECTIVE The objective of the analytical part of this study was to determine levels of perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS), and perfluorooctanesulfonate (PFOS) in fish and clams according to Protocol P0001131 (Appendix A). Exygen Research Page 12 of 132 Interim Report tflO-Analysis o f Fish and Clam Samples Exygen Study No.: P 0001131 3.0 INTRODUCTION ' ! This report details the results of the analysis for the determination o f PFBS, PFHS, and PFOS in fish and clams using the analytical methods! entitled, "V0001783: Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS." The study was initiated on November 05, 2004, when the study director signed protocol number P0001131. The analytical start date for this interim report was December 27, 2004, and the analytical termination date for this interim report was April 17,2006. 4.0 ANALYTICAL TEST SAMPLES Forty fish samples (Exygen ID C0048126-C0048165) were received on dry ice on October 23, 2004 from Tim Frinak at Weston Solutions, Inc. Fifty-six fish samples (Exygen ID C0049450-C0049505) were received on dry ice on November 6, 2004 from Charles Young at Weston Solutions, Inc. Twenty-six fish samples (Exygen ID C0056354-C0056479) were received on dry ice on January 13, 2005 from Tim Frinak at Weston Solutions, Inc. Six clam samples, consisting of whole clams from each sample site (Exygen ID C0056480-C0056485) were received on dry ice on January 13, 2005 from Tim Frinak at Weston Solutions, Inc. A whole Whiting (Family: Gadidae) was purchased from a local grocer by Ed Cams on December 20, 2004 and was used as the control fish (Exygen ID C0054391). The samples were logged in by Exygen personnel and placed in frozen storage. Sample login and chain o f custody information is located in the raw data package associated with this interim report Storage records will be kept at Exygen Research. 5.0 REFERENCE MATERIAL The analytical standards, PFBS and PFHS, were supplied by 3M. PFBS was received from 3M at Exygen on July 6,2000, and May, 13,2005. PFHS was received from 3M at Exygen on January, 20, 2003. PFOS was purchased from Fluka Corporation and was received at Exygen on April, 23,2003. The available information for the reference materials is listed below. PFBS and PFHS were stored frozen and PFOS was stored refrigerated. Exygen Research Page 13 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Compound PFBS PFBS PFHS PFOS Exveen Inventory No. SP0000252 SP0005726 SP0002401 SP0002694 LotJ 101 101 SE036 430180-1 Puritv (% ) 96.7 96.7 98.6 101.2 Expiration D 12/04/06 12/04/06 10/18/06 10/31/07 The molecular structures o f PFBS, PFHS and PFOS are given below: PFBS Chemical Name: Perfluorobutanesulfonate Molecular Weight: 338 supplied as the potassium salt (C+FSOjTC) Transitions Monitored: 299 -> 99 Structure: PFHS Chemical Name: Perfluorohexanesulfonate Molecular Weight: 438 supplied as the potassium salt (C6Ft3S03'K+) Transitions Monitored: 399 - 80 Structure: S 0 3* PFOS Chemical Name: Perfluorooctanesulfonate Molecular Weight: 538 supplied as the potassium salt (CgFi7S03'K+) Transitions Monitored: 499 -> 80 Structure: Exygen Research Page 14 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 6.0 DESCRIPTION OF ANALYTICAL METHOD i The analytical method "V0001783: Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS" was used for the fish and clam samples in this study. 6.1 Extraction Procedure 6.1.1 Sam ple Preparation Before the samples could be weighed for the extraction, they had to be processed. To process, the frozen samples were placed into a food processor and homogenized with dry ice. The samples were transferred to one-gallon Ziploc bags and placed in frozen storage with bag left open to allow the dry ice to sublime. After sublimation, the sample bags were sealed and remained in frozen storage until time o f analysis. Sample processing records are located in the Sample Information section of the raw data package. 6.1.2 G lassw are Preparation The 125 mL pear-shaped flasks were silanized by first rinsing the flask with 30% dimethyldichlorosilane in toluene solution. The flasks were rinsed once with toluene followed by three rinses with methanol. The flasks were allowed to air dry before use. 6.1.3 Column Preparation The 20 mL columns were packed at Exygen in sequence with 2 grams florisil, 2 grams silica gel, 2 grams o f carbon, and 1 gram LC-NH2. The columns were conditioned on the day o f extraction with 20 mL of methanol followed by 20 mL of acetonitrile. All washes were discarded. These clean-up columns were used to remove matrix interference and not to retain PFBS, PFHS, and PFOS. Additional details about the column packing materials can be found in the raw data package associated with this repori. 6.1.4 Sam ple Extraction A 5-gram* portion of fish or clam sample was weighed into a fiftymilliliter centrifuge tube for the extraction, After fortification of appropriate samples, 30 mL of acetonitrile was added to the samples. The samples were allowed to shake on a wrist action shaker for ~15 minutes. The samples were placed into a freezer for -1 hour. Freezing of the Exygen Research Page 15 o f 132 Interim Report #IO-Ana!ysis o f Fish and D am Samples Exygen Study No.: POOO1131 samples allowed the fats and proteins to precipitate. The samples were centrifuged for -10 minutes at -2000 rpm. The supernatant was then loaded onto the conditioned clean-up column fitted inside the silanized pear shaped flask. The eluate was collected in the silanized pear-shaped flask. This process removed the fats and proteins from the extract. Ten milliliters o f acetonitrile was added to die samples left in the centrifuge tube. The remaining sample in the tube was homogenized using a tissumizer for -30 seconds. The tissumizer was rinsed with ten milliliters of acetonitrile. The rinse was added to the sample tube. The samples were allowed to shake on a wrist action shaker for another 10 minutes. The samples were placed into a freezer for ~1 hour. The samples were centrifuged again for -10 minutes at -2000 rpm. The supernatant was then loaded onto the same clean-up column. The eluate was collected into the same pear-shaped flask. The column was washed with 20 mL of acetonitrile, collecting the wash in the pear-shaped flask. Approximately 3 or 4 drops o f 1-octanol was added to the extracts in the flasks. The samples were evaporated using a rotary evaporator at reduced pressure. The 1-octanol held the analytes in the pear-shaped flask while the solvent evaporated. Two milliliters o f 2% ascorbic acid in methanol was added to the flasks to make final volume. The flask was swirled to dissolve and mix the sample. The sample was transferred to a HPLC vial using a disposable pipet. Each sample was analyzed by LC/MS/MS electrospray. *Due to lack of sample, a one-gram portion of sample C0048133 (DMC-F01-MSW001 0-041216) was used. All other samples were weighed out in 5-gram portions. 6.2 Preparation of Standards and Fortification Solutions Individual stock standard solutions of PFBS, PFHS and PFOS were prepared as specified in Exygen method V0001783. The stock standard solutions were prepared at a concentration of 100 pg/mL by dissolving 10 mg of each o f the standards (corrected for purity and salt content) in methanol. From these solutions, mixed 1.0 pg/mL fortification standard solutions were prepared by taking 1 mL of each o f the appropriate stock solutions and bringing the volume up to 100 mL with methanol. By taking 10 mL of the mixed 1.0 pg/mL fortification standard and bringing the volume up to 100 mL with methanol, a mixed 0.! pg/mL fortification standard was prepared. A mixed stock standard solution of PFBS, PFHS, and PFOS was also prepared. The stock standard solution was prepared at a concentration of 1000 pg/mL by dissolving 100 mg of each of the standards (corrected for purity and salt content, if necessaiy) in methanol. From this solution, a 100 pg/mL fortification standard solution was prepared by taking 10 mL of the stock and bringing the volume up to 100 mL with methanol. By taking 10 mL of the 100 pg/mL fortification standard and bringing the volume up to 100 mL with methanol, a 10 pg/mL fortification standard was prepared. By taking 10 mL of the 10 pg/mL fortification standard and bringing the volume up to 100 mL with Exygen Research Page 16 o f 132 Interim Report # 10-Analysis o f Fish and G am Samples Exygen Study No.: P0001131 methanol, a 1.0 pg/mL fortification standard were prepared. By taking 10 mL of the 1.0 pg/mL fortification standard and bringing the volume up to 100 mL with methanol, a 0.1 pg/mL fortification standard was prepared. A set o f non-extracted standards containing PFBS, PFHS and PFOS was prepared in methanol. The following concentrations were prepared: Cone, of Fort Fort Solution Volume (ng/mL)1 (mL) 1.0 5.0 1.0 2.5 1.0 1.0 0.05 10 0.025 10 0.01 10 0.005 10 1of PFBS, PFHS and PFOS Volume of Fortified Sample (mL) 100 100 . 100 100 '100 100 100 Final Cone, of Calibration Std. (pg/mL) 0.05 0.025 0.01 0.005 0.0025 0.001 0.0005 The stock standard solution and all fortification and calibration standard solutions were stored in a refrigerator (4 2C) when not in use. Documentation of standard preparation is located in the raw data package associated with this interim report. 6.3 Chromatography Quantification of PFBS, PFHS and PFOS was accomplished by LC/MS/MS electrospray. The retention times of PFBS, PFHS and PFOS were ~ 4.7 mins, - 10.8 mins, and ~ 12.9 mins, respectively. 6.4 Instrument Sensitivity The smallest standard amount injected during the chromatographic run had a concentration o f 0.0005 pg/mL o f PFBS, PFHS and PFOS. 6.5 Description of LC/MS/MS Instrument and Operating Conditions Instrument: API 4000 Biomolecular Mass Analyzer Interface: Turbo Ion Spray Liquid Introduction Interface Computer: DELL OptiPlex GX400 Software: Windows NT, Analyst 1.4.1 HPLC: Hewlett Packard (HP) Series 1100 HP Quat Pump HP Vacuum Degasser HP Autosampler Exygen Research Page 17 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 HP Column Oven HPLC Column: Thermo Fluophase RP, 50 mm x 2.1 mm Column Temp.: -30 C Injection Voi.: 15 pL Mobile Phase (A): 2 mM Ammonium Acetate in water Mobile Phase (B): Methanol Time (min) 0.0 1.0 8.0 10.0 11.0 18.0 Total run time: -13 min Flow Rate: 0.3 mL/min %A 65 65 25 25 65 65 %B 35 35 75 75 35 35 Ions monitored: Analvte Mode PFBS PFHS PFOS negative negative negative 6.6 Quantitation and Example Calculation Transition Monitored 299 99 399 80 499 80 Approximate Retention Time (min) -4.7 min. ~10.8 min. ~12.9 min. Fifteen microliters of sample or calibration standard were injected into the LC/MS/MS. The peak area was measured and the standard curve was generated (using 1/x fit weighted linear regression) by Analyst software using six concentrations of standards. The concentration was determined from the following equations. Equation 1 calculated the amount o f analyte found (in ng/mL, based on peak area) using the standard curve (linear regression parameters) generated by the Analyst software program. Equation 1: Analyte found (ng/mL, wet weight) * (Peak area - intercept) x DF slope Where: DF = Dilution Factor, factor by which the final volume was diluted, if necessary. Exygen Research Page 18 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Equation 2 was used to convert the amount o f PFBS, PFHS and PFOS found in ng/mL to ng/g (ppb). Equation 2: Analyte found (ppb, wet weight) = TAnalyte found (ng/mL) x final volume (2 mLll sample weight (5 g) For samples fortified with known amounts of PFBS, PFHS and PFOS prior to extraction, Equation 3 was used to calculate the percent recovery. Eouation 3: Recovery (% ) " (analyte found (ng/mL) - analyte in control (ng/mL)) xlOO% amount added (ng/mL) Note: For the analyte recovery calculation, the "control" is the unspiked aliquot of the primary field sample. An example of a calculation using an actual sample follows (calculation is for PFBS only): Fish sample Exygen ID: C0049478 Spk E (Set: 061605E), fortified at 10 ng/g with where: peak area = 280750 intercept = 3070 slope = 13100 dilution factor =1 ng/g PFBS added (fort level) = 10 amt in corresponding sample = final volume (mL) * NQ (not quantifiable) 2 sample weight (g) =5 From equation 1: Analyte found (ng/mL, wet weight) - r280750 - 30701 x 1 13100 - 21.2 ng/mL From equation 2: Analyte found ng/g (ppb) = 121.2 nz/mL x 2 mLl 5g = 8.48 ng/g (ppb) Exygen Research Page 19 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 From equation 3: % Recovery = 18.48 ne/g - One/et x 100% 10ng/g = 85% 7.0 EXPERIMENTAL DESIGN For fish and clam samples designated as laboratory matrix spikes, PFBS, PFHS, and PFOS were added at a known concentration to the samples in the laboratory after the samples were weighed for extraction. The fish samples were extracted in thirty sets, five that were re-extraction sets. The clam samples were extracted in one set. Each fish or clam set included one control fish matrix blank and two control fish matrix blanks fortified at known concentrations. Two sets of fish contained three samples, one set of fish contained four samples, one set of fish contained seven samples, and twenty-one sets of fish contained five samples. Three sets of re-extracted fish contained ten samples each. One set o f re-extracted fish contained twelve samples. One set o f re-extracted fish contained nine samples. One set o f clams contained six samples. Accuracies were assessed for each sample by reviewing the individual QC results obtained for each sample. 8.0 R ESU LTS Analytical results and assessed accuracies for the analysis o f PFBS, PFHS and PFOS found in fish samples are summarized in Table I. Please note that the chromatography for PFBS was poor for this matrix. Exygen recognizes this and great care has been taken to integrate each data set consistently. Analytical results and assessed accuracies for the analysis of PFOS found in the re-extracted fish samples are summarized in Table II. Analytical results and assessed accuracies for the analysis of PFBS, PFHS and PFOS found in clam samples are summarized in Table III. The average percent recoveries standard deviations for PFBS, PFHS, PFOS in fish samples were 79% 13%, 94% 12%, and 85% 20%, respectively. Fortification recoveries for PFBS, PFHS, and PFOS in fish samples are summarized in Table IV. The average percent recovery standard deviation for PFOS in the re-extracted fish samples was 109% 20%. Fortification recoveries for PFOS in the re-extracted fish samples are summarized in Table V. The average percent recoveries standard deviations for PFBS, PFHS, PFOS in clam samples were 84% 10%, 93% 13%, and 73% 16%, respectively. Fortification recoveries for PFBS, PFHS, and PFOS in clam samples are summarized in Table VI. Exygen Research Page 20 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study N o.: P0001131 9.0 CONCLUSIONS The fish and clam samples were successfully extracted and analyzed for PFBS, PFHS and PFOS according to analytical method V0001783. 10.0 RETENTION OF DATA AND SAMPLES All original paper data generated by Exygen Research that pertains to this interim report will be shipped to the study director. This does not include facility-specific raw data such as instrument or temperature logs. Exact copies of all raw data, as well as a signed copy of the final analytical report and all original facility-specific raw data, will be retained in the Exygen Research archives for the period o f time specified in EPA TSCA Good Laboratory Practice Standards 40 CFR 792. Exygen Research Page 21 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: POOO1131 TABLES Exygen Research Page 22 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygcn Study No.: P0001131 Table I. Summary of PFBS, PFHS and PFOS in Fish Samples Exygen ID Client Sample ID C4 Sulfonate PFBS Pwftsenpbatew--trltenste Analyte Assessed Found Accuracy filetti retwt (*)%) CO Sulfonate PFHS PsHtewahareswallnnste Analyte Assessed Found Accuracy (nofa) Mint M. (*-% ) CS Sulfonate PFOS Psrltuoruoctrru wtirortsa Analyte Assessed Found Accuracy (rtf!) *"<(. (-% ) C0048126 DL3-F01-IPF001-0-041021 0.309 40 ND 30 5.18 30 C0048126 Rep DLS-F01-IPF001-0-041021* 0.318 40 ND 30 5.84 30 C0048127 DL3-F01-IPF002-0-041021 0.286 40 NO 30 4.18 40 C0043127 Rap DL3-F01-JPF002-0-041021" 0.382 40 ND 30 4.56 40 C004812S DMC-F01-IPF001-CKM1021 0.330 50 ND 50 7.40 50 C0048128 Rep DMC-F01-IPF001-0-041021* 0.366 50 ND SO 8.38 50 C0043129 DMC-F01-IPF002-0-041021 0.712 30 ND 30 14.1 50 C004812B Rap DMC-F014PF002-0-041021 * 0.792 30 ND 30 17.0 50 C0048130 DMC-F01-IPW001-0-041021 1.08 40 NO 30 70.0 40 C0048130 Rap DMC-F01-IPW001-IMM1021* 1.06 40 0297 30 58.4 40 C0043131 DMC-FC1-IPW002-0-041021 0.988 50 ND 30 34.4 30 C0048131 Rep DMC-F01-IPW002-0-041021* 1.34 50 ND 30 34.2 30 C0048132 DMC-F01-MSW001 -0-041021 0.218 30 0.524 30 270 30 C0048132 Rap DMC-F01-M8W001-0041021* 0.202 30 0.484 30 280 30 C0048133* DMC-F01-MSW 002-0041021 NO* 30 ND* 30 530 30 C0048133 Rep* DMC-FO1-MSW002-CF041021" NO* 30 ND* 30 546 30 C0048134 DL24-01-IPF001-0-041021 0.660 50 NO 40 3.45 50 C0048134 Rep DL2-F01-IPF001-0-041021* 1.00 50 NO 40 2.86 50 C0048135 DL2-F014PF002-0-041021 0.B16 50 ND 40 4.72 50 C004813S Rep DL2-F01-IPF002-0-041C21* 0.952 50 ND 40 5.04 50 C0043136 DL2-F014PF003-0-041021 0.247 30 ND 30 201 30 C0048138 Rep DL2-F01-IPF003-0-041021 ' 0.218 30 ND 30 208 30 C0048137 DL2-F01-IPF004-0-041021 0.348 40 ND 30 4 52 40 C0048137 Rep DL2-F01-IPF004-0-041021 * 0.330 40 ND 30 4.12 40 C0049138 DL2-F01-IPF00S-O-041021 0.338 40 ND 30 2.46 30 CO048138 Rep DL2-F01-IPF005-0-041021* 0.318 40 ND 30 2.80 30 C004813B DL2-F01-IPW001-0-041021 0.328 30 ND 30 5.88 30 C004ai3B Rap DL2-F01-tPW001-0-041021* 0.400 30 ND 30 6.12 30 C0048140 DL2-F01-MSF001-0-041021 0.333 30 ND 30 77.2 30 C0048140 Rap DL2-F01-MSF001-0-041021" 0.348 30 ND 30 #7.6 30 C004B141 DL2-F01-M5F002-0-041021 0.408 40 ND 30 94.8 40 C0048141 Rap DL2-F01-MSF002-0-041021* 0.308 40 ND 30 98.4 40 C0048142 DL2-F01-MSF003-0-041021 0.416 40 NO 40 41.6 40 C0048142 Rep DL2-F01-MSF003-0041021* 0.378 40 ND 40 40.0 40 C0048143 DL2-F01-MSF004434J41021 0.345 30 ND 30 S3.6 30 C0048143 Rap DL2-F01-MSF004-0-04102T 0.349 30 ND 30 63.2 30 "Laboratory D uplicate * 1.0 g of ample used for srtscttan Instead of5.0 g. ND Not detected at or aoowa Limit of Quantitation which Is 0.2 ng/g (wot weight) HD* Not detected at or above Limit of Quantitation which 1.0 ntfg (wet weight). NR Not reported due toquality control result failure. Exygen Research Page 23 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table I. Summary of PFBS, PFHS and PFOS in Fish Samples (continued) Exyoen ID C0048144 COO481*4 Rep C004814S C004814S Rep C0O48146 C0048146 Rap C0O48147 C048147 Rep C0046148 C0048148 Rep C048148 C004814 Rep C0048160 C0048150 Rap C0048151 C0048151 Rep C00481S2 C0048152Rep C0048153 COO48153 Rep C0048154 C0048154 Rep C0048156 C0048155 Rep C0048158 COO48108 Rep C0048157 C0048157 Rap C0048158 C0048158 Rep C00481S6 C0048159 Rep C0048100 C0048160 Rep C0043161 C0048161 Rep C0048182 C0048182 Rep Oiarrt Samp e ID C4SuMonat PFBS Analyte Assetsed Found Accuracy nam;m m . (+F%) C6Sulfonate PFHS ParflucrohcxwiMuWonnt Analyte Assessed Found Accuracy ("o/B)<etn. (*- %> CS 8ulf6nate PFOS Analyte Aseeeeed Found Accuracy (ntMixetva. (/-%> DL1-F01-IPF001-0-041021 0.432 30 0.261 30 33.4 40 DL1-F01-IPF001-0-041021* 0.428 30 0.252 30 32.8 40 DL3-FO1-IPFOOM-O41021 0.448 30 ND 30 3.55 30 DL3-F01-IPF003-0-041021* 0.440 30 0.228 30 3.36 30 DL3-F01-IPF004-0-041021 0.784 40 NO 30 0.382 30 DL3-F01-IPF004-0-041021* 0.900 40 ND 30 0.230 30 DLS-F01-IPF005-0-041021 1.63 60 NO 30 5.48 40 DL3-F01-IPFCXI5-0-C41021 1.07 80 ND 30 5.84 40 DBC-F01-IPF0Q1-O-041021 D6C-F01-PF001-O-O41021* 1.34 1.38 SO 0.682 50 0.98G 30 30 NR NR NR NR DBC-F01-IPF002-0-041021 1.43 50 NO 30 110 30 DBC-F01-IPF002-0-041021* 0.766 50 ND 30 109 30 DBC-F01 -IPF003-0-04 1021 0.868 SO 0844 30 NR NR O0C-F1-IPF003-0-041021 * 0.652 50 0.624 30 NR NR OBC-F014PW001-04341021 0.864 40 2.21 30 DBC-F014PW001434)41021* 0.5C8 40 220 30 NR NR NR NR DBC-F01-IPW002-0-041021 0.700 40 8.12 30 NR NR DBC-F01-IPWOQ2-0-Q41021* 0.738 40 6.8 30 NR NR DBC-F01-IPWOSO-C41021 0.738 30 6.76 30 DBC-F01-IPW003-0-041021" 0.780 30 744 30 NR NR NR NR DOU-F01-IPF001-0-041021 DOU-F01-!PF001^M)4I021* 4.38 4.24 40 1.02 40 0.872 30 30 NR NR NR NR DOU-F01-IPF002-0-041021 DOU-F01-IPF00M-041021* 8.78 8.28 40 0.702 40 0.720 30 30 NR NR NR NR DOU-F01-IPF003-0-041021 5.88 40 1.02 30 DOU-F01-IPF0OS4MM1O21* 6.86 40 1.02 30 NR NR NR NR DOU-F01-IPF04-0-041021 8.12 30 2.30 30 NR NR DOU-FG1-IPF004-0041021* 0.28 30 2.38 30 NR NR DOU-F01-IPF00S-0-41021 456 30 0.856 30 NR NR DOU-F01 -IPF00S4>-O41021* 4.36 30 0.382 30 NR NR DOU-FOI-IPW001-0-041021 2.69 30 0.948 30 NR NR DOU-F01-IPW001-C-41021* 2.81 30 0.964 30 NR NR DOU-FOI -IPW002-0-041021 16.4 50 3.69 30 NR NR DOU-F01-IPW002-0-041021* 10.3 50 3.70 30 NR NR DL2-F01-MSW001-04X1021 0-203 30 NO 30 120 50 OL2-F01-MSW14F041021* ND 30 NO 30 120 50 DL2-F01-MSW002-0-041021 NO 30 ND 30 102 30 DL2-F01-MSW002-0-041021* NO 33 ND 30 100 30 Laboratory Duplicate NO Not detected at or above Lim it of Quantitation which is 0.2 ng/g (wet weight). NR Not repotted due to quality control neault felturee. Exygen Research Page 24 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: POOO1131 Table I. Summary of PFBS, PFHS and PFOS In Fish Samples (continued) Exycen ID C0046163 C0046163 Rap C0O48164 C0048164 Rep C0048165 COCKS185 Rap C0049450 C00484SO Rep C0049451 COO40451 Rep C0048452 00049452 Rap CD049453 C0049453 Rep C0049454 C0049454 Rap C0049455 C0049456 Rap C0049458 C0C49456 Rep C0049457 C0048457 Rep C0049458 C0049458 Rap 00048459 C00445B Rap C0049460 C0049460 Rap C49451 C0049481 Rep C0049462 C0049462 Rep C0049463 C0049463 Rep C0048484 C0049484 Rep C0049485 C0049485 Rep Client S sm g i^ j^ ^ C4 Sulfonate PFSS Pufluorebutan--ulfow te Analyte - A lien ed Found Accuracy (no41wwt (>!-%) DL2-F01-WSW003-0-041021 DL2-F01-MSW003-0-041021* DL2-F01-MSW004-0-041021 DL2-F01-MSW004-0-041021* DL2-F01-MSW005-0-041021 DL2-F01-M3W0054J-041021* DBC-F01-M3WOO1-0-041104 DBC-F01 -MSW001-04)41104* DBC-F014t4SW0024W)41104 OBC-F01446W0024WM1104* DBC-F01-MSW003-0-041104 DBC-F01 -MSW003-0-041104* DBC-F01-USW004-0-C41104 DBC-F01-USW004-0-041104* DBC-F01-MSW005-0-041104 DBC-F01-M8W005-0-041104* DBC-FO1-M8F001-04)41104 DBC-F01-MSF001-0-041104* OBC-FO t -MSF002-04>41104 DBC-FO1-MSF002-0-041104* DBC-F014PF004-0-041104 DBC-F01-IPF00441-041104* DBC-F01-IPF05-0-041104 DBC-F01-IPF0054)4)41104* DBC-FOI-IPW004-0-041104 DBC-F014PW004-04)41104* DBC-F014PW0084W)41104 DBC-F01-IPW0064)-041104* DOU-F01-IPW003-0-041104 DOU-FO1-IPW003-0-041104* DOU-F01-1PW004-0-041104 DOU-F01-IPW004--041104* DOU-FO1-IPW006-04H1104 DOU-F01-1PW0054KI41104* DL2-F01-IPW0024MM1104 DU-F0HPW 002-0-041104* DI.1-F01-MSW001-0-041104 DU-F01-M 8W 0014M M 1104* ND ND ND ND ND ND 0.888 0.652 0.888 0.884 0.880 0.784 0.884 0.680 0.832 0.832 0.820 0.788 0.788 0.780 0.504 0.S20 1.60 1.84 0.484 0.444 0.572 0.532 3.12 ,3.05 3.40 3.11 2.97 2.90 1.04 0.960 1.20 1.24 30 30 30 30 30 30 30 30 30 SO 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 ., 30 30 30 30 30 30 30 30 30 30 30 C6 Sulfonate PFHS Partluorohman--utfonati Analyte Aaeeind Found Accuracy OWJHMtM (W-%) ND ND NO NO ND ND 8.32 8.32 3.43 3.35 3.82 3.64 9.72 9.80 Z77 . 2.66 3.58 3.48 1.94 -1.90 0.584 , 0.540 2.16 Z18 3.28 3.28 2.10 2.10 600 592 7.12 8.84 8.12 5.32 ND. ND 0.896 0.932 30 30 30 30 30 30 30 30 30 30 . 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 ; SO 30 30 30 30 30 30 30 30 30 30 30 C9 Sulfonate PFOS P+rtiuorooctan--uionafci Analyte Assessed Found Accuracy (nofoimwt < **> 92.4 30 91.8 30 88.0 30 94.8 30 168 30 174 30 NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR 142 30 152 30 NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR 8.20 40 8.60 40 NR NR NR NR ` Laboratory Duplicate NO * Not detected at o r above Lim it o f Quantitation which is 0.2 ng/g (wet weight). NR * N ot reported due to quality control laault failure* Exygen Research Page 25 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 Table I. Summary of PFBS, PFHS and PFOS in Fish Samples (continued) Exveen ID C0049408 C00494B6 Rep C0049487 C004947 Rep C0049468 C0049458 Rep C0049460 C049469 Rap 00049470 C0049470 Rep C0049471 C0049471 Rep C0049472 C0048472 Rep 00049473 C0049473 Rep C0049474 C0049474 Rep C0O49475 00049475 Rap C0049478 C0O49476 Rep C0049477 C0049477 Rep C004947B C0043478 Rep 00046479 C0049479 Rp 00049480 C0049480 Rep C0049481 0004948t Rep 00049482 C0049482 Rep C0049483 C0049483 Rep C0049484 C0049484 Rep Client Sample ID C4 Sultanat* PFBS Parfkiofotouan--u lfty iaf Anafyle Assessed Found Accuracy r<um)twi (/-S) Cfl Buttons* PFHS Prfluofphaxan--uffona Analyte Assessed Found Accuracy (nota)et *. (/-%) C t Sulfonate PFOS PwHu--o o c t a n --i Analyte Assessed Found Accuracy (nstfllsetwl. <*-% ) DL1-F01-MSW002-0-041104 1.30 30 1.72 30 IX 1-FOI-MSW002-0-041104* 1.15 30 1.54 30 DL1-F01-MSW003-0-041104 0.580 30 1.13 30 DL1-F01-MSW0050-041104* 0.528 30 1.14 30 DL1-F01-MSW004-0-041104 DL1-F01-MSW004-0-041104* 1.22 1.12 30 0.908 30 30 0.880 30 DL1-F01-MSW005-0-041104 1.22 30 1.18 30 DL1-F01 -MSVY005-0-041104* 1.20 3C 1.1B 30 DL1-F01-M3F01-0-041104 DL1-F01-MSF01-0-041104* ND ND 30 0.315 30 30 0 299 30 DL1-F01-MSF002-0-041104 DL1-F01-A4SF002-0-041104* ND ND 30 0.624 30 0.604 30 30 DL1-F01-MSF003-0-041104 OL1-F01-MSF003-0-41104* 0.203 ND 30 0.652 30 0.676 30 30 DL 1-F 01-WSF004-0-041104 DLt-F01-M6F004-0-041104* ND NO 30 0.532 30 30 0.538 30 OL 1-FC1-M SF005-0-041104 DL1-F01-MSFQ05-0-041104* ND ND 30 0.684 30 30 0.844 30 DL1-F01-IPW001-0-041104 0.772 30 0.566 30 DL1-F01-IPW001-0-041104* 0.870 30 0.744 30 DL 1-F 01-IPW 002-0-041104 0.468 30 0.536 30 DL1-F01-IPW002-0-041104* 0.472 30 0.618 30 DL1-F01-IPW003-0-041104 L1-F01-IPW003-0-041104* ND ND 30 0.209 30 0247 30 30 DL1-F01-IPW004-0-041104 DL1-FOHPW004-0-041104* DU -F01-IPF002-0-041104 DL1-F01-IPF002-0-041104* ND NO NO NO 30 0.238 30 30 0.246 30 30 0.222 30 30 0.247 30 DL-1-F01-IRF003-0-041104 DL-1-F01-1PF003-0-041104* ND ND 30 0.265 30 30 0.262 30 DL-1-F01-IPF004-0-041104 NO 3G ND 30 DL-1-F01-IPF004-0-041104* ND 30 ND 30 DL-1-F014PF006-D041104 DL-1-F01-IPF005-0-041104* ND ND 30 0.373 30 30 0.366 30 DL3-F01-MSW001-0-041105 ND 30 NO 30 DL3-F01-M6W001-0-041105* ND 30 ND 30 DL5F01-MSW002-0-041105 ND 30 ND 30 DL3-F01-MSW002-0-041105* ND 30 ND 30 NR NR NR NR 181 186 NR NR 293 307 NR NR NR NR NR NR NR NR 17.4 18.9 15.7 15.8 9.36 9.52 56.6 57.2 11.1 11.2 20.9 21.4 7.60 7.36 8.72 8.56 139 137 180 162 NR NR NR NR 30 30 NR NR 30 30 NR NR NR NR NR NR NR NR 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 ` Laboratory Duplicate NO Not detected at or above Lim it of Quantitation whlctt It 0.2 ng/g (wet weight). NR Not reported due to quality control m u lt failures. Exygen Research Page 26 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 Table I. Summary of PFBS, PFHS and PFOS in Fish Samples (continued) Exygen 10 Client Sample ID C4 Sulfonata PFBS RtrfluorotutMMuifoiMi Analyte Amssed Found Accuracy (ro*o)wN4. C0049485 C0049485 Rep C049486 C0049486 Rep C0049487 COC48487 Rep C0049488 C0048488 Rep 00049489 COC40489 Rep C0049490 C0048490 Rep C0049491 C0049491 Rap C0049492 00049492 Rap C0049493 C0049493 Rep C0049494 C0049494 Rap 00049495 C0049495 Rep C004949S C0049496 Rep C0Q49497 C0049497 Rep C0049498 CC049498 Rep C0049499 C0049499 Rep C0049500 C0049500 Rep C0049501 C004950IRep C0049502 COO48502 Rap C0049503 C0049603 Rep DL3-F01-MSW003-0-04105 DL3-F01-MSW0030-041105* DL3-F01-M8W004-0-041106 DL3-F01-MSW004-0-041105* DL3-F01-MSW005-0-041105 DL3-F01-MSW005-0-041105* DL3-F01-MSF001-0-041105 DL3-F01-MSF001-0-041106* DL3-F01-MSF0C2-0-041105 DL3-F01-MSF002-0-041105* DL3-F01-M3F003-C-041105 DL3-F01-MSF003-0-041109* DL3-F01-M8F004-0-041106 DL3-F01-MSF004-004110S* DL3-F01-MSF005-0-041105 DL3-F01-MSF006-0-041105* DMC-F01- IPW003-0-041106 DMO-FOI-IPW003-0-041105* DUC-F01-IPW004-0-041106 DMC-F01-IPW004-0-041105* DMC-F01 -IPWOOfWHH1105 DMC-FO1-IPWOO6-0-O41105* DMOF01-WSW003-0-041106 DMC-F01-MSW003-0-041105* DMC-F01-MSW004-0-041105 DMC-F0144SW004-0-041105* DMC-F01-MSW005-0-041105 DMC-F01-MSW005-0-041105* OMC-FO1-M6F0024MI41105 OMC-F01-MSF002-0-041106* DMC-F01-MSF003-0-041105 DMC-F01-MSF003-0-041105* DMC-F01-USF004-0-041105 QMC-F01-MSFC04-0-041105* DMC-F01 -MSF005-<M)41105 DMC-F01-MSF005-0-041105* DMC-F01-IPF003-0-041105 DWC-F01-IPF003-0-041105* ND ND NO ND ND ND NO ND ND NO ND ND ND ND ND NO ND ND 0.300 0.345 ND ND ND ND 0.313 0.233 ND ND ND ND ND NO NO NO ND ND NO ND 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 C6 Sulfonata PFKS P a rih m o rw u m u fo n rti Analyte Aeaeated . Found Accuracy CB Sulfoiuta PFOS Analyte Aaaeaaed Found Accuracy (nota)wM** t-N-%1 ND ND 0.285 0.282 0.366 0.341 ND ND ND ND ND ND ND NO ND ND 0.386 0.334 0.306 0.464 0.270 C.255 0.592 0.560 1.90 1.88 0.980 0.936 NO 0.387 NO NO 0.254 0.228 0.420 0.448 ND ND 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 148 144 198 199 173 168 116 115 118 111 75.6 72.0 130 134 81.6 83.8 61.2 63.6 68.6 696 64.4 62.8 354 330 259 269 NR NR NR NR 568 68.0 116 119 146 147 7.92 7.92 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 NR NR NR NR 40 40 30 30 30 30 30 30 ` Lb orw ory D uplicate NO * Not detected at or abova Unit of Quantitation wtiichk 0.2 ng/g (wet weight). NR N ot reported due to q u a lity oonbol reault failure. Exygen Research Page 27 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygcn Study No.: P0001131 Table 1. Summary of PFBS, PFHS and PFOS in Fish Samples (continued) Exvoonto C0049504 C0049504 Rep C004BS05 C0049505 Rep C0056354 C0056354 Rep C0066455 C0O56456 Rep C0056456 C0056456 Rep C005457 00056457 Rep COOS6458 COO56456 Rep C0066459 C0056450 Rep C0056480 C0056460 Rep COO36401 C0056461 Rap C0056462 C0056482 Rap C0066463 C0056483 Rep C0056464 C0056464 Rap C0056495 COO56485 Rap C0056466 C0056466 Rep CO056497 C0056487 Rap C0056468 COO56468 Rep C00584S9 COO56460 Rep C0056470 COO56470 Rep Client Sampls ID DMC-F01-IPF004-0-041105 DMC-F01-IPF004-0-041106* DMC-F01-fPF0054W41105 DMC-F01-IPF005-0-041105* DL2-F01-M8F0054KM1216 DL2-F01 -MS F005-0-041216* DL2-F01-IPW006-0-041215 DL2-F01-IPW005-0-041215* DOU-FOI -M3WX2-0-041216 DOU-F01-MSW002-0-C41216* DL2-F01-IPWOQ4-0-041215 DL2-F01-IPW004-6-041215* DOU-FOt-MSWOO3-04)41216 DOU-F01-WSW003-0-041216* OL3-F01-tPW004-0-041130 DL3-F01-iPW004-04)41130* DOU-F01-MSW001-0-041216 DOU-F01-MSW001-0-041216* DOU-F01-MSW004-0-041216 DOU-FOI-MSW004-0-041216* DOU-F01-M8W005-0-041216 DOU-FOI-M3W005-04J41216* DOU-FOI-MSF005-0-041216 DOU-F01-MSF005-0-041216* DL3-F01-IPW002-0-041130 DL3-F01-IPW002-0-041130* DL3-F01-IPW003-0-041130 DL3-F01-IPW003-0-041130* DOU-FOI-MSF003-0-0412t6 DOU-F01-M6F003-0-04I216* O0C-FO1-MSFOO6-t-O41216 DBC-F01-MSF006-1-041216* DBC-F01-MSF008-0-041218 DBC-F01-MSF005-0-041216* DOU-F014I4SF002-0-041216 DOU-F01-MSF002-0-041216* DL3-F01-IP W X 1-0-041130 DL3-F01-IPW001 -0-041430* C4 Sulfonate PFaa PartlucwliMaiwirifwurt Analyte Asboaead Found Accuracy ( -*) ND 30 ND 30 NO 30 ND 3C ND 30 ND 30 ND ND 0.206 0.227 ND ND 30 30 40 40 30 30 ND 30 NO 30 NO 30 ND 30 7.08 X 7.12 X 4.72 40 4.68 40 0.364 0.36 1.41 1.43 X X X X ND X ND X ND X ND X 1.72 X 1.73 X 0.220 ND NO ND 0.888 0.872 ND ND X X X X X X X X ce Sulfonate PFHS Pw terejglgaeg efoaato Analyte Assoaeed Found Accuracy (*P *>) C l Sulfonate PFOS ParltuM poaanaeiilfonata Analyte Assessed Found Accuracy (noro)'vel'M, <*>- *1 ND X 44.0 X ND X 456 X ND 30 46.4 X ND X 46.0 X ND 30 49.2 X ND X 51.6 X ND X ND X 13 2 X 133 X 1.72 1.70 X NR NR NR NR ND X 6 .X X ND X 880 X 10.4 X 10.6 X NR NR NR NR ND X 7.92 X ND X 752 X 15.4 X 14.2 X NR NR NR NR 21.7 X 22.5 X NR NR NR NR 1.34 X 1.41 X NR NR NR NR 8.40 X 8.44 X NR NR NR NR ND 30 12.0 X ND X 11.5 X ND X NO X 6.44 40 5.28 40 8.50 X 6.68 X NR NR NR NR 1.79 X 1.76 X NR NR NR NR 1.66 X 1.5 X NR NR NR NR 4.56 30 NR NR 4.56 X NR NR 0.246 0.216 X X 9.04 30 8.80 X Laboratory Dupflcste NO Not detected at or above Lim it o f Quantitation w titd i is 0.2 ngfg (vet weight). NR - Not reported due to quaflty control result M lurea. Exygcn Research Page 28 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Table I. Summary of PFBS PFHS and PFOS in Fish Samples (continued) Exvoen ID C0056471 COOS6471 Rep COO50472 COO56472 Rep C0056473 COO56473 Rep C0056474 C005474 Rep C0056475 COO56475 Rep C005647S COO56476 Rep C0056477 C0056477 Rep C0056478 C0056478Rep C036479 COO56479 Rep Citent Sample ID C4 Sulfonate PFBS PoaffeiMPOtMrtanooIfVoflito Analyte Asaaaeed Found Accuracy [rxyowatwl (/- %1 C8 Sulfonate PFHS Analyte Found (noArtwwwt Aaaeseed Accuracy (/-%> DOU-F01-M8F002-1-041216 DOU-F01-MSF002-1-041216* DOU-F01-MSF004-0-041218 DOU4F01-MSF004-0-041216* DOU-F01-MSF001-0-041218 DOU-FO1-MSF001-0-041216* DBC-F01-MSF003-0-041216 D6C-F01 -MSF003-0-041216' DBC-F01-MSF004-0-041216 DBC-F01-MSF004-0-041216* DMC-F01-MSF001-04MI216 DMC-F01-MSF001 -0-041216* DL1-F01-IPW005-0-041216 DL1-F01-IPW005-0-04I216* DL2-F01-IPW003-0-041201 DL2-F01-IPW003-0-041201* DL3-F01-1PW0C5-0-041201 DL3-F01-IPW005-0-041201 0.640 0.860 0.792 0.616 Z18 2.28 ND ND ND ND ND NO 0.253 0.249 ND NO ND ND 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30 4.88 4.72 2.35 2.22 10. 10.7 2.09 2.09 3.87 3.77 0420 0.456 0.844 0644 ND ND ND ND 30 30 40 40 30 30 30 30 30 30 30 30 30 30 30 30 30 30 CS Sulfonate PFOS Analyte Found NR NR NR NR NR NR NR NR NR NR NR NR 88.8 67.8 7.48 7.44 4.88 4.72 ASMMMd Accuracy </-%1 NR NR NR NR NR NR NR NR NR NR NR NR 30 30 40 40 30 30 ' Laboratory D uplicate ND - Not detected et or above Limit of Quantitation which it 0.2 ng/g (wet weighty NR = Not ivponod dut toquality control rerun failure*. Exygen Research Page 29 of 132 Interim Report #10-Analysis o f Fish and G am Samples Exygen Study No.: P0001131 Table II. Summary of PFOS in Re-extracted Fish Samples Exygen ID C0048151 C0048152 C0048153 C0048154 C0048155 C0048158 C0048180 C0049450 C0O49451 C0049452 C0049453 C0049454 C0049455 C0049456 C049458 C0049459 C0049460 C0049461 C0049402 C0049463 C0049465 C0049466 C0049467 C0049469 C0049471 C0049472 Client ^S am ptel^^ DBC-F01-IPW001-0-041021 DBC-F01-IPW002-0-041021 DBC-F01-IPW003-0-041021 DOU-FW-IPF001-0-041021 DCXJ-F01-IPF002-0-041021 DOU-F01-IPF005-C-041021 DOU-F01-IPW002-0-041021 DBC-F01-MSW001-0041104 OBC-F01-MSW002-0-041104 DBC-F01-M3W0030-041104 DBC-F01-MSW0040041104 DBC-F01-MSW005-0-041104 DBC-F01-MSF001-0041104 0BC-F01-MSF002O-041104 OBC-F01-IPF005-0-041104 DBC-F01-IPWG04-0-G41104 DBC-F01-IPW005-0041104 DOU-FOI-IPW003-0041104 DOU-F01-IPW04OO41104 DOU-F01-IPWC05-0041104 DL1-F01-MSW001-0-C41104 DL1-F01-MSWGC2-0-041104 DL1-F01-MSW0030-041104 DL1-F01-MSW005-0-Q41104 DL1-F01-MSF002-0-041104 DL1-F01-MSF003-0-041104 NR * Not (portad du lo quality control result failures Exygen Research CS Sulfonate PFOS P *rllu o n > o c ta n M i> ifo n a t* Analyte Found Assessed (rto lg ) W st W slght Accuracy(+/- %) 2080 30 3330 1660 30 50 596 30 838 544 1210 30 30 40 9480 7520 12600 3440 40 30 30 30 5240 30 5240 30 4780 608 1230 2620 NR 1740 30 30 40 30 NR 30 2130 968 708 AQf t 604 568 432 30 30 30 30 30 30 30 Page 3 0 o f l 3 2 Interim Report #10-Anaiy8is o f Fish and Clam Samples Exygen Study No.: P0001131 Table IL Summary of PFOS in Re-extracted Fish Samples (continued) Exygen IO C0M9474 COM9498 C0049496 C0056456 C0056458 COC56460 COC56461 C0056462 COO56463 C0056486 C0056467 C0056468 CC056469 00058471 C0056472 C0056473 C0058474 C0056475 C0056476 00048148 C0048150 C0M8156 C0048157 C0048159 C0049473 Client ^ S am p lel^ ^ DL1-F01-MSF005-0-M1104 MC-F01-MSW005-0-041105 DMC-F01-MSF002-0-041105 DOU-FD1-MSW002-0-041216 DOU-F01-MSW003-0-M1218 DOU-F01-M8W001-0-041216 DOU-F01-MSW004-0-041216 DOU-FO1-MSW 0 0 5 -0 -M 1216 OOU-F01-MSFM5-6M1216 DOU-F01-MSF003-0M1216 DBC-F01-MSF005-1-M1216 DBC-F01-MSF005-0-041216 DOU-F01-MSF0024J-M1216 DOU-F01-MSF002-14)41218 DOU-F01-MSF004-0-M1218 DOU-F01-MSF001-04)41216 . DBC-F01-MSF003-0-M1216 DBC-F01-MSF004-(M)41216 DMC-F01-MSF001-0-M1216 DBC-F01-IPF001-04)41021 DBC-FD1-IPF003-04M1021. DOU-F014PF003-0-041021 DOU-F01-IPF004-0-M1021 DOU-F01-lPW0014H)41021 DL1-F01-MSF00443-0411M NR * Not reported du to quality control rasult failures. CS S u lta n a t* P f OS P w t1uT>oe4anM ui1t>rut Analyte Found (no/o> WatWaktot A ssessed Accuracy (+A %) 740 30 560 30 620 30 28200 60 4040 30 NR NR 5120 30 496 30 720 30 848 30 NR NR 8960 SO 640 30 892 30 1820 30 2070 30 4640 30 9280 50 840 30 384 30 293 30 359 30 424 30 387 30 348 30 Exygen Research Page 31 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 TabJe III. Summary of PFBS, PFHS and PFOS in Clam Samples Exvoen 10 Client C 4 Sulfonate PFBS PwfluorotniUilWMH onett Anatyta Assessed Found Accuracy (no's) wet wt. <+/-%> C S Sulfonate PFHS A ertw orohsaonesullbnett Analyte Assessed Found Accuracy In g /fll w et w t. <+/-%) C l Sulfonate PFOS A erfluoroocttiiesiiltonett Analyte Assessed Found Accuracy (nu/a) w et w t. C0056480 DC3-I01-CFW001-0-041216 NO 40 ND 40 1.82 50 C0056480 Rap DL3-I01-CFW001-0-041216* NO 40 NO 40 1.82 50 C0056481 DMC-101-CFW001-0-041216 NO 30 ND 30 6.18 50 C0056481 Rap DMC-I01-CFW001-0-041216* ND 30 ND 30 6.12 SO C00664S2 DC2-I01-CFW001-0-041216 C00S6482 Rap DL2-I01-CFW001-0041216* NO ND 30 ND 30 ND 30 1.81 50 30 1.77 50 C0056483 DBC-101-CFW001-0041215 NO 30 ND 30 4.48 30 C00S6483 Rep DBC-I01-CFW0010041215* NO 30 ND 30 4.44 30 C0056484 DL1-I01-CFW001-0041215 NO 30 NO 30 8.80 40 C0056484 Rap DL1-I01-CFW001-0-041215* NO 30 ND 30 8.84 40 C0056485 DOU-I01-CFW001-0041216 ND 30 0.202 30 11.4 SO COO56485 Rep DQU-I01-CFW001-0041216* NO 30 0.205 30 10.9 50 Laboratory Duplicate NO = Not detectad at or above Limit of Quantitation which Is 0.2 ngAj (wet weight). Exygen Research Page 32 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples Sampte D L3^014PF0014W >41021 < o o o a i* S f* C, 1 k *> DL3-F014PF001-0-041021 (CDOteUS Spkr, foopp) DL3-F01-IPF0G2-0-041021 (CC04I1Z7 5 * D. 10 ftett) DL3-F01-IPF0G2-O-O41021 tC O M IItripk O, tOOppfe) DMC-FD1-JPF001-0-041021 {C0O4J1M |pk R, 10 ppfc) DMC-FD1-IPF001-0-041Q21 (O 00 4#i2 saiLtiit0 pte > DMC-F01 *IPF002-0*041021 (C00tei2f spkC, lapa) D M C -F01-IP F 002-0-041021 OBBiam flk N, too Pfd| D M C -f01-IPW 001-0-041021 (GOOttl10 Ipk 0 ,1Cppk) D M C -F01-IPW TO 1-04M 1021 (coouisoapki. tpepat DMC-F01-IFW0G2-O-O41Q21 (CO4I131 * * 1 . 1 0 **) DM C-F01-1PW 002-0-041021 (Cocxam * * j. toa * * O M C-F01-P4SW 001-0-041021 {0004413ZS**C. " * * ) O M C-FO I-W SW 001-0-041021 (C tw tiia Ipk F. lOOppk tpaa) DW C-F01-M SW 002-0 0 4 1 0 2 1 (C0O441 ate 0, M M frlp a *| DMC-FO1-MSW 002-0-041021A {C00441 9(4 M.100ppfc Ip to ) DL2-F01-IPF001-0-041021 (C8044134 lp k F .1 t pe***> DL2-F01-1PF001-0-041021 {C0D48TM Spt K. 100 ppk tp to | DL2-F01-IPFOC2-0-041021 <00044134 apk 0.1* ppalia s ) DL2-F01-IPF002-0-041021 (C0044ta6|pkL lopppa Ip i ) Amount E0kad C4 Stagnate PFES Amt Found Amount In dam pfe Raoouarad R tcovary W M waM tf. C6 S u * PFHS_______________ 9 Sultanat PTQ8 A M Found Amount A M Found Anwum InS w npl R acow ad R u eow y m Samp R n M n d F4eovy 1*1 . (% ) 10 0.30 100 030 6.56 62.4 03 62 NO 0.20 83 5.16 HD 732 73 5.16 12.6 78.4 74 73 10 0.205 6.52 63 100 0506 896 6 NO 7.48 75 4.16 NO 798 80 * .w 10.3 81 60.0 77 10 0.330 4.46 42 100 0.330 804 66 NO .5.34 S2 7 4 0 NO 74.8 75 7.40 9.96 61.8 26 74 10 0712 7.68 70 100 0712 76.0 77 NO 9.60 96 NO 96.0 96 14.1 14.1 10 1.08 7.60 SS NO 704 70 7 0 0 100 1.08 640 84 NO 800 80 70D 10 0.866 100 0.960 0.76 604 56 0 NO 7.98 80 34.4 NO 7 4 3 76 34.4 10 0.21 fl 0.78 96 0.S24 9A4 89 100 0218 812 81 0.524 932 93 270 270 so ND* 300 NO* 10 0.890 too 0.680 452 432 5 BO 46.4 90 86 $1 46 ND* 520 104 930 ND* 402 96 330 NO 658 86 3 4 5 NO 50.8 99 3.49 20.0 104 71.0 131 44.0 101 270 375 470 324 8.36 50.4 156 90 81 07 106 . 73 49 53 10 0416 5.94 46 100 0.01 644 64 NO 6.40 84 4.72 NO 764 7 4.72 8.56 724 36 00 "Snp iai*du# --c--da da aptUng levai atgaMcanty; IN a fc ft. an pccw te raoowaiy valu# cernt bacalculatad * I.O fl of Mmpla vaadforwilracitoot M a a lO liO g . ND Not datoctad at w abov* UmR r f Quantfiation vrtticH 0 2 nplg {W *t waigNt). ND* Not datacted at o r abova UrNt o f Q uantltatton h 1.0 nota (Wat walght). NR No* report*} dua loquaMy conbot raatitfaVuraa. Ra-anV^> fw its w * hown in Tabla I. ola: tin c a ttils suonmary tabla tow e rowtdad m u lte , roeovary velu may w y cilgW iy fiow th valuta In fea nm date. Exygen Research Page 33 o f !3 2 Interim Report #10-Analysis of Fish and Clam Samples Exygcn Study No. : P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) S im ple O ooiptlm 012-F01-1PF003-0-041021 (cooats *p * c. ta m * * * ! IX 2-H H *IPF003-0*041021 (CWUjiM *p*H. 1D0pp*pfc*| OL2-F01-1PF0C4-0-041021 (cooaaijr auk d. topo* s ***) D U -F01-IPF004-0-041021 P * * ia* tfp t U 00 pp* *p **| D12-F01-1PF005-0-041021 (O IM iU *p E, iO pp* *p*e| OL2-F01-IPFC6-O-O4121 (C00441M * * J. J00 pvii Sp*) DL2-F01-1PW001-0*041021 (Coomi aptt p. toppt tpm DL2.F014PW 001004 1021 {C0O4tiat **K. it t gp Spfta) DC2-FC1-USF0010 0 4 1 0 2 1 (O04B140 9pk 0 .10pp* >(#) DL2-FC1-MSF001-0-041021 (C0044140 fc* U 100 pp Sp) DL2-F01-MSF002-CMM1021 (0004*141 flpk c. 10pp*. *m ) DL2*fO 1-MSF002-0-041Q21 (C0M14) SpfcH. iCOppb 5g*e) D L2-F 01-M 9F003-0-041021 (C0*4*i4d<tpfc D. 10pp* 3**> DL2-F01-MSF003-00410Q1 {C*4*14& 8pk I. 1*0 pp* Sp*; DL2-F01-M 3R XM -0-041021 (CQ044.41flp* |,1Dpp& Sp*) D L2-F01-M 3F0)4-0*O 41021 (0004*143 Jpn J. 100pphKp*e) OLt-1-4Pf001-0-041021 (0004*144lpt1 F.10pp* lp*p) OL1-FOI-IPF01-0*041021 (0004*144 Sp* K, 1*0 pp* SpM) DL3-F01 *IPF0C3-O-O41021 (CtM*l4S 8pt 0.10 pp* Sp*) DLJ-F01-IPF003-0-041021 (C0040145 Sp*k. 1*0 pp* Opta) Amount Spiked C4 S u lfc n t P*=BS Amt Found Amount In Sample Recovered Recovery (V O -*-- C8 SuHorartd PFHS Amt Found Amount in Sdfitple Recovered Recovery tartiw ptw t- i* > CB S u lfo n ji PFOS Amt Found in Semple "t- Amount Reoovered Recovery (A rt)** * (*i 10 0-247 100 3247 7.00 B 2 ft 66 63 NO 7.66 80 2.01 NO 73-2 73 2.01 030 74.0 72 72 10 0340 100 3.348 6.64 07.6 66 7 ND 7.60 76 4.52 NO 76.6 77 4.52 113 88 60.4 78 10 0-338 6.66 66 NO 7.66 77 2.46 9 8 8 72 100 0.338 68*4 SB NO 60 ft 1 2.46 120 60 10 3326 7.04 67 100 0326 66.0 06 ND 7.60 76 ND 772 77 10 0333 736 70 ND 8.20 82 100 0333 646 64 no 77.6 76 10 0.40ft 100 0406 6.M 61.6 66 81 NO 748 75 ND 66.0 88 10 0A16 6.62 61 100 0*41ft 64.6 64 NO 6.7ft 66 ND 70.8 71 10 0345 7.06 67 NO 7.56 76 100 0 .3 71.6 71 ND a u 61 10 0.432 7.44 70 0.261 8.16 76 TOO <M32 67-2 67 0JK1 78.4 78 10 Q>U8 120 6ft NO 8.06 61 TOO 0440 992 60 ND 616 82 5.86 5.88 772 773 64.6 04*6 41 it 41.6 83.6 83.5 334 33.4 3.55 166 13.9 65.2 750 15B 101 156 412 104 91.2 167 40.0 108.0 10.8 600 60 79 61 * 61 62 83 73 73 76 mnttu* m m i S x hing km' thmmi* i. m u * raesvuy vu* eanvn b . catcuMad NO - Not dnXCMd I or bow. Umll cf Qixndwtkxi htati d 0.2 ngtg (wet uxtgM). MB * Not raporttd duu to qufdty oonM M ill Wurad NxxnaMo nouM ora ohoom In TaU* II Noto: n o M o ouirmory toMo otioara raundod raouM. rawtooiy voluoo nxy vwy odgMy tram 4 ualuaa In Mo raoi do. Exygen Research Page 34 of 132 Interim Report #10-Analysis o f Fish and Clarp Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) Sample Oaanption Dt3-F01-IPF004-0*041G21 (oooMwa Sp* c. paate a *) DL3^01-IPF004-(M M 1021 (C0044140 SpfeH. 0 0a Sato) OL3-F01 -IFF006-0-04 1021 (O0044U7 * * D. 10 ppaapk) DL3-F01-IPF005-0-041021 (Coaui7 * f* I. 40pp Iptoa) D6OF01 -IRF001-0-041021 (Ooo*aia aK. roppa dp*) D0C-FO1-IPRX) 14MM 1021 (C ooutts tpa j , leoppt M v ) D6C-F01-IPFDQ2-0-041021 IC00HU9 le p\ n pa* to*) D6C-F01-IPFO2-0-O41021 (C0O441pk K, 100f Ipton) D0C-FO1 *JPFD03-0-041021 IC004*190spk a, io ppasptoai D0C-FO1^PFOO3^O41O21 (cao4ais t ap i. lao ppa pto) OBC-F01-IPW 001-0041021 0044W1 tpk c. iopea$a*a) O0C-FO1-IPWO01-0041021 <0004411tfk M.100ppbOptoa) DBC-F01-IPW 002-0041021 (Caoaaiaz ta D, 10retapa*) OBC-FW PW 002-0041021 (C004I1&3tpfc 1,100ppbOpto) DBOFOI-J^W 003-0-041021 (C0M41S3tf E . 10 ppbtpfc) D0C-FO1.P W 0 0 3 0 0 4 1 021 (cao4iu tpfc j. ioo pat Spa) OOU-F01-4PF001*0-041021 (00040184 SpfcF, 10ppbBp4to) DO ll*F014PF0010*041021 (00041154 1 * K. 100pp# fc*a ) D O U -F01-1P F 0 0 2 0 0 4 1 021 (000401MfphO. tOppkOpto) DOU-FO1-1 P F 0 0 2 0 0 4 1 021 icooai6pL ioopeapto) DOU-F01-IPF0030041021 (000401508fkC1BppbMi) DOU-F01-IP F 0030041021 (C0040150Sf*H. 100ppp5p*) Amount 8p*cad C4 8 u *o n a la P fB 6 A M Found Amount tn Sampla Recovered Recovery C6 g u j g g tt PFK6 A n t Found Amount in Sdmpu R*co*<*d Recovery 1%) C> SuOogatg PFQ8 Am t Found Amount In S a fflp f* Recovered Rcoury r*i 10 0.784 0.04 01 100 0.704 60.0 00 ND ! NO 1016 t 00.4 . 1 . 0382 . r. b v. 0.382 896 012 86 01 10 1.53 b M 36 NO 8.10 82 5.40 117 62 100 1.53 60.4 56 NO 932 93 6.46 9 6 0 01 10 1.34 100 1.34 10 1.43 100 1.43 034 49 0.002 100 101 954 50 0.602 068 06 5.04 44 NO 936 94 7 50 NO 000 00 MR NR 110 110 NR NR NR NR 113 109 79 10 0.800 0.72 57 0.944 10.4 80 . MR 100 0.996 0.6 60 0.644 068 60 ! NR NR NR NR NR 10 0.064 7.00 02 2.21 0.60 75 NR 100 0.064 70.0 66 2.21 008 86 . NR NR NR NR NR 10 0 700 10 0.700 7.50 596 "10 100 10 ' 100 0.730 ' 32 0.700 . 70-8 4.36 436 10.9 700 00 i .1 2 66 0.12 7 6.70 70 0.78 05 1.02 72 102 162 81 90.0 . 07 16.5 \ 97 .i 00.0 ` 04 NR NR NR NR 0.40 06.0 05 85 NR NR NR NR NR NR NR NR NR NR NR NR NR NR 10 0.76 13.1 63 0.792 0.72 09 ioo 6.76 70.0 69 0.782 r 6 0 0 78 NR NR NR NR NR NR 10 S.00 120 9 1.02 - 10.8 96 NR, NR NR 100 5.66 ! W . ; ` 75 y i.be 00.4 07 NR NR NR 4lamptojwaldiien o n * tw pIMntov*i*l<jnHcanby.ttorafor*.anaccuratereooweryvaluacannotbacalculated ND Not detectedatoratxveUnyt ofQuantitationwhichI0.2 iVO (w*t weight). NR * Not reported dun to quality oonfrol reeult failure*. Ra-anatyah raauttn ara town InTabla H. Nota: Sines thla Nummary tabla shews reunded results. reoevery values may wary lighay fram tha vatuaa In tha raw data. Exygen Research Page 35 o f 132 Interim Report #10-Ana!ysis of Fish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) am ple ^ ^ o c rlp tlo n DO U-F01-IPF0044-041031 tC00*HS7 3p* 0. te pet tfO el DOU-F01HPF0044-041021 (oooaeiar apn i, laoppa Spe*) DO U-F01-IPF0064441021 iCOMtIM te E. 10pte Sptea) D O U -F 01-IP F 0054441 21 (OOOW1 te J. 100 pte dp*<) DOU-FOMPWOOt-0-041021 ICOW flte te F. te Pte M te) DOU-F01-IPW0O1 -0-041021 (Cotunsa te k. 100te te> DO U-F01-1PW 0024-041021 IC00441OO% Q. 10pte s#ie) DOO-F01 -1PW 002-0-041C21 (ca o te i te i. 1 0*-) DU-F1-M6W 0014-041021 (CCCteW te U pi* ttew ] DL2-FC1-M8W0CU-0-041021 ic o o in a i puJ. loope&apte) 0U -F 01-I48W 0Q 24441021 to m a tiz te k. te ppa P* DL2-FC1-MSW 0C24441021 (OOOteitt te U ite Mte M * ) DL2-F014 4 8 WC034*041021 IC O * pk c, ppe ptea| DL2-F01-M 8W 0034-041021 (Cnoteim pfc o. 100ppe Sptea) DL2-F01-M8W 004-0-041021 < coo 4 a i0 4 ite t.te p te te i DU*F01 44SW 004-0-041021 <C**41te te F, 100Pte 9 te *l DL2-F01-M SW O O 64441021 (cowEi te a, 10pp%$pw) DL2F01*MSHY00S*O*O410(21 (COOtette te H, 100 Pte 5pft*{ DBC-F01-M8W001 -0441104 (COteMM api c, 10pte 3pe) OBC-F01-USWO01-0441104 ICOMMOOP* .100 ppl % *) Amount Spfced O S u fonot* PFBS A m tF ouid Amount in Sample Rk o w m Recovery l% \ C8 Sulfonate PFK$ Amt Round Amount 1 * 3mpl Recovered Recovery (nteteMrt wt. t% ) j'l 1 C * Sulfoneta PFOS Am t Found Amount in Sample Recovered Recovery W o llte n (% ) 10 0.12 13.2 71 2.30 12.0 7 100 312 76.4 70 2.30 06.0 64 NR NR NR NR NR NR 10 4.50 <00 4M 122 76 0.066 112 102 70.0 71 0.268 05.6 60 NR NR NR NR NR NR 10 2JO 10 2 09 102 73 0.048 .7 72.4 70 0.948 03.6 06 63 NR NR NR NR NR NR 10 10.4 100 10.4 10 0203 100 0.205 10 ND 100 ND 10 NO 100 ND 10 ND 100 NO 10 NO too NO 34.4 w* 0.32 704 0.06 00.4 924 612 7.5 2.0 7.06 00.8 180 76 01 77 7 86 82 01 76 02 80 81 3 69 3.80 ND ND NO ND NO ND ND NO ND NO 132 802 6.66 602 9.12 3.6 9.06 S2 728 3.6 0.06 100 5 86 87 65 91 94 91 fi 73 94 01 HO NR NR 120 120 102 102 03.4 S2A 66.0 ex> 166 188 NR NR NR NR 126 177 07 110 185 63 103 162 90 75.8 166 78 135 234 66 10 0.060 10.3 04 8.32 17.6 98 NR 100 0466 00.4 00 8.32 113 106 NR NR NR NR NR 'Sample rw W ui Owe--te <ho apiktog level tegnMuir%; Ctoafare. an kOCUSt*recovery value carat* bo calculated ND - Not detected i t or above UmH a t Q uantitation which te 0.2 n t fj (*a t tveight). NR * Not reported (tea to quaiHy control reaull telkirea. R a-ardyelj ru rta armtown In Table H, Notes tin e a thla tum m ary tabla te o w t rounded rotuHw, recoveryvtfite may yory IlfM ly front tha valuaa In tha raw date. Exygen Research Page 36 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS aud PFOS in Fish Samples (continued) tem pi D a a a n g tjo r^^ D0C-FO 1-MSW0G2-O-O41104 (COWMS1 Spt c. 1 pv M a i 0 SC-R31-SWO2-0-041104 (COMM91 SffeP. <00pp* to ) DBC-F01-MSWOtKWWH 1104 <ca04M&2 t o G. 10t o to } DBC-F01443W 0C3-0-041104 (000432 t o M. 100t o M to O 0C-FO 1-M SW 004-0-041104 cow ! t o 1 . t o to ) 08C-F01-M SW 004-0-041104 (C004M91 t o j. 1 t o to ) DBC-F01 -MSWC06-0-041104 (C0MMS4 t o K. > t o to ) DCOF01-M5W C06-0-041104 (C004MM to U N to to > O0C-FCVM3FOO1-0-041104 |CUMH to<4. 10to to ) D B C -F01-M SF001-0-041104 (C004NM t o N, 100t o to > o e c ^ o 1-M 8F002-0041104 (C toM M toO . i to to ) D O C -F01-U 3F002-0-041104 (COD444Mt o P. WOt o to ) D 6C -F 01-rF 004-0041104 (CdQtosr t o c. w t o to ) D ec-F01-pf004-0-041104 (C0040457 t o 0. t o to ) D6C-F01-IPF00W H)41104 (CtoM t o i. Wt o to > DBC-FOMPF005-0-041104 icao4M t o f. ioo t o to ) DOC~FOVPW04-O-O41104 (C ffM M I t o 0 ,19t o t o ) DBC-F01-IPW004-0041104 (C to lto t o K 100to M ! DSC-F01 IPW006-0-041104 (C0B4WMt o I. Wt o to ) OC-F01-IPW0054K041104 icootatdo t o J. 1 t o to ) Amount 8*>kad M il C4 Sulfonate P fB $ Amt Found Amount in tem pi Ita o o w id Recovery ( * !> -- <%> 6 S u lto o to PFKS Amt Found Amount m Sample Recovered Recovery W l) -- <- PH C l 9dfonto PFOS Amt Found In tem p i Amount Recovered 4 V IIW W noo*ry {%) 10 0.900 8.00 91 343 126 84 NR 100 0469 04 6 84 3 4 3 113 110 NR NR NR NR NR 10 0 080 B SC 91 3 6 2 136 100 NR 100 0.900 660 85 362 104 100 NR NR NR NR NR 10 0.064 100 0.064 9 . 872 81 86 9.72 19.5 96 9.72 124 114 NR NR NR NR NR NR 10 0.902 8.20 74 2.77 102 74 NR 100 0.902 81.6 91 2.77 106 106 NR NR NR NR NR 10 0.820 8.60 89 3.56 13.7 101 NR ice100 0.920 8 -4 80 356 106 NR NR NR NR NR 10 0.769 100 0.766 10 0.504 100 0.504 8.82 90.8 10.3 84 j6 82 194 80 1.94 100 0.594 84 0.594 125 102 11.8 106 106 100 110 106 NR NR 142 142 NR NR NR NR 152 254 112 10 1J0O 104 99 2 1 5 12.2 101 NR 100 160 908 68 2.15 105 103 NR 10 0.404 9.72 82 320 13D 97 NR 100 0.484 87 7 328 104 I0 f HR NR NR NR NR NR NR NR NR 10 0.572 104 96 2.10 132 111 NR 100 CJ572 920 82 2.10 102 100 NR NR NR NR NR `te m p * tm kim --c--da lh> apiking yd i tyai c r i) ; * * , an accurate racovwy valua cannot fcacafcwiaftad NO - Nol d e te c ts X or oov* lim i of Q u a n lto o r *h ic h 0.2 ng'g (* l m it). NR Not reported due to quality oorrtro* ra a iit Murea. Raanatyali muMa a rt #hCMi In T M t I. Not: Sine thte summary M l ah*wa raundad pmvHs, f oovay touaa may vary lightly hm 1h valuaa ta t o raw data. Exygen Research Page 37 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: POOO1131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) Sample DOU-F01 -IPW0Q3-O-O41104 |C 0 0 4 * Bptt K. lO p p t t p * ) DO U-F01-IPW 0O KM V41104 (C0044401 SpfcL, lOOpoaSpAa) DOU-F01-IPW 004-0-041104 (cooeetiz aiefcc. toppaseeai DOU-F01 IPW004--041104 (C04MU ipa a t t ppe M w ) DOU-F01-IPW 005-CHK1104 tCoomma le a 1 10 pc* M * ) 0 0 0 -F 0 1 -IPW05-0-O41104 <000494*3 Spa F. 100 PC* !? * ) DL2-F01-IPW 002-0-041104 <00040444 apaG. (Oppa^pHe) DL2-F01 -IPW 002-0-041104 (00540444 Ip k H. 104 p f* 8p*a) O U <F01-M8W001 <0-041104 (OO44405 Spa i, to m t Seta) DL1-F01-M S W 001-0-041104 10004*44 ipfc J. 1 DC* ape#} D U -F01-M6W 002-0-041104 0004014 i * k K to ppbp4a| DL1-F01-MSW 0024M H1104 <cao4S4M Oak l ioo ppa spwi D L t -F01-MSW 00SO-041104 <00044447 Spfc G, 10 R * Ip * ) D L1-F01-M SW 003-0- 41104 (C00444S7 S |* . 100 POB < *) D U -f=01 -M 8W 004-0-041104 (0044444k BpfcE. I0 ppa <jp4} DL1-F01-MSW 004-0-041104 (C004S4# Opk F, 100 pp 44) D U -=01-M8W00-0-O41104 (C440444SpfcG. W ppfe Sp**| DL1-F 0 1 -M S W 0 0 0 -0 -0 4 1 104 icoo4M5i pa h, ioo pa spa*} DL1-F01-MSF001-0-041104 (00044470 9pk t ttp p tip k a ) O U-F01-M $F001-0-041104 (C0040470 tpa j, ioo ppa *#--) Amount S p ik e d in o /a ) 4 SiJfmato PFB8 Am i Found Amount in Sample Recovered Recovery infra) wet<*. C t Sulfonele PFHS Amt Found Amount in Semple Raoouww) Rcovry (nfrg)ert 8 Sulfonate PFQ6 Am t Found in Semple (flfrfl)v e ta t Amount R *cov<r*d ( H I * R a covxry 10 3.12 100 3.12 13.1 88.0 WO 65 6.00 6.00 16.3 103 107 101 NR NR NR NR NR NR 10 349 12.6 93 7.12 17. 103 NR 100 349 664 85 7.12 108 101 NR NR NR NR NR 10 2.97 100 2.97 10.5 93.6 75 61 5.12 13.5 64 b.12 109 104 NR NR NR NR NR NR 10 1.04 100 1.04 9.46 80.0 84 78 ND 104 104 820 NO 9 7 j6 9 8.20 14.3 1 124 116 10 1.20 10. 0.88 1 1 3 104 100 1.20 66.6 88 0.666 112 111 NR NR NR NR NR NR 10 1.30 9.7 65 1.72 12.4 107 NR 100 1JO 80. 79 1.72 100 96 NR NR NR NR NR 10 0 ,5 6 0 100 0 .5 6 0 6.92 65.2 64 65 1.13 10.2 91 1.13 9 0 4 96 10 0.900 8.52 7 0.906 9.76 69 100 0.906 Mi 84 0.908 99 96 10 1.18 6.60 7 1.16 104 92 100 1.18 82. 81 1.18 90S 9 10 ND 100 HD 7.7 76 0.315 9.52 92 76.4 78 0.31 S 90A 90 NR NR 161 18! NR NR 293 293 NR NR NR NR 201 286 108 NR NR NR NR 303 394 101 `Sampla raakta* sxoMdt #io apiklng level e^ lcarrty ; Bienefam. an ac o n to moovar/ vah* cannot becetolewd ND Nat detected at or above UmH of Quantitation hlch it 0.2 npfi (wi weight). NR Not rmparknf due to quality control naadt faluraa. M^enatywa reaufti am aiUMn In TaUa II. Note: since ttrii summary tibia ahowe rounded reealta, recovery values may vary eagMy fromtha vataaa In tea raw data. Exygen Research Page 38 o f!3 2 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) S vn p to OnecripUon DL1-F01-MSF002-0-041104 K. tO w * ** ! DL1-F01-MSF0024>-041104 (CM4Mn u io p p M l D L1-F01-M 8F 003-0-041104 *049473 9fk C. 1 * 6 M ) D U -FO I-MSF003-04)41104 (COOAM72 Apt D, 100 pp ftp* D L1-F01-M 3F004 -0 0 4 1 1 0 4 (C04M7t 89k . TOPPM l CX1-F01-M 3F004-0-041104 X04M73 M F. Ittp p M ) 0L1-F0 VM SF005-0-041104 (CWTO4T M a . Topp M ) DL1-F01-M 8F005-0041104 (00049474 3pk H. IDCppfi Ip ft DL1-F014PW 001-00411 04 (00048479 Ipk 1.14 PP3pM) DL1-F01WPW001-0-041104 (00048479 M Js WOpp Spft D U -FO U PW 002-0-041104 (C004847P tp X 10 pp9pfc DL1-F014PW 002-0041104 (00049478 M L, 100pp tpto) DL1-FC14PW 0034KJ4110 *049477 M C , 10PPP*; DL1-F01-PW 003-0041104 (C004W77 Ipk D. 100pp tp i ) DL1-FO1-IPW 004-0-041104 (C8049479ipfc C, 10 PP8p* DL1-FO1-IPW 004-0-041104 (CB04M7I tpF 100PPM > DL1-PD14PF002-0-041104 (00048479 m 0.10 pp Opta) D U -F01-IPF002-0-041104 (C0048478 M H. 180 ppM *> DL-1 -FO I -IPF003-4341104 {0004*4*0 M U * * * * GL-1-F014PF003-0-041104 <00849490M J. 100pp $*) A nou n i apilM d C4 S urent PfBS Am t Found Anrxxnt M 3*np Rcov9rd Rccvry CSultonalaPFH3 Amt Found Amount fei Sample n scovsrtd Rsooirary CW S iralrt. (X ) . C 8Jfenala PFQ3 Amt Found m Sampfc Amount Rsoovsrsd Rcovry V I)-->- (% ) 10 ND wo ND 0.60 88 0.024 10.5 99 M S 05 0.824 9 8 4 96 NR NR NR NR NR NR 10 0-200 0.70 88 0,652 10.2 95 100 0.203 04.4 84 0.862 96.2 96 NR NR NR NR NR NR 10 ND 100 ND 8.88 88 0.532 10.4 s e s 87 0.532 101 100 NR N NR NR NR NR 10 NO 100 ND 0.04 86 0.6M 10.1 94 824 82 0.684 8 4 0 83 nr NR NR NR NR NR 10 0.772 8.64 78 0-556 8.72 se 17-4 204 90 100 0.772 MO S3 0.556 978 97 17.4 113 90 10 0.438 Ut 2 0.538 8.96 94 18.7 2 7 4 117 100 0.468 78,4 76 0.536 88 0 . 87 15.7 100 04 10 ND 100 ND 10 NO 100 ND 10 ND 100 ND 10 ND 10O ND aoe 81 0.209 9.44 92 9.38 80.4 90 0.209 106 106 9.38 848 85 0.238 o.aa 88 71.8 72 0.2) 8 4 8 85 58.8 59.6 8.00 80 est 88 0-222 9.28 91 0.222 984 es 11.1 .i 8.80 M 0.285 10JB 103 20.9 78.8 78 0.2$ 804 so 20.9 18.7 109 72j0 140 109 108 309 109 n 100 80 7S 97 96 M aimdaiauairt rasas na panaimwtunicssy.aiarafcra.mcassiaracoranvaluscarmeSsratouisK NO - Not Uatsdsdat oratraca UmKd QuKTtltadcnwtdchk 0.2ng (watwalgtt). NR * No aportad dua lo quality consul raault la tu a a . RwanaEyas rasuSa ara taran lu Tabla ll. N on: Sisea INIs summary N bls straws rauauaS raauSa, rscovsty vsKras may ra y B ll^ rty tram EN. vaiuaa Is Sw raw data. Exygen Research Page 39 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 T able IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) S n p te D **ciipbon AiTioont Spiked C4 S u lfora t* PrQ 6 Amt Found Amount id Stampfe Raoovored Recovery O vS )** . C6 8uttonte F^H 8 Amt Found Amount in Sample /' covered Recovery |iy i) w e t*. (n&<e)cwt (%> C8 S o ifo n ie PfO S Am t Found In Sample {**> **. Amount R n o n re d I'M ) . R occvoiy (% ) D C -1-f0 1 -lP f0 04-0-04 1104 (C M <1f*K .10hj*e) 10 HO Dt*1 -F01-IPF004-0-041104 |C0044t1 %okL, V pp* tea) 100 NO 0.00 00 M l 65 ND 0.46 06 7.60 ND 98. 90 7.90 15.0 82 102 04 DL-1-FO1-IPF00<S-O-O41104 (COWOMJ M l C, Pppb Sp*e) 10 NO DL-1 *F01-PRJOSO-Odl104 (coo4etti7 pa o. 1JPpee tea) 100 ND 7.44 74 0.373 0.00 80 54.6 05 0.373 004 00 0.72 0.72 15.9 72 102 03 OL3W -M 0W O O1 -0-041105 (C004teC) ipk &. 10ppe tea) DLS-FOI MSW001-0-041105 (Cao4Mi Dpa9. tec POOtea) , 10 100 NO NO 6.3# 64 056 00 ND 920 92 138 ND 09.6 100 139 106 258 119 DL3-P01 -M8W002-<M)41105 (C00404I4 m * 3, Upptapfcei D L3-F01-M S W 002-0-041105 (CtXMMte SpkH, 100KiO te te ) 10 100 NO KO 6.44 04 004 00 NO 0.02 00 100 NO 106 106 100 150 . 283 103 D W 'O l-M S W O O ^O ^dl105 tcoteaeao M i. U pa t M e ) D L3-f01-M8W 003-(H>41105 (CWMaras ip * j. uppo te*) 10 100 ND HO 6.00 604 00 NO 0.56 00 146 ND 102 102 146 154 254 100 0134=101 -M 8W 004-0-041105 (C004e4M * * K. 10 Pte te*) DL3F01 -M3W 0O4-0-041105 <0M 4 tpkL. tOOpod teM ) 10 100 NO NO 8.49 06 0.286 9 .n 94 604 00 0256 102 102 100 19 225 301 103 DL34=01 .MSW 00SO4H1105 (coeeewj a * c. io p p t tea} DL3-FO1-M8W0C6-C-O411C6 fcaoitear spa o, no p?e tea) 10 100 NO ND O.S0 00 0.306 11.1 107 100 100 0.366 105 105 173 173 103 . 246 73 0L3-F01-MSF001-O4M110S |C0d4Qitee . pae tea) 10 ND 0.46 05 NO 114 114 110 D L 3-F 01-M 8F 001-0041105 (comMae M '.ia o p e e te te ) 100 NO 04.6 06 NO 109 109 110 D i-> fo 1-M ar002-0-041105 (ooMteao ip k a. 10 pea bptei 10 ND 9.68 00 NO 104 104 116 DL3*F01-M6F0Q2 -0-041105 (comm tea h. too peb tea) 100 NO 60.6 so ND 103 f03 116 130 234 118 989 * 21 100 DL3-F01-MSF003-0-04110B (CaMMOO SpaU 3 ppb te te ) 10 NO DL3-F01MBF0Q3-<H>41105 (COMMpe M J. n o ppete te ) 100 NO 0.32 03 90.0 01 ND 11.0 110 75.0 ND 107 107 79.0 02 0 190 114 "tem ple reeidue m cwd* f a apiUng aiw^nihr; ttw vfor m eccuvte leoovi?/ lue cannt be calculatad HO - Not delectad at or above Lim it of Q uantitation w tiirt te U 2 o( / q we* weight). NR > Mol reportad dua to quality control re a ilt teluro*. Ra-anNyaia reuutu ara ahutvn InTabla It. Note: no (tila lu m iH iy labte show rounded taeulta. recovery values neyv ;.y slifakdy hem die value In die raw darn. Exygen Research Page 40 of 132 Interim Report #10-Analysis ofFish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) a m p i* O M C rgjon D L3-F 01-M 3F 004-0-041105 fCOMMM ** K, 10 OS*90S*) D L3-F01-M S F004-0-041105 (000*<t1 %hL, 100p*****1*! D L*FO M *3F006-0441105 (0004*4*2 Spft C. 10p **lp*a} DL3-F01-MSF006-0-041106 (OD*4S4tf IffcO , 1*0pp* 1*4} DMC-F01 - IPWOO0-O-O41106 [0004*4*9 9 f* t , 1* pp* l*S) DMC-F01-IPW 005-0-041108 (0004*4*3 Ip * P. 100PS*!