Document npOgMk5O8njQOd0bGVQ3xLz9X

3M 3M Environmental Laboratory Project No. E05-0210 Interim Report #5: Analysis of West Morgan/East Lawrence Municipal Water Samples Study Title A n a l y s is o f p e r f l u o r o b u t a n e s u l f o n a t e (PFBS), P e r f l u o r o h e x a n e s u l f o n a t e (P F H S ), AND PERFLUOROOCTANESULFONATE (P F O S ) IN WATER, SOIL, SEDIMENT, FISH, C l a m s , V e g e t a t io n , S m a l l Ma m m a l L iv e r a n d S m a ll Ma m m a l S e r u m Us in g LC/MS/MS fo r th e 3M Decatur M o nito ring Program Data Requirement EPA TSCA Good Laboratory Practice Standards 40 CFR 792 Author Michelle D. Malinsky, Ph.D 3M Environmental Laboratory Interim Report Completion Date Date of signing Performing Laboratory 3M Environmental Laboratory Building 2 -3 E -0 9 935 Bush Ave. St. Paul, MN 55106 Project Identification E05-0210 E05-0210 Interim Report #5 Total Number of Pages 123 Page 1 o f 123 This page has been reserved for specific country requirements. 3M Environmental Laboratory Project No. E05-0210 E05-0210 Interim Report #5 Page 2 o f 123 3M Environmental Laboratory Project No. E05-0210 GLP Compliance Statement 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 Interim Study: Analysis of West Morgan/East Lawrence Municipal Water Samples Study Identification Number: E05-0210 This study was conducted in compliance with Toxic Substances Control Act (TSCA) Good Laboratory Practice (GLP) Standards, 40 CFR 792, with the exceptions listed below: ExceDtions to GLP comDliance: E05-0210 Interim Report #5 Page 3 o f 123 3M Environmental Laboratory Project No. E05-0210 Quality Assurance Statement 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 Interim Study: Analysis of West Morgan / East Lawrence Municipal Water Samples Interim Study Identification Number: E05-0210 This study was audited by the 3M Environmental Laboratory Quality Assurance Unit (QAU), as indicated in the following table. The findings were reported to the study director and laboratory management. May 6,2005 May 13,2005 & May 16,2005 In-Phase Data/Final Report Date Reported to M anagem ent Study Director 5-17-2005 5-17-2005 5-17-2005 5-17-2005 QAU Represe itive Date E05-0210 Interim Report #5 Page 4 o f 123 3M Environmental Laboratory Project No. E05-0210 Table of Contents G LP Com pliance S tatem ent..................................................................................................................................... 3 Q uality Assurance S tatem en t...................................................................................................................................4 T able of C ontents........................................................................................................................................................ 5 List of Tables..................................................................................................................................................................6 List of F ig u res............................................................................................................................................................... 7 Study Inform ation.........................................................................................................................................................8 Sum m ary and Introduction.........................................................................................................................................9 T est S am ples................................................................................................................................................................ 9 R eference Substances............................................................................................................................................. 10 Control Substance..................................................................................................................................................... 10 Method S um m aries................................................................................................................................................... 11 Sam ple C ollection.............................................................................................................................................. 11 Preparatory and Analytical M ethods............................................................................................................. 11 E xtractio n .............................................................................................................................................................. 11 Analysis................................................................................................................................................................. 11 Analytical R esults.......................................................................................................................................................12 C a lib ratio n .............................................................................................................................................................12 Limit of Quantitation (L O Q )...............................................................................................................................13 B lanks................................................................................................................................................................... 13 Solvent Blanks..................................................................................................................................................... 13 Method Blanks....................................................................................................................................................13 Trip Blanks........................................................................................................................................................... 14 System Suitability............................................................................................................................................... 14 Continuing Calibration....................................................................................................................................... 14 Lab Control Spikes (L C S s )...............................................................................................................................14 Sam ple D uplicates............................................................................................................................................. 15 E05-0210 Interim Report #5 Page 5 o f 123 3M Environmental Laboratory Project No. E05-0210 Surrogates......................................................................................................................................... 16 Field Matrix Spikes............................................................................................................................ 16 Data Summary......................................................................................................................................... 16 Statistical Methods and Calculations.......................................................................................................18 Accuracy and Precision Equations...................................................................................................18 Determination of Analytical Uncertainty............................................................................................ 18 Statement of Conclusion..........................................................................................................................19 List of Attachments...................................................................................................................................19 Signature Page........................................................................................................................................ 20 List of Tables Table 1. Sample Results Summary..........................................................................................................9 Table 2. Study Reference Substance..................................................................................................... 10 Table 3. Study Control Substance (Surrogate)....................................................................................... 10 Table 4. Sample Collection Information.................................................................................................. 11 Table 5. Instrument Parameters..............................................................................................................12 Table 6. Liquid Chromatography Gradient Program...............................................................................12 Table 7. Mass Transitions....................................................................................................................... 12 Table 8. Method Blank Area Counts.......................................................................................................13 Table 9. Lab Control Spike Results.................................................... 15 Table 10. Sample Results Location #1 : W M ELPW RW 01...................................................................17 Table 11. Sample Results Location #2: W M ELPW FW 01...................................................................17 Table 12. Sample Results: Trip Blank....................................................................................................18 E05-0210 Interim Report #5 Page 6 o f 123 3M Environmental Laboratory Project No. E05-0210 List of Figures Figure 1. Calibration Curve: Extracted PFBS Calibration Standards in ASTM Type I Water............. 22 Figure 2. Calibration Curve: Extracted PFHS Calibration Standards in ASTM Type I Water............ 23 Figure 3. Calibration Curve: Extracted PFOS Calibration Standards in ASTM Type I Water............. 24 Figure 4. Calibration Curve: ExtractedPFOA [1,213C] Calibration Standards in ASTM Type I Water. 25 Figure 5. Total Ion Chromatogram for PFBS: 0.0253 ng/mL Extracted Calibration Standard..............26 Figure 6. Total Ion Chromatogram for PFHS: 0.0263 ng/mL Extracted Calibration Standard.............. 27 Figure 7. Total Ion Chromatogram of PFOS: 0.0257 ng/mL Extracted Calibration Standard............... 28 Figure 8. Total Ion Chromatogram of PFOA [1 ,2 13C]: 0.0249 ng/mL Extracted Calibration Standard. 29 Figure 9. PFOS Total Ion Chromatogram: Representative Method Blank (050506a072; Day 2 Method Blank 3)....................................................................................................................................30 Figure 10. PFOS Total Ion Chromatogram: Representative Methanol Solvent Blank (o050506a052).31 F ig u re H . PFBS Total Ion Chromatogram: (Location #2; WMELPW FW 01, Sample ID82438, o050506a061)..........................................................................................................................32 Figure 12. PFHS Total Ion Chromatogram: (Location #2; WMEL PW FW 01, Sample ID82438, o050506a061)..........................................................................................................................33 Figure 13. PFOS Total Ion Chromatogram (Location #2; WMEL PW FW 01, Sample ID82438; o050506a061)..........................................................................................................................34 Figure 14. PFOA [1 ,2 13C] - Surrogate Chromatogram: (Location #2; WMEL PW FW 01, Sample ID82438, o050506a061)..........................................................................................................35 E05-0210 Interim Report #5 Page 7 o f 123 3M Environmental Laboratory Project No. E05-0210 Study Information Sponsor 3M Company Sponsor Representative Michael A. Santoro 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 W est Chester, PA 19380 Study Location Testing Facilities Exygen Research 3058 Research Drive State College, PA 16801 3M Environmental Laboratory 3M Building 2-3E-09 935 Bush Avenue St. Paul, MN 55144 Study Dates Study Initiation: 11/5/2004 Interim Experimental Initiation: May 4,2005 Interim Experimental Completion: May 8, 2005 Interim Study Completion: Location of Archives All original raw data, protocol, and analytical report have been archived at the 3M Environmental Laboratory according to 40 CFR Part 792. The test substance and analytical reference standard reserve samples are archived at the 3M Environmental Laboratory according to 40 CFR Part 792. E05-0210 Interim Report #5 Page 8 o f 123 3M Environmental Laboratory Project No. E05-0210 Summary and Introduction The 3M Environmental Laboratory extracted and analyzed municipal (raw and finished) water samples collected by Weston Solutions personnel on May 3, 2005 in Decatur, Alabama for 3M Environmental Laboratory project number E05-0210 (3M Decatur Fluorochemical GLP Monitoring Program). Water samples were analyzed for perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS), and perfluorooctanesulfonate (PFOS) using 3M Environmental Laboratory Method ETS 8-154.1 "Determination of Perfluorinated Acids, Alcohols, Amides, and Sulfonates in Water by Solid Phase Extractions and High Performance Liquid Chromatography/Mass Spectrometry" in accordance with Exygen Research Protocol P0001131 (Attachment C). The analytical start date for this interim report was May 4, 2005 and the analytical completion date was May 8, 2005. Sample containers were prepared at the 3M Environmental Laboratory. Sample containers for each sampling location included a field sample, field sample duplicate, low field spike (0.10 ng/mL) and high field spike (1.0 ng/mL). Each empty container was marked with a "fill to here" line and was fortified with a surrogate spike or an appropriate matrix spike solution containing the surrogate and the target analytes prior to being sent to the field for sample collection. 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. Quality control samples were used to establish the analytical uncertainty of the results presented below. Table 1. Sample Results Summary D escrip tio n 3 M L IM S S am ple Location ID Field Sample W MEL PW RW01 82756 Field Sample Dup W M E L P W RW01 82757 A veraa e C on cen tratio n W M E L P W R W 01 Field Sam ple W MEL PW FW01 82760 Field Sam ple Dup W M E L P W FW 01 82761 Average Concentration WMEL P W FW 01 mPFB S C oncentration (ng/m L) 0.0192 0.0210 0 .0 2 0 1 0.0263 0.0295 0.0279 mP F H S C oncentration (ng/m L) 0.0104 0.00990 0 .0 1 0 2 0.00955 0.00926 0.00940 <3>p f o s C oncentration (ng/m L) 0.116 0.0693 0 .0 9 2 6 0.0727 0.0782 0.0754 (1 ) Th e analytical uncertainty of the PFB S results is 10015% based on method accuracy and precision. The lower limit of quantitation (L O Q ) for PFB S is 0.0 050 6 ng/mL. (2 ) Th e analytical uncertainty of the P F H S results is 1009% based on method accuracy and precision. Th e lower limit of quantitation (L O Q ) for the samples is 0.0 052 5 ng/mL for P FH S . (3 ) Th e analytical uncertainty of the P FO S results is 10012% based on method accuracy and precision. The lower limit of quantitation (L O Q ) for the samples is 0 .0 103 ng/mL for P FO S . Test Samples W ater samples from two locations (3M LIMS ID # 82756-82766) were received on ice on May 4, 2005 from Tim Frlnak of Weston Solutions, Inc. A total of eleven containers were submitted for analysis. The samples were logged in by 3M Environmental Laboratory professional services personnel and placed in refrigerated storage after extraction on May 5, 2005. E05-0210 Interim Report #5 Page 9 o f 123 3M Environmental Laboratory Project No. E05-0210 Reference Substances Table 2 lists the pertinent information regarding the reference substance used for this study. Table 2. Study Reference Substance. R eference Substance PFBS PFHS PFOS Chemical Name Perfluorobutanesulfonate Perfiuorohexanesulfonate Perfluorooctanesulfonate Chemical Formula Identifier (1)Source Expiration Date Storage Conditions Chemical Lot Number TCR Number C4 F9S O 3 K* CAS #29420-49-3 3M 10/17/2006 Frozen 101 TCR-116 (99030-023) C6F13SO3 K* CAS # 3871-99-6 3M 10/18/2006 Frozen NB# 120067-69 TCR-83 (SE036) C bF 1 7 S 0 3 -K* CAS # 2795-39-3 3M 8/31/2006 Frozen 171 T C R -6 9 6 Physical Description White Powder White Powder White powder Purity 96.7% 98.6% 86.4% p)Solubility (1) 54,400 ppm No Information available Documentation regarding synthesis of the test substances is located at the source. 