* ) DMC-F01-IPWD04-0-041105 fC004*4*4 Ipk Q, 10 ps* ***) DM C -F01-IPW 004-0-041106 (004*4*4 IpkH, 110*** 9**) DMC-F01-IPW 005-0-041106 (C004S4QSa** U * pp* tpfes) DM C-Fd-IPW 005-0-041*>08 (0004*4*5 ip k J, 100ps* #*) DUC-F0144SW0C3-O-O41105 [C004S4M Ipk K. W P** Ip*) DMC-F01 -M3WDQ3*0-O41106 fCSM*4M Ipk U K pp* M M DMC-F01-M 3W 004-0-041106 (C004*4*7 Ip * C, 10 PS* Ip*) DMC-^Ot-M $W 004--041105 (0004*4*7 Ip * 0, 100b b t! 0M C-F01-M SW 006-0041108 [C0D4S4B*Ip * E, 10ppCSpiai) DMC-FO1-M 3W 005-0-041106 (C904MM s i* F. icopp* Iptts} DMC-P01-44SFT2-0-041106 [0004*4*0 Ipk C, 10 pp* tp *s | DMC-FO 1-M3F002-0-041106 (0004*4*0 |p * H. 1*0ppfclp*s) DMC-FC1-MSF003-0-041106 (0Q04M00 * * 1.1* |**S p*S ) DMC-FO 1-M6F003-0-04116 (COOMsaeip k J, too pp* Spt*) Arnesi Amt Found Amount Amt Found Amount A m i Found M o d In Ssm pt* R acow ad ftacovary in Sample R*cov*m d Racovwy In Samp i* (HM )M IM . - (S I Moti) a u t. (fl0f}W twt (% ) (ng/|)wst wt Amount Raoouarad I'W I - ' Rscovwry <%> to ND 100 NO 10 NO 100 ND 10 ND 100 ND 10 0.000 100 0.300 8.60 904 9.92 o&e 6.36 044 8.04 904 86 NO 11j0 110 130 so NO 104 104 130 08 ND 10.3 to o 01.8 66 ND 06.4 96 81.6 63 0.366 , 9.52 64 .0.386 8 8 6 81 . 88 612 81-2 77 0.3S6 . 8 J 9 , U J 80 0.386 104 104 66.6 144 236 97.2 161 768 150 107 166 106 88 86 80 10 NO 100 ND 0.86 so 0.270 66.4 66 0 ^7 0 10 NO KX) NO 8i64 86 0.582 824 82 0 502 10 0.310 100 0.316 9iS2 82 >4 82 62 1.80 1.90 10.1 96 105 105 11.0 104 106 106 12.1 102 101 ss . 64.4 64.4 354 364 258 290 75.6 153 9 389 . 484 110 258 313 94 10 ND 100 ND 3.16 92 0.S80 11.6 108 83.6 94 0.900 118 117 NR NR NR NR NR NR 1 NO 6.36 64 NO 104 104 NR 100 NO 0 7 .fi 86 NO 104 104 NR NR NR NR NR 10 NO 8.20 62 ND 10.4 104 68.8 100 NO ; 8 7 J 67 NO 110 110 66 J 108 184 137 *3wnpJJi*Jdu* M caadi 9m CpMop tam l ctpnOcantljc 4i*f*fem , *o accusalo vseowy vain* cannai be oicUM #d NO * Not d *t*c t*d at or abov* Lim it of Q uantitation M ch la 0.2 ngty (wat weight). NR Not nportsd duo to quslty oorrtrol rwuK W u m . r**Jts w * i Iu m In TabisJI. N ot*: Sino th l* summery is *** shows ourtdsd rssoHs, mmwsnf v fc j-- mayvsry tllg h tiy from ttw vstess In tho row 4 M . Exygen Research Page 41 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) tam o ia DMC-F01-MSF004-C-041105 (COD4WOI Cot K. 10 9pft X*4] OMC-F01 -M S F004-0-041106 (CM4IM1 CpfcL, lOOpptapa*) DWC-F01-MSPOO5-0-041106 (0004060 V C. 10 ppt M ) D M O F01-M S F005-0-041106 fuxuaaca Spt o. too a t* h m OMC-F01-JPF0O3-OO41106 |CW *|BQ3tpfcC.14pptapa) OMC-FO 1HPF003^W341106 (Ocotoo Spkr ?ao*ap*> DMG-FQ1-IPF004-0-041105 {COO*1804Spt G. 10 ppb 9pN) D M C -FO M PF004-04M 1105 (COOM8C4&|*H. lOOpp lp i | DMC-F01-IPF005-W 41105 (C00*0*0t8pfc1. tOppaSptt) DMC-FU HPFOfrO-04 105 (COWtO tpk J. 100C4W N ) D L2-F 01-M 8F 006-0-41216 (Coos030* M l. llw U p * * ) DL2-F01-IPW0C5-O-O41215 (C00MM4 it * F 100 f f t apa) DL2-F01-IPW0G6-OO41215 <CM44* tit C. t t fib <**> DL2-FOHPW 03-0-O 41215 (COOMPMOakD, 10Op9*M ) OOU-F01-MSW0Q2-0-O41216 <00064460 apt 1. toppa ttp*> DOU F0144SW 002-0-041216 (C00M464 tpk J, 1tO pa Op*) OL2-F01-IPW0G4-O-O41218 (CoavMsr apt c. io ppa %ia) O U *F01-IPVY0C4-O-O41215 (C00M4S7V * D, (00ppa SpN) DO U-F01-M SW 003-0-041216 {OoooMja apt k, ta ppasp*) DO U-F0144SW 003-0-04121 <00094456 d p t L. 100 pu t Sp) Amount M ud C4 ulfanela P fBS A n t Found Amount w Sam oa Recovered Recovery -- J C ulltanale PFHS Amt Found Am oitol to anpia Recovered Recovery (n rtiM ftw i. (%) C t ^P n a te PFO fi Am t Found in Sample ( * ! * 4 Amount Recovered Recovery |nOff)atwt. f% l 10 ND 100 NO 10 ND 100 ND 8.76 88 0.254 11.1 108 080 66 0.254 107 107 0.52 86 0.420 122 118 036 64 0.420 110 110 11 118 148 14 123 228 110 193 280 114 10 ND 7.82 78 NO 104 108 7.02 100 ND 820 82 ND 106 106 7.92 164 61 101 93 10 ND 7.92 70 ND 10.7 107 440 562 100 ND 832 S3 NO 107 107 44.0 134 90 K ND 100 NO 7.66 70 75.6 7 ND 10.3 103 4 8 4 NO 064 M 4vt 54.4 121 75 10 NO 100 NO 10 WO 100 ND 10 0.206 100 0.20 10 ND 100 ND .44 M .2 8.00 644 6.02 72 6.00 784 04 83 80 87 07 61 77 ND 10j0 100 402 ND 102 102 4 9 2 ND 115 11 lii NO 107 107 132 1J2 104 02 1.72 02jB 01 NR NR NO 10.4 104 .0 MO 96.2 95 6 60 592 142 22.7 121 NR NR 14.6 972 100 93 96 106 NR NR 80 to 10 ND 8.12 61 10.4 194 00 100 ND 084 80 10.4 10S 95 NR MR NR NR NR NR 'Sample raaldiia u m m Di apiklna laval n.fcarev. la n ln an tc a t i tt t ruxwaty valut carY'-.K M cSCHMad NO Not detected at or above Urn* of QuanWagon which a 0.1 n0g (iv rt winght). nr a Not resorted due to quality oontrol raaiit fwiuree. Re-enai/els neuRo ara W teo In Tama 1. Nota: M ica o u t evmmery tabla abowa rounded re a m . racevury valuat m ey. / a lo ft* Irani * * valiaaa in the taav data Exygen Research Page 42 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) Semple D L3-F014 PW 004-0-041130 (Commop* !, tee apaet DL3-F01 -IPW 004-0-0411 (C0QM4## t< * F, 1 ppa $pia) DOll-FO 1-MSWO01-0041216 tCMM* . o. *e M D O O -F01-M 6 WOO1-0 0 4 1 2 1 6 fcaoM4oo aaa h. leo ape si) DOU-F01*M5W0O4-{H)4121 6 ico pem i ana c. ie ppe #-- ) D O U-F01-M SW 004-0-041211 (0008#4#f tv# f. ieo ape *) DOOFT)1-MSW0SOO41210 6Mi(coo te r 6. to pp#tp tw i DOU-FOI-M6W00-OO4121 0 (COOO#42 Span, 100ppetpta) OOU-P01-|ytSF006-0-041210 <00000407 60* C* 10 ppetpMa) o,O O U -F01-M 8F006-0-041218 (C#oa#40) ip * teo ppe tpfta> OL3-F01-IPW002-CWM1130 <co*ea4 ope e. toppe own) DL3.F014PW 002-O41130 (CQOSP404 SpaF. 100 ppe tpiaa) DI3-FO UPW 003-0-041130 (cewspoo t t * 6, io ppe DU^F01-lPW 0W -OO41130 ICMM4PC apt h , too ppe v * * ) D O U -F01-M 8F003-0-041216 (00000400 tpa K. 10ppe Bpto) DOU-F01 -MSFOOWMW1216 (COQ50400Spa l iio mo tpAce) DDC-F01*MSFDO*VOl21d (C000407epk C, 10 ppeIp ta ) D B C -F01-W SF006-1-04121 (C0QU4018pk 0,100 ppb Bpfta) DBC -F01-M SF005-0-041216 <000004009pk X. 10ppe tp ta ) DEC-F01-MSF006-C-041216 (C0000400pa l. 100 ppeOMO> Amount Splkad C4 S o *o re f P fB S Amt Found Amount in Sample Recovered Recovery ( 1 wetft. C< SudoneTe PFH8 Amt Found Amount In Sample Recovered Recovery {"&*'* * (% ) C8 Sullbnele PFQ8 Am t Found A m o rt in Sample Recovered R ecoM ty 1%) 10 HO 7.52 75 HO 11.0 110 7 *2 17.3 94 7.92100 NO 664 66 NO 118 118 114 106 10 7.08 100 7.08 14.6 632 75 76 1 *4 2 7 3 121 1 *4 122.0 107 NR NR NR NR NR NR 10 4.72 106 61 21.7 20.3 76 100 4.72 724 66 217 104 82 NR NR NR NR NR NR 10 0364 7.08 67 1.34 8.62 72 100 0.364 72.0 72 1.04 67.6 66 NR NR NR NR NR NR 10 1.41 6.60 72 8.40 174 #2 100 1.41 72.0 71 0.40 944 86 NR NR NR NR NR NR 10 NO 7.02 7 NO 9.04 90 12.0 100 ND 612 61 NO 1KJ 110 12.0 166 46 106 93 10 NO 100 ND 7.06 60 64 A 4 NO 6.86 00 6.44 NO 102 102 6.44 12.3 92.0 69 67 10 1.72 00 73 6.56 14*3 77 NR 100 1.72 BO.6 66 8 . 107 100 NR 10 0220 6.36 61 to o 0.220 63.6 63 1.78 11.6 .9 7 1.7# 966 96 NR NR NR NR NR HR NR NR HR NR 10 NO w o HD 826 84 00.6 61 1.68 114 97 1.66 99.6 88 NR NR NR NR NR NR *3#mpe rendue MCWdt *># apdng Javl piflwWcarafr'; anaeoaale racrwery -atu canne* 0# celcued ND Not dotoctod at or aU>% Umtt of O uantitoiion Mhlch ta 0 2 n ^ g boat weight). NR * N o t reported due 0 q u a lty oonfroi re#uN f lv r . Ra-anaJya leaufte ereehO M i tn T#bl ||, Note: Since Oils summary ttb la stKW reundad leaulte, recovery valuae may vary tig h tly tram the value# In dm raw data. Exygen Research Page 43 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) Semole DOU-F01-MSF02-0-041218 (0090440 if * 0.10 api ip*a> DCMJ-F01-U S f002-0-041216 (coosataa Ba* m. iaoppt M ) DL3-P01-IPW 001-0-041130 (cooaat7otp * c 1 aabaaaa| DL3-F014PW 0014W M 1130 <cot4ro * o. tooaaatpM ) DOU-F01 -MSFOOO-1-041210 (C ooiatrt ipk t. toppa tpaw) OU-F01-MSFOM-041218 (0094471 8pk J. 100 a p i M j D O U -F O 1-M 3F004-0-041215 (COoaatTZ % * K. 1ppM pto) O O U -F 0 144SF004-04M 1219 ICMM47X 8pk L, 100 ppt tpfca) DO U-F01443f001-0041218 <oooa4nipkK. tappa api) D O U -F01-W SF001-0-041216 (C006447 a # F. 100 * 0 M } D 6C -F01-M 8F003-0-041216 (CO06A474Qpk a 1 ppbM u) D9C-F01-W3FO3-0-041216 (CCO&4474 Spk N, tOt ppb D B C -F01-M SF004-0041216 (oooaoaTi s p il toppa M w ) OBC-F01-MSF004-0-041216 IOOOH479 Ip t i. to t ppM p**) DMC-F01 M8F001-0-041218 ICOOBM <* C, '0 **% ) 0MC-FD1 -MSFO01-0-041218 (C00M478 9pk D. 1P0 ppa $pw) D L1-fO i-IP W 00M -041218 (Coaa#*77 ip t i. io ppa *p*a> DL1-F014P W 0Q & 4V O 41218 (C005W77 Spi J, tooppaspika) O L2-F014PW 003-0-W 120* (C00444T i ip k K, 10ppMpaa) OL2-F014PWOO3-0-041201 10004447 % * L, to p A * * * ) Amount Spiked (na'Q) C4 8ulloneU PF88 Am i FounO A m o rt In Sample Recovered Recovery 1%) 06 8ulfonel PFHS Am i Found Amount in S am oli Recovered Recovery (W fl)W ptW . (aM luatw i. -- I V ..... C S utfom l PFOS Amt fo u n d In S im ple (notti etL Am ount Recovered Recovery (non)eat4.... [%> 10 0.886 8.24 84 4 5 8 14.5 99 100 0088 816 83 4.56 104 99 NR NR NR NR NR NR 10 NO 100 NO 7.80 76 0246 9.6 96 82.4 62 0.248 101 101 9.04 9.04 172 62 102 93 10 0.640 920 84 448 14.5 98 100 0.640 2.0 81 4 j6 8 103 98 NR NR NR NR NR NR 10 0.702 7.78 70 2 3 5 M O 85 100 0.792 812 BG 2.36 66.6 63 NR NR NR NR NR NR 10 2.16 11.1 69 100 2.11 828 61 10.9 2 0 10.9 104 99 93 NR NR NR M l NR NR 10 NO 8.46 86 2.09 11.9 96 NR 100 NO 4.0 84 200 108 104 NR NR NR NR NR 10 NO 7.72 77 3 *7 12 90 100 NO 75.6 78 387 98.6 95 NR NR NR NR NR NR 10 NO 9.06 91 0.420 9.08 67 NR * ND 88.0 88 0.420 102 102 NR 10 0.253 100 0253 7,40 72>4 71 72 0.044 0.644 9.99 66.8 93 68 68.6 68.6 NR NR NR NR 83.2 145 76 10 ND 100 ND 7.04 71 jB 70 72 NO 9.04 90 7.4 NO 92.0 92 7.48 13.7 62 84.0 77 `S iw p tt fe a ifra me--fti H fM rtq tovol ignillcanty: tw a fo n , an accurata xpoovgry v * lt* mn d bo catculaiad ND * Not detected a i or sbovo Lim it o f Q uantitation which It 0.2 ng*o (wet weight). N R -N ot reported due to quality control rpaulttaluiee. R-ewly#*a alita w e eto *n ir TableIt. Nota: Sino# thla summary tabta ahowe rounded reeulte, rscevary viuaa mayvary lightly from the vafuoa In the raw Exygen Research Page 44 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: POOO1131 Table IV. Matrix Spike Recovery PFBS, PFHS and PFOS in Fish Samples (continued) 9 u n p i DL3-F01^PW 0O M M M 1201 lceoso47v S p 1, loppe Spa) DO-P01-IPW 00&-0-041201 (C *0B 4 7 * J.1 0 .p k*N ta ( Amount Spiked (f'tfo ) C4 S u lta n * PFBS______________ C6 3 u * c fH PFHS_______________ C8 S u lto m ltF F O S Amt Found Amount Am i Found Amount Am t Found In Sample Recovered Recovery in Semple Bocovom tf Rocmmry In Semple (rv^Jwatwt (nflifl)vatw l f%> <%) fns>g)wa( wt Amount Recovered Recovery r*i 10 NO 100 NO 8.18 82 85 NO 888 80 4.88 NO *7 2 7 4.88 14.0 94,4 91 90 A rw ig ii Standard Deviation: Tf 1J Averse; Standard Deviation: 84 12 ' Sample rM K k* Moeadc * piking (aval alpnlfleantty; th>arrfara, an accurst raooery value cannot ba calaJatad ND a Not detected at or above Lim it o l Q uantitation wtocn tt 02 *gfg wotohl). MR - Not reported due to quality control reault feduroa. Re-ana*da raautta are atiawn In T *4e II. No.- 8lo #! MMwnary labia allow s reunited reaufte, recevacy values may vary Hpftdy from the vatu la Via raw data. Averpa: Standard Deviation: 88 20 Exygen Research Page 45 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Table V. Matrix Spike Recovery of PFOS in Re-extracted Fish Samples S m p o _ D G flC 1 ti0 2 ^ _ D B C -F O H P W 00 1-0 -04 1Q 21 (00048131 SpkC, 1000 ppb) D 0C -FO H P W O O 2-O -O 41O 2I (c w * e i2 8 (* o. w oo ppb} D B C -F 0 1 ^P W 0 0 3 -0 0 4 1 Q 2 1 (C004163 Spk 6,1000 ppb) D O U -FO M P F001-<M >41021 (Co o u im $pk p. iooo ppt>) D O U -F 0 H P F 0 0 2 -0 -0 4 1 0 2 1 (C00*8155 8pkG , IOOOppb) O O U -FO I-IP F0O W M M 1O 21 (000*8150 Spk K 1M0 ppb) D O U -F 0 1 -IP W 0 0 2 -0 -0 4 4021 (000*8190 S p kl. IOOOppb) D B C -F01-M S W 001-<W 541104 (C0048430 Sfc* J, IQjOOOpp6| D S C -F 01-M S W 002-0-041104 (00040461 Spk K, 10,000 ppb) D 9 G -F 01-M S W 003-0-041104 (C0040462 Spk L, 10,000 ppb) D B C -F 0 1 -M S W 0 0 4 -0 -0 4 1 104 (C0040469 Spk MI 5000 ppb) D 0C -FO 1-M S W 0 0 5 -0 -0 4 1 1 0 4 (000*9464 Spk N. 10,000 ppb) D B C -F 0 1 -M S F 0 0 1-0 ^)4 1 1 0 4 (C0049468 Spk C. 10,000 ppb) 0 B C 4 -0 1 -M S F 0 0 2 -0 -0 4 1104 (C0049466 Spk 0 , 10,000 ppb) D B C -F 0 1-1 P F 0 0 5 -0 0 4 1104 (C004946B Spk E. 1000 ppb) 6 C -F 0 1-1P W 0 0 4 -0 -0 4 1 1 0 4 (000*9*60 Spk F, 1000 ppb) D B C -F 0 1 -IP W 0 0 5 -0 -0 4 1 1 0 4 (C049460 Spk G, SOOOppb) D O U -F 0 1 -IP W 0 0 3 -0 -0 4 1 104 (00049*81 Spk K 1000 ppb) O O U -FO I -IP W 004 -0 -0 4 1 1 0 4 (00049482 S pkl, 1000ppb) D O U -F 0 1-1P W 0 0 5 -0 -0 4 1104 (00049483 Spk J. 3000 ppb) D L 1-F 01-M S W O 01 -0-041104 (00049466 Spk K. 1000 ppb) D L 1 -F 0 1 -M S W 0 0 2 -0 -0 4 1 104 (00049*88 Spk C, 1000 ppb) D I1 -F 0 1 -M S W 0 0 3 -0 -0 4 1 1 0 4 (00049*87 Spk 0, 1000 ppb) D1 1-F01 -M S W O O S -0441104 (000*9*88 Spk 5. 1000 ppb) Am ount S p ike d M o) 1000 5000 1000 1000 1000 1000 1000 10000 10000 10000 5000 10000 IOOOO 10000 1000 1000 5000 1000 1000 5000 1000 1000 1000 1000 CS S u lfo n ete P F 0 3 A re l Found In S am ple (no/a ) w e t 4 Am ount R e covered (n g /o l w e t w t R e covery ( * ) ... 2080 2660 78 3330 8680 107 1860 3180 150 506 1780 118 836 1740 110 644 1650 111 1210 2810 140 480 23200 137 7520 16800 91 12800 23200 108 3440 8520 102 5240 16200 100 5240 14400 92 4780 15200 104 80S 1230 1800 2500 90 136 2620 8520 78 2270 2640 37" 1740 2740 100 2130 7600 109 988 1860 89 708 1780 107 606 1930 123 004 1680 106 "Not reportad da to quality control reault failurea. Valuee are not Included in the cutem aty ataltelica. Exygen Research Page 46 of 132 Interim Report # 10-Analysis of Fish and CJam Samples Exygen Study No.: P0001131 Table V. Matrix Spike Recovery of PFOS in Re-extracted Fish Samples S am ple D a acrip tio n (continued) C 6 Su to n a la P FO S Amount Splead (n fl/3) Am t Found In S a m p lt (nota) w at w t Am ount R covarm i (n o ta) w et w l R ecovery (% ) D L 1-F 01-M S F 002-0-041104 (0004M71 Spfc F. 10QC ppb) 1000 see -1090 112 D L 1 -F 0 1 -M 5 F 0 0 3 -0 -0 4 1 104 (C0O4M72 S i* 6.1000 ppb) 1000 2 1670 124 D L 1 -F 0 1 4 4 S F 0 0 6 -0 -0 4 1104 (QOO40474 Sc* H. 1000 p*X>) 1000 1 740 1830 109 D M C -F 0 1-M S W 005 -0-041105 (C0049400 Spk 1.1000 ppb) , 1000 560 1780 122 D M C -F 0 1-M S F 0 0 2 -0 -0 4 1106 (COO40400 S f* J. 1000 ppb) 1000 620 1500 88 D O U -F 01 -M S W 002 -0-041 210 {O00564M Spk K. 10.000 POO) 10000 26200 32700 45 O O U -F01 -M S W 0 0 3 -0-0412 10 (COO504M Spk L, 5000 ppb) 5000 ; 4040 10200 123 D O U -F 01 -M S W 0 0 1 -0-0412 1 (00056440 Spk C, 10000 ppb) 10000 7800 25000 174* D O U -FO I -M S W 0 0 4 -0 -0 4 1 21 (00006461 Spk D. 6000 ppb) 5000 5120 9880 96 DO U-FD 1 -M S W 0 0 6 -0 -0 4 121 (CO066462 Spk E. 1000 ppO] 1000 466 1780 128 D O U -F D 1-M S F 005-0-041216 (COO60463 Spk F, 1D0C ppb) 1D00 720 1820 110 O OU-FO 1-M SFO O3-O-O 41210 (COO56400 Spk o . 1000 ppb) 1000 846 1720 87 D B C -F01 -M S F 0 0 5 -1 -0 4 1 21 (00066447 Spk H, 10000 ppb) 10000 9320 32400 230* D B C -F 01-M S F D 05-0-041216 (CC066466 Spk 1,10000 ppb) 10000 6960 24100 151 D O U -F 01-M S F 002-Q -04121 (COO59460 Spfc J. 1000 ppb) 1DOO 640 1720 100 D O U -F01-M S FQ 02-1-04 1 2 1 6 (COO50471 Spfc K, 1000 ppb) D O U -F 0 1 -M S F 0 0 4 -0 -0 4 121B (C0056472 Spk L. 1000 ppb) 1000 1000 692 1820 1950 2700 128 B0 D O U -F 01 -M S F 0 01-0-0 41216 (C00504/3 Spk C, 5000 ppb) 5000 2070 7960 118 O B C -F01-M SFD 03-O -O 41219 (COO96474 Spk 0 . 10000 ppb) 10000 4640 15500 109 D B C -F 01-M S F 00 4-0 -04121 (00066475 S |* 6.10009 ppb) 10000 9280 24500 152 O M C -FO I-M S FO 01 -0 -0 4 121 (C0066478 Spk F, 1000 ppb) 1000 640 2040 120 O B C -FO U P FO 01-0-041021 (C0040148 8pk 0.1000 ppb) 1000 364 1350 97 D B C -F 01-IP F 00 3-0 -04 1021 (COO441SOSpk H. 1000 ppb) 1000 203 1340 106 D O U -F 0 1-I PFOOS-O-041021 (C0046166 3pk 1.1000 ppb) 1000 359 1400 104 "Not reportad dun lo quality control result fatante. Valuta a rt not Indudad In l i t summary talases. Exygen Research Page 47 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Table VI. Matrix Spike Recovery PFBS, PFHS and PFOS in Clam Samples Sample DLW01-CRIV001-0-041210 (CQOM4009pkC.tetete DI3-IO 1-CFW O 0UH M 1216 |C0M4S0kO,10Opptl) DMO401-CFW001-0-041216 {COW4S1 E, te POte DMOI01-CFW01-IW 4121H (00000411 5<* f . >00PP) OL24O1-CltW 0Ol-0-04121S (CC0CM42 3f* 0.10 pp) DL2-101-CFW001-G-041210 [IJDWB >qK 100CT*) DBOW 1-CFW001 -0-041210 {C K U 13 S *U n } OC-I01-CPWO01-0041215 {OOOOMtsapfcj, to o *) OU 401-CFW001-0-641216 lCOOdM145|*.lO pte DL 1401-C FW 001-0-04121# (OOOQB444St* L, 100pp) DOU-KJ1-CFWO01-0-041216 (COOMMflOSpaM. 10ppb) DOU-IOI-CPWOC1-0-041216 (CnoSMtf Spk N. tooppej Amount Amt Found 3p*ad in S arnie 10 ND Am outt R *W W d 606 Ami Found Racovary In Sump*. i* i 00 NO Amount RftOovervd 618 iw ND 02.4 NO 968 10 ND 100 NO f t 04 90 NO 02-9 03 NO 644 08.8 10 NO 100 ND 10 NO 100 ND 7.20 73 NO 036 04 NO 10 3 103 ND 04-4 04 NO 704 101 106 87.2 10 NO 100 NO 60S 00 NO 86.2 86 ND 832 103 10 ND 100 ND 646 86 0.202 8.32 044 84 0202 960 Amt Found hSam pte Am txjw Recovery <*) ( * ) 2 1.12 632 40 96 1.02 882 03 04 610 00 616 11.9 82-8 61 87 7 f 101 101 1.81 100 4.40 97 4.40 7.52 064 130 88.4 67 67 OS 82 3 600 103 600 16.Q 82 88 80 01 11.4 172 60 96 11.4 96.0 07 II i 04 A w f f: 10 Standard Deviations NO No detected at or abow Lim it at Quantitation wblch I 0.2 n tf0 w a ltftf). Note: Uno u irn w y tetite roundad faautt, wcovaey vaftia t may vary aghOy team tea vaiuaa In * raw date. 83 13 te fra o r. ttendprd Debatten; 71 18 Exygen Research Page 49 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 FIGURES Exygen Research Page 50 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 1. Typical Calibration Curve for PFBS in Methanol 0 9 ie o S E F M i.iU k (F F t}` U i< M r 1 1 0 0 4 x lJ T 7 *-0 0 3 (r- OWOS) A rea, c o u n t! Exygen Research Page 5 1 of I32 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Figure 2. Non-Extracted Standards of PFBS in Methanol, 0.0005 pg/'rnL and 0.001 fig/niL, Respectively 3 S 7 7 J 9 - P m (W U + 4 + n i) s m s -s sm p h 1 Of 30 from ttJCMEuMT A n s : S M Jeon# * * * * * J t t cp* *7! Il3 /2 - / * 0.61 8S7279 - PFBS C3tsnjf) 206 0 0 0 .0 am - Mmpi2 of 30 Vom MIBOOEjaiff A rts : 10037 *1.0.510 m f 002 tntensit. cp* Intensttp, cp* Exygen Research Page 52 o f 132 Intenetty, cp Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 3. PFBS in Control Fish Blank, 0.00625 pg/mL Fortified Control Fish Spk A, and 0.025 jig/mL Fortified Control Fish Spk B, Respectively `PfVS p--a*tf9m<0 9*f39tr*w iMKdir I ReegeH 3 * A PF<OC)200 .000 0 m u -u rn p n 10 e136 *m OIflE.Ntff Area:74474 count* Height 100. epe RT:<42m la 1000 4A2 000 0.59 n 1 23 6 9 10 11 12 12 14 10 I *I PF#*cac)MsmoBeunu.ui.fi. 11(mMMaeiaoeijTNimifel, min Area: 330820 oee fl* Height: 10000. opt AT: 0107 min 0.00 ln1ons%, cps In1*nv, cps Exygen Research Page 53 o f!3 2 Interim Report # 10-Analysis o f Fish and Clam Samples Exygen Study N o.: P0001131 Figure 4. Chromatogram Representing a Fish Sample Analyzed for PFBS (Exygen ID: C0049478, Data Set:06I605E) An: ! A3/WR7i -~mp* 2*3$*pm t Exygen Research Page 54 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 5. Typical Calibration Curve for PFHS in Methanol 061005E F i * * Um i ^ / W w aifM In* y - 2 M r 0 0 4 x * 34M *4<X 0(r- 04901) A re*, counta Exygen Research Page 55 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 6. Non-Extracted Standards of PFHS in Methanol, 0.0005 pg/mL and 0.001 pg/niL, Respectively 9Trat-pfm&--4*ni)3n,*u.$M* i<xnfr*m miux.wm Arm s: H l tt 1 4 k <p# R 7 i H J m fm 1032 litansitr, cps Tim*, min 1 07270 PFH0 (Otn<ar0 300 0 *0 ,0 im u -u m p li 2 ol 30 <tm OOlOOCE^iff Ak u : 33170 M untt 1300. Op *T : 10J min 1033 Intensttf, cps Exygen Research Page 56 of 132 Intensity, cps Interim Report # 10-Analysis of Fish and Clam Samples Exygen Study N o.: P0001131 Figure 7. PFHS in Control Fish Blank, 0.00625 jig/mL Fortified Control Fish Spk A, and 0.025 |ig/mL Fortified Control Fish Spk B, Respectively CM tr*/ p M lM t to f J9fr*m J R igni A . PfH$<OC)3BQ 0U80O im u &ampltt 10 of 36 from 001006E.wM Am j : 100711 counts N o li 13000. o*a RT: 10.3 irln 1032 A rts : 721071 o*unts H eight 30100. cpt RT: 10.4 ml* 3J0m4 2J004' 1j04 0 .0 - 1030 2 3 4 0 B 7 0 0 10 11 12 1 14 1fl 16 17 Tim, rain Inrtensttr. cps Intensity, cps Exygen Research Page 57 of 132 Interim Report #1 -Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 3. Chromatogram Representing a Fish Sample Analyzed for PFHS (Exygen ID: C0049478, Data Set: 061605E) vw mine ranc** im 11-t3Mmmirpk XofMfromKtdMEwMr Exygen Research Page 58 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 9. Typical Calibration Curve for PFOS in Methanol 10006 F ( PFO>- un! " 1 .l6 *0 0 4 x 3.96 003 (rO JM 09) A r *, count Exygen Research Page 59 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 10. Non-Extracted Standards of PFOS in Methanol, 0.0005 jig/mL and 0.001 pg/mL, Respectively I n r m -pfcx & * * * * ) *m -m p * i At m : > 3 I|< m w (i M tfg t d l l tp HT tlS m * 12.4ft Am i : 1 4 067 Wfitf Htlghl; 9 0 0 N RT: 12 min 12.4ft Intensi, cps Itrlersrt, cps Exygen Research Page 60 o f 132 Interim Report # 10-Analysis of Fish and Clam Samples Exygen Study No. : P0001131 Figure 11. PFOS in Control Fish Blank, 0.00625 pg/mL Fortified Control Fish Spk A, and 0.025 pg/mL Fortified Control Fish Spk B, Respectively I A M fw i C s w in /-PFOS A h : 7 3 m com m ta M i f i t S C H H Ji f l i m C mpto I o t l t t f m N W IM lir i> Vw * 1.1 I ae 3.41 a a sy 1 2346 .A4 87 R aagunt 6pfc A - R F O t (OC)400.OJ0OO m u 10 Area: 71600 oounK Mt9 h t 4940. 9M RT: 1 2 4 mm M 30 e(7 x y X . - ,ru , rysv *01 Tin, ml fom 001000E H r 12.10, .19.00 t700x 1741 liitanay. cds Intensity, cp i __ Intanstr.eps Exygen Research Page 61 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Figure 12. Chromatogram Representing a Fish Sample Analyzed for PFOS (Exygen ID: C0049478, Data Set:061605E) -p ro s fijtitrm m ') -ttm y j, H o titf n m H ttv iE .m tr A n t. M 2**W c o h (> t h t g t e U S S M iq p * t m H i m it 12.7 Intensity, cps Exygen Research Page 62 o f l 3 2 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 APPENDIX A Study Protocol P0001131 (Exygen Study No. P0001131) with Analytical Method, Protocol Amendments 1 and 2 and Deviations 2 and 3 Exygen Research Page 63 o f! 3 2 Interim Report #10-Analysis o f Fish and Clarn Samples Exygen Study No.: P001131 Exygen Protocol Number P0001I31 STUDY PROTOCOL Study Title: Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perflnorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program Exygen Protocol Number: P000113! Performing Laboratory: Exygen Roteateli 3058 Research Drive Stale College, PA 16801 Phone: (814) 272-1039 Sponsor Representative: Michael A. Santoro Director o f Regulatory Affairs 3M Building 0236-01-B-10 S t Paul, MN 55144 Phone:(651)733-6374 Page l oj 6S Exygen Research Page 64 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygcn Study No.: P0001131 Exygea P roioco) Num ber: POOO1 1J I DISTRIBUTION: 1) Jsisimh* Kesari, Study Director, Weston Solution! 2) John M . Fliherty, Principal Investigator, Exygcn Research 3) Michael A. Santoro, Sponsor Representative, 3M Company 4) Exygen Research Quality Assurance Unit Exygcn Research Page } o f65 Page 65 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygen P ro to c o l N u m b e r POOO1131 PROTOCOL APPROVAL Study Title; Anilysit of Perfluorobutaneiulfonete (PFBS), Perfluorohexaneaulfbnate (PFHS), and PerfluorooctanerulfonitB (PFOS) in Water. Soil, Sediment, Fish, Clime, Vegetation, Small Mammal Liven and Small Mammal Scrum Ueing LC/MS/MS for the 3M Decatur Monitoring Program Exygen Protocol Number: P0001131 Exygen Research P a g t J o f AS Page 66 o f 132 Interim Report #10-Anatysis of Fish andClam Samples Exygen Study No. : POOO1131 Exyfcen Protocol Numbet : POOO1131 TABLE OF CONTENTS TITLE PAOE................... .........................-- ........................................ .......... ...... ........... ...1 DISTRIBUTION..... . ......... .......... ....... .................................... ..................... ................... ...2 PROTOCOLAPPROVAL------------------------- ----- ------- ---------------- ----- ----------- ;.......................3 TABLE OF CONTENTS----------------------------------- ------ .................... ............... ........ ................ 4 INTRODUCTION--- -------------- ---- ----- ----------- --------------------------------------------------------- ,,..5 TESTMATERIALS-- ---------------------------- ---------------------------------- ------------ .....................3 OBJECTIVE .......... ........................... -- ........................ -.................................. TESTING FACILITY----------------------------------------------- ---------- ------------------- --- -------------- 6 STUDY DOBCrOR............................................................ .............. ......... ......... SPONSOR REPRESENTATIVE.................................. ...................................... -- ............. ....... 7 PRINCIPAL INVESTIGATOR------------ ----- ------------------------------------------------- ...... ...............7 PROPOSEDEXPERIMENTAL START AND TERMINATION DATES___________ __________ 7 IDENTIFICATION AND JUSTIFICATION OF TOE TESTSYSTEM............................ ___ _____ * SAMPLE PROCUREMENT. RECEIPT AND RETENTION _ ..... ............... ............. .................... 8 SAMPLE IDENTIFICATION______________ __ __________ ______________ ________ __ ANALYTICAL PROCEDURE SUMMARY........ ............................ .......................... VERIFICATION OPANALYTICAL PROCEDURE . .................... .................. _______ ___ * METHOD FOR CONTROL OF BIAS..... .......................... ................................... . STATISTICAL METHODS. .................................. .......................... ............... ............ ........ | 1 OLP STATEMENT___ ___. ... ........ ............. ........................................ SAFETY AND HEALTH______________ _____ _____ ____ ____________ AMENDMENTS TO PROTOCOL ...................... ......... .......................................... .....................13 DATA RECORD KEEPINO.... ....... ....... ................... .................. ................... QUALITY ASSURANCE................... ...................................................................... RETENTION OF DATA AND ARCHIVING........ .......................... ........ ................ APPENDIX L ANALYTICAL METHODS.......................................... ........ ...... . ......... ...........15 Pag* 4 of 65 Exygen Research Page 67 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E x y p a P rotocol N u n * e r P 0 0 0 1 U I INTRODUCTION The purpose of this itudy it to perform analysis for periluorobutanesulfonate (PFBS), perfluorohexanesulfonatc (PFHS) and periluorooctancsulfonale (PFOS) in water, toil, sediment, 3th, dam*, vegetation, small mammal livens and small mammal arum using LC/MS/MS for the 3M Decatur Monitoring Program. Ths study will be audited for compliance with EPA TSCA Good Laboratory Practice Standards 40 CPS. 792 by die Quality Assurance Unit o f Exygen Reaenh. TEST MATERIALS The test materials are perfluarobutanesulfonate (PFBS), pcrftuarohexanetulfonate (PFHS) and perfluorooctanesulionate (PFOS) and are all attpplied by 3M. PFBS Chemical Name: Pcriluorobutaneaulfonate Molecular Weight: 338 supplied u the potassium salt (C|FSO)K*) Lot Number: 101 Purity: 96.7% Transitions Monitored: 2 9 9 - 9 9 Structure: PFHS Chemical Name: Periluorohexanesulfooate Molecular Weight: 438 supplied as the potaasium salt (CFijSQ jTC*) Lot Number: SE036 Parity: 9S.6H Transitions Monitored: 3 9 9 -a 80 Structure: Pagr o fAi Exygen Research Page 68 o f 132 interim Report # 10-Analysis o f Fish and Clam Samples Exygcn Study No.: P0001131 Exyg*n Protocol Number: P0001131 PFO S Chemical Name: Perfluorooctanesulfonate Molecular Weight: 538 jupplied u the potassium aalt (CiFnSOiX*) Lot Number. 217 Purity: 86.9% Transition* Monitored: 499 - 9 9 Structure: OBJECTIVE The purpore o f thit itudy ia to perform analysia for pcrfluorobutancaulfonitc (PFBS), periluorobexaneaulftxiate (PFHS) and perfluorooctanesulfonate (PFOS) in water, toil, aediment, fish, clams, vegetation, amall mammal livers and amall mammal serum for the 3M Decatur Monitoring Program using the current versions o f the following Exygen analytic] methods: V0001780: "Method of Analyst! for the Determination o f Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781: "Method o f Analysts for the Determination o f Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" V 0001782: "Method of Analysts for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method o f Analysia for the Determination o f Perfluorooctanoic Acid (PFOA) b Fish and Clamt by LC/MS/MS" V 0001784: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method of Analysia for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" TESTING FACILITY Exygeu Research 3058 Research Drive State College, PA 16801 Phone:(814)272-1039 Pag* ( o f 6S Exygcn Research Page 69 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : P0001131 Bxygm Protocol N um ber POOOl 131 STUDY DIRECTOR Jaisimha Koran P.E., DEE Weston Solution*, Inc. 1400 Weston Way West Chester, PA 19380 Phone:(610)701-3761 Fax:(610)701-7401 j .keian@weetonsolutiom.coni SPONSOR REPRESENTATIVE Michael A. Santoro 3M Company Director o f Regulatory Affairs 3M Building 0236-01-B-10 St. Paul, MN 33144 Phone: (651) 733-6374 PRINCIPAL INVESTIGATOR John M. Flaherty Exygen Research 3038 Research Drive State College, PA 16801 Phone: (814) 272-1039 john.flaherty@exygoi.com PROPOSED EXPERIMENTAL START AND TERMINATION DATES It is proposed that die analytical portion o f this study be conducted from October 01, 2004 to December 31, 2005. The actual experimental atari and termination dates will be inclnded in the final report. Pagt 7o f65 Exygen Research Page 70 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study.No.: P0001131 Exygro Protocol Number: POOOL131 IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM The following art the tast aysterm for Otic study: Water (groundwater and surface water) Soil Sediment Fish Cams Vegetation Small Mammal Liver Small Mammal Scrum The sample* will be collected by Wecton Solution. The control samples will be purchased and prepared by the testing facility. Purchase and processing details for the control sample will be included in the final report associated with this ttudy. The test systems were chosen to access the environmental impact of PFBS. PFHS and PFOS in the Decatur. Alabama area. < - SAMPLE PROCUREMENT, RECEIPT AND RETENTION Water, soil, sediment, fish, cltm, vegetation, small mammal liver and small mammal serum sample* will be received at Exygen directly from Weiton Solution* The detail* o f sample procurement fix- this study are outlined in the 3M work plan entitled "Phase 2 Work Plan for Sampling Environmental Media." The number and type* o f simple* collected will vary depending availability In the field. The total number o f samples received turd analyzed for each matrix will be documented in the final report associated with this study. Water, soil, and sediment samples will be used a* received without farther processing at Exygen. These samples will be stored refrigerated at 2*C-SC. Fish, clam, vegetation and small mammal liver sample* will be processed according to the appropriate analytical method (tee Appendix I). These sample* will bo stored frozen at S -lCTC. Small mammal whole blood samples will be centrifuged in the field at the time o f collection and the serum fraction will be used for the study. Small mammal serum will be stored frozen at i -lCfC. The receipt and processing of the sample* will be documented in the final report and raw data associated with the ttudy. PageB o f63 Exygen Research Page 71 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygen P rotocol N um ber. PQO1131 SAMPLE IDENTIFICATION Prior to u u ly iii, each ample will be assigned a laboratory sample reference number. The reference number will be unique and will distinguish each laboratory sample that ii processed throughout the analytical procedure. Chromatographic data will be identified by the laboratory sample reference number. Sample storage conditions and locations will be documented throughout the study. ANALYTICAL PROCEDURE SUMMARY References: V0001780: "Method o f Analysts for the Determination o f Periluarooctinoic Acid (PFOA) in Water by LOMS/MS" VC00173I: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" V00017S2; "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method o f Analysia for the Determination o f Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS" V0001784: "Method o f Analysis for fee Determination o f Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001783: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" VOOOI786: 'Method o f Analysia for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" The above methods use analytical conditions capable o f separating the isomers o f PFBS, PFHS and PFOS. The final report will include the isomers summed into total PFBS, total PFHS, and total PFOS found. VERIFICATION OF ANALYTICAL PROCEDURE A laboratory control sample will be used for the preparation o f fortified control samples. The test substance will be made into solutions at per the method, and added to the matrices via a micropipette. For water sampling, Exygen will supply one bottle per sample collected. The bottles will be 500 mL precleaned Sci/Spec Premier wide mouth KDPE bottles. These bottles hive been routinely used for fluorochemical sample Page 9o f65 Exygen Research Page 72 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: POOOl 131 E x y ic n P ro io co l N um ber: POOOl 101 collection at the testing facility and have been shown to be free o f PFBS, PFHS and PFOS. Samples will be added to each container to a volumetric fill line at 200 mL A field duplicate, a low field pike and a high field spike o f each sample will be collected. The low and high field spike bottles will contain PFBS, PFHS and PFOS aa well aa perfluorooctanoic acid (PFOA) and U -1 3 C perfluorooctanoic acid (,JC PFOA). PFOA and l3C PFOA me included in the eolations used to spilce the samples. The results for PFOA and 1JC PFOA will not be reported in this study. Exygen will supply one field blank (control water) and two field blank ipiket (control water fortified with PFBS, PFHS art PFOS at a low art high level) for every twenty samples collided. At the testing facility, each water sample (excluding field duplicates and field spikes) will be extracted in duplicate and wilt also be fortified at a low and high concentration with PFBS, PFHS and PFOS and prooessed through the described procedure to determine method accuracy and to check for bias. For soil, sediment, clams, and vegetation, Exygen will supply one JOO mL precleaned Sci/Spec Premier wide mouth HDPE bottle per sample collected or a zip-seal bag. AH containerVbags used for sample collection will be shipped to the sample location. Samples will be added to each container or bag in the field. At the testing facility, each sample will be extracted in duplicate and will also be fortified at a known concentration with PFBS, PFHS and PFOS at both a low and high level art processed through the described procedure to determine method accuracy and Id check for bias. For small mammal liver, Exygen will supply a 50 mL polypropylene centrifuge tube. For small munitisi serum, Exygen will supply a collection kit for sack sample containing serum separator tubes (rad top), vaeutainera, needle holders and needles, transfer pipettes, and polypropylene tubes, At the testing facility, each liver and serum sample will be extracted in duplicate and will also be fortified at a known concentration with PFBS, PFHS and PFOS at both a low a r t high level and processed through the described procedure to determine method accuracy and to check for bias. Low and high spiking levels for each matrix arc defined below: Matrix Lew Spiking Level High Spiking Level Water SOOng/L 5000 ng/L Soil 4ng/g 40 ng/g Sediment 4 ns/a 40 Fish 10 ng/g 100 ng/g Clama 10 ng/g 100 ng/g Vegetation 10 ag/g lOOtlfi'B Small Mammal Liver 10 ng/g 100 ng/g Small Mammal Serum 10 ng/mL 100 ng/raL Pugt 0 ofss Exygen Research Page 73 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 E ry je n P rotocol Num ber: P O O O tlJl Recoveries are anticipated lo be between 70% Hid 130% o f the fortified level*; however, the exact precision and accuracy will be determined by the analysis o f the quality control samples described above. A statement of accuracy will be included in the final report. METHOD FOR CONTROL OF BIAS Control o f bias will be addressed by taking representative sub-samples from a homogeneous mixture o f each matrix from untreated control samples, and by analyzing at (ear two levels o f fortifications. STATISTICAL METHODS Statistics will be limited to those specified in the subject methods and to the calculation o f average recoveries, as applicable. GLF STATEMENT All aspects o f this study shall be performed and reported in compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792. The final report or data package (supplied to the Sponsor) shall contain a statement that the study was conducted in compliance with current and applicable GLP standards and will outline any deviations in the study from those standards. This statement will be signed by the Study Director and Sponsor Kepreacntativc. REPORT A final report will be prepared by the principal investigator or their designee at the conclusion o f the study. The report will include, but will not be limited to, the following: s The name and address o f die Study Director, Sponsor Representative, and of the testing fhciXty. A statement o f GLP compliance (any related documentation, such as chain-of-custndy records, must be in the study records). PUt U o f 63 Exygen Research Page 74 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygn Protoool Number. P000113] The signed and dated statement by the Exygen Retearch Quality AMunuice Unit regarding dale o f study inspections and dates findings were reported to the Study Director and Management. A description o f the exact analytical condition employed in the study. If the subject method was followed exactly, it it necessary to include only a copy o f the analytical method. Any modification! to this method w ill be incorporated into the report If the method ii photo-reduced, the project number and page number muat be included on each page. Description o f the instrumentation usod and operating conditions. AD results from all sets analyzed. Control and fortified samples w ill be identified and the data table w ill include sample number and fortification level. Representative chromatograms for each analyte in each matrix, including chromatogram o f a standard and a control sample, and a chromatogram at a fortification lev el The location o f the analyte peaks w ill be clearly identified in all chromatograms. AU circumstances that may have affected the quality or integrity o f the data w ill be documented in the report. Locations where raw data and the final report are to be archivod. Additions or corrections to the final report shall be in the form o f an amendment signed by dto Study Director. The amendment shall clearly identify that part o f the report that ia being altered and the reasons for the alterations. The amendment w ill be signed and dated by the Study Director and the Sponsor Represeotativei A ll applicable requirements for reporting o f study results as per 40 CFR 792.185. SAFETY AND HEALTH Laboratory personnel will practice good sanitation and health habits. Every reasonable precaution shall be taken to prevent inadvertent exposure o f personnel and the environment to the teat or reference substsncefi). f a t * 17 o f 65 Exygen Research Page 75 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 E x y jc n Protocol Number: POOUI131 AMENDMENTS TO PROTOCOL A ll significant changes to the analytical protocol outlined here w ill be expressed in writing, signed and dated by the Study Director and Sponsor Representative. Amendmenu usually will be issued prior to initiation o f study plan change. However, when a change ii required without sufficient time for the isaue o f a written amendment, that change may be effected verbally with supporting documentation signed and dated by the Study Director and followed with a written amendment as toon as possible. In this case, the effective date o f the written amendment w ill be the date o f the documented change. Copica o f the signed amendment* w ill be appended to ill distributed study plan copies. The original amendment w ill be maintained with the original study plum Any deviations flora the study plan or from the analytical method aa provided will be documented and reported promptly to the Sponsor Representative. DATA RECORD KEEPING Records to be maintained include the following (as appropriate): Sample tracking aheet(s) Sample receipt records, storage history, and chains o f custody History and preparation o f standards (stock, fortification, calibration) Description o f any modification* to the method flustrument run sheets, bench-sheet* or togs Analytical data Ublca All chromatographic and instrumental conditions Sample extraction and analysis dates A complete listing o f study personnel, signatures and initiala Chronological presentation o f all study correspondence Any other documenlation necessary for the reconstruction o f tho study Chromatogram- All chromatograms w ill contain the following: Sample identification, injection date, arrow or other indication o f the area o f interest, and injection number corresponding to the run. Additionally, fortifications w ill include the amount o f analyte added and the sample number o f the sample that w ai fortified. Analytical standard chromatograms w ill additionally include Ihe concentration (e.g., pg/mL). Pant Ii of 6} Exygen Research Page 76 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygen PrMoco) Number: roooi U l As part o f the documentation the following sheets w ill be included in each analytical set: a run sheet listing the sample* to be run in the set, and an instrument conditions sheet describing the instrument type and operating conditions. QUALITY ASSURANCE The QA Unit o f Exygen Research w ill inspect the study at intervals adequate to assure compliance with GLP'i, and w ill report the findings o f audits to the Study Director, Exygen Management, and the Sponsor Representative. RETENTION OF DATA AND ARCHIVING J . -!-*!. j Ail hard copy raw data, including, but not limited to, the original chromatograms, worksheets, correspondence, and results shall be included with the data package submitted to the Study Director. These w ill be archived with (he original study plan, amendments, final report, and all pertinent information from foe Sponsor. The tasting facility shall keep all electronic raw data and any instrument, equipment, and storage logs for the period o f time specified in 40 CFR 792.195. An exact copy o f the materials submitted to the study director will also be kept st Exygen Research. Exygen w ill obtain permission from the study director before discarding or returning samples. Exygen Research Fag* 14 o f rt,5 Page 77 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No. : P0001131 E xygen P rotocol N u m be r POOOl 13 \ APPENDIX I ANALYTICAL METHODS VQ001780: "Method o f Analysis for the Determination o f PerfluoroocUnoic A dd (PFOA) in Water by LC/MS/MS" V0001781: "Method o f Analyau for the Determination o f PerfluoroocUnoic Acid (PFOA) in Soil by LC/MS/MS" V0001782: "Method o f A nalytii for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method o f Aiialyiia for the Detennination o f Perfluorooctanoic Acid (PFOA) in Fiah and Clarna by LC/MS/MS" V0001784: "Method o f Analyaia foe the Detennination o f Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method o f Analysis for the Detennination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method o f Analyaia for the Detennination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" Exygen Research P a g tIS o ftS Page 78 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 Exygen Protocol Number: POOOIIJI ANALYTICAL METHOD M rthodNwb: V00I7IQ M M M a A ia lye k te r te t P U f n i M t f N i ftw n B itiM li Acw (PPOA) la Wawr ay LOMS/MS AmiyiicAl 7mtla$ Facility: B*yfen IU mmcJi 3031 K M w h D riw State ColUfa, FA id tO l Anmr-iBr. Paul CmmoI j TaefankaJ Lm Ur, LC-MS, b y j a i ioIxvAW Dm /J o ta F la b rty 7 ' V io P lfl WVO fllrtllIM, Xif Xm lLt Dm Exygen Research TUIPm ; 7 P att 16 o f&S Page 79 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Bxy|it Protocol Number: P0001131 ANALYTICAL METHOD M a*d o lA ju y m fta- d ^ I^ m iiid M orN rfuorooclM oic Acid (PFOA) m Watac by LC /M l/M S 1,0 Saopa Thia uOkail m te ffnpbjwd fi tin kokrioa u d qiantiMim of parfluomoeunoic cid bjr HJgfe M M io n liquid CfatonMoiptpfey copied 1o uedem Mm Spactomatnc D a te te (LC/MB/MB) in wate. 2j0 Siftty 24 Alwayt oteariaaft labontey p n o tko . 2-2 Conar il tha ^ ro p riM MSDO baten te u liag anyttem icai fer proper aatey ptw mriaai. JjO $m p l t 1234 AI Waat 40 anLoftaat aantpW te attrnettei. No M M pli p n o a w a t U oaadad te wmat aanplaa. U Stm pte owd ffctpar i sd te rn i< te idvwvd k> q iiilib n u n room 343.4 AU M iapitt OMMtte durM ffhly te x te to te M ag aamptad for macneiian Any M B yte rrte a ln iig partelaa rtou kt te c o m lA ifrt -J00 ry n for -5 m inute and te aqpamaiani aaad te lha attraction. 3.1 SarnyU t e t e t e n paoootem wiQ t e aparffiad in k amrplxng p in te thia F ** AO l i y F t e t e t e <1 W ate-K ?LC grate 4.2 M tem ni-M P LC ptea A } A a o te in A te te - A.C.B. te s te t O rtte 4A IteteM O octanoicteid-S tete-A idrkh 1.0 iM te B M and 1 ! A biffe parfeounca Mqsid tew nan^apli, cqpaMa o f pumping up io 2 aivaada aqaqppad wub a vwiabia v o te in je cte onpubln o f injecting S-2O0 pLcountead fte a n M m Spaoteonaaiar(LC/MS/M 3). 9 2 A dTries m msst nmr tel te peak ianfiratwm and qwaoiitation. 9.3 Aaalytenlbalu r ia t ^ ib ls a fm te g to 0.00001 g. 9.4 ^ tnLiiit e ^Wpofrpsopy1eM eatriltipa tubes. I I IS BLdteom hiopolyTropytea oaurifiign a b a t 14 D Lyw hle eteopIpM (JMOOuL. IOMOOuL I 17 123-snL LOFE jmiuumjmmOi tetttaa. I t 2 n L 4m HPLC vial k it 5.fi DUpoaabiapHMM. 1.10 A olopteua 000*1000 pLand 10-100 jiLX w i* dapoaafel ripa. I I I W ann Sqp Pak Vas 6a < l|) tC tl SPCcanridjat- p*nr2*'T Pag* 7 o f 65 Exygen Research Page 80 of (32 Interim Report #10-Analysia o f Fish and Clam Samples Exygen Study No.: P0001131 Exygen Protocol Number: P0001131 I AN ALYTIC AL M KTHOP I K i (PFOA)m * w r 5.13 sra n e i nalMd. 113 C ttn tiiV P C ip e b fce f^iD a ttf fO nlpeJyptepylenf etoeettJOOOcpm. 10 Omwoem*ielynam I I AM tyticel CbJuno: Nuopbaec KP{Kcyvtooe Scm lcV, 2. I m i * Je mm. $>* NH: 230S^S20) 12 KTC 13 Mobe N e (A ) : 2 m U A m m c won Aeeteie to Weier 6A Mobile Pbaae<B); Mnbaool 13 O raiieoi Prajpezn: D n tia iii) to Id K0 200 2*3 3LA 35 63 IS IS 63 New tom & (iL /rim i IS L 31 0J 75 0J 75 03 35 0.3 U b f*tto e V e lu n e : 13 L (n n b e rc e 4 toes mue* u 3 0 U C7 Q unUlUlwi: P w kA m -w lw a d e to ito rxicsiib r^io n cu rvw U Km T ine: -2 3 n tn u m TW eboe eeedte-- ttelnntoed n t fe to e m tn e y b e d in g e t in erber to flp tu e i to i HPLC tyvem . 70 M */M 39yttm 7.1 M o d i Siecemprey NegnJte MRM node. s e to a o riij 41) - f 369 < *l Tbe ebeve ceetottooi m Weeded n > guide m i may be eW *td n o kt io qwinfoe f l* MKMS lyeme I PraparnlQoefSolutoei K l Mobile N K U 2 eaM uuneoiun ernste le m e ie pnpared by adding 0.154 y o f ennwniim n e ttle le t 00Qm l * f w m . Exygen Research Page t i o f 6J Page 81 o f 132 Interim Report # 10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygea P rotocol Number: POOOl D 1 I ANALYTICAL M gTHO P~ Vatfux? o f w 5 y jiT ite ^Xyraiaerian o fPortiw LC/M&MS i Add (PPOA) n Watar by f . t fc w fo tlfta p rttiM t l S ta t ri SliNttPoitiAgatioa fohafon t . U P rapM oaM ekaotufiaa af-IO O jittaC -afP P O A byw dfhiiif 0 mg o faaalytkftf ataadard (eam ciai h r po tty) mvS dilu to 100 m l wi|h M te d k U frta L LDPB botile. 91-2 A I t p ffe L (betidcjbaa aohdoc o f PPOA lo paaparad by bringing lU caL of the 100 jtfta L totalion la a final valuti of 100 with mrhawt' h i B lU n L L D P fl boula. ? U A 1.0 |*|< M . rifio a tia a aohdanefPPQA la prepared by bringing in m l. o f Bm 10 pjtaL alutkn la a final v c lu c f (90 with mcriMnal in ft 129 ad. LDFB botila. 9 X A A 0.1 p jfitf. fim lftraflaa aofidoo ofP fO A ta prepared by W ln|m | 19 L o f (ba I. iig to L aftlabaa la t h a i v o la o f 100 wtfX atteM l in ft 125 mL LOPS botila. 9.1.1 A 0X>1 pgrinL fta fifio a t * aalufiae o f PPOA it pciparad by bringing IO m l o f Cm 9.1 jig ta L aaluboo id ft final voium of 100 with m f t a d a i IlinLLDPB battit. 9 !A Ybarioab--d f e r it o -- aolafiam la ba atdred la ft rtlH|araaor t Appratinafaty 4*C iad n tabla for ft n ftiin im period of b month IIw b (ha f ib ofprapvainL 9.2 Standoti C alibeato f i b t o i 111 t i -2 UVM&MS ca lib ra to ataadaada prapmed in HPLC water. The calibnfiaa andrai a n proaaaaad to u g h ihc attract-- procedure, <totfcalteaaaptM Tba feUewtag ic ft cypioal cample add itional ooncantntioaa may be ts=t Partfiaato Vataawof Calibration n S r i t a t o Vaftana P e rito Ornati Wafiaoftobl w 1 IO t 0 100 200 landa <aL) 40 40 40 * * * t! > * 0 25 50 - (ann*lt xcouSn-o XC-- Jdyy-1 XCrranddyy-2 10 400 tao 100 40 40 toc XCirmadyy-J 250 XCmnddyy-4 tor M0 _____ IK____ -- a s ___ 40 40 540 XCmnCayy-J IDO XCmmftdvY-ft * 11* mttocMd tmoooCldoa o fIfco aUbraUoc Mnasrd it q u i to J i m it! oooowttrott--, f u lo fco oo uuM nb o f it* Umdm fc a i f o o c tio n (SPEj Exygen Research Pagt !9 o fti Page 82 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygen Protocol Num ber: P000113\ Nyrtm VOOOJ' IO A N A L m U M . METHOD ktaltojo i to y tk h t Abiwimliwtntf Niftwipwanai Arid(tfO A ) inW wty L0MWMS MJ 9X4 9X5 A B a ia *i4 M )u ia a (iu |a u k h B lg u U t B w r i H i set * f amodarbi extracted. # lo n Ml M aided m A n tic n Standard in 15snL polypropylene lubes d 2*C 06"C, up to ten weeks. AHeneae retomes 4 cuwmfaetloo o f atentada may be prepend seeded. 1,0 B * Set Up 10.1 leeh Week e f Meples m eied (D fd e fty N sc le is eeel ioskde ei kesi see rrajant oortocl (metfcod Men* uaraf HPLC weMr) tnd iwo rangen omnote ftmUed at Ir a n i aencoendoiu (lee ceoirol apikd to vcnfy procdant raserary ft* the bate*. I0J AsquimBema for fle tf and laboratory duplieet* and ipMe w ill be apscifted to A * funkty a a n o n plan far tUa project 11.0 Sdnplefletteedoo 1 1 1 H at aree 4fl m l o f i m p le e r* perte e f sae^le dtoed re 40 mL w t water mae 50 mL polyprapylme ceetrifbf tubos (farbty es seeded, repisos lid and d ll re e l) 114 Contaos * C# IP * canrtdtei it g. mL) by pautan )0 mL metoanoi , fis lk *ta b y 5 m L Q rH P L C w iia r(-'2 ta pfres). Dp not let totoras nw dry 113 Leed etmpfs on eend itioeed C 8P fl esrthdpe. Discard sta te 1M Sluts w ith -<5 xol 100% M d S M l Celled 5 mL o f siuate into graduated 13 L potypnpylm a rentrifrgs m bit (final volume - 5 mL). 114 Aiui>c neapiae la ie f etavsepcsy LC/MS/MS. 12.0 Chemsmyspby 12.1 IsjecMhe amoenl e f emb atandmd, amaple m d Ib itifle d senpk into ihe LC M 8/US a jllie . A calibration standard muet precede and A llow aU -- freed s--| ilei. 124 StendtaeorW O Aoom apeedU sm uleui flveorm orteqnoeeirwuon levsli mast be tafcdad toes mntytfeai sec. 124 As e s ta eel o asead etibratien atandarda muet be included w ibc la p w e i esd at the end o f s temple M . Sanoied atontad* e ra be in tn p im d betw een every 3-10 raoplee. Ae an attsmasv, es ta ire ms ot M ond calibradon aadtaa may be bested at toe beflnnln* o f a *n firRosed by traded ilbnctoa n o t a i Im artfre md evtry 5-J0 emplee (le account far a reeoud ta o f atnctod endort*) In toar csee. extracted calibration cuitada muetbe lbs dm and leal injection in s ample aet. IX lira tim er a s t a emes fc f nanrtftfcm . L in a r a n a lta curras an gmwnlsd fce tbs e e ly is by lim er lepesrion eens l/a weighting o peak area P u * r0 yf6 S Exygen Research Page 83 of 132 Interim Report #10-Analysis of Fish and Gam Samples Exygen Study No.: P0001131 Exyten Ftctocol Number P00011J1 c analytical ftcgraop hl riho a t Aoifoeu to r * Drierm r otino c ffarfruarooctm oic Ac4 (PPA>u> Weierby LCAAW O VKM ciilbm tA rtendte aonounrorien iuta* MueLynx 3.3 (or eqivekni) M lb M tx ia . t U Sanate imporne m U m i w i l d r i t e t e reeyoooe. Any umpi tei i m i m u l t e teepanem tauri he tather rid ite end r--WyeW. 13.0 Afleepeeeenleria 131 C lim M y ffi-- N riw w i p--k o f a te n h w km et >49 era ftonv pironi of 413 mtm. H 413 em panni ooctMpqoi io ita PFOA aiion, while ik deaghter bm (3f> em) rtproenU te lo ti o fcaiboa dioxiic. 13-3 M eted bUaka m i noi Midan FFOA lavala frc tttr te n th LOQ. If ta n k onwhti-- PFOA et levali jr v rter to n SOn y l* tben new blwik wn^W n m herto rin ari end th ln a iiia iu b i ite in M d . 133 tonoveri f-- -- 1te tte m -- iw 0 m muai ho batweew TP.I38S o f te i* tnomm v * . If e eooeul *ido te aocaptee U rnii. te H it o f umptea ite u ri he w*wracted. Any rw trii apiko ouistc 70. 1M H te id he evahuMl by t e neij-it k i detonate if la-axtracion ix y errenari 13.4 to y calibrali m odini to te lo he a ie riu ic o l o n d i* by m a i te H at* Eh m Tori, nay ho enoludotf frodi to mlrnlorvoo o f th celibrade cerve. Howwo, te M al sumber o f oxmcled eeJjbntlon n m M i (hot eoeld be motte.u u luded m ori aot n e ri 2OS o f te M al Bombar o f extraote interri I U The renettaaa ooefBctet (1 ) te orito ariou enrvee tonerete m uri he hOitW (R1 *0i9tS ). I f e riteteoa iw ih M I em ide tee liana. te a appropria ape m uli he Uton lo ttju * marunmi opereiior, uni in m aterie a rte re ite ra m i o f tem p i* ebouM be rotiulyvotl. H A lu te in o te hetween m udarle end m ote m uri tot d rift more ihen 4 H witln M u u lytice lun. I f iwteotioii thne tk iil eneoeditire limM wilhin m u u iy tk d run duci tbe eei ouet ho reeoelyiod. 14.0 CifcnletMne IL I Uee te fe te te f quatte le c o lo rita t e aowooi o f PFOA fo lte (a np/L. betel co peak m a ) uetag te d a n te curve (linee* refreuto penametert) f * erfte d hy te Mm Lyn to te m p ro p *: n04hnl(k|lU DF O F - te to * hy hkh te fim i volume v ia diluted, Ifneceecery Exygen Research P age 21 o f 63 Page 84 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No. : P0001131 E x y *c a Protocol N u m be r POOOl 131 U n m t mmmk MVQDOmo I ANALYTICAL MKTHOP 1 K fe M e fA ju ly ib rtfc o D o M is itiflfttfP v fU iM Q O c U M te jU i4{P F O A )ia Wuerfcy LW81US H-2 For uaa plti lo rtifM v iA Itnewa mw mx o f FPA prior U M inoiion, um ti Nkrmug jurtica i ctkttlrts dm pmvm noo*my. IHvay(H)a | w l -- tyiohunrt(jm/L) > aaalytofcundin otm ttd (w T .)] aoriyodd^(b^L) * Exygen Research Page 22 o f 65 Page 85 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: POOO1131 Exyym P rotocol N um ber: POOOl 13 J ANALYTICAL METHOD katbod Vlunbcr V00P17H Matkarf a f AaaJyib to r tU Datarb Ju Umi r F i r Q W M t t M t i A ct* (FTOA) to Soil by LCMma AaoiyticjJ Tttof Focitkjr K30nSySfaHomt mwtncaHr dD>hv# JUfOCoOfft, FA litQ l Arpm ify. " P A C-JJL m r r m ii^r i Tadncat U ctor, LC-MS, feryM U n l i t Data / V k a ProafraL O pora**. b y iM M a^k. Exygen Research Total Pi#: 7 i o $ f i j o f 6S Page 86 of 132 Interim Report #10-Analysts of Fish and Clam Samples Exygen Study No.: P0001131 Exyfea Protoca]Number POOOJIJI ~ ~AK LynCA LM tTH O D erfAteyele far fae D o irwiii l im e f H r t e -- LCA0 /MS ic A t e (PFOA) in Soil by Ib i m ( M be --ftoyod fcr te ieolettaa d q-- itebe o f ptel*oreoctfk cid by High W w o m m U *iid C lm m io fra fh y ooupM H nodo* Maw f r ettio n iet fc Oe icior (tO W M g J fai kXI. 2.1 M ty 21 AJweye ebeevt t e Laboneocyprette*. 2.2 C-- Jt te te n g c lM USOS b e te b--dite* --y c h -rire l ferprapcriefciy k h Ub . 3.0 Jtropte ftequfeeaer 2.1 A llM D llftM ta M fIfik r tttiM iM . 3.2 No w p to p rm m rii h medid Jbr lol --qde. J J S i^ M la te re ftftte te ibcuM bo tt-- 4 IO e q u ilte * io n m M A ll M mpte iauet to (hetvafMy eixed be&ne b te g eerapled for ectrtction. yntetJ . i h n p ic odlMtian j-- ned-- will be p*aBo4 tu Ite umpfavg p i t e tfcis 4.0 m ite n b 4.1 W tlV -N T L O r^ 4 J M ite n i-H T L C ite e 4 J A e m o li-- A-- -A .C 1 . Ite fu iftto d e 4A F o ri-- fa A te - S i#---Aidricb 5.0 l-- oo--il m iBy --ra-- 5.1 A high p*tem iiM O Hquid du-- ctyepA e--it e o f pumpiag up w 2 te -- i HUippiO wit i t e t e vote-- iqjeaet o p t e o f lMecd| J-2uo nL-- le e i--deeM eiB pec-- er(LG M A 43). $,5.2 A d e ko e e e lle M n -- te fe ro e e kte e p e te ie iid q u R iM lu tio o . 3.3 Ateytfeil boi-re repohlo of nedfaftt 000002 5.4 -- mi *:- y f \ y | j` i n i i Jf i | r ni n 5 J 5t e s o t e li p o lyte0>tee -- ite it e . 5.5 Di-- t e - e r t e (50-IOOoL> I00-20CM,}. 5.7 125--L LO FI e -- oofa b o tte . S I Jw L cfae rtfF L C vU lU t. 5.9 Dupa--Me pipe--o. 3.10 Aeeplfeiie(10<M 00O |iL s e i IMOOnUb W itti diipoeebleiipe5.11 W tieiS 4p F ik V e le ( l| ) C ) l2PBeridte4 5.12 SF8 ve-- e --ifaid. 5.13 U t -- fcbelh. Pag1 24 63 Exygen Research Page 87 of 132 Interim Report # 10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E x y frti Protocol Nuoiber P0001131 fc w l i-- * AToarnc^MiTMOQ N i^ r VOOOI7tI MoiiJt A alym Sf tm D^mriinolwwo/PrOpoiooctiaow A d i (7F0A) inSoil by LCMMO 5.1f dubcr. H i CwvMUp capibti o f * umIa| JOm lpolypropylene tab at 500C rpitt. 6.1 pMB-- tiy p N i Systwn 5.3 Au|yticiCohau: Fhnpfcu V (Iryrtoaa Idrofilie).1 1mm x 50run, 5m tF 4 f:C 2 9 tt< O S 2 lJ0 J 42 T d p m : J0*C I S M o M k P lu o (A ); 2 m M A m n o a iu n A m a te lo W m 14 M akik Pb*M (B) ! Mecfcanel 4.3 O ndialP ra x is : Flow Rat* ILA iaUicm] 0.0 45 33 0.3 I J 45 35 0.3 10 25 73 0.3 20JO 25 75 13 22J 45 33 0.3 i i t i^ T r io n V d w P K H p * . ( c e b e i o a a w d w i u c e e 5 0 >lL ) 1 7 Q w oiuiU e: P w fc A m -o tm lm d o d o ilib n tu N o u rv . 04 JUnTtoo! -2 3 ateuM . MtMttteKPiCiyMn.TiM dborocoatfdeM roiiaiandedttafB iierodjM ybeehanfoiiiianfcrC D 7. M5/MS lyraoi 7.1 Mod; I k otrc^ w yNsiaave MRMnodo monitorial 413 - 349rki for PPOA. Tfea ite v c J d ttfe u m k m t k d a t * p u d ro d m ay be changed in ord a r u> pmm.o p d a J ie tte 3 tt Q 10 P nfm ey ofSotutiaae 11 KobUaPbMO 11.1 2 m M r o a m iu m acetato m w o t k prepared k y a t t u t i 0.1 S4 j a f tmmcmxm aeetata w>1000 m L o rw to e r. A tercato votone a * y be p re p u e l p#J*n Page IS o f6S Exygen Research Page 88 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygca Protocol Nmbr: P00011 1 itn im-- I AWa L TUCa L M C m O P vanirti I U J U i/V S J) k^dfnyMla 9.1 ntmmAmmi <jnv.fr/IUrri 9.1.1 9.1.2 ^B B C tK ockM luU M O f-IO O nirM iLofrP O A Vy w n jhm * 10 n * tm y i M * is rtiid (ceweoled IU yarity) m i dilato t 100 tL wtf> mediami tee tf$*m L LDPB beala. A lOpfiVaL fatMIceded aoJuonoTPFOA is pnpnr4 by brii^png IO mL a i * 100 taL M ie ta li lo Od) MlW M o f 100 v itti riamai Ina I2S a l LDPB botft*. : 9.1.2 A 1.0 p jria L fenifUarip n U i t a c e f PFO A i> p n p tW fry bowling i m L o ftlM lO ^ friiiJ, Rotolo t e finti v1u m o f 100 nth m rta ro l m a 125 otL L D fE beala. t.M A d i p ii'a iL fcrrtaatca M tasm of FrO Aiaprepued t>y b riri|in f IO L a fN >9 M H , saletfea a A w l v^urna e f 100 w i* weueeet a 125 mLLDPH ned 9 .IJ A OiOl m rtaL AiliSeedee aaloias a f fTO A ie pnywad by irin g ia * 10 mL o f dm l. l ftffu L k M m e a 0m J veto of 100 wNh m ttm m a 122 ta l LDPB botila. 9.1.1 T to Meok ita fertifieatoo asfalta* re la ba taoced ta r*iri#Rm<>r t ta y*n in ily 4 * C a d n n a b li 1br m n i m m pm o4 o f 6 m onta a de d * i e fpupim ioB . 