680 ppm (2 ) All test substances are believed to be soluble in w ater at the levels to be investigated. No visual precipitates observed. Control Substance Radiolabelled perfluorooctanoic acid (PFOA [1,2 13C]) was analyzed as a surrogate against a multi-level extracted calibration curve. A known amount of PFOA [1,2 13C] was spiked into each sample container in the laboratory prior to sample collection. Table 3 lists the pertinent information regarding the study control substance. Table 3. Study Control Substance (Surrogate). Reference Substance Chemical Name Chemical Formula Identifier Source Expiration Date Storage Conditions Chemical Lot Number TCR Number Physical Description Purity P F O A [1 ,2 13C] Perfluorooctanoic Acid C6F15[13C]F2[13C]OOH N/A Perkin Elmer 03/29/2009 Frozen 3507-195 TCR-744 White powder >97% E05-0210 Interim Report #5 Page 10 o f 123 3M Environmental Laboratory Project No. E05-0210 Method Summaries Sample Collection Samples were collected on May 3, 2005 in pre-rinsed NalgeneTM (low-density polyethylene) bottles prepared at the 3M Environmental Laboratory on May 2, 2005. Prior to sample collection, all bottles were spiked in the laboratory with a known volume of a surrogate solution and appropriate matrix spiking solution if applicable. Table 4 below details the samples collected and final spike concentrations added to each bottle. Table 4. Sample Collection Information F in al N o m in a l S pike C oncentration (ng/m L) 3 U L IU S Num ber S am ple D escrip tio n S am ple C om m ent V olum e P F O A [ 1 , 2 13C ] P F B S , P F H S , C ollected (m L) (S urrogate) PFOS 82756 W MEL PW RW01 0 050503 Location 1 Sam ple 200 0.050 NA 82757 W M E L P W R W 01 D P 05 0503 Location 1 Sam ple Duplicate 200 0.050 NA 82758 82759 82760 WMEL PW RW01 LS 050503 WMEL PW RW01HS 050503 WMEL PW FW01 0 050503 Location 1 Low Field Spike Location 1 High Field Spike Location 2 Sam ple 200 200 200 0.050 0.050 0.050 0.10 1.0 NA 82761 82762 82763 82764 W M E L P W FW 01 D P 050503 Location 2 Sam ple Duplicate WMEL PW FW01 LS 050503 Location 2 Low Spike WMEL PW FW01HS 050503 Location 2 High Spike W MEL W TRIP 0 050503 Trip Blank 200 200 200 200 0.050 0.050 0.050 0.050 NA 0.10 1.0 NA 82765 82766 W MEL W TRIP LS 050503 W MEL W TRIP HS 050503 Trip Blank Low Spike Trip Blank High Spike 200 200 0.050 0.050 0.10 1.0 Preparatory and Analytical Methods 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 Extractions and High Performance Liquid Chromatography/Mass Spectrometry". Briefly, approximately eighty mL of sample were loaded onto a pre-conditioned Waters C18 solid phase extraction (SPE) cartridge (Sep-Pak, 6 cc) using a Caliper Life Sciences Autotrace automated SPE workstation. The same workstation then used positive pressure to elute the loaded cartridges with 2 mL of methanol This extraction procedure concentrates the samples by a factor of forty. (Initial volume = 80 mL, final volume = 2 mL). As written, ETS-8-154.1 calls for a 40 mL and 5 mL extraction and elution volume, respectively. Analysis All sample and quality control extracts were analyzed for PFBS, PFHS, PFOS, and PFOA [1,2 13C] 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. E05-0210 interim Report #5 Page 11 o f 123 3M Environmental Laboratory Project No. E05-0210 Table 5. Instrument Parameters Instrument Name Liquid Chromatograph Guard column Analytical column Injection Volume Mass Spectrometer Electrode Ion Source Polarity Software ETSOIIIe Agilent 1100 Betasil C 1 8 2X 50, 5 um Betasil C 1 8 (2.1 mm X 100 mm), 5 urn 5uL Applied Biosystems API 4000 Q Trap Z-spray Turbo Spray Negative Analyst 1.4.1 Table 6. Liquid Chromatography Gradient Program S tep Num ber 0 1 2 3 4 5 T o tal Tim e (m in) 0 1.0 14.5 15.5 16.5 20.0 F lo w Rate (ftU m ln ) 300 300 300 300 300 300 Percent A (2 m M am m onium acetate) 80.0 80.0 10.0 10.0 80.0 80.0 Percent B (M ethanol) 20.0 20.0 90.0 90.0 20.0 20.0 Table 7. Mass Transitions A nalyte M ass Transition Q 1/Q 3 D w ell Tim e (m sec) <1)P F B S 299/80 100 299/99 100 (1)P FH S 399/130 100 399/99 399/80 100 100 (,'P F O S 499/130 100 499/99 100 499/80 100 P F O A M ,2 13C1 415/370 100 (1) Individual transitions were summed to produce a "total ion chromatogram" (TIC). The TIC was used for quantitation. A nalytical Results Calibration Calibration standards were prepared by spiking known amounts of stock solutions containing PFBS, PFHS, PFOS, and PFOA [1,2 13C] into 80 mL of ASTM type I water. Each spiked water standard was then extracted in the same manner as the collected samples. A total o fte n spiked standards ranging from 0.005 ng/mL to 2.5 ng/mL (nominal) were prepared. A quadratic, 1/x weighted, calibration curve was used to fit the data for each E05-0210 Interim Report #5 Page 12 o f 123 3M Environmental Laboratory Project No. E05-0210 analyte. The data was not forced through zero during the fitting process. Calculating the standard concentration using the peak area counts and the resultant calibration curve confirmed the accuracy of each curve point. Each extracted calibration standard used to generate the final calibration curve met the method calibration accuracy requirement of 10025%. Coefficients of determination (r2) were greater than 0.996 for all analytes. Limit of Quantitation (LOQ) The LOQ for this analysis, as defined in ETS-8-154.1, is the lowest non-zero calibration standard in the curve in which the area counts are twice those of the method blank(s). For PFOS, the LOQ was 0.0103 ng/mL as it was the lowest calibration standard to fulfill the area count requirement for all method blanks analyzed. For PFBS, PFHS, and PFOA [1,2 13C], the LOQs were 0.00506, 0.00525, and 0.00499 ng/mL, respectively. 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. Solvent Blanks Several methanol solvent blanks were analyzed to assess system contamination and/or instrument carryover. Analyte peak area counts in all blank samples were less than half the area counts of the calibration standard used to establish the LOQ. Method Blanks Over the two days in which samples, calibration standards, and quality control samples were extracted, several method blanks were prepared by loading 80 mL of ASTM Type I water onto a C18 SPE cartridge and eluting with 2 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 (glassware, SPE cartridges, etc.) Table 8 lists the area counts for the method blanks. Table 8. Method Blank Area Counts S am ple D escrip tio n D ay 1 Method Blank -1 Day 1 Method Blank -2 Day 1 Method Blank -3 Day 1 Method Blank -4 Day 1 Method Blank -5 Day 1 Method Blank -6 Day 1 Method Blank -7 Day 1 Method Blank -8 Day 2 Method Blank -1 Day 2 Method Blank -2 Day 2 Method Blank -3 Average Area Counts LOQ Area Count P F O A [ 1 , 2 13C ] A r e a P FB S A re a C ounts P FH S A rea C ounts P FO S A rea C ounts C ounts 5212 3285 2983 3110 2239 5510 3385 4119 1582 4036 2847 3483 81616 (0.00506 ng/mL) 5036 1352 1463 1603 984 5152 2132 5640 2931 2460 2761 2865 75010 (0.00525 ng/mL) 19444 17140 17746 15230 12305 20970 28896 37615 34842 19020 21958 22288 80616 (0.0103 ng/mL) 1907 793 670 485 572 1237 666 3283 1250 979 641 1135 25953 (0.00499 ng/mL) E05-0210 Interim Report #5 Page 13 o f 123 3M Environmental Laboratory Project No. E05-0210 Trip Blanks Prior to sample collection, one sample container was filled with 200 mL of ASTM Type I water, spiked with surrogate, sealed, and shipped to the sample collection site along with the empty containers. This sample was analyzed as the field/trip blank. 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. The resultant field blank concentration for all analytes was than the reported LOQ. System Suitability A 25 ng/mL solvent (unextracted) standard was analyzed in triplicate at the beginning and end of the analytical sequence to demonstrate overall system suitability. For PFBS, PFHS, and PFOS, the relative standard deviation (RSD) of the analyte peak area counts and the RSD of the peak retention times was less than 5% and 2%, respectively, for both opening and closing sets o f system suitability samples which met method criteria. For PFOA [1,2 13C], the opening set of system suitabilities met method criteria; however, the peak area RSD was 7.9% for closing set. A method deviation was written for this failure. 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 was still in control. All CCVs, for all analytes, produced recoveries within 25%, which met method criteria. Lab Control Spikes (LCSs) Replicate low (0.025-0.031 ng/mL nominal concentration) and high (1.25 ng/mL nominal concentration) lab control spikes were prepared and extracted each day samples or calibration standards were extracted. LCSs were prepared by spiking known amounts of the analytes into 80 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. Table 9 provides the LCS recovery results. LCSs were used to evaluate method accuracy and precision which was then used to determine analytical uncertainty for each of the analytes reported with the results in Table 1. Information on how the analytical uncertainty was determined is provided at the end of this report in the "Statistical Methods and Calculations" section. Two low level PFBS LCSs were outside the method acceptance criteria of 10025% (698% and 134% recovery). The PFBS LCS with 698% is considered an anomaly as only one spiking solution was used and the other analytes for that same sample produced acceptable results. Therefore, this value was excluded when the PFBS LCS statistics were calculated. The sample with 134% recovery was included in statistical calculations as its exceedance of method criteria was not as severe. When the high and low spikes are considered collectively for PFBS, the overall average recovery was 101% with a relative standard deviation (RSD) of 14%, which meets method acceptance criteria for accuracy and precision (10025%, RSD<15%). For PFHS, PFOS, and PFOA [1,2 13C], the LCSs show excellent accuracy and good precision, at both individual levels, and when considered collectively. E05-0210 Interim Report #5 Page 14 o f 123 3M Environmental Laboratory Project No. E05-0210 Table 9. Lab Control Spike Results. <1) PFBS m PFHS S am p le D escrip tio n LCS Low-1 (D ay 1) LCS Low-2 (Day 1) LCS Low-3 (Day 1) LCS Low-1 (Day 2) LCS Low-2 (Day 2) A verage Low (D ay 1 & Day2) LCS High-1 (D ay 1) LCS High-2 (Day 1) LCS High-1 (D ay 2 ) LCS High-2 (Day 2) A verag e H igh (D ay 1 8 . D a y 2) A verag e A ll S am ples S piked C oncentration (ng/m L) C alculated C oncentration (ng/m L) % Recovery S p iked C oncentration (ng/m L) C alculated C oncentration (ng/m L) % Recovery 0.0316 0.0316 0.0316 0.0253 0.0253 0.221 0.0307 0.0422 0.0240 0.0248 (2,6 9 8 97.2 134 95 98 0.0328 0.0328 0.0328 0.0263 0.0263 0.0343 0.0323 0.0382 0.0261 0.0248 105 98.5 116 99.4 94.3 1.26 1.26 1.26 1.26 106% 18% 1.32 1.22 1.15 1.16 104 96.3 90.9 92.1 1.31 1.31 1.31 1.31 103% 8.2% 1.37 1.32 1.35 1.35 105 101 103 103 95.8% 6.2% 10114% 103% 1.6% 1 0 3 5 .8 % (1>PFOS ID PFOA 11,2 13C] S am ple D escrip tio n S p iked C on cen tratio n (n g/m L) C alculated C oncentration (ng/m L) % Recovery S p iked C oncentration (ng/m L) C alculated C oncentration (ng/m L) % Recovery LCS Low-1 (Day 1) 0.0321 0.0294 91.7 0.0313 0.0291 93 LCS Low-2 (Day 1) 0.0321 0.0264 82.3 0.0313 0.0276 88.3 LCS Low-3 (Day 1) 0.0321 0.0338 105 0.0313 0.0365 116 LCS Low-1 (Day 2) 0.0257 0.0263 102 0.0249 0.0256 103 LCS Low-2 (Day 2) 0.0257 0.0235 91.4 0.0249 0.0236 94.8 A verage Low 1(D a v & D a y 2 ) 94.5% 9.6% 99.0% 11% LC S High-1 (D ay 1) 1.28 1.4 109 1.25 1.21 96.6 LCS High-2 (Day 1) 1.28 1.28 100 1.25 1.32 106 LCS High-1 (D ay 2 ) 1.28 1.26 98.7 1.25 1.23 98.4 LCS High-2 (Day 2 ) 1.28 1.13 88.5 1.25 1.14 91.3 A verag e H igh (D ay 1 8 D ay 2) 99.1% 8 .5 % 98.1 % 6.2% A verag e A ll S am ples 96.5% 8.9% 98 .6 8.7% (1) Table displays rounded values for all concentration and percent recovery values (3 significant figures). Recovery and RSD values may vary slightly from the values in the raw data. (2) Value excluded when calculating LCS statistics. Presumed laboratory anomaly. Sample Duplicates Because a field sample duplicate (separate container) was collected at each sampling location, dupllcate/replicate extractions of a given sample were not performed. Overall method precision was determined using surrogate spikes and LCSs. E05-0210 Interim Report #5 Page 15 o f 123 3M Environmental Laboratory Project No. E05-0210 Surrogates Although not specified in the ETS 8-154.1, PFOA [1,2 13C] was added to all samples and sample spikes as a surrogate to evaluate overall method performance. The final PFOA [1,2 13C] concentration was 0.0499 ng/mL. Surrogate recoveries are reported in the next section with sample data. Field Matrix Spikes Low (nominal concentration of 0.1 ng/mL) and high (nominal concentration 1.0 ng/mL) field matrix spikes were collected at each sampling point to verify that the analytical method is applicable to the collected matrix. Field matrix spike recoveries within method acceptance criteria of 10025% 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. Data Summary The tables below summarize the sample results for the two locations (Table 10: WMEL PW RW01; Table 11: WMEL PW FW01) as well as the Trip Blank (Table 12). An average concentration was calculated for the field sample and field sample duplicate. The relative percent difference (%RPD) of the two measurements is also provided. For Location #1 (WMEL PW RW01), both the low and high field spike recoveries for PFBS and PFHS met method criteria of 10025%. Reproducibility of the field sample and field sample duplicate (%RPD) for both of these analytes was less than 10%. Furthermore, the field sample, field sample duplicate, and field spikes produced acceptable surrogate recoveries. However, a low recovery was observed for the low field spike for PFOS (36.4%). This low recovery can be partially explained by the high RPD (50%) between the field sample and the field sample duplicate. Because LCS results for PFOS (Table 9) demonstrate excellent precision for PFOS, the discrepancy between the field sample and the field sample duplicate for Location #1 is attributed to variability in the collected matrix and not the analytical method. If the low field spike for PFOS is calculated using only the field duplicate concentration, then the field spike recovery becomes 59.2% which agrees well with the low PFOS field spike recovery (62.9%) observed for Location #2 (WMEL PW FW01). The high field spike at Location #1 for PFOS did meet method criteria with 87.3% recovery. Similar to Location #1, both the low and high field spikes for PFBS and PFHS and the high spike for PFOS produced acceptable recoveries for Location #2 (WMEL PW FW01). As mentioned previously, the low field spike recovery for PFOS was 62.9% which was outside the method acceptance criteria of 10025%. The low level PFOS field spike for the Trip Blank (Table 12) was 79.4%. Although this recovery technically meets method acceptance criteria, it is lower than that PFBS and PFHS recoveries for the same sample. This result may indicate a holding time issue with PFOS at the low end. If the low level Trip Blank recovery is used to correct the low level field spikes, the recoveries for PFOS increase to 74.7% and 79.2% for Locations #1 (WMEL PW RW01) and #2(WMEL PW FW01), respectively. Overall accuracy and precision of the surrogate (excluding Location #2 field duplicate) was 99.8% with a RSD of 16%. E05-0210 Interim Report #5 Page 16 o f 123 3M Environmental Laboratory Project No. E05-0210 Table 10. Sample Results Location #1: WMELPWRW01 <1)PFBS <1>PFHS D e s c rip tio n 3M LIMS ID DataFile C o n c e n tra tio n (ng/m L) % R ecovery Field Sample Field Dup Field Spike-Low Field Spike-High 82756 82757 82758 82759 0050506a057 0050506a058 0050506a059 0050506a060 0.0192 0.021 0.136 1.20 NA NA 114 116 A v e ra g e S a m p le C o n c e n tra tio n (nst/m L) % R P D 0 .0 2 0 1 1 9 % C o n c e n tra tio n (ng/m L) % Recovery 0.0104 NA 0.0099 NA 0.115 99.8 1.17 110 0.0102 4.9% (1,PFOS mPFOA 11,213C1 Surrogate D escrip tio n 3 M LIM S ID D ataFile C oncentration (ng/m L) % Recoverv C oncentration (ng/m L) % Recoverv Field Sample 82756 0050506a057 0.116 NA 0.0445 89.2 Field Dup 82757 0050506a058 0.0693 NA 0.0504 101 Field Spike-Low 82758 0050506a059 0.130 <2)3 6 .4 0.0485 97.2 Field Spike-High 82759 0050506a060 0.989 87.3 0.0485 97.2 A verag e S am p le C on cen tratio n (ng/m L) % R P D 0.0926 50% NA (1) Table displays rounded values for all concentration and percent recovery values (3 significant figures). Recovery and RPD values may vary slightly from the values in the raw data. (2) If only the field duplicate is used to calculate field spike recovery, the low P FOS spike recovery is 59.2% . If this value is then corrected for the trip blank (holding time) low spike recovery, the recovery is 74.7%. Table 11. Sample Results Location #2: WMELPWFW01 <1>PFBS <1)PFHS D escrip tio n 3M LIM S ID D ataFile C oncentration (ng/m L) % Recovery C oncentration (ng/m L) % Recovery S am p le Field Dup Field Spike-Low Field Spike-High 82760 82761 82762 82763 0050506a061 0050506a063 0050506a064 0050506a065 0.0263 0.0295 0.142 1.12 NA NA 113 108 0.00955 0.00926 0.113 1.05 NA NA 98.7 99.1 A verag e S am p le C on cen tratio n (n g/m L) % R P D 0.0279 11% 0.00940 3.1% (1)PFOS (1>PFOA [1 ,2 13C,' Surrogate D escrip tio n 3 M U M S ID D ataFile C oncentration (ng/m L) % Recovery C oncentration (ng/m L) % Recovery S am p le 82760 0050506a061 0.0727 NA 0.0504 101 Field Dup 82761 0050506a063 0.0782 NA 0.0134 2 6 .9 Field Spike-Low 82762 0050506a064 0.140 (3,62 .9 0.0672 135 Field Spike-High 82763 0050506a065 0.896 79.9 0.0596 120 A verag e S am p le C on cen tratio n (n g/m L) % R P D 0.0754 7.3% NA (1) Table displays rounded values for all concentration and percent recovery values (3 significant figures). Recovery and RPD values may vary slightly from the values In the raw data. (2) Surrogate result not included in overall statistics as it is not representative of the LCS data. Laboratory anomaly or artifact suspected. (3) PFO S low spike recovery = 79.2% when corrected for the recovery of the Trip Blank (holding time) low level matrix spike. E05-0210 Interim Report #5 Page 17 o f 123 3M Environmental Laboratory Project No. E05-0210 Table 12. Sample Results: Trip Blank (1)PFBS mPFHS D escriptio n m u m id Trip Blank Trip Blank Low Spike Trip Blank High Spike 82764 82765 82766 D ataFile C oncentration (ng/m L) % R ecovery C oncentration (ng/m L) `A R e c o v e r y 0050506a070 0050506a071 0050506a073 <0.00506 NA 0.102 101 0.913 90.2 ,1>PFOS <0.00525 NA 0.0985 93.8 1.03 98.1 <1)PFOA \1 .2 13C ' Surrogate D escrip tio n m u m id D ataFile C oncentration (ng/m L) % R ecovery C oncentration (ng/m L) "A R e co v ery Trip Blank 82764 0050506a070 <0.0103 NA 0.0416 83.4 Trip Blank Low Spike 82765 0050506a071 0.0816 79.4 0.0431 86.4 Trip Blank High Spike 82766 0050506a073 0.933 90.8 0.0439 88.0 (1) Table displays rounded values for all concentration and percent recovery values (3 significant figures). Recovery values may vary slightly from the values In the raw data. Statistical Methods and Calculations Statistical methods used to interpret sample results include averages and standard deviations. The Analyst software program calculated sample concentrations using resultant analyte peak areas and the established quadratic, 1/x weighted, calibration curve. None of the samples analyzed for this interim report required dilution. Sample calculations and equations used to report method accuracy and precision are described below. Accuracy and Precision Equations Calculated Concentration LCS/Surrogate Percent Recovery = Spike Concentration 100% _, S am p le _ Spike _ Recovery = -(iS-!pi-k--e--d--S--a--m--p--le---C--o--n--c--e--n--t-r-a--t-i-o--n----A--v--e--ra- 2g e- --C on--c-e--n--t-r--atio--n-:---F--i-e-l-d--S--a--m---p--le---a--n-d---F--ie--l-d--S--a--m---p--l-e--D--u--p--.-)- . .1n0n0oA/ Spike Concentraton , .. _ . . . . , standard deviation of replicates % RSD (Relative Standard Deviation) ---------------------------------------- ----------- * 100% replicate average , .. ,, , Absolute difference between sample duplicates . . nno. % R P D (Relative Percent Difference) ------------------------------------ --------------- r ~ r .