9 J Staodard Calibrata Batataaa 1 L C W M X callbnalaa h ta ta i are prapvta m HPLC water The m lltamim aiA edi ire pw r ran d dstngfa (he eabactfoa procedure. W J Tfe A ta w fe f a a fypse m tw y lr arfirinaal coaMrtratfona may be IST Cm mead fm i i m t m Vehamef Cn .ii m w a -e f CWibmwm m tM tm m m V a t u F n M C a u d Cfelilnaaa te n te ID . Sftejgjrti__ frU.....fr*rtrtiL ,l*44.ipg_ XCmmotyy.] He IM 40 KXCCamuwikMiyyyy.3* 250 XCmifKWyya IHe W< 4M_____ 5)0000Q XXCOanwmMdtyfyyy.5Q H ie M ia d eenoMKntieaQfA e eabbtadenaiteM is equal la I in untisi MmeoMrata^ ta * te the eoaoeanto o f the ewuUrd d ta n f ike traction (8P). MC adianid cattbradra federi. Pag* 26 o f te Exygen Research Page 89 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygen P rotocol Num ber: POOOl 131 ANALYTICAL METHOD of AwtyvU far tbaDaiteminteiim o f PartiuMooctenoic Acid (7FOAJ in Soil byTM IC /M *M S 9.2.3 %2A 9-2-5 A sare te n fo ld aolubcn (roagant b lin k} mux he prqw w t with e*ch M l o f atandanfo W M id . Star all m ated oaUhniioc ateodaade in 15-mL potyproprlwa tubes K 2*C 1 fi*C, op to two weak*. midAfoanutt votumM and caecanmtiaai o f atandxrda nuy be prepar'd u IdO Babe* 5 * Up 10.1 B itk bmcb o f awapUa tra c ts (lypecily 20 or le a ) mute include tenet one o nM I (rn tto d blank tin t 5 mL o f nitefcmtol) and two raapem oototoU fortified kmrmm erne*otm iwu (lab o m ro l apike) to verify prooadonl recovery b ttfit k ith . 10.2 Requunmaua far rtekt and tebocteory dn>Nc*tea end iplkc* w ill be u>ci(ld fa fan qaaiiiyaaem K* plan far tbaaproject 11.0 la o p te C n M id o 11.1 W ei|k 3 $ o i ample fate 10 mL poJypropyt-- oaoufapa tube* (fo rtify aa d m M . veylasu dtf a d mix wslQ. 1 U A id5rf,efow fam elM fatotooes w rite cd o n A ^ir f a r - ll m tiiute I I J l^ m fa r <betuba* to m tdaranoade bath and acucate far - 1J mwoMa. |M B rie f the volume ap te 40 mL wffa water in the $0 mL polypropylene iw d iitija Mhi 111 C totnfafafiv*lO teuivM ate-)O O O ipm . IU5 QandWan the C S M eartidfM (1 f. 6 mL) by paaalnf 10 mL metJuncI followed by 1mL ofKFLC wmar (* 2 cbopW:) Da notice column me dry U-? Lead (dacint) tea temple tm the oooditfanad Cu 8PE cartridge. Discard i )m m . I l l glete wtih *5 teL 10054 Mtehaeal. Collect J m L o f eiuau imo graduated IS aLpteypw py tie nrm ril i f tab {final vofeme SmL). 11.9 Anteyta te m p i- aafag eteeaw^ay LC/VQ/MS 12.1 fafect tbc aimc mnauat e f each teaodard, aampte and fortified sample mu> ihr L bdS/MS ayium. a catfbrutfan undard mute procd and follow all leelyaad aamptaa. U.2 h a lw d a ofPPOA iw r mpaerffaf te te leate Boa armera coooanoteiow l rh atebafaeludadfoaamadyticalM t U-3 Ax notera aat o f attracted calibration aundarda mute b t included die bapawfaf aad it (fcc m d o f a laeapli aat Extracted atandarda anni ba inteiipoiateibtewam mary 1-10 am ?!** Aa an alteraativa. an neirira ate o f Papa3te l P ose 27 o f 65 Exygen Research Page 90 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Bxyfeo Protoeoi Number P0001131 [ AN ALYTIC AL METHOD Mabad o fA ad)u tec fra D atoafaoioa o{ Pbi th w oectoaoic Acid (PFQA) in SoiI by LC/MS/MS sttMts oaftc iria aundoda m a j ba lajecfted m fee tagliarne o f t n i U tooed by am end oaUbnttoa h a ie * iM M ^ m d every 3*10 am pi (to m m I f a t M M lu t t f otorectod M o nadi), h eith n r extracted odihaiiee B a te * uat be (be Ant and iM t tiOectlao m a um m 124 Uk linear aandari curvai tor qiauietctiea. L im a etmdard ( w n *c ftoontfed ftr Am iualyto by Abmt iu fK ie a M ag t/x w fctiitf afpeafc artm Tenui calibrato* rta d a id uneaaairt tk * u tin f MtaaLy 13 (or equivaleri> often yuan. *13 myeoM abould rot sen d atad aid leayqoMC. Anjr sam pi dui Mc4 * an t d teafem m to-- H be iu tk ar diluted and reanalyzed. ILO Accorto* Caftait 13.1 CWeanio y o e amai dww peak a i a danfhnr bom u 369 snu ftean a peni ed413 oeu. Tba 413 anu pan nacropncirii tha PFOa aaoa w hik th decurtar leu ( H f a-- ) rap a mi! Ih tooe e l cobo tooKid. 13.2 M etto* M ori m wt wrt c a la i flPOA at ferale p M r tfc-- he LOQ. ifa bka aonieoa PPQA at levata praaic ttm t 30 ffT , th a aew hlank um p lt m m ha obuad m (ha n i M m be ( e m o ie . 11J fceooeer i e f oboi t y m 3 tori fta a treot he betweeo 7013OS o f (bar I r iw i velue. tf ream i * if e &Ue evtode th o --ptoblt linula. ih* t e m ed M opke sbotto be tcaxtnclad. A *y m irto apikt eutsxte TQ. 130H toouid be eveluaaad by h enalyat d ia m in e if re-csMcrien 114 A jty catfndea am m ftued lo be a toriato! c e lli by uain| toc Hupa ncr Te ceay ba escludati froas (ha calcelatto o f to t caJibmion curve. Howev, th to o l avm ke o f e icaoied calibratioa tn a d o d i d iti ccuM bc t noi e n d 20% o f th total nu n ta o f antraci) lu n titrd i 13.5 Tbe fatato aeeASnoa <R) lo calibeati curva perm uto vai he >0,992 (R* >0.9*3} ( f ealtoreboo rteuha ia ti eutaide thwa lim ila, ihen appartala at* u n M u b a i te adiva tnonunent operaix*. and toc codarda or th re fe --! aatef aamptoa ibcold be reenalyied. 13.6 le timae boweae ftndoda and aaoplea m uti net nfi more diari 4 s wittue a --d ytka l tuo Iftetoetton tona 4 a anal/ticel ra t in a tia aa Exygen Research Pogr 28 o f 65 Page 91 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: POOO1131 Bxyfea Protocol Number P0001131 ANALYTIC AL METHOD Method o fAnelywa for lb * Dewmiostioo o fN riluorooctuidc Add (PPOA) in Soil by LC/MS/MS |4 ) U*e (he fo lk rr f quation to kokte the mount o i PPOA fouad (in a*/L, breed on peek wee) seing die etandad c m (Ijmew re p tu ie n punm rtert] psendad by foi M r * Lpm eoftwue p r v g m : ,n g dope D F - Amor by which (be Sad rn tim r M l diluted, i f neoeeMvy. 142 f m w m flw foclifled w ith kn*we t o o m m o f PPOA pcioe lo extnctiou, u*e til following ofuaoc to c ilocln foe ywoent recortry. analyte edded<n#L) 142 Um (ho following e^uaboa I vmrtwi lbs amount o f PPOA found in iypL rflPP*). PPOA found (jp b )- ] g tw iriiftftlM l w l* w m ju (S g) H A Um I M ow ing eqaetioa tu eetetdeto the m o u it o f PfO A found in p beeeden dry weight. PPOA found (ppb)dry wwght - PPOA found (ppb> (I0 0 H / total eoUd*%)) Exygen Research Page J9 o f 65 Page 92 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E xjrfca Protocol Number: P0001131 ANALYTICAL METHOD I M M M a ta r. V00017IJ H e M * A a d jit t a t a lt a n k U H if F iif t t a m t i -- k Acta [t k ia i I le Mta ie l O y L O W IU AolyH ad Ttatta) Paciliqc Bxygn BatMrcIl X D IlM c k M o Aata C ollect. PA 16101 c jL Peel C a m ay I T e M r il Leader, L U O , Exytaofteeetacb / YicePieodect. Qpwedeiu. B ry fia teeetaeli __mWcN D e li *AUr D it Exygen Research Total Pi # * : 7 Pos* 30 o f63 Page 93 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygen P rotocol Number: P0001131 fe y p e ta w * 1 Muhod o f A im /g a te :L i VMO 17U /J a L Y T lC A L HOTHOO 1 ofP oiflaorooctanolt \ a ii (W O A) in Sediment by LDWWS 1.0 9kp* Thic n o te d if to ba anpbryad fcr te ioaletioa owl qiaotrtabo'i o f pofflukoocundc (Mid by W|fa Poribn&Moo Liquid Chm m alepivky coupled te a (andorc M a il S p d n n u Ite ra to r (LG M i/M S ) Mdimrail. 10 lifiC' 1 1 A h ijite n ie b lo k n n y in td M . 3 3 Caaouft t e qpuprieu M S botea h s U te | aey te n ia e ] t e paper aafoiy 10 Suapie teqairm oiat 3.1 A lloa 30$ e fte d am ple te a d m e n . I t NearaapScpcaoertiflfii needed teaedtaeani ample*. 3 3 SeapJaa naiad reihfcnaod abould be aNewed te equilibrate to room M poteC . 3A AM aaraplee ra te be te ra u ^J y rated betea being iirapled te ctt/aceon. 1.3 teapto o o lle ct procedural w ill ha * e a 6ed in te w ropnni plan for Urn 10 te m a a d to M i .1 W ear- H?LC prade 41 M etend-H FLC pado 4J Aflede Add - Mei^aat yede 4.4 AoMHoeeaa A c ia u - A.C.S. Koageat Orado 4 J Hrflaow octdwie Add - Jiy*-AM riok 3-0 fc iM a r a a d lr .a ^ -- 3.1 A aipb p n tirn m n i liquid d n n w ia p ^ b capable o f p w p ia g up to 2 drea d equipped w i a variable roJarae injector capable o f injectag 5-200 pL caw erad a te d e e M a il Ip e cte w ra r (L O W ta y 3.2 A deeiee to eelloct war data te peofc b le y u ion and qtunuuaon 3.1 f ao>tra* tetee a napabloe/raot e g la 0.00001 g. 54 30 e*L dwpqaeble polypeepylene eenarttogo tte e . 3.5 :< raL iteooobk pofrpaepyhae o te Hto y aiboa. 3.4 fryoeetoto w in op ^ m (30-lQOuL 100-200uL> 5.7 125-L LDPB u m a e e id bottlee. 5.1 1 oaL eloif KFLC rie l kk. 3.9 DippOtlbla pipO ff 3.10 Adopip t e l (100-1000 L m i 10-100 pLL * 1h dlapoaiblo ttpe 3.11 Weiara Sop 9 ik Var 6oo ( l| ) iC lt 391 eartridfae. 5.12 STB racvun audited. Pag* U o f 65 Exygen Research Page 94 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 E x y jc o P rotocol N um ber: POOOJ131 1 A H A tm C A L m T H O D 1 Uabod a l AialpU t o tfe D m ra iu ifa i oftm n w n o a m o i T c iT J ffS k ) i Stdimn bv 3.11 Va (M l l f C-- t f q i at e t q i m m t X m l yolyfny y ln c raba M MOO ip a . U> I A M lyO M l c a -- : T te ite t i [K>a w n iM U tftc ).1.1 m m t S O m n . S ffM IU B M n iM ) 't a r a w x re t t e n a i n 2 oM A a m a im Acatm la W iu r * U O nO N foinK S U 2U o j 35 0 3 14 55 35 03 l . l 25 75 0.3 20. 25 75 0.3 22.5 iS 35 0 3 * 6 m<cdM V o h n r 1 3 |& ( a b tte n u a d t a in u u 3 0 j.L ). 47 Qu rtm lm : f a t A m - a m nal -- dri M lik n tio i. cw vt (J la it e -U TW ito t a a W a i m t e M m |uM a d mwr te d u o p d i o d a eydatl ik HTLC qiaaa. 70 M S /M iM a a 7.1 Mto BafcM|N)f>l^lhf<10a<a<ite.nwniteigt) - HO a ft to ffOA. TW ito t* eaavete n t e M a pM w d a jr k * rttn p td i onto Mis (WMIMt WM. 9.9 Fnponnai if f a i d u n 1.1 Motikltoa 1.1.1 inMi w a t e i > U a 1 propam d Vjr tid in g 0.114 of t u lO O O aLtfwwr. Exygen Research P*tr)2of6S Page 95 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E xygcs Protocol Number: POOO113 1 I A J U iY T IC a METHOD | M ated o f A n iiy ril * r da D a toa dudacrf N rita ra a c m w ! Add (PFOA) in Sadlncn b> LC /M VM 3 U lOOOmLef*--r.12.1 IK m k add te a r ia pra te *) by adding lO m L o f acauc acid lo 9.9 -- fifftp a n tfo n 9.1 Stoewd 8te*fbelifi6@sSehiikai 9.1.1 9.1.2 Paap-- aaoofc*sliinf-lO O |i#*LofP F O A IO<n* a i m M (ao-- 4 flbf purily) aod dilua* to 100 red wash M0MK1 ia t125-m L LOFE boUe A LOji^M L ftnlfioadoo aatadao o ffP O A to pnparaO by b flfifk ii 10 w L oftha 100 u fta L a o h n im i to a foal yoIium o f;0 0 w ith raeihmul k a l2 5 m L U > rB W c k . 9.1.3 9.1.4 9.1.5 9.1.6 A 1.0 in M - h r tito lio u liiib u fP F Q ^ it |w p w 4 by bringing 19 a L o f tka 1 p fta L aobbaa I final vnlume ( 100 with ia ITSaLLD P ShoM to. A Oil mptaL JbtfUtoatim aohnion a f PP G A ii prvparad by b rin fitsf Itt L o ft I.O jtfta L so lu tio n a final voluma o f jOOwich m o m m ua a 135n L L D n hank. A M l p fta L fartficatian s o lo * / PPOA k f t i f m t i by b rin jia * 10 ad. o f m 0.1 pffaJL M iM ta ta a final volwm o f 100 wwS mttm t ) In a 135 mL LDP I W ftla. Tha-- dcan twttfiqtion aaktioM m t in ba ttond in i tafti gamertt PjanmmaMly 4*C and an M a Ibr mxJmuin pariod o r 6 monthi fo n t fi-- a fp ^a n d a o . 9 2 9-- dard C rfh a tir fiolH am 9 J .\ LC445/MS aolihration tandardi an pnparad in BMthaaoi vu diluiuMi o f-- 0.1 jtfto iL JbitMcatia aa--ton. 9X 2 71 fe lk w k f k a typkal sanqplr additional o m x n m iie a may tar C-- nMwi aTM -- * * --aOwnu 100 100 100 14 J MU 10 1 2 10 10 _____ i ______ ___ !S____ luted M tmL) 100 100 too too too too Ffewl raanatemton (n'toL) 100 1 24 1.0 0.J Si K |*4 mft Page JJ o f6S Exygen Research Page 96 of 132 Interim Report # 10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Eay$ca Protocol Number: P0001 t i l o fA n d y* fb rte ^INALYteALMrfBOP **V 90*7Z | BO fM tM C O O ltM dc Add (W OA) in Sediment by LC /M 1/M 3 3J ta n n il atKnn in aandarda i& e a L LOPS narrow-maiitli M in t 2*C * < :, > ala am riln. 9 3 d AHanat>aliinaaiirdcaM aM tfaM faaadardaaiaybapeaearadaa ISO B tuS S tU lp 10.1 B n k bank a t anapln n b a ind ftm o a lly M ar Ian) a n lachda tea aaOaand aaaaal and twa ndraalad aoatfola fc n ila d r. known aam am H aaa Qati aanaal Iftta ) m n rtfy ttnradiua l laoatary lb A t bam . 1*2 P nniia tn t u fc r Said aad lab arauay daailcawa and r> tk n w in ba uUIC ia Iba k ta llly aaamaaa plan * r tWa paojacr. 11. 111 W *b 1 | ad taenia Ia n $0 a l lypteyplaae eeoarilh|B aibea (fcm iy n a n d a i itfla e a lid 1 a n *aU)i 112 Add >1 n L a tm acauc acid, a n , a l a tad d a ta ea a wrist actioa maker lb r-* a ia u a a . 11.3 C a n iA n ta a A n -J 0 M n n ta -2 * n a n la a . li d Cbadidca dn C ,i IPS e n r l^ it [1 g, I n L ) bp in a u a j 10 n L aatkanal M ow adbylO m LetH P LC w aaarf-2ta(M ec). Da not ba oaluiaa nm dry 11.1 Load (daaaak) t a I t i> ll aa tb j cmdlrktaad Cw SPB enrndje. DlacanJ 11 Add 20 n L afaM b n allad ta ndiawa la * la In b o n o f tin 30 n L aaailafa tube C ^ , ta n a m i ahaka oa a a aotioa takas for - jo 11.7 C n ta d n t A a n b n K-lO eO cpn b r-d O nianaa. 11.1 D a a a d ta a ta a a le a *e a a a a a tra e n irtd *t. OoMectihe abnta. 11. T tib - b r - 1-------- l i t ` n l - fn - S n r l CaOaaidH a lm a aad add bra tbs > h f unllTiw t l a i t f 11J. :M 0 C oadiikaojH O BdC ii SFE n r*M 0t ( l t f mL) fry (MMiog 10 m l, methanol fblfewod fry M m l* fUPLC wafer (-2 tto p te c ). Do not lot dumn wo 4ry 11.11 M i tta m a rtin i 1 -200 > L t f wafer m i \ m i on tW fecend eondiiioned SM oanrU f. 11.11 &Jufe w (tt - i mL 100% m Um ( Cotkei S mL o f a h a * into c rtd w rt 15 * L polypopykoa eaatri&aB tefew (find volum e - 5 nvL) 11.11 A M lyw iM flM M iH ^K tre e p s^yU ^M S A tt. >*#. W 7 P ip e 34 q f65 Exygen Research Page 97 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No. : POOO1131 Exygen Protocol Number: POOO1131 SXypftJUHM* U S M N W M VQMI7I I AWa J-YTICa I K 1TBO D I l i risai ofAistlyNa ftp C S S B er it f Nrftuow>oiwc Acvd (B 5 ) I 5 5 mw ky UAOMi 0 9 Uannsaluaptiy IL I l4 a it ia tM ti< > ih re a tfca ld sia tri.a a tt U e a rife n ifb ie tsp l iM il> U D U W i l] im A aaUktiriaa n d a M anta atada assi b l low all M iljiii i^ r flw 13-2 S a r iftr i ofFPCA corr p iriin g to tt to n t Uvor m a n oowcmpU )##1< muM W notodad I t M M lytkal M t `.2 J An m n m i M a d d ftrad i m Urd man U iacluod u ftc Md M m f r i o f a M a*U Mt I ta n ll ta rn be iiM rf ra ri bmwrnn wwy J-IO NnfU. Am m dim itiv, n d m M t a f ta lilin tn n te lir d my l* k0M *d M ft# bfgm nhg o f * M t fc ow ri by calibrairart m darf t h i# f w il tm ry 5-1# npfe (t* oooool for w c o ri m i oi M riv d *). h M tW eam, caU krioa u r i* * am b# ft# Cr ri Im i iaj#ciica ia ampi# #M. 114 Um Haatr a rim i corv# fer quasCutioe. U n w s u riw d e w a re pftM #d fbr t * e w lyft by t iw r m p * eftos Ws weiphljna o f peak arca Varna aaHbaidati ita d a d eaneaknrio < M | MaatLyaea 3 3 (oa attivatam i d -- t r u aa, !2 J Baty l l iannat d w ilrf aal aaaaad atta i t t i raayanaaa Aay ampia [hat awaaal aan itp i raayaaaaa ahaa ll ha ftiOtaa d ista ti and rateaiy ia i U .9 Ataal aaia Cataria 11.1 Can n u lay ^ a aaual stow > peak a f tdaaigksa io aa 38 aia dom i pare e f il) aana. TW d l) aaaa pattai oosraapania io Um PFOA aoioo. wVU Die laopatariaa 0 ianas) tapaaaaaaad a laaa a f aeahoad1sirie. 1 U Maahai Maaka n a i agt aoaala rTO A lavala patta da da UXJ. I f a hfeak i natala i fTOA lavata jtaaaaa Qua 9.2 gftasL, dtan < naw hiaask atonia --0 ba aM alaaia n i Utaanatra aac-- baia a la to ti. D .) Kaeawaalaa a( tta a a l apfeni a l n a tia apkat m ia i >a ta tu a te 70-1W H af *w l auta vaiata IT aaakalaptba fcUt aalaida ita aooaptaMt luiau, tha attrita aaa n f alpha houli ka rwaxmeatd. Any n a n i aptkt nautde 7a. IJOH riaaM ka evala alari ky la t a tu lje t attenuine if ra^xkaciion ia A EU.VTIM Aay akUailna ataa tana Ika a i la ka ataacal a lita ky vaisi ika Huna Enav Tau. aaay ka e s a ltiti ka bcalcaianoa o f a ealbniMn I lavava, da aaaal aaaha n f oneaai ailbnaioa taaniaria itaa oould ka ava Iti a m a scaaC m H n f tha mesi aunhai a f assraslad K m ia iia NH* 13.) Tha aamlallasi aatA naal <JU fe i oaiibnslan oaevaa p a n a tivi asari ka kdP)3 CJt* fcO.MJ). I f aalfbailiaa raaufsa fe il snida (Saas lim .11. Irian pptopria tapa pana ha taiao le talleri inttivttiaaa aptnrinn. ani ika aaaniatia of d a rriavaai t r i a f ampia trin a li ka tu n a lyie ri ra tti ' F o tti a iti Exygen Research Page 98 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E x y jra P rotocol N u m b e r POOOl 131 AN ALYTIC AL M TTBO P ] U i A iA lytl k r a Dw m lim lm e t Pwuawom noie A nd (PPOA) in StdimaM by IQ IO A M U * -- Um Mm kaiM O) m a te ti M d ujaplM must um diiR mon ih n * 4 H ife l a iM b 'k * lm . f f HUMP H a d rill dt M il lim n within H l Uw tba fa lly w if oqar t oa ta r otoulwr m otm ofPFO A hun (iw if m t, b u a a s pa rt m a } iir o f M it M iM earva ( l i u a rapattion panmetan) d Vy tha M m Lynx u w m pw pam . fFQ A im d(a9*n iL )- i DP DF - to te r bjrvAfc K final 142 Fcr iw a iln fertU M *MU Inewa m guaw 0/ PFOA prior to traciioc, uw Hw feUowim t f Mhtm I calculate tha perm it recovery. wowory^i)" [ M alaoahu fcm rf (ajW L) - w alyw feuw diam ot(atfrm l,)I w w t> *U (i*fa L ) 14.) U w tfcabU av1*f quadoarew m w trtw ai 1o f PfO A in ia l in n**nL 10 P T O A Iw M {pp4)- MiBple w jfei (5 f> 144 U w dU fcltowk ofMdaw ( If m fbaad ppb to e d 00 y w atfht ) w calculai tho amount o f PFOA PPOA feuad (ppb) dry w eifet PPOA found <ppfc) x [100% f total aolida(%t) r fi7 W7 Pag* 36 o f t 5 Exygen Research Page 99 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 E iygtti Protocol Number: P0001131 ANALYTICAL METHOD M ated Number V00017 M u M TAM y v h fe r DaeermUadac r fF ir f t u n itiM dh A dd (fF O A ) 1 FUb M d CU-- by L O M tM S A u re a l T m t f F irjh ty: F t } f Jl -- b JQSt M m uch Drive State C o llie . PA 15#01 Approved iy yi-i. f - i l . Fed Oaecfilly I Tetteica} Lmta. LC-MS, Bxypa It--rii Aaia?/fe--qf ' Vet Dam *A A Exygen Research Total Pifea: P age 3 7 o f 65 Page 100 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: POOO1131 B x y je o P rotocol Num ber: P000113) hffM lM M W k c________ AN ALYTIC AL M ITH O O VW ITC .M odtodofA a a lyili f k k t Patanaioalkao^yarftaatooccanoic Ackl fPPQA) in H ahw T Clara by LC/MS/M3 1.0 Stop Tbia roatfcad l i to ka aptoyad far tba iacUboo and qunritotian a f parilueraocuMiaie M id by HSjb Pwfensanca U pud Ck omk Pgit pby couplad 1 tandtm M m Sprabm otrk D a to ** (U A I& M S ) in fidh and ctona. 2-0 Sbty 2 t A Jw yi i t i n w A laboratory practica* 2.2 OhmuII <to appropriai MSDS bate* handling any chacnical far prop salaiy S-- pi ia ^a in ia --l 3.1 At iM flt 20 f o ftoat I-- pl> Ibr nuraoian. 3 J Sm ph* IM U ba prW aaMd to * * xtracton. Plaaa tha t e u i aampla in a fcad paooaaaorand baiM |totsa till d iy mb. Mibb ka aaraplaa ui oonuiM n m i torra pan k k m a to n ft n w d ib t to *lkm fa - m b m dioxida utimatkfe I m I and plao tto aaaapiaa k k a tm toaapa until lima o f -- bka. 1 3 Iw p lt rcPartito procaduraa wiU to mariH ad la k * anplkg plan ter t k i 4,0 JkagMto tod Itoutaida 4.1 Watv-HPLCsrafe 4 J A w to o itrik -K P lC proda 4 J CmAct. (120 400n a to j - jUagart grad* U M ktow d-H FLC gioto A3 dliai pri (49-200 mach) -la a g ra t grada 4.4 M a tt <40lM naah)-lLaataat rad 4.7 dapaaclaaa LC-NHj - JUapmi pad 4 J ]-Oetan#l - KPLC pod 4.9 LAwwka arid - taaganl grada 410 D tokyldithkroailtoa - Rtogaot pada A ll T c k a o a -H a ifly d a 4.12 A w h t o Aeanaa-A . J. ta a |a to O fd a 4.11 Pargnamrrtoaraa A rid - Stpm^AMrich 3L0 A M |h partorcaano Ikidd ebramatograpfa capaM o f pumping up to 2 aolvMHf afuippod M l a r ar iaMa vote* kjaatac aapabk o f injecting 5-20o jiL eenaaetedla a tend Msea Speettoeto* (LCZMiyWS). 5.2 A Javkato oollaat tow data fcrpadh imay r t aai tod qaanntBiu. 1 J A M l)tieal bakncacapapk o f raadinp to 0.00001 g. n $ tJ ofI P n g r Jd o f i Exygen Research Page 101 o f 132 Interim Report # 10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 B xyg ra P rotocol N u o to : P0001131 ________________ANALYTICAL M ITTIO D _________________ S d artw To f Aurfywa fcr tbo 5 5 5 5 5 5 5 o fPwOwcocqUtwic Acid fPFQA) in Fii Clm abyLC/VtVM a 14 RaWy lu rn iu a i. 15 T lM M to r. 54 U m L m m upt A a. 5.7 SOmL polypropyWa w m N y tabu 54 15 mL diy a b lo p o l)p wpyiana w a iftfa boa 5.9 OupoMbU m icro*l*au (50 tOOuL. IQO-ttOaU 110 lHeLlDPB*mewmwdibotke. 111 S n L to H P L C v M Id t. jjy IM y ot~ia pij^ f 113 AumH mU (ICXMOOO pLaod 10-100 pL), wWi diapoaabl* Hpo. 114 O B Mb (30U {li*o ie o MI. no. N037I7T). 5.(5 WrM acttoa A M *. 3.16 CbBtetl# c ip h o fa pn oiftj 50 m L Qolyyfcpyt--n m b at 2000 qwn. J 6.0 O nm itognpU c SyvMni $.1 Analytici l C olm a: Floophw I f dUyMsot Sckm Ukk 2.1 pm x 50 mm, h> <WN: 42505^32130) 62 7** * * * * Y fC 1t J MMoobMilfiethPetamitC(BA)):! 2 m U A n e m Am m m Wu k Uemoi 44 C M d im fn p m : Plow SUn % A * 1 fmL/minS ac a 35 1.3 1C 65 35 U 10 23 75 14 204 25 75 44 324 65 35 44 64 ktiocboa V oiunc 13 L ( bo loaaaaad to much 50 L>. (.7 Q u a h liiu : h a k Ana - o u tm l tfaadord caUbration curve. 64 1 m T im -* 23 ounuaaa, Tha abowtconditiooa in Iw im rltl a fuido and may bo cbanfod <n arder to oMimlCT <boHPLC ayataaoL *o*JM Page 39 o f 65 Exygen Research Page 102 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygen P ro to co l N u m b e r P 000113I Mplk jii iif A lin ii far A N A LY T IC A l METHOD t t t m t M m a m M AcM ffT O A ) in Fil> ml O m n^rlC IW M M TO M&MS Sw an 7.J Mod: E lM o ^ n jr Nogaavo MKM mod monitoring 41J - 3* n#2 for FPOA.' H e ib *v * oondUeoi an tornado* u a pinto lad m y b* ttoanged in attar to wpUmiM* MSMS tymmn. 1.0 wC toM am i . i m o b ib n M 1.1.1 J mM M tu M Mlato to water to pnpud by adding OIM g o f -- imam w a H lO OOwLofwn. U lx tre o * Scfc&a 944 2 % *e *k e o d m d m o |ie iw i|ie n d b y d ie w lv w g 2 | f ie o A k rid to lO O m Lafam thtto] 112 30H D im totutdkU araiU o' to ioIm m to pt t f m d by bringing 3 m i o fdtomtogAlkUMMitoBC 1 A w l v d im e flO mL with toluene. AJtom tovohupeimey btpnpvad. M tapdariFtopm tofan 04 ttoartord H octofortifiedio B d le t #4.1 9.1J 9 4 .) t .lA 944 P iW ** to o * ohUkM d-100 tgfaL o f FFOA by weighing 10 mg o f ibdytfeaJ jfamdacd (ooeetad tor purity! u d dihit to 100 mL with meUtoaol to I2ShbL LO F t hotel*. A L Q n fm L ftriiJkoboa eoftattoa o f FFOA to pe+ *n4 by bringing * m Loftoe 100 ptomLeobtoM to e IbeJ volume o f 100 w ito m m o i toe 12) aiLLDFE battle. A M p ffe iL fcctillceiioa eolvito e f PFOA to prepend by bringing la m LofitM 1A eolation to * in to volume o f 100 w it* methanol in 125 o L U M t o * A 0.01 * !. Ibm flreltoa eohrtfan o f PFOA to prepared by briogiag 10 piL o f the O l pgfahL eolvttod to * Sod volume o f 100 with m etm elto 135 mLLDPB beetle. 71 Mek n d tortUfeeiloa rotettonrereto b e rto rid ijie re M g fre ie ra i toyvnrlmotoly 4*C m i ora otobto tor * maxJamre period o f 6 kmd 6om dm dote e fy ij ueeHaw- fm 4 ori 40 o f6J Exygen Research Page 103 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E xyfe a P rotocol N u m be r P0001131 /" CltOM by LC/M l/M S OA) in Pish d 91 mudare Q ilM o n SoM u 92, i LC/U&M 3 o Hhw tioq rtantarth tav pnpew d in aw thnol via hiiior mim I 4 p ^aiL fertJOeiUN otvtion. 9 J2 Th Mtowim typical auata: taditau] eoocMimboa may he ynpend h q n iM A ia te 14 V ib e JBIL . 14 ..a y . ito 0.05 liJ 14 am 9M> u 14 L0 II no too too loo 0425 0.01 0405 04035 l i M in 0401 to J 00 J S L T i) Stars Il oolibralioo ni Il ud i I LSaoL Lp m aaiTos-rnomb bollico al 1*C lo t r e . a* n A m a i. > A la rn e uslaraaa aal M taM ttnfioai of atandarda may bo prtpared sa !M r a k ta l^ 1& ] EaM b a d i a i la w p lil aaeaeM (tyfeaUy 30 or h a i) amai Includa Uaai a ia M e n n i e ro i a d n o e ia i l a n ta h n U M b a w i me n u Mon (lab ooaaal ebb i) e a rifr pwoodotal tsoovsry Sm a biadi. 1012 K afiainesM fc r in a a d labaiie ry l a 'ia a n d a ita i HI ba specified e De aualily iw e e e i b ln I * ia im lie L I L I 7 '* jti I | o f fio s s i n b baa 5C s L polypropylene m uriAipa tabu (Ibldiy sa eaeda4 ripeso Udani Mix w all) . 1.: iV-a 30 od. ofaanan itaaod a n ta eo tw ila i alias ababaa tor - 1 1 itim ela i l J r b n du stasa la a Asolar fcr - l boar. H A acfe n d oasdM en (ba SP8 ebae a d d la a la e tb a poasabapod fls e k i. I ! . ) ? * k s 30 oiL SPB aibaa ta aaqnaacc aridi 3 f D onali 2 9 lisa M l. 3 d tacbon ao 1 3 LC-NH). Candjlioo II Mlaaiaa s in 20 mL o f RioOsaol. boa 20 aC o f aaaaonMii. D in e d all vaabaa. Do noi lllo a . dia colami, *> aay. I l i lib a iM *h 13) sL f . e -i bl|i i l Seats by liaioa w ith dia J0H daistbyltiebt irr siin e ia (olussa solatio*. Uosa tba flask w idi lolaans u n , M e m d by it d im oi a trio fin a l), Dry d e naaka oootaldely balbrt uu. otta by aii>drylo| or id i a m a i o f e e o p n . P a g e 4 ! o f 6J Exygen Research Page 104 of 132 Interini Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 E x y p n P ro to co l N u m b e r POOOU31 I a n a ly tic a l m jc t h o p K o flw dofA niifa w forth Prtn u nfaoo cfparfhnooctoiioi Acid (fP O A ) in F i* wet Ctoou by LC/MS/MS *11.1 C n k lfa ff tto J n l p a l)fcapylM b ln (w u m m ^ m npto at -2000 /pm far -10 m inim , 1.I Xtoctal Uw a a c ln to coaditawd SFB ca la o * dttad iu id fa mouth o f fa t pcirHfaapad Ita * Coltocs It* ataato m & 129 m L iln r t pcrupa H i*. li. AM 11 a L W w M lib * a a p la i . Us 3 mL o m tiiA ijt III . . , Ha-- . -- llb ip iiO ittI m iriw m i . m r - M itaa i i l iicdnint dwdinmiawwfth-IOmLofiMiQBMli fato tiwtoba. 11-10 Shakefa a a a * d t f to r-1 9 minuto* o a t w il -K tia j faakar 11.11 n o tto n a to o b ifiu o o fc r-lta m rm o o . 1142 C n ai foy fa 50 m L poty f 14111-- tub-- o o w tw n f m p lt ft -2000 rptn 1143 Pacmd t m tn m a * Mm u r n i n tofana. Coltoci Uu olu*c in o ihr maw paw otoopod flw k n d cambia w ith *0 d a m Hom Hi initial U J d h a 20 mL o faaataastril Ocsufb to r IfE caluma n d cooU m lha hut hi to im a w p m is a fa ^ lto * IU J Add 3-4 (kopi o f l -cotonai too e rta c i fa to p a v -rta p tf fla k md avapamto w taduoad f r amm adag lo m y m a w * {m < 40*Cx I L I ! Maim fa fia ti a to m , by addin* 2 mL a f 2% wombw acid in atitotnnl to f a f tow id111* m d rM tl 10mfa/diootv. IU T TV-- dmfa i w a r n W HFLC fato m ie#dfapon b to p ip o * IU A m iyw tm m p * g u jd m tr o ^n c y LC/MS/MS. U.0 Cvnawmpwpay 124 fafaw <Wm m m ourn o f c a * m o to ri. imapto n d JtotUUd m apk m fa* IC M A Q I f a a A oMtomwa u d ir i oaul pucada n d torto* * Hulyaod wnptoo I t i ll * n li s f PAQA u Imi, ftv t or m ari ctwt^rirtC joa t r e li m b* iw bdM an a u lirte li act I t ] A . m m M o f calibralo# ataadank aauatba iiu lu M d ti it te p u n in j aad . a a l o f a caa^a Ml. 1lw i . l t m*M b . inm t-- tad bttw tan w r y SIC A i i l atin tUva. m w in M o f calibrali m M i may b* lai*** 0. b a p n f T 1 M MUowad by ca lib ra li*. ctandardi k ta n p a rtri ivo ry 5-10 i i r i j l . (ao a a a m l y c a a cm w f a a n flirti). I. M i cm , u A n . a lm M t mu M Ik * fin i and Iw iujcclw u in a m m ybw. I t i U n im t -- H im c a rn i M r w a ta m l i n a n d a ri c m an f i n m a d Mr dm analy. by llaaar reim ita wabi# li* t iib titj o f paak ana va ria i librarti ataw kri a-- Him aablf M m aLyn ) .) (or aviva i ) a rih ra m y n u . N m v .fi P a g t 42 o f61 Exygen Research Page 105 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Eayfen Protocol Number P00011JI McdadNaabctVCOOira I~ a n a l y t ic a l m it h o b M td a J a fAaaiyale 5 S tT S ttw S S S efrarfluotoacoftoic Acid (FFOA) ic Fufe and Clama by LC /W l/M t 119 lem |dt r y doaild oh e n d tornim i materna Any lam pi dui t u r n i to n im i m ot t a abatid bc lm iur dibuad tnd taanaiyrtd I I Aacaptaaaa Criban l . ' i Oib ii-- n yr a -- l ahaw i j m k t i i dc c jlif Iqu a ) t i i bw paran t i 41] a u . Ib a I I I r a pocm mnm otcdi *> <ba PfOA k k . K it ttc dmuhitr taaQd aaa) in m aa n ba lo n o f aataa diwuOt. bfatbad Meato wat aot acatar FPOA O Id v tli tretm r thac d LOQ. lf 4 bbak aaamiK FPQA at lvele pealar k i O] ppb, dan a aaw blanb aanpla -- n be obaiaad K d Iba agua Id mud ba ia K u ai la. 1JJ Raaovattat g d a c o o liflk K in d M o l to<lw O K I babatw iM r * - l] 0H o f dairkm xm tablee. Ifta a c a v l v ito M e autdatta eoccpubla Umita. Uw aadn tal afam ada atould b* m-aameaad. 13A Acy a U k u ta aricad fcuad u W a tm iamal u lia r by uaini iba H u * Em r Tael, may ba ciudad Aaa da oaku Utiao o f <kc calibruiua curve. Hattamr, Iba M al t ambar / calibrado K a riid i dial cnaid ba dudad a a a o * otear 20* a fd u m al m bar a f andada adecuad. IJ J Tba sartaWaa coeAlotaat (X ) fbr M itra n cwvm poanaad aaa bc O.992 (X1 bO Jdl). W aadbndaa maula M aauida h a Im ita. Iban tppm prfn M fc a a l ba a ln a adjuai la ia m in i apctition, tad iba Kadm di c ra c rcrraat m a f ic a p to i uaald ba lanalyaad. 13 6 KataodM taaa baramtc o m u i t K d eerapht o ta oat d rill mora iban 1 4 * ndda m acabtal m lf rattatlaa data d rill acocada Hila liar abbin k alyiical ma Una da a ta n o r ba aoulyacd. 14.1 Uaana to lira O f t|u a (a a n tataulaedaaaaam a f Pf o a twndO a baaad ac raab anu mua ha ito u la d a m a (lo ca r ram oneo paw iaien) a to a r by dta btma Lyox aoftoan p te y a : FPQA Ibaod l u / u L I IFaat ama - `-*--11- ) 142 \fte ito M aialai taMIioi le om eri da anear efFFOAfouad la n ita l. 'IW H rTO A feim dC K bl-ra g n a to u a d fa rn m n .Iir^ lv ^ ^ lm L ), DU m o la m im en DP-H a la rby Alab ito Jlaai buna wwdibUeA ifoacaaaaiy P a g t 43 a f 63 Exygen Research Page 106 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygcn Study No.: P000I131 Bxygon Protocol N u m b e r POOOl 131 !) -f> . va o o i'r, I " ' ` " a k a lv h c u . . rru o p M e ttio ilf A ol^i4 fof i c D iM rtilntScooiPirauw roocungic M u l (TFOa j J f im m l C bm ibyLO M V M S pmmJ O N t m l f a t l M w i* k a m a w i o f PPOA prior nineno*. U l Iba fcUowiD*ouboo Iw ltu (ho [ m u -l^ fa a u K o g lp u ly a m U M O o o lw n ] Exygen Research P o st 44 o f6 } Page 107 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygcn Study No. : P0001131 Bxyica Protoool Num ber: P0001 131 ANALYTICAL METHOD Madwrf Number V00M7S4 Mvtfiad r f AaaiytU U r A f l li li r i l M l lo i o f N rC u c a cct iiB lt AeM (P PDA) la VegNedai byLC/biS/Ml Amytioa) Teefeg Paoility: fix jfa i R atteret 3031 lU w m b D rx Sum Colley* PA 10101 Auhw o4 By. j5r=ik-- C^ALi----- PaUCouuJy 1 TacJuical L<*W . LC-MJ, B k y ftt Iw a n k .-iO jU lH Om ,rftn Flaherty Vice PreUtUrt. Operato, Dry* fcaaeaich Date Exygen Research Toul Fay*: 7 P ag* 45 o f 65 Page 108 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 E n y fe a P rotocol N um ber: P0001131 I ANALTICA!- MTBOP Molw J a fAnolytit fcc Ito D aw iniaoik ti cfN afljoaooam olc Add (P ro a ) i Vi by LC/MS/VS T t'i i i -*! ` I m di by ltb Upfe I p m riu m i P a n te r (LC /M axS ) i U lm , I l A tayi oboarva oadbIfeoaanry piauteo. 3.3 Cm wb * dpnyiita UTOS bodon In k U ru nydatatati far ynpm itftay y o toM fW niunM < 31 Al ta < 3 0 | (tMIMMvX * f PtoMtlo. 3 3 fciy b i hokd bi pwf m i l bafcro c an d o Plan dw N m templa ia Uto pomoar m i hin ip idai w ikdrrlaa Plato Dw tony1 I omoinan m i Imn opn i t m u i f ovare** *> o fo t atea dkaidt wblfeiifei Sao) m i phea dio mtplat lo bom a o nuil ihm of ataban13 l eo o li aoOatliao paooodino 411 bo poclHad k Iba oaoplm* p o fr tb it ta ita 40 a fe a n todlitodood 41 W ta n -ilK C p id t 43 AcokaomU-HTLCtraJ 43 Coat (130-400 n in fa )-d is ip a d a 44 H c a n a l - K7LC poda 43 Siliaa yol0-200 a a a ^-H o a d ta p n fc 44 HotiaB(0-l00 o o a o b )-d o a a a ta 47 Sipaaclaa IC-NHi - afean pida 41 l-Ootaoel.HnCpadt 4 UAacorticadd-dafeanped 410 DtotdiyblirMMaian - tajan pida 411 Tabana - aa*Mt podo 413 I mommio >nOai-ACA, tild a n Onda 41) Nr....... ruada Aak-MpaaAtbfct 5.0 5.1 A Uffa parlbnaaaoa H p k ah-- top fe h c ip b b o f pumpn* p o i Botana OQOidd td M o nrlobla rotuna infense a p 4 U oI kjacun* 5-200 bLooantoaed lo a ln d in Mmo i y im m o li (L 040/M 3j. 5 ) Advlooaoaollaotrawtaaferpaakiaaaplioaaadquaabtaiioo. 5 ) A nal)Ikal bofetea n a 4 b k fn o d ia |to 0.00001 * P agt 46 o f 65 Exygen Research Page 109 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Bxyfco Protocol Number: P0OO1131 ____________ A fU L im C A i MTgBOD M edi o f Aagiyrt K r im n tfim a n tica i o fPrfluojooctancic Acid fPFO Ajin Vageianon by LC/M3/M3 14 IL tfvy e n p a n ttr. 5.5 125 iL i w ft*. 5.4 50 iL d i^o --bic polyproyylme trm iriR nr tobe 5.7 13 i L f liy n ahkpofrpfofyfriw oaogi& po tubo. 5. - f if a t i (5<M00uI* KKWOOuLX 5.5 125-mL L D tt ntinw-twouih butin*. 5.10 2 anLol--n KPLC vial kit. 5.11 Dlipeaabl* 5.12 A m ip frd -- f l 00-1000 fLanlQ .100 fL ). whfa dkpoaaW Upa. 1.11 IfB w M iO O m L) (S eH esm t W.N057177). S.i4 W m tictiae attic--, 5.13 C-- riflifa t a bl i o f ofMafag 50 n L fd ypropykna 13000 ipm 40 CariajjUpafhio l y t -- 6.1 Analytical C oiem : PloopMae t ? (Kaytlooe SetwbflcX I-1 mm * 50 mm. Syi C M fct2505-052130) 42 Twnf-- um: 30% 43 MeW*fbnan<A): 1 nM Amoenium Am MI h Wcmt M MobUnPtamtB): Madunol 41 O ndim i Prapran; XIb r Mr) 00 1.5 1.0 200 .5 S tA 45 45 35 25 45 P to* Rata taiL/m m l 35 0.3 35 0.3 75 0.3 73 03 35 0.3 46 Mfacftaa Vo(m m : 15 pL (o n bn fcnnaaad tona much an SOpL). 47 Q u m tita te Patir A m a-arta-- I atia d -d natiWaboa mrva. 4 t Ran Tim e ~ 2 3 ia tia -- Tba a b ra eonditiac* an intandad a i guidi md nay U cbanpad in ordnt la ofimina ih HFLC lytaw, 7.0 M 3/M 9Sytia 7.1 Mod B lactioticiyN etactvt MOM B O tm ordm lfit *1 3 - 3 4 9 n ft ftv PFQA. Pa$t 47 o f6 i Exygen Research Page 110 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygea P rotocol N u m b e r POOOl 131 Matotd Nunter V0OITM AN ALYTIC AL XETHO P Malhol e / Avajyi ter Um Oetemiiaoon a fPrfiw nooctw ie Acid (PFOA) r. Vernation by LGMS/MS Tb* obon M o te tte n Imanted a* a |ei4 *M d n u yb o Ju n f*d ln ordarto ptmda* fcoMKMS ayvten. .9 Pw fira ti-o /S ffa io M 1.1 Mobil*Ttnc I.M 2 m H M uxw diin ocoom in wm I prepared by ateU* 0.154 | o f tm m m m m e te lOOO mL U w*ir. U labeeten Solution 1 .2.1 2% aaeoafateM id in BwOMUtol i* prtpved by diaaolvinc 2 1 o f acortxc and in 100 o L e iM kasoL 1.32 30% DinatkjIdteM om tilteto in totem * m pny ed ly brinjpnf ) mL ofdteM teytdkfctofniteavtoaffBaJ ve lw ite e f )9 ic L wifeuluO Tt. A to m * voltnn** te*y b* pnpamd. 9.0 S iM to r fte fm te 9.1 SMdMdSttKtoTrtlActktt Motten 9.1.1 Prepare t M k aotattoaof-100 ^ fta L tfP P O A by watching 10 tu* f analytical ttn daU (oormctod I m jw rily ) and dilute tc 100 mL wnh mtihanal te t IXSenL LD fB bettle. 9.1.2 A 1.9 tertofcmtoa whdfen a flW M f f * ad by briefing 1 L * f te* 110 lt# to L Boteaten to e fin d volume o f 100 walla m*ot tea LU biLLO PBboote. 9.1.9 A (XI pgtoL tertaHrtoinn aotedoo o f FFQAiapwpared by to in p n i 10 m Lefdw 1.0 p jM L aehiboo m ih ilw lim * a f 100 rite atitoaal in t 25 toLLD Fe boote. 9.1*4 A 0.01 | i | U fMdficattee lohnte o i PFOA te p p in < by bnftgwg 10 ML f te* 0.1 uteteL aoludon to final votant* o f 100 wie* eetenm tea 12$ mL L D H bete*. 9.1.1 Tteatof& M dfcrtU katiaaaoittttaaiB ratob* taam dmaraAtfacteor at appaoaiMBtety 4"C and are nabl* ter m axinun period o f I monte io n te* datomipreparation. 92 StandardCabtetoteai Solution 92.1 LOMS/M8 ctefbradon tendante ae |--f n ail te piatiuool via dilution a lte * 1.0 pfbeL fertiAcatioe toiutfcn. lift * m' P tlft U Ofti Exygen Researoh Page 111 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygen P rotocol Num ber. POO1131 I M ito d f ""a NajL Y I U A L MIlIU O D 1 & tta D ^ M k a lio n ofP arfoonw cteok Add (PPOA) in V*jm i"cm byLC /M S/M J 1Z.Z 71 M tow kg le i TypkeI w m p lt: additional eaneetxmicMu nuy be peiMMd a s M id ii .o--tk e e *TPjrtiJfat**B VtkflM OUuMdM f --I Cm m Mo* *-- l. f 1.9 0.M ^ T 2.5 1.0 10 -- a*-- 100 too 100 QS OjIIU 0.01 0.003 0.021 10 100 0.0023 f . l to too am l 0.003 w 100 ____ _____ 9*b* a caJW uM lMdkde k 125-aiL ID P B narrow-moedi boni at 2*C I S*Q KU i l l moatba 9 X 4 AfcUP m h-- m d u n nratradaoa a f m ndardi nuy bo pteperrd aa ICC fit*i l* if 10.1 Kwb boMb o f w tp lia named (typically 20 or leaa) mtiat include least 4M m ^TTlr* aokroJ and Mo uclmeed oonuolt fortified at known eonoM natiw fleheenbel apik)to ra rity procedural recovery for beech 10J K f d m m for field Md labariaary duplkecee end spikea w ill be specified imin i quality aaarato* plan fee h i t proje t 11.1 TCalafc 3 g e f to a m aatnpU kao 50 mL polyprapyieor centrifuge tubes (fo rtify aaseeded, Nplece Ud and n k well). 11.2 Add 39 m!. o f weudirile aad kaioa oa a wrist aotioa abetter Ibr -15 minute* i).3 C*4rtfega (fee 90 mL potyprosylMr bee eoauiabii aaraefc at ^2000 onn fbr^dan eaaa. .4 ? a c k m k o M ilfe a M J n b fe a flM d c ilM iu the pcerebaped fluke. 11J Peak the 20 mL f iJ t tobea k mqMtice wbb 2 g flarm l. 2 g attics ( l U | eben. Md 1 | LC-NHi Coediaia tbe ro ll--ma with 20 aiL o f methanol na 20 b L o f aoetootfttUa. Dteovd a ll weaboa. Do noi allow the column ia ** I U 9Um Im daa 12S mL geeMlwped 0 u k t by nnaing wirb II JO* dfaadbyWiehlaroaUaaw k tokMsesluCfee. Rinse (be lleek whb toluene one*, followed by nwettanel (fibre lim a ). Dry (ho fleeke ecrapUeriy before uee, either by rir-dkyieg or with t atream e f mkofeiv. 11.7 Deem foe extmet a to meoaditjaaed SFB eoUuaa teed inside Ike mouth o f the ji war akyeil Beak. Collect lbe skate in the 122 mL sHaoiaed pear-elmpe (M e. P a g * 4 9 c f S Exygen Research Page 112 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 B x y tra P rotocol N u m be r P0001131 U M hw n vuoci7u 1 a w t i<.m . m it m >d 1 Aualt for (ha O M m Um iM f FarOuorooaucu AmJ ( W A i n V ,,n u M n b jrL Q M M a 1 1 .1 - AU4aO .L<rfM Uni>Tm a -- p l. i. fr u ttin i. fra n to . I 1J H u be* w upleieslu fcr-1 6 eieuN ioee w ritir# w e * l r. l i t i - Domai Uh tu e t OBI dH t n SPB cO lun. Coltaci (la a iu itt into die \ N M p r chapad t m k ad Bambina wfch the luam to rn dH inJil 11.12 IHM IM toroogb U .I1 M a ilt t l . l l AM 3-4 topa mt l-oUnto to Iba o h m ia toa p a r topad flask and aw poroltm ratoiead pww m uaiaj a saury tvspocasof (at < 40*0- 11.14 Mato toa flto votola , by addisi 2 mL o f 2% aacortlc acid in matoanoi toa pomtomad doto mA mHrtto mhMisaolv. 11.12 TVmufa to tocto lo HFLC vala m p topaaabU pipeto 11.14 AttJya lamplia itondilntoimny UVMl^Vtf 12.0 Cbrafaatofnpby 12.1 bda * a mmoamaeto a f * otowlaidL aeeple e d tortifla d ampie imo ih* LCto<S/MI Qffima. A UbmHan standard imito prosada and IWlow all m a lya il tompha ; a . fisnim ds a fffO osempeaiing to to boto (Ivo cr m ari aantonaraiian lavala amai ha auJitdad saaa anstytab ac ' 1*3 An to t ato o f bacato e s ita to Maliarda ma be tocludto ito b *ta a *g d al toa aai o f a ampia at. Extractad an todi musi b* la m p a n a i btovmm rm ry 9-10 -- U. Aa m attarudv, m tira m or Mamad fsltaH lna standard* ay to Iniootod Ow bepto ii i o f a ai fottmmd by to rta i catiteada itandaids Intorapcnid tv try 9*10 Mmplm (toaoccunt tor a aaacmd ao f Pacaci standard). la etOwr ta n . espaciad ca lta a tm atotomds mutoto toa fila i sai lato oacto io a caspia m 12-4 Uaa knaar atom i marna far quaatiUdori. llm if standard ewvsa a * pmiratod t ir to t M ly itiy (tosarrafraaaiooustai Irti m a i|)c iii| afpaak ma t o Uhcartoci candar canemeslod utong M aaaLyu 3.3 (oc equivalenti to sse sytoam . 12J In o p i* ri f l ifeeuld no* asad toamlato mapeoaae. Aay samptoa tluu aaad toandmd mapeaase abaald ba Mtoar dthitod and reanalyaed. 130 Aoocpunoe Otaria 13.1 O mm atoy onto barn a paab mi a dwfhtoc ioo at 340 mnu from a panai o f 413 uno* Tba 413 m p a ra to cmiw p uudi a Iba P70A io n . wtoila ito daughter ito <349 amu) wpwaala Iba Um a fardendioxid* P a i* SO / 65 Exygen Research Page 113 of 132 Interim Report ^10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygen Protocol N um ber POOOI13 1 1 a n iv t ic a i te rr a o p I M atyo f Anelyai ftr& eD ainniw dqoB fP iriW aeoctaiaia A o id(P fO A )ia Vegetation U jL O M IA O 11.1 Meda*M antaataatieiaaaainPPOA alenali r e a * dtae 4 UX}. Ila M ak osanna PPQA a lavata ulta duo 03 ppb, t t a i m tieni ampia n u l b# obum U m (b Prtlnr m m t\m iv-vilnHctad. I1J Hmov * t wafcol t^ikm m m U >f& mtM to b*tcn 70-1W% twir kMwa vaimi, If a n a n i afilo bili outaida Mocpuble Umili, th Aut rf Awid ba iHidnMd. 13.4 Aaj a lite a k a modale Itnaat io ba > aatloieel o u il by eaint ita Hugt Boar Ttnt, aey ba adedad te a la gakulatNn o f Ita calibntion o e u Itow ear. la M i M anta a f calibraeion lanHirda M noeti be eoludad aael M m e t 10H a/Ua M al aaabera(nuakM ti iqaaiae. 13.5 firn eanlarioa iaaffic ieee (K | kti cebhveOoa aurea paneretad ohm! ta OPPI ( l ' M lt lt ll lf colIta lia re a ** Ik ll o n d a ikdad lim o . Orni tpKUriaa a * n anet ta tana a ed|wt ta n a n a i opemion. ani d m a M a a Ha M im a i M ofaaapla ibouM ba leeaatyaee. 134 P aM ieo tma baaaat a l e a end a m p ia m uti eoi O rili aure am 4 % riifeia ea aelTboal m I l m a rin a rima del* ete a a d e a ie lin ii ritMn a aalydeal M ia ite M B u tb a remaiyrad 14.0 14.1 la i Iba b lW f genian a t all lriaM ha m ra la f PTOA bum! (ia n^eiL, ila r i aa pmk m a i a la ) ita a it a i a rra ( Ib w nom ai] p a a ia ia n i yaaM ed by ita Meet Lina eodtoare p ro z ia : PPC3A kamd Inata LI - IPanh area - rinarrarti elope 14J Uae la Mleapinpedattaa a eoaveet d a moan o f PPQA tbund a opimL in M ttP b ). PPOAaad(pyri)- D f - k m by Mck ta Had eetaaa m a d ila ta , if m enavy. 14J P a arepla M i e d ridi M a v aaiunw aI PfOA feiar a actration, m d a fcOoerind gaarino a aalaalea d a p a n a i m oney. eaovayiX) [ a a laalylafeaadU M l) - enalyaltliiidaoanO nl In o lili eailya added (n*H) Pag* i l o f i i Exygcn Research Page 114 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygan Protocol Number POOO1131 ANALYTICAL METHOD MtftodNiaitar VQOOPIJ AMJpbfartoft'UmUuulMoINrflMroMtMofc Actt(rPOA)Ja liaaJI Mm w I U n r fryLC/MS/MI AMlytlMl Tadof P^aUty: Batyiao RaMirab 3W HHw rdi D ** StacC<rfk#p, PA 16WI A ffe o v ^B y ; C -JiL ___ Pul CcnwUy I Tcd&ucal U m ltr, LC4Q, B xypn g m infc o //n d - / / ________ ' VicF red a * O p n k m . tx y tm i M w t Dot -* A A Dew Exygen Research TW1PlgM: 7 Paz* S2 o f dJ Page 115 of 132 Interini Report # 10-Analysis o f Fish and Clam Samples Exygcn Study No.: P0001131 E xygcn P ro to co l N um ber: POOO1131 C O fto w MatMC NUMHT VttOQi>15 ! A A .*l.y n C A L M T tt0 8 Vaikid^AjaIy>l3fodM&MDMiMiatKiJlu<nQcaaaKA<U(VroA) nSmt M m > i Lhm by H a v a n a I 1. Supe ~ i-t T I-V 1 il l i III n iln in J fu (til I ililr n i1 i ninna in iT| n r ......... n n arid by H I* M r -- n liq u id O tw lu g ie pfay ww qiU to a tandem M au Speesrtraie Detectar(LC/M8/M 8) in h m JBcant] liver, 20 fr * y 11 hraj ebtmve ta it laboratory practice. 2 J Conralttita *prapri-- fa tfM befara h ittin g y chemical fee proper u ftly 3j0 Sample Requirente^ 11 A t Ira rl 5 gefteetw tap ii b r o W tim 12 taephe abraU be procu rad before M elici. Ptaee the frasea imnple in a bed prooeeeor wA heraepra rtc w ith fry lo t Place Ifaeemplee in conminen a d lerae e p a fai b a a atara* evaraigbt I allow for carhop dwania b itta taira Seal a d piece foe empiee fai fre o n i orape until time of aalyfoe. A b m airiy, I f th a t ia m tra u lld ra t anorat o f ample Mece than 3 g). then no peooraelag it M reewry a d the am ple can beuaed aasupplied 3.3 Seraph ealotaira proendaree o rili be pacified in the eeeqpliof pira for this pact 4jS fteagrata rad tendente 41 Wera--HFLCjede 4 3 Malfammi- KPLC pada 4 3 AcetonitrileKPLC grade 4 4 Aametaeet Acetate- .CS. Ksafrat Orafo A S N ritom ootraaie Add - SifpnakAMKefa 3.1 A k*4> perforaran | * ld ebraraetagrapk eapebfo o f pumping up M 2 cahoraa equipped e rf* a variable vallane fapeotor eapable o f injecUny 5-20(1 j i t r erarta ri m irara Mem fry i i n f ie i tC/M M Q . 3-2 A device teoelhel (tardata for paifcfotagralJco and qrandtatten 3.3 Analytical balancepablaofnadtag le Q.O001 g. S A S idL ditporabh polyprapyfcm orairifogr tab. 5 3 13 mL tapciab lt pcly y opyhne oratrifrige tubes. 3 6 Duperabh em pipen (50-lOOnL. lOCMOOnLl 37 125mLLDPCn nm etetah botile*. 3.1 3 mL leer HPIC v iri k it Exygcn Research Pago S3 o f65 Page 116 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exyjcn Protocol Number: POOOl 131 Bxypai feMKk Ui i i i iUMUiVWinil I ' AN ALYTIC AL U lT M O P M M rfA a > l) i< lb rifa > D < u a n lu ttiia rM li> in o c tu iH c AcM (7FOA) in Small Mh m I lim b r LC /M V M i I ' 110 A M tppt e (1CD-10O0 itf. and 10-100 uL). w tt tV p te ic lip t 5.11 W aM raSapfekViri t ( l|) C J llr t c a ilr i4 p 5.11 jU v a c t e t e f t ld 5.1) T liM iiH r . 5.14 W tiai-oailoo te a r . 5.15 C tO rtth fi w yfch o f q ifw iag 15 a t po)ycopy1oM iitlo s i 3000 rym. 4.0 Q m m f p j q k k B ya te 41 AootyVol CaOraw fe wfihm R I (K^ytano S cigrtjflc), 1.) mm > }Q mm. 3^ (T ^ tW O W n iW ) 42 Im + m m m - WC 4 ) M otel* F t e (A ): 2 a M Anoncrtiima Am M i in Water 4 4 Mobile fea<B>: M ateo ! 45 Ck*4tni ?vofnBiu m Ik t i) ao 1.0 40 340 22.5 ZA 45 43 15 25 43 FlowXMo HB 35 0.3 35 4 ) 75 43 75 43 35 43 44 J o i c ti iV o te : 15 |4 .(c w V is e te d in atonch aa 50L}. 47 t ^ if iiit h w feafc Anat - ostanaJ tfn d o rt calrtnccn curve 33 Hun Ite r . - 23 wi t iiHa. T V aV*aM d0fcM art banded u i pride m i rnojrV dumped in order to o p h ite to HPIC ayteeo. 7.0 M IO A V 7.1 14>4c BUdMOorayN a ftftv rM R lf t e c , noortoriot * U -e 340 n i l for rfO A . I V above oooditircti in bunded at t p rid i and racy bo chanted in order to p a te tv msmb n te t Exygen Research /to ff 54 o /65 Page 117 o f 132 interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygeo Protocol Number: P0001131 | MCTROO_______________ _ MKKad o i Amiyte far flw Paanuialkw e fVwfhofoocKmok A rid (9FA) i'n Souli ~ Mw u m I U w by IO M S/M S lo m p itdoe ofS oto tea i i u& urm 1 U 3 m H -- w h n m m In w tt la puparad by addtaj 0 t54 t o f P IM -- IM M 1000lL<fwt>C. Altefuaa veivmea may b* pmpeaai 9 0 S w M N ftr ttin .! StmdaH StocfeVarttflcatian Salum 0.1.1 tapera attack n M m i o f-100 ttfta L o f PFQA by w a i|h in | 10 m* .rm ly tk a i M a te d ( t m i t u d iiW purity) and dilate m IOC n L with T J d h d h a !2J - L L D fftb o k . 11.3 A 14 |*#fcaL kmcaan teluOeu cfPFO A ia prapurad by ta n * * I u L o f m 100 iipftaL aoludon a float volume o f IO0 wnk nuuooi ia 125 teL LDPB bottk. 9.13 A 31 p fftn L featiie tektt Mlitean a t PPOA ia praparad by h rin fin f 10 ttL Wtba L0 |fmL aoMtioo lo ft flnal volume o f 100 w ill) im iterai in ft 125 L L C P l tend*. 0.M Tb#toAandtoerttneo*aolutteoa t t* tto iW in rcigexakx al appffteteiatey 4*C ad an atabie f tr a n u imum period o f 6 montte H m tea 4Mb o f pnpanduu. 9:2 Standard CbUhntka Sohdaot J.I LC/MS/M3 oaUbndon ite d u d i ftn pnparad In methanol via dilution Tdw a i p j u L Jta U kfliM in W m llta ftowtn typical am ple additional eoncantatiooa may te peaparadaa needed. O otim rteaa ffF a te flfttiaa SohuiaaCnataU 100 too too |Jb 2.1 1.0 <|>U ii 10 10 to 10 10 B btedio - ta U -- .. w too 100 100 too LOO fatal Caaocnmion iite W .) ` ^ :d 2.0 10 0.2 0.1 0.1 at 1*C la 4*C up la rf* mouth 9l2A Alternate volume* and coneantntiaaa o f stanrfanla may be prepared *t f t p * a ft Page 55 o f 65 Exygen Research Page 118 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P 0 0 0 131 Btygcii Protocol Number P0001131 iqpatm aap* VMDlTfj i a s a l y t k a l M rrx o p Matt* oAaatya torD*<imiadoofPorfWooounok cld<?FOA) in m*B Mammal L lm b y LC/MS/MS 14.0 BambSaiUp 10.1 Back |M k o f m a im beu-- ad (typically 20 or lam) muai belud* bi Wan c m u a ta M a to l m i lwo la m ua J M aooli lU tilld u known am eaM loM (tab aoolmJ apba) Io vaviiy FOMdural rooowwy b r ba bMck. 10.7 B*qmraman tar Bold and tahrnmnry dapheama m * ik w ill U ip iW . ta Iba quality aoaonnuiptao ta ibia projaat IU 0 Irn fW IO M M a 11.1 Wai|fc 1 1 o f ampta tato 1 50 mL potypopyliM canrnAma tabo (fo n ify m Mtadad, mptaoa M and atax wolQ. Mola (hat ahantat* w w jh ti o f livor may b* m wweA itapmdBM * m ^ ta ata i* r as. 1 IJ Add w a s ** Mmaompta ta rt to o l * ) * iQ m L 1 IJ W ana>aaim awaftauaU>Htaaaamloarfcr-l Etaauio. 114 T no aflf l m L o f Mm mmpta uaaof a dt^o ariila pipoM Imo 15 mL di^om blo o a o to flifi tub*. I l J AA dfm LofaataaaiiiiloaBdihaktfcr^O m im taaaoBiawriaM gtiooalM km li Ca*MAi#a b * tataa a<*-3000 zfH is r-S arrad**. ' 11.7 Domes Um a w a rE M tato 59 iL dhpomtata o a n tifc ft uba wd add 35 ` m Lfwatar. U l CoariioE Mm Cu SPE oaflndpaa ( t * t mL} by y a a ilw 10 L m attend WtowM ky 3 L o fHFLC waiar ( -1 roptaac). Do ool los eolumn n df> ' IM loodtteauplooE eoudittauadC u SPBeaitaUfO. Diacord aliuw. 11.10 Eta wfth -2 mL o f naumiL Coact 2 mL o f bji* im o a graduated l5m Lp^f00pytaM oaD rtJb|oftibo(fijuivotm aw 2m U I U I AmbiEoaamplastHtafotaetaoopaayLOM 8/MS. 17.0 C fc w tamapky 17.1 bgaat tte m m amrnmt o faoh afcmdM, amqpta ad Ib rtifa d aunpb into <h* LClMSMB sj tawt A ooltartM aswited muta frem d* m d fallow an i|m d a a fffi-- Z 2 Staadwrta p ffF O A o n m n n d im o H -- tfly g c o w a o oonoanimlon W vtli amat W tactodad ta be analytical m l IX ) Ae ante* aalo f aoMbrati atandaada muaito iacludad ta Iho te g m n iri and dm and o f a carayla aat. Btaodud* muai W mtanpmood teiw tan ovary S-IO aamsptaa. A i mi allarm livo; an o m i* aat o f caiibrotiw ataadarda may be tajadad i l dm M pantaf o f E am fWlowod by calibrate* atmutank taiarapanad army J-IC amnpteo (Io aocooitt tat a aaoomd an o f aiandardii in *fMmr c m , calibration m uted* (Mat bo Mm Eml and lia* injouiun in a unptaaat 1X4 U t* fiooor w i nd curvao tar ymmn rm Linear standard carvaa am to-- *ad tarste aEolyta by Uoaar mgroaaltm ataaj \J* weighing o fy a k a w Pox 56 o j 65 Exygen Research Page 119 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No. : P0001131 Exyjen Protocol Number: POOOl 131 _______ _ _ _ _ _ _ _ _ _ ____________briba i M itear V000I71) 1 A hA L v iic u . tr r H o i> M a te d riA te te te te l!> a > iw te tte fP rflio T i rn n o lo A te (frfO A )iii8j ' MaoMHil Livw by LOMJ/M S a a lili JteaLyroi J.3 (or QotvaleiM e ftW M jM k H i ta ip la nopcesa ihoul* a rt excoad bedani N ^ o b m . Any triple* that a a cte banted foy ana ii dwnU be fla te c ditata* and nanelyzed. 13.0 AccaptaoM Crdaria 13.1 C ta a te o p a m a t abaw a paa o f a 4a|btar ma al 369 amu fa a panni f 4 )} n i Tbo 413 umu p in o t nncrponda io t e PFOA anion. wbils ihe ria g h n r oa (349 mbm) wyw iw ii Dio lo o f o tte * dtoaida. M I M dkod M uda a ia aot oocted FPQA lavati in a la r than te LOQ. If a U nlc ot e il KPOA lavali p a ia tkm lO o p 'j, te a a m w blank t ampie m ia ib o o b ia ite m tbe e n te aat ostribo*ainotad. 133 U m or i o f control aptoa od m u te y w t a r i bo b r in a i 7<M30H of te irk s rw vate*. If a onte apte fella te d i (K i acoaptable U n iti. th a b te aat o f > ln te u U ba i m m ay n o n o tk * oyudc 70130% dwuU ba m te lad by te aoalyal fio detenerne if ra-catracdoo io U Aay calibratiao aodaad b a i io ba a ariatiod ondi by ueing (he H*pe Bbar Tasi, ap ba ct e ad t e * (ba eaUMktto* o f (bo olibnoon cerve Ib w w r. te total naaaW o f o tlb n te n abndaadl te t eould ba occludo! m a i A ra te M K o tte M ai nuotar a fa tte n d a in io c b i U J D m ooiwlatioa aabBcaal (R) far aalteabaai o rin a gaaantod nwiat be 90.993 (R1 b O .tti). I f ca b ririo a m alte M I irid e teee lim i*. bea q y rupriab anpa aat bo (a ria (fe ti te riria n t operaiwn. and ihe e b a tedt or te ta o t o f aonpba te u ld bc re a a riy te . 13.6 Mi r i irim te baram i dan tedi t e a m p i m a l noi d rtft more ihin 4 * r it t e aaalyboai a . tfre ia miuu timo d rift cnoaedethu lum i within analjaleai mm A t e t aaot ba nanalyiad. 14.9 Calauladana 14.1 U n te M a rin i apaaoa lo calcata te a m o u a tc/ffO A 9^nd(m optnL baaad a * pad a) uaia| te andari arra (baiar ragna* paraiaaten) p a a n rid by te A te i L y n aoftwan p o p m : FFOA r ite (B |te L ) a fl L* DPt aliquot frcior U f m teaorby teaite t e t e i lu n a wm d itu te , ifnonraaory. Aliquot b o ta r- 10 Papef? Page 37of6S Exygen Research Page 120 of 132 Interim Report # 10-Analysis o ffis h and Clara Samples Exygen Study No.: P0001131 E x y | n P ro to co l N u m b e r P 0 0 0 1 131 M x < w iM i.v n iiii I A N A L rilC A L M tT W O C M ed B d rfA M fcn lian b tp M iito lM B aaarN rflim o w M ael A ld(W O A )lnSiiuill M to u u ) Liver by L O M S M S K J Pot wtoIm M M triti b wuo n N o t* gfPPOA prier to roctfon. uk 4 M h ir t to U t t a llito l l i M tm vw y. b M iy ( |> I te o l M m a M lM e p L ) -- ly Bund le cannai (m o n ti) M lyIM ritaddV W -} * 14.3 U to to W t o w i i erpwlin i Ip c m v w t to m u n i o f f f J A trnmd in nipniL to M d iM ). pro* fcM (h*)- mmiitoriiBrtiL) liai reta fml.il tomptawtotfK<t) Exygen Research Paga M o /U Page 121 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygcn Study No.: P0001131 E x y p n P ro to c o l Nucribar M O O II) l A N A im C A L METHOD VMOI ? M a ta rt a fA u lp ia h r ha D aear^adee a# h r f lw r w w lr AaU (P fO A l la S u it M u a a l la m a by L C M IM ) AnafyScAl TaacSisfaculty: Icytaa Kaaaarch 303 Paaaarch DWva OtaCaC U hfiL PA I<01 AppAM Br __iri=A--C.~lLa___ P u l CjwpoBy T advkal M a r. U > M I. Ity p a Raaaaidi a / L j ________ Jm tP lalM dy / Vjo* Pnwklaoi, Opwetpors, t t j g m Mmamdk ___ L V lu ,/0 '< Data Exygcn Research Pag* 59 of65 Page 122 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Bxyaen Protocol Numb: 1*0001131 )*. VW0I7U I A tlA L V n C A i. M I TMOP Mft4toolal)l 4It tNantuGian afN tloorooctonolc Acid (PrOA) *jaSmiil M a in a i Senn by LC/M 8/MS Tbb in ten d tow be m p b y d fe r ft* IsaM m and qwftte<fcnerpertui>i'ioaano cid by H^t H H m m o a liq u id C fcrwuw tnpfey * a taadam Maw SpecfteoMw On oacr h i m a il rnanuml earaot. 10 M M y 2.1 A hm yob--1veaaJbfcbeanarypmriow. 1 2 Ctem k tee nnm prim M S D J betel* bandliaf any tecm ical fer p o p * ealcty pawmmrlrau 30 Sample Itetywenant I.) A lle m lm L n fia e tM n p la fe ra M ie n . J-2 Ne w agte f u iw jii j to needed tor eeewa a m p in . However, frozen arum a n p ta i a m e atiewad com plm ly ih a w te io o h i le q im m w b e ih re u * . 3-3 Im p 1a m O ndo* p qceter w w ill be eped it e m tea aanpltei plan tor 1K11 p u l . IQ 4ft, 4 S lu U irii 4.1 W ator-HTLC grWo 4 J M id u n o l-H 7 U :ru 4 i 4J A w w M b -lffL C p * i t A o B M diU K A M ia i-A C S RatputO nO o 4 J rm * m n cM m m r Ao4 - SiyuoA M riM 30 h n m a t> U W I 3.1 A high p d b n a u c . MfttiO ctaauautm O Cftfftbto of ponuauu v 1 2 o tu a u i f ii p ii l M i ft vatoklc ulano tu l to r MfOhto o f iujocun 5 2ot> y l n u ftftr ftiil i t i m l -- >1-- i i uctu (LC/MS/MS) SJ A Co to M tlftol to 4 , far rak tnU pulion ond ouftndMion. 3.3 A m lp m l W m u p t * o fn k Im 0.000011. >* 30 o L jiu ftu M ft rolypcoQyl--. W Btofcn tub 3J 13 n i 4ift0lft4l4 fB typrryyimm ojftn4j* uiltoft. 34 ItoQ O tftlil, in a fiQ . (30. lOOul, IOO-2QOuL>. I I 123-toLU>MMnowtoioudl**. 3J 2 i eUar HFLC ill IdL j,9 DbpotoW apiyte 110 Aitoopipad (100-UH0 pL and 10-100 pLJ. wite dkpoeihlc ripe. I I I W orn SepPakVa 9 oc( If) C lI SPE cwtridpn. 112 B r t m m M iM l 112 Vartw ar f>tr 2rf7 t o p 60 o f 6S Exygen Research Page 123 o f 132 Interini P.eport # 10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 Exygeft Protocol Number: PQOO131 MbuJ <rf VD00I7* ATW VnCAL M1THOP fc D ttrm autiaa a f Pirfiaocoocttnoie Acid (PPOA) m Small lbyLO M SA 4$ J .H U S C n frtfip tttp iU fe a rin g I S ra L p ly p riy y ta im b it 3000 ipm 0.0 Cwgma#rtfttciy+ ta i l A u ly tic a l CaIu b t Wooptami IP (Kayttom Scientific), 2.1 mm x 50 mm. %i <7fc *2305452130) 4 2 T m p m o n : XTC J M obikPkoM (AJ : I h M A o BMBUBAm ata i t Wh h i*4 U iM U rW (B > : U ittm n f 5.5 Gmcfatf Program: flm a/m m l 0.0 14 1.0 20.0 2U %A 03 45 25 25 05 PbwRm 5kJ Im l/m in l 35 0.3 35 0 J 71 0.3 75 0.3 35 0.3 * 4 latoegtoe V o h m IS p i (ow \* m o m m i to muck m 50 u 0.7 Q u a rta ta ; ta lc A n a -m o m * m wlard oriitaetum ce m . 4 1 SmxT o ; --23 miaou*. Urn en w w tidow eo wm*d a gtidM nd my b# cArnigsd in order e#dk <eHFLC e je w t 7.0 M S M S y 7.1 U o 4 r EUettWFay N a tiv e MEM moie, m w ihortiis^ Il- T ltiv i Ibr ffO A . Th$ ito v i oooO iooo M io f dml u t faida Ad nuy bo ehaefod io ordir io a p O ra x iU M S M S o n n . 10 P n p m la a irM u d ia i 1.1 M e tti* Domi 1 1 1 le M e w w ie ie e m ie fa wemr iijw o p tte d by eddtns0.1S4 s o f wnrmlum K M k hi 1000 mLofwmw Aharamivotame*nuy b* pwjwrwl Fagw 6 / o f 6J Exygen Research Page 124 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.; P0001131 x y |* a Proeocol N um ber: POOOU31 M ^M N uo^rVO dO TK, ANALYTICAL m e t h o d tfA u iy ri for th Deunefautte e /ftrfle e iw x tte tf fe tt (PPOA) in Snull Mm b ih I Serum hy LC/H3/MS 94) 3 \m m tn n m n H m 9.1 Ic te a id Stodc teriflctec SohUte 9*1*1 P n p w e lD *e l*i is f-r|Q O n j*a ,s f 9FO Aby w eijhins 10 * * oimmiyMcti t e lte d [oomeiM for purity) and diluN to t 00 n L with i m ette! ia 123-mL LDPK bor, 9.1 J A l.0 |i|W . fortilU edo* oludos o f PFOA I jreperN hy bringing l L o f te 109 u ttm L te te io fite volonte e f igo with evteM i U tiM i f , u w Wuri 9 1 J A 9.1 f f t e . fortU cfdee ohrioo affP O A 1ywy w d by b rit\g ^ IM L if t e 1.0 M0ta L a o h M * IU * J vcleraco/lOQ withmwtunoi in US b L LDPB bodlfc 94.4 Th aleck t e foatifo ette aoJuboee at lo be etored ta t*fn% tnux m p fte te te y 4* t e m ste te for t maxknuM p e rte f 6 m onte te te 4m e fi fip -- iica. M S te te C e fite tte f te * * 2 1 L O M M f l te fo re te te rnie m o p r ^ te ia nwafamol via ditutio oft e 0.1 pftaL forUlaert estate. 92.2 Th foltow taf 1t tfdcd u a p ta eddlrtote ooaocntmioa raey N , pnpped Bteed. CteMsndte w ifm u a m V te * --------- F5SJ------ D ilute ie C oM entitte O e te a ste M .) ( U (eriJ 100 W m .J * :j 14 T 3 S m 3.9 10 *4 1.0 100 10 14 190 1.0 11 100 0.3 H 100 0J H 190 0.1 Jt ca ib n tm n > l23-vL )P 8 narrow-mote) holt1m M 2*C le 9*C, up to ht t e * . 92 .4 AMmimK voUm i t e ooGoeatmUena o f a tte s ri May h p*eperei s e s te OO te c h t e Up UM lm b beta r f w f l sm ote (typoity 20 or te) m u te ilte t Im t m M N te aon te t e te te s t e om irol fontted known ooMXrtraoo (Ufa p te o l ^At) to verify prteur) iseovsry for te bewh 10.2 Requirement! for M d and te o rte ry fo h lte t t e spikaa w ill be ipecifiaJ ia t e quality aeemiee p lu for foi project. r r *( i Page 62 o f 65 Exygen Research Page 125 o f 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Exygea Protocol Number: P0001131 fcOQU I W OK t Mua V00017I4 I A W A trn C A L M IT B O P M a t l< rfA i^ iim lb rik > D M n B > u ija a ^ M w im n i> w A tU (rF O A )ifiS a u ll M ira n t I m ty LC/M3/THS 1 1 . ro d ) 11.1 M w w 1 L f I |il M o t <0mL polj Em n 'lm fu n lfti m bo (fanify d m M . n p k e t U d a b m UX Wo duu b c n u <raluma o f leruni m ty bo mm* n i dopmdan m the ewayk * * availed! for um. i : 2 AJd M a ris tto in a p t* for a fo u l o f 20 o l. C a ^d |h tly li J Vorte* fo r - l Mcmfe. U.4 T rw ttm t u L o f it w y W v ia * * ap<Mble y/poni late a 1$ * L rfftTTTltVH M fe ih H tube. H i Add i m l o f eMtoaibiW cad hoio fo r-30 miaialoo on a w ti-*ctioa shaker. U 4 C M d fo it foofoheoi-3000 rjw for ^ m ia a ie i 11.7 Locaw foe n y in r in t Mo * 50 m L dupoubk ocoorfufe tube u d iM 33 Lofw M cr. H A Cbodhko fo i C O T e n td ie (1 s *<U W p w tiH IQ mL m tihw el Ibli&wodVy JaLfHFLCwoio((~2dVQpfcee). D * oo< 1 olumn ran dry iU > U altbem ptooaoondU taM dC uIF E CM bidjc Discardehiaie. ll.W EhOa with -2 mL f Mtrtbaaal. Collect 2 nL o f eJua* IS BiLpdlyy topyfoa cantrifop tuba (l& a l vcIuhm 2 tnL). H.L1 / iflly a eemplei u tia i deettoepney LOMSAC8. graduated 3L0 Clw a i nyepby U -i byact the m bm -- w ot o f--ch kmdard. eaatyte and fcwiflod temple into foe LC/M2/MS ) * . A uM btttigo f doid nonet procode end follow all ia tl)a k u | ili, 12.2 Staodcrde o f FFO Attircapaadlai to u keci five or n o n ooneoftfntLon level act be included ie n a a ly tlta lM L 12.3 Aj eabn M l o fadihraiian OMMbrtl oaiat be iaefodad k (ha iM finnin^ and m Ik i cod o f a am ple mi. Staadarda muat bo iatenponod betwee* ary 3-IO MO^rioL Aa an ahomaiiv, m aatin BCt o f caJibmtiee tandcrck m iy 0 injected be boftanfaf o f m i followed by etllbru ion eianderui lakrapcned ovary 5-10 aeaapko (to aooouai for 0 eeoond M l o f etandrde)> 1 etfhar oaaa. e e likn tm C a a te li a m t bo (he Ant and foot isjecuon in a u n p ie eat. \2A rJM Him m kaodn4 ourvae for quantitation. Linear fU nU nt curvvi an ptM M ttd for foe wwlyte by Haoar reyeeaton uobtfl l/x w etfK ing o f peak ore venue oUfonbea standard Ouncantraxieo uciag M a a L yu 3.3 (or oqwivalootl OOCOOMCJMMI. )2 -i Saoqte iMpqaae eheeld not w ood alandard reapcocca. Any uaiptw that CKeeedsMadafdraeponMe Mould be fathe r dihrted end ttanclyred Exygen Research P a g e AJ q f'43 Page 126 of 132 Interim Report #10-Analysis of Fish a id Clam Samples Exygen Study No.: P0001131 B xyg ra P rotocol Number: POOOl 131 ftU a l l HiA ______ ___________________ MtAkd kWWr VO0017H 1 A K A ty tie A T iw iiT iio f f S J ? 3 ^ J l r S B 5 5 3 3 7 K 3 C S S g l? 3 3 T ro ) i!i5 M anual S a m ky LC/MS/MS 1 1 1 Q w iw m y w i awn w> pw k o f tu p to w too at 360 amu from t paraat o f413 w i. T to 415 M p m ) ngwrapmA to t o PfO A muon, tu * Uk da tt^tf oq (M 9 > rapraranta to Ira i * f e vto n drastda. J 2 J MafeOd tto fca sum mm comma FPOA at lavU eater than (to LOQ if a M u k io to -- FPOA at k v ili p u to A m 10 pffe tL . Am now Wank em plasrw ttoebtaaw dm J to * * rt* M \)*t toia-cxtractae. 1J3 la crm ira a f acato! f i t o m d ratoia qwka m a l to betw een 70-130Hof tto ir b w w vtoira. V a uoartiul wpk* lUi enteide Uto aocapubl lim n , tte to a m i a f m ft o Mould to la m tm tod Any a m i lite oauide 70PC% aiwuM to evtoeaied ky tto malyar m I m m m If rcra tra ctia * IM Aay to to tm Amdacd bund ta to a w e a l outtor ky ra in * tto H uf* Knor T to , ay to m cto M to n tto calculation o f tto calibration carve H to to r, tto total m nbor o f m I M m adarda dut could to eluded nalnut mooed 20H fto load M ttbra o fatorada o yn tto 13,3 Tto ooctoUboo o o a flkivtf (R) to ealibraaea n m famrmtad mutt to JO M 2 (K] a a s u ). I f n llb n d o a M tk M l oulalda dnae k a u . then pprapnata apa mu to latoa ta adjuat tomunral operation, rad the emtoda or (to relevant rat o f aaniplat Mould to ram tlysed, 13,S X alandoft tana t o m a madaiO in f a m p in mum not d rift n o n than * 4 % Uhls m analysed ran. If ratration tima d rift aerada Ais lim it within m analytical ran than tto aat m to ranalyud- ! 4oO ra lra lilin m K 1 Ura tto A U ovia i qufeu i calculara (to amount o f FPOA fraud (in nffm L brand en pato ana) uehtf Ao a d ia d curva (toara ra^aaclwi parwoaura) Sananiad by to MaasLynx aetwara praflraai: fTOA toad (a iftitf.) iE ito to to d llto M L XDP atow* lhaor to p DP to ra rb y h ito to 1* 1 raluioa n w ddutad, I f osowenryAliqaot h o o t - 20 142 Par M apIm fcrtiflad M i known m a t t o f FFOA prior to attraction, oac t o M ia w liif to to io n A cttlpitoa Aa paraant (icorary. to to m ry (% ) [to ld m aty* to a d (n ifa L ) - m ibao to n d in control(flp to L )] k]Q0 ________________ to y added (oatoL)________________ * KpTaf? Pagt 64 o f Exygen Research Page 127 o f 132 interim Report # 10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 Bxyfeo Protocol Number: POOOJ13] ItyfMltM M Mk M iti< Kumt VOM17 I /Q t/v irn C A l METHOD ~~ Method t f A m lyiia Ibr A t OwNwaiiwdoa o f N rfW w u w ic Acid (PPOA) in W ll UammJ Im w n by LC/WS/MS I4 J Ute * t JbUowtac tqiMon io couvert Um D tuni f rfO A te n d m n^mL io PPOA IbuaJ (ppb) - fl Exygen Research Pg* i 5 o f 65 Page 128 of 132 Interim Report 0-Analysis of Fish and Clam Samples Exygen Study No.: P0OOU31 EwggI T RESEARCH 3058 Research Drive Phone: 814-272-1039 State College, PA 16801 Pax: 814-231-1580 PRQTQCQi. AMENDMENT Amendment Number 1 Effective Date: 01/18/05 Exygen Study Number POPP1131 Cileni Study Number Pe1oM DESCRIPTIONPFAMENDEDSECTION 1) Analytical Procedure Summery V00017B0:SecUon 9.1 2 ) Verification o f Analytical Procedure _ tte 0 2 _ AMENPfeP.Ta 1) Add to Section 0 .}: Section 9.1.8, Alternate weights of atandarde may be used to prepare altemeto concentrations of stock solutions as necessary. Aftem ata levels of fortification solutions may tfao be prepared. 2) Low and high spMng levels of the aratytealor each matrix mey be altered depending on sample size available for extraction andfor to cover analyte concentrations expected In the samples. RATIONALE 1) Htghsr concentrations of standards need to be prepared In order to spike the sample bottles at higher leads. 2) The sample fixe svelablo for small mammal I ver and serum was smaller than pectod . Spiking at the pre-detarmlned lavala In ths protocol puts ths spiked ooncenketlon tower then the detection ImK. Also, the analyle levale In the ground w ater samples are enpneted to greatly exceed the predetom tined spiking levata listed In the protocol. When Ihe levale to the samples greatly exceed the spiking levels, an accurate recovery value cermet be cafcuM adforttw Q C (am ple. Htyher spiring levala In the bottles e li covar ths analyts concentrations expected In the watar sam ple*. IMPACT ON STUDY The LOQ la 100 ng/g for a 0.1 g sample of small mammal liver and it 1000 ngAnL for a 0.01 mL sample of sm al mammal serum. Higher level* of spiking for the water samples w it ensure that more Q C recovery data can be used. E ff***-* Exvosn QAU Review I t f o t U i if LIBRARY IKW H0012JM v ... ADMINISTRATIVE FORM Exygen Research Page 129 of 132 Interim Report #10-Analysis o f Fish and Clam Samples Exygen Study No.: P0001131 3054 Rosearch Drive Phone: 414-272-1039 State College, PA 16801 Fax: B14-231 -1SSO PROTOCOL AMENDMENT Amendment N u m b e r _____ 2 Effective Data: 03/07/05 Ex>vr Study Number P0001131 aient Study Number fto o la M None DESCRIPTION OF AMENDED SECTION 1) Report png* 11 of 05 2) Test M aterial*, pepe 8 of 66: PFOS tranettlon monitored 499 -> 99. AMENDED TO 1) Instead of one Unni report, Interim raparla wAI be issued. 2) PFOS transition monitored may alto ba 499 -> 80. RATIONALE 1) Due to the enoearive atoaa of toa data seta, Interim reports will be Iseued to altow toe dient to receive data In a Umettarreeeen ''*'* e t/U A r r 2) The API 40 LC/M /M S systems dataci the 499 > 80 PF/1 ranriitors with greater ensWMty toan toe 499 96 transition, impa5 t 5 n st u d y 1) The d e n t w be abta to wcalve and revtaw the data more qUeMy. 2) The 499 -> 80 transition can be detected with greater sensitivity; bettor ohnxnotogrephy. giving Exygen CAU Ravtsur. UBRARY ID: V OG0122W l ... ADMINISTRATIVE FORM Exygcn Research Page 130 o f 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 W-HI-tM! Frce-IffJTN SOLUTIONS n RESEARCH 1 305* Research Drive Stale College, PA 16801 . ,. t' T-114 P.001/104 F-417 Rhone: *14-272*1039 Fax: 114-231-1560 DEVIATION FORM pronai X Project Spedflc Deviation PadRy Deviation ... Date cf Occurrence: 08/29/05 Exygen P r o je t# : Oient Project#: P 7B tV P iW NA D eviatio n#: ili R eference#: & z l l Realitten Driver: Devistfori Type: /IndudTvSTrmwthodM and GP) OMP GLP Other None 8efhi>fp P H g M a m X Protocol _____ Method VW; NA Notebook reference: NA ______ s o p Log in #: U 2S 4A 4258 Container: . 00086460-86 L o t# : NA iM ir U K P f Pevletfatai The protocol states that control and fortified control aamplee of each matrix will be analyzed; however, control dam was not obtained. Fish was ueed ee the control for the aneyds o t fteee six clem samples. Cauce: Preparation___ Analysis____ instrument __ __Client Request X Other Im pact: No negative Impact on this study. C o rre ctly Action: Deviation Investigator f}y~ + Quality Aeeurence m Date Exygen J Sponsor Representative Jk. Sponsor Management ste Dels D e is Exygen Research Page 131 of 132 Interim Report #10-Analysis of Fish and Clam Samples Exygen Study No.: P0001131 M-tO-tOO! 02:35p* froe-WSTOK SOLITTIQMS n RESEARCH T-JI4 P. 004/1)04 M 3T 30S# Research Drive Phone: 814-272*1039 SUte Collise, PA 16601 Fax: 814-231-1580 Ganarai: X Project Spedite Deviation DEVIATION FORM Facility Deviation PWLoM Date of Occurrence: 12/27/04 Exygen P ro ject#: d ie n t P ro je c t#: P780/P1131 _____N A D evletfan #; 3/3 Reference # : oS'-fS ^ Regulatory D river. Deviation Type: floct/de V tith r methods a /x f SO Pal QMP CLP O ther None 1 5 5 Pawnotfan: Login#: NA ______ Protocol _____ Method V * 202-20 Section 5 2 .3 Notebook reference: NA Container#: Lot#: X SOP NA Summaryj f Drvfttfgn; SL2114 (30% OimethyWidUoroellane In Toluene) wee gfcen the expiration date o f 02/13/05, but RE544 (Toluene) need to make S L 2I14 expired on 03/28/04. SL2114 was ueed to jia rtied glaHw are prepared for the fish extraction from 1227/04 through 01/07/05. Cause: ____ Preparation _____ Analysis_____ Instrum enta____ Client Request X Other Impact: No negative impact on the study. Toluene was used only ee a solvent for the gbesw are preparation. Dimethyldlehlorosllana. which is the oaab'ng agent, was not expired. C orreettve A c tio n : Deviation issued. Study Director Q yC *. S id L tK Q uality Assurance Oats mb la tt( f/it/o r Date -- Exygen t^Mi^gwnent & _______ Sponsor Representative Uh Sponsor M eragem ent Date Dele Data Exygen Research Page 132 o f 132 INTERIM REPORT #23 Analysis of Sediment Samples STUDY TITLE Analysis of Perfluorooctanoic Acid (PFOA) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program DATA REQUIREMENTS EPA TSCA Good Laboratory Practice Standards 40 CFR 792 STUDY DIRECTOR Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 Phone:610-701-3761 INTERIM REPORT COMPLETION DATE May 2, 2006 PERFORMING LABORATORY Exygen Research 3058 Research Drive State College, PA 16801 Phone: 814-272-1039 STUDY SPONSOR 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 Phone: 651-733-6374 PROJECT Protocol Number: P0000760 Exygen Study Number P0000760 Total Pages: 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 GOOD LABORATORY PRACTICE COM PLIANCE STATEM ENT Exygen Study Number P0000760, entitled "Analysis of Perfluorooctanoic Acid (PFOA) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program," conducted for 3M Company, is being performed in compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792 by Exygen Research, with one exception. The laboratory control and control spikes for the direct injection and alternate SPE methods and the spiking procedures for the surrogate were not recorded promptly at the time o f extraction. J b h n Flaherty ' Princrinpal iInnvestigator Exygen Research Jaisimha Kesari P.E., DEE Study Director Weston Solutions, Inc. Date Exygen Research Page 2 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 QUALITY ASSURANCE STATEMENT Exygen Research's Quality Assurance Unit reviewed Exygen Study Number P0000760, entitled, "Analysis of Perfluorooctanoic Add (PFOA) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program". All reviewed phases1 were inspected for conduct according to Exygen Research's Standard Operating Procedures, the Study Protocol, and all applicable Good Laboratory Practice Standards. All findings were reported to the Exygen Principal Investigator and Management and to the Study Director. Phase 31. Final Raw Data Review and Interim Analytical Report Review Date Inspected 5/2/06 Date Reported to Date Reported to Principal Exygen Date Reported to Investieator Management Study Director 5/2/06 5/2/06 5/2/06 'Note: All in-lab inspections will be documented in the QA statement for the final analytical report at the conclusion of the study. This QA statement involves only the review of the interim report and associated raw data. Exygen Research Page 3 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 CERTIFICATION OF AUTHENTICITY This interim report, for Exygen Study Number P0000760, is a true and complete representation of the raw data. Submitted by: Exygen Research 3058 Research Drive State College, PA 16801 (814)272-1039 Principal Investigator, Exygen: Date Exygen Research Facility Management: fi> Z Richard vy<jj President Exygen Research Study Director, Weston Solutions, Inc. Jaisrmha Kesari P.E., DEE Weston Solutions, Inc. Sponsor Representative, 3M Company: Date Date Exygen Research Page 4 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 STUDY IDENTIFICATION Analysis of Perfluorooctanoic Acid (PFOA) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program PROTOCOL NUMBER: P0000760 EXYGEN STUDY NUMBER: P0000760 TYPE OF STUDY: Residue SAMPLE MATRIX: Sediment TEST SUBSTANCE: Perfluorooctanoic acid (PFOA) SPONSOR: 3M Company 3M Building 0236-01-B-10 S t Paul, MN 55144 STUDY DIRECTOR: Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 STUDY MONITOR: Michael A. Santoro 3M Company 3M Building 0236-0 l-B-10 St. Paul, MN 55144 PERFORMING LABORATORY: Exygen Research 3058 Research Drive State College, PA 16801 ANALYTICAL PHASE TIMETABLE: Study Initiation Date: 11/05/04 Interim Analytical Start Date: 04/27/06 Interim Analytical Termination Date: 04/30/06 Interim Report Completion Date: 05/02/06 Exygen Research Page 5 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 PROJECT PERSONNEL The Study Director for this project is Jaisimha Kesari at Weston Solutions, Inc. The following personnel from Exygen Research were associated with various phases of this interim portion of the study: Name John Flaherty Karen Risha Paul Connolly Christine Edwards Mark Ammerman Amy Sheehan Brittany Kravets Scott Crain Frances Crespi Devon Ingram Christa Gallant Title Vice President Scientist Technical Lead-LC/MS Technician Sample Custodian Associate Scientist Technician Technician Technician Technician Technician Exygen Research Page 6 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 TABLE OF CONTENTS Page TITLE PA G E ................................................................................................................................ 1 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT................................ 2 QUALITY ASSURANCE STATEMENT................................................................................3 CERTIFICATION OF AUTHENTICITY.............................................. 4 STUDY IDENTIFICATION....................................................................................................... 5 PROJECT PERSONNEL.............................................................................................................6 TABLE OF CONTENTS.............................................................................................................7 LIST OF TABLES........................................................................................................................8 LIST OF FIGURES...................................................................................................................... 9 LIST OF APPENDICES............................................................................................................10 1.0 SUMMARY.........................................................................................................................11 2.0 OBJECTIVE........................................................................................................................11 3.0 INTRODUCTION............................................................................................................... 12 4.0 ANALYTICAL TEST SAMPLES.................................................................................... 12 5.0 REFERENCE M ATERIAL...............................................................................................12 6.0 DESCRIPTION OF ANALYTICAL METHOD............................................................ 13 6.1 Extraction Procedure For Sediment................................................................................13 6.2 Direct Inject Extraction Procedure For Sediment......................................................... 14 6.3 Additional SPE Extraction Procedure For Sediment....................................................14 6.4 Percent Solids Procedure For Sediment........................................................................ 14 6.5 Preparation o f Standards and Fortification Solutions............... 14 6.6 Chromatography...............................................................................................................16 6.7 Instrument Sensitivity...................................................................................................... 16 6.8 Description o f LC/MS/MS Instrument and Operating Conditions............................. 16 6.9 Quantitation and Example Calculation...........................................................................17 7.0 EXPERIMENTAL D ESIG N .............................................................................................19 8.0 RESULTS....................................................................................... ;...................................19 9.0 CONCLUSIONS............................................................................................................. 20 10.0 RETENTION OF DATA AND SAM PLES................................................................. 20 Exygen Research Page 7 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table I. LIST OF TABLES Page Summary o f PFOA in Sediment Samples........................................................... 22 Table II. Summary of PFOA in Direct Injection Sediment Sam ples.............................. 23 Table IH. Summary o f PFOA in SPE Extracted Sediment Sam ples................................ 24 Table IV. Matrix Spike Recovery o f PFOA in Sediment Samples.................................. 25 Table V. Matrix Spike Recovery o f PFOA in Direct Injection Sediment Samples...... 26 Table VI. Matrix Spike Recovery of PFOA in SPE Extracted Sediment Samples........ 27 Table VTT. Surrogate Spike Recovery of UC PFOA in Sediment Samples.......................28 Table VUL Surrogate Spike Recovery o f 13C PFOA in Direct Injection Sediment Samples........................................... ......... ........ ................................................... 29 Table IX. Surrogate Spike Recovery o f UC PFOA in SPE Extracted Sediment Samples....................................................................... .......................................... 30 Table X. Total Percent Solids in Sediment Samples.................................. ...................... 31 Exygen Research Page 8 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Figure 1. LIST OF FIGURES Page Typical Calibration Curve for PFOA in M ethanol............................................33 Figure 2. Extracted Standards o f PFOA in Methanol, 25 ng/L and 50 ng/L, Respectively.... ..................................... .,............................................................... 34 Figure 3. PFOA in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively..................................... 35 Figure 4. Chromatogram Representing a Sediment Sample Analyzed for PFOA (Exygen ID: C0172892, Data Set: 042806D)................................................... 36 Figure 5. Typical Calibration Curve for ,3C PFOA in M ethanol.....................................37 Figure 6. Extracted Standards o f 13C PFOA in Methanol, 25 ng/L and 50 ng/L, R e s p e c tiv e ly ......................................... ..................................................................38 Figure 7. ,3C PFOA in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively..................................... 39 Figure 8. Chromatogram Representing a Sediment Sample Analyzed for 13C PFOA (Exygen ID: C0172892, Data Set: 042806D ).................................40 Exygen Research Page 9 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 LIST OF APPENDICES Page Appendix A Study Protocol P0000760 (Exygen Study No. P0000760) with Analytical Method V0001782: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS", and Protocol Amendm ents............................................... 41 Exygen Research Page 10 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 1.0 SUMMARY Exygen Research extracted and analyzed sediment samples for the determination o f perfluorooctanoic acid (PFOA) according to Exygen Method V0001782 (Appendix A). The limit o f quantitation for PFOA in sediment was 2.0 ng/g. The standard sediment method (V0001782) was originally used to extract all of the samples. Use o f this method yielded poor spiking recoveries for P F O A Because o f this, a direct injection method (Section 6.2) was used, which also yielded poor recoveries. Finally, an alternative SPE column extraction (Section 6.3) was used to analyze for PFOA and ,3C PFOA. Analytical results for the analysis o f PFOA in sediment samples extracted according to the method based on dry weight are summarized in Table I. Analytical results for the analysis o f PFOA in sediment samples by direct injection, based on dry weight, are summarized in Table II. Analytical results for the analysis o f PFOA in sediment samples by alternative SPE extraction, based on dry weight, are summarized in Table III. The overall average percent recovery standard deviation for PFOA in sediment samples based on wet weight was 73 16%. Fortification recoveries for the analysis o f PFOA in sediment samples are summarized in Table IV. Fortification recoveries for the analysis o f PFOA in direct injection sediment samples are summarized in Table V. Fortification recoveries for the analysis o f PFOA in SPE extracted sediment samples are summarized in Table VI. The average percent recovery standard deviation for 13C PFOA in sediment samples based on wet weight was 97 27%. Fortification recoveries for l3C PFOA in sediment samples based on wet weight are summarized in Table VII. Fortification recoveries for ,3C PFOA in direct injection sediment samples based on wet weight are summarized in Table VIII. Fortification recoveries for 13C PFOA in SPE extracted sediment samples based on wet weight are summarized in Table IX. The total percent solids for sediment samples are detailed in Table X. 2.0 OBJECTIVE The objective o f the analytical part o f this study was to determine levels of perfluorooctanoic acid (PFOA) in sediment according to Protocol P0000760 (Appendix A). Exygen Research Page 11 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 3.0 INTRODUCTION This report details the results of the analysis for the determination o f PFOA in sediment using the analytical methods entitled, "V0001782: Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS." The study was initiated on November 05, 2004, when the study director signed protocol number P0000760. The analytical start date for this interim report was April 27, 2006 and the analytical termination date for this interim report was April 30,2006. 4.0 ANALYTICAL TEST SAMPLES Three sediment samples (Exygen ID: C0172892-C0172894) were received on wet ice on April 22, 2006 from Tim Frinak at Weston Solutions, Inc. The samples were logged in by Exygen personnel and placed in refrigerated storage. Sample login and chain o f custody information is located in the raw data package associated with this interim report. Storage records will be kept at Exygen Research. 5.0 REFERENCE MATERIAL The analytical standard, PFOA, was purchased from Oakwood Products, Inc. and was received at Exygen on March 17, 2005. The surrogate spiking standard, 13C labeled perfluorooctanoic acid (13C PFOA SP0004184), was received at Exygen on April 15, 2004 from the 3M Company. The available information for the reference materials is listed below. PFOA was stored ambient and 13C PFOA was stared frozen. Compound PFOA n C PFOA Exygen Inventory No. L ot# SP0005444 203 SP0004184 3507-195 Purity (%) Expiration Date 99 03/31/07 97 03/29/09 The molecular structure o f PFOA is given below PFOA Chemical Name: Perfluorooctanoic acid Molecular Weight: 414 Transitions Monitored: 413 --> 369 (for quantification) and 413 - 219 (for confirmation) Structure: Exygen Research Page 12 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 13C PFOA Chemical Name: 1,2-13C perfluorooctanoic acid Molecular Weight: 416 Transition Monitored: 415 --> 370 Structure: 6.0 DESCRIPTION OF ANALYTICAL METHOD The analytical method "V0001782: Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" was used for the sediment samples in this study. 6.1 Extraction Procedure For Sediment Before the samples were weighed for the extraction, they were mixed thoroughly by vigorously shaking the container. A 5 gram portion o f sediment was weighed into a fifty milliliter centrifuge tube for the extraction. After fortification o f appropriate samples, 35 mL o f 1% acetic acid in water was added to the samples. The samples were vortexed and allowed to shake on a wrist action shaker for ~60 minutes. The samples were centrifuged for ~20 minutes at -3000 rpm. The supernatant was then loaded onto a C u SPE cartridge conditioned with 10 mL o f methanol and 20 mL of water. The eluate was discarded. Twenty milliliters of methanol was added to the sediment samples left in the centrifuge tube. The samples were vortexed and allowed to shake on a wrist action shaker for another 30 minutes. The samples were centrifuged again for -2 0 minutes at -3000 rpm. The supernatant was then loaded onto the same Ci8 SPE cartridge. The eluate was collected into a 500 mL Nalgene Bottle. The column was washed with 4 mL of methanol. The wash was collected in the same bottle as the eluate. Approximately two Exygen Research Page 13 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 hundred milliliters o f water was added to the bottles. The samples were mixed by shaking and loaded onto another Cis SPE cartridge conditioned with 10 mL of methanol and 20 mL o f water. The eluate was discarded. Approximately five milliliters o f methanol was added to the cartridge. Five milliliters o f eluate was collected into a graduated 15 mL polypropylene centrifuge tube. Each sample was analyzed by LC/MS/MS electrospray. 6.2 Direct Inject Extraction Procedure For Sediment Before the samples were weighed for the extraction, they were mixed thoroughly by vigorously shaking the container. A one-gram portion o f sediment was weighed into a 15-milliliter centrifuge tube for the extraction. Ten milliliters o f 1% acetic acid in methanol was added to each sample. The samples were then shaken by hand, vortexed, and sonicated for thirty minutes. The samples were then centrifuged for -1 0 minutes at -3000 rpm. Each sample was analyzed by LC/MS/MS electrospray. 63 Additional SPE Extraction Procedure For Sediment The samples that were prepared in 1% acetic acid for the direct injection method were used for this extraction. Five milliliters o f each sample was aliquoted into a 50-mL polypropylene centrifuge tube and the volume was taken to 40 mL with water. The samples were then centrifuged for -1 0 minutes at -3000 rpm. The supernatant was then loaded onto a Cis SPE cartridge conditioned with 10 mL o f methanol and 5 mL o f water. The eluate was discarded. Approximately five milliliters o f methanol was added to the cartridge. Five milliliters o f eluate was collected into a graduated 15 mL polypropylene centrifuge tube. Each sample was analyzed by LC/MS/MS electrospray. 6.4 Percent Solids Procedure For Sediment Percent solids were determined using the procedure indicated in Exygen method V0000427. Approximately 20 grams of sample was weighed into a pan. The weight of the sample plus the pan was recorded. The samples were then dried in an oven overnight at 104 2 C. Then the samples were transferred to a dessicator and allowed to cool for -15 minutes. Each sample was then weighed again, including the weight o f the pan. The percent solid for each sample was then calculated. See Table X for percent solids results. 6.5 Preparation of Standards and Fortification Solutions A mixed stock standard solution of PFOA and n C PFOA was prepared at a concentration of 1000 pg/mL by dissolving 100 mg of each o f the standards (corrected for purity and Exygen Research Page 14 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 salt content) in 100 mL o f methanol. From this solution, the following fortification standards were prepared: Cone, o f Fort Stock or Fort. Volume Solution (mL) (pg/mL)' 1000 10 100 10 10 10 1.0 10 0.1 10 1o f PFOA and ,3C PFOA Final Volume (mL) 100 100 100 100 100 Final Cone, of Fortification Std. (pg/mL) 100 10 1.0 0.1 0.01 A stock solution o f only 13C PFOA was also prepared at a concentration o f 100 pg/mL by dissolving 10 mg the standard (corrected for purity and salt content) in 100 mL o f methanol. From this solution, the following fortification standards were prepared: Cone, o f Stock or Fort. Solution (pg/mL)1 100 10 = 1.0 1o f 13C PFOA Fort Volume (mL) 10 10 10 Final Volume (mL) 100 100 100 Final Cone, o f Fortification Std. (pg/mL) 10 1.0 0.1 A set o f non-extracted calibration standards containing PFOA and ,3C PFOA was prepared in methanol, as specified in Exygen method V0001782. The following concentrations were prepared Cone, o f Fort Fort or Calibration Volume Solution (mL) (ng/mL)1 100 1.0 100 0.5 100 0.2 10 1.0 5.0 1.0 2.0 1.0 1o f PFOA and 13C PFOA Final Volume (mL) 10 10 10 10 10 10 Final Cone, o f Calibration Std. (ng/mL) 10 5.0 2.0 1.0 0.5 0.2 Exygen Research Page 15 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 The standards at final concentrations of 10, 5.0, 2.0,1.0, 0.5 and 0.2 ng/mL were used for calibration. Extracted calibration standards were prepared following method V0001780. The stock standard solution and all fortification and calibration standard solutions were stored in a refrigerator (4 2C) when not in use. Documentation of standard preparation is located in the raw data package associated with this interim report. 6.6 Chromatography Quantification of PFOA was accomplished by LC/MS/MS electrospray. The retention time o f PFOA was -11.1 min. Peaks above the LOD were not detected in any o f the reagent blank samples corresponding to the analyte retention time. 6.7 Instrument Sensitivity The smallest standard amount injected during the chromatographic run had a concentration o f 0.2 ng/mL o f PFOA. 6.8 Description of LC/MS/MS Instrument and Operating Conditions Instrument: API 4000 Biomolecular Mass Analyzer Interface: Turbo Ion Spray Liquid Introduction Interface Computer: DELL OptiPlex GX400 Software: Windows NT, Analyst 1.4.1 HPLC: Hewlett Packard (HP) Series 1 00 HP Quat Pump HP Vacuum Degasser HP Autosampler HP Column Oven HPLC Column: Thermo Fluophase RP, 50 mm x 2.1 mm Column Temp.: ~30 C Injection Voi.: 15 pL Mobile Phase (A): 2 mM Ammonium Acetate in water Mobile Phase (3): Methanol Exygen Research Time (mini 0.0 1.0 8.0 10.0 11.0 18.0 %A 65 65 25 25 65 65 %B 35 35 75 75 35 35 Page 16 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Total run time: ~18 min Flow Rate: 0.3 mL/min Ions monitored: Analvte PFOA PFOA Confirm Ion 13C PFOA Mode negative negative negative Transition Monitored 413 -> 369 413 -> 219 415->370 Approximate Retention Time (min) --11.1 min. -11.1 min. -11.1 min. 6.9 Quantitation and Example Calculation PFOA concentrations are quantitated in nanogram o f PFOA per milliliter o f extract and then converted to nanogram per gram. The PFOA concentrations were quantitated in nanogram o f PFOA per liter for Set #3 (Set 042806D). Fifteen microliters o f sample or calibration standard were injected into the LC/MS/MS. The peak area was measured and the standard curve was generated (using 1/x fit weighted linear regression) by Analyst software using seven concentrations o f standards. The concentration was determined from the equations below. Please note that the equations in this section are presented in a different order than the equations in the method (V0001782), but this has no effect on the final calculations. Equation 1 calculated the amount o f analyte found (in ng/L, based on peak area) using the standard curve (linear regression parameters) generated by the Analyst software program. Equation T. Analyte found (ng/L) = (Peak area - intercept! x DF Slope Where: DF = Dilution Factor, factor by which the final volume was diluted, if necessary. For samples fortified with known amounts of PFOA and 13C PFOA prior to extraction, Equation 2 was used to convert the amount o f PFOA found in ng/L to ng/g (ppb). Equation 2: PFOA found (ppb) = fPFOA found (ng/L) x SPE volume extracted! [sample aliquot volume x (sample weight / direct inject sample volume)] Exygen Research Page 17 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Equation 3 was used to calculate the percent recovery. Equation 3: Recovery (%) = (analyte found (ng/g. wet weight-) - analyte in control (ng/g. wet weight)-) xlOO% amount added (ng/g, wet weight) Note: For the PFOA recovery calculation, the "control" is the unspiked aliquot of the primary field sample. Equation 4 was then used to calculate the amount of PFOA found in ppb based on dry weight. Equation 4: PFOA found (ppb) dry weight = PFOA found (ppb) x [ 100% / total solids(%)] An example o f a calculation using an actual sample follows: Sediment sample Exygen ID CO172892 Spk D (Set: 042806D), fortified at 4 ng/mL with PFOA where: peak area = 236588 intercept - 4630 slope = 420 dilution factor = 100 ng/g PFOA added (fort level) = 4000 amt in corresponding sample (ng/g) - 586 SPE volume extracted (L) -- 0.04 sample aliquot volume (mL) =5 sample weight (g) =1 direct inject sample volume (mL) = 10 total solids (%) = 66.54 From equation 1: Analyte found (ng/L) From equation 3: PFOA found (ppb) - 1236588 - 46301 x 100 420 *= 55200 ng/L 155200 ne/L x 0.04 L) [5mLx(l g/10mL)) = 4420 ppb From equation 3: % Recovery = (4420 ng/g - 586 ng/g) x 100% 4000 ng/g - 96% Exygen Research Page 18 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 From equation 4: PFOA found (ppb) dry weight = 4416 ppb x (100% / 66.54%) = 6640 ppb NOTE: This value may be slightly different than that of the raw data due to rounding. 7.0 EXPERIMENTAL DESIGN 13C PFOA was used as a surrogate for all the samples. 13C PFOA was added to the sediment samples in the laboratory after the samples were weighed for extraction. For sediment samples designated as laboratory matrix spikes, PFOA was also added at a known concentration to the samples in the laboratory after the samples were weighed for extraction. The sediment samples were extracted in one set. The set included one reagent blank and two reagent blanks fortified at known concentrations. The set contained three sediment samples. For each sample, two laboratory matrix spikes and a duplicate were extracted. 8.0 RESULTS Analytical results for the analysis o f PFOA in sediment samples based on dry weight are summarized in Table I. Analytical results for the analysis o f PFOA in sediment samples by direct injection, based on dry weight, are summarized in Table II. Analytical results for the analysis o f PFOA in sediment samples by alternative SPE extraction, based on dry weight, are summarized in Table III. The overall average percent recovery standard deviation for PFOA in sediment samples based on wet weight was 73 16%. Fortification recoveries for the analysis o f PFOA in sediment samples are summarized in Table IV. Fortification recoveries for the analysis o f PFOA in direct injection sediment samples are summarized in Table V. Fortification recoveries for the analysis o f PFOA in SPE extracted sediment samples are summarized in Table VI. The average percent recovery standard deviation for l3C PFOA in sediment samples based on wet weight was 97 27%. Fortification recoveries for 13C PFOA in sediment samples based on wet weight are summarized in Table VII. Fortification recoveries for 13C PFOA in direct injection sediment samples based on wet weight are summarized in Table VIII. Fortification recoveries for I3C PFOA in SPE extracted sediment samples based on wet weight are summarized in Table IX. The total percent solids for sediment samples are detailed in T able X. Exygen Research Page 19 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 9.0 CONCLUSIONS The sediment samples were successfully extracted and analyzed for PFOA using an alternative SPE method, which yielded acceptable QC data. 10.0 RETENTION OF DATA AND SAMPLES All original paper data generated by Exygen Research that pertains to this interim report will be shipped to the study director. This does not include facility-specific raw data such as instrument or temperature logs. Exact copies o f all raw data, as well as a signed copy o f the interim analytical report and all original facility-specific raw data, will be retained in the Exygen Research archives for the period o f time specified in EPA TSCA Good Laboratory Practice Standards 40 CFR 792. Exygen Research Page 20 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 TABLES Exygen Research Page 21 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table I. Summary of PFOA in Sediment Samples Exygen ID C0f72892 C0172892 Rep C017283 CO172893 Rep C017284 C 0 172894 Rep Client Sample ID DAL-SO-BPPO VO-OOOO D A L -S D -B P P 0 1 -0-0000* DAL-S D-BPP02-0-0Q00 DAL-S D-B P P02-0-0000* DAL-SD-BPP03-tHK)00 DAL-SD-B PP03-0-0000* Analyte Found (ppb, ng/g) Dry Weight C8 Acid PFOA Perfluoreoctanoic Acid NR NR NR NR NR NR Assessed Accuracy ( '*%> NR NR NR NR NR NR 'Laboratory Duplicate NR = Not reported due to quality control result failures. See Table II for re-Hnatysls results. Exygen Research Page 22 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table II. Summary of PFOA in Direct Injection Sediment Samples Exygen ID C0172892 C 0 172892 Rep C 0 172893 C0172893 Rep C0172894 CO l72894 Rep Client Sample ID D A L -S D -B P P 0 1 -0-0000 D A L -S D -B P P 0 1 -0-0000' DAL-SD-BPP02-0-0000 DA L-S D-B PP02-0-0000* DAL-SD-BPP03-0-0000 DAL-SD-BPP03-0-0000* Analyte Found (ppb, ng/g) Dry Weight C8 Acid PFOA Periluorooctanolc Acid NR NR NR NR NR NR Assessed A ccu racy (+ /-% ) NR NR NR NR NR NR ' Laboratory Duplicate NR * Not reported due to quality control result failures. See Table III for re-analysis results. Exygen Research Page 23 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table III. Summary of PFOA in SPE Extracted Sediment Samples s=lSSSi^= 00172662 C0172892 Rap CO172893 C0172893 Rep C0172894 C0172894 Rep 'laboratory Dupllcste Client Sample ID Analyte Found (ppb, ng/g) Dry Weight CB Acld PFOA Perfluorooctanolc Add DAL-SD-BPP01-0-0000 QAL-SD-BPP01-0-0000' DAL-SD-BPP02-0-0000 DAL-SD-BPP02-0-0000' 881 945 627 823 DAL-SD-BPP03-0-0000 DAL-SD-BPP03-0-0000' 2390 2380 Aaaatted Accuracy <*-%> 30 30 30 30 30 30 Exygen Research Page 24 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table IV. Matrix Spike Recovery of PFOA in Sediment Samples Sample Description DAL-SD-BPP01 -0-0000 (C01728M S pk , 400 ppb ap t ke) DAL-SO-BPP01-0-0000 (C M 7 2 M 8 p k E, 4000 ppb Spike) DAL-SD-BPP02-0-0000 (C0172SM S pk G, 400 ppb Spike) DAL-SD-BPP02-0-0000 (C0172M3 spk M, 4000 ppb Spikft) DAL-SQ-BPP03-0-0000 (C01728M spk J. 400 ppb Spike) DAL-SD-BPP03-0-0000 (G0172804 Spk K, 4000 ppbSpike) Amount Spiked (ng/g) 400 4000 400 4000 400 4000 Ami Found in Sample (ng/g) wet weight C8 Add PFOA Amount Recovered (ng/g) wet weight NR NR NR NR NR NR NR NR NR NR NR NR Recovery f*) NR NR NR NR NR NR Avaraga: Standard Deviation: NR NR `Sample residue exceeds the spiking level significantly ('is spiking level); therelore, an accurate recovery value cannot be calculated. NR - Not reported due to quality control result failures. See Tabi V for re-analysis resulta. Note: S in e * this sum m ary tab le shows rounded results, recovery values may vary slightly from the values In the raw data. Exygen Research Page 25 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table V. Matrix Spike Recovery of PFOA In Direct Injection Sediment Samples Sample IOBscftj)tjor^_ DAL-S D-BPP01-0-0000 (C01778B2 Spa D, 4000 ppb Spills) DAL-SD-BPP01-0-0000 (00172192 8 pit E, 40000 ppb Spike) D A L -S D-BPP02-0-0000 (00172993 Spk G, 4000 ppb Spike) DAL-SD-B PP02-0-0000 {C01723M S pk H. 40000 ppb Spike) DAL-S D-BPP03-0-0000 (00172894 Spk J, 4000 p p b Spike) DAL-S D -B P P 0 3 -0 -0 0 0 0 (C0172S94 Spk K, 40000 ppb Spike) Amount Spiked ...W O ) 4000 40000 4000 40000 4000 40000 Am t Found in Sample (ng/g) we* weight C8 Acid PFOA Amount Recovered (ng/g) wet welqht NR NR NR NR NR NR NR NR NR NR NR NR Reoovery (%> NR NR NR NR NR NR Average: Standard Deviation: NR NR 'Sample residue exoeeds the piking level significantly (3x spiking level); therefore, an aocurate reoovery value cannot be calculated NR Not reported due to quality control result failures See Table VI for re-analysis results. Note: SI nee this sum m ary table shows rounded results, recovery values m ay vary slightly fro m the values In the raw data. Exygen Research Page 26 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table VI. Matrix Spike Recovery of PFOA in SPE Extracted Sediment Samples 1 11 1 Sample Description DAL-SD-BPP01-0-0000 (C0172SS2 Spk D. MOO nfl/B Splka) DAL-S D-BPPO1-0-0000 (CM 7JIS2 Spk E, 40000 n r t Splka) DAL-SD-BPP02-0-0000 (C M 7 *S Spk G, 4000 np/a Splka) DAL-SD-8PP02-0-0000 (C0172893 S pk H. 40000 n t f j SpiX*> DAL-SD-BPP03-0-0000 (C9172904 Spk J, 4000 ng/g S p lk*) DAL-SD-BPP03-0-0000 (C0172994 Spk K. 40040 ng/g Splka) Amount Spiked (ns/gl Amt Found in Sample (fin/a) wet weighl CB Acid PFQA Amount Recovered inc/g) wet weight 4000 40000 586 586 4420 25600 4000 40000 4000 40000 504 504 1060 1060 3860 25100 4140 23200 Average: Standard Deviation: Readvary {*> 96 83 84 61 77 55 73 16 N o t: S in e , th is um m ary ta b ), ih o w t rou n d e d rM u lts , recovery va lu e * m ay vary s lig h tly fro m the v a lu e . In the raw date. Exygen Research Page 27 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table VII. Surrogate Spike Recovery of ,3C PFOA in Sediment Samples Exygen ID Sample Description C0172892 COI 72892 Rep C 0 172892 Spk C CO172892 Spk D C0172892 Spk E C0172893 C 0 172893 Rep C 0 172893 Spk F C 0 172893 Spk G C0172893 Spk H C 0 172894 CO172894 Rep CO172894 SpkI C0172894 Spk J CO172894 Spk K D A L-SD -B PP01-0-0000 D A L -S D -B P P 0 1-0-0000 DAL-S D-BPP01-0-0000 DAL-SD-BPP01-0-0000 D A L-SD -B PP01-0-0000 DAL-S D-BPP02-0-0000 DAL-S D-BPP02-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-BPP02-0-0000 DAL-S D-BPP02-0-0000 DAL-S D-BPP03-0-0000 DAL-S D-BPP03-0-0000 DAL-S D-BPPQ3-0-0000 DAL-SD-BPP03-Q-0000 DAL-S D-BPP3-0-0000 Amount Spiked (ng/g) 4 4 4 400 4000 4 4 4 40.0 4000 4 4 4 400 4000 1sC-P FO A Amount Recovered Wet Weight {ng/g) NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR Recoveiy (%) NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR Average: Standard Deviation: NR = Not reported due to quality control result failures. See Table VIII for re-analysis results. Note: Since this summary table shows rounded results, recovery values may vary slightly from the values in the raw data. NR NR Exygen Research Page 28 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table VIII. Surrogate Spike Recovery of 13C PFOA in Direct Injection Sediment Samples Exygen ID Sample Description C 0 172892 C 0 172892 Rep C 0 172892 SpkC C 0 172892 Spk D C0172892 Spk E C0172893 C 0 172893 Rep C 0 172893 Spk F C 0 172893 Spk G C 0 172893 Spk H C 0 172894 C 0 1 72894 Rep C 0 172894 Spk I C0172894 Spk J C 0 172894 Spk K D A L-S D -B P P 0 1-0-0000 DAL-SD-BPP01-0-0000 D A L-S D -B P P 01-0-0000 D A L-S D -B P P 01-0-0000 D A L-S D -B P P 01-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-8PP02-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-BPP03-0-0000 DAL-SD-BPP03-0-0000 DAL-SD-8PP03-0-0000 DAL-SD-BPP03-0-0000 DAL-SD-BPP03-0-0000 Amount Spiked wg) 40 40 40 4000 40000 40 40 40 4000 40000 40 40 40 4000 40000 1sC-P FO A Amount Recovered Wet Weight (ng/g) NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR Recovery (%) NR NR NR NR NR NR NR NR NR NR NR NR NR NR NR A v a ra g a : Standard Deviation; NR = Not reported due to quality control result failures. See Table IX for re-analysis results. Note: Since mis summary table shows rounded results, recovery values may vary slightly from the values In the raw data. NR NR Exygen Research Page 29 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table IX. Surrogate Spike Recovery of ,3C PFOA in SPE Extracted Sediment Samples Exygen ID C0172RS2 C0172892 Rep C0172892 Spk D C0172892 Spk E C0172893 CO172893 Rep C0172893 Spk G C0172893 Spk H CO172894 C0172894 Rep C0172894 Spk J C0172894 Spk K Sample Description DAL-SD-BPP01-0-0000 DAL-S D-BPP01 -0-0000 DAL-SO-BPP01 -0-0000 DAL-SD-BPP01-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-8 PP02-0-0000 DAL-SD-BPP02-0-0000 DAL-S D-B PP02-0-0000 DAL*SO-BPP03'0-0000 DAL-SD-BPP03-0-0000 DAL-SD-BPP03-C-0000 DAL-SD-BPP03-0-0000 Amount Spiked (ns'a) 40 40 4000 40000 40 40 4000 40000 40 40 4000 40000 1lC-PFOA Amcunt Recovered Wei Weight ("aw 52.0 51 0 3750 25800 47.8 51 a 3500 25200 45.2 40.3 3060 22100 Note: Since this summary table show rounded results, recovery values may vary slightly from the values In the raw data. Avaraga: Standard Oavlatlen; Recovery (%) 130 128 94 65 120 129 86 53 113 101 77 55 07 27 Exygen Research Page 30 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Table X. Total Percent Solids in Sediment Samples Exygen ID CO172892 C0172893 CO172894 Sample Description DAL-SD-BPP01-0-0000 DAL-SD-BPP02-0-0000 DAL-SD-BPP03-0-0000 Total Solids (%) 66.54 54.37 44.46 Exygen Research Page 31 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 FIGURES Exygen Research Page 32 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Figure 1. Typical Calibration Curve for PFOA in Methanol 0 4 2 9 0 0 0 Sd. SPE.rdb (PFOA> " U M i r Rg*ion f 1 weighting): y 4 2 0 4 C 3 r K ) 0 3 O WOO) Area, counts Exygen Research Page 33 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Figure 2. Extracted Standards of PFOA in Methanol, 25 ng/L and 50 ng/L, Respectively 1 X 0426*4-1 - PFOA (S ta tt* n)) 41X 400.4 n u r -aampi* 2 o f 4 t from U T tO taw fff A n * : 112M *o**t* Hofgrr. 1 .1 0 4 it* icp * KT: 11.4m m 11.01 1106 Exygen Research Page 34 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Figure 3. PFOA in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B Respectively I R t a n t C o n tr o l-P F O A < ftoktow w )4n.M SS% am a - ta m p I t 1 a f J i f t o m M 2tO CO .tr/ff a o t fO lU H t) ( <->. cU a> H I 37A rt : 3 0 2 9 9 count* Hoth t: 2 o-KX op HT: 11.0 min 11.00 2.04 caLn> f 10. 4 C &c 0 0 -------- 1 -- T I I - - I 12 3 45 ---------- r 07 11.00 Time, min Exygen Research Page 35 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Figure 4. Chromatogram Representing a Sediment Sample Analyzed for PFOA (Exygen ID: C0I72892, Data Set: 042806D) C t m n 2 PFOA $ to t * c v r * ) #1 1 1 X 0 0 1 * J N ( - t t m p l * l 9 f 4 * f r o m frO M E C L M tr A m : 35421 c c u j t r H tJ ftt: T tt*003cp * RTi 11.0min 1 1 ,0 5 intensity, cps Exygen Research Page 36 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Figure 5. Typical Calibration Curve for ,3C PFOA in Methanol 0 4 2 0 0 0 0 S ta . S P E .rib C13C P F C A i U n til" R tg rta le n / y wtlgM Jrg): y 480 x + 0 .0 0 10 (i - 0 .0 0 0 0 ) Area, counts Exygen Research Page 37 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygcn Study No.: P0000760 Figure 6. Extracted Standards of 13C PFOA in Methanol, 25 ng/L and 50 ng/L, Respectively X C U 1 U U 1 . 13C P F O A (SU rnfm/tf)411$G70l9 mmu - s i m p * 2 o f 46 front V42$0VXwM Aron: 1Td01 cnnnta f h ig A t : 1 . tf * * C 0 3 c p r f t 7 : 11.$ m in 11.02 tntensiti'.cps Are: 26099 oounfc* Height: 2.*t0e+QQ3 op RT:11.1 min 11.00 Intensrly, cps Exygen Research Page 38 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Figure 7. l3C PFOA in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively R to g o a t C o n tro l t3 C P f O A (Uokmowm) 4 0 .0 1 3 7 * 0 am u -oa mpfo t o r n fro m M2WCXwtW p o a k ( fo u n d ) 10 3 a T im *, min I R f j g a n t Spk A - 13C PFOA (OC) 413 .0/370 0 j it h j - l a m p l t 10 o f 40 from 042 8000 w ltf A r* j:2 7 2 9 G oounts Ht i g h t 2.02 +03 c p f RT: 11.0 min 1 i^J1 1UD7 Exygen Research Page 39 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No,: P0000760 Figure 8. Chromatogram Representing a Sediment Sample Analyzed for 13C PFOA (Exygen ID: C0172892, Data Set: 042806D) Intensity, cps I 0*72*92 * 13C PFO A (Uhttow*) lir a -s a m p b 2 lo f4 6 T r* m O tttiia w tfT A rt* : 3M 937 co a n tt H tlgAL Z 9 U + 0 9 4 tp i RJ; 11,1 m in 2.9*4 2 0*4- 1107 2 4*4 2,2*4. 20*4- 1 -8*4 - 1.6 *4 1.4*4- 1 .2 *4 - 1.0*4eooo.o OOOOO 4000 O 2000.0 - 0 . 0 ---- ----- 1---- ------r---- ----- .---- ---- ---- ---------------i---- -----1---- ------r1234607B Tim, mil Exygen Research Page 40 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 APPENDIX A Study Protocol P0000760 (Exygen Study No. P0000760) with Analytical Method and Protocol Amendments Exygen Research Page 41 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P0000760 STUDY PROTOCOL Study Title: Analysis of Perfluorooctanoic Acid (PFOA) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program Exygen Protocol Number: P0000760 Performing Laboratory: Exygen Research 3058 Research Drive Stale College, PA 16801 Phone: (814)272-1039 Sponsor Representative: Michael A. Santoro Director o f Regulatory Affairs 3M Building 0236-01-B-10 St. Paul, MN 55144 Phone: (651)733-6374 P agt I o j S5 Exygen Research Page 42 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Num ber: P0000760 DISTRIBUTION: 0 Jaiaimha Keiari, Study Director, Weston Solutions 2) John M. Flaherty, Principal Investigator, Exygen Research 3) Michael A. Santoro, Sponsor Representative, 3M Company 4) Exygen Research Quality Assurance Unit Exygen Research Page 7 o ]6 i Page 43 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygco Protocol Number P0000760 PROTOCOL APPROVAL Study Title: Analysis o f Perfluorooctanoic Acid (PFOA) in Water, Soil, Sediment, Fish. Clams, Vegetation, Small Mammal Livers and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program Exygen Protocol Number: P00007i0 APPROVALS JaismXa Kcsaxi, Study Director Weslon Solutions ' u LytKa Shaffer, Technii /gen Research Quality Assurance Unit Date g f? -< 3 X S lO O 'J Dan P a fu 3 o f 63 Exygen Research Page 44 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: ROOOG700 TABLE OF CONTENTS TITLE PAGE-------------------------------- ------- ------------------ ....... - ..... ................... - ......... ....... 1 DISTRIBUTION_________________________ ___ ____ ___________ _______________ 2 PROTOCOL APPROVAL--------- ----------------------------------------------- - --------- ------- -- .......... 3 TABLE OF CONTENTS _______________________ ___ ------- ------------- ---- ......... .......--4 INTRODUCTION-------------------------------------------- ....... ....... .... ........... ....... ....- ...................5 TEST MATERIAL........................... .................... .................................. ........... ......................- 5 SURROGATE FIELD SPIKE COMPOUND__________ __ _______________ ........................... J OBJECTIVE________ __ __________ _____ ___ ______________________ ________--6 TESTING FACILITY____________ ________ ______ -- ------------------ ------------- ............-.6 STUDY DIRECTOR................................................ ..... ..................-- ----------------------------- --- 6 SPONSOR REPRESENTATIVE.... _.............- ................. ...... ........-- ......... ............................... 7 PRINCIPAL INVESTIGATOR......................... ................................................... ............. ...........7 PROPOSBD EXPERIMENTAL STARTAND TERMINATION DATES........................ ......................7 IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM.............. ................ ........ ........ 7 SAMPLE PROCUREMENT, RECEIPT AND RETENTION -- ......................................................... 8 SAMPLE IDENTIFICATION................................ ........ ............. .......... - ........... - ...................... S ANALYTICAL PROCEDURE SUMMARY__________ __ ____ ____ _____ - ..................... ....... VERIFICATION OF ANALYTICAL PROCEDURE...................... ................... ......- ......................5 METHOD FOR CONTROLOF BIAS---------------------- ------- .......................................................11 STATISTICAL METHODS___________________ _________ ____ _______ ____ ______ 11 GLP STATEMENT---- --------------------------------------------------- --------- ------------------------------ 11 REPORT----- --- -------- ------------------ ------------------------------------- ----------------------- ---------- 11 SAFETY AND HEALTH____________________ __ _________ _______________ __ ___ 12 AMENDMENTS TO PROTOCOL----- --------------------------------------------- ..... ......... ............13 DATA RECORD KEEPING -- ___ ______________ -- ...... - ........... .............................. 13 QUALITY ASSURANCE____ _______ _________ ___________________ _______ ____14 RETENTION OF DATA AND ARCHIVING______________ __ ________ ____ ___ _______ 14 APPENDIX I. ANALYTICAL METHODS------- --- ------------------------ --------------------- ----- ----- - IS P a0 4 o /6 3 Exygen Research Page 45 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygoe Protocol Number: P0000760 INTRODUCTION The purpose of this study is to perform analysts for perfluorooctanoic acid (PFOA) in water, soil, sediment, fish, clams, vegetation, small mammal livers and small mammal serum using LC/MS/MS for the 3M Decatur Monitoring Program. The study will be audited for compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792 by the Quality Assurance Unit of Exygen Research. TEST MATERIAL The test material i8 perfluorooctanoic acid (PFOA) and was purchased from Sigma-Aldrich. PFOA Chemical Name: Perfluorooctanoic acid Molecular Weight: 414 Lot Number. 23116HB Purity: 97.64% Transitions Monitored: 413 -* 369 (for quantification) and 413 -219 (for confirmation) Structure: SURROGATE FIELD SPIKE COMPOUND The surrogate field spiking compound is l,C labeled perfluorooctanoic acid (a C PFOA) and was purchased by 3M from Peridn-Ehner Life and Analytical Sciences. " C PFOA Chemical Name: 1.2-13C perfluorooctanoic acid Molecular Weight: 416 Lot Number. 3507-195 Purity. 97% Transition Monitored: 415 -*370 P agt 5 o f65 Exygen Research Page 46 o f l 10 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P000G760 o b jec tiv e The purpose of this itudy it to perform analysis for perfluorooctanoic acid (PFOA) in water, toil, sediment, fish, clams, vegetation, small mamma) livers and small mammal serum for the 3M Decatur Monitoring Program using the current versions of the following Exygen analytical methods: V0001780 V0001781 V0001782 V0001783 V00C1784 VOM 785 VOOOl 786 "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" "Method o f Analysis far the Determination o f Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Fish and G am s by LC/MS/MS" "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Smell Mammal Liver by LC/MS/MS" "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" TESTING FACILITY Exygen Research 3058 Research Drive State College, PA 16801 Phone: (814)272-1039 STUDY DIRECTOR Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way W est Chester, PA 19380 Phone:(610)701-3761 Fax: (610) 701-7401 j.kesari@weston*olutions.coen P agi 6 of65 Exygen Research Page 47 o f 110 Interim Report #23 --Analysis of Sediment Samples Exygen Study No.: P0000760 BxygtmProtocol Number P0000760 SPONSOR REPRESENTATIVE Michael A. Santoro 3M Company Director o f Regulatory Affair* 3M Building 0236-01-B-10 St. Paul, MN 55144 Phone: (651)733-6374 PRINCIPAL INVESTIGATOR John M Flaherty Exygen Research 3058 Research Drive State College, PA 16801 Phone:(814)272-1039 john.flaherty@exygen.com PROPOSED EXPERIMENTAL START AND TERMINATION DATES U is proposed that the analytical portion o f this study be conducted from October 01, 2004 to December 31, 2005. The actual experimental start and termination dates will be included in the final report. IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM The following are tho test systems for this study: Water (groundwater and surface water) Soil a Sediment Fish Clams Vegetation Small Mammal Liver Small Mammal Serum Tho samples will be collected by Weston Solutions. The control samples will be purchased and prepared by the testing facility. Purchase and processing P ag* 7 v/05 Exygen Research Page 48 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P0000760 detail* for the control sample* will be included in foe final report associated with this study. The test system were chosen to access the environmental impact o f PFOA in foe Decatur, Alabama area. SAMPLE PROCUREMENT, RECEIPT AND RETENTION Water, soil, sediment, fish, clam, vegetation, small mammal liver and small mammal scrum samples will be received at Exygen directly from Weston Solutions. The details o f simple procurement for this study are outlined in the 3M work plan entitled "Phase 2 Work Plan for Sampling Environmental Media." The number and type* o f samples collected will vary depending availability in foe field. The total number o f samples received and analyzed for each matrix will be documented in foe final report associated with this study. Water, soil, and sediment samples will be used as received without further processing at Exygen. These samples will be stored refrigerated at 2C-8C. Fish, clam, vegetation and small mammal liver samples will be processed according to the appropriate analytical method (fee Appendix I). These samples will be stored frozen at i -ICC. Small mammal whole blood samples will be centrifuged in the field at the time o f collection and the serum fraction will be used for the study. Small mammal serum will be stored frozen at -10*C. The receipt and procesaing o f the sample* will be documented in the final report and raw data associated with the study. SAMPLE IDENTIFICATION Prior to analysis, each sample will be assigned a laboratory sample reference number. The reference number will be unique and will distinguish each laboratory sample that is processed throughout the analytical procedure. Chromatographic data will be identified by the laboratory sample reference num ber. Sample storage conditions and location will be documented throughout the study. Page 8 o f 65 Exygen Research Page 49 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen P rotocol N u m be r POOOCT760 ANALYTICAL PROCEDURE SUMMARY Reference: VOOOI7SO: "Method o f Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V00017B!: "Method o f Analysis for the Determination o f Perfluoroocianoic A dd (PFOA) in Soil by LC/MS/MS" V0001782: "Method o f Analysi* for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method o f Analysis for the Determination of Perfluoroocianoic A dd (F?OA) in Fish and Clams by LC/MS/MS" V0001784: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Vegetation by IX/MS/MS" V0001785: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" The above methods use analytical conditions capable of separating the iaomers o f PFOA. The final report will include the isomers summed into total PFOA found. VERIFICATION OF ANALYTICAL PROCEDURE A laboratory control sample will be used for the preparation of fortified control samples. The test substance will be made into solutions as per the method, and added to the matrices via a micropipette. For water sampling, Exygen will supply one bottle per sample collected. The bottles will be 500 mL precleaned Sci/Spec Premier wide mouth HDPE bottles. These bottles have been routinely used for PFOA sample collection at the testing facility and have been shown to be free o f PFOA. All containers used for water sample collection will be shipped to the sample location containing 100 ng of C PFOA (equivalent to 500 ng/L in the final sample). Samples w ill bo added to each container So a volumetric f ill lin e at 200 mL. A field duplicate, a low field spike rod a high field spike o f each sample will be collected. The low and high fiekl spikes will contain PFOA as well as perfluoiobutanesulfbnste (PFBS), perfluorohexanesulfonate (PPHS), and perfluorooctanesulfonate (PFOS). These compounds are included in the solutions used to spike the simples. The results for PFBS, PFHS and PFOS will not be reported in this study. Exygen will supply one field blank (control water) and two field blank spikes (control water fortified with PFOA at a low Page 9 o f 65 Exygen Research Page 50 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exyfttn Protocol Number: P0000760 and high level) for every twenty samples collected. At the testing facility, each water sample (excluding field duplicates and field spikes) will he extracted in duplicate and will also be fortified at a low and high concentration with PPOA and processed through the described procedure to determine method accuracy and to chock for bias For soil, sediment, d im s, and vegetation, Exygen will supply one 500 mL precleaned Sci/Spcc Premier wide mouth HDPE bottle per sample collected or a zip-seal bag. All conlainen/bags used for sample collection will be shipped to the sample location. Sample will be added to each container or bag in the field.' At the testing facility, each sample will be extracted in duplicate and will also be fortified at a known concentration with PFOA at both a low and high level and procened through the described procedure to determine method accuracy and to check for bias. n C-PFOA will also be added to each sample in the laboratory prior to extraction at the level specified below. For small mammal liver, Exygen will supply a $0 mi- polypropylene contriiUgc tube. For small mammal serum, Exygen will supply a collection kit for each sample containing serum separator tubes (red top), vacutainers. needle holders and needles, transfer pipettes, and polypropylene tubes. At the testing facility, each fiver and serum sample will be extracted in duplicate and will also be fortified at a known concentration with PFOA at both a low and high level and processed through the described procedure to determine method accuracy and to check for bias. l3C-PFOA will also be added to each sample in the laboratory prior to extraction at the level specified below. Low and high spiking levels for each matrix are defined below: Matrix Low PFOA High PFOA "C-PFOA Spiking Level Spiking Level Spiking Level Water 500ng/L 5000 ng/L 500 ng/L Soil 4 ng/g 40 ng/g 40 ng/g Sediment 4 ng/g 40 ng/g 40 ng/g Fish 10 ng/g 100 ng/g 100 ng/g Clams lOngtg 100 ng/g 100 ng/g Vegetation 10 ng/g 100 ng/g 100 ng/g Small Mammal Liver 10 ng/g 100 ng/g 100 ng/g Small Mammal Serum 10 ng/g 100 ng/mL 100 ng/mL Recoveries we anticipated to be between 7094 and 13094 of the fortified levels; however, the exact precision and accuracy will be determined by the analysis o f the quality control samples described above. A statement of accuracy will be included in the final report. Priot to sample analysis, a laboratory control spike study will be conducted in water to justify the use o f 13C PFOA results to establish precision and accuracy of PFOA samples. The following samples are to be prepared: Page 10 oj 65 Exygen Research Page 51 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P00OO76O Two control water blank* Two control water samples fortified at 250 ng/L with UC PFOA Two control water samples fortified at 250 ng/L with n C PFOA and at 250 ng/L with PFOA. Two control water samples fortified at 5000 ng/L with n C PFOA and at 5000 ng/L with PFOA. The above samples are to be extracted and analyzed according to method V0001780 entitled "Method o f Analysis for the Determination of Periluorooctanoic Acid (PFOA) in Water by LC/MS/MS.