-- - ---------- 100% average sample concentration Determination o f Analytical Uncertainty Both the accuracy (percent recovery) and precision (%RSD) of the lab control spikes are used to estimate the analytical uncertainty for a given analyte. For example, the overall accuracy and precision for PFOS based on LCS results was 96.5%8.9%. The measured precision (%RSD) is then used to determine the spread of the accuracy. Example: 96.5*(0.089) = 8.59. 96.5 +8.59 = 105; 96.5 - 8.59 = 87.91 E05-0210 Interim Report #5 Page 18 o f 123 3M Environmental Laboratory Project No. E05-0210 Thus, LCS accuracy results range from 87.9% to 105%. The absolute difference of the low and high ends of this range, when compared 100%, are then calculated. 100%-87.9% = 12.1% 105%-100 = 5%. The most conservative (largest) absolute difference is then used as the analytical uncertainty for the given analyte. Therefore, the analytical uncertainty for PFOS is given as 10012% for this set o f data. Statement of Conclusion Sample results listed in Table 1 are considered accurate within the analytical uncertainties listed. Spike recoveries for low level (0.100 ng/mL) PFOS field matrix spikes may indicate a holding time issue; however method accuracy and precision demonstrated by lab control spike data indicate the method is control and results for the two sampling locations are valid for all three analytes. List of Attachments Attachment A: Sample Chromatograms and Calibration Curves Attachment B: Extraction and Analytical Methods Attachment C: Protocol, Protocol Amendments and Deviations E05-0210 Interim Report #5 Page 19 o f 123 3M Environmental Laboratory Project No. E05-0210 Signature Page We certify thaj/this report is a true and complete representation of the data for this study: Jaisimha Kesari, P.E., D E E , Study Director S b Jo f Date 'S /z p / slocS William K. Reagen, Ph.D., Testing Facility Management Michael A. Santdfo, Sponsor Representative Date Date E05-0210 Interim Report #5 Page 20 o f 123 3M Environmental Laboratory Project No. E05-0210 Attachment A. Sample Chromatograms and Calibration Curves E05-0210 Interim Report #5 Page 21 o f 123 E05-0210 Interim Report #5 Figure 1. Calibration Curve: Extracted PFBS Calibration Standards in ASTM Type I Water. Concentration. ngSmL 3M Environmental Laboratory Project No. E05-0210 Page 22 o f 123 E05-0210 Interim Report #5 s)uno3 'eajv Figure 2. Calibration Curve: Extracted PFHS Calibration Standards in ASTM Type I Water. Concentration, ng/mL 3M Environmental Laboratory Project No. E05-0210 Page 23 o f 123 E05-0210 Interim Report #5 s)uno3 'eajy Figure 3. Calibration Curve: Extracted PFOS Calibration Standards in ASTM Type I Water. Concentration. ng/mL 3M Environmental Laboratory Project No. E05-0210 Page 24 o f 123 E05-0210 Interim Report #5 Figure 4. Calibration Curve: Extracted PFOA [1,2 13C] Calibration Standards in ASTM Type I Water. Concentration, ng/mL 3M Environmental Laboratory Project No. E05-0210 Page 25 o f 123 3M Environmental Laboratory Project No. E05-0210 Figure 5. Total Ion Chromatogram for PFBS: 0.0253 ng/mL Extracted Calibration Standard. |S a? 8* sss 300OO0000Q0Q0Q00QQ ' ` K8 n I 88 88 88 ` j sd3 `^|SU8|U| E05-0210 Interim Report #5 Page 26 o f 123 E05-0210 Interim Report #5 Figure 6. Total Ion Chromatogram for PFHS: 0.0263 ng/mL Extracted Calibration Standard. Time, min 3M Environmental Laboratory Project No. E05-0210 Page 27 o f 123 E05-0210 Interim Report #5 Figure 7. Total Ion Chromatogram of PFOS: 0.0257 ng/mL Extracted Calibration Standard. Page 28 o f 123 E05-0210 Interim Report #5 Figure 8. Total Ion Chromatogram of PFOA [1,2 13C]: 0.0249 ng/mL Extracted Calibration Standard. 0059500 13C PFOA 414.1! 369. (U iltitm n t) 414.3/363.9 a n - sample 11 o f t 5 from O tStSM a.w itr Time, min 3M Environmental Laboratory Project No. E05-0210 Page 29 o f 123 E05-0210 Interim Report #5 sdo 'A)|suaiu| Figure 9. PFOS Total Ion Chromatogram: Representative Method Blank (050506a072; Day 2 Method Blank 3) 05050i72 PFOS TIC (U nknow n) 49*.W it now ,43./M lnm n,49.m s.*m n tn m p tn n o t S trom O tS tS tS n .w lff Tim, min 3M Environmental Laboratory Project No. E05-0210 Page 30 o f 123 E05-0210 Interim Report #5 Figure 10. PFOS Total Ion Chromatogram: Representative Methanol Solvent Blank (o050506a052). 3M Environmental Laboratory Project No. E05-0210 Page 31 o f 123 E05-0210 Interim Report #5 Figure 11. PFBS Total Ion Chromatogram: (Location #2; WMEL PW FW 01, Sample ID82438, o050506a061). 3M Environmental Laboratory Project No. E05-0210 Page 32 o f 123 E05-0210 Interim Report #5 Figure 12. PFHS Total Ion Chromatogram: (Location #2; WNIEL PW FW 01, Sample ID82438, o050506a061). Time, min 3M Environmental Laboratory Project No. E05-0210 Page 33 o f 123 3M Environmental Laboratory Project No. E05-0210 Tim, min E05-0210 Interim Report #5 sch '/usuami Page 34 o f 123 3M Environmental Laboratory Project No. E05-0210 Tim*, min E05-0210 Interim Report #5 Page 35 o f 123 Attachment B. Extraction and Analytical Method 3M Environmental Laboratory Project No. E05-0210 E05-0210 Interim Report #5 Page 36 o f 123 3M Environmental Laboratory Project No. E05-0210 3M Environmental Laboratory Method Determination o f Perfiuorinated Acids, Alcohols, Amides, and Sulfonates In W ater B y Solid Phase Extraction and High Performance Liquid Chromatography/Mass Spectrometry Method Number: ETS-8-154.1 Adoption Date: 28 A p r 2000 Revision Date: 5 May, 2003 Effective Date: 5 May, 2003 Approved By: William K. Reagen Manager O S ^ > s -^ > 3 Date ETS-8-154.1 Page 1 of 17 Determination of Perfiuorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 37 o f 123 3M Environmental Laboratory Project No. E05-0210 1 Scope and Application This method was validated for the collection, extraction, and analytical procedurefor the determination of Perfluorooctane sulfonate (PFOS), Perfluoroodane sulfonylamide (FOSA), and Perfluoroodanoate (PFOA) in groundwater, surface water, and drinking water samples. This method may also be applied to the determination of other perfluorinated adds, alcohols, amides, and sulfonates in similar matrices, as long as the defined QC elements are satisfied and with the understanding thatthe method is not validated for compounds outside the scope of the original protocol This method is based in part on the report "Method of Analysisforthe Determination of Perfluoroodane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluoroodanoate (POAA) inWater"(see Sedion 17), as developed and validated by Exygen Research (formerly Centre Analytical Laboratories, Inc.). 2 Method Summary Water samples are collected from a site of interest and shipped cold to an analytical fadl'rty. Perfluorinated acids, alcohols, amides, and sulfonates are extraded from 40mL water samples using C18 solid phase extraction (SPE) cartridges. The compounds are eluted from the C18 cartridge, using methanol. Separation, identification, and measurement are accomplished by high performance liquid chromatography/ tandem mass spectrometry (HPLC/MS/MS) analysis. High performance liquid chromatography/mass spectrometry (HPLC/MS) may be used if the defined QC elements are satisfied. The concentration of each identified component is measured by comparing the MS response of the quantitation ion produced bythat compound to the MS response ofthe quantitation ion produced bythe same compound in an extracted calibration standard (external standard). 3 Definitions 3.1 Analytical Sample A portion of an extracted Laboratory Sample prepared for analysis. 3.2 Calibration Standard A solution prepared from the Working Standard (WS) and extracted according to this method. The calibration standard solutions are used to calibrate the instrument response with respect to analyte concentration. 3.3 Duplicate Sample (DS) A DS is a separate aliquot of a sample, taken inthe analytical laboratorythat is extracted and analyzed separately with identical procedures. Analysis of DSs compared to that ofthe first aliquot give a measure ofthe precision associated with laboratory procedures, but not with sample collection, preservation, or storage procedures. 3.4 Field Blank Control Sample (FB) ASTM Type I water placed in a sample container in the laboratoryand treated as a sample in all respects, including exposure to sampling site conditions, storage, preservation and all analytical procedures. The purpose ofthe FB isto determine iftest substances or other interferences are present in the field environment. ETS-8-154.1 Page 2 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 38 o f 123 3M Environmental Laboratory Project No. E05-0210 3.5 Field Duplicate (FD) A sample collected induplicate at the same time as the sample and placed under identical circumstances and treated exactlythe same throughoutfield and laboratory procedures. Analysis of FD compared to that ofthe first sample gives a measure of the precision associated with sample collection, preservation and storage, as well as with laboratory procedures. 3.6 Field Matrix Spike (FMS) A sample collected induplicate towhich known quantities of the target analytes are added inthe field at the time of sample collection. Alternatively, the known quantityof target analytes may be added to the sample bottle inthe laboratory before the bottles are senttothefield. A known, specific volume of sample must be added to sample containerwithout rinsing. This may be accomplished by making a "fill tothis level"line on the outside ofthe sample container. The FMS should be spiked at approximately50-150% of the expected analyte concentration in the sample. Ifthe expected range of analyte concentrations is unknown, a lowand a high spike may be prepared to increase the likelihood that a spike at an appropriate range is made. The FMS is analyzed to ascertain if any matrix effects, interferences, or stability issues may complicate the interpretation ofthe sample analysis. 3.7 Field Spike Control Sample (FSCS) An aliquot ofASTM Type I water to which known quantities ofthe target analytes are added in the field at the time of sample collection (at an appropriate concentration to be determined bythe project lead) or inthe laboratorypriorto the shipment ofthe collection bottles. The FSCS is extracted and analyzed exactly like a sample to determine whether a loss of analyte could be attributed to sample storage and/or shipment. A lowand high FSCS may be appropriate when expected sample concentrations are not known. 3.8 Laboratory Control Sample (LCS) An aliquot ofASTM Type I waterto which known quantities of the target analytes are added in the laboratory. Two levels are included, one at the LLOQ (approx. 25 pg/mL), the other at a concentration of approx. 100-250 pg/mL or another concentration to be determined by the project lead. The LCS is extracted and analyzed exactly like a laboratorysample to determine whether the methodology is in control, and whether the laboratory is capable of making accurate measurements at the required method detection limit and higher. 3.9 Laboratory Sample A po rtion o f a s a m p le received fro m th e fie ld fo r testin g . 3.10 Limit of Detection (LOD) The LOD isthe lowest concentration of an analytethat can be measured and reported with 99% confidence that the analyte concentration is greater than zero. If required, the LOD may be determined in several ways, including signal-to-noise ratio and statistical calculations. 3.11 Limit o f Quantitation (LOQ) The LOQ for a dataset is the lowest concentration (LLOQ) or highest concentration (ULOQ) that can be reliably achieved within the specified limits of precision and accuracyduring routine operating conditions. Note: For many analytes, the LLOQ analyte concentration isselected as the lowest non-zero standard in the calibration curve to simplifydata reporting. Sample LLOQs are matrix-dependent 3.12 Matrix Spike (MS) ETS-8-154.1 Page 3 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 39 o f 123 3M Environmental Laboratory Project No. E05-0210 A matrix spike is an aliquot of a sample, to which known quantities of target analytes are added in the laboratory. The MS is extracted and analyzed exactly like a laboratorysample to determine whetherthe sample matrix contributes bias to the analytical results. The background concentrations of the analytes inthe sample matrix must be determined in a separate aliquot and the measured values in the MS corrected for background concentrations. 3.13 Method Blank An aliquot ofASTM Type I water that is treated exactly like a laboratory sample including exposure to all glassware, equipment, solvents, and reagentsthat are used with other laboratory samples. The method blank is used to determine iftest substances or other interferences are present in the laboratoryenvironment, the reagents, or the apparatus. 3.14 Method Detection Limit (MDL) Determination A MDL isthe statisticallycalculated minimum amount of an analyte that can be measured with 99% confidencethatthe reported value is greater than zero. One of several processes that may be used to establish a LOD value is found in 40 CFR Part 136 Appendix B. 3.15 Sample A sample is a small portion collected from a larger quantityof material intended to representthe original source material. 3.16 Spiking Stock Standard (SSS) A solution preparedfrom stock standards used to prepare the working standard. 3.17 Stock Standard (SS) A concentrated solution of a single analyte prepared in the laboratorywith an assayed reference compound. 3.18 Working Standard (WS) A solution of several analytes prepared inthe laboratoryfrom SSs and diluted as needed to prepare calibration standards and other required analyte solutions. 4 Warnings and Cautions 4.1 Health and Safety The acute and chronictoxicityof the standards forthis method have not been precisely determined; however, each should be treated as a potential health hazard. Unknown samples may contain high concentrations ofvolatile toxic compounds. Sample containers should be opened in a hood and handled with glovesto prevent exposure. The laboratory is responsible for maintaining a safe work environment and a current awareness of local regulations regarding the handling of the chemicals used in this method. A reference file of material safety data sheets (MSDS) should be available to all personnel involved in these analyses. 4.2 Cautions None 5 Interferences ETS-8-154.1 Page 4 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 40 o f 123 3M Environmental Laboratory Project No. E05-0210 During extraction and analysis, major potential contaminant sources are reagents and solid phase extraction devices. All materials used in the analyses shall be demonstrated to be free from interferences under conditions of analysis by running method blanks. Parts and supplies that contain Teflon should be avoided due tothe possibility ofinterference and/or contamination. These may include, but are not limited to: wash bottles, Teflonlined caps, autovial caps, HPLC parts, etc. The use of disposable micropipettes or pipettesto aliquot standard solutions is recommended to make calibration standards and matrix spikes. 6 Instrumentation, Supplies, and Materials Note: Brand names, suppliers, and part numbers are for illustrative purposes only. Equivalent performance may be achieved using apparatus and materials otherthan those specified here, but demonstration of equivalent performance that meets the requirements of this method is the responsibilityof the laboratory performing the analysis. 6.1 Instrumentation Balance, analytical (display at least O.OOOIg), Mettler HPLC/MS/MS or HPLC/MS system, as described in Section 10. 6.2 Supplies and Materials. Sample collection bottles--LDPE (e.g., NalgeneTM) narrow-mouth bottles with screwcap. Note: Do not useTeflon bottles or Teflon lined caps. Coolers for sample shipment. Ice for sample shipment. Vacuum pump, BGchi. Visiprep vacuum manifold, Supelco. Sep Pak Vac 6cc (1g) tC18 cartridges (part# WAT 036795),Waters. 50mL disposable polypropylene centrifuge tubes, VWR. 15mL disposable polypropylene centrifuge tubes, VWR. Disposable micropipettes (50-1OOpL, 100-200pL), Drummond. ClassA pipettes and volumetric flasks, various. Hypercarb drop-in guard column (4mm) (part# 844017-400), Keystone. Stand-alone drop-in guard cartridge holder, Keystone. 125mL LDPE narrow-mouth bottles, Nalgene. 2mL dear HPLC vial kit (cat# 5181-3400), Agilent/Hewlett Packard. Standard lab equipment (graduated cylinders, disposabletubes, etc.), various. 7 Reagents and Standards Note: Suppliers and catalog numbers are for illustrative purposes only. Equivalent performance may be achieved using chemicals obtained from other suppliers. Do not use a lessergrade of chemical than those listed. ETS-8-154.1 Page 5 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 41 o f 123 3M Environmental Laboratory Project No. EOS-0210 7.1 Chemicals Methanol (MeOH), HPLC grade, JT Baker, Catalog No. JT9093-2. Ammonium Acetate, Reagent grade, Slgma-Aldrich, Catalog No. A-7330. ASTM Type I Water, prepared in-house. Sodium Thiosulfate, Reagent grade, JT Baker. 7.2 Standards Potassium perfluorooctane sulfonate Perfluorooctane sulfonylamide Ammonium perfluorooctanoate Others as required. 7.3 Reagent Preparation 250mg/mL sodium thiosulfate solution-- Dissolve 25g of sodium thiosulfate in 100mL reagent water. 40% methanol wash solution - Measure 400mL methanol and adjust volume to 1.0L with reagent water. 100mM ammonium acetate solution (Analysis)--Weigh 7.71g of ammonium acetate and dissolve in 1,0L of reagentwater. Dilute the 100mM solution by a factorof 50 to make the 2mM ammonium acetate solution used for mobile phase A. Note: Alternative volumes may be prepared as long as the ratios of the solvent to solute ratios are maintained. 7.4 Spiking Stock Standard (SSS) Preparation The following standard preparation procedure serves as an example and may be changed to suit the needs of a particular study. For example, pL volumes may be spiked into volumetricflasks when diluting stock solutions to appropriate levels. 100pg/mL each PFOS, PFOSA, and POAA SSSs--Weigh out 10mg of analytical standard (corrected for percent salt and purity--i.e., 10 mg C8Fi7S03K purity90% = 8.35mg CaF^SOa") and dilute to 100mL with methanol in a lOOmL volumetricflask. Transferto a 125mLLDPE bottle or o th e r s u ita b le c o n ta in e r. P re p a re a s e p a ra te so lution fo r e a c h a n a ly te . S o lu tio n s m a y b e sto red in a refrigerator at 42C for a maximum period of 6 monthsfrom the date of preparation. 1pg/mL mixed SSS--Add I.OmLeachofthe 100pg/mL SSSs (from 7.4.1) to a 100mL volumetric flask and bring up tovolume with methanol. 