* METHOD FOR CONTROL OF BIAS Control o f biaa will be addressed by taking representative sob-samples from a homogeneous mixture of each matrix fiom untreated control samples, and by analyzing at least two levels of fcrtificatious. STATISTICAL METHODS Statistics will be limited to those specified in the subject methods and to the calculation of average recoveries, as applicable. GLP STATEMENT All aspects o f this study shall be performed and reported in compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792. The final report or data package (supplied to the Sponsor) shall contain a statement that the study was conducted in compliance with cunent and applicable GLP standards and will outline any deviations in the study from those standards. This statement will be signed by the Study Director and Sponsor Representative. REPORT A final report will be prepared by the principal investigator or their designee at the conclusion of the study. The report will include, but will not be limited to, the following: Page I t o f 65 Exygen Research Page 52 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen P rotocol Num ber: P0000760 The name and address of the Study Director. Sponsor Representative, and o f the testing facility. A statement o f GLP compliance (any related documentation, such as chain-of-custody records, moat he in the study records). The signed and dated statement by the Bxygea Research Quality Assurance Unit regarding dates o f study inspections and dates findings were reported to the Study Director and Management. A description of the exact analytical conditions employed in tho study. If the subject method was followed exactly, it is necessary to include only a copy o f tho analytical method. Any modifications to this method will be incorporated into the report. If the method is photo-reduced, the project number and page number must be included on each page. Description o f the instrumentation used and operating conditions. All results from all sets analyzed. Control and fortified samples will be identified and the data table will include sample number and fortification level. Representative chromatograms for each analyte in each matrix, including chromatograms of a standard and a control sample, and a chromatogram at a fortification level. H ie location o f the analyte peaks wilt be clearly identified in all chromatograms. Alt circumstances that may have affected the quality or integrity o f the data will be documented in the report Locations where raw data and the final report are to bo archived. Additions or corrections to the final report shall be in the form o f an amendment signed by the Study Director. The amendment shall clearly identify that part of the report that ia heing altered and the reasons for the alterations. The amendment will be signed and dated by the Study Director and the Sponsor Representative. SAFETY AND HEALTH Laboratory personnel will practice good sanitation and health habits. Every reasonable precaution shall be taken to prevent inadvertent exposure of personnel and the environment to the test or reference substances). Page 2 o f 62 Exygen Research Page 53 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P00007&Q /-- AMENDMENTS TO PROTOCOL All significant change to the analytical protocol outlined here will be exprciaed in writing, signed and dated by the Study Director and Sponsor Representative. Amendment uiually will be iaauod prior to initiation o f study plan change. However, when a change ii required without sufficient time for the issue of a written amendment, that change may be effected verbally with supporting documentation signed and dated by the Study Director and followed with a written amendment aa soon aa passible. Id this case, the effective date of the written amendment wilt be the date o f the documented change. Copies o f the signed amendments will be appended to ail distributed study plan copies. The original amendment will be maintained with the original study plan. Any deviations from the study plan or from the analytical method as provided will be documented and reported promptly to the Sponsor Representative. DATA RECORD KEEPING Records to be maintained include the following (as appropriate): Sample tracking sheet(s) Sample receipt records, storage history, and chants o f custody History and preparation o f standards (stock, fortification, calibration) Description o f any modifications to the method Instrument run sheets, bench-sheets or logs > Analytical data tables All chromatographic and instrumental conditions Sample extraction and analysis dates A complete listing o f study personnel, signaturea and initials Chronological presentation of all study correspondence Any other documentation necessary fbr the reconstruction of the study Chromatograms- Ail chromatograms will contain the following: Sample identification, injection date, arrow or other indication o f the area of interest, and injection number corresponding to the run. Additionally, fortifications will include the amount o f analyte added and the sample number of the simple that waa fortified. Analytical standard chromatograms will additionally include the concentration (e.g., pg/mL). Pag* /J o f 65 Exygen Research Page 54 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygec Protocol Number: P0000760 As part of the documentation the following sheets will be included in each analytic! set: a nut sheet listing the samples to be run in the set. and an instrument conditions sheet describing the instrument type and operating conditions. All applicable requirements for reporting o f study results ss per 40 CFR 792.185. QUALITY ASSURANCE The QA Unit of Bxygen Research will inspect the study at intervals adequate to assure compliance with GLP'i, and will report the foldings o f audits to the Study Director, Exygen Management, and the Sponsor Representative. RETENTION OF DATA AND ARCHIVING All hard copy raw data, including, but not limited to, the original chromatograms, worksheets, correspondence, and results shall be included with the data package submitted to the Study Director. These will be archived with the original study plan, intendments, foul report, and all pertinent information from the Sponsor. The testing facility shall keep all electronic raw data and any instrument, equipment, and storage logB for the period o f time specified in 40 CFR 792.195. An a c t copy of the materials submitted to the study director will also be kept at Exygen Research. Exygen will obtain permission ftom the itudy director before discarding or returning samples. Exygen Research P o g t 14 0 / 6 ) Page 55 o f l 10 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 APPENDIX I ANALYTICAL METHODS V0001780: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781: "Method o f Analyaii for the Determination o f Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" V0001782: "Method o f Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS" V0001784: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in 3mall Mammal Liver by LC/MS/MS" V0001786: "Method of Analysis for the Determination of Porfluorooctanoic Acid (PFOA) in Small Mammal Scrum by LC/MS/MS" Exygen Research P age I ) o f 6S Page 56 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 ANALYTICAL METHOD M rtM dNw nfer VW017SO M c*fc<4fAJyib fo rth * Dl*nijiaa<iaa a t F n flu a ra a ctw o k Acid (7TOA>la Water kyLC M M M i Analytical Turinj Facility: E*y#en Jlcaeatcii 3051 Jbacarcfc Onvt Sm Coliefe. PA 1001 Approved By; ___C -- Faul Comolly ' T i c t o a l Laad*, LC-WS, &&ypn RaaMcdi ________ 'Oohn Flaboty / Vi Prendente OpmioM, Rxyf Rwaardt Dale Exygen Research Total ? : 7 Page 16 o f 65 Page 57 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number. P0000760 BxyfnXiaaepch Nwabar VOOOl 7*0 | ANALYTIC A l M im O D | Mcfood o fAmUyaU for tfce Dotenmaadao o f Ndluerooclaaolc Acid (PFOA) in Water by LOMS/MS 1.0 Scop* H ii matted i* to te oefleyad for 1b latec and quanbtatiaa o f jm fW ooctaocic add by Hijto ftrfo m u iK * Liquid Ctmoutography coupled to a tandem M m Spectromctric Detector (LC/MS/M5) io wamr. 2.0 Saitey 2.1 Alw ayiobavviM /blaboraiotfyprM tioM . 12 Cooeuit foe appropriate MSDS before handling any chemical for proper safely 3.0 SampleR a iu ira a o i 3.1 Alto 40 eeLeftaai aanplf for traction 3.3 No lairqila prnrow im j la bad to tor aamplaa. JJ Samples band refrigerated hoold bo allowed to equilibrate to room tcm jxratvt, 3.4 A ll m plea moat be thoroughly mixad te lb n being sampled for extraction 3.3 Any um pire contatoteg pm fc* should be ceoorltogcd at -3000 rpm fbr -3 ninutH and tba nq>cvn*Uftt Mad for tba traction. 3.0 Sample coDacUon procedures w ill be pacified io the itm pling plan for din petyart 4.0 Raagcnta aid Shndasda A l W ater-H?LC grade 4J MathanoJ - HPLC grada 4J Axnnoemm Acebde-A.CJS. Reagent Gral <4 Pcrfloorooctanoic Acid - Slgm*-A2drich 5.0 Imtnimeot "d Equipment 3.1 A high perfoamtnee liquid chromatograph capable o f pumping up to l aotvnxa equipped with a vih a b lt votuoa elector capable o f injecting 3-200 hL eeoMCtad to a tandam M an Spectrometer(LOMSA4S). 3.2 A device to colie d raw date for peak Laiegrstwa and quarutmiion. 3.3 Analytical balancecapable o f reading l O-OOOflI g. 3.4 30 raL diipoeaH apoljpropyteaa cenmfcge m b. 5.3 13mL dispoabla paljpropylane caotrifega tubea. IS Di^eaabla tecrcfcpau (50- lOOuL 100-200uU 5.7 123-cnL LDP morow-taeutb bcmlaa. 3.1 2 mL laar UPLC vial Ut. 3.9 DR o uble pipcttac 3-10 A iitopifcec (IQO-IOQOpL and 10-100 pLX with diapaaabla tip, i l l Waten SepPak Vac 6 cc (lg ) CI8 SPEcartndgaa j d Pag* 17 o f 03 Exygen Research Page 58 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 / " * Exygen Protocol Number: P0000760 fxya^lM M K* ANALYTICAL METHOD | Method o fA ja Jp k ft* ttoD m rm lD tfioe oiN riluofoocunolo AddCPPOA) m Watorby U7M 3/M M 2 SPB YICUUOOBMBlibhL 315 O a ttifc f coprirto o fip iir a f 30 iLgolypcopyton tub-- * 3000tpm 4.0 4.J Analytic*! Column: Pluopfca* 8? (Keyrtoae SciartiBc), 2.1 mm * 50 I, 5p (WN: 12505-0521301 4.2 Tampa m e: 30*C 4.3 ttobD oPhuo(A ): 2 mM Anreociaci AcetaW 1 Wab* 4.4 M *W aPha*a(B): M c& moI 6.5 Gnutiml N npvn: S ttilU B ) 0.0 1J) 12) 200 233 &A 65 65 25 25 45 Flow Rat* XJL Im L/tpin) 35 0.3 as 6.3 75 03 75 63 33 0.3 6.6 InjectJon Volume: l5 p L (c b e in c i8 e d t o jw Ji* * 5 0 jiL ). 6.7 Quartitabeo: PeakA m - external itiodm k calibration curve. L I RwTSme: - 23 mwuet Thoobovocoodittanoroiatwidodiaiguld fldjnayboclm ijediaortC T to optimise the HPLC tytttm . 7 0 MS/MS 7.1 Mode: Bkctn?fay Negrv* M TO moda, mmitoring 4J3 - O O P mil. Tbc tfeov coodibccu M imended ta k guide m d may be changed in oeder I opnme du MSMS eyttem. 1,0 PnpcraiiiB of Sdutioiu 1.1 M obile Mum .M 2 mM KWVOonnn acetate is water j prepared by adding 0.154 g of Id lOOOaLofwKler. Automata vchjanaamay be prepare!. Exygen Research J>02* 18 63 Page 59 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 [ ANALYTICAL METHOD_______________________ Method o f Anaiymii ftr the D^wm iaatiw o fPerfUiorooduoic Add (PFOA) in Water by LC/MS/MS 9.0 SoufamlPneMntoti 9.1 Standard Stoek/Furtificsbon Solution 9-1-1 Ptcp i nec* eahboa o f-100 u*rtL oiPPO by w a ijh irj 10 m * o f sndyticaJataodenJ (cecmted fiu p * ity ) and Iute to 100 mL with nwfasnol 1 125-uL LDPB botti*. 9.1.2 A 10 igAiiL fortification aodotionof PPOA i l prcpwed by tainiing iu m i o f Ih* 100 jif'm L solution lo fim i volume o f 100 w iih nxSwnoi ifl a 125 nsL LDPB botti. 9.1.3 A 1.0 pifferi. tonificato solution o fPPOA i l prcparod by brlnflng n i m Lofttw IO HV'rol' dution lo final volume o f 100 with mcihwral ni 1 125 mL LDPE botti. 9.1.4 A 0.1 fottifjcaon aohrtion o f PPOA i* prepered by b o n jjijj In m LaflSa | .0|i^S )Lelutiotttoe ilo) volume o f 100 wiifameihmol in 125 mL LDPB bottlei 9.1.5 A 0.01 jtfito L fo ttifieaiioa boIouqji o f PFOA 1 prtftared by hH rifio* LO mL o f tbc 0.1 \ilmL olutioo to i (bui volume o f 100 wiih u in i in a 125 mL LDPB botilo. 9.1.6 Thestock md fortifloation acheim i are b>b* rtc ttd in refiigeracor it ppfcxlm atr ly 4*C ad u t sublo fot maximum parrai o f 6 wnihs &tmt ih * d a b e fpreparala. 9.2 Standard Calibratimi Solutions 9.2.1 LOMS/MS ariihralkirt lu o d iftl* ire preparai in HLP1X wattr Th* eatbralkn standard* are pcoceaaed tbzough Ih* tn c tio n procedure, identical to i rapite. 9 2 2 Th follow in* ii a typicei rnplcc idditkm el cooce&trailoiu may be prepariti a i needad. Pinal Caaoemraoe fortiScatioo Voiture of CanoaMitknof Calibrano* oTFortificatici* Volerne Portifind Coevo1 GaUbsabee Staoderd E> Solution (wb) ___ -1) 00 10 1D0 3aiddtar (example) *0 0 XCramddyy-0 40 35 XCamddyy>1 IO 200 40 IO 400 40 JO XCamddyy-2 100 XCtnraddyy) 100 100 40 230 XOantidyy-4 100 200 40 300 XQaiaddyyl _____ 122____ eoo _____ !S_____ 1000 XCtwaddyy-6 * The ttOraolBd Muwntralioo o fth calibralirm ftanderd l* aqua) to Sx ita m itili cooeeniralkic, cimato checoncentrationoflh*standard durin| ih traction PE). XC extracted caJSxmMi irjndird. Page 19 o f 65 Exygen Research Page 60 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 BcygenProtocol Numkcj; P0000760 fc fjli I w w t MAodN*MerVW>ClW> / ANALYTICAL h tfT tfQ l> ~~ [ MMbod o fAnalytu IbrtN D m nnnw ticia ofPrflocro>ctnic Acid (PPA) in Watar by LDMS/MS 9J-J 9,2.4 925 K *m indmd aotatfo* ( m in i btafc) n.ar tTM with ich Mt o faodarda cwm tati. Sten all trtOWd catfbraticn ataotoda in 15-mL poiypropyie* tute M 2*C to 6*C, up to two weak*. A h vctuxaaa n d oeftetntnuiecii of itiadarda my b prepared u J ii HO Rate* 1 1 * IP.1 Each N id i o f Mnpka tttn c td (lyytfciy 20 or h fc ) mu include lent M TMgMA coftlaol (tm bod blank uains KPLC water) mb two mgem Hnnml tortiHad at known corwantnuiani (lab confrei pile) to verify procedali r*w w y to fee batch. HU Reqitamc* far field and laboratory duplicata* and pike* w ill be apccilicd in IN quality aafurtoia plan far feia pm pet I. 1,1 4naia 40 L o f p ia era portion o f smtpLafelutod to 40 raL wife wafer inn 50 mL potyjvopykaa c*nirife|e tub (tbrtJiy m needed, replace Kd and aweU2 I I . 2 Comliwo fan Cm SfE cn ld p M 0 l> 9 mL> by pnuJng 10 mL methanol fallowed by 5 mL of HPIC water 2 dropiNc). Do not let column run dry 11.3 Load mmpk cp renditiorud Ctt STB ofUidpc. Discard chiata. 11.4 Bute with -5 m l 100% mafeatol. Collact 5 mL o f ehiato into mdunicd J5 mL polypropy l fl oentzilUfe tnfcee (final vtxhnna StaLX 11J M a ly uBQf tlecroepmy LC/MS/M$. 12j0 CNcxmtopupfcy 12.1 fejaet fee a amount o f each andari. sample and lo tti(led aampia imo ihe LC/M g/M S ty ib w i A calibrati rOndard rm ut precede and foftow al/ analysed urnptae- 12.2 StandanfaofPPO Aazm ioceidicg le n t IWo or mocMcoiieiniJetien levels muatN included k an umifyUcnlaat 12.3 An tir u t o f nctraciad calibration andarti muat N included at fee bapaofaf and at fee end o f a am pia mL Extracted tandarta m u* be Ip la t^ m ad batwMe every $-10 lamplas. Ac an alternaiiva, n entire et or anatri cahbndco atifldnrda may be acted at fee N fanainf of a act followed by merected calibration ata&darda iotortperad every 5-10 empiei (to icttram E* a lactrnd tat o f aettnclri itandarda). In ito ti cam, axUacted calibration andarti m at tw fee fin t and li lt injection in a ampie *cl 12.4 Uab linear tandawl ca va far quamiutlcn. lin e andart curve art paneraiad far fee anafyta by linear rapreMkai uaipi 1/X weighting o f peak im i P o g * 2 0 o f 63 Exygen Research Page 61 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Bxygen P rotocol N um ber. P0000760 h n w U m rtl MH*ed N un** vqOO1710 ]______________________ ANALYTICAL METHOD_______________________ o f Analyaii fcx tboDetermiaatioa o fParfluorooctaoaic Acid (7 F 0 A )ii Water hy LC/MS/M5 v e rn i onlibraaoa m im t easommoo using MisxLyax 3 3 <w equivalent) lOftWIl WfXMML 12.3 SarapU rwpoaee aborti nat exceed stradari rwpcBiee. Any samples ehm xoeed standard ra p o o icj shouldhe furtherdtluttd and manalysed. 130 Acceptance C rlttrt* 13.1 C hM aito^aiw iit A o w ftpe*k*f*d m ^itrQ < iB t3 6P m v6a(n* pernii o f 413 mdil The 413 u u prmt oomapoada lo Uni PPOA tiojv whilc ihe datigbtcrkm (349 uau) rspracnls Drc loia o fe v ira i diax de. 13.2 VMhod blaniu n w t noi conta PFOA et levale gnau? ih i the LOQ. Jf bUck coetalni PFOA al teveii p m tr thaa 50 agO, tlwn new biaak (ampie must bc and th atavi M mustb tn c N il. 13.3 Recoueriee o f contool spaine m daiatr sp ira i m ini bc bctwaao 7M 30S or ttmh tornare vatuas. l i control apike A lti wm# tbe captatole lim ili, th o lir* M i o f am pie* ould ho lerartraotod. Aoy m etri* epdce outsde 70- 130% aborti be evtteefed by th UtoJyet te detenni i f re^xtraetm n wansmed. 13.4 Any eeiibrdinn standard Ibend to be * stat istics! outlier by w ing the Huge Encv T a t. may be excluded font (be calculation o f lb * calibration carve. However, the total number o f extracted catibnticet eUndenle Uui could be excluded nmet m i exceed 20% o f (be total numb- o f extracted landardi injected lie MrrelalUM coefficient (X) far califeeiticn eurvee jcaarxtod must bc 20.993 (R1 24L99S), If cslibratioo Torulti M outside Jibm bmite, then appropriate rtepe owai be taken to fttu i irtatranent operation, and the tandode or the relevant act o fin a p la ebomld be reanalyzed. 13.6 Retmtkua (me between standards and sample# must not d rift more than 4 % within an m alyticti run. I f retantua time d rift exceed* thia lim it wiihm m analytical roc then the act muetbe reenalyved. 14 0 Calcolisi004 14.1 Um the Allow ing equation to esfculste Um mumm o f PFOA found {in ng/L. bated ob peak aree) using the standard curve (tinenr regreauon pervref> f tnoetedbythaMasa Lynx software propina: PFOA found W L3 - ffn k araa - inlCTDgnri x PP slope DF Actor by which the final volume wee diluted, if neceeaary. Exygen Research Pofe 21of 65 Page 62 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygm Protocol Number P0000760 1 M a lia ) <rf A u l y w fc r * mwwmiNbwii v a h id A fiA L v n o i. w rru o o <tfPk Hiu t m o iu m U A d d (? P O A) in W t* r by L C /M S /M S 142 fix M iyki fbrtUBMd with known amourt i o f PFOA prior to bvetion. me Ite foUowinf cqaatao to ca lc u li* ite percent recovery- Jtewwwy 1%) }iPttlM l)tejb>uidCiifiI> -analyU f a m i in control {nyT.)] ^ ^ i d i ) ) added (e$ iT.) Exygen Research Page 22 o f 65 Page 63 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygea Protocol Number P0000760 ANALYTICAL METHOD Method Number VOGO1711 Method o fA aatyjb 1er the Datormlaados o f forfluorooettsM c Acid (PFOA> la Sou by LC/MS/MS Analytical Testing Facility: Exyfon Raaearct 3G5SResearch Drive Stal College. PA 16*01 Approved By: "ft-X C -jL ____ Paul Connoly * Technical Leader, LC-M5, Bxygao Research .- js Iia H Date rohn Flaherty ' Vice President, Opcntiotul, ExygeaReeooroh Exygen Research Tool Pages 7 Pag* 23 o f 65 Page 64 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Bxygen Protocol Number*. P0000760 fixypttkmeftftfe Mi*odMtmr VOCOl7l | ANALYTICAL METHOD Method of Analysis fer tfw Desenaiaalio e f PerfluoioociaMic Acid (PFOA) in Soil by LQWS/M8 1.0 Scop* Tha msiftpH is to b asployad fi lha isolation and quantitation o f perileorrxxrU/iwc add by High Pw&smsaee Liquid Q nwnaipfnphy counted to s ndetn M us Spectromatric Deactoc (LC/M3/MS) m soiL 2.0 Safey 11 Always obaorve ufi laboratory pradicea. 21 Cbnault tbe appropriale M5D3 cibi hautting any cheocal fot proper safety prscM itiant. 10 Sample Raquirecuat 3.1 A lle t* S | ftM lte m p ia fi artractiofi. 3.2 Ho sample promoting ia maded fix- od samples. 3J Samples mored letgsnoed toonid be allowed lo squilibro to room ta p m ta t . 3.4 AD samples m tt bethoroughly mixed before feeing sampled for extraction. 3.5 Sample colletto procedine w ill be specified in dia saanpliat plan fcr Ih projact 4.0 Reagents and Sbnteda 4.1 W r- KPLCfmU 4J Methanol - HFLC grade 4.3 Ammosriua Astiate - A.CJ . Reagnat Grade A4 ParftooDootaaoie Acid - Sigma-Akkieh 5.0 Indnimeot sud Equipment 3.1 A high petAnnaacs liquid ebromtiofraph capable o f pumping up to 2 arrivami equipped with a variable volarne injector capable of injecting $-200 MLconcerted to a tandemMass Spectrometer (LOMS/MS). 5.2 A dorine to eoUoei raw dam ft pack integration and quanimtion 3.3 Analyticalh1* * *" capable o frta d in | to 0.00001 g. 3.4 $0 uiL dilpeaabla polypropylene cnrOriJVgt tubes. 5.5 15 mL dlapeeable polypropylene eentriAigc lube*. 5.6 Disposable ndaoprpets (50-lOOuL, 100-200uL). 5.7 H S -aL LDPB asirow-mcwih torti. 5.1 2 mL dam n m : visu a l 5.9 Diqpoeablc pipettes. 5.10 Aulopipebae(KKMOOOpL and 10-100 pLX with dtspoaable tip i. I l l WatsreStpPak Vec6cc41g)tC1ftSPB cartridge*. $.12 SPBvacuma manifold. 3.13 U hnw dc bath. Page 24 o f 65 Exygen Research Page 65 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen P rotocol Num ber: P0000760 ty|w t o l r t V00017II f ~ ANALYTICAL M1THOD J Method o fAoalyti fee tie Determination o f ParfluoroocUnoic Acid (PFOA) in Soil by LC/MS/MS 3.14 W rin-icdo ibakar. 5.15 CcotriAtt* capable o f ginning 50 mL polypropylene tub ai 5000 rpra- 4.0 O tTKnjognpbk yitem 4.1 AndybnCohiBMi: FloofhaMRP(Ke>v4oe$oiMbfiBX2.] numx 50 mm. 5^ (P/N: 12509-032130} 12 Tbmpmpu-r: 30*C 13 Mobil Ftuuw (A ): 2 mM AnanoaMan Acetate in Water 14 MotHaN 6.5 QradUot prognm: T in ffrw a ) 00 1.0 IX) 210 22.5 2 65 65 25 25 65 PiOV Rate 2 k fl fm lA pinl 35 03 35 0.3 73 0.) 75 0J 35 0 J 14 k ftc tu Volim e: 13 j&L (eia be fascreMOdto m much u 30 >iL). 6.7 Q u aatiM ta; Poak A m -c x ts n u ] steadardcahbratioa eurre. 6.8 Run Tune ~ 13 m im t*. Tb above oanditiMWtro intended m a jxide end may be changed in order to optimize (be HFLC tyvtom. 7.0 MS/MS Syilesn 7.1 Mode: Blectroeprsy Negative MBM mode, monitoring 413 -* 369 m fr for PFOA. Tba bow condition era bounded u a guide and may be changed in order to cpUraix* the MSMS ayitem. 8.0 Prapcilecior$ebCWM ( .) Mobile P lu * 1.1.1 2 mM anunenJua acetate in water Je prqpimd by adding 0.154 g o f ammmexn acetate to 1000 mL o f wata. A ta n tt# Toluroa ourybe pttpwed. Page 25 o f 65 Exygen Research Page66ofn0 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 IxyftaftjNM c M fM N w w VOMICI T ANALYTICAL MXTH OD j Muhod o f Anifyai fbr thaDetanainato w o f Ew flaarooftaittic Acid (PPOA) in Soil by IC/M3/MS 9.0 Standard Prpanoj* 9.1 9a odm i ftett/Fortiftah Soliden 9.1.1 h e p n t stock ohMia o f -100 p jfa iL a f PPOA by weighing 10 mg o f tndytoal mndard (eeneefed far purity) and dilute te 100 mL with * W J ina 125-mL L2R botti. 9.1-2 A 1Q faiticaiiQ Hution o fFPOAia prepared by bflcgln* 10 mL oftbo tOO pgtaL solution to a fin d v o ta o f 100 w ith nctbarol m a 125 aoL LDFS boati. f.J J A LO pgtaL fo rtific a ti lo lulloa o fPPOA is prepared by bringing 10 aaLoffac 10 A fta ! aotaon to a final vo ta i /100 with m W ei in A 123 AbL LD fB AotQt. 9.M A Oil pptaL fertifleedos soluto o fPFA ia prepared by t r f ic i 10 m Loffae 1.0 pg/mL aetaikxi to a fatal v o lin e of100 with m stW ci in a 125 mL LD fB book. 9.15 A <M)I jjp'm L fortificato olution o f FFOA ia prepared by bringing 10 mL * f th* tt.1 uptoaL solution to a fin d volume o f too wiih mitiw il la i 125 mL LPFB botto. 9.16 Tha dock m i fbrtiftcadoa otaon ara to be stored In a refrigerator it ppwxaatoly <TC aad 0 ittb k ft a u u d iaun period o f 6 memtka fio o the dal* o fpreparation. 9.2 Standard Calibrati Solution 92.1 LGMS/MJ calibradcn standards sa prepared In HF1C water. The cabbrtao standards aro prQCMad through fa extraction procedure. IdaetiMJ to eamplse. 9.22 Th* Sowing is a typical example; additional concentrilioca may be pnpared aeaoaded. Final C e t i * iti u t PW t&MUH V e lu m e f Cboowinatiaa o f Calibration ufFortificabcn V e t o fw a lle d C--b e l Calibration Standard ID Sobrio* ippfa iu L l S a n s J tie iL l Standard (M l* (c u ird ri 0 0 40 0 XCmmddyyO 10 1 40 I 200 40 25 X C m io id l XCinrniUJyy-2 I t 400 40 i n too 40 too XCbuoddyyO 250 XCbuaddyy-4 too W 40 >00 X C tw oddyy-l ______ !S2_____ e 40 1000 XCm tnddry-i The extracted oaaeaniratloa o fthe calibrato standard it equal to 5* Ila initial eoacanirato^ due fae eooocntratfc offa# atandard (taring * e extraction (SPE) XC-extracted oritecioii standard. P a& 26 o/6$ Exygen Research Page 67 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygrn Protocol Number. P0000760 H yy ww> voOHfl ANALYTICAL METHOD | M aUiodofAsalyM dm D M nm n*iiw o f Podtuoroectaoolc Acid (PFOA) in Soil by LC/MSLM3 9.2.J 9 .;.* JJ A m o tfaadwd BokiSao (raetaas bl&a)0 nn b ro-gpirgd with eh Mt ofMndardi xtnctwi. Stai* a ll atncvad calbratios eieadarda ifl IJ-atL polypropylene tube* at 2*C U0*C, i f to two wwk*. Alw eatevohim 4 ceneealrsdcmeafsUndod* maybe prepared *1 Beaded. tao laidi SMUp K>.1 Sack batch o empiee extracted (typically 20 r icaa) nwat includa al Icaat ooe in ij<1 contro] (ttaftod blank ualnj 5 mL o f malfaoaol) and two reaycoi cottoli fcetifiod 1 known coooamndoni (lab control ipilce) io vcniy proeedural recovery fo rti beach. 103 IcqwreacnU ter add md bibontovy (hpbcacoe and spdtcs w ill be specified in Lbapatty atauraoce pian tee thia pspQjoe* 114 Saaape Sbroca* 11.) W eijh 5 f o f sample into SO mL polypropylene cantziAipa mixta (terrify ax a--dad. raptaca Ud and n ix wall) 113 Add5mL ofmethanol n d ifaalt oo awriit action ahak*r f o r - | 5minutes. 11.3 Tnawfer Iha tutoa to aa ultraaooic bath and sonicate fo r-15 minute*. 11.4 B hnf the volume up In 40 mL with walar m Iha 50 aiL polypropylene canCrifrfB tuba. 11-5 C W nftifo fa r"ID mijuno* at '-3000 rpn. 11.6 CoodJttoa the C * SF5 cartrldfat () f. 6 mL) by palling ID mL methanol Allowed by 5 mL o f HPLC water {- 2 drop/aac). Do not lei column run dry 11.7 Load (docim) the ample on the conditioned Cu S? cartridge. Discard eUialm. 1L I Slut* with ^5 m L 100S mdhwmt Colled 5 mL of eluale into graduated 1$ mLpoIypropyteaa coalriteso tubes (Dualvoluma - 5 m U. 11.9 AnalyaaaampkoMltttelectroapay LC/M&MS. 12.1 faucet 0k som amount o f each randasd, sample and fortified lam pi* imo the LC/MS/M? ayrtam. A calibration atandani moat precede and follow all analyacd ip lw 123 Standard o fPPOtA cocraapoodiof lo al lead Uva or more coooantration levali auwt h* iaclodad im an analytical le t 12J An emiro ace o f extracted ce&tndoa m ndaria must be Included at (he bagnam i aad at tfaa rod of a u a y ie M t Extracted tuodarda rovai ba InttHpanadbetween ovary 5-10 aacaplaa. A t m aUanativ. an mbro aat of Psg* 27 o f65 Exygen Research Page 68 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygra Protoool Number P0000760 Mario Nuoto V00017II ANALYTICAL METHOD Mediad o f Anafrr fio tto D n-- MUtfcaio fPteflaoreoctnoow Acid (PPOA) in Soil by LOMS/MS extracted caM ratka andad may be U^ ootod at fee beginning of i %m JbQewwl by extracted M tibntiea andada intertpcned vary 5-10 an diti (te accrual f tr i Mooad to o f OTected Brindad). I* wthar c m , attracted atibntio sentente n u t bn tbs fin i and lu i injection in a sample act 12.4 Um Sato n ta d cerva fb# quantitation. Linear xwterd curva u t ta unted fix 0a m ly to by linearfcgrrioa M in t U * weighting o fpeak we* ram a coUbntrao toodwd ooncartratk using Maeelynx 3 3 (or equivalent) software system. 12-3 ta p i impania should boi msmdl standard iq iw . Any sample* that caed Brindar Tcspoaec*should be Anther diluted and reanalyzed. 13j0 Accapianca Criteria 13.1 C toonoofran m ia daw a peek o fa daughter ion at 369 emu from a ptroni o f 4)3 aim). The 413 tmu parent corresponds to the PPOA anion, while the daughterion (369 amu) m am m athe kma o fcarbon dioxide. 13.2 Method blanks must not contain PFOA at levala greater dun the LOQ. If a blank Denuina PPOA at lavala fre tta r than 30 t\g/U man a oew blank sample ro u t beobtained and ibe entire a * must here-extracted. 13J Recoveries o f control spihw and matrix apikaa must be between 70-130% of fe rir known value. If a conimi rike fella outride fee acceptable lim ila, the M tire to o f ungile iheald be r*-xincted. Any matrix spike outside 70IW % should bo evaluated by fee wialyal to dotevtmae i f re-attraction is warranted. 13.4 Any calibration standard femd to be a staiLiticai oiilKar by using fee Huge Enr Te may be excluded from the calculation o f fee calibration carve However fee (otai sambar o f extracted eahbraiicw standards that could he excluded mtut not exceed 20% o f the (odi number o f extracted standards injected. 13.5 The oocnlabon ooefliciant 0 t) fer catibrthon curve generated must be MX992 (R2 20L9B5X I f calibration result fe ll outftide (hene lim its, then apprty iaSe step touri be taken to adjust instrument operation, and ihc andante at the relevant to o fiim plns feouid be reanalyzed. IW Lm otion tine toxwean atandardi sod saanplee owat not d rift more than 4 % within an analyticalran. I f retention tins d rift caroaedathis Umh within n riyti Jn m then toe item iuu hareanalyzed. ru tto ri P ag* 26 o f65 Exygen Research Page 69 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 IiUmlMUfdi V0OOPII | ANALYTICAL M IT S U I) Method o fA u ty i fbr fa t Deunmnuien o f PerflviMoocttneic Acid (FFOA) in Soil by LC VS/M 8 14.0 Calculating 14.1 Um Um following equation to ctfrulet t the amount o f FFOA fined (in ngiT, booed peak im ) wing the atndwd eurve (linear regraeaxw parameter*) ganvated by the M u * Lyat eoSwire propani; FFOA fount f /L i - ff-- kr-- . imfoepil x DF feh>)M DF * factor by which Iho final volume w u diluted, if noocMary. 14.2 For umpies fb rtillid with known tmouma o f PFOA prior to caVKiion. u x the following oqniiinn tocalculate the peroau recovery lUcovcty (S ) - [ totalanalytefire d ifit-) - enelytp found io control(a yL )] nutate added(a*/L) 14.) Uee the following equation io convert iha amour* o f FFOA (bund in ng/L to o^fiFP b). FFOA end (ppb) * fffQAfamd ( m l ) 4 JtafcMIgglEinl IQ.ML1) ample weight (5 g) U.4 Uk (bo A B o w in g qtutioa to calculate the acMiiat o f FFOA found in ppb beeed on <ky weight. FFOA found (ppk) A y weight* FFOA found (ppb) x [100% / to lti *lida{%)] Exygen Research Pag* 2? o f 63 Page 70 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P0000760 analytica l m ethod Method Nuubar V00017I2 H N M i f Aaefyiia fa r tk* U m m M lim er F a rflu ro e e tifto fe AcM (TFOA) la Sedim ut by L O M B /M Aaalytjcaj TMting Facility: Bxygua Reaaarch 30SI Raeawdi Drive SUMCollag*, PA 16101 Approval By. C JL Paul CoaooUy Tectowel Leader, LC-MS, Bxygec Rawareb Um r f U / / Jobe Flaherty Vioi a P~rrmd`mt OpcraOooa, Bxygen m arch D ai* Data Exygen Research TceaJ Pag; T Page 3 0 o f 65 Page 71 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 I i ea I n m t l Mafted Master VOOOl'l i 1 AN ALYTIC AL METHOD 1 Method of Ajielyala kr lha Pf m inatiau of Pm flporeocfoio Acid (PPQA) in Sediment by LC/MS/M3 T V i method i l Mb*employed to the laofcdoo to quintunoo o f perHuoooctmoic add by H i# fwlbnnaa; Liquid Chronuacgraphy coupled to undent M ur Spectrocutric Detecte* (LC/MS/M5) in aedimeat 2.0 3mh*y 2.1 Ahvtyv obwrve mib laboratory practica. 22 Coamll ite pptupdiU M3DS before hmdlixqi soy cheoucsl foe proper aefeiy 3jO Sample Xoqtmancel 3.1 A t k e it 30 f o fUH Miopi# for eatnetioa. 3.2 No eample petceeetag i i needed to esdioieat wraplee. 3.3 ampie itoiod prfrigeriHd ihould bo allowed to equilibrai lo room tempeauJure. 3.4 A ll aampleenuut be ifaofouihly mixed bdbrcbcm f lampJed for jctrution 3.3 Simple collection procedura# w ill be rpeciSed in the lamp ling plan for tb it ptaytet 4.0 Raajentt and Standard! 4.) Water- HPLC grade 4.2 Methanol H7LC grad 43 Anade Acid-Raagsmt grade 44 Ammonium A ca ule- A.C.8. Peepw l Greda 4$ Perfltmrooeuaoic Add Sigm^Akthch 5.0 inatnoncm end Bquipmera 5.1 A high parfaemmes liquid cfcrtanitnfreph capable of pumping ip lo 2 m Nuju equipped w tt t nriabla votene injector capable o f injecting 5-200 pL eem eued to etamtam Maw pctemete (LC/MS/MS). 32 A device to collect rv dida 5 peak integrati) and quantitation. 3J Analyticalbalance cqpsblo o freading to 0.00001 g. SA 50 m Ldpoeabltpolypsropyiteeentriftije tutea. JJ 15 mL dU^tteitbU polypropylene centriftjgs tube. 5A Dpeeeble mioaoppcU (30-1OOuL, l0C-2O0mL). 5.7 115-m l LDPH usov-m outb bottles. 5.1 2 ad. c M HPLC vial k it 5.0 Dlapeuble pipettes. 5.10 Amopipettai (100*1000 iiL m d 10-100 fiLXwidadbpoaalilfl tips. 5.11 W stes Ssp Pak V tc 6 c c(lq ) tC l ISPE cartridges5.12 3 P ncuuan numfbld. Pagt J o f 65 Exygen Research Page 72 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen P rotocol N um ber P0000760 layfesfasraeb 1 ANALYTICAL METHOD | MedaodofA w ly tli fc f tb t Dsterminsdcn o fPsrfluorooctaooic A dd (PPOA) in Sediment by 3.13 V ortosr. 5.14 W rirf-sctios shaker. 5.15 C m lriftjfa capable o f nwming SOmL polypropylene tubra t 3000 rpm. 60 C hw stefrapbicSy4 6.1 Analytical Column; Pluopbus RP (Xsyiteos Scientific), I- t nun 50 nun, Sy (M i: 12305452130) &2 T<ffHM r: 30*C 6.3 Mobile Phase(A ): 3 mM Ammonium A estiis io W se 6.4 Mobile Phase CB): M rth ro l S J C kadieot Program ; X jn fi n i 0.0 I jO 10 20.0 22.5 JU 65 43 25 25 65 Flow Rale 5LB tm l/p in l 35 0.3 33 0.3 73 1.3 73 0.3 35 0.3 64 Iqjscaiea V o lta s: 13 jtL (canbs ncreusd l ts much * 30 pL). 6.7 QuaotitztioTL Peak Aras-stecnal ataadiKj ciXbradoa eurvs. 6.6 BLtmThnr -2 3 ramu*. T b t *c v oomtttkns ra foUMfcd u a guide sod may be changed in ertfcr X) optim a* thJ H?LC i>*an. 7.0 MS/MS System 7.1 Mode Sketroapny N sfstiv* MUM medo, moataciBg 413 -360mtz ftr PFOA. TM above ooBddau a iatadad u a golds u d may be changed in ordsr to opdmfes On M3MS syttsra. 80 FrapantkmoCSoluHoai 1.1 Mobile Phis* t.1.1 2 snM anonhw u m m n snow Is prapetsd by adding 0.154 x o f ttnonium acetate to 1000 <L s f water. Exygen Research Page J2 o f65 Page 73 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P0000760 MmfeX Mutfcar VOM 171} ANALYTICAL MXTHOD " M tcVrf o fA natytii fix- * neuraunition o fFw ftwaoociinok Acid (PFOa >ln Sediment by LCYMS/MS 12 Euncdoa Sofctioai $J l IH aoeti u id ca Witar it prtjwred by idding 10 mL of acrtic acid k> lOOOmL ofw ftar. AkcnxU vqJucto* mnjr b* praparad. >0 Standa-dJcapantio 9 .i Suadard Sled/Peitiflestioe Sotuoe 9 .L I h9r a Maok miulaoD of^lOO ks^ qCPFOA by w ith u i| |0 itik *f xaalyaktl cfmdart (onwiad fbr ptuity) aa4 ditoe w IW m l wnK wucnol in a 125-mL LW B bottl*. 9.12 A lO jiffa iL fortiltaA oo aohfae a f PFQAia pnpartd by brin*m it 10 mL o f tb* 100 p ftn L vahnkni to l Saal volume o f IOOwuh nwihinM in a C25 mL LUPE bettle. 9-1.3 A 1 0 p fta L farbficat* Mlution i PPOA 1 prtparad by brinjin# ID b*L o f Uu 10 pj/m L tutioo to aflaal wolume o f 100 with methawi in a 125 mL LDPE bottk. 9.1.4 A 0 .\p ffa L (brtiftteoe aefatitn o f ffO A i l prcpertd Vy tainging 10 mL of Th 1.0 m jrtn i toluon to a ftxuJ volume o f 100 v ith nutiunoj in a 1Z mL LDPH bottla. 9.1.3 A 0.01 p *i L artkadoc solitioc o f PPOA i i prtpared by brm*ir\g 10 mL of tfao 0.1 M ftaL MJatiott to a EueJ rolumc o f IDO with natkMel m a 123 mL LOfBbodU. 9.1 iS T V ock cd ta tifie ih o aolution an io be M oral in ar tfrlftm it appeojdmataly 4*C ind in stable fix a m n in u n poried o f 6 month ftcaa tfaa dato o fpnparttioo. 92 RtmdaidCalibeatioaSeloticme 9 2.1 LOMS/MS cabmjm t f r i i i c ra ptaporod ia mothanol via dilution e fib e a i pffaLrd M tkaaohrtiin. 922 I V foilew fat ia a typJca) xarapl* additional ooocantoaiioiu tnay be of Fcrt/baden Schitw (ngrtaL) 100 WO 100 10 i l V okiina D O jttd ( *-) 1 ( U 100 S 100 2 IOO 10 KO 10 100 ___ IS---------_____ m . final CowaonUon [MfrU 10.0 so 20 10 OS 0.2 Page 33 o f 65 Exygen Research Page 74 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number. P0000760 I ANALYTIC AL METHOD Method o f A m lyri* for the PeteanJnalk* o f JWluorooctanoic Add (PFOA) in Sodimont by UVMS/N8 9.2.3 Bier 111ttlfo n fa n ttandudt la IZS-mL LUPE narrow-,mouth bot|Jt . r e f*C , up toaU m ontili fZ * Alternato waluma and ooocrti*oa o f atoftdarla may be prepared u 10.0 Btch Sat Up la i Each batch i f templet extracted {typically- 30 or Io n ) n w i include at Icut ad two untreated control* fortified at known (lab control <plba) to verify procedural tacovery foe the batch, for Held and laboratory duplicate and tfU m w ill bo specified in therjuahty aamn cca pkn for this project. 110 IL1 Weagb 5 1 a d B fle into 50 mL polypropylene n trifu ^ i tuba (fortify u M e d * replace lid n d adz well). 11.2 Add. 33 aL Of IS Made add, cap, vortex and ifaak* on a w rial aedon shaker 11.3 Cdiifoa the tabaaal-3000 ip n for-OOmimUu. 11A CoodHScn (be C m S W ubibid pea (1 g. 6 raLJ b y passing 10 mL medtanol feUowadbyiO m LofK P LC w itor(-'*2 drvpfwc). Do not let column run dry 11.5 Load (dacant) foa atonali on & * conditioned Cm SPE oartndja Discard alitata. U .I Add 20 m L o f medtowol to the aedimsei left is the bottom o f ihc 50 mL Clolrifoga tab*, p . varia* and baie on a w rial action shakir for -30 11.7 Cmtnfiage fonta b * at -3000 rpm for -20 stonata. l i . I Decant the methanol no 0 tuna SPEcartridge. Collect die eluat*. 11.0 Wash (be cotam with 4 mL of methanol. Collect foe e lu te and add ii to the fatalacollaoted in ap M .t. 11.10 Q m d iiln a MOOndCit STO cartridge (t g. 6 L ) by p ew oi 10 mL methanol followed by 20 mL o f HFLC water 2 d ro p '**). Do t k* column ran dry 11.11 Add (ha methanol to -200 ntL o f water and load on the second conditioned SPB cartridge. 11.12 Bhde wife ~5 mL IDOS meffcaool. Collect S raL o f thiat imo graduated 15 mL potypwy y tma p tn fiig o tubea (final volume 5 mL). 11.15 Analyse sample uain| ebetroeprty LC/MS/hdS. ?#5fT P age 34 o f 65 Exygen Research Page 75 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 VQC0I7I2 \ _______ ANALYTICAL MTTHOD [ Method c f A i Jyfii r 1 D f m lj tIo ft o fPer^uorcoctuxKc Acid (PPA) In Sediment y LCTMSfMS 12j0 Quonuu>frq>hy 12.) lq)M Ihe M int n* ouM o f aecfeecsadnrd, M afe end fbrtified urople imo the LOWS/MS tyrt J. A u lb n tio o tundard must precede and follow il l analyzed aamplae, 122 Staad*d ofPFOA coo*^cadin| toaUeerf fiveoravore oonoetratien le*xl* m ut b* included in an am ytkj m i. 12.3 Aa M ir* K t o f extracted calibration gUnderdi raiiM be included at tftr bagUmlai and & and <rf a m pla m . Standard wet be intcrapcrad between wf^y $-10 umpUa. At an eanativ, aa M ire m o f calibration Uandarda may b irgectod at the bctum irif o f a at followed by calibration (teutardf nsenpereed w ry 5-10 Mmplee (to acoouni for a second m i o f m cdarfiX Ifl eatfxr caM. ealibnttaa naadard m ini be toe f t * and LX rzOCtiG m sample set 12.4 U * linear stand4 curve ft* qaartiw jon. Linear ttinderd curve* are fm antad <br be apatyta by 11m m ram w icn ualft* lf r w n gininj o fpeak area veraua calibrmtioc ataadud CMuentradon via% MeatLynx 1.3 (or equivalent) software tyetanv. ]2_i Simple Impouie efarnid not rawed o u tla id reepanaaa. Any tam pki tbw exceed nanrfard reeponsee should be Airtfa diluted end reanalyzed. 13.0 Apong CrttarU 13.1 Cbranatopram mut xxv a peak o f a d iu g b t* ion at 369 amu firtwo a purent o f 413 m u- Tbe 413 une parent correspond to lhe PFOA anioa, while the dsugbur ion (340 mu) topraeata tbe Ion ofcsrbcn diootide. )32 Matbod bbMo MM MM ooMMn PPOA m Jgveig jm to r tban the LOQ Jf t UmA wt*fcu PFOA ai k v e ii poster tban 0,2 itjptaL, thon a new blink ampla tmM be obtetaod and lhe e rtir* Mt mut be m-axtneted. 13.3 Reoovebea o f oaol ndlw* ad raaaix plksa rouai be betweee 70-1>0% of c know value. I f oootoal gplke fille ovtsk the acceptable limita, ihc jtu * M t a f m ptot ahould be ro-eatracted. Any matrix ip d u ovtods 70130H ahould be cvakrafcd by tbe anatysl dtermine if rc-catractkro it aminetn i 13.1 Any oaJnnbn u*ndart fcund ta ba a rtaedcaJ otia t by ualng the Hugt Errer Ttet, may be bcUkM wn tbe oakwladon af (ba calibrsilon curve. Nowavar the total number o f xtracted caUbntioa itandarda that oovld be ecludid amie am axcaad 20% o f tbe total number o r extraclad auodardi tqjwtsd. 13.5 Ib e oDoelatk coeffloen: (R) for oaHbnJxm ctnvei fenenued muet bc 20.992 (R3 2D.91S), I f calibnoon raeulta l ouUide thaee lim ita, then appropriele tapa nuul be tadoD adjum inetruioMil operation, and lhe rafeoda or the refovani m o f aamplae ibouid be reaiwJyxed. Page 35 o/6S Exygen Research Page 76 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 VttXlTO ANALYTICAL METHOD Mvlhodo f AnsJyt tat Qw Dotaraum kn o f farfhtoroocUnoic Aud (PFOA) in Sediment by LC/MS445 13A litm flo n tfanM tatwaan Standard ic d samples o n * not d rift man ihui 4 *w iifc fc iw analytical tub. I f im m rign d m d rift qaeead did lim it wtihin m analytical tub & <twset puul ba naaulycod. 14 0 C a k u h tiw 14.1 U * thclbllowm gtipum oo tckeJete t * mount o f PFOA found (io nf/rw l. btoed a peak arsa) m io | tfea standard curve (linear wgwask* panmeierti) taoaatcd by 4 M ax Lyra aoftwan program: PPOA fctmd (ngAnL) - ib ekiB A ^iB lesaB tiL X DP 4l0(M DP - freier by wfefck dw flu ] volume w u dilutnet if necessary. 142 Par m p ta i Jbrtifiud with known mount o f PPOA pdw to e*tractkx>. use the fcHowwijgnaden t* eaieulsts e peresm recovery. Rooov*y{%) " (to ta l-- lytofeaod (nyfriL) - adyte found in control (ng/tnL)] ^ JtuJyt* added(n fmL) U J Usettw ib ile w in| gustion to eoevset to mount o f PFOA found in r*6nL u> o^K(ppb). ppQAfeoad(ppt) * fpfQA&upd,(ciftni, final Yfllumc3..aL>J . svepie w tajjrt (i g) HA Um the foflowm j equation ( If aooeaury) to calculate tha amount o f PPOA found in ppb Ueed on dry w |ju. PFOA found (ppb) dry W M|bl- PPOA found (ppb) x [10OH / total aeiidiO l)] Exygen Research Page 36 o f 65 Page 77 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen P rotocol N um ber P0000760 ANALYTICAL METHOD Method Nim twr: V00017JJ forMttlMl *f Aa]yab IM o/Frtl*rwctww4 Acht (PFOA) ko71*1 d CImdj by LC/MX/MS Analytical Tattinf Pacity; Brygcn Bjaearoh 3051 RMe*oh Dtive SIAM CQlUp, PA 16101 Approved By: . C i L _______ Paul Crauolly Tedukal LmkUc. LC-W i. ftxT jm fcwwarcii M uM Cto g L *? / / JrnbFFWaacty 'r V>ie<a*Pw OpwBtkmi, Ex/g*n R*m reb jJ d r . D* Exygen Research Tatet ftaM: t P a te 37 o f 65 Page 78 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Bxygan Protocol Number P0000760 ____________________ ANALYTICAL METHOD_____________________ Method o fA aal/fla for (heDerearmaetica o fRacflwooctoaoic Acid (PFO A)tafieb and ClunabyLC /M S /M S 1.0 Sooft D iu wuflind Ifto W --ployed & rth # Violation nad quantitation o fparfloorooctinoic add b y HJfh fo ta a ia x Liquid C brom etojripliy coupled to tandem M u t 9pee*omafric Dtu:tor(LQ ,M3/M5) in fiah and e lm . 2.0 Safety 2.1 A l-- y ob-- vb ufe laboratory practical. 2.2 Cooroh the appropriate MSDS befere handling ln / chemical Torproper w foty 3.0 Sample Racuironu m 3.1 Ax 1-- 120 %of M temple Cor atfraction. 3J - Simple* dmdd be pro*--id before atiactiaoL PIbob the fio u a sample in a feed prod-- or m t ho aopenlt * wUh dr>> tec. PUca the templee in coounnat* and l-- r* open in feoeen a te n ft overnight Id allow for carbon dioxide wbHnudon. Seal nvl place the taotple* in froxea iterate until lime of iM lyaU. 3.3 Sample oolleoBan prooedunr w ill be petrified m the tampling plan for thia 4.0 Magcote and SlanrtenJa A ] W iter-W U T pade 4.2 Acetates-ile-HPLC grade A3 Carbon (120-400merh) -Reagent grade 4A Mdbaaol - HPLC grade AS Silica pal {&-20Q meah) - Unapnt gradi A6 R o d iil (O-IOOmMtO - RMjt*Qtsnda 4.7 Superoteaa LC-NHj - Reegwn pad A t 1-OcUncd - HPLC grade A9 L-Aaoocbic acid - Reajpcx rede A ID Dim cthyldkhkroiiUae - Reagent trade A ll Tolaene--Raagmt grade 4.12 Ammoolm Acetat - A.C3. Reagwtt Onde A 13 Perfl-- oecunofeAnid-S ipna-A M iidi 5.0 kfla -- Mat and B q u ip m o t 5 ] A high performance liquid chromatograph capable o f pumping up to 2 eolvaote equipped with e variable volume htfectoe capabJr o f injecting 5-200 pL ooamieted eai tandem hdeeaSpectrometer CLC/MS/MS). 5 2 A device tg collecttaw datafor peak intoyatioB ad qiacritetioo. 5.3 Anelpical balancetu fu b ite f reading to O.OOODl g. t t y 2 ura P a g e 3 8 o f 65 Exygen Research Page 79 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen P roiocol N um ber P0000760 * _____ I A W A LV TiC A LM rraO D Machod o fAaalytl* fa w P^armiaiittaT o f N rfiuow w iw Cknu by LC/M3/MS I M a d N ia a b VOOQ171] Aad <PPOA) in Pinh w j 3A JUm y wwfwmor. 3.5 9j6 I2SaLpM M h4N dflM kA SJ 30 tL diipo--b k polyyn^yfauy mvitugi mb, 54 15 g>L diryoiabU polyyropytaoe coctrifotc tubes S.9 DiopMMbk m ianfi (SO-LOQuL, 100-100uL). 5.10 12S-L LD.PBnurow-m oaft bottko. 5.11 in L o k w H T L C viU u 3.12 pjpM N . 5.15 Aw^fnpittM(IOO-tOOO^Laod 10-100>U.) w ilhd iipottbletipa. 5.14 SFB tk (JOroL) (Supdca c*L na h/0$7177). 5.13 W rijl etiasutuk*. $.16 C m trifajp aprilo o f opinata* 50 n L polypropylow tubo it 2000 rpm. 6.0 Clnuuirtupqthic 5y*m> 61 Analytic*! Column: Fluophiaa RP (JUytotw Seiendftc). 1 men i (F/N; 12505*052130) 62 Tempasraro: 3(TC 63 Mobile Pfaue (A) : 2 taM AjuiTMuuiun Ac*Ukis Water 64 M obil* Pham (B) : Methanol 6.5 Q m di Profrim : T ifltf irc ifl} 60 1.0 1.0 20.0 213 54A 65 65 25 25 6$ Flow Rate fmIVmin'1 33 03 3$ 0.3 75 63 75 0 3 33 a s 66 liyoetlon Votum * 13 pL (cm bo o te m -d to a much M 50 uL). 6? QkUtmtoban: Pak Am* - tornai itaodiH calibradoc cum 6 * Rm Tnte - 23 nula Tb bv cooditioat&f intended u p guida and may b* charged morder to ojnnza th* HPLC kMofl. h fr > fl Pu%t S9 o f6$ Exygen Research Page 80 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number P0000760 kypntoM K k \ A N A tm C A t MSTBOP Cisco* by LCVM3/MS Mcto<t Haw Vw)?AJ ] A c ld (tF C iA )k F .ii( n<l 7.0 MJfMS3)Nai 7.1 Mod B lN feM payN sptiv* MRM mod, m acftariiif 4 1 3 -0 6 9 mfz ft* PFOA. The ebov* am dtikw w iohndid i s guide id d my b< chened ie eider le a ftu a n a * MSM5 gy*am . 1.0 FnptnAanoffefeutoM t x M obili PW * w . l 2 sflM M M B IB ortafe I* v m v pmporod by sdding 0.15* %of acMaoahanM M to 1MOn L fwater. 1 2 Extraction Sohitios* 1.2. t 2% SSOiiWc Arid inm dhino) isp ttptred by diw olvim 2 gofascorbie cid i 100 n t o fmctiWol. 1.3.2 3OH DinwtylricbkuoriLicM io loluono 1 prtpnvd by b ritigm j > m l o fd in 0orUidi)oMlMlo B fln ilvo liU M O f lO m Lw ith lolueni- AIM m m volume* m y b tjn p m i. 9.0 Sttudted pnpeeKke 9.1 Susdinj StodcTorticacioJi SoJunon 9.1.1 PnfNm mstuck safridim of-1 00 jiy 'm L o f P70A by wadghinf |0 o f sasJylwa] Uodxd (correaod fix purity) And dilu io 100 mL wkh RMthmxJ m o 125-a L LDPE botti*. 9 1 3 A 1.0 p^m L forti(Variod tunor o f FFOA u pripuod by bnn^ing 1 m i o f tc 100p fta L Aokion to fim i volume o f 100 w ilh m Uu n I in * 125 mL LDPE botti. 9.1.3 A 0.1 p ffa L fo ttiflc tlic n AOlAxm o f PFOA iip n p tro d by b rin im i IO a L o fChe 1.0 pffrsL tulltw fc>a fin ii volume o f 100 witb rariJwwi in s 125 mL LUPE botila 9.1 A A 0.01 ligAnL RxtlflcAdon sotutk o f PPOA i l prepored by bringiivg IO m i f 0m 0.1 up^pL solution M i fim i volume o f 100 wilh m xhoiiil in A 125r o l LU PI botti* i l i The Meck *ad ibnificotion KtoUan er to b itorad in i rc friiw iio r pyteximetety 4*C Ani ara *Ub)a fbr * maximum parod o f 6 marni* CroioIhe d a tio f pnpiratioe. P n tio ri Pogt 40 o/65 Exygen Research Page 81 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Bxygen Protocol Number: P0000760 Mwfcod VOCOI?I3 ANALYTICAL M ETHOD Method fAnalyst* fix lb * Dateontaibon fPsrfluocwoctandc Acid (PfO A) a ftab and Clima fcyLC/M SAtif 9.2 Standard Calibration Sotedooa 9.2.1 LCM8AIS sodard ar pcreparad la mtfhanol via dilution O f lk 1.0 pffaiL f o rtific a ti solution. W 2 Tha fvUowiaft * lypwal auntplo; additional coocsatratioiu may be p w y a d aaaaadwl effareficaboa V o te Diluted l Piasi Canoasnation 1.0 It 10 05 OlW ai 0JX>3 h^> 5.0 2.S 1.0 10 10 to 10 fru -j 300 100 100 100 too 100 100 9W 0.025 O il 1X00? DD02J OOOI 0.0003 9Jt.) Steal U mbbratke standards is 12S-mL JJDPE narrow-mouth botttt* U 2*C te V*0, up to m BtettQn. 9.14 fiJtoraate volume* and oQOCCOtrtrion* o f itandardi may be prepared as needed. 10.0 batch Sta Up 104 Back batch o f aaraptai enacted (typically 20 o r leas) u u i include ai lean one uotraatad ooMvol and two untreated coctroli ft* tjfieri known eoocentnboita (lib contra] ipOw) to v irlfy procedure] rooovary fw the batch. 10J ior Said and id e d to ty ffcpbcatsa and apakaa w ill be specified in the quality u ra u c e plan for thia pacjeCt N.D Sample Extraction 11.1 W rjh $ | o f fcesne M ttp k into 30 L polypropylene cemrifugs tubes (fo rtify ai needed, replace lid and mix well). I I 2 Add 30 m l o f atwwauxik and dtakaoajiWTtM action shaker for 3 minuiM. HO B*the tabea to a b a te r far ~1 hour. 11A Pack and condition 0N SFE rebaa and jOanizB lb pear^haped flaafca 11-5 Pack the 20 a L 9Pfi tubaa in tatpiaiKie with 2 g fio riu l, 2 g silica p i. 2 j caibaa, end l g LC-KH,. Condition the column* w ith 20 s a l o f methanol, then 20 mL o f acetorutrik. Discard ail washes. Do not ibow the column to Ay. 11-6 SUsnii tha 125 mL peaMdupad flu k * by rtoainj with the JOS dmte&yidkhWvedlw* io te)ucsa aotetkm- Rime the Bask wUb telsane once, foUowad by aM tbMri <ihraa t o n ) . Dry tha Casks completely before use, o th v b y slf-drykfl or with a atman o f B toot. Page 4f o f 65 Exygeti Research Page 82 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Bxyjen Protocol Number P0000760 r t M etfodw w voim i3 ANALYTICAL METHOD Mofaod oCAnalysis fa tbe Detenroutic ofPtrflwtoocwneic Add (PPOA) In Pith md a in b y L O W W 3 11.7 Caainfoge foe 50 mL pctypropylro* tubes ooeteiniag sample m -2000 rpra I I M Deetrt (M atme* n to ondikienM SPG cohutm fined inaide the mouth o<` Uw H u t Ceiled the lud in the 123 mL uluuM d pw-hap* flask. 11.9 Add 10 mL e f eoetaaltrilc t tbe sample tbe 50 mL ca>tfifug* ubc. Horao|Hza the 0<ot> tl pfc*e uatng * lUsumw for -30 seconds md rinst the tissuraasr whb -10 mL o f acetonitrile int the tub*. 11.10 Stube tue u n ( b *#*fe*~10 nuwsUe 9 s wrisl-*ction shaker. l l . l t PtaeeUwtubeeieewfor-|beurm o 11.12 Ceatribge du 30 mL polypropylene Tube* conlaimiig u n p le et -2000 rpm ftr-tO m inoU i. 11.13 Decnt the extract onto du tun SPE oolumn. Collect tu lust* ist ihe n u t pesr-ehaped Disk and combine with the ttuam frtxn the initial extraction. 11.14 Psu 20 mL >fte uodoil* &rout du SPE column and combine the aluite m thcsanw poN tiqw dfluk. 11.15 Add 3-4 drops o f 1-oocud I tbe tu e t ln the pam-sbaped flute and npoeU f M redMPfdp N m u iu reduy evaporate (4 < 40"C). L1.16 Mai Ibe foul volume, by adding 2 mL o f 2% ascorbic seid in methanol to Uwpear-ebeped ftaek and awiri to mi*A*aotv. 11.17 ftasw farlM axirim toHTLC vieJaceinf dUpeeabJepipata. l l . l t AnalyzeaamptaewngcotrorprayLOM5/MS. 12.0 Chion&ato yaphy 12.1 Iqjaet Uwum amount o feach standard, sample and fortified aampieiato Ihe LOM5/MS system. A cailbralit Standard raust precede and follow all aoaJjMd mph 12.2 StandanUofPFOAeerrupoading tik a fiv to r mors eonctotm kn kveli buWb inclodod in awanalytical m l 12.3 An OfTtiawsd c f cabbrao* stsadads must be included tbe bofmnmj and ti tu end o f t aonpU aat. Standard mutt be wKeupemd batw ur every 5-10 samples. A S aherenove, u e oiirt set o f oaJibcatkm standards may be averted at tbe bepmnag o f a set followed by calibration standards Inariperaed away 3-10 sarapJss (to aacnant for second aw o f standard!, lr other am, caflbratu* standards meat be ttia fleet and Ian injection in a o n y fo u L 12.4 Use linear sundwd curvza for quantitation. Linear standard corvee sm isatciated far tbe analyte by Uneesregmiion using 1/x wrighting o f peak area veraua cabbcabon itandard oosMntrttioQ using MaasLym 3.3 (or equivalent) ofoeve eyatam. Page 42 o f t i Exygen Research Page 83 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygon P rotocol Num ber; P0000760 >-- irh M rte d M uster VOTO17U ! a n a ly tica l w r r o o p Method o fAnalysis & : fro DatancliuuJoB o f Perfluoraoctanole Acid (?FOA) in Fiih #nd Duna by LG14S/MS 12.5 Soaepte ratpona* tfw ild not oxeoed andarti rropcnion A ny samples ih n tx m standard rwpooaea should be A rth e i diluted a n ! reanalyzed 15.0 Acceptance Criteria 13.1 Qumaatograa nnnrtriicw apeak o f cfaugbter ion at 569 amu from a pareni o f 413 m u. Tfco 413 emu panm corrc^cnda to Du PFOA anion. while the d*U|Mor ion (369 amu) rtproaama (ha low o f carbon dioxide. 13.2 Matted blazfci muat not contain PPOA at )*vls po tte r than the LOQ. If a Manic ommdm PFOA al lavds p y u n than 0.5 ppb, then a now blank sample muat beobtained and the edr art muat bo ro-extwctW. 13 3 Rocovarioa o f control gpikti and matrix ipiko* muat be between 70.J30% of thoir known valuta. If a ootmol stake frifa outride cho acceptable limit. the oatni aot o fMinplos Aouid bo rwwiraciod. U A M y callbraboo atoodard frond to bo iw fc o c il o tu li by oaiof the Huge Em Tart, may bo mcfalod from fro calculation o f the calibration curve. However, the total number o f eatibratm standards feat could be excluded emutam teaod 20%o ffro total Lumber o fataodarda liyoctatL 13-5 The cottvbboa oeffirianl (X) tor oaUbrnkw curves fsoeraied runt be (X.1 WX3I3). I f calibration results fa ll outiida these lim its. then ifpfo^riaco stop tnuit bo taken to adjust instrument operation, and the rtaadaadaor tfaa raUvaat art o f M pM s ihould bo reaMlyxod. 13i5 Kmntkm tone* betwomt itandarda tad tampLas muK not drill more then 1 4 % withm wt analytical nm. If retmition limo drift exceeds this limit within an analytical run lb die art must bo leanalyxed. 14 0 CaJculaitooa 14 1 Um Um frOowtsgoqusrimi to cakolato the amount offP O A found (in ng/mL, based on peak area) using tbo standm l curve (luusax regmarion parameters} gn*f*udby tfcoMso Lynx software prepare: PFOA ftxad tnata Ll " ( Peek arte to u ra y t) slops 1*2 Use the following oqoaoa to coovart tbs amount o f PFOA found in rtg/nL to (!**> PFOA found (ppb) v [PFOA faund fn rtn L ) final volume IreL) x DF uanple woight(g) D P * factorby which tha final voluto was dlluwd, if nctoary. Page 43 o f 65 Exygen Research Page 84 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: POOO076O f c w f a n w * __________ ____________ -- _________________________ ________________M m d U t a + i i VOOOl7t3 I ANa LTO C A L M tT H O D | M e ta i fAntlyd im du DoenaiudM fftofluuoocMaoic Atid (JTOA) in Hw ird CtuubylC/MS/M* 14.3 Pt mmpi* fistiflad with known amount* o f PPOA prior le extrtctioa, u * die folfewiu# MfosttoD to O ikuiiie the percentro o m y . R e co vw y(S ) ^to u lia ly ^o fb u fld ^n ^l) - nalyU found in opm rol(pfi'g)] MkU)4eaddtd Om/ i > Exygen Research Page 44 o f 6i Page 85 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygca Protocol Number: POOOtHiO ANALYTICAL METHOD M rtlwdNum b: VOCOlTW M ate d ( Aaalyab ta r ta r D tn ra la n ta a ad P irfla a ra M lu a fc Add (PFOA) la VacdaUu by LC7MSAU Analytical Tortue F iirty: Exygw JU w rch 3051 lU taan* Drive Stale Cocfe, PA l4 f0) Approved B y p*u!CbwaoUy TacJuucal U k r, LC-MS, B x y fu Rjmm tJj g / / n f/U Y 3 5 * fSS^T ' VinoetaPPnwTxiWtiiUart, Opmboat, Exyjcn RatM rtb Orta Exygen Research Tout Pcm : 7 Png< 45 (if65 Page 86 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number- P0000760 UMaO M veoei 7|. I A .'V A l.V TlC A l.M iTH O D MMkxt o fA u ly tli fbr Iho Drtcm uuiran o /^rttu o ro M U n Acid (TfOAJ in V(0taiiar kyLC /M I/M S 1.0 Scope H i etho a ta be aployad (br A elatfo sed quantitation o/parfluoroocUAoic oid by Jiilfe Fcribroaoc Liquid Ctaroaacopnpiiy coupled to a tarwUm Mau SjW tnaaatrifl Dateetor (LC/MS/MS) ia vcfotaiicn. 2.0 3b*y 2.1 Abo,) obacrvcsafo laboratory practice. 2.2 Conaalt lb appropriate MSDS be&w handling any chemical for proper safety pncntica. 3.0 Saspla leqoinaM ot 3.1 AilaM tjgaTlH M apleJbrm &aotta!. 3-3 Soptes should he piu /asead before extractioe. F la n the fiozae templa in food f m m m md bom ofmos with dry ico Ploec ih ampies m containers d le * * op b boceo tarap n n ii|h t w aOow for carbon dkmta subHmaHnr Seal aad place (lie sample lo fin a n atorase until time o analysU. 33 Sample Section procedures w ill be specified bt m sampling plan for ihn pPCQKt 4.0 AcpentemdSemtvda 4.1 W ater-HPLCjrode 4 2 Accwototte - HFLC prade 43 Carbn (130-400 M el>)-fU iaa1 preda 44 Metaeool-HFLC prado 4.3 Silfos pel (60-3t)0 m uh) - Ixagar* prada 46 FknaU(M 00 tamh) - Reap proda 47 S u p a d LC-NHj 4.1 i-Octanol - HPLC jwia grade 4.9 L ^ico tb ia acid - Rasgad pad 410 Ple0iyldfohtareeilea~ Oaapeoi prade 4.11 Token - Raagent prtde 412 ABHnoolum AJW taie-A.CS. Keaati Onde 413 Heflueroeetaeeia Aaid - Siproa-Al*ob 5 0 Instrument md Equprasd $.1 A bipb perform ance liquid cbaoroatoprtpb capable o f pumping up io 2 antonU equipped with a w e ll volume liqoctar capable o f Reeling 5-20 pL eowaecled to a taoufam Mesa Spactromf'sr (LC/dS'M S) 3.2 A devieata collect raw data for peak ietagnuioB U xiqiuntiudott 3.3 Aa*l)Acai balance eapabte o freadinp to 0.00001 p. Pogt 46 q f 65 Exygen Research Page 87 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 ixrtM lM M Kk c ANAL VT ICAL MTTHOD Method o fAnalyei (or the DaMrzmiuliOfi o fPrfborooctenclc Acid PFOA) in Vegetation by LC/MS/MS R tury rv ^o n to r. 3.3 133 mL jmsj-shaped flu K 3-6 3flniLdiifO M bl*pely]prc>pyiuM cim fiif>bet. 3.7 l5 m L dispoeeblepolypropylene orntriftigrtobei. 5J D U yoe*l*ieaepipett(50-t0(XjL 100-200uLX 3.9 li5-fiiLLI>PEa*ffowHnot^fa bottles. 5.10 SmLclNT HPLC vini Idi. M l D ii^oitb2ipipivi> 3.12 AntopfOKM <100*1000 pL ad 10.100 pL), with disposal! lip*. 3.13 JPE tubo (20jiiL>(Sup*loo c*L oo. NO57177), 3.14 W rl*aeiloo*hak. 3.15 Centring* capable o fspinning 50 mLpohnP'Qpyfent'Ubeesi MOO ipm 6.0 OxOGUKCf*?fe 5yUHQ (.1 Analytici l Column: FVayhsea RP (Keystoco Sciam ile), 2.1 mm x 30 nun, 5m <P/N: 12305-0I JO 42 T n p n ta r*; 30*C 6J M A tli Phase (A ) : 2 mM Ammonium Acetate in Water 64 Mobil# Phase (B) : Motfanol 6J Qmdiajt Progrtm: Tima fm jftl OO 1.0 to 200 215 3L 65 63 25 25 4 Fkrw Ra 5Lfl fm Lftnkii 33 0.3 35 0.3 73 0.3 75 0.3 35 0.3 3.6 fctfoctloe Volume: IS pL(ee* b o ta n e * to aauracli u 50 6.7 Quantitt*: Peak A n a - external standard calibration curve. 6.1 Ran Tims: - 23 minuto. Tbs store Modtfoo are intended ae a paid* and may be changed in order to pennies the HPLC syttam. 7.0 MS/MS System 7.1 Mode: BtectaHpnyNaptiveMRM mode, monitoring 413 - 369 m ti foi PPOA. P ag* 4 7 o f 6$ Exygen Research Page 88 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygcn Study No.: P0000760 Exygen Protocol Number P00007M & yg*l MMirb ' _______________________________Msfeod N tdm VOW 71* 1 ANALYTICAL M1THQD M * S d 5 r 5 j5 fcr ttmB S SSm SS?o f PW borooconoic Asid (PFOA) in Vmumioh by LC/MS/MS Th bovo oonditiooa m intans u a gnide in ) jn *y b changed in order 10 o ptim al MSM1 system. f I- t.O fraontinof$oh(*am t.) M otte phi. 1 1 1 2 mM astOTmfttm steal b ctar Ja pf*pwd by siding 0.154 g of omnnwini HOM 0 10COmL o f watw. 1-2 BxtrgsJioA Solutions 1.2.1 2% aaeottio acid tn m s&iool it prtpvad by diw oivint 2 | o fa*codc cad i* 100 snLo f Datbaxk). 1-21 30% DUtehflddlcbJoftMltao ta toluene is propl by bnngiag 3 mL oddbnsthyMfchlocosiUiKta sih a lvo lu o M a n O ra L w ith ioIumc. Alternai votamem iy b* prepared. 1.0 Standard Preparation 9.1 S tu d v d Stoci/Portificatio Solution 9.1.1 9.1.2 9.1.3 9.1.4 9.1*5 Prepare a ccfc solution o f-100 >ig/mL oTPFOA by wdghing lOmg e /j*i)y d a d noted (ormted %e parity) tnd dilute to 100 mL with Stttasnol In 123-wL LOPB boari*. A 1.0 n g t o i ftm iflte ia o s o lltion o f PFOA is prepared by b ro ^ in j l m l. a f dm 100 og/m l solution itasi volani o f 100 with mctWol tn 125 m LLO fH book. A t ti |i4 *aL fcrtiiicstiaa solutia* o f PFOA is pregiami bybririgung 10 m L fto 1.0 pg/mLsoiutioo to i iu l VoluaMoflOO with mtdunol in 125 m l. LOTE batto. A'OOI jig ta L fbrtiAtefcdb solution o f PFOA is preparai by bringing 10 mL o f tbs 9.1 pgriaL solution to finn! vaiamo o f 100 with mtdmml La 123 m L tW 'S booif. T1 stock and fortification solutions act to b* itorad in s rofrigsrstor si Ifpraxiouedy VC and m nable for & maximum period o f 6 months to o th dais o fpreparali*, 9.2 Snodarti Calibration Solution 9J-1 LC/MS/MS oalibntioe s ta te d i ire prepared ksmeOvswol via dilution of 1.0 p tfn L fortification solution. Pig4ot ' P a g t 48 o f 65 Exygen Research Page 89 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 d Number VOOftP AiSAJLYTltAL M t THOD Method o fA n ily ii far Ih r Dcnrean rPerftoorooctwoic Aod (FPOa ) m Vegetation by LC/MS/MS 922 Tb ftttow ing U t typkeJ example: addi(tatui ooooiraQja p ity be prepared needed. Ceucaotratwo offatifieehoa Vofatoe DiloMd to Pin) OaBtentreliM Seiut h ta fo ilj _ (mL) IP JO OnL) 100 (W **iL 0 0J)J 1. 13 100 M 1.0 100 90S 10 100 n o n W IQ0 0.1 ID 100 a # 10 100 0.023 *01 0005 00)035 0.001 000 92-i Stan t il ealftnitaa -- dardi la lZJ-reL LDPE aurow-nwuth borila tf 2*C to 6*C to fix booCu . 0.2.4 Ah-- m vcJam and ronem ti'i timi1 o f atandeid maybe p n yw d aa Beaded. 10.9 BeachSet Up la i Badi bad* o f uraptaa vxtnctad (typically 20 or Ism) m iui include at leas: ty-- M b -- ad astool a d tw e -- H eotdrtU fott& ed 1 tow n oeec--ndoa* (kb contrai qnke) to ri procedural recovery for the batch. 10.2 Require-- Sor field and laboratory duplicala and qxkea w ill be pacified Bthe quality eaamnee plan ka dm project 11.0 S n pie Bimettan 11.1 Weigh 5 g o r fru c a M spie iato 50 toi polypropylene ceturifogc tubes (fcrri/y u ended, replace lid aod o ix weil). 1 U Add 30mL iatatauaH t and ik k on a w rirl action lU ic r ftu - lJ minutes. 11J Ceotrfugc the 50 mL polypropylene tubes containing (ample at -2000 ipm far -lOnnBBSa*. 11.4 fm k and oxidUiro the SPB tub kid nlaaixe (he pear-duped fluke. 11J Pack (he 20 ib L SPI tob im aequeace with 3 g Donati, 2 g eilte gai. 2 g carton, a d 1 g LC-NHj. Coaditioo the ooluraoa with 20 mL of methanol, than 30 mL o f acetonitrile. Discard a ll waehe. Do not allow the column la dry. 11.6 Siimi (be 125 n L peanhaped flir t* by riming with the 30% dimethoidicliktfotiLaae io toJium eotutioa Rine* the Quit with i )u m nee, followed by g rOwmcl (three cimai). O y the flatL i completely before ute, dthw by ilr-d ry tB i or wUh aatmes of nurogMv. 11.7 DocantQ Urect w to 1 couhtioned S?B caJuaaa flttad ioaide (ha motnb of tha pearehiyed flaak. Collect the eliuae in the 12J mL eiUrdaed pew-ihop fi* Page 49 o f 6i Exygen Research Page 90 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygcn Protocol Number P0000760 Nntaw V0001714 ANALYTICAL .MLTHOD Method e fAnalyvi fl dw Dctram raaiuttafPviliKnooclaook Acid (FFOA) ui Vetaon byLC /M *M S 11.1 Add 30 m L efaoetnailriletD tb r ra s p le in & 50 aaLeaacrifijf tub*. 11.9 Ihaka thr rarapte again feg ~lDm inut--oa owriet-actiac ihafcar. 11.10 C aatrfflifc da SOmL pofrpr opytwM xubee oQotaintaf ampk ai -2000 rpm 11.11 Derate: da Mate onto da h m flPE aohunn. Collrc* the chute inte the atme pearetuped fla k a d coatbina with da oUiant from da in itial tm ctlo*. 11.12 Jtapaat H jpt t l . l through 11.11 agata. 11.1) Add M teopa f lf 1-oOanl a da atm et Ihm p w ahaped Huk a d evaporate Mreduced pacaaura Qtaef 1 rotary evaporator (at < 40*C). 11.14 M ik f the final vohae, by adding 2 n L o f 2% wcovtio id in im Om o I to the yardbnped Oak and w ill a nrix/dtaeolve, )U S T ta ita r fra ctrats io KPLC viele iteog dUpoaebla pipet*. 11.16 A ialyw u np ka stag eketrorpray LCMS/MS. 12.0 12.1 1*2 12.3 12.4 12-5 lopect da Moa oun iU o f each etradard, M u^tk Mid fortified aarnplo into the LC/MSAdS xyetrarL A calibration standard must praoada and fiallow all Aaljcod um pia. S ta w lM d i o f PTO A e e rc a p o n d in i ta k s e l fiv e o r m ore ccn ccn tra tio n levels BHiM ba included in a tra ty tic a J a t. A n a n lira art o f extracted c a lib ra tio n atm d a n la im u t ba in cluded at Die b o g w w ftf and a t tha and o f a a m p le a t. E xtracted im d a rd i co uii ba m tenperoed between every 5-10 aaraplM . A * an a lte rn a tiv e . an enure te l of re tra cte d cabfcraiioc uadanta m ay be iqyected a t th e begjm ucg o f a te l fo llo w e d b y extracted ca lib radoa a ta n d a n ii in te u p n s o d rve vy 5*10 samples (to aecaunl fo r aoeond ite o f d u c te d ta m b fd i). to a O w ew e. extracted c a iilr a ln i standard! m a t ba th a fin ta n d la stn ye cxio e in a o ja p la a rt. Um lin e a r M andaid om vaa fo r tp ia o d ta b im . L in e a r standard c u rv e t arc generated fb r tia analyte b y lin e ro frw te o ta g 1/x w rtg h tin g o f p e a l area v n m cabV afco* ita n d a rd ra c ra tra iw c ostag M a n L y iu 3 J (o r eq u iva le n t] aoflware ytteoL Sam ple reaporaa should s o t exceed atandard m p n iK i A n y sam ple* that B K ceed standard raapaoeu abould be fa rth e r d ilu te d and re a nalyzed. 11.0 Acceptance Criteria 13.1 Chromatogram rauet how a p a rt fa daughter ioo at 30 ana from a parent e f 413 amu. The 413 anui parent oorraapoads I (be PFA muou, while the daughter ion (369 m u ) lepneanta the lota o feaifeon dioaids. P ag * SOo f 65 Exygen Research Page 91 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 /-* E x y fe o P rotocol N um ber: P0000760 tiy iw R j* --* r___________ a x a iy t ic a i M MKtodNurito Vixcvm r r g q p ________________ Method fA rn d y *to tt D tf* w tie a o f PerfhweoetaaatoAcidTOA^m Vsgetxtion byLC /M SA 13.2 Method blanks am t not conodn PFOA st levels prater Lhan the LDQ. If a Wank ocnUuu PFOA at levels paster than 0.$ ppb. then i new black ample bjm* be obtam d o ld the entire Ml m u* be i<x(noted. 13J R owwise ofcoorol y flew u d matrix *riksa anal be between 70-130% o f (heir known valuas- I f t eoalra) Spike U li outside the eeooptsble lim its, the t in m i o f *M n p lM tfw u id bo it-sotimrted. U .4 Any nalihnuba standard to be * stotinicAi outlier by u *n * the Huye Bate Teel, awy be secluded from da cafculsdon o f the csUbrauon curve. However, the to o l aurabw o f calibration standards d u i couki be excluded m u* notexceed 20H o f Ua total numbero f standards uyeeied. U J The oorrelukn coeflkao* (R) to caHbntttco o rv o i generated must be m.991 (R2 20.915) I f cehbfsiktfi results &JI outside these lim its, then ^greprtats eteys must be ttlo a te edjo* ioabvaenx operation, and the huidm& oribe relevant m i o f samplesshould be reanalyzed. 13.6 RatsstiM dm si bettvew stendads and renplM musi not d rift more than 1 4 % within an analytical nm. I f retenlion time d rift excoedstlui lim it within in u ulytke l nut thee: the setmust b* reanalyzed MO Calculstuns 14.1 Used* follow ifl^ eqpat ce caleuto the aoootjss ofPFO Aftw vJ [in rtgtaL. based on peek *) uclng (he standard curve (linear regression psrwnstws) fpoeraiad by the Maas Lynx software program: PFOA found (a f'mL) - fPaak area in itc u il stops 14.2 Use da following pisdoa to convsrt the none* o f PFOA found in itg/mL u> a^(ppb). W Q A ftm irt h l rF P O A ftnmdrns/mi w final w ilia n fn L U OFT sample weight (g) D P -fc d c f bywfakh the final raiatiM W w d itto e d . if necessary. 14.3 f t r sanipto fe tifie d wifo known amounts TPFOA prior to cntsuclion. use a follewkg equation lo nefeisiato the p a ra recovery Reootery (%) - [ total snafrta found(n ffr) snslyte found ip control (ng/g)] [W a u fy te td d a d fo f'g ) h ir i 7 P age 5 / o f 65 Exygen Research Page 92 o f l l O Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 ANALYTICAL METHOD M ifrorfN om fer V000PI5 M e to d i A oalpfc llv tte D tfv m lu d c i tfP trflB c rto rtM c k AcM (FTOA) la S a il M a u l Liver t 7 LC/Mg/MS Analytical Tm df Facity. E xy* SU*Mrck 3051 H in t b Drive StMCoU|a.PA 16101 Apprevcd B y v t A C--fL Pw CPanoUy Technical Laadac, LC-M3, Bxyjm Research lU tfc fM Date a //ft / / / V & n F kh a rty / Vice Preshkni, Dpardioae, Hxyjcn Research D ite Exygen Research Tout Pajm : 7 Page 52 o f 65 Page 93 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P000Q760 Exygen P rotocol N um ber: P0000760 lejFSW lUm itil Nuotar V000I7JJ 1 ANALYTICAL METHOD Method o fAjadyaUfotboDctcnniiudioii afPtrduorooctsomc Acid (FFOA) in Smell U W n aJ Uvar by LC'MVWS 10 Scop* Thaa mttb-od i tobn ooptoyod fo the iuJMieo tod quabUtion o f parfluoroacianpie acid by Hash PwJfaraunM Liquid Clcuiutagrapby conpletJ to a tandem M iu gpcotnuttflric Potoctor(LC/M S/M 3) n u ll m ir in i liv *r. 2J> $afey 2.1 Alweyi obcnrv* afe laboadotyptnctioea. 32 Conduit Ou appropriata M&DS b e fo t fcaadlmg u y (toomical for proper iifc ly pnoubaoL 1.0 Sample X/kjsbranavi 3.1 A i Ica* 3 g o ffra t i --p it fo c rtra c tk a . 1.2 SsapUo tbauU be ptoccwod W o rt traction. Pfeoo Ih* fixraen temple in a food pioceuor m l ta m o f'd J t with dry io*. Place 0 ram pici in ooiuiim it ad vt. fi a frozen atonge ovcangki to allow for carbon dioxide sublimation. Seel and place toc ampie* > frocen ttorage until lime of aelya. Alternately. I f there it u> Laaufficicnl amount of ample (-! > Hum 3 iX *ao no proooaoinj it necaaaary and die ample can be used ai supplied 3.1 Saasfk collectioo procedura w ill bo vpoolfled in ibe runpling plan for this jwojart. 4.0 tcegoata od Standard* 4.1 W afer-HPLC gode 4.2 Metoanoi-HTLC grada 4.1 Aoctonibile --HPLC nde 4A A nutttthsa A D *tttc-A .C JS .fo4fni Grado 4J Rnfluorooetanoie Add-Sigm a-Aldrich 3-0 InatmmaBtaadlqviiptom 3.1 A high perforatane liquid ehroatOjfr>i capable o f pumping up to 2 olventi quippad with a vanablo velane ipjeetor capable o f injeciing 5-20O liL oottifecfed to a tanfcn M ia i Spactmnator 0/M S/M S). 3.2 A device to coHoot row data for peak integration and quantitation 3.2 Analytical balance capable o f reading lo (1.00001 g, 3.4 30 mL dbpecafcl* pelypropyto* corttiAtgo taUa. 3.3 13 mL diapotabl pcdyprcipykiw ntriftigo tubar. 3.6 Diapwablemleropipew (JIMOOuL, IOO-2QOuL). 5.7 125m L LDPB aarrw-mtrarh botti#. 3.1 2 *L tlaar KPLC v ia lto t Page J J o f65 Exygen Research Page 94 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygea Protocol Number: P0000760 fc/ F U iM *m*VWOl7|J ]| AKa Ly ITCAL METHOD ^ 5 n fA a iy ^ ^ tt Dv*wuiiiJ>tioBo/HrfliwQocttBOM; Add (FFOA) in Spall M a n u l Urm by LCA4MS 10 Qfcpoaafai pipiM*. 1 )0 Autovlpen000.1000 iiLmd lO-lOOtiU wi*b di*>obletipa. i n w s ^ P a k V M 6 o c (i* )ic n s n ( rtrid fF . 112 3PC new m nunllbld. 112 Tiuuamztt. 5.14 W rii w w abak. 113 CacxriiU|*E4fB)>kafiy>nvk 15 cnL polypvpfyttir tubat M3000 rpm 6X1 O n a u td ^o U e S yM * 1 ) Ae*iytic<lC oht:PhioflM W R P(Ks)ietooe9oiatilic).2.1 nam x 50mr>SH (WH: 12900432130) 6 2 Tkn^M ha: XTC 0.3 Mobil PfcaM(A) : 2 m if A nnnaiian Acetate ca Waler 6.4 Mobile fbMC (B ): M a tte d 15 Gladiant PiOjjjmn: Tima 0. 1.0 10 200 22.3 FW Re SLA a * (lal/mia) 65 35 0.3 63 35 03 23 75 0.3 25 75 0.3 65 35 03 6j6 Inject* VohmM: 15 jrL (c*a b h n ric d 1 M much u 50 ^L) 6-2 QuacotJtior P*k An - e*tenJ jundnni odihratjera curve I t Run Ti m : *25 minuftu Tb* abowooditioa < iotsndod *ai juxi m l nay b changed ln ordw io opdciiMthe HUC lysenn. 7.0 U/M5 5y te 7.) M odr Blacltp^iniyNoptivwM RM node. oaitori*$ 413 - 369 itirt for PPQA. . Tb ik o n oondixk arv minded at t pud* and may bs dunged in der to optiviriiatha MSMS lyttera. Exygen Research P ag* 54 0j 65 Page 95 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 Exygero P ro to co l N um ber: P0000760 B>)nw 1iMim^ dN lobn VBOOITIJ ANALYTICAL M tT H O P ~Medwd oftody ftrtb e Determination o f forfliwreocttD oic Add (PPOAl in Small M aram i U v by LCAfS.'MS t.O PuparM ioa ofSotoSom 1.1 Mobilaffan I hccUUe In water ia pn?nd by W it 0-154 f of to 1000 mL o fwater. Alternate volume mmy be prepared. 9.0 to ad ied Prcpando 9.1 Standard Stoclc/Feriiflcation Solution 9.1. Prepare a deck aohtim of -100 p jfa L o f PPOA by weighing 1 0 mg o f M ly ftN i Kaacard (oeooeted fcr purity) *nd dilute to 100 mL with mtbaaoJ m. a 12J*mL LDPE bottle. 9.1.2 A 1.0 fertifii*ticm eolation o f PFOA it prepared by bringing I mL o f the 100 p jfa L eoXeioo to ftuU voiumr o f 100 w ith methenol tn12SmLLDPZ bertda. 9.1.5 A 0.1 ptffaL fbcttftcaiionaolurioaof PFOA la prepared by bringing 10 m L o fiM 1.0 |ig fa L u?u lo a final volume o f 100 w ilh mtthaaol In a (2$ m L LOPBbottle. 9.14 TbcrtocJc tod ftalUkrtum eohuwxu ere lo be ttord in t reftxgeraror il ippcoxlmaialy 4*C end an ttabi* for a maximum period of 6 month from the duo o f preparation. 9.2 Standard Calibration Sotattax 9.2.1 9.22 LC/M&/MS calibration findardi an prepared ia m oth*)] via dilution o fthe 0.1 pg'mL xtiUcahon aoUatwu. T V following i i a typical n im p fc: additional concentrations may be pRpared u oMded. CoocetuntLflft Fimi wfftxtiAcalks Votame Dthriadto Concentration SoJuuon(n*L) i* U 190 14 (m l) 100 0MiLJ J.O ICO 2.0 100 2.0 loO 10 10 1.0 5,9 10 109 20 10 100 1.0 10 100 05 0l2 Li 92 J Sion i l eafibrabon etaadarde la 125-mL LOPE narrow-mouth beides at 2*C to 6*C, if> to rix mondi. 9 2 4 AJksvut vomm and oonoeotratioos o f Kandarda may be prepared at mML P ip 4 rt Page 55 o f 65 Exygen Research Page 96 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 B*ygen Protocol Number P0000760 LfTWi|M w tt VQQC1713 I . ANALYTICAL liUtTMOP Mefeod o fAnalymi forth* Deacaninafiofi o f Perfliiomootaaoio Add (PFOA) in Smmll Mammal Uvar by LCVM3/M3 10.0 Batch M U |i 10.1 Each beteh o f Mediae axtrecmd {typically 20 or k u ) muu include at leesi on* m treated uw trol and two untreated controll Fortified at known B a nw aW k- (lab control ^ok) la ra tify procedure) recovery for the batch 101 kequiniM B li (Wr field and laboratory duplicates and apikea w ill be specified in Ox quality aMracc plan Jbribupaqjeci 11-0 Sampio Extraction 1U Weigh 1 o f Mmpi iK> a 50 nal polyqwopykna cmfcifege lubes {tovCy m needed, Mpbco lid and mix well) Not* that stornate weight! oMivr m*y be measured depending on Iba maple lire available for ut>. 11J A id waMrlo 0m tempi* ft* a Anal volume o f)0 raL. 1 U Kflmogamxeaaa^Ja M ngam am ier Hbf-1 minute. 11.4 Transfer I n L o f the ample ruing t disposable pipette into a 15 mL epeeebtootwtaiagelabe. 11.5 Add >mL o f Ketooitnie and ibsk*for-3Q minute* oti aw rin-sction ihakar 11.6 Cm trifltge the toben at -3000 rpni for ^3 ruinate*. 11.7 DecaaMb* mpemaiant mto a 50 mL diapoubk centrifuga tub* aad add 35 n L a fw a iv . U J Conditioo the C it SFE rtdg** (I fo 6 mL) by pusing 10 tL methanol ibowedby Sm LofHPLCw aM r(-2 daop/tec). Do not Jet eohonn nut dry 11.9 Ltmd the samplecat conditioned Cn SFE cartridge. Discard aiuac* 11.10 BkOs wide 2 mL o f methanol. Collect 2 mL o f doatt into a graduaied L3 blL polypropylcni ocatoifuge tube (final volume 2 mL). 11.11 Analyse Jaopleuin|ol*ctaoq*ftyl/M 8/M S - 12.1 Inject the awn* amount o f each dm fardi ampio nd fortified ample into Ihe IC /M W t lyataca. A dfbratioa atandart must precede and follow ill analyzed aampls. 12.2 gtaBK&wdeofPFOA oearceponding to at ieeat five or more concentration levels auut be toetodad m amuulytfeaJ set. 12J An entire aet o f m ilbreion ataodaida moat b t Included at the begiim iiig and n the ml o f a am ple mL Standard rauet be intanperaed between every J-1ft emplee. A en sltornave. an anti aet o f calibration uendardi cruy be rejected M the begtzsiiog o f i M followed by calibration pandarck inbuepmaed every S-I emplee (to account for a aecond aei o f etaodarda) In fiber cei, calibration standards muat bo tba flrai and last injection in a Sample eat. 12.4 Urn lo o r atanrlsrri eurvua far quantifaring. Linear atenderd curves are generated for 0 aosiyto by Uooarregreeawc using l/x weighting o fpeak area P age 56 o f 65 Exygen Research Page 97 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 EiQUftJU-- di__________ MrttfdKMbwVOOOJHJ I a n a l y t ic a l m e t h o d Method o f AnsZyeisi x Dstom teatieii n fPerflimooctaooic AbId (PFOA) in Small Mammal Liver by LOMS/MS vwaua oifantHB ttJDdvd coocwtfntoca w ing M atiLynx 3 3 (o* quivalent) itA w m iy ita a . 115 u ifle reepemae dxxiLd dm *ted lUndsrd rwpoMM. Any iim ples ihi exceed U ftfardttapauM UwU b farther <tthu*6 md raarulyicd. 13.0 Am ^ U mm Criteri* 13,1 ChrowMo^wn muri bow * pak o f t daughter ion at 309 m u frxxn a peroni o f 413 * doul Tba 413 tmu parent MrreepcaU to 01* PPOA anion, w h il* tlu daughter lm 0<9 m u ) reperente Uslo u o f cwbco dioxide. 131 Method blrake m wt not ontani PPOA at W vsli jw a lcr than ih LOQ. 1/ blank coatau PFOA at levai* p e e l than 10 nf/g. then a new blank ample m m be obtained m l the oath* ret nuiet be ix u n a e d . 133 Reeovwita o f eotUrol if ik * d n u lrix e p ik* m iK be between 70-11Q% of (hair koowo valuta, I f a eottm l iptic* fella oohide th* taceptabte lunata, fee attire le t o f samples ihook) be re-axtrectod. Any matrix pike ouuhJq 70U 0H afauld be evaluated by 4m analyst te detannine if re-extrawioc it 13.4 Any cittueriM i Modtad Sound to be a atarimeal outlier by using tbe Huge Error Test, may be euntodad to m the cakelcttae o f the calibration curve. Howaeat, (be total cumber o f cahbntiod standard* feat could be excluded bum net exceed 30H f fee total awabar o fataadarde h\jecied. 13.3 The cenelatkm coefficient (R) for calibration curves general! mu be 0,992 (R10.943). I f calibration rceuLtt fe ll oalsidr ihete linnu. then M prcpiitte etey* m ist be taken to adjust ioftnenent cpration, and the rtaediidJ or Ike ratovant a * o f um pfcedwuld be reanalyzed. 116 Retention tknes between aedanls and u n ifie s m ini not d rift more than 1 4 W within ananalytical run. If retention time d rift exceed this lim it wiihiu ananalytical me Ok* the m i must be rw ulysed. 14.0 Cakuleriona 14.1 Uto Italbllew tofoquaiiaa to calwlato the amotmc o f PPOA found iin ntfm L W d on peak aree) wieg fee standard curve (linear regression paruntcn> genaratad by the Maat Lynx ioAwert program: PFOA feimd (nytavL) - freak area - Intercept) s D F r atiqwol feetor top# OP -factor by which the final volume wasdiluted, if necessary. Afiquol fector 10 Exygen Research Page 57 o f65 Page 98 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study Mo.: P0000760 Exygen Protocol Number P00007&0 V*nMkMMc MwbtdMi VQOQtTtJ r AN ALYTIC AL METHOD Method o fAiMyiU 6 c S DSnSda c fPort)oorooettnein Acid (PPQA) in Small 14.2 For unuaa fcatifttd w ii tarara traoums o f PFOA prior to u tm tio o . u the folkw og equtiAO to cak*)* ibe prol m xm ry. Rtoovory , [toJralyl fan d (affati) - ndyte fated in oootroj (nftnL)] taulyt added (affanL) M 3 Um th* IbD ow iif eqnidaa lo ooo w i the intougt o f ?FOA found in nj/m L io f t o a !6<iad(pp)ffrQ A to a d in ^ m U x C oli t t ia n c im i. copi weight (f) Exygeh Research Page 58 o f 65 Page 99 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 E xyte n Protocol Number P0000760 A N A L Y T IC A L M E T H O D k M u iN B * < r VOOOIT16 Mtfted ai AMutrik h r h t D rlv a k u lM a i F a h n n w l u a k Ada (PPDA) li in ali .VlmUMJ I m a bjr U M I /M I AtAjyocil Tcacmjt FwiUty: Rrytr. K***iti 3058 R otearci) D riv * Strt CoUca, PA I 101 A p p ro vi By: Pm U C oonotty Tooknicai Lcmm, IC*MS, Exyjon Roaetctfc q //7 ? / & / _________ .M ttH ifcacty ' V ica P la id a n t, Op a rtic a * , E xyp m P.eaaroh D at D tf* Exygen Research T o lti P u fe r 7 Page 59 o f65 Page l OOofl l O Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 xyg e n P ro to c o l N u ro b o r: P 0000760 Ixype Uw eib MtffaodMmfor VBOOI7I6 | ANALYTICAL M THOP ~~ M tS ot AaSJeu K S ESSmS S m ofParfliMMoUiiou: Acid (PPOA)S Small Mammal 9nun by LC/MS/WS 1.0 Scope T u* method it to be employed for (he ieoleticn md quantitation o f perfluorooctuioic K id by High Perfoemexe Liquid Chromatography coupled to t tandem M u t Spectrum Oetecw {LC/MS/MS} ia m ull m m au] eerum. 2.0 Siftry 2.1 O weyt obeeve u ie labontory practicee. 2.2 C ocidl the appropriate MSDS before b s x fl i eny chemical for proper n fc iy pwamiem . 3.0 S n ple Requinmaol 3.1 At ta d I o L o fla o t saiqdsfijrutm e tiaa 22 Ne eemplo ptoeeeezig Isnowfod for * u m w n plw . However, ftomn aerum aofoleefoU to allowed to aw pJrttlytfaow to mom teanpanture before use. 3.3 Sample ralkrtw n procwLrw w ill be ipocUiod ia the w npling plan for thus AO R m fm li aad ftaadarde 4.1 W ater-HTLC ptd o A2 Methanol - H TtC grade A3 Aeetonitrile - H H C gnda ^ 4.4 Aanaorimn AceUfo-A.C.S. SUepot Grade A3 Partuoraoctanic Acid-Sigm i-A M rlch 5 0 lACtroaxot modBqeipmcal 3.1 A high perfonaaaca liquid ctawutograph capable of pwrgring vp to 2 olveata equipped w ith variable volume Injector capable o f qjectntg 5-200 yL eooMeaed loaUndem M ua Spectrometer (LC/MS/MS). 3.2 A device to collect rear data for peek tatopvboa and quanatarian. 5.3 Aaeiyticj] balance capableo / raadiagt 0.00001 g. SA 50 mL diapoeabk polypropylene cm irifoge tube*. 5.3 IJmLdifoxMabfcpolttrQpytaDacenfflAtgf Mbee. M Disposable mkeopipet* {50-100*L, 100-200uL) 3.7 IlStakL LD Pfi tunoar-iboiili boclea. 5.1 Im L e fc tf K F lC viahdt. $.9 Diyoe^vle papcho. 5.10 Autopipcttce (100*1000 pL B id 10-100 pL), w ith disposable tip i. $.11 W atm S epP akV ae6cc(lx}tC ll SPBomtridgM. 112 SPB veeuum m anifold 115 Vertexer. P ag* 60 o f6$ Exygen Research Page 101 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 E x y je n P rotocol N u m be r P0000760 IxyjM k M K fa M rih x l V um M i VOCIO17 1 AN g y n C A t METHOD Methodo f AivysU for the DotonwoMion /Perfluofooetanoic Acid (PPOA) in Small MtmmaJ Strum by LC/ML8/MS 5.14 W ilaioctioo akakar. 5.13 Ccutriftiga c^poM o f ipim unf 13 mL pclypnrpykn tuba ai 3000 rpm. 30 CbroraMorapble Syitem (.1 Analytical Column: Fhiopbaa HP (Xeyeton#Scientific), 2.1 mm * 50 ram.5p (F/N: 8250S-Q52130) 62 Tctnpcranyg 30*C 6 J Mobile PtuK (A ) : 2 raM Ammonium Acciai is Water MbM.lvP hicefB ): Metiunol Cradiost Program. 'TW fa.lnS ao 1.0 1.0 20.0 125 s a 65 65 25 25 63 Flow Rat SUI ftn l/m in 35 0.3 35 0.3 75 0.3 75 0.3 33 0.3 6.6 bwfcon Volume: 13|LL(om beireoloa noch 50 oL). 6.7 Quantitation: Paak A rto - XMmlJ Mandatd calibratimi curva. 6.6 RenTim: 23 niouiac. Th above conditicai* ire iiundadao guide and maybe changed in order to optimise Am HFLQ lyateaa. 7j0 MS/MS Syncro 7.] Und iaetnwprayNogative MRW mode, momtaring 413 -* }69 m /i for rFOA. . The bove ooodrtwni an intended u t guide ad m*y be changed in order in optim a* the WSM3 yearn. 8 .0 P rq ra tio o a fS o te tio iu t . l M o b il* T h a n 8.1.1 2 mM ammonium set! io ter i l prepared by adding 0 154 g o f anvaonum acetate lo ] 000 mL o fter. Alternata vohuoM may beprepared. r*f< >or- Pagt 6 ! o f 65 Exygen Research Page 102 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 fa y j- ram rJi " Mrftod Wwbw V00017>0 | AN ALYTIC AL M ITH Q D ketbod o fAoaiycij Cbr lb Defannrwticn orPcdlooroocUooic Acid (?f 6 a ) in Small Mnrvmai S o w by LCTKtS^MS 9.0 Standard Prepariti 0.1 Standard SlocLTxtiflcMiOtt Solition 9.1j Prop* a Hack solution o f-l pjpW . o/P fQ A by w ciihinfi JOmy o f MuJyoci) m lan i (oorm ud far purity) and dilu to 100 mL uh modund ia a 125*faL LDPB b o tili. 9.1.2 A 14) p fm L fiutifuation solution o f PFQA i* prepared by bringing 1 mL o f tb 100 H ff'nL tutina io fim i volum i o f 100 w ith mcUunoi a a 12$ m l U > n botti. 9.1J . A ftl|^M Lfb (bfiu H oB obtioa afP F O A ip rp w 4byb rirtsin || IO mL o f Bm 1.0 \iffaL relink io fttu volume o f 100 w lib nwirniot in % l25m LlXrPEbok. 9.1.4 Tb* ito c i and factiOcatiai aoluliooi ra to b farad in a refngcretor At approximately 4*C and mmaUbfc far a. maximum period o f * mootris firm the date fpreparata. 9.2 Itaodiad C alibrati Sohkfare 9.2.1 LC/MS/MS cahbcthon ffanriairia arep repared in wlhanot vi# dilution o fSia 1 p j/o iL femficedon solutaa 9-2.2 Tb folfawfag ia a typical m n fk i: additional luulcjju arim i may be fn p n d aaMtded. Cdnoceoaboa fin a l fPaitilLtttfan Votum D ihM dtt ConMnmciofl Sohkco (aatnU 100 too ICO indO Ja 2-0 ID i U 100 100 100 (n j/m i) 5. 2.0 ID 3D 10 100 2D 10 100 ID 10 100 J 0.2 0.1 9J.3 Store afi edibreta itaodudt fa 12S-mL LUPE narrow^noudi botti ai 2*C to 6*C. ap iix a u t k 9.2.4 Alternata vdajor and ecnecntration* o f standard* ay be prepared a* ttoodod. 10.0 Batch S#i Up 10.1 Each brtoh o f maple simed (typically 20 or fare) nut include at lean oat mumtad control and two ntTM! controls Unified at known CODMOtrattoni (lab control apik) to verify procedural recovery far the botch ]D.2 ftcquriamBta far field and laboraaoiy duplicate and ip ik i w ill b specific*! fa iha quality aaaunee* plan far (M l project. Page 62 o f6S Exygen Research Page 103 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Nurobcr; P0000760 Exyyii***aucl * * * * * N Wt VQ0QPI* I A W /UtnCAL MSTHOD lUtsoi 9tAalyB fcnks orftriuwooeunoic Aciii (?POA> in Smill Monuooi3nim V/ LG/MS/MS [ ll.O Strap]* Sxtracdan 1 U M u m 1 ttL b f sample into a 50 mL polypropylene ccottifog tubes (fortify m nwied, teplac lid and n ix weti). Not* dial alternate velum** of mum may b* measured dapondini on th* aaaaple tile available for use. 112 A idw trtoibss pfo for* fosaJvoium safiDiuL. CapUgMIy 11.3 Voctex ib r - j mmut*. 11.4 Tranafrr 1 mL o f tfca atapU usfag a dbpcaabl* pipttta into a 13 mL tfasposaola caasrifon tub. 11.5 Add 5 m Lof accionidik and diaka for -20 m inalu on a wriJt*aciion shakai. 11.6 Ctoixifos* tJmtub at *3000rpm for -5 anna. 11.7 Decant ifce nrrnTaiU info 50 tnL dippoaablt cntrifk|a tube and add IS mL ofwatar. I I . I QfidiMo*. tho Ci 5PB cartririgaa (1 g, 6 m L) by paasiog 10 mL mothanoi ibilow id by J L o iM K C wfow ( - J drtp/H c). Do not l# i coiumn nm dry 11.6 Loed ths am iyk oondkiowcd Cu 5PE cartridge. Discard duatc. 11.16 Blet* w iib -2 aiL of medunol. Collect 2 mL o f ekua* into * graduated 15 BiLpotypropyHens esantnfoga tub (final volum e- 2 mL). 11.11 Analyze sampim wring ekcdoqpray LG/MS/MS. 12.0 Chraemnynphy 111 Inject tbc tame amooetofacta standard, sample and fortified sample into the LGWS/MS system. A calitnatjnn standard muat procod aod follow all analysed uzapJoa. 122 Standards o fP fO A conrwpoadlng to at loaat liv * or note eoneentrsiran levels ra n t beincluded in pa anaJytical a. 12J An eitinc soTofcalibration standards ntuet be included at the beginning and at the cod o f asample ML Standards meat b intsrtporaod between very 5 -10 ampler. /ts an alternative, m entire set o f calibration standards may be injected M Che bagtaaring o f a set followed by calibration standards interspersed rm y 5*10 sample# (to account for asecond w t o f standards). In other case, caHbratkn standards must be tba Aral end las! injection in M fk lA 12A Um faaar standard surras for quantitation. Linen ilacdard curves are grentad far the relytc by linear regrcaaioo using V* weighting o f peat area rareua calibration standard w oemfratson wing MatwLyitt 3.3 (or equivalent) wftwar* tyrtani 12.5 Sample respout ahtrald not exceed standard rrspomej. Any samples that excreri ataidacd napoo*** should b* fartherdiluted and raaoJy*d. ftp so r' Page 61 o f 65 Exygen Research Page 104 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Sxyj.ec Protix! Number P000DT60 f r y f l i --wb_______________ -- _______ ________ Mated Nwabvi VOOOI7H j r ANALYTICAL HKTFIOO ] f t f C S S n S S w o fPriflaotooctmcc M M (P fO A ) in Smal1 ~ ................. - M im nal Serum by LC/MS/MS 13 JO A u B f t v m Ctfiarfa 13.1 O v o m o p ia muriabow peak o f adaujhtar ion 1 W amu boro pareri . Df413 tcm . T ht 413 amu parane ra p p o rti io ta PFOA amen. wtidc u * dMRffatjr ja {39 am i) npfW M ttu to ofoubM dtoxidL 13S . M attici bim bi miu t ara fltaa PFOA lavala g ro ri* thm Um LOQ ir a biadi contala* PPOA al M a M tuo 10 a |/ L tbcn a tM bUnk uop fc bum ba obulft! andtb# aulire a * muai ba ra-axtracrsd. 13.3 Rpwvwiaao f oortrol palata and B rina tpika m uri ba btrw etn 70-130% o f Ihair V n iw i vaiata. I f a m ira i aptka M ia aotiida (ha aooaptable linai, ih .potira aa> f rampi rbovid ba va-mrtraoted. Aay aiW rit *pike outiide 70130% ahonU ba vafeatod by (ha am i) lo driamain ir nc-em ctwn i> w a nutad 134 Aoy calibrado ataofenj fbund ab * atadriical c a d * b> ujhj the Hufc Erro Taat, mxy ba xclnded ftcao fb calrulriio n o f tha Cmlibniioo curva. Howevar, tha lotal number o f t librario ttasdand* t ir i couid tx excludcd BW KBOlcacaadJOHefdataUlpunbvofaiaDdrids byacied. 133 Tb* corralriioD cocfBdraf (R) far catibniioo curva gaasvted muri be 0.992 (R2 0.905). I f M iibrriHB rondi {ll outridc th*e lim ili, thcit appropri!* * 9* m u t ba (aitali to ajuct kurnm unt operano, ad ihc itm d v d i or ib i raiavaat n i o f am plia ahould ba raanilyzcd. 13.6 Rriantion lim ai batwra aUodarti and Menata im ut noi d rifl m ori than ' 1 d % witbioo analyiic*) ru. Ifreaantlao l ncdrift e*cecd dii* lim ii wiihjn mi loalyiioOl nm Umd dia aatm ori be roani)yd. 14.0 CafcetaiJoiw 14.1 Uratha foUowing quatto io calculated) mount o iPFOA found (in rvjrinL. baaad a paak m ) unni fca m drid curva (lim ar n p w iW ! panimitara? tta ta m by fl Mata Lynx oftwira program: PFOA fbmd (agfmL) - (fa ti H l intercept) x DFx aliquri factor dopt DP - fiatar by whiditbo linai votame waa dihuad, i f neooarary. Aliquot fc n r 20 14J For la m pi* fortified with known amara o f PFOA (vier to extraction, use dafollowing aquation to oakiolri* the porreoi rocovray. Recovary (H ) [ loto! rariyN txm d (ngbmL) - anal)forato atoaOol <ag/mL)) >1Q0 _______________ -- lV taaddad(otW _______________ ! _________ P age 64 o f 65 Exygen Research Page 105 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 Exygen Protocol Number: P0000760 Hcftta U m r * M tb D d N a n te iV O O o n a * r A N iq V T lC A L METHOD Method o fA n d yib fa th iD d e n m n lio D o/Parflviim ocUaoic And (PFOA) in Sm*H Mwnraal Seno by LC/M3/MS 14J Um An foltarrap equation Io convert th* araonatofPFOA found in nj/m L to ppK ro fU m fe fld fm /m lA /b a l volume (mLW Mmpl vtriunw (mL) Exygen Research r**' *' Page 65 o f 65 Page 106 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 3058 Research Drive Phone: 814-272-1039 su te College, PA 16801 Fax: 814-231-1580 EF*QTQQ1--AMENPMENI Amendment Number 1 Effective Date: 01/19/05 Exygen Study Number PQ001131 Client Study Number. Page 1of 1 ESCBIPTIQNJDF AMENDED SECTION 1) Analytical Procedure Summary V00017SO:Sect)on 9.1 2) Verification of Analytical Procedure None AME N P 5 P T9 1) Add to Section 9.1: Section 9.1.8, Alternate weight* of standard* may be used to prepare alternate concentrator of stock solutions as necessary. Alternate levels of fortification solutions may also be prepared. 2) Low and high spiking levels of the analytes for each matrix may be altered depending on sample size available for extraction and/or to cover analyte concentrations expected In the sample*. BAffimS 1) Higher concentrations of standards need to be prepared In order to spike the sample botttee at higher levels. 2) The sample size avileWe foremen mammal Iver and serum was smaler then expected. Spiking at the predetermined levels In the protocol puts the spiked concentration lower than the detection limit Also, the analyte levels In the ground water samples are expected to greedy exceed the predetermined spiking levels listed In the protocol. When the levels Inthe samples greatly exceed the spiking levels, an accurals necwvery value cannot be calculated for the QC sample. Higher spiking levels In the bottles win cover the analyte concentrations expected In the water samples. IMPACT ON STUPY The LOQ is 100 ng/g for a 0.1 g sample of email mammal liver and Is 1000 ng/tnLfbra 0.01 mL sample of small mammal serum. Higher levels of spiking for the water samples will ensure that more QC recovery data can be used. LIBRARY C:W tXM122frS / Exygen Research ADMINISTRATIVE FORM Page 107 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 3058 Research Drive Phone: 814-272-1039 State College, PA 16801 Fax:814-231-1580 Amendment Number. Effective Date: Exygen Study Number PROTOCOL AMENDMENT 2 03/07/05 P0001131 Client Study Number: Page 1 cf 1 Nona DESCRIPTION OF AMENOED SECTION 1) Report, page 11 of 85 2) Teel Materials, page 8 of 65: PF08 transition monitored 499 -> 99. AMENDED TO 1) Instead of one finaf report, Interim reports will be Issued. 2) PFOS IransWon monitored may also be 499 -> 8b. B&HSNALS 1) Due to the excessive stees of the data sets, interim reports wll be Issued to allow the dlent to receive data in a Umelier sm r " ' *At/Vr 2) The API 4000 LC/MS/MS systems detect the 493 -> 80 PFOS transition with greater sensitivity than the 499 -> 99 transition. IMPACT ON STUDY 1} th e dent will bo able to receive and review the data more quickly. 2) The 499 -> 80 transition can be detected with greeter sensitivity, therefore, giving better chromatography. S fw e w S lc n q w * (V re q iilrs d ) O L 8 R W Y IO: V000122S-S ; DM * Exygen QAU Review XI O s its ifS AD U IN ISm A TTVE FORM Exygen Research Page 108 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0000760 3058 Research Drive Phone: 814-272-1039 State College, PA 16801 Fax: 814-231-1580 DEVIATION FORM ____________________________ __________________________________________ Pag 1 of 2 General: X Project Specific Deviation ____Facility Deviation Date of Occurrence: 04/28/06 Exygen Project # : P7B0/P1131 Deviation U : 5 Client Project # : ______ NA______ Reference # : 06-076 Regulatory Driver: Deviation Type: (In clu d e VU h r m ethods a n d S O P s) _____ X _____ ____ GMP GLP Other None Sample Description: X Protocol ___ _ Method VU: NA Notebook reference: NA ____ SOP Login#: L0006191 Container#: NA Lot#: _________NA Summary of Deviation: The three sediment samples In L8191 (C0172892 - C0172894) were originally extracted using the sediment method V0001782. Poor recoveries were obtained for PFOS, PFOA and ,5C PFOA. 8ecause of this, the study sponsor requested the use of an alternative extraction for these compounds, as follows: Direct Injection Method: Before tea samples were weighed for the extraction, they were mixed thoroughly by vigorously shaking the container. A one-gram portion of sediment was weighed into a 15-mitMrter centrifuge tube for the extraction. Ten milliters of 1% acetic acid In methanol was added to each sample. The samples wsre then shaken by hard, vortexed, and sonicated for thirty minutes. The samples were then centrifuged for -10 minutes at -3000 rpm. Each sample was analyzed by LC/MS/MS elact/osprey. Using this method acceptable data was obtained for PFOS, but the recoveries for PFOA and ,3C PFOA were still poor. Another alternative method was then used for PFOA and 15C PFOA. a follows: Alternative SPE Method: The samples that ware prepared In 1% acetic acid for the direct injection method were used for this 'extraction. Five milliliters of aach sample was eliquoted into a 50-mL polypropylene centrifuge tube and the volume was taken to 40 mL with water. The samples were then centrifuged for -10 minutes at -3000 rpm. The supernatant was then loaded onto a C,f SPE cartridge conditioned with 10 mL of methanol and 5 mL of water. The eluate was discarded. Approximately five mUlilters of methanol was added to the cartridge. Five mHllllters of eluate was collected into a graduated 15 mL polypropylene centrifuge tube. Each sample was analyzed by LC/MS/MS electrospray. Cause: ___ Preparation___ Analysis ____ Instrument____ Client Request X Other More usable date was obtained. LIBRARY ID: V 5001040-6 hxygen Research ADMINISTRATIVE FORM Rage lUy o 11U Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0000760 CHEH EHSft 2 3 6 1B 651 733 1958 Q M i-to o e s t a in F r w - iifT c n s c u rriO B 0 5 /0 1 ' 0 6 1 3 :4 6 NO .106 0 3 /0 3 + m m p . r n / n : F - ll STa CK * > KJ58 NCTMrchDrfvt Statt CoUcg, PAtWH . P h o n rlt+ lT i-i? FM 1 + tti-ll* . b a p tfc rm e im Q H r tk > ' * 4f '! ' J L IwiNaptdSeOMWon ' PtflBfrPiMbn OtfcWOurmrtcB: !OWMI 1 O h n tn n fto tl: "523255531" M M o n lH U ttf. . tu ' ' W J d jfB w o r UHMNY ID: V9C9M404 >s M t on ' 7 * / * Tm V - to r & ` D rta ttm * n t . r, \v * /, ' * . * O , *>^ * N * ` ; . >. .. * * . ' ' * '. ~ -i . >*, \ - O - i s > .V ' \ ' . ' /v._3>vr\ *V:;'v<5./1. ~> r -.. . \' * ' - s. j1 : >.s'>*'* Vj . . >.w . 1 .. . S - V ' A - ! - : . ! * '.` , , . . . . .# V `.k ' i s. ^ ( . ::Ci />*'' /'* / . . ` *V * r ` ' " " ' . .f O - 1. .* Exygen Research Page 110 of 110 INTERIM REPORT # 23 - Analysis of Sediment Samples STUDY TITLE Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program DATA REQUIREMENTS EPA TSCA Good Laboratory Practice Standards 40 CFR 792 STUDY DIRECTOR Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 Phone: 610-701-3761 INTERIM REPORT COMPLETION DATE May 2,2006 PERFORMING LABORATORY Exygen Research 3058 Research Drive State College, PA 16801 Phone: 814-272-1039 STUDY SPONSOR 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 Phone: 651-733-6374 PROJECT Protocol Number: P0001131 Exygen Study Number: P0001131 Total Pages: 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: PCQ01131 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT Exygen Study Number P0001131, entitled "Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sedimept, Fish, Clams, Vegetation, Small -Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring, program," conducted for 3M Company, is being performed in, compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792 by Exygen Research, with one exception. The laboratory control and control spikes for the direct injection method were not recorded promptly at the time of extraction. . ... Weston Solutions, Inc. Michael Ai. Santoro Sponsor Representative 3M Company T Exygen Research Date Date Page 2 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: POOOl 131 QUALITY ASSURANCE STATEMENT Exygen Research's Quality Assurance Unit reviewed Exygen Study Number POOOl 131, entitled, "Analysis of Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfhiorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program". All reviewed phases1 were inspected for conduct according to Exygen Research's Standard Operating Procedures, the Study Protocol, and all applicable Good Laboratory Practice Standards. All findings were reported to the Exygen Principal Investigator and Management and to the Study Director. Phase Date Inspected Date Reported to Date Repeated to Principal Exygen Date Reported to Investigator Management Study Director 25. Final Raw Data Review and Interim Analytical Report Review 05/02/06 05/02/06 05/02/06 05/02/06 Date lNote: All in-lab inspections will be documented in the QA statement for the final analytical report at the conclusion of the study. This QA statement involves only the review of the interim report and associated raw data. Exygen Research Page 3 of 110 . Interim Report #23 - Analysis of Sediment Sample? Exygen Study No. : P0001131 CERTIFICATION OF AUTHENTICITY This interim report, for Exygen Study Number P0001131, is a true and complete representation of the raw data. Submitted by: Exygen Research 3058 Research Drive State College, PA 16801 (814) 272-1039 Principal Investigator, Exygen: Date Exygen Research Facility Management- President Exygen R< Study Director, W eston Solutions, Inc. k Jaisimha Ke& P ^ ., DEE Weston Solutions, Inc. ' C t. Sponsor Representative, 3M Company: Michael A1 Santoro Director of Regulatory Affairs | Exygen Research Date % 06 Date Page 4 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 STUDY IDENTIFICATION Analysis o f Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) in Water, Soil, Sediment, Fish, Clams, Vegetation, Small Mammal Liver and Small Mammal Serum Using LC/MS/MS for the 3M Decatur Monitoring Program > PROTOCOL NUMBER: P0001131 -- EXYGEN STUDY NUMBER: P0001131 TYPE OF STUDY: Residue SAMPLE MATRIX: Sediment - -- , ' -- TEST SUBSTANCE: Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) SPONSOR: 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 STUDY DIRECTOR- Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 STUDY MONITOR Michael A. Santoro 3M Company 3M Building 0236-01-B-10 St. Paul, MN 55144 PERFORMING LABORATORY: Exygen Research 3058 Research Drive State College, PA 16801 ANALYTICAL PHASE TIMETABLE: Study Initiation Date: 11/05/04 Interim Analytical Start Date: 04727/06 Interim Analytical Termination Date: 04/30/06 Interim Report Completion Date: 05/02/06 Exygen Research Page 5 of 110 Interim Report #23 - Analysis of Sediment Samples. Exygen Study No.: P0001131 PROJECT PERSONNEL The Study Director for this project is Jaisimha Kesari at Weston Solutions, Inc. The following personnel from Exygen Research were associated with various phases o f this interim portion of the study. Name John Flaherty Karen Risha Paul Connolly Christine Edwards Mark Ammerman Amy Sheehan Brittany Kravets . , .Scott Crain v Frances Crespi Devon Ingram Christa Gallant Title.. Vice President Scientist Technical Lead-LC/MS Technician r Sample Custodian Associate Scientist Technician Technician Technician Technician >, Technician Exygen Research Page 6 of 110 Interim Report #23 - Analysis of Sediment Samples Exygeri Study No. : P0001131 TABLE OF CONTENTS Page TITLE PAGE......................................................................................................................... 1 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT........ ..................... 2 QUALITY ASSURANCE STATEMENT..... . ..................................... ;...................... 3 CERTIFICATION OF AUTHENTICITY.......................................................... .................4 STUDY IDENTIFICATION..................................................................................................5 PROJECT PERSONNEL.......................................................................................................6 TABLE OF CONTENTS............................................................ 7 LIST OF TABLES................................................................................................................. 8 LIST OF FIGURES..................................................... 9 LIST OF APPENDICES......................................................................................................10 1.0 SUMMARY............ ............................................. 11 2.0 OBJECTIVE................................................................................................................. 11 3.0 INTRODUCTION......... ............................................. 11 4.0 ANALYTICAL TEST SAMPLES............................................................................ 12 5.0 REFERENCE MATERIAL.................................................... 12 6.0 DESCRIPTION OF ANALYTICAL METHOD......................................................... 13 6.1. Extraction Procedure For Sediment.................. 13 6.2 Direct Inject Extraction Procedure For Sediment...................................................... 14 6.3 Percent Solids Procedure For Sediment............. ......................................................14 6.4 Preparation of Standards and Fortification Solutions............................................... 14 6.5 Chromatography.... ........................................ 15 6.6 Instrument Sensitivity.................................................................................................15 6.7 Description of LC/MS/MS Instrument and Operating Conditions............................15 6.8 Quantitation and Example Calculations.................................................................... 16 7.0 EXPERIMENTAL DESIGN............................. 20 8.0 RESULTS.....................................................................................................................20 9.0 CONCLUSIONS^............................................ 1...... :................................................ 20 10.0 RETENTION OF DATA AND SAMPLES.............................................................. 21 Exygen Research Page 7 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Table I. LIST OF TABLES Page Summary of PFBS, PFHS and PFOS in Sediment Samples......... ..................2 3 Table II. Summary of PFOS in Direct Injection Sediment Samples............................2 4 Table m . Matrix Spike Recovery of PFBS, PFHS and PFOS in Sediment Samples....25 Table IV. Matrix Spike Recovery of PFOS in Direct Injection Sediment Samples......26 Table V, Total Percent Solids in Sediment Samples.....................................................27 Exygen Research Page 8 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study N o.: P0001131 Figure 1. LIST OF FIGURES Page Typical Calibration Ctlrve for PFBS in Methanol........... .............................. 29 Figure 2. Non-Extracted Standards of PFBS iri Methanol, 0.2 ng/mL and 0.5 ng/mL, Respectively ............... ...................................................... 30 Figure 3. PFBS in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively............. ........................... 31 Figure 4. Chromatogram Representing a Sediment Sample Analyzed for PFBS (Exygen ID: C0172892, Data Set: 042606F).................................................. 32 Figure 5. Typical Calibration Curve for PFHS in Methanol.......................................... 33 Figure 6. Non-Extracted Standards of PFHS in Methanol, 0.2 ng/mL and 0.5 ng/mL, Respectively.................................................................................. 34 Figure 7. PFHS in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively......................................... 35 Figure 8. Chromatogram Representing a Sediment Sample Analyzed for PFHS (Exygen ID: C0172892, Data Set: 042606F)...................................................36 Figure 9. Typical Calibration Curve for PFOS in Methanol.......................................... 37 Figure 10. Non-Extracted Standards of PFOS in Methanol, 0.2 ng/mL and 0.5 ng/mL, Respectively................................................................................. 38 Figure 11. PFOS in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively......................................... 39 Figure 12. Chromatogram Representing a Sediment Sample Analyzed for PFOS (Exygen ID: CO172892, Data Set:042806B)...................................................40 Exygen Research Page 9 o f 110 interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 LIST OF APPENDICES Appendix A Study Protocol P0001131 (Exygen Study No. P0001131) with Analytical Method V0001782: "Method of Analysis for the Determination of Pcrfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS", and Protocol Amendments.............................. Page ,41 Exygen Research Page 10 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0Q01131 . 1.0 SUMMARY Exygen Research extracted and analyzed sediment samples for die determination of perfluorobutanesulfonate (PFBS), .. pprfluorohexanesulfonate (PFHS), and perfluorooctanesulfonate (PFOS) according to Exygen Method V0001782 (Appendix A). The limit o f quantitation for PFBS and PFHS in sediment was 0.2 ng/g. The limit of quantitation for PFOS in sediment was 2.0 ng/g. ' The standard sediment method (V0001782) was originally used to extract all of the samples. Use o f this method yielded poor recoveries for PFOS. Because of this, a direct injection method (Section 6.2) was used to analyze for PFOS. Analytical results for the analysis o f PFBS, PFHS and PFOS found in sediment samples extracted with the original method are summarized in Table I. Analytical results for the analysis of PFOS found in sediment samples by direct injection are summarized in Table n. The average percent recoveries standard deviations for PFBS, PFHS, PFOS in sediment samples were 85 19%, 99 29%, and 93 6%, respectively. Fortification recoveries for the analysis of PFBS, PFHS, and PFOS in sediment samples extracted with the original method are summarized in Table III. Fortification recoveries for the analysis o f PFOS in sediment samples by direct injection are summarized in Table IV. The total percent solids for sediment samples are detailed in Table V. 2.0 OBJECTIVE The objective of the analytical part o f this study was to determine levels of perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS), and perfluorooctanesulfonate (PFOS) in sediment according to Protocol P0001131 (Appendix A). 3.0 INTRODUCTION This report details the results o f the analysis for the determination o f PFBS, PFHS, and PFOS in sediment using the analytical methods entitled, "V0001782: Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS." The study was initiated on November 05, 2004, when the study director signed protocol number P0001131. The analytical start date for this interim report was April 27, 2006 and the analytical termination date for this interim report was April 30,2006. Exygen Research Page 11 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 4.0 ANALYTICAL TEST SAMPLES Three sediment samples (Exygen ID: C0172892-C0172894) were received on wet ice on April 22, 2006 from Tim Frinak at Weston Solutions, Inc. The samples were logged in by Exygen personnel and placed in refrigerated storage. Sample log-in and chain of custody information is located in the raw data package associated with this interim report. Storage records will be kept at Exygen Research. 5.0 REFERENCE MATERIAL The analytical standards, PFBS and PFHS, were supplied by 3M. PFBS was received from 3M at Exygen on May, 13, 2005. PFHS was received from 3M at Exygen on January, 20, 2003. PFOS was purchased from Fluka Corporation and was received at Exygen on April, 23, 2003. The available information for the reference materials is listed below. PFBS and PFHS were stored frozen and PFOS was stored refrigerated. Compound PFBS PFHS PFOS Exygen Inventory No. SP0005726 SP0002401 SP0002694 Lot# Purity (%) 101 96.7 SE036 ' 98.6 * 430180-1 101.2 Expiration Date 12/04/06 10/18/06 10/31/07 The molecular structures of PFBS, PFHS and PFOS are given below: PFBS Chemical Name: Perfluorobutancsulfonate Molecular Weight: 338 supplied as the potassium salt (CiFgSOsTC*) Transitions Monitored: 299 -+ 99, Structure: " . F FF F F! FF PFHS Chemical Name: Perfluorohexanesulfonate Molecular Weight: 438 supplied as the potassium salt (QFnSOsTC) Transitions Monitored: 399 --80 Exygen Research Page 12 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No. : PCOO1131 PFOS Chemical Name: Perfluorooctanesulfonate : Molecular Weight: 538 supplied as the potassium salt (CgFnSOjTC*) Transitions Monitored: 4 9 9 --80 Structure: . 6.0 DESCRIPTION OF ANALYTICAL METHOD The analytical method "V0001782: Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" was used for the analysis of PFBS and PFHS in sediment samples. A direct injection method, detailed in Section 6.2, was used for the analysis of PFOS in sediment samples. 6.1. Extraction Procedure For Sediment Before the samples were weighed for the extraction, they were mixed thoroughly by vigorously shaking the container. A 5 gram portion o f sediment was weighed into a fifty milliliter centrifuge tube for the extraction. After fortification of appropriate samples, 35 mL of 1% acetic acid in water was added to the samples. The samples were vortexed and allowed to shake on a wrist action shaker for ~60 minutes. The samples were centrifuged for ~20 minutes at ~3000 rpm. The, supernatant was then loaded onto a Cis SPE cartridge conditioned with 10 mL of methanol and 20 mL of water. The eluate was discarded. Twenty milliliters of methanol was added to the sediment samples left in the centrifuge tube. The samples were vortexed and allowed to shake on a wrist action shaker for another 30 minutes. The samples were centrifuged again for ~2Q minutes at -3000 rpm. The supernatant was then loaded onto the same Cf* SPE cartridge. The eluate was collected into a 500 mL Nalgene Bottle. The column was washed with 4 mL of methanol. The wash was collected in the same bottle as the eluate. Approximately two hundred milliliters of water was added to the bottles. The samples were mixed by shaking and loaded onto another Cis SPE cartridge conditioned with 10 mL o f methanol Exygen Research Page 13 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 and 20 mL o f water. The eluate was discarded Approximately five milliliters of methanol was added to the cartridge, Five milliliters of eluate was collected into a graduated 15 mL polypropylene centrifuge tube. Each sample was analyzed by LC/MS/MS electrospray. 6J2 Direct Inject Extraction Procedure For Sediment Before the samples were weighed for the extraction, they were mixed thoroughly by vigorously shaking the container. A one-gram portion o f sediment was weighed into a 15-milliliter centrifuge tube for the extraction. Ten milliliters o f 1% acetic acid in methanol was added to each sample. The samples were then shaken by hand, vortexed, and sonicated for thirty minutes. The samples were then centrifuged for ~10 minutes at -3000 rpm. Each sample was analyzed by LC/MS/MS electrospray. 6.3 Percent Solids Procedure For Sediment Percent solids were determined using the procedure indicated in Exygen method V0000427. Approximately 20 grams o f sample was weighed into a pan. The weight of the sample plus the pan was recorded. The samples were then dried in an oven overnight at 104 2 C. Then the samples were transferred to a dessicator and allowed to cool for ~15 minutes. Each sample was then weighed again, including the weight o f the pan. The percent solid for each sample was then calculated. See Table V for percent solids results. 