0.1pg/mL mixed SSS--Add 10.0mL ofthe 1.0pg/mL-mixed solution (from 7.4.2) to a 100mL volumetricflask and bring up tovolume with methanol. 0.01pg/mL mixed SSS--Add 10.0mL of the 0.1pg/mL-mixed solution (from 7.4.3) to a 100mL volumetricflask and bring up to volume with methanol. Storage Conditions--Store all SSSs in a refrigerator at 42C for a maximum period of 6 monthsfrom the date of preparation. 7.5 Calibration Standards The following standard preparation procedure serves as an example and may be changed to suit the needs of a particularstudy, provided the concentrations are calculated correctly. ETS-8-154.1 Page 6 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 42 o f 123 3M Environmental Laboratory Project No. EOS-0210 100pg/mL each PFOS, PFOSA, and POAA stock standard solutions--Weigh out 10mg of analytical standard (correctedfor percent salt and purity) and dilute to 100mL with methanol in a 10OmL volumetricflask. Transfer to a 125mL LDPE bottle or other suitable container. Prepare a separate solution for each analyte. Store solutions in a refrigeratorat 42C for a maximum period of 6 months from the date of preparation. 1pg/mL Working Standard--Add I.OmLeachofthe 100pg/mLSS solutions (from 7.5.1) to a 100mL volumetricflask and bring upto volume with methanol. 0.1pg/mL Working Standard--Add 10.0mLofthe 1.0pg/mL mixed solution (from 7.5.2) to a 100mL volumetricflask and bring upto volume with methanol. 0.01pg/mL Working Standard --Add 10.0mLofthe0.1pg/mL mixed solution (from 7.5.3) to a 100mL volumetricflask and bring upto volume with methanol. Storage Conditions--Store all WSs in a refrigerator at 42C for a maximum period of 6 monthsfrom the date of preparation. Calibration Standard--Prepare calibration solutions inASTM Type I using the following table as a guideline: Final Calibration C oncentration Volum e o f Standard Volume, m l. , of WS, pg/mL WS, pL of ASTM Type I Water Final C oncentration of Calibration Standard. pg/m L, in ASTM Type I W ater 0.0 0.010 0.010 0.010 0.10 0.10 0.10 0.10 1.0 1.0 1.0 0 100 200 400 100 200 300 400 100 400 1000 40 40 40 40 40 40 40 40 40 40 40 0 25 50 100 250 500 750 1000 2500 10000 25000 The standards are processed through the extraction procedure (Section 11), identical to the laboratory samples. The concentration ofthe calibration standard in the final extract is equal to 8X the initial concentration, due to the concentration of the standard during the extraction process. Storage Conditions--Store all extracted calibration standards in 15mL polypropylene tubes at 42C, for a maximum period of twoweeks from the date of preparation 8 Sample Collection and Handling _______________________________ Note: Sampling equipment, including automatic samplers, must be free of Teflon tubing, gaskets, and other parts that may leach interfering analytes into the water sample. Automatic samplers that composite samples over time should use refrigerated polypropylene sample containers if possible. Sample bottles should not be rinsed before sample collection. Labeling: Each sample bottle mustdisplay information regarding the collection ofthat sample, the individual collecting the sample, and any matrix spike that has been added to the sample. ETS-8-154.1 Page 7 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 43 o f 123 3M Environmental Laboratory Project No. E05-0210 This includes the volume and concentration of any spiking solution added and the volume and identification of any preservatives added inthe field. Spiking: The spiking scheme will be clearly outlined in the sampling plan, including whetherthe samples will be spiked inthe field or in the laboratory priortothe shipment of the bottles to the site. If spiking isto be performed inthe field, materials and specific instructions will be included in the sampling kit. Be sure to dearly label each bottlewith spiking information if applicable. Tap Water: Open the tap and allow the system toflush until the watertemperature (1510C) has stabilized (usually about two minutes). Adjustthe flowto about 500mL/min and coiled samples from the flowing stream. Ground Water: Purge the well of standing water using a pump or a bailer. Colled the sample directlyfrom the pumporfrom the bailer. Surface Water: When sampling from an open bodyofwater, fill the sample containerwith water from a representative area. Sample Dechlorination: All samples should be iced or refrigerated at 42C and kept inthe dark from the time of collection until extraction. Residual chlorine should be eliminated byadding 200pL of a 250mg/mL sodiumthiosulfate solution to each tap-water sample and assodated FB and FSCS (which may be placed in each bottle before leaving for the sampling siteor done in the field.). Holding Time (HT): Results of the time/storage studyof all target analytes showed that the three compounds are stable for 14 days in water samples when the samples are dechlorinated and stored as described in the previous section (see also references in section 17). Therefore, laboratory samples must bieextracted within 14 days and the extracts analyzed within 30 days of sample collection. If the HT exceeds 14 days, great care is used when evaluating field spikes to avoid misrepresentation ofthe sample concentration. 8.1 Field Blanks Process a Field Blank Control Sample (FB) along with each sample set (samples collectedfrom the same general sample site at approximatelythe same time). At the laboratory, priorto sample collection, fill a sample containerwith ASTM Type I water, seal, and ship the FB to the sampling site along with the emptysample containers. Return the FB to the laboratorywith the filled sample bottles. When sodium thiosulfate is added to samples, use the same procedure to preserve the FB. 8.2 Field Duplicates Collect a Field Duplicate (FD) for everyten (10) samples collected or per each sampling set, if less than 10 samples are collected. Separate FDs must be collected for each type ofwater sample (ground, tap, etc.) collected. Collect the FD immediately afterthe sample. Preserve, store and ship FD using the same procedures as usedfor the samples. 8.3 Field Spike Control Sample (FSCS) A Field Spike Control Sample (FSCS) must be preparedfor each sample shipment. If multiple coolers are used to ship a set ofsamples, each cooler must contain a FSCS. At the laboratory, fill a sample containerwith 100mL ofASTM Type I water. Seal and shiptothe sampling site along with the empty sample containers and FBs. Samples may either be spiked in the field or inthe laboratory priorto shipment. The method employed should be consistent throughoutthe study. If the samples are to be spiked in the field, be sure to send appropriate supplies and instructionsfor the field personnel tofollow. ETS-8-154.1 Page 8 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 44 o f 123 3M Environmental Laboratory Project No. E05-0210 Seal and gently Invert the FSCS to mix. Store and ship the FSCS using the same procedures as used forthe samples Provide information on sample collection, preservation, shipment and storage. List applicable holding times. Include sample stabilityand extract storage requirements. Reference the method used for sample preparation, if applicable. 8.4 Field Matrix Spike (FMS) A Field Matrix Spike (FMS) must be prepared for each sampling location. One unspiked sample fromthe same location must accompanythe FMS to determine endogenous levels in the sample. The samples should be dearly identifiable as being from the same location. Samples mayeither be spiked inthe field or in the laboratory priorto shipment. The method employed should be consistentthroughout the study. Ifthe samples are to be spiked inthe field, be sure to send appropriate supplies and instructionsfor the field personnel tofollow. 9 Quality Control and Data Quality Objectives Analytical results ofthe FB, FMS, FD, and FSCS should be evaluated at the condusion of the studyto help interpret the qualityof sample data. Analytical results for these control/duplicate samples must be reported with the sample data. 9.1 Solvent Blanks Solvent blanks are analyzed with each sample set to determine contamination or carryover. Aliquots of methanol representthe solvent used for the standard curve and the sample extraction. Solvent blanks should have area counts that are less than 50% ofthe area count of the lowest calibration standard. Solvent blanks should be analyzed priorto andfollowing each calibration curve, each set of system suitabilitysamples, and after no more than 10 unknown sample extracts. If instrument carryover is a problem consecutive solvent blanks may be necessary. In this case the area counts of the solvent blanks should return to <50% of the lowest calibration standard priorto the injection offurther standards or samples. 9.2 Method Blanks A method blank consists of an aliquot of ASTM Type I water, equal In volume to the samples, and extracted in the same manner as the samples. At least two method blanks should be prepared an d a n a ly ze d e a c h d a y th a t ex tra ctio n s a re p e rfo rm ed for a p a rtic u la r stu d y or p ro ject. W h e n analyzed the area counts of these samples must be less than 50% ofthe area count of the lowest calibration standard. 9.3 Sample Replicates All samples, including field spikes, trip blanks, etc., should be extracted at least induplicate, and in triplicate if difficultieswere encountered in the sampling and/or holding conditions ofthe samples. The relative percent difference (RPD) of duplicate samples or relative standard deviation (RSD) should be less than 15% for the precision of sample preparation and analysis to be considered in control. 9.4 Matrix Spike Matrix spikes are prepared for each sample type and analyzed to determine the matrix effect on the recovery efficiency. Matrix spike recoveries shouldfall within 25% of expected values. If the matrix spikesfail, evaluate the lab control spikes. If the LCS are within acceptance criteria there may be matrix issues in the samples. Discuss these in the final report ETS-8-154.1 Page 9 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 45 o f 123 3M Environmental Laboratory Project No. E05-0210 Matrix spike duplicates are prepared periodicallyto measure the precision associated with the analysis. Analyze a matrix spike and matrix spike duplicate (if prepared) in the same run as the original sample. Matrix spike and matrix spike duplicate concentrations should fall in the mid-range ofthe initial calibration curve or should be prepared at 1.5-5 times the endogenous concentration ofthe analyte. Spike concentrations shouldfall in the low-range of the initial calibration curve if extremely low-levels are expected. Generallytwo or more levels are prepared, one in the low range ofthe curve and one in the mid-range. This avoids the need to pre-screen unknown samples prior to preparation. 9.5 Laboratory Control Spike Lab control spikes are prepared for each studyto ensure recovery of the target analytes. These should be prepared at a minimum of 2 levels and in duplicate ortriplicate. Recovery ofthese samples should be within 25% of expected values, and the RPD (or RSD) be 15%. If recoveries fall outside these limits the samples should be addressed in the final report 10 Calibration and Standardization 10.1 Instrument Setup Note: In this example, a MicroMass UltimaTM Liquid ChromatographyTandem Mass Spectrometer (LC/MS/MS) is used. Other brands of LC/MS/MSs as well as single quadrupole mass spectrometers (LC/MS) may be used as long as the method criteria are met. Brand names, suppliers, part numbers, and models are for illustrative purposes only. Equivalent performance may be achieved using apparatus and materials otherthan those specified here, but demonstration of equivalent performance that meets the requirements of this method is the responsibilityof the laboratory. The operator must optimize and documentthe equipment and settings used. Establish the LC/MS/MS system and operating conditions equivalent to the following: Mass Spec: Micromass Ultima (Micromass) Interface: Electrospray (Micromass) Mode: Electrospray Negative, Multiple Response Monitoring (MRM) Harvard infusion pump (Harvard Instruments), for tuning Computer. COMPAQ Professional Workstation AP200 Software: Windows NT, MassLynx 3.3 HPLC: Hewlett Packard (HP) Series 1100 HP Quaternary Pump HP Vacuum Degasser HP Autosampler HP Column Oven Note: A 4 x 10mm Hypercarb drop-in guard cartridge (Keystone, part# 844017-400) may be attached on-line after the purge valve and before the sample injector porttotrap any residue contaminants that may be in the mobile phase and/or HPLC system. HPLC Column: Genesis C8 (Jones Chromatography), 2.1mm x 50mm, 4pm ETS-8-154.1 Page 10 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 46 o f 123 3M Environmental Laboratory Project No. E05-0210 Column Temperature: 35C Injection Volume: 15pL Mobile Phase (A): 2mM Ammonium Acetate InASTM Type I water (See 7.3.1) Mobile Phase (B): Methanol Time, Percent Mobile Percent Mobile Flow Rate, min Phase A Phase B mL/min 0.0 60 40 0.3 0.4 60 1.0 10 40 0.3 90 0.3 7.0 10 90 0.3 7.5 0 100 0.3 9.0 0 100 0.4 9.5 60 40 0.4 13.5 60 40 0.4 14.0 60 40 0.3 Note: Other HPLC gradients may be used as long as the method criteria are met. It may be necessaryto adjust the HPLC gradient in orderto optimize instrument performance. Columns with different dimensions (e.g. 2.1mm x 30mm) and columnsfrom different manufacturers (Keystone Betasil C18 etc.) may be used. Ions Monitored: Analyte Primary Ion Product Ion ____ Approximate Retention Time (minutes) PFOA PFOS FOSA 413 499 498 169 99 78 5.0 5.2 5.8 Other product ions may be chosen at the discretion ofthe analyst, although m/z 99 is suggested fo r P F O S . U s e o f th e su g g ested p rim ary io n is reco m m en d ed . R ete n tio n tim e s m a y v a ry slightly, on a day-to-day basis, depending on the batch of mobile phase etc. Drift in retention times is acceptable within an analytical run, as long as the drift continues through the entire analysis and the standards are interspersed throughoutthe analytical run. 10.2 Tune File Parameters The following values are provided as an example. Actual values mayvaryfrom instrument to instrument. Also, these values maybe changed from time to time in orderto optimize for greatest sensitivity. Analyte PFOA PFOS FOSA Dwell, sec 0.2-0.4 0.2-0.4 0.2-0.4 Collision Energy, eV 10-25 30-60 20-50 Cone, V 20-30 50-80 30-60 ETS-8-154.1 Page 11 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 47 o f 123 3M Environmental Laboratory Project No. E05-0210 Source Capillary Hexapole 1 Aperture 1 Hexapole 2 Source Block Temp. Desolvation Temp. Analyzer LM Res 1 HM Res 1 lEnergy 1 Entrance Exit LM Res 2 HM Res 2 lEnergy 2 Multiplier Gas Flows Cone Gas Desolvation Pressures Gas Cell Set 2.6-3.5kV 0.5V 0.2V 0.8V 100-150C 250-400C Set 12.5- 15.0V 12.5- 15.0V 0.7V -2 V 1V 11.0V 11.0V 1.0V 650V Set 150Uhr 700L/hr Set 3.0e-3mbar ETS-8-154.1 Page 12 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 48 o f 123 3M Environmental Laboratory Project No. E05-0210 10.3 Calibration Curve Analyze the standard curves priorto each set of samples. The validated method specifiesthat the standard curve should be plotted using a linearfit, weighted 1/x or unweighted. However, the standard curve may also be plotted byquadratic fit (y = ax2+ bx + c), weighted 1/x or unweighted, using suitable software. The calibration curves may include but should not be forced through zero. The mathematical method used to calculatethe calibration curve should be applied consistentlythroughout a study. Any change should be thoroughlydocumented in the raw data. If the calibration curve does not meet acceptance criteria perform routine maintenance or prepare a newstandard curve (if necessary) and reanalyze. For purposes of accuracywhen quantitating low levels of analyte, it may be necessaryto use the lowend ofthe calibration curve ratherthan the lull range. For example, when attempting to quantitate approximately 50 pg/mL of analyte, generate a calibration curve consisting of the standards from 25 pg/mLto 1000 pg/mL rather than the full range of the curve (25 pg/mL to 25000 pg/mL). This will reduce inaccuracy attributed to linear regression weighting of high concentration standards. High and/or low points may be excluded from the calibration curvesto provide a betterfit overthe linear range appropriate tothe data or because they did not meet the pre-determined acceptance criteria. Low-level curve points should also be excluded iftheir area counts are not at leasttwice that of the method and/or solvent blanks. Any curve point may be rejected due to a bad injection orfailing to meet accuracy requirements of 25% (and 30% for the LLOQ). Justification for exclusion of calibration curve pointswill be noted in the raw data. A minimum of 6 pointswill be used to constructthe calibration curve. 10.4 Continuing Calibration Verification (CCV) Continuing calibration verifications (CCV) are analyzed to verifythe accuracy ofthe calibration curve. Analyze a mid-range calibration standard, one of the same standards used to construct the calibration curve, at a minimum after every tenth sample, not including solvent blanks, with a minimum ofone per sample set. Calibration verification injections must be within 25% to be considered acceptable. The calibration curve and the last passing CCV will then bracket acceptable samples. Multiple CCV levels may be used. 10.5 System Suitability A minimum of three system suitabilitysamples will be injected at the beginning and end of each analytical run. Typicallythese samples are run priortothe calibration curve. The system su itab ility in jectio n s m u st h a v e a re a co un ts w ith a n R S D o f 5 % an d a reten tio n tim e R S D o f 2 % when evaluated independently. 11 Procedures 11.1 Extraction Scheme Allowsamples to equilibrate to room temperature. Thoroughlymix samples bygently inverting the sample bottle. Measure 40mL of sample into 50mL polypropylene centrifugetubes (Spike the Matrix spikes as required*, replace lid and mixwell). Note: * Samples may need to be prescreened to determine an appropriate matrix spike level (typically 50-150% of sample concentration). Alternatively the samples could be spiked at more than one level, allowing forthe inappropriate spike level to be eliminated. ETS-8-154.1 Page 13 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 49 o f 123 3M Environmental Laboratory Project No. E05-0210 Condition the C18 SPE cartridges (1g, 6mL) by passing approximately 10mL methanol followed by approximately 50mL ASTM Type I water (flow rate approximately 2 drop/sec). Do not let column run dry. Note: Forthe following steps, maintain a -1 drop/sec flow rate. Do not allowthe column to run dry at anytime. Load the analytical sample onto the C18 SPE cartridge. Discard eluate. Ten mL ofthe 40% methanol inwater wash mixture is passed through the C18 SPE cartridge to rinse away potential interferences and then discarded. This step must be omitted if perfluorinated compounds with chain lengths less than C8 are targeted since these will be lost during thiswash step. Elutewith exactly 5mL of 100% methanol. Collect eluate intograduated 15mL polypropylene centrifuge tubes. This is the target elution fraction (final volume approximately4.5 mL as not all of the solvent will leave the SPE column. This will not affectthe calculations in any way sincethe curve is also extracted). Analyze a portion of the target elution fraction eluent using negative electrospray HPLC/MS/MS or HPLC/MS. Note: Samples are concentrated by a factorof eight during the extraction; Initial Vol = 40mL -> Final Vol. = 5mL. Samples are stable at room temperature for at least 24 hours. Analytical samples may be stored in a refrigerator at 42C until analysis. Standardization of C18 SPE columns--If poor recoveries are observed, it may be necessaryto standardize the C18 SPE columns in the following manner before analyzing samples. Use a standard with an analyte concentration between 1000 and 4000 pg/mL. Repeat the extraction scheme from the beginning up through the eluting with ~5mL 100% methanol. After the eluting with ~5mL 100% methanol step, collect an additional post-elution fraction by eluting with an additional 5mL of 100% methanol. Analyze bothfractions by HPLC/MS/MS or HPLC/MS. If the targetfraction contains a minimum of 85% ofthe respective analytes, it may be considered acceptable. If the wash contains significant standard (>15%), eitherthe wash volume or percentage of MeOH should be decreased. If the post-elution fraction contains significant standard (>15%), the target elution volume should b e in c re a s e d . 11.2 Sam pie Analysis Set up analysis sample queue. Inject the same volume (between 5-25pL) of each standard, analytical sample and blank into the instrument. All samples with a concentration > ULOQ must be diluted and reanalyzed. If dilution of the final extract fails to produce acceptable results (e.g. poor MS recoveries) dilute the original sample and re-extract. 12 Data Analysis and Calculations Calculatethe analytical sample (extract) concentration from the standard curve using the following equation: ETS-8-154.1 Page 14 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 50 o f 123 3M Environmental Laboratory Project No. E05-0210 Extract Concentration, pg/mL = (Peak area - intercept! (slope) Calculate the percent recovery ofthe FSCS using the following equation: FSCS % ree. = (FSCS cone., pg/mL) x 100 (Cone, added, pg/mL) Calculatethe percent recovery ofthe MSs using the following equation: MS % rec. = (MS cone., pg/mL - Sample cone., pg/mL) x 100 (Cone, added, pg/mL) 13 Method Performance Note: Any method performance parameters that are not achieved must be considered inthe evaluation of the data. Nonconformance to any specified parameters must be described and discussed in any reporting of the data. If criteria listed in this method performance section are not met, maintenance may be performed on the system and samples reanalyzed, or other actions taken as determined bythe analyst. Document all actions in the rawdata. If data are to be reported when performance criteria have not been met, the data must be footnoted on tables and discussed in the text of the report. 13.1 System Suitability A minimum of three system suitabilitysamples will be injected at the beginning and end of each analytical run. Typically these samples are run priorto the calibration curve. The system suitability injections must have area counts with an RSD of 25% and a retention time RSD of 22% when evaluated independently. 13.2 Quantitation Calibration Curve: The coefficient of determination (r2) value forthe calibration curve must be greaterthan or equal to 0.990. Each point in the curve must be within 25% of the theoretical concentration with the exception ofthe LLOQ, which may be within 30%. Demonstration of Specificity: Specificityis demonstrated bychromatographic retention time (within 3% of standard) and the mass spectral response of unique ions. 13.3 Sensitivity Solvent Blanks and Method Blanks: Solvent and method blank area counts must be < 50% that of the lowest standard used in the calibration curve. Limits of Quantitation (LOQ): The lower LOQ (LLOQ) is the lowest non-zero active standard in the calibration curve; the peak area of the LLOQ must be at least 2X that ofthe extraction blank. Bydefinition, the measured value ofthe LLOQ must be within 30% of the theoretical value. ETS-8-154.1 Page 15 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. \ E05-0210 Interim Report #5 Page 51 o f 123 3M Environmental Laboratory Project No. E05-0210 13.4 Accuracy CCV Performance: Calibration verification injections must be within 25% to be considered acceptable. The calibration curve and the last passing CCV will then bracket acceptable samples. Multiple CCV levels may be used. Matrix Spikes: Matrix spike percent recoveries must be within 25% ofthe spiked concentration. If matrix effects are suspected, evaluate the LCS results to determine if a matrix effectsare present and if the method is in control based on compliant LCS results. Discuss all results in the analytical report. 13.5 Precision Reproducibility: Reproducibilityof the method is defined bythe results of duplicate or triplicate analysis of samples. A RPD or RSD of 15% will be considered acceptable. System Suitability: The system suitability injections must have area counts with an RSD of 5% and a retention time RSD of 2% when evaluated independently. 14 Pollution Prevention and Waste Management Sample extractwaste and flammable solvent is discarded in high BTU containers, and glass pipette waste isdiscarded in broken glass containers located in the laboratory. 15 Records Each data package generated for a studymust have the following information included: studyor project number, acquisition method, integration method, sample name, extraction date, dilution factor (if applicable), and analyst. Printthe tune page, sample list, and acquisition method to include in the appropriate studyfolder. Copythese pages and tape into the instrument run log. Rot the calibration curves as described in this method, then printthese graphs and store in the studyfolder. Print data integration summary, integration method, and chromatograms and store in the study folder. S u m m a rize d a ta using s u ita b le so ftw a re an d sto re in th e s tu d y fo ld e r. 16 Attachments None. 17 References "Method ofAnalysis for the Determination of Perfluorooctane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluorooctanoate (POAA) inWater", E. Wickremesinhe and J. Flaherty, Study Number 023-002, Centre Analytical Laboratories, Inc., State College, Pennsylvania, January 2000. Validation reportfor the "Method ofAnalysis forthe Determination of Perfluorooctane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluorooctanoate (POAA) in Water", E. Wickremesinhe and J. Flaherty, Study Number 023-002, Centre Analytical Laboratories, Inc., State College, Pennsylvania. ETS-8-154.1 Page 16 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 52 o f 123 3M Environmental Laboratory Project No. E05-0210 18 Affected Documents None. 19 Revisions Revision Number 1 Revision Number Updated to the new format. Changed Title. Section 1: States the validation o f 3 analytes, removes reference to EPA document that's no longer applicable. Section 2: Provided for the extraction o f more than the 3 validated analytes, allows the use o f a LCIMS system, not only the LSIM SIM S previously mentioned. Section 3: Revised definitions for field matrix spike, field control spike, LLOQ, method blank, and MDL. Section 5: Reworded the interferences, added recommendation to use disposable pipettes. Section 6: Recategorized and pared down. Section 7: Changed storage time to 6 months. Added more calibration points to the table. Section 8:Added statement addressing labeling requirements and spiking procedures. Expanded section 8.8. Section 9: New Section Section 10: Changed some o f the parameters in the tables. Allowed for use of different instrumentation. Added information from section 12 o f previous version, extensively revised. Section 11 (section 9 in previous version): Clarification o f wash step, stated exact volume o f eluate is 5 mL, revised standardization process, rem oved requirement to use LCIMSIMS. Section 12 (section 13 in previous version: no changes Section 13 (section 14 in previous version): Extensively rewritten. Section 14 (section 15 in previou s version): no ch an ges Section 15 (section 16 in previous version): Minor changes to recording requirements. Section 16 (section 17 in previous version): Removed attachment. Section 17 (section 18 in previous version): Removed reference to EPA document that no longer applied to this SOP. Section 18: N ew section. Revision Date 7 ETS-8-154.1 Page 17 of 17 Determination of Perfluorinated Compounds in Water Using SPE and LC/MS. E05-0210 Interim Report #5 Page 53 o f 123 3M Environmental Laboratory Project No. E05-0210 Attachment C. Prototol and Protocol Amendments E05-0210 Interim Report #5 Page 54 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 STUDY PROTOCOL Study Title: PFeMirsfhlau,moCArmlnoaaaomPllcyestSrsa,eifnVsrlueueosomgfurePlotMfUaheorstenoiifxoannlauntgiento,oLer(SrPsoCmiubnF/lauMgOfoltlPSaSnMnr)/aMoeitangesSmur(WalPmffmooFarantHleatrShLt,e)ei,Sv(3aPeoMnriFld,aBDSnSede)dc,Saimmtuearnllt, Exygen Protocol Number: P0001131 Performing Laboratory: Exygen Research 3058 Research Drive State College, PA 16801 Phone: (814) 272-1039 E05-0210 Interim Report #5 Sponsor Representative: Michael A. Santoro Director o f Regulatory Affairs 3M Building 023 6 -0 1-B-10 St. Paul, MN 55144 Phone: (651)733-6374 Page I <>J65 Page 55 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 DISTRIBUTION: 1) Jaisimha Kesari, 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 E05-0210 Interim Report #5 Page 2 of 65 Page 56 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 PROTOCOL APPROVAL Study Title: Analysis o f Perfluorobutanesulfonate (PFBS), Perfluorohexanesulfonate (PFHS), and Perfluorooctanesulfonate (PFOS) 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: P0001131 APPROVALS JaisimhatiCeSan, Study Director Weston Solutions Michael A. S fioro , Sponsor Representative 3M Comparfy u HO ate ohn M. Flaherty, Principal Investigator Exygen Research sident, Facility Management Date 20 -o r- Date [Shaffer, Techni; ead, Quality Assurance Unit E^/gen Research / U WDaOte) E05-0210 Interim Report #5 Page 3 oj 65 Page 57 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOO1131 TABLE OF CONTENTS TITLE PAGE........................................................................................................................................................ 1 DISTRIBUTION................................................................................................................................................... 2 PROTOCOL APPROVAL....................................................................................................................................3 TABLE OF CONTENTS......................................................................................................................................4 INTRODUCTION................................................................................................................................................. 3 TEST MATERIALS.............................................................................................................................................5 OBJECTIVE..........................................................................................................................................................6 TESTING FACILITY...........................................................................................................................................& STUDY DIRECTOR.............................................................................................................................................7 SPONSOR REPRESENTATIVE...........................................................................................................................7 PRINCIPAL INVESTIGATOR.............................................................................................................................7 PROPOSED EXPERIMENTAL START AND TERMINATION DATES....................................................... 7 IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM..........................................................8 SAMPLE PROCUREMENT, RECEIPT AND RETENTION...........................................................................8 SAMPLE IDENTIFICATION............................................................................................................................. 9 ANALYTICAL PROCEDURE SUMMARY..................................................................................................... 9 VERIFICATION OF ANALYTICAL PROCEDURE........................................................................................9 METHOD FOR CONTROL OF 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........................................................................................................15 E05-0210 Interim Report #5 Page 4 of 65 Page 58 o f 123 3M Environmental Laboratory Project No. E05-0210 Exgen Protocol Number: P0001131 INTRODUCTION The purpose of this study is to perform analysis for perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS) and perfluorooctanesulfonate (PFOS) 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 o f Exygen Research. TEST MATERIALS The test materials are perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS) and perfluorooctanesulfonate (PFOS) and are all supplied by 3M. PFBS Chemical Name: Perfluorobutanesulfonate Molecular Weight: 338 supplied as the potassium salt ^FqSO'K*) Lot Number: 101 Purity: 96.7% Transitions Monitored: 299 - 99 Structure: F FF PFHS Chemical Name: Perfluorohexanesulfonate Molecular Weight: 438 supplied as the potassium salt (CFuSOj'K*) PLuortiNtyu:m9b8e.6r%: SE036 Transitions Monitored: 399 -- 80 Structure: F F Page 5 of 65 E05-0210 Interim Report #5 Page 59 o f 123 3M Environmental Laboratory Project No. E05-0210 Exgen Protocol Number: P00011 ? I PFOS Chemical Name: Perfluorooctanesulfonate Molecular Weight: 538 supplied as the potassium salt (CsFnSOj'K4) Lot Number: 217 Purity: 86.9% Transitions Monitored: 499 --> 99 Structure: OBJECTIVE The purpose of this study is to perform analysis for perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS) and perfluorooctanesulfonate (PFOS) in water, soil, sediment, fish, clams, vegetation, small mammal livers and small mammal serum for the 3M Decatur Monitoring Program using the current versions of the following Exygen analytical methods: V0001780: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" V0001782: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method of Analysis for the Determination of Perfluorooctanoic V0001784: "AMciedth(PodFOoAf A) innaFlyissihs afnord tChleamDestberymLiCna/MtioSn/MofSP"erfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method of Analysis for the Determination of 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 Page 6 of65 E05-0210 Interim Report #5 Page 60 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 STUDY DIRECTOR Jaisimha Kesari P.E., DEE Weston Solutions, Inc. 1400 Weston Way West Chester, PA 19380 Phone: (610) 701-3761 Fax: (610) 701-7401 j .kesari@westonsolutions.com SPONSOR REPRESENTATIVE Michael A. Santoro 3M Company Director of Regulatory Affairs 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 DPRAOTEPOS SED EXPERIMENTAL START AND TERMINATION It 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. E05-0210 Interim Report #5 Page 7of 65 Page 61 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 IDENTIFICATION AND JUSTIFICATION OF THE TEST SYSTEM The following are the test systems for this study: Water (groundwater and surface water) Soil Sediment Fish Clams Vegetation . Small Mammal Liver Small Mammal Serum The samples will be collected by Weston Solutions. The control samples will be purchased and prepared by the testing facility. Purchase and processing details for the control samples will be included in the final report associated with this study. The test systems were chosen to access the environmental impact o f PFBS. PFHS and PFOS in the Decatur, Alabama area. SAMPLE PROCUREMENT, RECEIPT AND RETENTION Water, soil, sediment, fish, clam, vegetation, small mammal liver and small mammal serum samples will be received at Exygen directly from Weston Solutions. The details o f sample procurement for this study are outlined in the 3M work plan entitled "Phase 2 Work Plan for Sampling Environmental Media." The number and types of samples collected will vary depending availability in the field. The total number of samples received and analyzed fsoturdeya.ch matrix will be documented in the final report associated with this 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 (see Appendix I). These samples will be stored frozen at < -10C. Small mammal whole blood samples will be centrifuged in the field at the time of collection and the serum fraction will be used for the study. Small mammal serum will be stored frozen at < -10C. The receipt and processing o f the samples will be documented in the final report and raw data associated with the study. Page 8 of 65 E05-0210 Interim Report #5 Page 62 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 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 number. Sample storage conditions and locations will be documented throughout the study. ANALYTICAL PROCEDURE SUMMARY References: V0001780: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781: "Method o f Analysis 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 o f Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method o f Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Small Mammal Liver by