6.4 Preparation of Standards and Fortification Solutions A mixed stock standard solution of PFBS, PFHS, and PFOS was prepared at a concentration of 1000 pg/mL by dissolving 100 mg of each o f the standards (corrected for purity and salt content) in 100 mL o f methanol. From this solution, the following fortification standards were prepared: Cone, of Fort Stock or Fort Volume Solution (mL) (UK/mL)1 1000 10 100 10 10 10 1.0 10 0.1 10 1of PFBS, PFHS, and PFOS Final ' Volume (mL) 100 100 100 100 100 Einal Cone, of Fortification Std. (pg/mL) 100 10 1.0 0.1 0.01 Exygen Research Page 14 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 A set of non-extracted calibration standards containing PFBS, PFHS, and PFOS was prepared in methanol, as specified in Exygen method V0001782. The following concentrations were prepared: r' "'i '- Cone, of Fort Fort or Calibration Volume Solution (mL) (ng/mL)1 100 1.0 100 0.5 ' ' 100 0.2 10 1.0 5.0 1.0 2.0 1.0 1of PFBS, PFHS, and PFOS Final Volume (1^) 10 10 10 10 10 10 Final Cone, of Calibration Std. (ng/mL) 10 5.0 2.0 1.0 0.5 0.2 The stock standard solution and all fortification and calibration standard solutions were stored in a refrigerator (4 2C) when not in use. Documentation of standard preparation is located in the raw data package associated with this interim report 6.5 Chromatography Quantification of PFBS, PFHS and PFOS was accomplished by LC/MS/MS electrospray. The retention times of PFBS, PFHS and PFOS were ~0.56mins, -9.4 mins, and ~11.4 mins, respectively. Peaks above the LOD were not detected in any of the reagent blank samples corresponding to the analyte retention time. 6.6 Instrument Sensitivity The smallest standard amount injected during the chromatographic run had a concentration of 0.2 ng/mL of PFBS, PFHS and PFOS. 6.7 Description of LC/MS/MS Instrument and Operating Conditions Instrument: API 4000 Biomolecular Mass Analyzer Interface: Turbo Ion Spray Liquid Introduction Interface Computer: DELL OptiPlex QX400 Software; Windows NT, Analyst 1.4.1 HPLC: Hewlett Packard (HP) Series 1100 HP Quat Pump Exygen Research Page 15 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 HP Vacuum Degasser HP Autosampler HP Column Oven HPLC Column: Thermo Fluophase RP, 50 mm x 2 1 mm Column Temp.: ~30*>C Injection V oi: 15 |aL Mobile Phase (A): 2 mM Ammonium Acetate in water Mobile Phase (B): Methanol Time (min) 0.0 TO 8.0 10.0 11.0. , 18.0 Total run time: ~18 min %A 65 65 25 25 65 65 %B 35 35 75 75 35 35 Flow Rate: 0.3 mL/mm v>- . '>: J - i , ' . Ions monitored: Analyte Mode PFBS PFHS PFOS negative negative negative 6.8 Quantitation and Example Calculations Transition Monitored 299 -> 99 399 80 499 -> 80 Approximate Retention Time (min) ~0.56 min. -9.4 min. -11.4 min. Fifteen microliters of sample or calibration standard were injected into the LC/MS/MS. The peak area was measured and the standard curve was generated (using 1/x fit weighted linear regression) by Analyst software using six concentrations of standards. The concentration was determined from the following equations. Equation 1 calculated the amount of analyte found (in ng/mL, based on peak area) using the standard curve (linear regression parameters) generated by the Analyst software program. Please note that the equations in this section are presented in a different order than the equations in the method (V0001782), but this has no effect on the final calculations. Exygen Research Page 16 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Equation 1: Analyte found (ng/mL) = (Peak area - intercept) x DF Slope Where: DF = Dilution Factor, factor by which the final volume was diluted, if necessary. For samples fortified with known amounts of PFBS and PFHS prior to extraction, Equation 2 was used to convert thfe amount o f PFBS and PFHS found in ng/mL to ng/g (ppb). Equation 2: Analyte found (ppb) = fAnalvte found (ng/mL) x final volume (5 mLl] sample weight (5 g) Equation 3 was used to calculate the percent recovery: Equation 3: -. Recovery (%) = fanalvte found (ppb) - analyte in control (ppb)) xlOO% amount added (ppb) Equation 4 was then used to calculate the amount of PFBS and PFHS found in ppb based on dry weight Equation 4: Analyte found (ppb) dry weight - Analyte found (ppb) x [100% / total solids(%)] An example of a calculation using n actual sample follows (calculation is for PFBS only): Sediment sample Exygen ID: C0172893 Spk F (Set: 042606F), fortified at 4 ng/mL with PFBS where: peak area intercept : ^" . p 253819 , . = -592 slope ; = ; , 42300 . dilution factor 5`?-,,K.*.-:, 1 ng/g PFBS added (foil level) 4 amt in corresponding sample (ng/g) :=r 2.41 final volume (mL) ;. - 5 sample weight (g) - 5,;, total solids (%) * 54.37 Exygen Research Page 17 of 110 Interini Report #23 - Analysis of Sediment Samples Exygen Study No.: POOO1131 From equation 1: Analyte found (ng/mL) = f2 5 3 8 1 9 -(-592)1 x 1 42300 * 6.01 ng/ttiL From equation 2: Analyte found (ppb) = (6.01 ng/mL x 5 mL) 5g = 6.01 ppb From equation 3: % Recovery . . = (6.01 p p b - 2.41 ppb) x 100% 4 ppb - 90% From equation 4: ' Analyte found (ppb) dry weight = 6.01 ppb x (100% / 54.37%) = 11.1 ppb NOTE: This value may be slightly different than that of the raw data due to rounding. A slightly different set of calculations was used for the direct injection samples analyzed for PFOS. . Equation 1: Analyte found (ng/mL) = (Peak area - intercept! x DF Slope Where: DF *=Dilution Factor, factor by which the final volume was diluted, if necessary. For samples fortified with known amounts of PFOS prior to extraction, Equation 2 was used to convert the amount of PFOS found in ng/mL to ng/g (ppb). Eguation 2; Analyte found (ppb) * [Analyte found (ng/mL) x volume extracted (10 mL)1 sample weight (1 g) Equation 3 was used to calculate the percent recovery. Exygen Research Page 18 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen -Study No.: P0001131 Equation 3: Recovery (%) = , (analyte found (ng/g) - analvte in control (ng/el) xlOO% amount added (ng/g) Equation 4 was then used to calculate die amount of PFOS found in ppb based on dry weight. Equation 4: . .. r Analyte found (ppb) dry weight = Analyte found (ppb) x [100% / total solids(%)] An example of a calculation using an actual sample follows: Sediment sample Exygen ID: C0172892 Spk D (Set: 042806B), fortified at 400 ng/mL with PFOS where: . - ' peak area = 152371 intercept slope dilution factor = 3720 = 22700 - 100 ng/g PFOS added (fort level) ana in corresponding sample {ng/g) volume extracted (mL) = = :. = 4000 2360 10 sample weight (g) =1 total solids (%) a* 66.54 From equation 1: Analyte found (ng/L) = H52371 - 37201 x 100 22700 = 654 ng/mL From equation 2: Analyte found (ppb) = (654 ne/mL x 10 mL) From equation 3: % Recovery ' - 6540 ppb ' ` = (6540 n e /g - 2360 ng/g) x 100% 4000 ng/g * 105% From equation 4: Analyte found (ppb) dry weight = 6540 ppb x (100% / 66.54%) = 9830 ppb Exygen Research Page 19 of 110 Interini Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 NOTE: This value may be slightly different than that of the raw data due to rounding. 7.0 EXPERIMENTAL DESIGN For sediment samples designated as laboratory matrix spikes, PFBS, PFHS, and PFOS were added at a known concentration to the samples in the laboratory after the samples were weighed for extraction. The sediment samples were extracted in one set. The set included one reagent blank and two reagent blanks fortified at known concentrations. The set contained three sediment samples. For each sample, two laboratory matrix spikes and a duplicate were extracted. Due to poor recoveries, the three sediment samples were extracted and analyzed using a direct injection method for PFOS. This method is detailed in Section 6.2. 8.0 RESULTS Analytical results for the analysis of PFBS, PFHS and PFOS found in sediment samples extracted with the original method are summarized in Table L Analytical results for the analysis of PFOS found in sediment samples by direct injection are summarized in Table IL The average percent recoveries standard deviations for PFBS, PFHS, PFOS in sediment samples were 85 19%, 99 29%, and 93 6%, respectively. Fortification recoveries for the analysis of PFBS, PFHS, and PFOS in sediment samples extracted with the original method are summarized in Table III. Fortification recoveries for the analysis of PFOS in sediment samples by direct injection are summarized in Table IV. The total percent solids for sediment samples are detailed in Table V. 9.0 CONCLUSIONS The sediment samples were successfully extracted and analyzed for PFBS and PFHS according to analytical method V0001782. The sediment samples were successfully extracted and analyzed for PFOS using an alternative method, which yielded acceptable QC data. Exygen Research Page 20 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 10.0 RETENTION OF DATA AND SAMPLES When the final analytical report is complete, all original paper data generated by Exygen Research will be shipped to the study director. This does not include facility-specific raw data such as instrument or temperature logs. Exact copies of all raw data, as well as a signed copy o f the final analytical report and all original facility-specific raw data, will be retained in the Exygen Research archives for the period o f time specified in EPA TSCA Good Laboratory Practice Standards 40 CFR 792. Exygen Research Page 21 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 TABLES Exygen Research Page 22 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Table L Summary of PFBS, PFHS and PFOS in Sediment Samples Exypen ID C0172892 COI 72892 Rap C 0172883 CO1 7 2 *8 3 Rap CO I 72894 C 0172894 Rep C 4 S u lfo n a te P F B S C * S u lfo n a te P FH S CS S u lfo n a te P FO S dent S am ple ID D A L-S D -B P P 01-0 -0 0 0 0 DAL-SD-BPP01-O-OOOO* D A L-S D -B P P 02-0 -0 0 0 0 D A L -S D -8P P 02-0-0000 * D A L-S O -B P P 03-0-0000 C A L-SD -B PPO 3 0 -0 0 0 0 ' Analyte Found ipp*>. *o7a) Dry W eigh! Aweeaed Accuracy < '-* ) Analyta Found (P pb.ntfg) Dry Weight 3 .1 4 4 0 8 4 .3 3 1 0 4 0 86.1 4 .4 3 30 7 4 .5 4 .8 4 30 6 4 .7 9 .8 7 8 .7 3 30 30 139 123 Aaeaeaed Accuracy W -*> 30 30 30 30 50 50 Analyta Found (P P h .nyg ) Dry Weight A im u d Accuracy ( /-* ) NR NR NR NR NR NR NR NR NR NR NR NR U M x n to ry OupNcat* NR = Not reported due tc quaitty control rmuK lailunsc. See T ti II far re analy tic rcau*- Exygen Research Page 23 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Table IL Summary o f PFOS in Direct Injection Sediment Samples Exygen ID C0172892 C0172892 Rep C0172893 C0172893 Rep C0172894 C0172894 Rep Orient ^ S am p le !!} DAL-SD-BPP01 -0-0000 DAL-SD-BPP0* -0-0000* DAL-SD-BPP02-0-0000 D A L-S D -B P P 02-0-0000* DAL-SD-BPP03-0-0000 DAL-S D -B P P 03-0-0000* C8 Sulfonate PFOS - AerfluorooctanMulfonat Analyte Found (ppb, ng/g) Dry Weight Assessed Accuracy (+/-% ) 3550 3520 4160 3840 11900 12000 30 30 30 30 30 30 ^Laboratory Dupficate Exygen Research Page 24 of 110 Interina Report #23 - Analysis of Sediment Samples Exygen Study No.: PQOOl 131 Table III. Matrix Spike Recovery of PFBS, PFHS and PFOS in Sediment Samples Sample oeacriD&oo C 4 S u d b tiite ETBS W atW e lflftt in c u o i Ami Found -A m ount SpHwd In Semple Recovered Recovery ( " ) (n o )-- w t (O titi) -- ! <%> SuW w fr PFH8 W et W eight C IM c m PFQS W et W eight AmCFound in to n e * . Amount Amt Found Recovered Recovery In Sample Amount Recovered Recovery <%> f u r t i m . (%> DAL-SCAPPO 1-4X0 (Ctmaas sta c.4 -- ape) QAL-SD-8PP01-0-0000 jC W IM liR f tt ilit a tip-- 1 QAL-SD-8PP01-0-0000 c a ra ra t i e.a m -- s d M 4 400 00 2.09 . . 4 47 2.09 202 2.00 2070 06 ? - ? . 561 56.1b 464 , 5100 SO 12 , ,, NR .,, NR NR DAL-SD-6PP02-0-0000 |cai7aaas9pkP,4ppa9pa^ DAl-SXPPQZ-OOOOO (caniiaa ta k a, 4 -- a-- QAL-SO-BPPG2-0-0000 (OtitataM ita N,-- 3 d ta i 4 400 4000 2.41 2 41 .2.41 0.02 374 3020 00 > oaf ' 96 - - - . 40.5 - 346 40.3 5000 . 70 ' _ 126 NR _ 1 NR NR OAL-SD-BPP03-0-0000 *CW7taa* t a U t a ***** DAL-SO-PP03-0-0000 lOaiTaaa apa a 4ae-- s d M DAL-SD-3PP03-0-0000 (Cairaaa a t* k. m -- i f -- 4 400 4000 430 4.30 4.30 7.71 390 4640 83 , 8 - 81.8 121 6 1 J 203 4600 . 65 m _ . . NR NR . _ NR 0 Standard Deviation: Iti til Standard Devtetfcm: 2 9 San-- -- aedi tteepMnf level Jgnifeendy (2* spiking lavati; tieretai. an accurate recoveryvtau# cannot be edcuiated ND * Not detadadattyataiw iw Urn* DiQuanltaaon<LO<^ o<02 no^twelwtagM) NR Not reported due id oueMycof result tamm . 3oa Table IV for re-andywnatta. Nota: tanca Ihta aunmiary table atiowe rounded lesulta. n w n y v laaa nyvory aBgltdy frcm tbo votaoe tata m u data. A v a ru a : NR Stand-- Dotation: HR Exygen Research Page 25 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Table IV. Matrix Spike Recovery of PFOS in Direct Injection Sediment Samples Samp DMCription DAL-SO-BPP01-0-0000 (C9172t92 Bpk (X4M0 ppfeSpite) DAL-SD-BPPO1-0-0000 icemaea a* e. aeoeo pph Spew) DAL-SD-BPF02-04000 (CST72tM tffc G, teOOppbSpfct) DAL-SD-BPP02-0-0000 fc e in tn sph h. aeooo ppb Spa) DAL-SO-BPP03-00000 (C0172tMpk J, MOOpppSpIks) DAL-SOBPP03-0-0000 01726*4 S (* K. 400*0 p i* 5p*) Amount Spaced (PPb) C8 Sulfa rude PFOS Wet W eight Amt Found Amount in Sample Recovered (rrg/fl) wet w t (ng/g) wet w t Recovery (%> 4000 2360 40000 4000 t,, rsr 2360 -. 2260 40000 2260 6540 40000 .4 5950 38000 105 94 92 89 4000 40000 6280 5280 8770 41200 87 90 Average: Standard Deviation: 93 6 'Sam ple residue exceeds the epking level aignrfcsrtly (3x spicing level); therefore, sn accurate recovery value cannol be aatculaled NO - Nol delected 1 or above the lim it of Quantitation (LOQ) o f 0.2 ng^j (wet neigM ). Note; Since thte summery table shows rounded results, recovery values mayvary eHgtiSy from the values In the raw data. Exygen Research Page 26 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Table V. Total Percent Solids in Sediment Samples Exygen ID C 0172892 CO172893 C0172894 S am ple D es crip tio n DA L -5 D - BP P 0 1-G-QOO DAL-SD-BPP02-0-0000 DAL-SD-BPP03-0-0000 Total Solide (%) 66.54 " 54.37 4 4 .4 6 {, Exygen Research Page 27 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 FIGURES Exygen Research Page 28 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Figure 1. Typical Calibration Curve for PFBS in Methanol O42S08F S *ilin*r* lib (PFBS? UMJ*1SHWiltifl ("1 / X- y 4.23*004 X - -OBZ ( r - 0.0000) Area, counts Exygen Research Page 29 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Figure 2, Non-Extraeted Standards of PFBS in Methanol, 0.2 ng/mL and 0.5 ng/raL, Respectively -P F B S < S U ln t* ri} 7 t* .* m .* firm - n m f t t f 1 ra m A r H i t n t M N t i Mtf'fM? I H M l R v t R T ; $.154m i* * 4 }U t* ,w m Inisnaiy. cps A m : 20091 M W k H *)Jht 1 . * 0 0 0 q * RT: 0.990 rail Intensity, cps Exygen Research Page 30 of 110 .Interim Report #23 - Analysis o f Sediment Samples 1 Exygen: Study No. P 0001131 Figure 3. PFBS in Reagent Blank, 0,4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively C e n tro i-P F B S f M m m ) p f t Mat fc r n d } 9 v f 43tr6m MMMFdwTT '., i >- v ',. 13.S3 Intensity, cps A /.jr 19076 s*ant> H a l,tit 1.00 +003 opf RT: 0-667 min 0.60 intensity, cpe A m : 168227 K W b HaigM: 1 .1 1 O M f t RT: 0.667 in 066 Intensity, cps Exygen Research Page 31 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Figure 4. Chromatogram Representing a Sediment Sample Analyzed for PFBS (Exygen ID: C0172892, Data Set: 042606F) I C 41T2*92-PfVS(U t*< nm ) M H I I m i -ir m p lt J o f 43 from H H H F m M An a : r t O M f i e HMgtC AT: tS ftm m 0,87 2900- 2000 WOC 2200- 2000- T lm . m in Intensity, cps Exygen Research Page 32 of 110 Interim Report #23 - Analysis ofSediment Samples Exygen Study No.: P0001131 Figure 5. Typical Calibration Curve for PFHS in Methanol 042900F d iau n tid b (PFHS^ "U ntil* R t f w p i h w dfM iii): y - 706^ +004+ -1.1*008(r>0.9M 6) Ares, count Exygen Research Page 33 of HO Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Figure 6. Nan-Extracted Standards cf PFHS in Methanol, 0.2 ng/mL and 0.5 ng/mL, Respectively I S S ttttS M PFMS V ( r 77774 lom* * 30* tt $4M rJSi4<W 2^r RTi W m i * wW 0.40 Intensity, epa Ama: 33083 e tn a * Haight 1 73*+003 tp* RT: 9.40 min 0.40 intensity, epe Exygen Research Page 34 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: PQOO1131 Figure 7. PFHS in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively t o f u CwKn/ P f m p tr J t m otto*w 4) M m fiJ t i a m h t m tC 2 S * V .w iH ; to 1800Ao 1000 * R 900& .. c j - i................ 01 2 3 4 6 9 7 0 0 10 11 12 13 14 16 10 17 Time, nW* I Raogant Sp*< B P f MS (tX ) 3 0 0 O 0 D im u sanipl* 10 af 45 from 042008F.-4# A ft* 278070 eoonti Hai^htt 1.80*4004 epa RT: Q.40 min 0.40 Tima, min Exygen Research Page 35 o f 110 Interim Report #23 - Analysis o f Sediment Samples * Exygen Study N o.: POGOl 131 Figure 8. Chromatogram Representing a Sediment Sample Analyzed for PFHS (Exygen ID: C0172892, Data Set: 042606F) e r r r a m PFHS g H ta o a m ) 3 M .M M .P m - ta m p f H c f a s / n m M H M F M I T A n a : 3 * 5 3 9 w a i t H H * * b 2 .2 3 a M 3 ( p a X T ; mm Intensity, cps Exygen Research Page 36 of HO Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P001131 Figure 9. Typical Calibration Curve for PFOS in Methanoi 0428008 S td . Dl 14b (PFOS): H.g'*K>n p i tyr iMigMIfte): y 2 ,2 7 # t0 0 4 x + 3T2*+ 003 (r 0 0046) Area, counts Exygen Research Page 37 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Figure 10, Non-Extrscter? Standards of PFOS in Methanol, 0.2 ng/mL and 0.5 ng/mL, Respectively I S S H U S X P FO S ( S t r a t a * ) *SL*ML t *m **m ot* 1 o f 45 fro m M 2 M & ip flr A m : 19759 co* at* HajAt: l i f e ^4*2 cp RT: 11.4m i* 11.43 Intensity, cps Intensity, cps SSOD14696 PFOS (Standard)400 0/00.0 inw ample 2 45 Wm 042900 Ai m : 12226 oovnfc H i|h t 7.72+0C2 cp t RT; 11.4 min 700000$00- 11.43 Tima. min Exygen Research Page 38 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Figure 11. PFOS in Reagent Blank, 0.4 ng/mL Fortified Reagent Spk A, and 4.0 ng/mL Fortified Reagent Spk B, Respectively A M fM f Ctrtrrt-PFOS 4MUMM n r tampM*ot4$tbm *4Zm8.wtff Art*: S3*ce*w<* HUgM: im+091cp* STT: 11.5mi* 11. R axgaatS ffc A - P F O S (00)400.0490.0 m d s sam ple 0 o< 46 from 04 2 8 0 0 6 * 4 * Aroa: 10000 oonatr H aight 0.2 1 1+002 p B t: 11.4 mi -- * t*4 6 000 400> fc a 200- 0 1 2*4507*0 I1 0 4 ^ J iL 10 11 Tima, mi* Roeg%t 9pfc B * PFOS (O Q 4GOXU0O0 am - *am ple 10 ef 46 from 042 908B .w i* Aran: 03701 o m 4 H aight R.07*+003 <pt R T :1 1 4 m i* Exygen Research Page 39 of HO Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Figure 12. Chromatogram Representing a Sediment Sample Analyzed for PFOS (Exygen ID: C0172892, Data Set:042806B) C t m m - pros (Uataawa) I M I I i h - t a m p h H a t 4 5 tram O M C & w c ff Araa: SWT caw rtt H U g k tit J^M 2qpr XT: t t S a / i a 11. Exygen Research Page 40 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P000113 1 APPENDIX A Study Protocol P0001131 (Exygen Study No. P0001131) with Analytical Methods and Protocol Amendments Exygen Research Page 41 o f 110 Interim Report #23 - Analysis of Sediment Samples -Exygen Study No.: P0001131 Exygm Protocol Number: P000113) STUDY PROTOCOL Study TiD e: A nalysis o f P erfluorobutanesulfonate (P F B S), P erfluorohexanesulfonate (P F H S), and P erfluorooctanesalfonate (PFO S) in W ater, S oil, Sedim ent, F ish , C huns, V egetation , Sm all M am m al L iver and Sm all M am m al Serum U sing L C /M S/M S for th e 3M D ecatu r M onitoring Program E xygen P rotocol N um ber: PO001131 Pe r fo r o Exygen Research 3058 Research Drive State College, PA 16801 Phone:(814)272-1039 Sponsor Representative: Michael A. Santoro Director of Regulatory Affairs 3M Building 0236-01-B-10 S t Paul, MN 55144 Phone:(651)733-6374 ftljp. / o j OS Exygen Research Page 42 of 110 Intrim Report #23 - Analysis o f Sediment Samples * J - - Exygen Study No.? P001131 Eiyieo Protocol Number: PSOOI 13) DISTRIBUTION: 1) Jaiiimha Kesah, StudyDirector, Waaton SoJuto* 2) JohnM. Flaherty, PrincipalInvwtiittw, ErygeoResearch 3) Michael A. Santoro, Sponsor Reprcacntadve, 3M Company 4) Exyccn Research Quality Assurance Unii Exygen Research P a g r i o f 65 Page 43 of 110 Interim Report #23 - Analysis of Sediment Samples Exygcn Study No.: P0001131 Exygeo Protoeoi Numb P000113I PROTOCOL APPROVAL Study TMw Analysis ofPerQuonAutaoeculibiuUe (PFBS), Perfluorohexatiesuifonate (PFHS). and PerflucrooctMiesulfcnate (PFOS) in Water, Soil, Sediment, Fish, Clan, Vegetation, Small Mammal Liven and Small Mammal Serum Using LC/MS/MS for the Decatur MonitoringProgram Exygcn Protocol Number: P0001131 APPROV. iaiamha ^ L t o r Weston Solutions Exygen Research & - c tr - w y - Date Pape 3 dS Page 44 o f l 10 Interim Report #23 - Analysis o f Sediment Satriples Exygen Study No.: P0001131 Bxygcn Protocol Number. P000U31 TABLE OF CONTENTS DISTRIBUTION________________ 1___________________ ____ _____ ____ ____ PROTOCOL APPROVAL-,.-,. _-- .......... ........- I ----------- ---------------------------------- TABLE O f CONTENTS-------------------------- . -- .................... ........ - ---------- ----------- INTRODUCTION_______ ______________ __ __________ __________________ TEST MATERIALS-------------------------------------------------------------------------------------- OBJECTIVE------------------- ....___ ___ ____ _ ____ __ -- --_______ ---- ........ TB8TINO FACULTY--------------------------- -- -------------------- ------------------------------- STUDYDIRECTOR.__ _________________ _____ _____ ________ ___________ SPONSOR REPRESENTATIVE---------------------- -------------- . . ----- ----------------------------- PRINCIPAL INVESTIGATOR----------------------------------------- _______ _______ PROPOSED EXPERIMENTAL START AND TERMINATION DATES________ ........... ... IDENTIFIGVnON AND JUSTIFICATIONOFTHE TBSTSYSTEM_____________ ____ SAMPLE PROCUREMENT. RECEIPT AND RETENTION----------------------------------- ----- SAMPLE DDENnFJCATXW_______________________ _________ _________ ___..... ..... . . 9 ANALYTICAL PROCEDURE SUMMARY-------------------- ------------------ ------------- ------- VERIFICATION O f ANALYTICAL PROCEDURE_________ 1___________________ ______ 9 METHOD TORCONTROL OF BIAS________________________________________ STATISTICAL METHODS------------------------------------------- ---------------------------------- QLP STATEMENT_________________________________ :________ ______ ...__________ I I ________ I I SAFETY a n d HEALTH-------------------------------------------------------------------- ------------ ________12 AMENDMENTS TO PROTOCOL___________ ___ __________________________ ________ 13 DATA RECORD KEEPING--------------------------------------------------- ----------- -------------- ...............1 3 QUALITY ASSURANCE______ ________________ ___________________ __ _ ...............1 RETENTIONOPDATA AND ARCHIVING ..L .TM .TM .,...................... ............ ________ 14 APPENDIX L ANALYTICAL METHODS------- --- --------------------..................______ . . . ________ IS Page 4 o f63 Exygen Research Page 45 of 110 Interim Report #23 - Analysis o f Sediment Samples .. Exygen Study No.: P0001131 Exyfen Protocol N urtw r: POOA1131 INTRODUCTION The pmpoao of this study is to perform (tuiJysii for perflucrobutanesulfonate (PFBS), periluorohcxanesulfonate (PFHS) and perfluorooctancsulfonate (PFOS) in water, soil, sediment, fish, clams, vegetation, small mammal liven and small mammal serum using LC/MSAfS for the 3M Deratin' Monitoring Program. The study will be indited for compliance with EPA TSCA Good Laboratory Practice Staadsrds 40 CFR 792 by the Quality Assurance Unit of Exygen TK>f-iB, CaiCtk. TEST MATERIALS The lest materials are perfluorobutaneaulfonate (PFBS). pcrfluoiahcxmcaulfonate (PFHS) and pecihorooctsneeulfonste (PFOS) and are all sigapliod by 3M. PFBS Chemical Name: Pecfliaorobutancjulfooate Molecular Weight: 338 supplied as the potassium salt (CtFvSOj'K'1') Lot Number 101 Purity: 96.7% Tranritions Monitored: 299-99 Structure: PFHS Chemical Name: Perfiawobewmesuifoaate Molocubff^Weight: 438 supplied as the potassium salt (CiFnSOj'K*) Lot Number $3036 Purity: >8.6% Transitions Monitored: 399 -* 80 Structure: Pa* S o fiS Exygen Research Page 46 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen S tudy Ko.: POOO1131 Exygen Protocol Number: POOOl 131 PPOS Chemical Nunc; Perfluorooctanesulfboale Molecular Weight: 538 supplied as the potassium n it (C1F17SO3K*) Lot N um ber 21T - r Parity: 86.9% ri--- ^ ~ r; Trsantioa* Monitor^: 499 -P 99 Structure: J"x - OBJECTIVE : . it' The puipoec of this tudy is to perform analysis for perfluorobutancmlfoiuto (PFBS), perfhkMohexaneeulfoaate (PFHS) and perfluorooctanegulfonate (PFOS) in water, toil, sediment, fish, ctsma, vegetation, small mammal livers end small mammal serum for foe 3M Decatur Monitoring Program using the ernrent versions ofthe following Exygen analytical methods: V0001780: `Method of Analysis for foe Determination of Perfluorooctanoic Add (PFOA) in Water byLC/MS/MS" V0001781: "Method of Analytie for foe Determination of Perfluorooctanoic Add (PFOA) in Soil by LGM&MS" V0001782: "Method of Analysi* for foe Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LOMS/MS" V0001783: "Method of Analysis for the Determination of Parfiuorooctanoic Acid(PFOA) in Fish md Clameby LOMS/MS" V0001784: "Mathod of Analysis for foe Determination ofPerfluorooctanoic Acid (PFOA) io Vegetation by LC/MS/MS" V0001785: "Method ofAnalysie for foe Detenninalioo ofPerfluorooctanoic Add (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method of Analysia for the Determination of Porflooroocuooic Add (PFOA) in Small M m sid Sesum by LC/MS/MS" TESTING FACILITY Hxygen Research 3038 Research Drive Stale College, PA 16801 Phone:(814)272-1039 Page 6 o f 63 Exygen Research Page 47 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 ProtocolRombar. P0001 111 STUDY DIRECTOR Jaisimh* (Cestri P.E., OEE Weston Solution, Inc. 1400 Weftoc Way WeatChester, PA 19380 Phone: <610) 701-3761 Fax:(610)701-7401 j.keari@westontolutioni.com SPONSOR REPRESENTATIVE Michael A. Sanano 3M Coroptny DirectorofRegulatory Aflairs 3M Building 0236-0! -B-10 St Paul, MN 53144 Pitone: (651) 733-6374 PRINCIPAL INVESTIGATOR John M. Flaherty Exygen Research 3058 Research Drive State College, PA 16801 Phone:(814)272-1039 john.flahefty@erygec.coin PROPOSED EXPERIMENTAL START AND TERMINATION DATES It if proponed that the analytical portion of tins study be conducted from October 01, 2004 te December 31, 2005 The actual experimental nan and termination date will be included m the final report. Page T o fti Exygen Research Page48 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Exyjea Protocol Number: WOO1131 IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM The following are (be test systems for this study: Water (groundwater rod surface water} Soil Sediment ; Hah ) Clam* Vegetation Small Mammal Liver Small Mammal Sent The ample* will be collected by WeetOQSolution*. The control sample* will be pmcbaaod and prepared by tbe testing facility. Purchase and processing detail* for tbe control ample* wiH.be inctnded in foe final report associated With this study. The teat system* were chosen to access the environmental impact of PFBS, PFHS andPPOS in the Decatur, Alabama area. SAMPLE PROCUREMENT, RECEIPT AND RETENTION Water, soil, sediment, fish, dam, vegetation, smaC mammal liver and small mammal serum samples will be received at Exygen directly Born Wests Solutions. The details ofsampleprocurement for this study are outlined in the 3M work plan entitled "Phase 2 Week Plan for Sampling Environmental Media.** The number rod types of sample* collected will vary depending availability in foe field. The total number of sample* received rod analyzed for each matrix will be documented in foe final report associated with this tody. Water, oil, and sediment sample* will be used a* received without further proceaaing at Exygen. These samples wifi be stored refrigerated at 2C-8C. Fish, clam, vcgetttioa and small mammal Ever samples will be processed according to foe appropriate analytical method (tee Appendix I). These sample* will be atond frozen ait S -10*C. Small mammal whole blood sample*will be eentrifliged in foe field at tbe tune ofcollection and the serum fraction will be used for the study. Small mammal serum will be stored frozen at i-tO^C. The receipt and processing of the samples will be documented in tbe final report andraw data associated wifofoe study. P a ieS c/6 5 Exygen Research Page 49 of 110 interim Report #23 - Analysis o f Sediment Samples Exygen Study No. : POOO1131 E*yjeo Protocol Number: P0CC1131 SAMPLE IDENTIFICATION Prior to aruiysii, each sample will be assigned a laboratory sample reference number. The reference cumber ill be unique and will distinguish each laboratory sample that ia processed throughout the analytical procedure. ChrocBstopsphic data will be identified by the laboratory sample reference number. Sample storage concStints and locations win be documented throughout the study. ' ANALYTICAL PROCEDURE SUMMARY References: V0001780: "Method of Analysu for tbe Determination ofPerfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781; "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sofl by LC/MS/MS" V0001782: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" Y0001783: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS" V0001784: "Method of Analysis for the Dcternrinitron of Perfluorooctanoic Add (PFOA) in Vegetation by LC/MS/MS" `V0001785: "Method of Analysis for tbe Determination of Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: '`Method of Analysts for the Determination of Perfluorooctanoic Acid (PFOA) in Small Mammal Serum by LC/MS/MS" The above methods use analytical conditions capable of separating the isomers of PFBS, FFHS and PFOS. The final report will include tbe isomers summed into total PFBS, total PFHS, and total PFOS found. VERIFICATION OF ANALYTICAL PROCEDURE A laboratory contra] sample will be used for the preparation of fortified control samples. Toe teat substance will be made into solutions as per tbe method, and added to die matrices via a nncropipette. For water sampling, Exygen will supply one bottlo per sample collected. The bottles wilt be 500 mL precleaned Sci/Spec Premier wide mouth HOPE bottle. These bottles have been routinely used for fluorochemical sample Pngr 9 o f 65 Exygen Research Page 50 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study N o.: P0001131 P reaacm N u n b e r POOOt U I collection it tbe toting facility and bavebeeo shown to be ires of PFBS. PFHS and PFOS. Samples will be added to cadi container to a volumetric Gil line at 200 mL. A Geld dplicate, a low fieWspike and a high field spike of each sample svill be collected. The low and high field spike bottle* will contain PFBS, PFHS and PFOS aa wellaa -perflucrooctanoic acid (PFOA) and 1.2-13C perftBoroodanoic acid (IJCPFOA). PFOA and ,5C PFOA are included in the solution wed to spike,the sample. The result! for PFOA and C PFOA will not be reported in this study, Exygtn win apply one field blank (control water) and two field Monk spikes (control water fortified with PFBS, PFHS and PFOS at.a low and high level) for every twenty samples collected. At file testing facility, each water sample (excluding field duplicate and field spikes) will be extracted in duplicate and will also be fortified at a low and high concentration with PFBS, PFHS and PFOS and processed through die daacribed procedure to determine method accuracy and to check forbit. For soil, sediment, clams, and vegetation, Bxygeo will supply one $00 mL precleaned Scv'Spcc Premier wide mouth HOPEbottle per sample collected or a zip-seal bag. All contamcn/bag* used for sample collection will be shipped to the sample location. Samples will be added to each container or bag in the field. At the tewing facility, each sample will be extracted in duplicate and will abo be fortified at a known concentration with PFBS, PFHS and PFOS at both a low and high level and processed through tbe described procedure to determine method accuracy and to check for bias. For smafl'manimal liver, Exygen will supply a SO mL polypropylene centrifuge tube. For small mammal serum, Exygen will supply a collection kit for each ample containing serum separator tube* (red top), vacutaioen, needle holders rod needles, trawler pipettes, and polypropylene tubes. At the tertinfi facility, each liver and serum sample will be extracted m duplicate and will also be fortified at a (mows concentration with PFBS, PFHS and PFOS at bods a low rod high level and processed through tbe described procedure to determine method accuracy and tocheck for bias. Low and high spiking levels for each matrix are defined below: Matrix Law Spiking Level Hieb Spiking Level Water SOOog/L 5000 ng/L Soil *ng/g 40 na/a. SedioM Pt *oa/i 40 ng/g Fish lOng/g 100 na/S Clams 10 ng/g 100ng/g Vegetation lOng/g 100 ns/s SmallMammal Liver IQmtte 100ng/g Small MammalSeran 10 ng/mL 100 ng/mL p* lO o fS S Exygen Research Page 51 o f 110 Interim Report #22 - Analysis o f Sediment Samples Kxygen Study No.: P0001131 Exygm hmocol Number: F000I 131 Recoveries we anticipated to be between 70% and 130% of ihe fortified levels; however, the exact precision sod accuracy will be determined by the uniyxia of the quality control samples described above. A statement of accuracy will be included in the final report. METHOD POR CONTROL OF BIAS Control of bias wifi be addressed by taking representative sub-samples from a homogeneous mixture of each matrix from untreated control samples, and by analyzing at lea two levels of fortifications. STATISTICAL METHODS Statistic wifi be limited to those specified in the subject methods and to the catrulatkm ofaverage recoveries, as applicable. GLF STATEMENT All aspects of this study shah be perforated and reported in compliance with EPA TSCA Good Laboratory Practice Standards 40 CFR 792. The final report or data package (supplied to the Sponsor) shall contain a statement that the study was conducted in compliance with current sud applicable GLP standards and arili outline any deviations in the study from those standards. This statement will be signed by the Study Director and Sponsor Representative. RETORT A final repoit will be prepared by fite pdndpsl investigator or their designee at the conctuahm offite study. Thereport will iuclude, but will not bc limited lo, the following: e The Tarne and xVhesa of tim Strdy Diractor, Sponsor RepresenUtive. and ofthe testin* faeihty. A Statement of GLP compliance (any related documentation, och as chain-of-cuatodyrecotda, must be in the studyrecords). Pajir I l o f il Exygen Research Page 52 of 110 Interim Report #23 - Analysis o f Sediment Samples ' Exygen Study No.: P0001131 Exyssa Protocol N toabcr. P0001131 The signed and dated statement by the Exygen Research Quality Assurance Unit regarding dates of study inspections and dates findings wore reported tothe Study Director tad Management. >y'. A description of die exact analytical conditions employed in the study. If the subject method was followed exactly, it is necessary to include only s copy of the aoalyticai method. Any modifications to id method will be incorporated into the report. If the method is photo-reduced, the project number end page number must be included on eachpage. ' Inscription oftlWinstnbweutstk used and operating conditions. All results from all sets analyzed. Control and fortified samples wiD be identified and the data table Will include sample number and fortification level. Representative chromatograms for each analyte in each matrix, inchxlmg chromatograms of a stimdard and a control sample, and a chromatogram at a fortification level. The location of the analyte peaks will be clearly identified in all chromatograms. All circumstances that may have afibetnd the (polity or integrity of foe data win be documented in the report. a . Locations where raw data and the final report are to be archived. Additions or conectiona to the final report shall be in the form of mi ' amendment signed by foe Study Director. The amendment shall clearly identity that part offoe report that is being altered and foe reasons for the alterations. The amendment will be signed and dated by the Study Directorand foe Sponsor Representative. All applicable requirements for reporting of study results as per 40 CFR 792.185. SAFETY AND HEALTH s Laboratorypersonnel willpractice good sanitation and hcakh habits. . , fjL ~ ' . - : . > Bvcry reasonable precaution ball be taken to prevent inadvertent exposure of personnel and tire environment to the test or reference subttsncefa). P a st H o f 65 Exygen Research Page 53 of 110 Interim Report #23 - Analysis of Sediment Samples . . Exygen Study No.: P0001131 Bxygm Protocel Number: POOOU31 AMENDMENTS TO PROTOCOL Alt significant change* in die analytical protocol outlined here will be expressed in writing, rigned and dated by the Study Director and Sponsor Representative. Amendments usually will be issued prior to initiation orstudy plan change. However, when a change it required without sufficient time for the sane of a written amendment, that change may be effected verbally with supporting documentation .signed and dated by the Study Director aad followed with a written izneriffteent at soon as possible. In this case, the effective date of foe written amendment will be the date of the documented change. Copies of the signed amendments will be appended to all distributed study plan copies. The original amendment will be maintained with tbe original study phtru Any deviations from.# study plan or from the analytical method is provided will be documented and reported promptly to the Sponsor Representative. DATA RECORD KEEPING Records to be maintained include the following (as appropriate): * Sample tracking sheets) * Sample receipt records. Stonge bistory, and chains ofcustody * History Midpreparation ofstandards (stock, fortfiewion, calibration) Description ofsoy modifications to the method < Inrtnnr'cmt run sheets, bench-sheets or logs * Analytical data tables All chromatographic and instrumental condition * Sample extraction and analysis dates * A complete listing of studypersonnel, signatures and initials * Chronological presentation of an study correspondence * Any other documentation necessary fbrthe reconstruction of tbe study Chromatograms- All chromatograms will contain the following: Sample identification, injection date, row or other indication of the srea ofinterest, and injection number correspondmg to tbe ran. Additionally, fortifications will include tbe mount of analyte added and foe sample numberoffoe ample that was fortified. Analytical standard chromatograms will additionally include the concentration (e.g.. pg/mL). P ag* 13 o f65 Exygen Research Page 54 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Stdy No.: P0001131 Exygea Proloco) Number P0001131 Ap*t of the documentation rhe Allowing sheets wiMbe included ha each analytical act: a ran sheet Hating the samples to be run in the set, and an instrument conditions sheet describing the instrument type and operaring conditions. QUALITY ASSURANCE The QA Unit of Exygen Research will inspect the study at interval* adequate to assure compliance with GLP'a, and will reportthe findings of audita to the Study Director, Exygen Management, and the Sponsor Representative. RETENTION OF -' D^ -A-T- `A AND ARCH, IVING - 4- '} . . . . - AU hard copy raw dais, inchading, but not limited to, the original chromatograms, worksheets, correspondence, and results shall be included with the data package submitted to the Study Direct'. These will be archived with the original Rudy plan, amendments, final report, and ad pertinent information from (be Sponsor. The testing facility shall keep all electronic raw data and any instrument, equipment, and storage logs far tbs period of time specified in 40 CFR 702.195. Ah exact copy of the materials submitted to the study director will also be kept at Exygen Research. Exygen will obtain permission front the study director before discarding or returning samples. Exygen Research F a te 4 o ft* Page 55 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Btygeo Protocol Nomixr. P0001131 APPENDIX I ANALYTICAL M ETHODS V0001780; "Method of Analysis for the DeterminMion of Perfluoroocttnoic Arad(PFOA) in Waterby LC/MS/MS" V0001781; "Method of Aralvsi* for the Determination of Perilixirooctanaic Arid (PFOA)in Soil byLC/MS/MS" V 0001782; "Method of Analynt fra- the Determination of PerfluoroocUnoic Arid (PFOA) in Sedhnent by LC/MS/MS" V0001783: "Method of Aradyris for the Determination of Perfluorooctmoic Arid (PFOA) in Pish and Claras by LC/MS/MS" V0001784; "Method of Analyst for the Determination of Pcrftuarooctanok Arid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method of Analynt for the Determination of PerRnorocctanoic Arid (PFOA) in Small Mammal U m by LC/MS/MS" V0001786: "Method of Analyait for the Determination of PcrfluorooctaaoM: Arid (PFOA) in Small Mammal Seram by LC/MS/MS" Exygen Research F f t IS o f O Page 56 of 110 Interim Report # 2 1 - Analysis o f Sediment Samples Exygen Study No.: POOO1131 Exyjeo Protocol Nwnber P000ID1 AMNAMLYTMICAL MV0EMT1H7OMD ^ iiB ilir ilH M ii < fh r f i i r i i t tt tite A tM <W OA) h W u tr ... . by LC/M &M S Amlytk] Tating f *ir* Bypw I -- wH >031 RanarA Pnv 9tM i CoSnf*. PA )SW1 AppwdBj: Exygen Research P*2 * 6 c f AS Page 57 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study N o.: P0001131 E x y je a P rotocol N u m b : P 0001131 bihb y^MMtervoidim ANALYTICAL m i t h o p I Mefto d e fA rfrw s far tog Eetom ku too orfyeriharoocucoic Acid (?FOA) in W er by LG W M 3 1.0 Seope im n i i be iwytoyed to ri tootocoqicdgicM itei ii o f ptfluatpocaooie add by Hifb M m m m UpCd Ctowwnftipfry ooepfed * t tandem Mu> I pw iw w L it Da*ttc(LCVK3'MS) ia M r . 2-0 W bty 2.3 Atowyt abu r w td b lebotdofy pramkee. 2 J Go*! toe appwprim* M 8D I beta bwrlUnt u y cAamcaj tor proper tatoiy fn a u fa * . 3.0 SmnfJi Jta^dmacai 3.1 A lto e e t4 ta L ofto* eerapto torttnetien. 3.2 Ke temple piccmriii# needed far water m ptea 3J -- r ~ towed m filfCMKd dacuJd be Mowed mom 1--yerim w . 3A ^ ia a p te * mast be &M0|fely mixed betort bwofwm ptod forexmeuoa. 3.5 A s / tem pi ccwnrintog p ric k s should bo c a w lfu *ri tt '3000 tpm for -3 tts a lM and fee Mpcnuamt wed toethe mxnctim. IJb Semple cdfcotioa paceriurt* wiH be pacified to tie aampttof ptac tor ibu 4.1 W tom -W LC trade 43 MaUaeoi- HPLC pride 4.3 As m u m i Ah u m - A C S . KM toat Qnde 4.4 TedtogiawlaEeto Acid - S i* * A ld rk i 5.0 la c n n m i acd Eqtopnect 5.1 A togb ywrtocounee ttqiiM ftooumroyrli capefck of pmapto| up to 2 aotvwt popped ntt e variable vptane injector capable of injecting 5-200 pL esonected m AtaademMi Spectrometer (L 048A O ). U HHnrrfim n0.0000)|5 .2 A !-- id m fleet *-- * d d i & r M k n t c n it o t end fu m u im w n i t 5 !. an ^ n otntrtitu atoai 5.5 15 m LdlipH M >H n>nn>lM <aaii< ttfn u b m . !JS Di^waMr Wniflf lOOJOOuL). 51.17 tamt^LLcLkmD WUTaVanCwiU-ntkiM. kM aM . *5.V1 D<hIpfeei(Nllfllf(10e0w-1.000 M0^UwtttocwU5. 5J1 W > M n 9 * M tV e ea0 e< C U trB cM U (n . Pa& 7 a f i i Exygen Research Page 58 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E xyg m P rotocol N u m b ; P 0 0 0 U 3 I IH ! I I U a M K a M V tn iW I ... - ... 1 A b A tm e -U . METHOO HKiatlfAMrtymIhihDmrmmlcrtfrwIbionocumoKteklt?FOA)tsWturty ............ UQMSIMS t ; 1U A ' ' . V4_. _ O Crrm a rp taMr lym m q w M QV) a L p o ta n c y ta t m m it 3000 tym -x -6 2 M .c M 45 CMtOMMW T M p a ^ n : y fC M ih n M (A ): 2 aM Aw nflnhm i c a ti* k Water y w o y (g ) : H litM ir1 O n M Piogm u Xm Ub A oj %A 6i 1J *, U 25 *3.0 25 r x 324 *5 P W J tm tm i/m ia l 35 03 35 03 75 03 75 03 >5 03 66 ll|d .(e * W ie * w 6 lo * a * li* $ irU . 6J l a i -13-- I6.7 (fm tta U m : t* k A tm - k m la U M iC fa iO a a r c oytaatu6wOTLCfjoMM.TU* e n * * ienubC |6i64 06y W d u n |* l im u d ir b 7.0 7.1 M c d rOli o r p iir H i p tw M R * n o te <acomrinCH l i M oW b n iM --- ------ - -*----11 1 1 1 6 -- n il HUB la -- r i i pnpm 6 0? irtfa * 0.IJ4 | o< (-- 1 A 6-- n n h fi H -- y l i iK pcitd. - ' - -Vs*' Page l i q f t i Exygen Research Page 59 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E x y | Protocol Number P000113! a H A IT T IC A L m it h c p _________________ ______ o f Aaoya jbr ifal ft o fRcriJuorooctmwifl Acid (PFOA) m Watar by ijcnisjm 1* ' , - S ^ed e d S w V N k d tad ice lo lilk m 11.1 Tmpam t mock aofctka ai-1QQ P |M . Of P?QA by w m *huif IO mg ai aaaytkal M tri (aanaeta tot pm kfi and Alme io 100 mL w * n t m u a t 12M .L D P B tool*. t j i A 10 mfad iW tiihadic otaioo o f PfOA kprqpw eri by bricvgiai IO m L o f tw W M s h * w kE tw le e wd vekinw o f W ih * medw-t M 123m L LDPB basto. M J A 1J3iQfad* btScattm ooktim of^PO A * preparai by b rin d a i 10 mL o f te 10 eohtfiM lo fim i v h tm o i 100 ri* arti** m * 1 B A LLO T E boa*. 11.4 A 0.1 |*im L fc rtifia u k * aoM oo ofPTOA * preparai by b ria |M | 10 iRlUyiM kmL aitoa LO pf'm L d e tta to E A tti volume ai 100 W t* mciaaol io 1 1 3 O H p jta d , fo rtific a ti Botam i d f PFOA is preparai by bnopLnu IO mL v i ma 11 npmL otatta lo fin nXanc o i 100 with a d w l te l L L I boote. 1 1 3 Thonodr M tf ta iiflc a tta Mtatm m e b t ta ra i io e rcfh fe rtto r et ryjwtwimoOity 4*C a d ma tu tto far maximum parta o f 6 moetta Lom Iba ta o fpnepantion 12~ StmidosdC d ftn d M M H iN t 4X L C M M d l c fb m tta andarti ma poponi la HFLC wamt Tbe ta b n dao fi n i v gra n at a tom*& the em etta procedue. IdEodod tornala*. fnptamstaei.1X1 Ib i JaJhmnng k a p ia i taampia- a d d ita ti m ccM rttttM cuy bc Csmmmtta (M W M M tM 0 io 10 ir Mb m .. m na .Fortfierwa V otataci CaaoOfOntonai Yaamt fo ita d C arta CaMntiso aL -- K flE l * * + (ppV 1 40 $ m 40 2J 29$ 40 J0 m 40 100 19$ 40 330 m 40 300 m 40 ... > ..... . CeMWh m i Standard JD X C w tayy-C XOa-- djryl XCmmOn-I XCtamrtyyO XCmmtrH XCfcmddyy^J XCtmm+r^* O tm m M k , dm r a t a a f ta t a t a dario *m cunetta <SRf> XC e a n c t a a f f e t t a t a t a . Paga 19 o f 65 Exygen Research Page 60 o f 110 Interim Report #23 --Analysis o f Sediment Samples Pxygen Study No.: P900I131 E x y fs P ro to c o l N u m b e r, FOQO1131 *******_ j A P tA lY T IC A L METHOD M h > d t^A a ti> e ta ftrll Dstanateatk ofPardaoiPOceaelo; Actd(7TOA)a> Wsiar Sy 9.13 *14 sAatafj 904 sohtoca (reapni btattaj i ut 15-otL polypropylene tubes o f aadarda aay be prefsaitd as 100 * SatUp tea Each baaefc a f mpfos sx&atesd fo pfra iy M as km ) m m include ai laas< m f f tm cawfcil (mathod Manic u m | W LC water; and two s p a t scatiota fartifiad si known cotMntmaona (tab cia trol $ * k ti to venfy pgacadartl iw o m y lo rta ik M ik I0JZ R|aam i tar Seta w d tatan--Irwy t a i p l i n i m and sptitsa w il ba spsofied ta tiw qaairy aw ic e s ptac fenhfe M ar t. 11.9 II.) > t a m < O n l a < w y k g r t p t i w t f i ia f ) B i k in l O n L w id i * kw Inlp <0 L pajypnpylsiia ti-- iik p trim U b rtifr m Beaded. repines M and ndxwad). 1 IJ CoBdUMM the C 8PB e*trid#M (2 |, 4 o rfj by p a o ia f 10 mL methanol fcdowudby J L o fH ? L C w m (--2 4 t***c > . Do bm tai coteaa run dry I I J leadoanpla omw dihrad C{, S F fearfcidp. Diecvd sfaala. 114 Bate wdh -5 mL t09% nathm ol. CoHact 3 mL a f June ana padutted 1 5 ^ p o b ca f)lm aaolRft* tnlaa (1m 1-oliicna 3 14. H i Ana lyta pfaouatt slactaeapsiy LCAdS/M S, I l l C haiw aioy^by tajeel iB * * *a *ie fe a d iita e d a a d , w n p lo rn d fcrtiJfled UBiota ime tke LCA&V8 syatan. A wShn**i *mdm4 m ust prscade id fo ik rw ,u i Ijis d n m d w . 113 SMMtardfWWPOA serrasponding to at taaat five or mom aonesMndan fevsh a m i ha leotadad ta m analytical sat 12J A sarirp. sot s i Attracted caHWatw andarts must be inefaded ihe tu ffa n fa f m i at tbs id o f a sample sat Extracted standard* aw * be 9-10 aarspfea As m abarm rira, w enure sat o f taandarda may ha kyaoed u the begawiac o f s m i tatatand hy sxtm tod o M w standards bwersps n ad w r y *-to *m p *s Op ataaaad far |,asoeed ssa a f srWaateri standard. ta (her esas, xtrscisd rshhnrtna itandisrta ntaha the But i d lota aasettee s w u p k aat 11* Uaa linaar staadasd ansae far fsanthatitei. Linear standard c*rvss w for f a Mudytahy tiasariafraasloa M ia f t/X w d jtK te f o f pcaksies P a p 20 0/ 65 Exygen Research Page 61 o f 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study N o.: P 0001131 Exygea Protocol Number: P0001131 tn n ta N K li i o A cn lytit fie Medndi it a ir veatiTM AN ALYTIC AL M XTHOP | OoiCEdc^ttan ofPnrlucrtCtnnoic Aci (PPOA) in Waler by IC M Sft wlwwiy--v ahrftfen -- rWrl oocartrna a i m a g M atLjt J J (cr aga mlm) U.% Snap* mpoMi Am U net seed nadad le^pooan. Any naspla ttax caoeed ttm itr rtpooet ibonld be Anher Owed m i m vm iynd. >3j0 Anwytaae Criada U.l ClnwMlapM t i w t p n > 4mb*r im a 3 9 m e bwm pertrt o f 413 am . tte d M a m p a a 4 oeU M poJi H M PTOA MfaB. dfc dnagbtar fat (3 0 any) mpreaaia&e lena efcabos dio* idc. 13-2 b ftte d b te ta ^ net a t e * WOA a leve* penar fm da LOQl ir * Hunk motada W OA M Im ta p o te tfew 30 i^ L , iferat i new bhnk wmpte mmk be obtained and ibc andre --t m iat be re-eatfrarted. I3J Kncmam of nente *to t e nutriz p i t a mal be between 70-130% of fta r knew* vate*. Ifnoe ntrol *fd t* U ta antnde tbe aceptable limits, he entile Ml of imples sbauid be le-extracted, Any merit spike outside XK 1 3 M Amid be m n u d by tbs analyst to d a w d a if ite iiM ia n is 1X4 Any c ilftwrton Mudad to te to be * atadcnl oottfer by mj the Huge Enor tea, any be ewtodid ta n dw wdculetwm of da calibrado* w ro Monetr, da teal somber of netneted catteuioo standards Am tedd be retorted m at not toned 20% of the total umber of ax&ncied ttandwdt / A - 13J The s u lta n coefBdm *) f tr e ttibntloo earns penaetted muM be &JW3 (R2 %&9ftS> I f anfibita n rem it* fa il eotnde b o e bimits. the* ^pm pnato m ps most be la ta to a fu a hnstnuneDi openrion, and the andad oe Ae niew nt a t o fsamptoa abte bn reanaJyead. 1J4 Putei no t e a bntwnea a te a f ti aid am pia mu* not drvd n on itua 1 4 % w4bln m nulydcto rm I f a n sie n te e d ri ft accondilldn b a il te d * W walydeel a n Ann An na a a lb e mnm^md. Hkl U t fte U b m rftf n p ita n to ta k te * (be m o te o FPOA ibttnd (in ntfL. I en peak am ) am g fla mndnrd curve (Jiawr repwaten pemnieten} rf by fte M b* Ijm x a o ftv tr program: pfQAftmdfarD- I a DP OP - b e a r bp abWb tto fiM l v o te * ww dUnad. 1/oeenavy. &***n Pag* t o f6$ Exygen Research Page 62 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: POOOl 131 E ly * P ro to c o l N um ber POOOl 131 vmmo F A A A im C A L METHOD | f A m ly d fc rtlw P rtan r i* W g ^K ir^ o to o ct< B O fe A d d <?PQ AJ m W * g by t u. fftatwMpolMa atk^riaitlmkat i*aMta>Mla amtkanynmMMioctsYafvOwAyp.an nncH M . it c*>- Ota] Exygen Research hm77 P at* 22 o f65 Page 63 of 110 interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Bxyjen Proocoi Number: F0001131 AKALTITCAL METHOD MMtodMaster VWOmi M M M * s T U tfra h fc r i AjMtyticiJ T o tta f F idliQ r * F M ir n H M r tf A M (TFOA) l i S-- >y ucM vm Bxypw 0-- r r t 3051 Kwntfob D riv* 8 1010mm C tlfef*. ?A ] AppKWMOBy! H A C-J.L PttA C M noihr * TtdMUeM Uafe,LC4<8.fa>9 Dm /Vice ! V i i * i t . C y-'T w . ft)y > Exygen Research 7<sri*VK7 Pfe73<?/di Page 64 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E iy jt P rotocol N um ber PO O O llJI __________ _______ vtA LYTiC A t M rrn o n ~ bCAH'VBW w hodrfA arfyie fee Hwr iia w iie rtiiiq fP irflarwm'ru i f A rid (PFQA)iw Sail by ' T b it m4md n to ba woptaytd h r lb tonI to t i iod n in tittrin w o f parfarowatanoic ad toy H ifh y w fa a m Li*tod C teoatonpty caupfed to a Mede Mass ' 2.0 SOKy *;. 'it" /flpattto ral t f Do wlor (LC ftC /M B) in a 11 A)tyfcBvitfrUMtstorypi*ctiM*. 24 OoaaaH beappropriate MSP9 baftra bsadBn u y chuateal Tor proper friy p p c rita m . 3.0 5toapk JUqatn w ir * '*A 1.34 falcali IS to ftm w n pMkofdoardefxtttnwcRm. piti, JJ faa^be Mud n iW K < llM nld ha O o w d le .f H n * n e ri J 4 AXavallai m m be feanu04y jailed badbeab r | aouplad fereaoaebM1J S tm k idifrtaa iw r iim in be apceSed la the aanehng plea fee U n PM - 4. H im -- m i T ltlir i 4.1 W a w -IM C in d a 4J M adjeaal- FtC Fade 4 J A-- w jfc w A e u m - A .C J . m a Giada M FvtiK M O ctH nc A nd - Aldbefe 11 to lto * wd E q u f d S.l 1144 1-4 JJ S4 5yy11.i7Jno 5.12 513 A kpfc p a rto rito la liquid etom am fM pt capatole o f pump up to 2 i l I m ay ip pad wO t variatola votapw tofrctoc capatola o f 3-200 pL o M M d te a m im Maaa Speetratater fLC/M &M S). A divtoe flU w iw Ibr IwegmieB io d qoeitiaatoa. AuJjticaliMbM w b l a of Midb to aM M ! f. M bU,diapeutofc palyp-apytena cew tritoy tab. 13 mL itwynaAto patyi--pytaa a a b ifttf mb. Dtapaaatot m ienpipato (30-lOOoL, 100-2OM.). )23-aL LDPB n i m r - m i i battiti. 2 *LctaarH FLC ria lto * Diapnaahlr pipa--a. A n p if f (100*1000 pL --d 10*100 mU m A pa. Water. top P tk Va* i m ( 1*) tC ll 8 canidfM . S n 'm pnm m m M . V b w n fc b a d t P ill 24 o f 65 Exygen Research Page 65 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Exygen Protocol N im ber POOOl 133 ffAl-->_____. _____________ VMBITtl | ANALYTICALMXTMOP J Mwtod A--frm ks omZimmtt!ion ofPtrfhiottaocUftok Add OTOA) to ta lly ic /u a /u 944 W eHsrioe Arit*. S4S Cilw lu i of iptBMOf90w i p o frffpyi--w>1m H5000rpm. U . r .b iM p y h ii tym m , IJ r 42 ^M m ijui Cefac*. PWcm *? QLxy*Bm Sca^fcbck 2.1 c i 50 wm. Ht <r*fcl2MS-to2l3C* Th p m m tm WC UaMk>PlHe(A): 2mM Aanonfam Atietaw in Wmr MM k ht e lid) : Mtotewi IB M S3 tft US 2.D 224 Pbwftwe 5LA u 4$ 35 49 35 29 75 25 75 49 15 04 0J 04 03 04 6.6 bfrwtaVchner lipLfcMbefacmeaedtoMsieobMSOpL). 6.7 QuiMtMtoe.' P a k M - ttiiu l ta fid ii4 e ttib c i< im c t v e . U kaTtaw ~23wta*_ TbpcAbneiHiircwo}Q*lCigosi)ertHt ie.mtfedm 6prid be dw ed io eider TO M&MS t a 74 MAr- fflecooaptay Negativ M UM wod*. w u to rii> i4 l3 - 369 mfi for PTOA, H w iiv co n itto M BFP law ew J i * * p a d * o d nw y b e c h*>f frd m ord er to LO PrfewatiaoafSofetOew 1.1 MM ePhi 14.1 2 wM -- iw lH w n te w H r t i y n p m d by jding 0-1541 of a w m im acebae to 1000n L o f water. A ta w ita M B o r to iN p iiii n w j f 22 65 Exygen Research Page 66 of 110 Interim Report #23 - Analysis of Sediment Samples fixygen Study No.: PflOOl 131 ftq rg e n Protocol N om ber: P0001131 . . . - ______ AJU1.YTICXLMtTRO U i t r t <,!***!>* ftr a Dwntfcttioc OrdIwfMKmok AcU (PFOAl Sell kr io n * ms 9.1,1 h*peB iladtioBti8*ef-tOO hS`* L ofFTOA ky wd^kiae 10 w* My k l, fw dlT d (comotad for purity) and dilute 106 tnL with f i n l k > 123-mL L D fg b o le . ' > /U A ltp ^ fe n U S e ttiM io tiitiO T ^ m A ie p n p i^ iy h rijH P H ^ mL of ht 100 pi* iL siutii to toxi votame oX 900 with methanol * 123oiL U>PS txxrte. *.1 J ' 1.0 vtfmL fer r i! W i n la tuftoa oXPFOA prepared by ira ifv r^ U L mTIta 10p jflriL mtuttCn to a t a il volume o f 100 w ith methanol in ft U lm llO fE te ftk . 4-4 A & lM jfta > L fra i6 c a tia i o to io * fF F O A jfp n p a n d b y b ria ftfl| 10 AL oTifei i^ rL ftOMtM 10 ftaai votftttMoX10CWia reethMf 10 * 125 m LLDTBfcnW . 1 L 5 A 301 p jfe L firiJ e a iiM m totka o f PFOA a prepared by s n r ftfi v S.l.fi 10 mL o f (fca 31 p*riaL ftokoioe * Coal votoow o f 100 wjCi w # --rik 1 2 3 m l.L D ? flto ttie . TIm ta x i tad tortjficto* tthxioci art to b* nortd ft refrijw*ior & appraristaltiy iK m w m ttah fer a mmkbmbu period o f 6 month# fern* fha 4M*o f preptniioa. 913.1 tC A tt/M fl cafibrftltoo m fe d i u prepared k HRLC nvmt The idcatkaltOBingrimLttSkmOom it f t d r t ar wocftftftftd th in g Ox trira tia n procedure. 932 Tie ftdrirwinf Qipkei m itf lr ftrthtem l MMDftitM may he Fini C iM H fm V fth M ttf Calbrattea riT Xf i l l i i Vaiarne N riife i C anal GaMaatim fe k M ta fn f) <aU Ira d a ln D h m im ito r StaadaadID (example) 0 4 Jto-- K n ^ 1 0 40 K 4 ! 40 40 m 10 40 23 XQmtMyy-l SO XCmnddyy-2 ISO XCaawidyyO 230 XCmeadiyy-4 m 30 40 IM 4C 40 *0 XCmwtfrrJ 100 XCmraddyy.4 TTwftrtacftftdicaBMRiDftaqXfo rittftftckft*adirttftiyftlio5x taka**] coanariaiife. feafto feft a a n n trarin o ffe r i - d m d tto f e g * vegeti-- (& Pt) x c exoaow c n a a ifti M aun. f age 26 o f65 Exygen Research Page 67 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E xygcn P ro to co l N u io b c r P 0 0 0 IL 3 I | A H A IY TIC A L HTTHO P j liW ^ia J rfA w ly w ilfc -llu P M w iiB B C T H ^ ^a n o m n lirfil1Add (ffO A ) ip S o ilty LC/MS/MS . * W .i A m o r a lard roturio* (u n p m Wank) -- * tu p n p tm w ia each m o f danim -it msaaeted. 92A Stare aS agrietad at&n m Bandard! to 13-aiL polypropylanr tWw M2*C <S*C, sp in two waak*. W J AJwtMM and oaaoanfri riona o f toadarda may bo prepared is MMkad 15.0 Batak SatUp 111 la ck bm b o f aanpha o in n o d (lypicaMy 30 or laas) m a t tactoda a k a il is scaffSM M attel (M ttkad Wank aw| 3 m L o f n d b n o i) and two reage Httrott JM fla I known cdneanbntiaas (lab control rp & t) * v tn fy p n to n l xooomy fc r nr kntek HU U ^ana nra tt fcr 1WM ta4 laboratory Jupfioa(a and (pike* w ill >c tpaeifkd in ton qaatky -- m nm plan far iha fncpact. * 1U in 'm |lJ l' 5u7 11J U> W a ifli J | a f m apk bu 50 n L pofr yc p y l-- < * * * * atoa (fcc/y aeaifed, Malaca t i n i a i x ttdl). AM 3& sfnwhanol eadehakaanwrit! actiacahitar fcr-13 roinutw. ro ta te ikataWa to MlW tnoakMOi od awdcala fcr->l)iiW m a a b u t ft* M httM M to M oaL wkk w aft* in Mb SO m L polypropyl cartridge (be. C ntes& ft fcr ^W nriaaitaa* *-2000 rpM. Ctawhka J Cu SPE camttM <1 f. 6 eaL) by p M trif 10 OL cmUim o I feflowt by 3 mL afHPLC wat 2 #pp/M c). Do MU tet eoKuan ron dry rhfliflfload (daca*} Iba --ip lo a* Uta conditonad Cm SPE cartridge. Dtacard Hato wsfc- ~5 a i 100% metbw&aJ. C o ito* 5 m o f stow inte graduated 13 w L p y7aap)fcn ec o triftg fl tWm (final yobnwa - Sn L ) Alyeto araplw w ing lecttiapiay LC/MS/MS. 111 kpntt fcnm * u rn a * a f red rieded sample and fcrtifted sampk me tt LC/MS/MS lya ton A catJbnna standard n u patada and fcdow a 1 2 2 S tM riM i o ffQ A nw e^ ie a i f i l 10 M k m t fla n o r n o ca canait m fcn *v K m a t bn iackadad in an anaJyricai m l 12J A afea M t o f eibaetod etafbaattoi nnH m fc nuat ba chided at * * and t i ON and o f a iampia m , extactod andarda anu be ry 5-10 eani|ii-- Aa an a im a ttva , an m ito tex t i Page 27 o f 6 ) Exygen Research Page 68 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: PQ001131 Rxygm Protocol Number TCC01131 to m to--to . ' to tlO K to u VtoM m 1 V - : AM ALVTICAI- M TH O D M bted OfA M lto tl fc f d a ;n a il III Hill ftfHrfooaooc-- ole Awd (PTOA) in Soil by ____ LC /M W ! n ftm lT in x tm u mag to w r-- l it ito h if t o ^ o f t m I k)r <toxto<l totonaioa m to to toununod n q i 3*10 mmfim (to -- . t o n t o w t o t o f to tra cto ) to ih rrt l to a to ir cto e. in 11tU.*tHUottooUelnltnotootrTnrtwofnetortoiritorll)mttooutroraftocMafneteoflymrM^ttootoniohlmnonMa tmoUt*fllAtorwIcUifamfedarnpdfaoofwmp.eeukl mrnt t owiMKnBJen uttof MeaCytu 3 3 (or o^mritewi Any m tpl Im hould to fkrtbor diluted n d reanalyzed 43.0 r ftpafc of daifM* 00*M 9 m u tern i p*mx o f 413 m l Tto 4 P IBM y tto t ra n t | toll a cto ffO A traoo. Wfaik te d w fb a ri y (?d$ to o ) repo--n tj ito km o fcc to a diuxide. M atted Mm bju* no ffrtam PTOA at k re l peater than fee LOQ if * Ueafceim aiM rFOA et te a k p e e ta rte 50 ii^ L , fe a s o blank-nipte -- beetod--ri w d tlw w iifH O ia iw N ^ o iw o id d ttutnx y ito e m a t be between 79*130% of [f a ooofcri pik M il aide fee acceptable tont. ic tto m o f -- pin tborid be in w d Ay aM a pte eudt A>130H A m M be -- In-- d by 9m Kuyac to d-- note* if n-extracriop is idfcnM i i(m M ta a d to to a a to ire l cwtfeer by m m 9m H u y S tm T M r a v be totaled te a d a a to ll-- o f d a ceitbcidcn curve. r s f t t i-.tr* ciUbniiQA Mtndsids hot could be t m i M ead 20% of th* toud m otor o f n a id M dorfe T to je n ito tie ii m flu ia (Jt) to coKb a ilo t w m i foa m e d must be iO-MS f it 1 kQ JiS). I f oatibratiM rtauto to t outride ttoae to iirt, then ; to a im to d a iMtruawit operation. and ih r w la rrim m a iw f m ^W e ito e ld to T W to y eo d . l between tto tto rd t eod u n p in a w oo< driA mow te n 4 % ato to im ly d ri1 n a . Ifl--ulm *drifte-- to*toutw ton b analytical (SMt o t tto a t a m t o m m ly s o d ' Exygen Research Page 28 o f 65 Page 69 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Kxyjtm Protocol Number: P0001131 U yfN ft vooomi -- A2iXL~mCM. M X T ftP D | M o tto * tf A a tly th forlbc Dtionafattlkm ofP*rfluroect*k Ae*4 (PFOA) in So l fay LOM SM S M.I 1Vf * 4 * fae-rtf Muotoc to ealwiUH B aaount cf fPOA fc o * (n cii/L. fMMwM*M0dpboyafdcttimM)w'Lrijtw#iIttotlhUlvfftfwprdopersraa:(tmttr rvpeukm picaiiwim) r PPOAWifrflL)" D f- W fa y w to d tto aal nmm W J For w o g lci fw lU M M i known kdotoU o f PFOA prior le attraction, uso (fat M o ia n t f a t a t e oofcutat ito n e t raeovay. n it r ii tir in i f ip l i 14.3 Un ite fe&owiag tananai n n a w t In m ona af PFOA fan4 io nfrt- to **> WPA * m i l~>3-fWQA. * m lMfl.1 H an. M m l 1041-31 144 U n * M o % aquation to ntenlau *4 a m o f M C A b o a t a fpb Omm irfwom m . m u b o n d (M b )d ry aH M -P T O A Ihum S[fp*)< |IO O H Ib o 1io M o(H )] r*o>ion P age TV c f S i Exygen Research Page 70 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.. P0001131 Exygeo Piotoeol Number. POOOII1I ANALYTICAL MBTHOD ; V O M ITO Antybcai Testine Fiaty; lyL O M M I EMiI*JUmrcfeDrtvt 5 * C o0 h .PA. tM O l Awvni*r J^Oelo)tXy - c j L Tk M m I U r te . L C -M *. E > n a b r n n k Oew /vJa*otmPlmr*-'i--yM / *U r Dece Exygen Research P a g t JO 0/ 6} Page 71 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 E x y ft* P ro to co l hAanbor: POOOJ 331 NwtoVWI1713 ___ ___________ ANALYTIC AL MXTHOD | o f Pftvfoavooctftccic Akl (PFOA) in SdnwM by LO M SAO *I 1.0 r :- Scope 7M m ttko u to o a p to yto fc r toe k W t t oad ^ow b*otio o o f porflvoM ocm oic cid ks- ? ! U qm i C to n rto g ra p fe y ooupiod k> ft utoto ni M m 3 p e ar lr ii OcUctor (L C H S /M I) k d to n t 1 0 SoAty 2.1 A lw y otom u Mb b to n m y pn ctoift. XX Q wiw fc Sto w yriftM MSDS to fe rtlm ilf t ii ic y ckmmc*l fee proper fifa y p n c o tiio a t 3.0 Dftinpli ftr ir r'i^ n n l XI At boot 30 $ o lxot i--y k f a gxfrwtioo. 