LC/MS/MS" V0001786: "Method o f Analysis for the Determination of 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 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 fluorochemical sample Page 9 oj 65 E05-0210 Interim Report #5 Page 63 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOOl 13 1 collection at the testing facility and have been shown to be free of 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 spike and a high field spike of each sample will be collected. The low and high field spike bottles will contain PFBS, PFHS and PFOS as well as perfluorooctanoic acid (PFOA) and 1.2-13C perfluorooctanoic acid (13C PFOA). PFOA and l3C PFOA are included in the solutions used to spike the samples. The results for PFOA and l3C PFOA will not be reported in this study. Exygen will supply one field blank (control water) and two field blank spikes (control water fortified with PFBS, PFHS and PFOS at a low and high level) for every twenty samples collected. At the testing facility, each water sample (excluding field duplicates 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 the described procedure to determine method accuracy and to check for bias. For soil, sediment, clams, and vegetation, Exygen will supply one 500 mL precleaned Sci/Spec Premier wide mouth HDPE bottle per sample collected or a zip-seal bag. All containers/bags 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 and processed through the described procedure to determine method accuracy and to check for bias. For small mammal liver, Exygen will supply a 50 mL polypropylene centrifuge 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 liver and serum sample will be extracted in duplicate and will also be fortified at a known concentration with PFBS, PFHS and PFOS at bdoettehrma ilnoewmaenthdohdigahccluervaeclyaannddptoroccheescskedfotrhrboiausg.h the described procedure to Low and high spiking levels for each matrix are defined below: Matrix Low Spiking Level High Spiking Level Water 500 ng/L 5000 ng/L Soil 4 ng/g 40 ng/g Sediment 4 ng/g 40 ng/g Fish 10 ng/g 100 ng/g Clams 10 ng/g 100 ng/g Vegetation 10 ng/g 100 ng/g Small Mammal Liver 10 ng/g 100 ng/g Small Mammal Serum 10 ng/mL 100 ng/mL E05-0210 interim Report #5 Page 10 of 65 Page 64 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Recoveries are anticipated to be between 70% and 130% 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. METHOD FOR CONTROL OF BIAS Control of 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 least two levels of fortifications. STATISTICAL METHODS Statistics will be limited to those specified in the subject methods and to the calculation o f average recoveries, as applicable. GLP STATEMENT All aspects of 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 Representative. 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: The name and address o f the Study Director, Sponsor Representative, and of the testing facility. A statement of GLP compliance (any related documentation, such as chain-of-custody records, must be in the study records). E05-0210 Interim Report #5 Page II of 65 Page 65 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 The signed and dated statement by the Exygen Research Quality Assurance Unit regarding dates o f study inspections and dates findings were reported to the Study Director and Management. A description o f the exact analytical conditions employed in the study. If the subject method was followed exactly, it is necessary to include only a copy o f the 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. The location o f the analyte peaks will be clearly identified in all chromatograms. All circumstances that may have affected the quality or integrity of the data will be documented in the report. Locations where raw data and the final report are to be 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 o f the report that is being altered and the reasons for the alterations. The amendment will be signed and dated by the Study Director and the Sponsor Representative. All applicable requirements for reporting of 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 test or reference substance(s). E05-0210 Interim Report #5 Page 12 of65 Page 66 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMENDMENTS TO PROTOCOL All significant changes to the analytical protocol outlined here will be expressed in writing, signed and dated by the Study Director and Sponsor Representative. Amendments usually will be issued prior to initiation of study plan change. However, when a change is required without sufficient time for the issue 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 soon as possible. In this case, the effective date of the 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 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 chains 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 of study personnel, signatures and initials Chronological presentation of all study correspondence Any other documentation necessary for the reconstruction of the study Chromatograms- All chromatograms will contain the following: Sample identification, injection date, arrow or other indication of 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 sample that was fortified. Analytical standard chromatograms will additionally include the concentration (e.g., pg/mL). Page 13 of 65 E05-0210 Interim Report #5 Page 67 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 As part of the documentation the following sheets will be included in each analytical set: a run sheet listing the samples 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 will inspect the study at intervals adequate to assure compliance with GLP's, and will report the findings 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 wchirthomthaetodgartaampsa,ckwaogrekssuhbeemtsit,tecdortroetshpeonSdtuednyceD, iarencdtorre.suTlhtsessehwalilllbbee ianrcclhuidveedd with the original study plan, amendments, final report, and all pertinent information from the Sponsor. The testing facility shall keep all electronic raw data and any instrument, equipment, and storage logs for the period of time specified in 40 CFR 792.195. An exact copy o f the materials submitted to the study director will also be kept at Exygen Research. Exygen will obtain permission from the study director before discarding or returning samples. E05-0210 Interim Report #5 Page 14 of 65 Page 68 o f 123 3M Environmental Laboratory Project No. EOS-0210 Exygen Protocol Number: P0001131 APPENDIX I ANALYTICAL METHODS V0001780: "Method o f Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Water by LC/MS/MS" V0001781: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Soil by LC/MS/MS" V0001782: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Sediment by LC/MS/MS" V0001783: "Method of Analysis for the Determination of Perfluorooctanoic Acid (PFOA) in Fish and Clams by LC/MS/MS" V0001784: "Method of Analysis for the Determination o f Perfluorooctanoic Acid (PFOA) in Vegetation by LC/MS/MS" V0001785: "Method of 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" E05-0210 Interim Report #5 Page IS of65 Page 69 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMNeAthLodYNTuImCbAeLr: MV0E0T01H7O80D Method of Analysis for the DeterminbaytLioCn/MofSP/eMrfSluorooctanoic Acid (PFOA) in Water Analytical Testing Facility: E3S0txa5yt8geeRCneoRslleeeasgreeca,hrPcDAhriv16e801 Approved By: ___________________ TPaeuchl nCiocnalnLolelayder, LC-MS, E'xygen Research _ !0 |zaM Date V'JiochenPFrlaehsiedretynt, Operations, Exygen Research Date E05-0210 Interim Report #5 Total Pages: 7 Page 16 of 65 Page 70 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygcn Research Method Number VOOO1780 _________________________ANALYTICAL METHOD_________________________ Method ofAnalysis for the DeterminatiLoCn/oMfPSe/MrflSuorooctanoic Acid (PFOA) in Water by 1.0 Scope STacphieidsctmrboyemthHotridgiehisDtPeoetebrcfeotoermrm(apLnlCocye/MedLSif/qMouriStdh)eiCnishworaolatmetirao.tnogarnadphqyuanctoiutaptlieodn otof paerftlaunodroemoctaMnoaiscs 2.0 Safety 22..12 ApCrloewncaasyuuslttiootbhnsese.arpvpersoapfreialateboMraStDorSybperafoctriecehsa.ndling any chemical for proper safety 3.0 Sample Requirement 333...123 33..54 3.6 pASmNSAtAeraanomlitomlmnyjlspueseppaaceatsllemmtsraee.satmspptc4aulplo0neerslledsetplm.oesmtrrchLocetuecoidoosenstsfnutsbrtapeeipeinefnsrrgrttoiihnngcsioagaseetrmdranopauneupatrteeglreedudthsiesclfeydwolsedrhmifsleoofliosxruxhbrtwlerodetadhaucsetlbtbepdieoereefxbnscota.eirrafmeailcelcpbeodtnlieweotiisrnnen.id.gfuthsgateeomdspaealmqetdup~ill3fiion0brg0rae0pxteltrarpanmtcotfiooforrnor.toh~mi5s 4.0 Reagents and Standards 4444....1234 MAPWemearfttmhleuaroon-nrooiHulom-PctLHaACnPcoLegitcCraaAtdgeecr-aiddAe-.CS.iSg.mRae-aAglednritcGhrade 5.0 Instrument and Equipment S.l $55...324 AspSAAoOLnldvamhceleiovyLgnnithtcidnsceieaespctlpqoetbeourcafdsioolpaatlrpbonlmeelceadcaetpntwcarocaanilewtpydhapeldbirmaqoaleutpvMaiyaodrflafeiosarncrsebhepalSredecopaeimvnekncogtatirlnrttuioooftmuegm0ggre.eare0atpi0entthuir0joeb0(nccLe1atsCao.pgnr/a.MdbcaqlSepu/MaaobnSlftei)t.paotuifmoinnp.jiencgtinugpS-to2002 5.5 IS m L disposable polypropylene centrifuge tubes. 5555S5......17869110 2DDAW12miiusas5tpptoL-eoomprscssiLpaaleSbebLaetllerDtepeHmpsPPiPE(apic1kLern0toCVta0epr-avsri1p.cio0aew60lts0k-cmic(pt5.o(L0ul-gat1hn)OdbtOCo1ut1t0L8l,-e1sS10.0P00Ep-2cL0a)r0,turwiLdi)gt.hesd.isposable tips. Page 2 of 7 E05-0210 Interim Report #5 Page 7 of65 Page 71 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Hxygen Research Method Number V0001780 _________________________ANALYTICAL METHOD_________________________ Method ofAnalysis for the DetenninatiLoCn/oMfSP/eMrfSluorooctanoic Acid (PFOA) in Water bv' 5S..1132 CSPenEtrviafucgueumcapmaabnliefoolfds.pinning 50 mL polypropylene tubes at 3000 rpm. 6.0 Chromatographic System 6.1 6666....3452 TGAMM(Pern/oomaNabbdlp:iiyillee8teenir2caPPt5athhPul0aarr5Coess-ee:og0lr((5u3aAB2mm01))n:3C:::0F)l2MuomepthMhaasnAeomRl mP o(KnieuymstoAnceeStactieenintiWfica)t,e2r.1 mm x 50 mm, 5p Tim2208e120....(.00050mini %6622655555A %3733755555B FfmloLw00000./....33333mRiant)e 666...786 QIRnuujeancnTttiiiotmanteiVo: no-:lu2Pm3eeam:ki1nA5urtepeasL.-(ceaxntebrneailnsctraenadsearddtocaalsibmrauticohnacsu5rv0ep. L). TophteimabizoevtehceoHndPiLtiConssyastreemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 Mode: Electrospray Negative MRM mode, monitoring 413 -369 m/z. TophteimaibzoevtheecMonSdMitiSonsyssaterme .intended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 Mobile Phase 8.1.1 2ammmMonaiummmaocneituamte taoce1ta0t0e0imn Lwoaftewr aitserp.repared by adding 0.154 g o f Alternate volumes may be prepared. Page 3 t>f ~ E05-0210 Interim Report #5 Page 18 of65 Page 72 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOOI780 _________________________ANALYTICAL METHOD_________________________ Method ofAnalysis for the DeterminatiLoCn/oMfSP/eMrfSluorooctanoic Acid (PFOA) in Water by 9.0 Standard Preparation 9.1 999999S......t111111a......n615234darmTmaAAAmmmaafAoi1Pdnrp0hrfeo1eLLL1Spe00ta1e1ma2t2hprt.m.h.0noo155oo0oas01aaattnc1fxfL2frpmhmnopplkoteti5ytgehochmgglhp/LLt/oakFlme/e/diiemgammncfoiLaL/astL1na1Lmr1ettnLLaDDt.lt0eol00haidLyLcfsfPP01efofpoipDtk1foo2EEca4gfrofpg2r5ranP0to/pstrtg/bi5bmt.iCdoitmErfrmi1f/fo-iotieoalmifimiLcuttLcpbcLirfnattacdiptaalLalLonsciteeasStLtgroitdoaii(.o.toatsLoo/onDcillttlmnooeuilannDoiuouPoo.lnrtrtLusnPitsesnEirfioosootoeGo.i~solnslcnblsonutuou1lotboantutetltt0oilbtooiuitdouto0otiltltoanoteateinnfilopn.oeofoaffoonsgi.nironfnff/farpoamaiPPPtrnullfaoeFFFLarvvPOlimtOOootoaFoyvAlAlAaOuf)uobxfmmlePAiiaiiiusnssmnFesemapdpptOiluoosorerrmAedrefefvpelpoppiOold1raaapbfuel0rOrrueytpie10eeenrmad0ddiwwwrota0eeobbbdeiidrtwtyiyyheoh1gofib0bbbhftrfmmyh0rrrii6gniiiee1nnnmbmegtt0hhgggmrr0eLaaaiiii1ntnnnonnth0owggoogwgnarilltmniinhH1t1taomih1gh0ngstl1i 9.2 Standard Calibration Solutions 9.2.1 9.2.2 pTciLdarhCeeleinp/bMtafrirocaSeatldil/looMtanwosSissnntacegamaenliddpisbealedrarasd..ttsiyopanircesatlparneoxdcaaemrsdspseleda:rteahdrpodruietgpiohanreathdl ecionenxcHtreaPncLttriCaotniwopnarstoemrc.eadyuTrhbeee, Final Concentration Fortification Volume o f Concentration o f Calibration o f Fortification Volume Fortified Control Calibration Standard ID Solution (ppb) (UL) Sample (mL) Standard (ppt)* (example) 0 0 40 0 XCmmddyy-0 10 100 40 25 XCmmddyy-1 10 200 40 10 400 40 too too 40 too50 XCmmddyy-2 2SO XXCCmmmmddddyyyy--34 100 200 40 500 XCmmddyy-5 TcXohCnec=ee1xne0ttx0rratarctaitcoetnde,dcdocunaeclietbon4r0tatrh0taietoincoonsntoacnefndthtareradct.i4ao0lniborafttihoen ssttaannddaarr1dd00di0sureiqnugatlhteoeX8xCxtrmiatcmstiidnodintyiya(-Sl6PE). Page 4 of ' E05-0210 Interim Report #5 Page 19 of 65 Page 73 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1780 _________________________ANALYTICAL METHOD | Method of Analysis for the DeterminatiLoCn/oMfSP/eMrfSluorooctanoic Acid (PFOA) in Water by 9.2.3 9.2.4 9.2.5 anAsASetetlote2tzedroeereCfrndoasa.tlttolasetne6avdxnoatCdrrlaaud,crmsudtpeeedxsstootcarlaanutwcldtitiboeocrdnowa.nt(eicroeeenkansgst.retaantntidobanlrsadnoskfi)nstma1nu5ds-mat rbLdespmporlaeyypparbroeepdpyrlweepnitaehrteeudabceahss 10.0 Batch Set Up 10.1 10.2 ipREocnornaeontecqhcthuereroiderbqlueasaumrgtaacefellhniontrtryetotscicaffofiosseovsnaduertmrrrfyoaiapenltfllceod(eskrmapntehneloxatedwthnrbolanafadcobttccreohbodtrh.lanait(cnsoteykrpnyprtiourdcajsauetilicpnoltlyg.nics2aH0t(ePlsoaLrbaCnledcswossna)pttiremkore)lussatswnpiidnilklceltbulwedotesopareetvcaielgferieaienlsdyit 11.0 Sample Extraction 11.1 11.2 1111..34 11.5 AEMCmfLi1no5loonitluexoalnamatodldesw5wyLius0ztweeraipeloedmmilot)ns4hbl.paLy0ymtlp-hepm55rpeooolmLnlepCymsyLcpouLulorfeoosnnSsipf1dnaePHy0imgtcE0liPeeope%nnLlncleeetaCercmrdoicttfrwerreuCointagdahstuetgprapeierntSofrausiorP(yibtl~g(Ei.e1Leos2ncCtCg(aud/fo,rMbroitnfoerl6lpiaSssedal//mcgs(MmvfteeLoocp.S5rl))l.ut.eDimbmfdDyiysLeicolapu=asonatre5sdfnodsetemeienlltleuoduLgtaae)4c.tdt1ee0o,0.lrmiuenmmpLtoLlnawcgrmieurtanheldtidhduwraayaanttenoeddlr 12.0 Chromatography 12.1 12.2 12.3 12.4 Ue(cgAfLmbiISantonxaeneotCsntuljlantgaeeleir/snoabeliraMctynedscrwrctlabzpnnacaitStetenteeetiorheii/ndddreduorMdeisgsaennnsebrsfcSocatdyosaasalmfftsumnrelbaoPtesipdtdtanberyehxFlentdresoteawdOaaadrtasfeatmta.arsieAimcdnnoreeeottdscanhne.xcauoldmeontynnecrsArttauvautdcreeacoensearnnbtvstscarlfdeidpeyyeablbedytaoslearloi5tronaibcidct-nfcfdthhrfIasioeaaOioeleanalrstinxmibrtgfsosatsiqrrearanrsntaaeuasomtytmdtgataciasrnopantapbetertdnnltadldseeiaeldnts,searidss.asditlosttaniaasasaenorsjtmAnntdent.iufddcsnp.iisanatvtlmieareerjenEdddenrugLasocsxsspnta)irtatlin.reldmt/omaterxenpcsfaIutroterohnwrnseierdrentadeceetstieiiaecfitbbtgdvaisvohseeehtneeenaeagdt,drrmcniiiynanasaendcrpgannnncaad5mldltirsoeu-rnede1apfndcgss,tf0lupeteioeieotdrerom.lsxvnealiafonteukarmwtslsasateoeatcpvtrtatltseeeahbheorealdleseeesilf Page 5oi7 E05-0210 Interim Report #5 Page 20 of 65 Page 74 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1780 _________________________ANALYTICAL METHOD_________________________ Method of Analysis for the DeterminatiLoCn/oMfSP/eMrfSluorooctanoic Acid (PFOA) in Water by 12.5 evsSoxeafcrmtsewupesadlercseatraseliynsbspdrtoaaerntmdiso.ernessshptaoonundlsdaersdnsochtooneucxldecnebteerdaftusiotratnhneudrsaidrndilgurtMeesdapsoasnnLdsyernse.xan3Aa.3lnyyz(oesdra.emqupilvesalethnat)t 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.5 13.6 Ra2asTHeiAtewERbm1odMCnhtnpx0nelh3aarf4oeajnheaupc.terteun0aecrw94iynrieoslrt%crnrdg%roouh9ont1kareteaahcvmcdekob23ptnvlriowTatcedendsryetodesealorreiad.eeothi(nirrstairodimnbttbRsoe,weocthiob.ttmlersgoloaJu,utaaitatanrinrhnlailnfo.utmtmandst2iieirvkhs(afmonotssu0e3Tetaasneanenctbn.spl6yPom9hdomnuremas9tFtse8enboecnahpbatuluO5stoeeatlaaeeln4emrs.s)etvmnelvnoA1detw.tnysafdueal3xustuIfttnleaxsnishaihchusf)mcotecrchatpteeIoeadaarnitmlosfeibeetwlbluskeeeecudcpnefueedrcnsedtvooolrrtutaatedsteo2ennuapnimldotlbspr(0tifntnaaa.sbeabRrefye%uksdndrofienrgeaaIsn)ersadelatrmneofktttnrxsrteohostdmPrtmibeftapopoeteosmr-fFcehttbtalianefrooeeekOetttucrexaarahhrnrseetnscntArlireeetataraoxtseedaadihafdndhossslblaacatcsaisyasupjonuoelbtntsuactpllsoleauotgayrslatsrtitadtlcnla5smikhezfltiedmlo.utids0e-tetcvibpoteuelsfiodbsaneeiarnnalrxtcn.Aeraluetsmtlsgddtbsaiiotlismrtornoi/dlreorauLeocginynutoetnbcfmaosunerh,mttuetneruraetedmoutertvmhaatbeshdtisxsflltePeaotnie.oe3iyetiacsdtexnFttnr6zrnfaehberineiOe9ceddxetgeaoeodhbcdaetAetxpait.wneayh.srcsnftmednpeaeprdeetraasuwtlrshiuarreniatekcesaibithnbitf-tsieitfborheeotenleralellnad7xnogdoiaitmtLm,tmi0m,unlrostiO-wcaktoihmimnt1astotcaarQse3ihnsetuiunwd,ci0itda.psldoslHeuimd%atett,tanhhrruhIt7rvptetaehbgfbidhoh0enlnnineeeeseeseaft 14.0 Calculations 14.1 bgUaessneeedtrhaoetendfopblleyoawtkhienargMeaea)qsusuasLtiiynongnxtthsooefctsawtlacanurdleaaptrerdotghcruearmavme: o(luinnteaorf rPeFgOreAssifoonunpdar(aimn entge/rLs). PFOA found (ng/L) = (Peak aresalo-pientercept) x DF DF " factor by which the final volume was diluted, if necessary. Page6 o f7 E05-0210 Interim Report #5 Page 21 of65 Page 75 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Retearch Method Number V0001780 _________________________ANALYTICAL METHOD_________________________ Method of Analysis for the DeteiminatiLoCn/oMfSP/eMrfSluorooctanolc Acid (PFOA) in Water by 14.2 tFhoerfsoallmowpliensgfeoqrutiafiteiodnwtoithcaklcnuolwatne tahmeopuenrctsenotfrePcFoOveAryp.rior to extraction, use Recovery (%) - [ total analyte found (ng/L) - analyte found in control (ng/L)] Qp analyteadded (ng/L) E05-0210 Interim Report #5 Page 7 of 7 Page 22 o f 65 Page 76 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 ANALYTICAL METHOD Method Number V0001781 Method of Analysis for the DeterminaLtiCon/MoSf /PMerSfluorooctanoic Acid (PFOA) in Soil b'v Analytical Testing Facility: E3S0txa5yt8egeRCnoeRslleeeasgreeca,hrPcDAhri1v6e801 C-jLApproved By: PTaeuchl nCiocnalnLolelayder, LC-MS, Exy>gen Research Date Date E05-0210 Interim Report #5 Total Pages: 7 Page 23 of 65 Page 77 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1781 ________________________ANALYTICAL METHOD________________________ Method of Analysis for the DeterminaLtioCn/MoSf P/MerSfluorooctanoic Acid (PFOA) in Soil by 1.0 Scope TaScphieidsctmbroyemthHeotirdgichisDtPoeetebrfceotoremrm(apLnlCocye/MedLSif/qMouriStdh)eiCnihssoroolialm.tiaotnogarnadphqyuancotiutaptlieodn otof paerftlaunodroemoctaMnoaiscs 2.0 Safety 22..21 pACrolewncasauuyltstiootnhbses.earpvperosapfreialtaebMorSatDoSrybperfaocrteicheas.ndling any chemical for proper safety 3.0 Sample Requirement 333...132 33..45 pSAAtNSeraamltoomlmjlpseesppaacaellmmtsree.atspptc1ullo5eerslestplg.oemrrcooetucfidosettesnsbsretipenfsrrtgaohimgcioseerpdronlaueuetreegfeddohselrydwesmxhtioltolriruxabslcedeotdiioslbbnpsee.eafcmoirafpeillelbeodsew.iinendgtshateomspaelmqedupilfliionbrgreapxteltarantcotfioorrno.tohmis 4.0 Reagents and Standards 4444....1342 APMWemeartftmhleuaroon-nroioHluo-mPctLHaACnPcoLgeiCtcraaAtdgeecr-aiAdde-.CS.Sig.mRae-aAglednritcGhrade 5.0 Instrument and Equipment 5.1 555555......725634 D5AA1sAp1o052Linlds5vmmahpce-leimoovyLLgnnsitLhtciaddnscebiieLaesspltclppeDqoetbooeurmPcafdssiooEplaaiatclrpbbolnmnreelloceaeadcapertppnrtiwcaroopocaanlweleiwyytpdths-ppaemldbrr(miaqoo5aloeup0ptvuMai-yyoatd1hlflrafeeoiOsarbnncrsOeboehepaluStredcectLopleaei,emvennkns1cogtt.ait0rrlntruiit0ooffotmuu-egm20gggre0a.eeer00pati0ettntuhuui0rjLobbe0(n)ceeLc1.ssatCao..pgnr/a.MdbcalqSepu/MaaobnSfltei)t.paoutifmoinnp.jiencgtinugp St-o2002 55..98 2DmispLocsalebalreHpiPpLetCtevs.ial kit. 5555....11111320 AWUSPulatEtrotaepvsroisapncSeuietcutpembsPa(tamh1k.0aVn0i-af1oc0l6d0.0ccp(Llga)ndtC1108-1S0P0EpcLa)r,trwiditghesd.isposable tips. Page 2 o f7 E05-0210 Interim Report #5 Page 24 o) 65 Page 78 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygca Research Method Number VOOO1781 ________________________ANALYTICAL METHOD________________________ Method ofAnalysis for the DeterminaLtiCon/MoSf P/MerSiluorooctanoic Acid (PFOA) in Soil by ] 55..1154 WCernistrt-ifaucgtieoncasphaabkleero. f spinning 50 mL polypropylene tubes at 5000 rpm. 6.0 Chromatographic System 6.1 6666....5423 MTM(GAPernoo/maNabbdlpii:yilleeee8tnir2caPPta5thhPul0aarrC5sose-ee:og0lr((5u3aAB2mm0)1):n3C:::0F) tM2uoempthMhaansAoelmRmPo(KnieuymstoAnceetSacteieinntiWfica)t,e2r.1 mm x 50 mm. 5p Tim2208e120...f.0.0050ain) %6622655555A %3337755555B FfaloLw00000/.....m33333Riante) 666...876 RIQnuujenacnTttiiiotmanteiVo: no-:lu2Pm3eeam:kin1A5urtepeasL.-(ceaxntebrneailnsctraenadseadrdtocaalsibmrauticohnacsu5rv0ep. L). TophteimdirzoevtehceoHndPiLtiConssyastreemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 MPFoOdAe:. Electrospray Negative MRM mode, monitoring 413 -+ 369 m il for oTphteimabizoevtehceoMndSiMtioSnssyasrteemin.tended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 Mobile Phase 8.1.1 a2mmmMoniaummmaocneituamte taoce1t0a0te0 minLwoaftewraitserp. repared by adding 0.154 g of Alternate volumes may be prepared. Page 3 of" ' E05-0210 Interim Report #5 Page 25 of 65 Page 79 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1781 ________________________ANALYTICAL METHOD________________________ Method ofAnalysis for the DeterminaLtiCon/MoSf /PMerSfluorooctanoic Acid (PFOA) in Soil by 9.0 Standard Preparation 9.1 9999S99......t111111a......n615342daraTmfAmamaAAmmiA1oPdrnp0hrofe11LLeSLpe00aet11ma22thrpt.m.ho.0nSo13oo0osa01aatfatn1cxfL2fuhrmomnfpplkoei5tghtyecomghglph/LLo/kteaFlmd/e/ieigmammncfoiLaLas/t1Ln1amL1eitntaLLDDt.0telolh00adiLyLcff1sPP0oepffiopDtkf1oo2cEE4gaforfgp32rra0Pntpo/srtt/gibb5tm.CiiodtEmrrmfi1/ffooi-etiolamfiimLicuttLpcbLcniafrttacpidtaaLallonLsciteeasSLttrgotidoaii..ot(atooLs/ooDnillcttlmnooeuailnniuoDouPoo.nlrtrtLsuintessnPEifrioosootoeo.Ei~noslsllbncsonutuoul1otaobntutetltt0obiltoouiitdoutoot0iltltaooentateinniflonp.oeoffaofoonsi.gniornfnff/faropamaPiPPrtnlulfaoeFFFaLrvvPOimtlOOoootaFoyvAlAAalOufb)uoxfmmePilAiaiiinusmssnFseaempdtppOiulooosrrermrAedeeffvpepoppiord11lpaaabfeul00rrerupyit0e10eenermad0didwwwroat0eeobbbdeiidrttwyyiyeohhg1ofbibbbfh0rtfmmyhir0rrig6niiiee1nnnbemmgtt0hhggmgrr0eLaiaaiii1ntnnonnnth0ogwoongggwarilltmniniht111atioihgsnhng00t0l 9.2 Standard Calibration Solutions 9.2.1 9.2.2 TpciLdarhCeeleinp/bMtafrirocaSeatldil/looMtanwosSissnntacgeamaenliddipsbealrdearasd. tt.syiopanircesatlparenoxdcaaemrsdspseleda:rteahdrpodruietgpiohanreathdl ecionenxHctreaPncLttriCaotniwonpasrtoemrc.eadyuTrbheee, Final Concentration Fortification Volume o f Concentration o f Calibration o f Fortification Volume Fortified Control Calibration Standard ID Solution (ppb) (tiL ) Sample (mL) Standard (Dot)* (example) 0 0 40 0 XCmmddyy-0 to 100 40 25 XCmmddyy-1 10 200 40 50 XCmmddyy-2 10 400 40 100 XCmmddyy-3 100 100 40 2S0 XCmmddyy-4 100 200 40 500 XCmmddyy-5 * cXTohCnec=ee1xne0ttxr0ratarctaitcoetnde,dcdocunaeclitebo4nr0tatrh0taietoicnoonsntoacnefndthtareradct.4iao0lniborafttihoenssttaannddaarr1dd00di0sureiqnugatlhteoeX8xCxtrmiatcsmtiidnodintwia(-Sl6PE). Page4 of7 E05-0210 Interim Report #5 Page 26 of65 Page 80 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygea Research Method Number V0001781 ________________________ANALYTICAL METHOD________________________ Method of Analysis for the DeterminaLtiCon/MoSfP/MerSfluorooctanoic Acid (PFOA) in Soil by 9.2.3 9.2.4 9.2.5 AnsaASetetltoet2zedoreereCfrndoasa.ttltoalsente6avdxnoatCdrlruaad,crmsudtpeeedxsstotocarlaauntwcltdiitbeoocdrnowa.nt(eicroeeenkansgts.retaantnitodbnalrsadnoskfi)nstma1nu5ds-atmrbdLespmporlaeyypparbroeepdpyrlweepnitaehreleudabcaehss 10.0 Batch Set Up 10.1 10.2 RocpiBnorneaonteqchcthurereoeidrbqaleusagumrtaeacfelnlhointtrryetoctscioaffofisnesovstadurremorrfayliapen(tlflcmeodeskreapnttenhholxadeowtnrdblnaafacbobttlccroeahodtrnh.ankit(csoteuyrpnsypritionrdcajgauetlipc5lotyl.nimcs2aL0te(lsooarbfanmledcesostsnh)ptairmnkooelusls)tswapiniinlkdlcelb)tuwedtoesopareetvcaieglferieiaenlsdytt 11.0 Sample Extraction 11.1 111111...243 1111..56 11.7 11.8 11.9 EAceLCfABCTnIWoSleeloronuredlunaeiealnanamdnttodnidtlterdggewyresiL5ihif.fdztw(feueimptdeu,odh5grioegrtnsLeetehcbleahgypatyomtevupf-nlhooafob5r5tpteocur)fmlelmuepeb-.Csmte1msyeahlLtiislm0LhsuedeeoastnpmSoanisf1uelnnaPoeHa0ipgmdnclEn0Piueea%mtpnuLntolncleteltiaeoCdtsrxcmrr4itatsfa5orww0reuhsto0nitogdaea-shmnlke3tpgmatleihe)re0ntLrc.aLuseo0o(ybbl~0nw(.pace1Lootsr2aihCptlnCg(hwdym/fda,rpMoinrion.wltri6dilspaoSoeatl/p/mscnstMavoyeteecLonrlcdSte5li))i.uncoi.nCmbaemntDyueLcestohehfp-neSoaonatkrP5sfor5es-iEtemfi01rnullf5euLcggomtaa)emcrt.rL1e-ottir01ulniuidbp5unmmgeottemoselLnys.i.(pgrnfmurrouoaDnertdpettidhiuyssfra.calyyenatenroaddels 12.0 Chromatography 12.1 Inject the same amount of each standard, sample and fortified sample into the 12.2 12.3 bmSaiALnentCnutagaens/lirMytnedszbnpnaSeeietrni/ddrrMisgsensecaSodaslmfunebPsdpdteyFeltesowdOatsfete.iAmneetnh.axcenoterArvaareecnenrtsacyedplady5oloit-nbic1fcdra0aaiallntisibgsoasremnatato.mtpisoalptetnalseln.esdatsAasaetrnstd.fdaiavnmreEdauoxslsttretrmmranpcuoatrsetreitedcvecebsdo,etenaanncineadenncanldtrurtdiadrstfeeioodmslnleoutalwsteotvftebhalleesl Page5 of 7 E05-0210 Interim Report #5 Page 27 of 65 Page 81 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VQ00178! | ANALYTICAL METHOD Method of Analysis for the DeterminaLtiCon/MoSf P/MerSfluorooctanoic Acid (PFOA) in Soil b'v 12.4 12.5 sSgveef(cUootxxaeeaosfllnrctmletirseoeawbaupercwrclsaadlaciteretentoceesiddeauodtrsaalennbiynrfcsbtyosspdarfttsrloaaeaoitetrtnmbanrhxidnsordte.aeardnraateaarsiscsdsonrehpttdscnaeaoooldmnuyncnsdltsuutcddeaaearsrnsvbstldnideeybsbotacshertlroaooidtnntefuhfsioexcoleedraecnmxrfnebtqietrsrarerueasdtyaatgacftnuntarisbeoedtnrtadeistnadthnsarsieuddiltotnairssaaonjsidrnenntdiuidcn.gilsnutatreierjMtendeedrsLgscdsappt)iasoian1.eston/nerLxnssdaItyhenerwindrsenee.exsaaieitbtgnav3hesAnhae.aeg3ldtrnmryiiyannyz(crpngoead5lisdrseo-nae.1ecgfms,0quepeuptroe.sxvilavfaetekrmassaalacepthrtnalseeearetdacs!tt 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.5 13.6 RsaTHwAEeiteRdbmMCo1nthp0xnalaher3foeanhjeaup.certuntee0c9rwy4riinoeslrtcrdgr%roouh9ot1nkareteahcvmcdkeo2p3nbtvrToaitcdeednsrtedoeesaleorrid.aeeohir(rstanriodnbmbtsoR,eewoti.botmtlresgloou,uaattJaatnrhirnailno.futtmmadntsieiivkhsm(foonst0se3Tataeseanncteb.sylp6Po9mhodmnumers9tFs8eenboecabptauulO5satotaeleelen4mrs.s)nemtvevAod1tne.twsfdaueal3xuusfIntlaxsenisahhcfsum)torcecahttpeoIeadarintmlobsfeietlewubkeeeeccduenpfdeuncdevstooorltraeatdtseonen2upaidoltlbrsp(t0tnfinaaasbearRebfye%ndksrdofinegraeas)nrdeealatronkemttnxrsrtehoostPmmtdieftaotppoeormsf-FchbttaalinefroeoekOettcetuaxaahrrhrsetnsctnAlirreeeataroxtdseeaahdfdadhsssbalalacsctaysisjupouleotbancuttpslsaouleogtslaarstritatlcdnl5askhemtlfiedloui.tsd0et-tcvbiotuelepssifobnaeiarnnarxtlcnAraeultsetmlsgdtsabiiotlimsrtoorni/ldeorauoeLcgnnyutteobncomsaunehr,mtuetenurratemoedturvmthaatshtbdeisfsltetPlaoen.eieio3eayitsdtextnFnt6rnrzafhberineiO9ecddeaxtgeoeobhdcaeAtextpaite.wnayrc.hsnfmtedpnaerpeederasalutwrsiuareiarnetcksebaithnbit-tifieftrbhoeneeolteraeall7dnxogndotiatL0m,itmm,unlrots-iOwcatkhoimm1ntsatocaaeQr3shintsnuuied,ci0itpad.dsloHsleu%dmateta,tnrhuhrrIv7ttpetaeghdbbfhoeh0nlinnoeeesese.aeft an4a%nalwytiitchainl raunnatnhaelnyttihcealserut nm.uIsftrbeeternetainoanlytizmede.drift exceeds this limit within Page 6 u i7 E05-0210 Interim Report #5 Page 28 o f 65 Page 82 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1781 ________________________ANALYTICAL METHOD________________________ Method of Analysis for the Determination of Periluorooctanoic Acid (PFOA) in Soil by 1 J LC/M S/M S ' 14.0 Calculations 14.1 bUgaessneeedrthaoteendfopblelyoawtkhienarMgeaea)qssuuaLstiiynongnxtthsooefctsawtlacanurdelaaptrerdotgchruearmavme: o(luinnteaorfrPeFgOreAssifoonunpdar(aimn entge/rLs). PFOA found (ng/L) = (Peak aresalo-pientercept) x DF DF " factor by which the final volume was diluted, if necessary. 14.2 Fthoerfsoalmlopwliensgfeoqrtuiafiteiodnwtoithcaklcnuolwatne tahmeopuenrctsenotfrePcFoOveAryp.rior to extraction, use Recovery (%) * [ total analyte found (ng/L) - analyte found in control (ng/L)] ^ analyteadded (ng/L) 14.3 nUgs/egt(hpepbf)o.llowing equation to convert the amount of PFOA found in ng/L to PFOA found (ppb) " fPFOA founsda(mngp/lLe )wxeivgohltu(mSege)xtracted (0.04D1 14.4 bUasseedthoenfdorlylowweinigghte.quation to calculate the amount of PFOA found in pph PFOA found (ppb) dry weight - PFOA found (ppb) x [100% / total solids(%)] Page 7of 7 E05-0210 Interim Report #5 Page 29 oj 65 Page 83 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMNeAthLodYNTuImCbAeLr: MV0E0T01H7O82D Method of Analysis for the SDeedteimrmenintabtyioLnCo/fMPSer/MfluSorooctanoic Acid (PFOA) in Analytical Testing Facility: SE30txa5yt8egeCRnoeRslleeeagsreeca,hrPcDAhri1v6e801 Approved By: PTaeuchl nCiocnalnLolelayder, LCC-MSJ, Exly.gIen Research _D_at_elo lb M / y4dVoiZhcJ'ilne PFflr?laehsiedretynt, i/. Operations, Exygen Research Date E05-0210 Interim Report #5 Total Pages: 7 Page 30 of65 Page 84 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V000I782 1 ANALYTICAL METHOD ] Method ofAnalysis for the DeterminatioLnCo/fMPSer/fMluSorooctanoic Acid (PFOA) in Sediment bv 1.0 Scope TaScphieidsctmrboyemthHeotirdgichisDtPeoetebrcfeotoremrm(apLnlCocye/MedLSif/qMouriStdh)eiCnihssoreoldamitmiaoetnongta.rnadphqyuancotiutaptlieodn otof paerftlaunodroemoctaMnoaiscs 2.0 Safety 22..12 pACrolewncasauuytlsitootnhbses.earpvperosapfreialtaebMorSatDorSybperfaocrteicheas.ndling any chemical for proper safety 3.0 Sample Requirement 333...312 33..45 tpNSASAeraamlotomlmljspseeppaaeaclmlmrseetatsppt3cullo0eerselstpgl.oemrrcooetucfdisoettesnssrbeitpenfsrrgtaoihmgcioseerprdonlaueeutreegfeddhoselrydweshmxfilootlriruxablscedeetdidosbibnpmee.efeconiraftelilesboadewmiinenpdgletshsat.eomspaelmqedupillfiionbrgreapxteltarantcotfioorrno.tohmis 4.0 Reagents and Standards 44444.....12543 AWMPAemcearetftmhtleiuarcoon-nArrooiHluco-imPcdLtHa-ACnPRcoLgeiectCraaagAtdgeeecrn-aitddAge-.rCaSd.iSeg.mRae-aAglednritcGhrade 5.0 Instrument and Equipment 5.1 5555....4235 5AAAsp1o05LnldmvmahceleiovyLLgnntihticddnsceiieaspesctlppqoetbooeurcafdssioolpaaatrlbpbonlmleelceeadcaepptntwcaroocaanlleiyywptdhppaeldbirrmaqoolaeuppvtMaiyyaodrllfafeeiosarnncrsebheepalSrecdecopeaeimvennnkcogttatrriltrniiutooffotmuugem0gggrea.eeer0ptai0ettnthuuri0jobbe0(nceeLc1asstCao..pgnr/a.MdbclaqSepu/MaaobnSfltei)tp.aoutifmoinnp.jiencgtinugp 5t-o2020 5555S55.......819176l20 1D2ADS12Piimuss5WEtppLo-oomvpascstaiLaepalcerbebuLastlluerteDSemHpsmePipPE(mpi1cLPean0rtCaonta0ekpir-svfr1iVop.oi0alewad0ltc.s0-km6i(pt5.ocL0ue-tal(OhnlgdbOo)u1tttL0Cl-,e11s108.00S0pP-2LE0)0,cuawrLti)rt.hiddgiessp.osable tips. Page 2 of 7 E05-0210 Interim Report #5 Page 31 of65 Page 85 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001782 ANALYTICAL METHOD Method ofAnalysis for the DeterminatioLnCo/fMPSer/fMluSorooctanoic Acid (PFOA) in Sediment bv' 555...111354 VCWeornritstert-ixfaeucrgt.ieocnasphaabkleero. f spinning 50 mL polypropylene tubes at 3000 rpm. 6.0 Chromatographic System 6.1 6666....5234 (APn/Nal:y8ti2c5al05C-o0l5u2m1n3:0F) luophase RP (Keystone Scientific), 2 GTMMeroomabbdpiiilleeeenraPPtthPhuaarroessee:gr((3aBAm0)):C:: M2 emthManAolmmonium Acetate in Water .1 mm x 50 mm. 5jj Tim2208e120....i.00050minl %2626655555A %3737355555B F(mloUw00000.....m33333Riant)e 666...876 QRInuujeancnTttiiiomtanteiVo: no-:l2uPm3eeam:kin1A5urtepeasL.-(ceaxntebrneailnsctraenadseadrdtocaalsibmrauticohnacsu5rv0ep. L). TophteimabizoevtehceoHndPiLtiConssyastreemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 PMFoOdAe:. Electrospray Negative MRM mode, monitoring 413 -- 369 m/z for TophteimabizoevtehceoMndSiMtioSnssyasreteimnt.ended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 M obile Phase 8.1.1 2ammmMonaiummmaocneiutamte taoce1t0at0e0imnLwoaftewraitserp.repared by adding 0.154 g of Page 2 of 7 E05-0210 Interim Report #5 Page 32 of65 Page 86 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001782 [ _________________________ANALYTICAL METHOD__________________ j Method ofAnalysis for the DeterminatioLnCo/fMPSer/MfluSorooctanoic Acid (PFOA) in Sediment bv' 8.2 Extraction Solutions 8.2.1 11%000acmetLicoafcwidatienr.water is prepared by adding 10 mL of acetic acid to Alternate volumes may be prepared. 9.0 Standard Preparation 9.1 99999S9......t111111a......n564132darmafTmamAAAmmaoi1APdrnp0hrofe1L1LeLSpe00te1am1a2t2hpr.tm.h.0oo1n55ooo0sa01aatatfn1cxfL2pfrmhmonpplotekti5gytechohgmglphL/Lt/okaFelm/e/diiemgammcnfiLoaLas't1Lna1mL1tietnLLDatD.l0etoh00ladiLLycfsfPP10oepffpoitDk1ofo2EEc4agffogrp25rra0nPotp/sttr/bgb5imt.CiodimrtErmf1ifof/o-iteoilaimfimiLctutLcpLcbinrafttacpditallaaLonLsiecetasStLtgrotdioi.a.io(tatoLo/osoDnicllttlmonoeualinniuDouoPoo.nrltrtLsuisetnsPnEirfoiosoooteo.E-ionssllnlbcsonu1tuoulotaobtnutt0elttoilbotouiitdout0ootiltltaoonetateinnfilonp.oeofaoffoonsgi.inronfnfff/arpoamiaPPPrtnlulfaoeFFFaLrvvPOilmOtOootoaFoyvAlAlAaOuf)uboxfmmePliiAiaiiunssmsnFesemappdptOiulooosrrremAedereffvpeppopio1r1dlaapabfuel00rrreuypit00e1eenermadd0diwwwroat0eeobbdbeiidrttwyyiyehhog1ofbibbb0hftrfmmyhrr0irigi6iniee1nnnbmetmtg0hhgmggrr0eaLaiaiii1nnotnnnnth0ogwogonggwarliltmniniht111aiotihgnsnh0g00tl 9.2 Standard Calibration Solutions 9.2.1 9.2.2 TLoCfhte/hMefoS0l/.l1MowpSignc/gmaliiLsbrfaaottriytoipnfiiccsaatatlinoednxaasrmdoslpualteiro:enap.dredpitairoendalincomnectehnatnraotliovnias dilution may be Concentration o f Fortification Solution (ng/mL) 1to0o0 100 10 5 2 Volume (m L) 150 2 10 10 10 Diluted to (mt1o0Lo0) 110000 100 100 Final Concentration (ng/mL) 10.0 5.0 2.0 1.0 0.5 0.2 Page 4 of 7 E05-0210 Interim Report #5 Page 33 o f65 Page 87 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOO1131 Exygen Research Method Number V00I782 | ANALYTICAL METHOD Method of Analysis for the DeterminatioLnCo/fMPeSr/fMluSorooctanoic Acid (PFOA) in Sediment by 9.2.3 9.2.4 nSaAtetloe2tedrereCndaa.tltolec6vaolCilbu,rmuapteisotonanssidtxacnmodnoacnrdethnssti.rnati1o2n5s-mofLstLaDndPaErdnsamrroayw-bme opurethpabroetdtleass 10.0 Batch Set Up 10.1 10.2 RociEnonaentecqhchueeiunrbqentaurmttaarcetlehiianottytonesdsaffs(soslacaurmobrfanipcentlorlcedonesltapreonlxaaldtnnsrdlapafciobktrteoewdtr)haotit(osotyrvpuyperniordctijraufeeylpcaltyltp.iecrd2oa0cteecsodoruanrnltaerdlosslrsse)pcimkofevouserstrtiwyfiiinefloldcrlbutedahteespbaekatcnticlofehiwae.sndt 11.0 Sample Extraction 11.1 11.2 1111..34 11.5 11.6 111111...987 11.10 11.11 11.12 11.13 WWSAACEfAmeeCDcILnfCCAfoSoollePeloodneuuoedeaeildulnrenlEnnadalcmnisaanddtotdo-gudhtleadtttrd6wcryeewerhtLtin3i2iiheifa.tzdt0wtf(te5fcuhe0isripeeu5d,uodoottd.geimoeghrmrgtnslmnmeigbehlcbleeceadiyeLpayanoeymLtgtmtcotp-lnhhtuhlus2aet52ohrupfteeteeebeoco.)0ea0mlftdsceeepsthfntmCamo.utm1us.nyiaohlmmnn%btibLlunLlCeLdwtedeepesostnsasiiaSoattlooslC1tneephencaaaPhf0ofpg,aue-dtmctH0EniHn2t4e-n-ev1%itmSpo3P3n0cPotl1ocmolePl0tL00Li.aerratc8mxEtL0h0itCrCcSte.fomtore0w0eOiurxcoodwnitwgsLaterdhsr,fmhaprpealpagaactlmtmeomtnmhr)tnerLateea.iudsefeopedrsrybtflf,pewg(h(S(.ooecsLd1o~e-varhsPoarilCont(2a-ymn-2EC(ge1ro2k/2fpdt,rMdoiedlece0ri0ng.nlrtroxa6alaioS,otmrepnmooClp6tp/amcydnnrMliv/ni/oteilnesmnsdLodelfleSdeuual5ogtnl)Lcect.utsaeee)ce)iC)mhwmbd..n.stsacb.y.urLetCkDoeDyithhsennpoeoortoSpetalorStealPnfsnaihenblssfoEoceucmesuiotttalntaigiutLnslcgotltteewaeeeohga)nttc.tmerrte1atco1cuirn0sesoon0ibhitdldollnduemaumuagftsaakmcomecLtLdtete.o(hidnnrf.gonmeomrnrdirrfaDtueueito5dtintstntir0fihuhohsoydadaaca-ntmnratnrk3haeeyyrooeLesd0ddllr Page 5 ot` 7 E05-0210 Interim Report #5 Page 34 o f 65 Page 88 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOOt782 | ANALYTICAL METHOD [ Method ofAnalysis for the DeterminatioLnCo/fMPeSr/fMluSoiooctanoic Acid (PFOA) in Sediment by 12.0 Chromatography 12.1 12.2 12.3 12.4 12.5 egvsUSmbbsiSAssaILnonxttteeneeataCasanjufjacnrgtameennnetwssne/leiwccdMdudytnpeedrtetalasazaadbinnlaeiSorerrtetrntiercndhdedsenin/ddearMtsdresisgsaeaseln)eisiynn.rsfsmvoSciabaoasndpslmearfntusmaertaoareIPetsdydstpyrttnmnareyhadiFelnssoeosmbae5.dOpeaerdntste-feemeia.apiA1tssmnrstnrhle0hiohsptdenaexa.ecaoueoljrnssdotnuynenrceaAdreclattscautmrdeeant.ceaenorevrdtsvpbssacddfeneepleyalsdeoeory,osclhaaysitltofnotbci.iccncnfdeuarh5aatoaeacxlhil-lAinldartesi1csebibgtrnases0obrarqebrmteantnaerteuaodet.satdgipnaigfatosaoarulnismitnereotnantrtadasitnlplnnthsts,nsealeisdeieteuadsntotartariassasan.gnonrtmindradnnd(dfdouitta.pgSilioasvufvrlmtriredMetaeenadeasn,usLgosadcapdssnairmcstoeaa1nndmsomtnrn/euLxudpsduafeofsynsroenroswtrerntsrttelecmt.eilxbsiarofeictbfeeonugadwi3oeeAanrhse.ntsedt3ldhtcneydaiiaenneabtsyz(rangncoeebsodmfldetsdryuoirfaci.raepndcmsofcctfqlteutinpaaoeeupodrllderlaviilinlsiabvnbeonepkassradrlwtsetaeaeolraettvttsliirhantteaooeheahoartlsdnlance)settlf 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.5 sTieEHeaARstwdbMCo1nhta0nxpalh3rafeoanjhaemptc.eruene0cr9wi4yrniotlrcrrgdo%orphu9o1karetehamvccleokdp23nverTaotedsdnecrtedsaelerraie.mdeoohir(trasirdbmtbosRe,nweotiu.otlmregsooltuu,aJtansaarrhnnol.ftuttiamdtsieinvkb(hsfmosot03Tsatsaeaeencntb.6lypom9homunmoPer9sts8eeneobcapFbtuul5staaoettll4eeOmrs.ase)mnveov1tetni.sAfdaunela3xusuIfnlaxsnesihfcsu)mortcacdhtpeaoIaadrtimloefistbewetbuakecedcupenfnudelcedotoolrrteaeddasnen2oupvdlotbsptt(t0anifaeabahreRbfyenr%ldksoifnesereanad)reelatnkotmtesnxrthgosoPmtmeinftotporepor-FcfhtebtaiaenlifrookOeaertttcxeaahrhrestettnlcrAiereeeroxaadseesaarfadedsshsaallcctassisypttujoulotbathtpculsoemolaugtslraiatstaldnctlakhsfletnuidlo.uidtestctvbisoueltssofinabet0iarnrtacArnulaetsm.tblsdbsai2iotimlteoonirdeloruocgenyntuternbonsuoeanrhemeutgenrarutemd-orhmvat/etbasteismftleatoxePlen3ioeiaiytsdxtenFtLt6rrnafrzhbeirieOa9,cdedxteegeochcbdeatAxetattpieway..rchhsefnmtepdnapredeeaertsaulrinsu.enaaritkecseabhnbtit-teiifotarheelenoreae7nldaxogonditL0nt,mmtiu,lro-eiOtswtacm1mhiwntsatcoaQa3ehisnitudun,0tiicp.dssblldoeH%uadet,ltanrrahuI7trtvetnfehbghbdoh0neinkeeeeeseasft Page 6 of 7 E05-0210 Interim Report #5 Page 35 of 65 Page 89 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research MethodNumber V000i 782 I__________________________ANALYTICAL METHOD__________________________] Method ofAnalysis for the DeterminatioLnCo/fMPeSr/fMluSorooctanoic Acid (PFOA) in Sediment b'y 13.6 aRne4ta%ennatliwyotinitchatiinlmrauennsatbnheaetlnwytteihceeanlsersuttanmn.udIasftrdrbeseteranentadinoanslaytimzmepdel.edsrifmt uesxtceneodts dthriifstlimmoitrewitthhainn 14.0 Calculations 14.1 bgUaessneeedtrhaeotenfdopblleyoawtkhienagrMeeaaq)susuasLtiiynongnxttohsoecfatslwtcaaunrldeaatperrdtohgceruaarmmve:ou(lnint eoafrPrFeOgrAesfsoiounndp(airnamnge/tmerLs). PFOA found (ng/mL) - (Peak arselaop-eintercept! x DF DF ~ factor by which the final volume was diluted, if necessary. 