1 2 H o -- p fa p n i -- 'f ie -- r fto fa rw d to n M m iMft. 1 3 - Sftppto -- t o nM gftaiad i M t t o O m i qutbtauc io roe 1 4 A I iftm p iift m u* to tto tto g U y to rn ! to A n r hang w n y frd fo r w tn c to n . 19 Supple coJIootian procedure* nfl to ipecito io tto im p ta * p in for Ms 4.1 W are--I7 L C p o d * 43 H iiW iM flC p to 4 J A n to A a d ^ fw ^ o ly to ft 4.4 A a mcwimw A c-- - A .C 3 . lU tg ir t O n to 4 J f a ftm o c c tw ii A d i-S ttm A M rid 10 bew m eeiw S Etpmmee I I A M flb lif c iM M M *dd f t t o M f i p l cepehk o f p m p w f * to 2 a to m A f ru in in to wtife t T r tM i in lm n o in jw ito r r y l l i o f jggciing 5-200 jlL co-- rtftd te ft P to M ilfc M flp e -- 52 A wrio io ooHeeln w d m far peak hto p ilin m l y p a ifttio 5 1 Afttoptort tto -- rip ib ic e f rcoA f ip OXWOOl Q. 1 4 S O m L ^ ip M to fftly to K to i o o n ftrtftiift Qtboo33 13 L d to ft--M* p o frp rc y y lw t --to fr p m oo. 5 J Dh p-- Mft M n o p y eti (5<MQ0bU 10C-200o14 1 7 ' 4 M . L D P l- o w - n r a * b a to n . 5 J X aL dm H rtC vtoto. 5 3 Dip o ot o li pip-- 3.10 A a r-- -- (100-1004 p& 0 10-100 hU w f> *p c w M o bps. 112 m \*ftcanMnmJbMI I I W M n Sep PftfcV at 4 ec ( If) (CIS SPZ n rtrid fm Exygen Research Page 72 of 110 Interim Report #23 - Analysis o f Sediment Samples '!i- 1 Exygeh Study No. : P0001131 E x y |c n P ro to co l N u m be r P00C1131 If I ~ ,, . AtmjfTKAt wrracB ) M c M ttA m H H I* 4 o U m !W in V tribtrxxaam * A a i ;PFOA) w S H ra n hv LO W W ;,.xill. wl lilwiMiw r , U l n il ll n |a | M a i^ j i | 3 0 L clj^ r y )rt-- *MMCOnnj. 6 0 O ogiili r ^ r ty-- 6.1 C e b n * r r 6 M. W Qtwyuam J c w d E c ),X I man x W m . Ji* , , , <WN: H>5-S3t30) 43 T o fr tR V C 4 ) M oH iltaK A ): m il A a tta W n w M H M A k ( : Mi n i 6.3 O M m W w " s r 1.0 mo 22J S4L5A 65 25 25 45 PlowR*rt S3L5A faaL/m ^l o3 35 03 73 0.3 75 A3 35 A3 ' `t . 't i i hi|T~iinr " r 1------15 <<*be acrMM d to as ouch as 50 gE). 6.7 Q w d W h e : N tk A m - la c o id itw d w d o*fe6n *i c curw . 4.1 **Tfar ~23non. T h lw ^ e e e 4 W e * w h ttB d e d w a g iA k * d *Ey b e e tw iy d ie e4w opikafamifaB HPLC 17* 0 . TO M M a S yM M 7.1 I^B)KliomNip^M]P4a^JWiBn^4ta-36t>^ifcr r. FPOA- TW a^v* MiWMi atM ddai m 1 pud* asi flttQrte taanpd tk r to o p tia ta to M B A jfM a U J T n p M tiM b fS p k tk M 1.1 ItoW U f% m 1.1.1 r h pmftnC Y f adtfinf 0.154 g o f Exygen Research P tt& U cfii Page 73 of 110 Interim Report #23 - Analysis o f Sediment Samples . Exygen Study No.: P0001131 Exygra Protocol N r o ix r POOO! 131 MS? Aaatjoo fc AffA TYT IC A L METHOD 1 of ?wi*iionaMoic Add (PFOA) MSediiww by LOWiB U Cl&VQaB MlNfSnf 12.1 !%Mctkaedia wtftrit pnfiradfcyddtet tOmLof lie Kid to ' Iis L o f m r. AJHm>H r o lt m y be prqwnri. 9f Stufitta&M&cMiamSmkvAam , '9 /j J r fM p a m o d ta d N tiM a f^ !* Q * j^ ftfF K M b y ifh i|i li) * a ti a ily tioi l --nOPd (om nttad <w panajr) and dilm c to too mL ri* n h ^ n lk i 125-nL LD W ioM k. 1 U A H a fta L ftalH katioa ie l to tttfP P O A i> im i id b y la ri*p n f to u L f * 101 p y te l aokidca * float v t o w a f KW w ife n i2 5 is ilX * 9 VM* 11.1 *, 1J>w ^aL ^ficiiioo otokno/PfOA pnparvd ty bnaging 10 mLtim H ufteL k linp la a fa d vofana of IOCwkhaMfeaool in I 125 LLOPfibmrta. 1 1 / A M w # *L fcnUcainaaotartaa o f rP O A * pwpattd b yfrin n a g W n L o ffa 14 iffc L tutina a fe d volana oflOO w id autenoi ft 23 LLD TSbatite. IL S A 0.01 n y M . ia tif a tin o aahticn o f FPOA b prepaid i>y brqpm 13 Ir o f f a 0.1 p jfa L aohation to a B in i vobuna o f 100 with esafaal * a 125 L LD fB boote. 1 1 / Tfa acteandfud Lartoa la tin t t i ba wofad tea tatef! x ej^ CTiw u hr^ ^ d -- iM a flirawaKkwMupgiot/iPOHhi i n ili dateoipKfwaHtxv 92 3fad Qtfbwtfa lotedr 11 LC A ttfaf casual Piallate a te il toafaool fa dtteai. a fttiU rnfaLterideaticnaototioa 92-2 U h fttb w to t I * typical M arnila: id dhit n l eoftccatraliona may tx tlnafaari W at) tot ICC too 10 5 2 Vatina rLi 10 5 2 10 W D IM to teL) 100 100 100 100 100 100 final O w n niaa 100 JO 20 10 0 02 PofSS 9f6} Exygen Research Page 74 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E x y g t* P rotocol N um ber. P0G01131 T itfU L y n c A L x tT H O D LC/M/MJ AoM (PTOAj in fedincM >y S x n iB oHMto * i . m is IBVoL U Y E ooarcm ooa.'i tooba IJ.4 end A iM M r iM N Md nparar v i ad H M aaeybe prepared et mM 1(U> M d M U ) 1IU lo o k M t f IH |I nibnii l f t paaeUy or M t) n a Miede heu fl M M ooM oL IM Wo M M erdiot feetified n r o i o m m b M o O M M M t M M o o M Q r p n o o M n i n c a 7 fe rilM M c h . 1 J t i i 0M i..i Ib r fielt iM M o n w y t + h n f md fa a fa e w * bo y o c ifto d n O w .% -- p e r-1p M fc rM a yaej ar t ) U W efakJ o fw w to M JO m L wtypeopya w h i i oabee Uanif m m W . M M M Mb w . ) U A M 35 a -e f t t osobe oed, < w rode oa abate M m W actio aabar Mr-dO aaeaoaoe t u Cwari<UMa*aM~XIMiwi(br-aMMat. L U C a M U w M C S carMdaaa 0 ^ 6 M .) by paaaiat 10 n L iw uiaool M eiw b)r20 oa./H T LC w *aH ro rW c). Do aol le< alaeae m dry L U Uod M X ) Lbo aunpl. o LM coodinoori C ,, WB carmdie. Diecaxt L U AM 30 M . f M u n t M d M M M I Mk bedon of fee JO b l e M lifa fe a d r i. w w d Lato a a eerier ooo itaker far - X 11.7 Cwriftp * M i t eJOOOtpo* * - aiiMdil L1J DacaM fee eriMoel onto the aw e 8 H cartridfe- CdieelfaaaMd LL. WiMfao rntaa edibaal.adifawnl Cid w iba ettieteaod add eio d MM cotactari io a 111 LI M CaadibeiiaaooMCIVTaaMdp(l 1 6 oaL) by pea!* 10 m l aKfeMet ftlloinJOy30aaLofHfLC paar(~ 2 jaog/aoe). Doaacletoebaaaraadiy 11.n AM Mo audkaol M - JC0 m l o f alar aad Laad ao cb aoeood eoardaaaoeed s ra M arita. 1111 Sbao arLM-5 aal tOOH fau l Ctroa J l o f ofaot do mfejerad I) ad. peiypaderMa oaaoaltaga LoM (Mal m a a a - S otL). LU J taly a aampma aaio ilKboapcoy LCfatlritH Exygen Research P a f* H o f 65 Page 75 o f 110 interim Report #23 - Analysis o f Sediment Santoles Exygen Study No.: P0001131 Exygen Protocol Number P000113) _________ A h i^ m C A L M XTH O D_______________________ j o fAaalyfo foe t e Dete Mit tihoa n fParfleoreocteck Add (PPOA) Sediment fry LOM&MS 12-0 O soBm So^kjf 1 1 1 mm m n n i o f te c k andari, a a a p le and fb rti& e d M ia p le in to tb * o r in A catto*tkm anted n u t pancotto and follow all 113 ihar to da a fffQ A tnp o q ondiegie at IcaM fiw or now concgntmion leveto MMt j9 teeto tal In n analytical m l 124 An entire ta l o f m anosi edibraitan andarti must be incfodcd at the bcffeataf and at tta oat o f & V npte ML S tu M O m u t be intenponed Mwean w ary M Q opto*. A * an aHaroadve. m wan; o f teto m n u i anto d i may be weeded ttw beftoamg o f i aat M owed by cajbratkm anted* inim ipnnaif every 5*10 a n r tn (to account for a mcqo let of H 4 Use Smmt anted carte l te ^luntitatkm . Linear d a rti curve arc filM to rf for toe te iy to by Sneer w raM om ii^ 1/a enac t ing o f peak area verme tte bn tx a a ttu ta i oonceotntkM nog M icsLyra 3.3 (or oqu*v*rt> a o tw n ) ttua. 124 fo rn ito nap foni ibeuld net eeeeed Hm M > Any 13.1 ActfW W a CBMnt 111 444131 t e s t e r tan (3 o n ) npnema foeleu ofcartondioxide. Mttbod Mack* ttnet aot oeaain FPQA at Uweia gnater teen Am LOQ. If a btak nertttoe WQA b e b te c 04 ogfeL, te n i mm Wank aettftea * heattuned and t o aneto a ttu ta be M -uM *d. S13 Reoowadaa nf consol qdkae and unsi giikaa am* be between 70-130H of ifowr favM vaine. Va control apflba fiHt die acecpeable Uniti, the III Ifill *MlN W r<CTCla* Any nam x <p&> o a tiid t m. 130% d im ! la w iitM i kr Oa m ly tt ranni it n -m n e tk m a 1 M iI bji u g n ili B im ini Hum r I . > m ila i il m ill, f im iii, ttn llup-r A ra T in , r a ba * la * d te n te a lw ln ln i o f On caUWaOon t u n B n m , t i m il M atter a t m e n calibnnoa a n te * te i maid be -- i-*--1w in a rt tacaad 20% o f fea 1olal aamtwr i t nn cttd teadards laanad. IX Tte a o w ln lw a a m d w l d ) te cakbradac o rra a |a a a a d awn la 20.992 OL1 20.9!). it ca lInalca nani M amalda ite la lim ai. c m i la lakw adlan uminmm ofam iaa. aad ike t relevant aeto f te n p in ttrrvH be n m ly n d - P a g r i! o fSS Exygen Research Page 76 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.iPOOOl 131 Exygen Pnxcco] Number: P0001 131 [M O tb i o f ............. A M L Y n C M . M tTH O D | P to E in iiiliiw ofP irttooioocim ak A rid (tV Q A ) ut Stdmmnt by LOMVUI D i 1 -- Boa i M b m M WriiwH md nmptoa hmn* m i drift non rfcmi 4 4 %-4wH k M im ly lirol . I f mriptioa tarn d rift g o --do ftM to n t --thin | | J U--itofeltaHo#oq--tiM to criwbto of I o*dybmyktom.kmfc}.wLfjetf Am Ultimi m m (How rofmokm m M in ) ra id o f FPOA prior to cxtroctio, we to ralw k to the p--m it m over?. 14J U-- t iqpw io W w w l fc * -- a t trfF T Q A ta w d ittn^M Lia P T C A O m U p f^- B ia p to m W O 144 u w ^ iU b w iM mwriwi Ofi i w Tlio fta iio a k w of rfOA M i l Pfk tend ao dry fFQAfcwiipM*r-^-ProAfairtij*)[IOOH/ll*(Hi) Exygen Research ftrtj J Page 77 of 110 Interim Report #23- Analysis of Sediment Samples Exygen Study No.: P0001131 Exygen Protocol Number. P00Q1131 ANALYTICAL METHOD MpfcodNumber. V0001713 i Arid (PFOA) lmFlsJi Am kfaal Tmtia*TwSSf. Sxyita ftwnecft JO ttXam m tfcD riv* Sum CollefB, PA 16101 ApprovedBy: C -Jd L ____ M iM ir 1 TactuMi La4ar. ic -* a , e w tMMnb g/w f Viona PriwfiiiftlMn . O yidom 8 n y r w rt Dste pM r am T a ta lH fM : I Exygen Research Page 78 of 110 Interim Report #23 - Analysis o f Sediment Samples " ' Exygen Study No.: P0001131 Exyien Protocol N m abtr ?0001131 ------ I ......... -- . __________ I ANALYTICAL m ethod .* ' Clam# by LOMS/MS M a M ta a b vV fM lT n | 1J Sny.e S-i.b ': . S .-i TVw aaoiaod Sot W oMpioyod fat tho imhaom m i y --r ifttion o f porOuorooctacoK acM by Hi** fa iiiu n B c i Liquid Chromatography coapM u> a tandem vu & Spe-- rfo Potato* (LC M O M fl) in t e and ffa m . *4 Safcb 2.1 A h n y i obm Tf l i f t fa ftM ta y y tid iM 13 0 -- ft dm appropr iate MSD6 befbec ti--fla g --9 cftem kri for proper wfcty 3.1 A lftm t 2 0 | f|M M fQpieibreKmctiotx 3 2 : f --f li b --Id W pfP -- ad W Jta aataadfaa. PS-- t a ft-- nw pte ia a feedpw cataK ia d tioftiEi i nho irttft try k *. Pt-- daaam pksin ccatauren ad Sqm op-- .hi ftps-- atimy ovareqfti la a lte r fa r carbon djetod* anSiftwinn t e l ro d pile die --ftit b frcaan d o u |t w rit tim e o f -- lyfc 3 2 Sampb u t e n p-- do m bo ly cfftod in tie mmtptog pi-- for tbi% X*4*l 423 . lS i jM if P t e 4.1 Wo-- -H P L C jid o 44 A m tm ttik - HPLC pado 44 Cub-- <120-400 me--- Ita g --t padn 44 T ifrtte o l - H7LC sad 43 S ite foM dM O O m a*)- te g M * pad* 4.6 PSwiril (S6lttJmeh) - R atfM ii grado 47 Suporcieac LCNBa - H eap p id o 4 4 i-H P L C p o d * 4 ft I^A -- t e a o d - I --p a c jrid a 4.3ft D ite b yM rfclm o iil-- - R - f t- t |a d c 4J1 Tab--o- gMj--1frida 412 AnamooriMl A--laia - A.C4. Roogmt Grade 433 ftaflaa r-- fcA pd*-pa'A U rid> S4 loofti-- l SqaipoioMf 3.3 A U |ft p--bm --oo dqmd cJaanatopaph capable of pwopto* up n 2 io p m-- q a i ^ i l --fa a w ia M a vot e a Lajeotor capable of ife tm g 42D u jrL -- tadla a taadaa MbaaSp-- o ur (L C /K 3 A I3 ; 14 A d--S-- 1 o c te t war data ftr poafc iitie y iti-- aad p --m af iaa 5 4 A--lyfiiW b a l m rapabla of mad* 40MP1 p e--r ) r t M t>/ 65 Exygen Research Page 79 of 110 Interim Report #23-Analysis of Sediment Samples Exygen Study No.: P0001131 E x ffe a Protocol Number P0001131 [ "U rtw dot A juiyJ. I b n U Mate* *-/f t* i?C3 AW ALYTKAL METHOD offtH lu arao ctiB o ae AoU (HPOA) i* F*h in 4 ClaoKky tO M V M S f M tiM 7 fn v n lK J J T k M a in r. u ia L p w -if^ flM U 5.1 15 t f t> y rn jty jw iw rtrifc jr o k --. 3.5 D i o w b ls rt e p ip e {S M e L 1 0 M 0 0 iiL ). 5.W m -W .U y g (W > M l Im ld M rW L C r it t t it . 5 il3 P |^ g n |l| p^pMlML 5.33 (IMMOOOiLa i 10-105 p i). w ik dbpnwMitip. 514 9 1 tab (2 tn L ) (Sapdcw m l m i. 7*37177). 5.15 W titf t o * * * . 516 r w 4 ifM >ii<5*M irf<piaB iaf 30 n L poftpapyt tufc H 200C rpp. 6.0 C!w tujwjhicSywaw 51 pkyfad C n k w Ftaphiw K> |Kytca Sekwiflc). 2.1 tarn <50 ctm . (M fc ftS ftM ttU lB ) 5 2 ' im u w r 30*C 53 U dtkM w M (A) : 3 M A m m * A<W*t* W *r 5 4 wwtoJo PJn m ( ! ) : M xTW rt 55 G nicD tm m K How Rmc % A i l (M ijm m ) a & 35 03 LO <5 35 03 50 25 73 03 250 25 75 03 7ZS 45 35 03 6 4 fcfrctjaa V o ta* IS pL (c m he iocttwod to to mefa*a 5 p i) 5? QtvcMtttatka: * * a i An** - u m h ) sttn cta l caliboo* curve. 4 J RuoTfJM: -23msuw- Tht atart ijowliltox ire iwmirinri aa tfaid a 4 a n j bec W y d ia order to tptinta o EfcoUHX ijfiN w . Exygen Research P ag* 39 o j 55 Page 80 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: PG001131 Exygon h o to c o ) N u m b e r POOOI131 -fa y y h w w li AKAJUmCALMlTWOO ] Mib a i t f A n lym fcr dw D-- In r ica o fT w flo onw -- ok: A rid (ffO A ) '* F ull and .... ,, Q w M riyLO W M S 7.0 MMOIjilw > .i M fe B m w p n y N tu ltw fcClW mode. w a u ic n i 413 --> 149 arvir tin FFOA. ! TX b H tu ftlw t mo kmadai m > >wd m i nay bo ri-- gM la outer to optoMa ih* MSMS q m t U r.i.i 2 1 is or-- r it jnperad by addbf 0.154 j o f a w o t 1 tMO WkLa tw bm . i l l 2% -- ariwc Madk -- th u n l U | * f arid by fliw olw -- 2 g o f -- frc --Mu \W m L a tn m *m *i 1 2 2 >0H M -- I p --chk)-- to loin-- prtpercd by bna* 3 m l . p fthm-- yVfcrhWn)-- M a final vote-- o f) 0 w L w ith toluene. A lto n-- v-- n nwybo prapond. 9 # S tn to d fa y -- Aon 0.1 8-- te d fte d iffa fd fi-- iaoSofamoo fe U to -- x o a k io to k a af~100 >>^aL < f PPQA b y -- erf ly tk a i MmuM J (ec-- 4 fcr parity) m d dftut to 10C mi. with aw4 m * * * a IWhoL LO W M * 0 .U A 1.0 mpML A ftt&codc l ehrttai o f PfQ A ie propered by W w pot : ad, r f t o lO t W haL w tafea to t f in i volum o f 100 wito n o te *! A t 125u L lD F t to ftfe A A l p jfe L farffleedoo *c lw k * o f PPOA It pttp-- d by brinfW f 10 * i 9 f t e IX) jtfta L b M ob 1 I n i v o k n to n o o w ith jw ttaooi in - fttiS H fc L O K b --b: 9. M A U>1 p jfo L b tflc u lo a aoJaboH o f PPOA * prepared by tanking 10 mL ef Dm 0J p tfe iL wM iow to fiaol A o f 100 with -- m e lh i 125 mL LDFBbo-- p. 0,1J The -- * n d fc n il-- ioo *ju tic m -- to be -- i* i frig -- tor * -- no-- ty 4*C nd -- itthlo ibr s Ew dm e* period of6 nwwh* fie n h td M ittfp n p n M t. Pagt 40 o f6) Exygen Research Page 81 o f 110 Interirr. Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Exyfcn Protocol Number P0001131 a w a i. \ w c j . Mimico ] Om m W UVM&MI M i. MAcaooaolvmo.t.2 J LO M fW il caKbraban tfa ia id s >v jw pnW in methanol via dUtoon TW fainw wg * Sf it t i Lancio: id dlhanai M M M d o a i maj bc ili fa -- VbIm M m in to U <ntonU 1** u HO 949 14 l i Ito 9.M9 14 14 Ito 0*1 45 I I Ito 0.909 u m 19 ito 0.0025 l i 1 Ito 9.901 tu ra 1 ____ 12282____ J J Sfai all caUbnimiawM) 1 2 W . " ti2 T < > *C .w < o w H ta . A24d.2.4 u m votem i a 4 teeu a u to ct a f la p lg d i im y te ? w w d c ,0.4 a w fa S a il* m i t e t U d rtwmftm m tM (lyyo i y 3* or X) m t tuchd ut t e i oh uni--14 M I d tu* eunuca murai lo tiM * kisira mKkll{ldi a m i faffcd lo vntiyprendami ftoovety ferite Me H U ftaqavnaiit far M 4 m bbontory O upfatn m 4 ip ilN i wiH te peeifiad fa ita om lky n u tea t ir *4t pnricet. 114 3o*i b m a c te t i . l V a lili J I * t team a o p h it t i ad. polniropjrkar eaw nfe|t u b o (M im d a d . p to M n d a i voli. 1*2 M i io mi. o(acBenilrJe a d iteka od%wnal ICIien itefcK { - I l r r .o a o I l i . fia t* fa tute l i a A m ar ftv *1tenir. ,1U . t 4 r-- lliiK lla g g n if c M il flu ii U w ^B fH lm w ! B o ta I I J P U Uh 24 wL STB tuta q m c * w ifa 2 g flo rid i, 2 g a lia * al. 2 t *0 . 4 I I LC -N H . Co4it>M Ila III III Wjilb 20 m L o f sbctaanci.. - , fan 70 ' c / Manrmill DImte di a V i a Do noi dtow a calai a *T. t u M a fa 135 hL p in fa ip il u Vf oTM k tl !0H m id i f a - tir n ifa ii fa ta lo n i lo d . Xjm afac f l i t wM itotuonce, fals a rli 7 l i li I l ( t m tia n ). Dry fa A nte nnuplf lr ty M are o c . f a r te dHPjfafl fa ri* team f muoprt H14l P agt i l o f 6J Exygen Research Page 82 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: PC001131 Bxyprn Protocol N unher P0001131 ..... ,, fatrnek . , M a ta ttta a r VtttHTO - I- ....... AKAIVTICAL M T H O P V . MMhorfo t ft. d r>iCTiMiLiii a fH rth cn o a n ire AM ffFOA) in . . . . .......... .................. q w n b y u a io /M i It'.T O ttriffefi 0 a ja tffK fflm t ubm em--imma unei. .2900 im fe ih 1 M ->B*eta to t M lo t e s ta tfa ta STB co tta fitted aids th* m#t> te te* p w t e p il la te . C o tta Ove tete in ite 123 mL aita te } pearrtapc ta te U 3 A d i 10 a*L t e a ta o o te ifc t i t e ta p i* fa Dm SO m L oM Cnlbfe t a c I fara o j te t e ( te t e t e t e > t e t a f a t 1-- t a t t i - 3 0 corva ano reme te tri -- h ir t a l -1 O rn ite ta ta Jfa h imo ta ta . lf.1 0 S ita t e t t a i Ifa fa te r-l m fa ttm ** w riM ta fa * fa ta *. 11.U Vite rim fakm fa a te o u r te *'1 te o r mom. 11.l i Cantrifta te* 51 mL ptejpanprl tate* aoniaufaf um ple -2000 pn 11.13 D ie ta tea t w ctao te * o 3FB fa r o C o lta te Jum imo te M i ilip w l fiate ad cernit a ta te dm fa firn * tto 11.14 P ta S te m L e ra o ta ta rifa ta aIta S fE cofano ori m atta* ita eterne fa dmmm*p*mmiu|mri fiate. IU 5 ta ri * 4 ta te or l-octata io I e ra a ta peartaped la te fa vpontem ra ta ta pm aMm ta ta *< > ra y a te (*> < 4 0 *0 . I L V ita te * fita ta w , t a t a 2 m L te 2% aacoteieafad am ttata tte p ta ta u r i fiate t a lt a l fa ta te m o * 11.17 Tnmrffa teaunce HHC vita u 11.11 *m ym -- mte M ta ilcriraapny LC teW IS . 110 12.1 fai rima am ata te s ta i andarti. ampia and tentead sample imo ri* LC /M M 4 I yaCara. A ta a tfa a mandate m ta peata* t a fallow all 1U I t a fate* teTTOA fa la ta firn or fc v ti* 0 3 fai fata afatetalfacatkm modarda a ta fa ioefarite tea tafanair^ufa al Dm t a t e a a ta p l* ML H a ta * m ta V* fafaripew d bmwee every S-10 aaaripta. A * M m fata an ta ta am o f etlArat standard! suy oc fajamri te* ta fin r f o f * *ai feifawed by fa ta to * Utoml ri ver? M O sample (to aetmmt far in c o a d M ite N atante) la m b* ti Eras sto im inject fa 0 4 * Um 1 p m n ta fiarte* anofaaeV?rimar rapmte tuta* l/x t a it a s te paak acca - vanm i tritatilo tanfate -- otratioQ tufa# MaaaLynx 13 (oc affavate*) P a g t 42 o f 65 Exygen Research Page 83 of HO interim Report #23 - Analysis of Sediment Samples Exygen Study No.: PCOO1131 ExyfeoProtocol Number: P0001I3I to ie m v e e o in j I jllTTTCAl, MTHO I ums^ofAi>b)wixfcPe>mB^<rrr<lucnimicA<i4(rF0A)mawl CtaMbyLOMSMS O S Seml IH M *w lM aoe w arn* um turA ttm mm m ey m u * agmUaadm* jaiponan ahoutdU tettar dihfd andrxm iyxd IM ( V w a n p w w w tilw if ip d it fi iih iir k a ^ X lw f a B ip M it a i tm m U m 4U mm p n a c etaaapoo lo Un T O A me W t ih dngter too(30ami h*m * te tat ofwrtoidtatl^ 1X3 M aUwrf hfanfcs n o * m i r tt fc PPQA at lorde p w f * m Mm LOQ. IT a taak contai W OA at lw d p w lw Uno tX3 ppk, fa v i s new blank > f te onar W oM oaU awMbc d R N tn n e I iMRizacaed. 1 3 4 M im w io tfm V B l IfflCR 4 M i x to ta m a I t a n n m 70-130* ut fa k a m n fa e . i f t M utatotastatoli*om totofaoceettH c k a ta . il* U n m Iaaamhe ilw M to immfemad. H A Any c a n a to * a d o ri Sein b e im e liti w K Oy m y ito Hu*e Boor tu e , n u r to ttoMed d m Om atoctoai iti u t dm a ftm ia n curve. I to . , Su la i m it o u t o a ttn b o i andarti ita a rid m ila ri m a to m l JDK o f* ta l atadur e f a d ir ti iajacid- 1JJ I t a so Io w to fc lr t l (X ) fee U M m i ta u n te d man to M c f kOSdl). IT Iton alm tm toa N I m aid to rn Smite ito Hfa ef r ieu Mpa n a I u lta adjiut iuemmum tpem ua. m l the mandarti Su (S M B u t m m fim fa to d to taamlyaad. 13 u n ii ito m t o n n o d a rit a d eamptaa m w m i i l noi th a t H fa to m m i a t ran. U n K aS w ikM dnS w cceS tthii lim W ie I m ijlit i Ita ta m m be reanaiyMd. 144 Citatoiota 141 U f a I b t o i i t a t e s m l m na tami m a r n i fP F O A f a n d (in t*L . to m i ob fm k m ) m Su u n itar i c u m (S aar i w o o pm urcuiw fm a ito b ito tam L in to h rtn ilo sm : ltoO A Ih--I ^ H . S t o m . t a m l l tom 143 Ita fa ta to m o lla Boom * d u drfWOA kamt ionytaL m*<r+\ 7TOAI u U tin ta U> W fa tta torfato f a fa to w ta m m dthtad. i#mamm e/. H m iutt Pttgt 43 o/6S Exygen Research Page 84 of 110 Interim Report #23 - Analysis o f Sediment Samples ' Exygen Study No.: POOO1131 Exygen Protooot Number P0001131 b w U w k .- n n --l.i vceciTt) | ~ ~r A K A i.Y T C U .ItC TM O P | >IO i_'l o_^A_M_<w_.lt,^ rw ^ Cm'liirlnl7(<Ur iiUilu>WmM_.--__k_V_td_(?_T_O A_)_i._l=_nt__l I O h i tmoptm M W n t i know i t u i a ! PVOa prim to cM neiiM . me tfw iHMrwt 1p ctfruli fc tfap w a ii ngpvtry, *>**<*) l* " Exygen Research **4 P *$*44of61 Page 85 o f 110 Interim Report #23 - Analysis of Sediment Samples. Exygen Study No.: P0001131 Exyjcn Protocol Number P0001 ]31 ANALYTICAL METHOD M * d ) W r VOC017M M M T M b * h r M n U a W V ra rn ttM ty L O M tM t t A cftH ?rO A )to Aaftfertfeal Tarttaf PadHtjr 8 xy|w lU aw c6 W llw filiftiy i StatoCaOa. FA 14101 kL^Vuo<nti-vX c a Lr ',100*3, r /*> f h / y ^fZ*L.OrK*lem.Zrypm*Mm** DMt Exygen Research TotelH^K 7 o/*J Page 86 of 110 Interim Report #23 - Analysis o f Seditnen. Samples Exygenj Study No.: PC001131 CryjEQ Protocol Niaober: P0001UI XMmltfAnboUfcrtka A ' A i. 1 T IC A 1 M tT IIO D j f N r ib m u c W M i t A c id (P F O A ) io V n o u tw b rb C M sa a . .. l <* TU-->Cliio>oiMpbjoCfcr>laHttooto4iMriotin f>iUoipoconwc m ii by fmliMnmi i L if r i fto-- n y aph;j oouplad to a laadcro Mm Spar t i -- tiki P i t ta (LOM3/M3) in mpt irinH- LO & ^ty LI Ahnya h i m ato labwdpej prato L 2 O -- iO toe p p a y ito c M 3D 8 h to b ca la c d fa if my cO oetoal fo r p ro p e r w fe ty XD S w pla tu to r -- 1.1 asicc 2Q$fwiNnpii tor -- doo. 1 2 S o ffr i toc^ U p roc-- d btobretrawfeo. Placa the f a n m npk ia to ti j* m e r e itv|iito p B wito * y ice. Ptoo dw uieptoe i* eeeduner* to to m papi fa -- * w p m o to to tom tor emtoa dioxide tomato.M h m Om . S to to ptoet too aanpk to t o o M n p mil turn o f 3.3 S M fh D toirtoB pro d ig w ill bo ipw t t ad to too lam pitaf to ** ...... * . 4.0 Raafato M Staedatoi 4.1 w * - a n c ^ e d * 4.2 Acalentriie-HPLCpto J Cboo (120-4QQ iato ) - 1 gadc 4.4 Metowel - HPLC yada 4 J filila to <60-200 -- to ) - Ramato ptoa 44 FliHrttfO-itO--ab)-IUaf-- pad 4.7 N y - cf-- lO M H i - t r a tto ! ptd$ 4 J lO c tta to - HPLC trade 4 L-A fliabw asd - Roopmt prao 4.30 DiliOgrltocMnniil-- - O i i f i pad O l ToIm m - Raapoat pad 4.12 A-- to-- Ac-- - A X J . R a if* Prado 4.13 Ptoto-- -- i ,Ritolto 31 b-- a to Byop-- X I A tofh ; to-- am B fdd t o a ii p if l rapali o f p-- p to f op a 2 n l a to ajwppad w ito rm bt r t U m h jtc to r e n able o f i^ c c tn f 5*200 p t -- i -- lio to to -- I f -- tpooO-- a O C W lto . X2 A d ito c i to a-- -- d-- farpoto Intog tea to partoarine 5J Aaljtoaaiba-- acapaMa o fa a d to f C.00001 9. Hm2*n P ag* 46 o f 65 Exygen Research Page 87 of 110 7nteriir Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 Btygjen Protocol Number: POOOl 131 **rt>*M*Tvgoo]7tf ______________________ A N ^ m C A L METHOD ^cthpd M fa. o/FwfoowedaoQic A4 (PPOA) m Yepettt* bjrLC/MS/MS 5.4 J U ttry m p c rm r. 5.3 s5,t7 it>rj2-5mnrL p--MM^b^raf'dtrfyHrfartdtfr.y1-h-ii-t'-ft-ffliftit U D iip iiilli te -- jjp te fffr lO O e l, !>0>200eL)c 5.4 l 2 * a i U B m**w-wm*hhottkm. I l f JurK FLC vM kit 111 Dhpo n M * p ipette. I K A M p^M ()O 0>lW 0pL ai4l0']C 0^itk4M pou6Jetip. 113 rPBtatee (2Qm LJ (Iqpefe* eel. *&. MQ57177). 1 H V rirfeaite abater. 3 .i> C te rk ifll eapafek o fapinnlpp 30 iL poiyjnjpytea tnbe a*3000 ipn f > n . alyteel Chlin r H f fc-- HP (Kayttcm ScimtiOcX 2.1 m > a 50 ran*. 3p (P*fc 505-0531*) T ta p & M iK 3" C u Mobile m m (A ): le M /M o o a m A te ftio W M e M : Hw teeil 3.5 O n d iw N p n a . H a tia ta i M 1.0 LO 20.0 3L3 *A 33 69 23 25 33 FlewlUae Z M UbU b ib } 33 0.3 33 cu 73 o .j 73 OJ 35 0.3 3.3 Inaction Velum : 13 pL (cee bebicruaad B>at tm eftae 50 p i *,V Qi--ititerjn Peak A m -H em al iteadard aettbretipe aerve. 6J te n T te *-2 3 te n *m T te atew eotettiooi an imancM as i pude ami s *y be dianjed in order to apte tw ite feflC yetea. U M W M jb a 7A Mad* Elergem *XafCtva fcfltM mode, moaUo/tei 413 -369 VZ fet PFOA. Page47cf6S Exygen Research Page 88 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Exyjea Protocol N m tb tr . P000I131 I - AWA^mCAl MITMOB 1 W Aj i It m t t * Oiw npim o o fi m tlmxooa m m A m i (?TOA) in V caam ^ T ...............kyLO M /M S T h a itim n--i lili i i k Inamil al aa | i iili u n i im j in i ili- n iiii hi m ill i n 1.0 P i y i a i ofTlntiim 1 1 M M in 11.1 l a M --waa acstaa m m m la p ra p a il by a rtirg 0.1)4 I o f I acetote to 1000 w L o fn u r . t t .1 . 12J u U fc n O o L *fs M 3CH D um O vktkU * ^dP M ^yttoU oroil ttfld f r yt ip f i d by dfraolvTog 2 $ o f --oortnc law# h leheo p n p e i* by b h u b is 3 mL p 1 fla il vofcMM o f 10 n L w ith to to n * . AJn f.l HI t.L 2 dU . 9.1A 91-3 h * ^ *a io c fc ta liM O f~ IO Q ji ta L o frP Q A b y * ia f 10 ilw iw t fcemaied fc r pwriry) wW *le t 1 1 inL w i* jm Om m I in 1 123-oUL LDPB botfift. A 1.0 p fftn L ibnUWtoto w fcbon o f PPOA k prmpmd by briAgutf I n L i t 100 ji<b L aototM* to a te a l votom o f 100 wKk nwrtMoJ i&A L2JLU >PBboote, A O il njrtw L fc n ila rtc i cbif lw fH F Q A ttpfptosO bybriw ftof l l toL fdw 10 ttfta L tofeJttail to l final vofaaue o f 100 w ith aacttattl m t 123 (. ID ffl bMOfc A W A flfeL Ifc rtiU M iif --kid aa o f F fO A it ptepflrtd by brm*m* 10 a l. * f a i (i^ n L astoriaa to a Am i vote* o f tQO vkh M lD a l tf.u n ta a ]ft 1 b **B d ra d fe 4 fle a tio *ttte tin ia to b ' io p 'd a n 6ifa a r ^ e K w ite ty 4 T e d are seM i 6r * maiBMaa period a f 6 w a tlu h a i iO ln f jiii|iiiilh ii. 9-2 f t a M C M i t i M a i m calibCTdoo O wtodb r> w ~ o fte lD M ta L M lS e a tic B a o lM tto a t a a wflMwcJ m d ik tio r Pug* *6 o f 65 Exygen Research Page 89 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 E xyg e n P ro to c o l M uxnben POOO] )3 t M itiwd o f A a rtjd a ta d AN ALYTIC AL M fcTttOO o f: b? LOM3A4S Milked Nwwbw VtXM lTX A n d (P P O A )io Vt|o<atMn U J Th Mowing 1 tpptaal XMBSdK addiriocal cancaniou m*y be *a j, -Jiy. .. . \* . \Jt. . C4S OXB* 41. - W - _____ 10y0 ______ - " S r -- LS 1D0 14 100 .01 10 100 1005 10 100 tm ii 10 100 a jo i J , IN OXIOOS 12.2 S ta n d ] e e d w N id 12 5 L LOPE MfTO*MI r c c s*c. up ix b U 4 o f tapferta way bf prepared 10.0 Batch St Up .1*1 Ek * M d , o f au^iaa m a d fypfcay 20 or M ) sutt tnriudfc ar Km mm uaimNMl oootrei and twa im m alid conoofe fom *d u known - oaaNatratiant 0abcontrol jptke) io verify prroaduMl recovery ft*r du huch. i0 4 haqumaqdo dar M d and tahonaofy dqd*coi and *itaa wdl ha apaoTito IndM quality verarme plan Ihr ttta priced. ILO Saspla Bxdractian (LJ IU M 1J W a |b 3 I Of t a a u n p ia hfto 3* UL polypiopytnne caUrifuge tubei (ibetify at aaadad, repboe lld and ix w *$ , Add 30 L a faaatoaitttia ad A nte n aw *i*t etwa taker fo r-15 rmauus. CmOifcga tha 50 n L pafypopytaat ta te t c w ta d f aampk u - 2CC0 nun * 1L4 IU -v ^ #. 114 IL7 fa & M e a n d h ica fca8?F*fceam d atom m thepcar*ahapad fades. Faak da 29 L SFE tubca b H-- w v A 3 | floriaiL 3 g afcea fd . 2 g M ta h j tad 1 1 IC-N*. Condition dia eotam * w idi 20 m i o f m * a t L d m 20 m l o i aoalaeikita. Oiacard an waabta. Do im aow tha oobeu io Sibatn the 123 mL prf hipnrt ! k ib by m n | w ith the 30H . Rfraa d a flm kw idi lotwcnc ooce, felluwaa hy M h -o l (ttw a M aa). O iy dM teaks eompMely before aat, adM kpah M fcybim w idbatkia a p fo itwip-- Omans dar tm a m a oadfabaod >PE eowmo flnad bette * m w A * ' fta paar-obapad te te . Codaa tba im fe tha 123 L lmiaad pe-a*pe ab Nt ij w - r'agw 9 o f t ) Exygen Research Page 90 of 110 Interim Report #23 - Analysis of Sediment Samples'! ' Exygn Study No- : PQ001131 E x y fe s P rotocol N w ito e r POCOl 131 I - - - ----------- - A ^ U T t C M M E T t l Q | MatoodWAeafrtfta4t>a<amfrMdqnafPatfVMtoocTanric A c ^(P ? ^A ) h Vagatacor ..................... ................ ... feLQM SM S 114 jM A ^ ^ o M iM H C a il9 te in p i io iB 5 0 B L M K T iA ^ K fe & 1 U M e d to ito ^ w ta H O e to B to < M *ila *e c tfe a M i*r I I . Cm tr f t * toe 9 mL p c iS fte fjtm t to ta l c it a iin t tx r + h at <2000 rpm IM I I STS wfawn. CoUaat ita ala ts*a th* m Um wh the akm t to rn tta hmiI 11.12 114 11 .111 11.12 A M 5-4 tan wt M to to ta ll m to* ] tto forato to w taci d pmiMw a a f a wtory g* if ow tor (to < 40*C>. 1 U 4 W ltatatiieltotoi.hr lid ia s 2 * L o f 2% toeoibicscto to toeiw D i to 11:15 T ta to ta ta toOtota to v U i ta g d i^ to a M p ifta . Mrntrm i i ^ I i i u t ii m x m n * , U U M IH B. k4W H aM M of K k t a M , n s p la ao hniflM M<npU > Ik. L C A tM > )< M . A lA a la a ta M mum f a a t t m CoHow .It d u ly djawpha ofPfOA wBMpoBtiat to< tom ftm or n>ottooocwmoti kvcto iatoludad h analytical wL A * cadre a o f extnwaed a k h n iia a atananLa anal be iacJoda at lha t f t a t a f m l a iito to d o f u n $ k w t, Extm lad sttotianfe on* to t a t a j u it it a dta ta tiy M l t it ip lt i A l m i -- t iie , aedm at o f n o to i oaWtaoa itMdjgda n ty ta injected at the ta fira to f i f a m i fcftnewd Vjr hartad caibaatl aa lOwnlai It itottoapftiad evwy $ -11 la a y lc l (to iia w t ih r a m w ti aat o f whentod atatotarta). h r ita r caw. a n c to ataodiap attoaAf^a aaiaita dto Sim toad hat iaiactioa to e atosple act U to ta w fta tito i o n to fb r f v iM t o Lieaae atondatd cnavta aae p to to p d -S f h a p ily t by Hm k m w lo a t o n I f t watghtwn a fpeak am 2 *sta { M oaLyiu: 3.3 (cr $eraleatt 12J d e ta eel anirrT*f atoa Any aaawpha toe am aai ta a d ifd (totototo a ta tiita ta ia a r toda id tototiyecd. 15.1 CVa--h i ^ i pMl ticm a yeta a f t danjtotf to it 36 m u taae a p a n t e M tS to ffc The 411 -- i p o t nanwpondi to toe PPOA ion white lie * 'd itlfM ( X 9 iw i) N |M tii toa haveratoboo dioxide. ^ dT ' PmV SO o f15 Exygen Research Page 91 of 110 Interim Report # 2 3 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Exy|*o Protocol Number POOOIUI I mf n u li M M M M ito vot|7ta ______________ A A lA H C A l.M T H O O ________________________ iB S 3 n 5 5 ) iin k S t5 B 5 r o 5 f? S S S i S iS AoM (PPOA) m V*fe-atin b flM B M B 112 MWiMd IM O imm ani Piati* PPtM m Ovtia placar thm ihe LQQ. X . Maalc w a ttitc FFOA rn la te im m tk m 03 ypb, than a --w bttn wnpte M M te o b ttlte M id te tira te s to lte la -a a tra o n d . 133 JU co 'tea trfaa teo JipiteB aK iaB teA q pfeB pumi k b P w m KM 30H v f te ir b m vate*. V a c a M pte M b w taid* t e acevpteW M i. ite M tw iO fi r < u t e n t i te ro-acm cte. 13<4 Aay Mlihaiii s te r i fiate b i i adatte * ite r by M nt t e Hagt firur T t V te n e f r ite t e t e terateio n a f t e ctibcte curve. H iw r a , t e i t e mate af cafflm* utoted t e i oaid bm u te c d i--n*t cete 30%*fte I t e ubar fat e t e la Uyaciad. 1JJ Ite c a n ta te c a r t a t e (R) t e alitante carmi fa ta n te atte te 20,993 (R* 20-995). I f o te d o u m alta Ib i a tte di tim i l i t e beo a ffid a te 91 a te te te sa la te jjte inteunent ofw ntoa, ad thr t e a * or te te r m i a o f am pia* t e d i te m te y te 113 In d ir a i (te a fra te -- d n itin ia and m iglia a tte w t d rift o r* te a 4 % v ite * m teJytor-J il I f fa u s te da B cpeate dei Kant u rite b rin i m it e r i* u t sntat te raaotlyrad. HO C a lc ite -" 161 U a c te ifrjo y ia g a y id o lo euteufaaa ha m a te o f 9K>A faaad (jjm p s L te n d a ptafc m a ) --lag daa a n te ! curva (M m tr n g n a io t panem et fauteata fry r i* M as L jm o lb v a n i p ro fe ti*: FPOA f a t e <te*L) Ite 14 2 Ito ria riD aw rig an sa te laoa--t e ria a--oo--a fP I'QA t o t e * ffo iL t* te id te > P fO A h a te te ) * rffP A h s r iia rta D i fin a l to t,U iU i D fl n p la w te M tl) D f te la r by Webite it e votane waa Aluted, U naceaaary 14J f m a a p b r te fiW w ill b ia v i m o te o f FPOA pbor lo axwacrion. am a t Mtowiai p ia te to s t e n t e parconi recovoy. t e w d y ffW * [ tote aaaly**tote (te i) - te y to fasti in copavKiyi)| f |QC te m e t e Page 31 o f 6 i Exygen Research Page 92 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study N o.: PQOO1131 Exyfen Protocol Number POOOJ131 r ANALYTICAL METHOD . . .................. ... ............. M ** M otor VD0017M k A*M(FFOA) b Sm H -* _ftflaMMl Lhrtrby L O W M I *'* A M tjM T M tatridttr tpr : n, -* ` *.*v . .' v . trnm tm m ok 1W RnwnfcDrix ta d CtJtapB,PA 16M1 A* * * * * * ..VI 'v t A C _ J lL fm irm M tf 1 Tijftwjud Ud*r,1C XI BxytesIn iw ii DM j& M ' ------------------------------- / V ie t FwpdM, (> m . h | y a ItMMfck _____ DM j i .. . . T m lfa p * : 7 P a tti! of6 i Exygen Research Page 93 of 110 Interim Report #23 Analysis of Sediment Samples Exygen Study N o.: P0001131 Exygen Protocol Number POOO1131 bnalM ck M g M M Vi*ll7tS a is a a . 1 T T C A i M E T H O D Midw*WAalehito w SwI^sawwiieB elHdlooiwetiwk Aeid (>FOA) ia Smdl Mammal U w by LOUS/HS 1 I.0 Scspa T a t anfad u tp b t wq laye t r *> iioli im md rpm Hiatio of pttfhmoicaaoc aci by lfigb IPwfi-- hi U*4 Q -- npiphy ceeplcc so M ann Mw gpaMFOB**Dwartar (LCiMBOdS) ia aaefl m i m ] S v . t y a h a m ya i fa r pro p e r sa fe ty UO 3.i At toat* >g ofHat tnpfe kr txtnetim . J2 " flapplai Hsottldbaptoctn ad befere mebon. Pface tbt h m nmpk *mj fail poewoe m d faw pafae wrifcdry Ipa. Hape A> u n fa w comiiism afa iiwftr pur ' f a w m n ji veatifat to aftow for aibcm dfasfa --MfafacH. t a t HM pot 4M Hfafa in ftaem nortf* nnl iww f m ytit. AHansafay, ifA m ii m fattlNcfait mooum of sample (-lau tfaer 5 # fa u o p o e fa itf iew y hMtfa sample c n be wsd MrupplMrl 3J Sm pit Ofactiat pceaaJum wil St epedfM in Ae mptotf plan fa fa* PWJfa' 4,0 UmpHW w tO m ufafa Al W far-H H T paia 42 Mifhi il-HTLCyad U AiHnMhtli-HFLCjwfa A4 U m n --n f a n s - A.CJ. K n e w Onfa iS> 1 1 A hrjfa iw iiw e t fa tid cferanatapapli capfah a{ pum ptnj w 2 m liftr oqmpped vMS a v ia b le y o k u t iajeetor sapablt # f injeotintf 3-2J pL caoMcaed to aM ates M m SpecttoawMr (LOMSA4S). 5-2 Adewieot*eii hw * H fa-peik faesntfai ad ^Hutitatioo. 5.3 Analytfad balancecapable ifiM db| l i 6.00001 p. M $0 dLf a y l l a paly^niyfaa llllllfll^tabw. S5 tS ail ^ e irtli palyp epylBM omrifluctabaa. 10 PHponlla H t f a i iOO^CQwU 5.7 123-enLLDPBaaaae moeHWH. U 2 mL dm HPLC ial Irit ?Q%* 5J q f6 * Exygen Research Page 94 o f 110 Interim Report #23 -A nalysis o f Sediment Samples Exygen Study No.: PC001131 ExyfttaPrtrtocol Number. POOOI13I _______ IM U M V O M W I . . ... A A u lv rtC A L A irrH O D ^ __________ U M l l i w b r L O U I M t 3.9 Ph p p i^ ip ip lB MO : Antapip#*## (100*1000 pU. m d >0-100 *L ), " tik OUpo#*]* Up#. 5.11 W ei*ipphfcVic<ee(HJ<niS?ec*toi* M 3 O T v m w M w itifc l SJ3 T iu n m ln r SJ4 WMt*#dMc ifcitar. 3.15 Cwmifcj <^xbkof ym ctof 13 mL poJyycyykitt oAx * 3000 rpm. 5.0 aeaewoenpOic S e tte r M Aad y u ril Cat-- a: FlappO-- t> (Kay-- Sat-- MkK lla w n S t mm. ( M t O S 4M 62I:) U T p p r t . 30*C 6J JUobiW f t -- (AJ 2 04 Aw o m A ttd k 14 U c k M U m <g): FV nrX m &4 * 1 . 6$ IS OJ 65 35 0J 25 75 0J 30.0 25 75 0J 215 65 35 OJ 64 ^ K tk a V o iM 15 pL (cm ba Im-- m4 to at -- cli m 50 jtLV 6.7 Q w etiw ica: FvakAxia-am sm Janndanl caliknbonpxv. 64 X n T q m - 23 1111 TWah-- madrtr>i ntmmdii p 6 Q i l --yfrt cimfcil inw tltri o u fttU P L C ij m l 74 W SMS! 7.1 t iM c l mU. <13 - . u ftfc . n i < may > chapped in art* Exygen Research P a # 54 o/6S Page 95 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Exy*en JnXocoJ Number POOOl131 4 y ^ e e H ie tir V I T i A .\/b L v n c A t in im fO D ; fcf tto O p tA to y d a fcrdwr ^WrmintifliofPw^ i'x i tmui Ad4(PfOA) m&mdt U rm b y LZ'SC J S 1J M i t J W 1 U 2 mM MOMMtMi Mite M acqr ia pwpiw i adding 0.154 g oi' MMMton aut 1000 mLofwtor. m y to p M ffmitoii iv^im<ii 9 ) . `U M torl T lrrtT n rl-flrT ln fi ^nlm i-a 9.1.1 Pn pan * MOck oiutwci rf -100 o f PFOA by weighing '0 *>% o fm ttjm itl p a M (ocwnat tot pMy} Md * U t io 100 mL wit RMdiaoi ta i 125hrL LDP* beto. 9J.2 A U fc rlifitttio a aoW oaofP fO A ifpropaw d Oy brm ftn| I L o tto 100 w teLioH tioB * <qb1 votuaie o f 100 w i* mcuimoi * ft 125L LDPE bock. 9.13 A a i pgtaL fo ft& rio n ta ti f PFQA i i prapM ) Oy k ia g n f 10 mL o t to 1.0 n f m L n h itio a to i Seal volume o f1OOwith nwftanot m * 125 I LDfl bettle 9J4 Tb*look Md JbctUJeati lolaooo n n k t stored ut Prtipntor u ^ p w to to ty iod an tobli fcr mexien pehod of 6 modtbi A m t o t o i e fy r^a m ie *- 93 Stotodl Catibnbo* SohtioM 9 2 J LCM B / M coUhnMi ttMdirrfc epnpared tu stopp! UQiteUsc o tto 9.1 jm L JbsbSoat* t o n t o 9 2 2 V m b to i m g typical m p lr t ddOoto coecomrodao may x fMpendaaneedcd- Dtocd -J f f -- -- - 190 24 ICO 20 110 1J 101 \x> - JjO 10 ,00 OJ 1* 10 100 12 ' Ml IO 0.1 * rc* rc, a oftoavtod* ey be prepared as H+4r> Pogt 55 o/65 Exygen Research Page 96 of 110 Interim Report #23 -A nalysis of Sediment Samples f Exygen Study No.: P001131 E x y g ra P ro to co l N u m b e r P 0001131 f ". - J . ! ----- .oaLyn eA i w rraoc 1<t4 <afAiitlTa W d w Dtiam fairinatfyttfliiaiTOittMit A a ld (T fO A )ii iU w rb rL O M S A H 1U M e l S a l* ' 14.1 ' CaMi ta *fc ( ia ^ J a t a k a M [lyp ica ly 30 ar lata) m m 1k M > a Im o h m |m M m M a d a m atad catxM a tortiflad k a * n doemUlWiflwi (h t partial y frr)l to verify paoordafil xccvary far itw. N fft 103 ................... ha i l d M am m y < 1 t4 iii m i a fa t "U b t tp tn lU t a t p M s r attaraat* aka torM * arejao. ll.o S m la fim iib _ , 1U HJ rj a i# 1 e fam pia (ata a 50 la l potyyaopjiam c tm n ttft M *a <ibWy ja a i lit T^ltn l a l t a aag] Hala 1 tttanm a waapMa flivtr mar 0 m ti ` ` i - " i ,* -- i* i - :----- if1i `[` f i n i d d w < a m ^ a fc rifc a la tt-- a flO iL . O K a a m i lan a li ia i|l a ia t n > t - l n a da -rj : 11.4 T r tttT r 't T - f - i i i'- ' ~n V i j 'i i 'I i m i a l ' i il ' " im > I. -- M apaOa. " 1 U A d 0 S L tfB B i a iO fltta 0 m a far-CO a ta la a c p t m ita artice dlafctr n . C ^ t ^ ^ t^ m m -y j d t ' II.? D m *e r n im 90 oL rttoorabir g a m ito * tube m i add 35 J -* V y -r. IL1 C M U t C IPE mrip (l t. # mL) by pmnof nL amhaoai fc liiin iU j Tin! iflT T I " M u 1 H |i1 n .i Pn III l 111 I ib ..... Ill V f t * iW iw e w v te o a c e id U m d C ^ ^ E c e im lg * . Dtocaitietom li.Hk Btm w tt-2 ^ L e f r n m e L Cefleci 2 L of d m too fndumd lS ^ y iijy o r,jie a c ea%itobr(flaalyotame-2ml . 1U 1 Aadym * * * * * --i r t m ^ w y LOM&MS 12J , O m m k y nd pw t< * and falto* ill trfP P O A o m p e A d fa g to a t le a d A v a r m a rt o o o M M n tw a le v e l* mm >edeluded tom meydcaJad. O J A a m e *ta fe e t n ii i*d im d e m w tb e o d a d a l6 *b s w n s d a toe and e f t seemte eet a m denfa m m be Mu pwn d baimea r m y 3-n> w n jid A i m dN nm hn, actira m i o f celfere*oq w d ir ii nay Im fajadwi d toa he$dato| o f a eel falto* ed by ceiibcdicw u n o n k f 5*10 mwpl ee(to acco e i far a amend m o f mndu d n la addde aiaet be fee fin t and fad iecaiea in a 1X4 Ifca J tor (b e a d y * by Itoaer am 1* o fpeek area a ^s r? P o fe itf c / J Exygen Research Page 97 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No. : P0001131 Exygen P rotocol N u m be r POOOl 13 J EtW lm - r t________ ______ ____________ _________ MtoaOtoto rV flM lm ; A M A LV U C a I. m k t w o c B8 3 BE^5CSBnS55C35rsy53E55iS 5 n i3IWS7!SS' tteaatf LtvaayLOM&MS *a p g a iM in i m a te * M acaontfot te n t M te L y a i JJ ( a a d ite m i n ta w a a y a a n . t t i S te tte m a l t e t e m i ncM d M a te * .............. Any aamfUcs lU m t m i a ar ia * napaia i at e w k t H a im te n te m i w i ly n t. 13. A n te a s CaMm l i t C lfte M p M anaetea a|aak (dautbna taa a S m m te a a y a rn f *tS aaj, t t a 41 a m m b -nnaapcnla t e TFOA m oa. hila t e *M (h M la * O f t te a ) K fM aaa te >Mt * f arta* Ma I l l Ma te * teak a n a ta t coniaa TTQA a Irrita y a te n Ua L o g . L' a M a t c a te n ! 7T0A k ite g li a te a 1* t e l. Baa a raw Mank ample M H tS ifM M d m i the tu tto set n u t be (M xhacad. tS j Aeeavame o f santol pikee od ra e rk ^D ok m u be between 70*130% o f te w hmnwi w liw 1 a eotoxd ^ke & & ouuide te jccoptabie lia iit*, to atto to t o f raspile ito ttiA b t M -exm atd. A *y a to m pte wrtwte XV 1)9)4 ik to ld be evaluated by te enelyat to dcteranac i f n ra tn c b o i i 13.4 Any ffik b n lk e i anted fe u d to be totoirbeal outlier by ueinf tbe Hu? B m r Ten, raay be enchila d fie toe eekittobee a f te cabbntiM ourve However. toe total w a te r o f ca&rtoM it t u t o l i f i* coat be ciudad aran o asead 30H o f toe toral ra b e e o f a o d ite bdocted 13J Tbe ceneltoke coefficient (A ) tor calibration corvee unerased muto be toXMQ (A* 2&9tS). I f rsHtettem raaults to ll autxide tocar lim ile, toen tofeopritoe epe ran be taken to aJimt te tru m m operette, end the B * * or toe retoviento of eenptoetoMttld be rtieoiyted. )M Rettorie* tonto betoni* to te * and H elp let a m era dob more toen 4 % wilbte. M enalytal nm. I f rateatimi bene drib caceede tins lim it whin m e ly ite ran te e (be etoam* be reacaJyted. 140 CM ratei-- 141 Ubetoe M M | i t e c i lete n ta le the aarnot o fPPOAtouto (ia e t* iL baeed to peek area) u k f too --rieri! curre (linear ra^ouicn parameter*) ftow ted bjr theb te s lyax Hftwve pofieai: Itotbk tonal f tm t o n - f f te -- .tot--mpA T toty* totter te p e DP toraceby wklcfc too toeel v o te wee dim ed. I f eeceeeery. A tenei te ta r* 10 Exygen Research Pa** S 7 o f6 S Page 98 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 Ecyje Protocol Number P0001131 f l lHIU. . . ... - A K A IK I K -'A L MCTKOO | K*Ji>l'W ro t r lU p i Ml 111 O tofhOT. n iro fc A A ttffro A ) Smelt' . . . . H n llr ttrlO B K l U J P aarofki fa tfM w tarn m m u MrfPPO* priw to cxmctlon. o b o M M a ffq M iia i n k a iM i bojereau m every x w W : ..... J w W h lilr o k fta ftiU - tu to r f a t i i co H l(m lm L )| | t r I4 J U M io a to u t r o t a t i t o r n o i m o u n to fP T O A fa ti inni. rafH n o A O-- 1 ^ - n w f t A f c -- t / . ^ . t t , 0 .^ ,0 ----- L iv i M a w ttoto (o) -, ------- Exygen Research P t f i S t f 3 Page99ofll0 Interim Report #23 - Analysis of Sediment Samples Exygen Study No. : P0001131 Exygm Protocol Numlier P0001131 ANALYTICAL METHOD Ifa M N M fcr VC0017W M N M A - t f * for & D M m i M d M t f H r f M M M k KM tFTOA) hi Sm JI illim ^ lC M A iS Analytical M i r v M r ExySM KJMVC MM RaHvok MV* M > ( < > . PA 16*01 A ffK O td Bk te U L C _JtL ItllA fM Dm s iZ /p p ^ j/ SU *TM **! VT"S5frttiJ jfc . G ftrjifo . B x rfw Xaacwtb r * /' Dm Exygen Research Page 100 of 110 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 E x y fc a P ro to co l N u m be r P 0001131 _____________________ a w A iv n c A t w it m o c Mi t o d M A !* tm tm Dm iM in r.n f fM flw ro c a m k A rid CTTOA> m SxmuT U W H ! n i by L C /M M tt S o f . t . TIM a id u d w W --pl]n< fcr flu i-- m i -- imUta o f jx f lB a w a i rid by f i i f H r-- n-- a Liquid O y u m ry iy b y caspM t taadu. Uau t pwwnnniyDiliaw <LOMaoB)Bi-- U-- mill -- 2.0 gWMy 2J A h w y nM m a M h l-- ty pe-- -- . U Cl -- a -- i -- 3Q n8 k t n 1-- itlm m y rl-- in i fc r jto p n fc ir lift 3.1 U Ha n n yli yfno-- b n Ip -- d id fcr i -- ta aoKBin-- no---- tyt-- to rn U 3iw p it id -- in n iw i w fil b t --n jf liJ la tb * -- pda* plan fcr 4m U ) fa-- * 3 -- 4i n mi--u i-m e >id 4 3 ........ M il-Ill-H F IX f-- 4.4 Am -- d-- -- - A X .3 . fa -- 0 -- > 4J PwOn i l tlaioic Arid-S-- aAldridl iJ> k m rm m m U fd p -- 3.1 A biyk ya fc i-- I-- d m -- aupfc cip M l i o f puapias a K 2 ioh"-- -- f id -- r irid h li rata-- a iicie r t -- tU o f -- ciia 3-246 p L rn a a ira d i-- M i S a a ia a (LO M SM 53 j.j A ww# n t frf ytifc trttfnrtHn trrf nwmtiiartcm 5J A M ly in l ImUmih a 0-00001 5J 13 t l i ipMhV potyyopylcae W 5.< Im fifiW <30- 10O -teL). 3.7 125-mL LDPB Mnvw-ttoutk book*. 34 ZnLtiM rHPtCvidkiL 5j n ^ iiw lih p ip n ri 110 lnjiM ii <100-1000 p L s rf 10-100 n U * * 3.) I W M n O p h k V it6 ( l|) C ll3 r E ( M U ^ . 5.12 O if i- iM lO M , 5.13 Y n w . t* * P m ^iO o/65 Exygen Research Page 101 o f 110 Interim Report #23 - .Analysis o f Sediment Samples Exygen Study No.: P0001131 Kxygen H rotocot N u jn b e r POOOI131 I /H W V T IC A i. METHOD ] tfA -x ty ru tbc r*3 trm /u iK x ffw ftatn m m m ie Aod frrO A ) S ir^ l 5.14 vngMatoillwMr. 3.15 C--I n in i cip a b k o fIpwn4>t 1 5 n l,p ly; iF )4 M liit i m 3M 0 h i. ',j< * ri u a- -<i ' O eg^nripnpM o 9y$t*m .f fi .M 6J ; M ..* 3 Am k f m l CM-- F lw p b u i I f r QLwymn t t S n g m ic ), %i mm a SOa m . Sp ( m *23*5-05213*1 T m n t iw 30-C fcloW t FIh m (A ): 2 A M * is W ttt M pW te|taM (V>: M M G w iiiy r iy i^ 31 31 *4 B <5 55 <U u> 15 35 0J 25 75 03 2& J 25 73 03 ^ 22J 65 35 03 A f N |W < if Vl ht lC 15 p i ( *0 iDCTMMd to M OMCk M 50 ^ L I QaaMtatiae; Peek A m - rtm a l ti&dird n tib n to c eunw. 6.1 Am T1m k ~23jm m M . TheobevoM nttlM oO fobtfefidedM ifokleaM jm ey bechanged ie orderte 7.0 M S M S 4 9 M 7.1 M ole; B toftvneoy K osovo MRM owd, nw niM rini 413 - 369 n tz for _IfO L ' ^ __ T he above coaM om a o lo a d e d m p rid e lid m ay be c h a n p d in order to 0F b s l M S M S y to . U favenM eefS otetiene M Mobile Phow % \A l m H noiin m w i h wmm >o pw poead by adtag MS4 | o/ iTiowm aconOe to IQOOmi ofwoMr Exygen Research f tr je 61 o / dJ Page 102 of 110 interim Report #23 - Analysis o f SedidtoitSamples 0 ' Exygen Study No:: POOO1131 E xygftn P rotocol N u m b e r POOO113} MtowdNuterVOPOink . A im y n c A L m m w o H eflwf o fA--frrie fc r fcaD am afcatieii t f Pit f t ii iooettpofc Aa4(PPO A) in Small Mo m I S a ra by LC/MSA4S 9jQ CwtoeNfnpKado* 9.1 fciafcwi Sfcck4apflcaon S a ^ e c 9.1.1 Prpp-- awpek e&laOQof--ICO pftaL efPFOA by waijftm* 10 mg of wwlyUrd rtaadato [eoeraeted fcr parity) mA dilute id 100 mL with tnartamrt 1 125-at LDPE botflo. 9 .IJ "A l-O pytaL ftitiflaaiienat*lM H t>f?PO A It prepared by brioting ; mL o f ft 108 ppM * eaialwa ID a float vol-- e o f 100 with a**mvi id i LISmL LDR] boOfc. f .U A 0.1 njhad. fertLScako# iotMk> o f PPQA. u prepared by On*j f I* a L o ftte 1.0M M . tu d o a float voioconO O w M iM ttiao!i 125LLDMWfc. 9.1.4 T lie ia d c n iJ b tttik a iM ia lo )e o a a n *b fto e d fa *i ftif*n e rK lyjam-- Ij 4*C wri ira liable f a a nmnvm panod of 6 rworJ* h o fltd a n ffn p a ilQ n , 9j S H a M ra llriiio a SoM m m 9 a 1 LCM SM S cafibnuiM a c ria i* an p n p * to methanol via ettunon o fthe Oil jkgmL flrtflc O M . K | a ln 0i a The M w h | b a typical c u n p te a ttM cft*] (M ta to ja a f may be (n p a e fo a n M . Ftoal n S E ta l Yah. M uetfto H X ntaW .) M J tP* J (n x a U IN 54 IM 54 toO , . 10 w -i 14 24 >ttoo4 100 1W K* 100 24 1-0 05 4 14 100 .i 113 f t -- tol^abbnbo*. ttartoerde 135-mL LDPB MwcnT a*cmh bottle* tf2 *C to ^ C ro to m n o n **. may baprepaoC u Seech SatUp m flo fc b a ta h ^ -- p i* eatocwd (Typfcafty SO of le ) ro w iocM e tea roe M a M rewrol aai two irw e J eoatofe t e m lM a known ira o -- flcaeQ ft eeoW part vw ify pei r aduwl recovery far die batch M a , flip ftm m fi far CaM tad lalom ury dapifcatot end ipakn w ill be speeded fcatpu tity a-- nee pfco fcr (Mepeoyeet. 4W ? P tg t 62 o f 65 Exygen Research Page 103 o f l 10 Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: P0001131 B xyje a Protocol N a cie r P 0001131 an i iw n I " * A tU L Y nC A L M flH O P H ffe te A nt r i O r ti Dt t wmma i-- r f M w w t t w ic Add (W P A ) in S ia ti M ttn d Sem by LC/M3/M5 ] 11.0 S a w lt a n n e tta : t u . M-- n t L o fpie ! 1 petnnpyfc ceoPiAip* id m (fivnify n t m h i nph n M a i l IQ . H o iH m nw rchnees o( w n m a o jfb * M n d p n ilio f Ib rjM a p te rjie m rk O ie fc rtiK H.S A M a m fc fto M n p lo fe ri& B le o h a to f !. C pptipM y J O V e rt l t i - l o e e IW Tleeer ! mL t t tm p ie ip p * P o > l plpeae i* o e 15 t filp irillW j fiM T fllft Alb*. IU - M i edorfeeteietofeafid fatte fee ^ jfiN u M a o e ri H K ii toter . fc .llA C te te iq p |U w te s *fc3000rpaifer-4 n u u i . I I J Dees* tbe p r -- Mf teo t SO L iip m llt e a tn ft^ i take M i add 35 w L itvm m . a t U C te A k e C n ir B tW id |tiO * t L ) Vy pee*<*$ 10 L ethanol fc A i te k y J ttlfin L C te rfr-2 d rcp>tee). Denot le tcoton* pma<ky lt : * ic te f t r fTwple m condfaccd C>| S?E c trid je . D tecfttetfM *IL IO Hte -1 mL * t mettuuwL C o lte i 2 ta t o f ehm to o jrteeette M jre p y te * coemAf tate Ctbui retane - 2 mL). IU * A firiy l-- ptci y rie f tto tttc ^ w y LC fld&ldS. >- . '- 17? O w r t efw pky 111 t e f if llt e n n i-- wir feeA M u te d . a ^ m < fcrfated -- pe m ftfc LM JO I Q m . A tbm k M a A d mm* pnoede sta (W ie* ad 112 11? 13,4 alW S lta d e rte rfT F Q A tW n ^ B n & n tu ta d ftr e o r tn o ra cap cem ra m * te re fa ta te in s ite te fe a n m ly tifa ita L A i es tri afcatibaoa um d tt s muet be MCtaJte it fee ata fs - r f i u n p l* set. Studente m u l te isteepened between every 5-14 l iw pl A i M ta rn te n , a* M in M l o f Befifarttioa tenderli nsey be H * M i C UM hniw rtng W I M foflowcd fcy caJibretx>* tUftdanU w tcapvvte every 5-10 aaflfke fio aocowt fi* I cond m i o f uMtaria). In ter c m l cnlibncm. stnted a must be (he firn u Inat in^eatoo a m Mopom. TJa* Sm v tenderti auree h r quantitation. Linear standard curve art jfia a r tte l te r Ite u a ty te b y Bnaar r p y te a n nso>g Ir i. m ite n i o f peak w i venne cafifenden stendine concatenatela i m i M neLyiu 3.3 (et cqwrelcM) n f t x i* statere,., . MTj5Me *M dd eoe ce te tee ie ri rcafoneee. Any i -- p lrr IM Mioete fim k ifd ieepeoeeheoklbe AnUMrdUeete f d f f el>t d. Exygen Research Pag* 63 af65 Page 104 of 110 interim Report #23 - Analysis of Sediment Samples Exygen StudyNo.: P0001131 Exjrjca Pmcoral Numba: raw 1131 ....................... M M 4 tente r VOW 1 A W A i-m cA L M rra p p _________________ > 4 ^ 4 o f I h i & t t i r w i n i i i i o f^ r lm n i n n i i A o d(rT O A ) A 5 w .ll ky L O M & M 3 13.0 > 1 >1 n O il III 1 1 1 C lm m A y w i O M * r > u tk o f u t n n m a M * n tram i ' mC4 l3 M O L T b e 4 1 ) m i p n o l iu ij--| * iu i| to t e PP O A m u c h , w J u it dur te ^ n a r t e ( 3 ft ^ le g m a n * t e k>ano f carbon r a te . 1 3 3 M a te d M ateu n u t n o t c a n to n F F Q A v In rto e gnamr ta n tW L Q . If U o fe r B tw `u ' F fO A to M A m 10 a g riJ iL te n a m v b ln r* im tp to in H tb e tM M n l M l te n te r a al b o m bn ra-antraoed. 1 3 3 Kacovtetoa o f w a te rs rm m i rnumtx tfkm max be te w -- 7 0 -1 3 0 o f - Ib a irlte w n rn to ne. t f a t e I y t e fc lb o n tii do tw aocepm bto tin te . tha --' te i 'in - f a ijiin a tfconSrf be re tr a c te A mymmaria y b o n ts d e 10- ***. I30H iftoW tf 6e v tn n tf by Km wtoyw to iTaMmim if re-tertr*ct*on it IM Amy o liM w ite to rri te n d M b * otite rai e n te r by nates te Huge B M T ? te n a y b aatetadod ta ra the r a irte rin n 1 t e ca JA rstto n s r*e Hnwnm; te a M te r o f eaftkB &n M d irfc te c could bn exchtoed caaaraM m aned 3QN t e t e te a l o u te r o f a n te d * te a c te d . 1 3 3 T I * n W M ooar tN am (1 ) f t * nnM ra d o n o w w j generated ra *j*t be 9AJ93 <*> M J K ). I f o tA n d o * m a l M * llw K u . a apprapd? M g M O I be taken to a d yn t kM b a n cn t o p e ra tio n , and the r d w n iM M to f a ra p ie i A c u te te m M ty z o d , 1 3 3 R a ta n te Omm a n te d * a n t srcpiec m a t not d rift mart te u , 1 4 H w ite i m m n lj r ic ii r t tfm a n tte mm d rift w m a A a h m n w kb te m mmtfkml mmte tfeo M m a t be m m alyzed- 1 4 ft r rto a fln m 141 Ueatbe AAewtef ofPfOA te te (a aprin-L x D F x a liq u o t facm r DP - te w by te c b te te a l n M ne to te * i f aeeaaaary. A Jkp M fiK to f-iO * 1 4 3 P v m a p lie r it in * rid i know n m a im a iP K )A peu to e x tra c tio n , ac te te in a r tM i rpi te rrn to c a k te to t e p a rc a n f oeovexy, T naan y O q _______________ -- W i- M w i^ a i_________________________ N r s in P v t* 6 4 < tfii Exygen Research Page 105 o f 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: POOO1131 B xyge P ro to co l M u n ta c r P 0 0 0 1 1 3 ] lo w m n __________ i. A>Ai.rncAi. wrraoo '* T 5 S 5 'cf A w lyiity teO M x iiifcM d jeam rilucn oc m o Agd(PTOAl jS m U U a m l S o n hy LGWMrfS MS loStwii f i p rtw c--vn 6m mmamt o f PPOa kmo4 io nymL m Exygcn Research h a > ra n f a t e 65 o f 65 Page 106 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: P0001131 & Q II'bR E S E AR C H 3054 Research Drive Phone: 814-272-1039 S ta ti C o le te , PA 16801 Fax: >14-231-1580 PROTOCOL AMENDMENT Amendment Number 1 fW *c tt^ D M r,. .Q lflg fiS ,, Exypsn Study Number P0Q0113T O tart Study N u m b er, Pegel o ft Nana P E S C ftTfifol P f AMENDED 1) Analytical Procedure Summary V0001780ec8on 9.1 2 ) Verification o f Anatytcai Procedure AMENDED TO 1) Add to Section 9,1: Section 8.1.8, AftemelBweigftta of standards may be ueed to prepare alternate concentration! o f slock solutions as necessary. Altem ete levels of fortlllcation solutions may afro be prepared. 2 ) Low and Ngh spMng levels o f the anaiytee for each matrix m ay be altered depending on sample size available for extraction andfer to cower analyte concentrations expected in the samples. RATIONALE--------------------------------------------1 ) Higher concentrations of standards need to be prepared in older to spite the sample botttoe at Ngher levels. 2 ) The sample s in avalabte tor small mammal h e r and s e rin twee im adar than expected. SpMng at the pradetonrthed Must In the protocol puts the spited concentration lower than ie detection fcnit Aleo, the analyte levels In the ground water samplaaara expected to greatly exceed the pre-deteimlned spiting levels feted In the protocol. When the levels In fta samples g raaly exceed t ie spBdng levels, an accurate recovery value cannot be calculated for the QC sample. H ghar spiting levels h the botttee w l oowor 1he analyte concentrations expected In the w ater samples. im pact p w a n a The LOQ le 100 ngtp for a 0.1 gsam ple of sm al mammal Buer and Is 1000 ngtonL for a 0.01 mL sample o f am ai mammal serum. Higher levels of spldng for the water samples WB ensure that more Q C raoovery data can bo used. LIBRARY DtW S00122s4` . Exygen QAU ReviewJ jL L f lik k ADWreSTRATTVE FORM Exygen Research Page 107 of 110 Interim Report #23 - Analysis of Sediment Samples Exygen Study No.: POOOl 131 Am endment Num ber Effective Date: ~ Exygen Study Num ber 305* Rescindi Ortwe Phone: 14-272-1039 State Cottage, PA 16801 Fc 814-231*1580 H' PROTOCOL AMENDMENT 2 (&7/P5 " P0QQ1131 ~ C fe n t Study N u m b er Pagai <*1 None .IX :I:if.',T. 1 ) Report, page 11 o f 65 - 2 ) Teet M aieital, pepe 6 of 88: PFOStnrerttonrnonSored 4 8 6 -> 90. AMENDED TO 1) Inetead Of one linei report, kdettim raparle vM L b efn ued/ 2 } P fO S transition mcnftorod may eleo be 489 -> 80. RATIONALE 1 ) Due to the eMceeeiv* e in s of tie data et, interini report wM be issued to alo w the cfient to receive data in a Umettar m e m * o tjn /ts **W 2 ) The A PI 4000 LCM S/M S systems detect lie 499 80 P F 0 8 transition wUh greater tensidvHy tie r* toe 490 -> 80 transWcn. tMPACT ON 5TUO Y t ) The d e n t writ be able to receive and review the date mora quiody. 2 ) The 498 -> 80 t a neBon can be detacied e tt i greeter eenaiavty; therefore, gMng better chromatography. O L IB R A R Y O c V C 0 0 1 2 Z M ; Exygen Research A D M IN IS T R A T IV E F O R M Page 108 of 110 Interim Report #23 - Analysis o f SedimenfSamplss Exygen Study No : P0001131 3058 Research Drive 1 Phone:814-272-1039 State College, PA 16801 Fax: 614-231-1580 DVIATION FORM S s n e o li X Project Specific Deviation ~ Facity Deviaon' Exygen Project# : P760/P1l3t -- -- -- ---- -- -- -- Ctent P ro je c t#: NA Date cooccurrence: 04/26X16 Deviation#: R eference#: 5 1-" J` - 064)76 Raoufatorv Driver: X _____ GMP GLP Other None Seeipte Description: Deviation Tvoe: lin d u d a V# for methods a n d S P s ) X Protocol . ,, . . Method VfoNA Notebook reference; NA - -- SOP Login#: LS008191 Container#: NA List#: _________NA SummaryelDevlehon: * - ... The three sediment sample In L0191 (C0172892 - CQ172884) were originally extracted using tie sediment method V0001782. Poor recoveries were obtained for P fOS, PFOA and ,JC PFOA. Because ofMe, the study sponsor requested the use of an atematfve extraction for these compounds, as follows: Direct Injection Method: Before the samples were weighed for foe extraction, they were mixed thoroughly by vigorously shaking the container. A one-gram portion of sediment was weighed into a 15-mi*terc#nlr9ufle lube for the extraction. Ten mMilHersdf 1% acetic acid in methanol was added to each sample. The samples were then shaken by hand, vbrtexed. and sonicated for thirty minutes. The samples were then centrifuged for -10 minutes' -3000 fpm. Each sample was analyzed by LC/MS/MS eiectrospray. Using this method acceptable data was obtahad-for PFOS, but the recoveries for PFOA and UC PFOA were stffl poor. Another aftemetfva method was foen used for PFOA and ,5C PFOA, ss foSows i -.4 4. Alternative 8PE Method: The samples that were prepared In 1% acetic acid for the direct injection method were used for tils extraction. Flue-m*l*iers of each sample was alfcjuaied info a 50-mL polypropylene cenWuge tube and the volume was taken lo 40 mLwth water. The samples were then centrifuged for -10 minutes at -3000 fpm. The supernatant was then loaded onto a Cn SPE cartridge conditioned with 10 mL of mefoanol and 5 m l of water. The eluale was discarded. Approximately five m utters of methanol was added to the cartridge. Five m utters of eiuate was collected into a graduated 15 mL polypropylene centrifuge tube. Each sample was analyzed by LC/MS/MS eiectrospray. Cause: ___ Preparation___ Analysis____ Instrument____Chent Request X Other Im pact More usable data was obtained. LIBRARY ID: V000164CMJ hxygen Research ADMINISTRATIVE FORM l'ge UW ol 1 1U Interim Report #23 - Analysis o f Sediment Samples Exygen Study Ko.: P0001131 CHSH EH5R 2 3 6 1B 651 733 1958 c-ti-iai tmu? hwiutw tairriow STAUCH 0 5 /0 1 '0 6 1 3 :4 6 N 0.1 06 0 3 /0 3 m m r.jn/ra f*m * > : i. ^ * -I ,r f - . .S\ > * 1 ' " * ** B S B M I * --------BkUlow > * .. V -K :ii " j ' j 1 */>.r/ . * > . A: pajr ChfMBOi QuM yW vu UKW* r. J i fgHig * ^ //h ^B q p W tH lM ty . ,, ; ' *. ` . . . ; *V1. ' % * `` v '- j f ' 1 f .. ; /. t ' . .. . ' . / * . A * \* **. * *"*. \ V ^V > : > ' ., * . V -: V* - . ! : ' - ? v . ' ' ' ' ..? . . y * '*{> * {5* ' s : % V V* * ' v . \ > v.-f .. ? ' . ** '-{ T * ` .. ' ' - ", - V .A ''.s * V * i V mi * ' V>: * . * '.*u. 1. . <I * * ' . -A _ ** V . N l/t ' Exygen Research Page llOof 110 v Interim Report #23 - Analysis o f Sediment Samples Exygen Study No.: POOO1131 Exygen Research Page 110 of 110