14.2 Fthoerfsoallmowpliensgfeoqrutiafiteiodnwtoithcaklcnuolwatne tahme opuenrctsenotfrePcFoOveAryp.rior to extraction, use Recovery (%) - [ total analyte found (ng/mL) analyte found in control (ng/mL)] ^ analyte added (ng/mL) 14.3 nUgs/egt(hpepfbo).llowing equation to convert the amount of PFOA found in ng/mL to PFOA found (ppb) " fPFOA fousnadm(pnlge/mwLei)gxhtf(inSagl)volume (5 mLll 14.4 Ufosuendthien fpoplblobwaisnegdeoqnudartyiowne(iigfhnt.ecessary) to calculate the amount of PFOA PFOA found (ppb) dry weight ~ PFOA found (ppb) x [100% / total solids(% )j Page 7 of 7 E05-0210 Interim Report #5 Page 36 of 65 Page 90 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMNeAthLodYNTuImCbAeLr. MV0E0T01H7O83D Method of Analysis for the DeatnedrmCilnaamtisobnyoLf PCe/MrflSu/oMroSoctanoic Add (PFOA) in Fish Analytical Testing Facility: E3S0txa3yt8egeCRneoRslleeeasgreeca,hrPcDhAri1v6e801 Approved By: " P - l C_ LTPaeuclhnCiocnalnLolelayder, LC-MSj, ExygIen Research l/r) fiU / 'VJoichen PFrlaehsiedretynt, Operations, Exygen Research Date Date E05-0210 Interim Report #5 Total Pages: 8 Page 37 of65 Page 91 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Resetrch Method Number VOOQ17S3 [ ________________________ ANALYTICAL m eth o d_________________________ Method of Analysis for the DetermCilnaamtisobnyoLfCPe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 1.0 Scope TaScphieidsctmbroyemthHeotridgicihsDtPoeetebrfceotoremrm(apLnlCocye/MedLSif/qoMuriStdh)eiCnihsforioslahmtiaaontnodgacrnlaadpmhqsyu.anctoiutaptlieodn otof paerftlaunodroemoctaMnoaiscs 2.0 Safety 22..12 pACrolewncsaauuyltstioothnbses.earpvperosapfreialtaebMorSatDoSrybperfaocrteicheas.ndling any chemical for proper safety 3.0 Sample Requirement 33..12 3.3 pSaaAfsSounrnaatoobamdmjldlleeyippmclapsleltsieer.aastostv.2cicesoo0ehnlslogo.seupocolertdfSinaotebnenaisdenltpphsraraoofnormmcodcezpeodesplguenslereafencdossietzrboweeertxhaiwfltogelrireatbehsceatoedimvxosretpnyprre.alniecccisitgefi.iohienntdP.tlfioanrPcoleaztahlectlehneoewstshastmaeofmropfarrplgoilnezecgesanuripbnnslotaaicnlnmontpdfitolmiaeoriexnitinhedoriessaf 4.0 Reagents and Standards 4444444444444.............11157816914231320 TAPFDMSSLAC1Weloiu--miacoleArOmplairefrutcmbelhitsteceauseoorcatotincornnhoganlnle-riyreneo(H(otibl-6laru1loodiin0(Pm2-cllRci6e--c0LtLa10HeahH-A-c0CCa-n4lP2iPocg0Ho0d-L0ereLgNi0Pmo-tc0nCrCaHLsamteAtmRidegCslgigeaceehrer--rsnaiaagas)hddegdhdARr-)ea-ee)eR-e.-dnC-aSeeRtRg.aRiSgeegegre.anmaaeagRtgdnagegeee-netnraAntagtgdtlgrgedaegrnrradraitdaecdGdeheerade 5.0 Instrument and Equipment 5.1 55..32 AAspAoLnldvahceleiovygnnithtcinsceeaeptclqoetbuercafdiooplatrlponlmeecadcaetntwcarcaaniewptdhaeldbimaqaleutvMaioadfrafoisarrcsebhpalSredeopaimvnekcogaitlnrttuoootmegm0gre.ear0atpi0enthi0rjoe0(ncLc1atCaopgnr/a.MdbcaqlSepu/MaaobnSlftei)t.paotuifmoinnp.jiencgtinugp5t-o2002 Page 2 o f S E05-0210 Interim Report #5 Page 38 of 65 Page 92 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOOP83 | analytical method Method ofAnalysis for the DetermCilnamatisobnyoLfCPe/MrflSu/oMroSoctanoie Acid (PFOA) in Fish and 5555555555$55.............11111115894671653420 C2DWASR5TD11150P2e2imiouisrss5nE5ttmmisppLoast-umroottrmpLLmiuycssafiLbplLauaddcieezbebegtipiLvaissestlleeoartepDepre(apn.cHoo2psmrPoass-0isPE(rpspaaihm1acLhabebatn0rbatlColLkotea0eplere)per-.eppsvrri1(.d.ooopoiS0awlelfuf0lyytslp-s0kppapmesirr(iptlookSn.ocLsppOnou.yyi-tanclllhnOeeagdbnntO.5oee1u0ntccLt0olmee-,e.nn1sN1Ltt0.rr000iip0ff5oupu-72lgLgy10ee)p70,ttr7uuuwo)Lbbp.i)eety.hssl..edniesptousbaebslaetti2p0s0. 0 rpm. 6.0 Chromatographic System 6.1 6666....3254 MTGMA(Pernoo/maNabbdlpi:iyilleeeet8nir2caPPt5athhPul0aarrC5oess-ee:go0rl((5u3aAB2mm0)1):n3C:::0F) lM2uomeptMhhaasnAeomlRmPo(KnieuymstoAnceetSacteieinntiWfica)t,e2r.1 mm x 50 mm, 5p Tim2208e120....(.00050min) 66622/.55555A %3355B IS I3S5 FfmloLw00000/.....m33333Riant)e 666...768 QRInuujeancnTttiiiotmanteiVo: no-:l2uPm3eeam:kin1A5urtepeasL.-(ceaxntebrneailnsctraenadseadrdtocaalsibmrauticohnacsu5rv0ep. L). The above conditions are intended as a guide and may be changed in order lo optim ize the HPLC system. Page 3 o f 8 E05-0210 Interim Report #5 Page 39 of 65 Page 93 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001783 ________________________ ANALYTICAL METHOD_________________________ Method of Analysis for the DetermCilnaamtisobnyoLfCPe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 7.0 MS/MS System 7.1 PMFoOdAe:. Electrospray Negative MRM mode, monitoring 413 -> 369 m// for TophteimabizoevtehceoMndSiMtioSnssyasrteemin.tended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 Mobile Phase 8.1.1 a2mmmMonaiummmaocneituamte taoce1t0at0e0imnLwoatfewr aitserp.repared by adding 0.1S4 g of 8.2 Extraction Solutions 8.2.1 8.2.2 oa32c0%fi%ddiamisDnceiot1mhr0bye0iltcdhmiyacLclhdioldiocfrihmnolosmeirltoahesnatihnelaaotnnol.oealiinfsintpoarlleuvpeoanlreuemdisbepyordfepi1sa0sroemldvLibnwygib2thrgitnoogluifneagnsec3.ormbiLc Alternate volumes may be prepared. 9.0 Standard Preparation 9.1 Standard Stock/Fortification Solution 9.1.1 9.1.2 9.1.3 mmaAomiAPnrfL1eLe0a1at2p.h.on51o01aaaff2rmnplpte5tyhoghgLtamle/fiemcriLsnnL1a1tLDL.lo0a0LcsP0ffpDkt1oEoagp2rnPrst/g5btiodmEif/-oflamimiutLcbrcttdalLLoaiestotti(.otiLsoncloloenoDuno.lrtusPifrsotoeEo-iclno1lutubne0tttooidi0totootanflnpeoafog.orin/ffpfmiaPnuPlFLarFvliOtOooyvAlAf)oumPlaiusinFesmdOppoerAderfeoipl1pbafu0aryt10reee0dwwdt0obeibwtiyhg1yi0hbtmh0birniernmgimtnhgeLagi1tnnih0nwogaglmniitoinhgnIl 9.1.4 A 0.01 pg/m L fo rtifica tio n solution o f PFOA is prepared by bringing 9.1.5 maTf1rp0hoepemthrmosattxnLhoioecmlokidanfaatetnatehldy1eof24fo50prC.tmr1iefpiLacpanaLgrdtaDi/tmoaiPonrLeEns.osbstlaooublttutilloeetino.fsnoarrtaoe a fin a l volum e o f 100 w ith mtoabxeimstuomredpeinrioadreofrfig6emraotonrthast Page4 of 8 E05-0210 Interim Report #5 Page 40 of 65 Page 94 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygea Research Method Number V0001783 ________________________ ANALYTICAL m eth o d_________________________ Method of Analysis for the DetermCilnaamtisobnyoLfCPe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 9.2 Standard Calibration Solutions 9.2.1 9.2.2 poTLrfhCetep/hMaefroSe1ld/l.Mo0awpsSignnc/egmaelidiLsberfdaao.trtiytoipfniiccsaatatlinoednxaasrmdoslpualteiro:enap.dredpitaiorendalincomnectehnatnraotliovnias dilution may be S99oo..C22lfuo..F34tnio00co0r.0.en1t110.0ianAn.S0...1f020(00tte5itu5r5lcoe2tafaedrttt/ierieCmonodnanLa.tlto)lec6vaolCVilb(u,om25r11m111lua...u00005L00ptemi)osteonanssudteacnmodnoDancrid(elt111um111h1sn000000t0steL000i0000.rdna) tti1oo2n5s-mo fLsCtLaoDn(n00Ud000cPF.0.00.0gea..0E.i00.n00/r0n00m20d1t02a51r55nsl5L5aat)mrioroanyw-bme opurethpabroetdtleass 10.0 Batch Set Up 10.1 10.2 ociREnonaentecqhchueeiunrbqnetaurmttaarcetlehiianottytonessdaffs(soslacaurmobrfanipcentlorlcedonesltarpeonlxaaldtnnsrdalpafciobktrteoewdtr)haoti(tosotyrvpuyperniordctijraufeeylpcaltyltp.iecrd2oa0cteecsodoruanrnltaerdolsslrss)epcimkofeovusrestrtiwyfiiineflolcdlrbutedahteespbakeatcnticlofehiwae.sdnt 11.0 Sample Extraction 11.1 11111111....5342 11.6 dtfedSPcPP(AWhofiiariaaltmodyellarchcelarbn.dcokkieentogeiwtr3i2fhanhzttyb00ehny,hedyedaleSmmadsanbt2citLLuingchdoy0rbhe-neoodeelo1mmdfosdrfifyargeLte1aioicitnd2cofnhseLrS,e5nigaatolCtrPonaozetofnE-hnoernpmrNieeneliltwtetarLHrS(uiizcsintlibPlhjeeetae.hertErmp.aolesifaenlCedptouaDudislrnoraeebtti--sirnnnsems1hseddceisheaanaeiaqmhsamkttrspioounode)oiel.eduoxnuodan5rftsn.lwc0itDnliohelaeifawnretmllywrnw.alao)cisLsi.zgrtokRthhiehelsseieupnttnsh2m.oa.sfbeclelnygaytDptsipshoekfrownelaosrorinpifsr-nctlyoishhsaoshltaisimenal2kka,npge0plewle2rloedmifwcwtgtoeehfLrlilnstatyt-hihtsoo1lrekiilbf5cfusceuat.mehfomgnogleeeeuritenehmlout,a3unutnn02ebcso%seeetl.go,s,, Page 5 o f S E05-0210 Interim Report #5 Page 41 o f65 Page 95 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P000113l Exygcn Research Method Number V0001783 ________________________ ANALYTICAL METHOD_________________________ Method of Analysis for the DetermCilnamatisobnyoLfPCe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 11.7 11.8 11.9 111111...111120 11.13 11.14 11.15 11.16 1111..1178 TAMAtetPsetDCAHPfSDCtffhhhhaovlxoarlhndaeeeeedoeemaeaaratsrsaancdrnmcdkncpps-skt-aklpaettaiee1aso1eyr.cr2soenen3fmiiarz0t1st00gaftteftiaph-treuh0uuorhree4-tmettemm-megegnsnshhetaspmhhae.etiieehidaLrifsuenznmaez-itaaLrtaebuetspnueeohopehmrprhxetaxteretp-xeeeefelsahlotdstdpwesdstrsarpheri5vfsa.ulS.afcnaieeac0ofOcltocfeucdapalhtreaalttafstcsuoemodmogssi~ekofdmznnkafnrlttL1pnLel-gooaei.ifae0rnnttotnelnreos,oeHappczidmCikfsflslobtoteoeasPaetkraoslyLlhructri.yLnwylacltteprf-lhepopraonoCeoe1ihorrdlrrncdrsfooua0tolsdvtadoa-uppsstptmiiicotmoeicgrnaytyhne1aitolhegluleohimtsgeyemnthosnehtn2SnueiuoanihLexbeenelsPtuiedusrmCi/eitgEonrdanrtatseS/uSiuLmitemagtMxlsaPoebPbcestrtpdonEeooerHSioywiialrissslnnf/lesaeucscMevictp.c2tcumtootwveohooih%omSl.anlntursnienu.hpnia.imttstmozeatabh1athesri-nlit2etnnaCecanehru5tiocaiebfoopnfnentor5imeliirgbgtldlopr0.ptue(iLeen~eacsecscdtamata3ntosssarmm.0<timhi-Ltcnlhsapaips4sbhdkfenleil0criaeedeincoepizereoenmC.naelaenutttdtd)tmhrdah.~i-etstpfee2e2hfuetlam0e0eghaanil00saenroudk-n00tiuasonotrthtuirhiearlatnptpbnihptsaomimedoneeel.f 12.0 Chromatography 12.1 12.2 12.3 12.4 esgvsUsmtiiAILaShnnaoanieentCtsnmejtumfjnraaheeeetss/nelweecreppcMuytrdstrnntlllsaabzepaeeitSdttredneecirscheesr/ddrea.aoMdeeeisasssnltseafiye.rssfcaSobdtA,aoesalmrftusmtreasecstPstdoptvtmaahaeyhiFelafnelmoeese.dinaOrdcnbtsympeaa.airAbmnlasnlar5oeietdltab-a.atcuigle1sornnoyniearc0Andnrtntttuare.aniasoenrorsiatvbscadfnntiSmpaaevylgeytncsolseaapilttodn,bicnailonncafedhrndaoaecrasfidandnlraetsr(aistgrnsdtoaraeqoetrdsnntnmreuotsaadtgmasitucsamearnritcusroeetttadtousisnlnt,steussabfdituesanbtooeaaitssbtelnormitlefdnnootfuiohpgw.niifsrvnelmtieeecMecandralufLgaosusilasnrpieirdstsb1nbdcesemtr/eroyLdxpasfanoatyroerwainedrnrdoctecdtexansistebicftlhegdiai3loeteebsahnet.ntdros3wtdbacatifnaenaseet(irgasdnionegodntmiandtrannojrrneaepdncefcdtfqlsiuvipotseanuoiretelrogvianmlirdavonnnyeksaaawtlsd)enoly5.aiaevdn-rrante1aedhbrItalla0neaee)sstl Page6 of 8 E05-0210 Interim Report #5 Page 42 of 65 Page 96 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001783 ________________________ ANALYTICAL METHOD________ *________________ Method ofAnalysis for the DetermCilnaamtisobnyoLfCPe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 12.5 eSxacmepedlestraenspdoarndseresshpoounlsdesnsohtoeuxldcebeedfusrtathnedradrdilurteesdpoannsderse.anAalnyyzedsa.mples thai 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.5 13.6 R2amasTAHbmteEMCodRhtnp0nlahearf4heonuauepet.rutneacrw94yinsioestr%rgondrohtt1o9nkreehmaaccvnokb3pt2vlraiocTtswedonreyoaedeelorriaeotittr(irnsairtonbmboitsR,eweetcihobolttrgesoxl2aaua,titanatfrrinnhclani.ottmamsnti2seeiikvrhm(oafaontseu03eTsaaemennnecdtn.s6Pylpoh9dmomurap9s2Fstete8nbecnabalhtu0ulO5estaaoelnae4eemrs%s.sm)ntelvAdon1ttwn.ydfuseal3usIufntttohaexsnihihahsf)motrcoceafctptIoeedaritanumltfsbieewluhblskelceecpnefddueeerncesovtoorrtuatetastneetbounpaindomoltslprt(entnafiaas.aterRebfednardksuorifngreasanl)Ierdaceslatenkroftm-trnanstoestePdtmmriflbuoaptpoexiomsFchebmbttaitlntfrooeOkeeerrteturaabaharrrnseescnAlirrcteeataotaxdteesiadithfrodnhsobessalacasasoisnujapdonulebtaunstpllofal.ougasrylstritlstcsnalsOkihmetfezitdomtutsd-aetec.vaiSutepelsfnosidbaneieanaxtrlcndr.epldstetmlsdtbsaiioialrpsatoordlroarrurcgebinnudteocdnfomusahr,mtstustenuraedteotuevhtaahtdtbieitsfxsltnolePehe.n3eiieytcsjdxentaFtr6ezahebrietnOc9ecdeeategotohdbccdeAteetpatnwea.oy.hcsdnmenepuae.edeatrwutllshuarernaidtseabiithnbiifsitfborneotlerballe7nlogdniaetiLmm0,mimn,lo-Otiwekoihm1minatxtaQre3shnsuceiw,c0ita.pdlsslHuu%miaettt,trdhhruhIttvpetaeehbgfhihonelnndneeeeeaeft 14.0 Calculations 14.1 gUbeasnseeetdrhaetoenfdopblleyoawtkhienagrMeeaaq)susuaLstiiyonngnxttohsoecfatslwtcaaunrldeaatperrdtohgceruaarmmve:ou(lnint eoafrPrFeOgrAesfsoiounndp(airnamnge/tmersL), PFOA found (ng/mL) = fPcak aresalo-pientercept! 14.2 nUgs/egt(hpepfbo)l.lowing equation to convert the amount of PFOA found in ng/mL to PFOA found (ppb) - fPFOA foundsfanmg/pmleLw) exigfihnta(lgv)olume fmL) x DF1 DF - factor by which the final volume was diluted, if necessary. Page 7 o f 8 E05-0210 Interim Report #5 Page 43 o f65 Page 97 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V000178.' I ANALYTICAL m e t h o d ~ Method ofAnalysis for the DetermCilnamatisobnyoLfCPe/MrflSu/oMroSoctanoic Acid (PFOA) in Fish and 14.3 Fthoerfsoalmlopwliensgfeoqrtuiafiteiodnwtoithcaklcnuolwatne tahmeopuenrctsenotfrePcFoOveAryp.rior to extraction, use Recovery (%) = [ totalanalyte found (ng/g) - analyte foundin control (ng/g)] ^ analyteadded (ng/g) E05-0210 Interim Report #5 Page 8 o f X Page 44 of65 Page 98 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMNeAthLodYNTuImCbAeLr: MV0E0T01H7O84D Method of Analysis for theVDeegteetramtiionnatbiyonLoCf/MPeSr/fMluSorooctanoic Acid (PFOA) in Analytical Testing Facility: 3SE0txa5yt8egeRCnoeRslleeeagsreeca,hrPcDAhri1v6e801 Approved By: PTaeuclh"nCViocnailnLolaelayder, LC-cMS-,JExiyLgen, *Research Date ' JVpoihcne/ zPFrlnaehsiedret/ynt, / / ____________ Operations, Exygen Research E05-0210 Interim Report #5 Total Pages: 7 Page 45 oj 65 Page 99 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V'OOOI ?#4 _________________________ ANALYTICAL METHOD__________________________ Method of Analysis for the DeterminatbiyonLoCf/MPeSrf/lMuoSrooctanoic Acid (PFOA) in Vegetation 1.0 Scope TaScphieidsctmrboyemthHeotirdgichisDtPeoetebrfceotoremrm(apLnlCocey/MedLSif/qMouriStdh)eiCnihsvoreolgametitaoatntoioganrna.dphqyuancotiutaptlieodn otof paerftlaunodroemoctaMnoasics 2.0 Safety 22..12 pACrolewncasauuyltstiootnhbses.earpvperosapfreialtaebMorSatDorSybperfaocrteicheas.ndling any chemical for proper safely 3.0 Sample Requirement 33..12 3.3 pfasaSSAounnraatoobadmmjldlleeiyppmclapselltsierea.astost.vc2icsoeo0ehnlslgoo.seuopcolretdfSinaotebennaisdenltpphsraroofanormmcodcezpeodesplguenslereafnecdossieztrboweeertxahiwflotgelrireatebhsceaotedimvxosretnpyprr.ealniecccisitegfi.iohienntdP.ltfioarnPcolezatahlectlheneoewstshastaemofmropfarrplgoilnezcegesanuripbnnslotaaicnlnmontdpfitoilmaeoriexnitinehdoriessaf 4.0 Reagents and Standards 4444444444444.............11118967123541320 WALTPFSSDACM1-euil-oamaicOloAermpltirefrutecmbcelshtiereauostorcaaotcni-ononhgntnnleriryeeoH(otoi(b6-arllu1llodiiPn0(m2c-l-cRi6eL-0ctLa10HeaHh-AC-cC0a-n4lPiP2Hog0co0dg-L0LrNeei0rPmo-c0CnatCHLsamdteAmRitgCe]slggeeacerhe-rrs-aniaaag)shAddgeRddrh-)eaee-e.-e)-CdnR-aSeRtRRg.eiSgeaegeer.angmaaagRetggdaengeeee-ntnraAnatgtgtdlgregdagernrradraitadecdGdeheerade 5.0 Instrument and Equipment 5.1 55..32 AAAspoLnldvahecleivoygnnithtcinsceeaeptclqoetbuercafdiooplatrlpolnmeecdacaetntwcarcaaneiwtpdhaeldbimaqaleutvMaioadrfafiosarcrsebhpalSredeopaimvnekocgatiltnrtuoootmgem0rge.aer0pati0enth0irjoe0(nccL1atCaopgnr/a.MdbcalqSepu/aMaobnSlftei)t.paotuifmoinnp.jiencgtinugp5-to2002 Page 2 o f 7 E05-0210 Interim Report #5 Page 46 o f 65 Page 100 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygeri Protocol Number: POOOl 131 Exygen Research Method Number V0001784 _________________________ ANALYTICAL METHOD__________________________ Method of Analysis for the DeterminatbioynLCo/fMPeSr/fMluoSrooctanoic Acid (PFOA) in Vegetation 555555535553............111117156894153240 CADWS25DR111250P2meuioir5s5snEtmtmippLo-astmmrootptrLLciuyssaifLLlbpauaddceebeebgtpiiaLvissstlleeorateppeDea(Hnpcoop2msrPoass-si0P(Erppaasihm1aLchbbaeat0nrabtlloLkCotee0aplere)per-v.eppsrr1i(d..oiooopS0allwefuf0yyltlskp0-sppapmeisrr(iptkoolSn.ocLsppOnuo.yyi-tanclllhnOeeagdbnntO5.oee1u0ntccLt0oleme-e,.n1nsN1L0tt.rr000iip0ff5opuu-72lLggy01ee)p07,ttr7uuuwo)Lbbp.i)eety.hssl..edniseptousbaebsleatti2p0s0. 0 rpm. 6.0 Chromatographic System 6.1 (APn/Nal:y8ti2c5al0C5-o0l5u2m1n3:0F) luophase RP (Keystone Scientific), 2.1 mm x 50 mm, 5p 6666....2345 TMMGeroomabbdpiiilleeeenraPPtthhPuaarresosee:gr((3aBAm0)):C:: M2 emthManAolmmonium Acetate in Water Tim2208e120....(.00050mini %6662255555A %3337755555B FfmloLw00000/.....3m3333Riantei 666...687 IRQnuujenacnTttiiiotmanteiVo: no-:lu2mP3eeam:ki1nA5urtpeesLa.-(ecxatnerbneailnsctraenadseadrdtocaalsibmrauticohnacsu5rv0ep. L). TophteimabizoevtehceoHndPiLtiConssyastreemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 PMFoOdAe:. Electrospray Negative MKM mode, monitoring 413 -> 369 m/z for Page 3 of? E05-0210 Interim Report #5 Page 47 of65 Page 101 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO17S4 | ANALYTICAL METHOD Method of Analysis for the DeterminatbiyonLCof/MPeSr/fMluoSrooctanoic Acid (PFOA) in Vegetation TophteimabizoevethceoMndSitMioSnssyasrteemin.tended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 Mobile Phase 8.1.1 2ammmMonaiummmaocneituamte taoce1t0a0te0 imnLwoaftewraitserp.repared by adding 0.154 g of 8.2 Extraction Solutions 8.2.1 8.2.2 o2a3c0%fi%ddiamisDnceio1tmhr0bye0itlchdmyiacLclhdiodliocfihrmnolosmeirtloheastanhinelaaotnnol.oealiinfsintpoarlleuvpeoanlreuemdisbepyordfepi1sa0sroemldvLibnwygib2thrgitnoogluifneagnsec3.ormbiLc Alternate volumes may be prepared. 9.0 Standard Preparation 9.1 Standard Stock/Fortification Solution 9.1.1 9.1.2 9.1.3 9.1.4 9.1.5 AmTafmaommA1PAirnp0hrofe1eLLpe00atem1at2hpr.m.hon.15o0osa10aaattf1nxL2frhmonptlpoe5iythecogmphlgLtokamel/idige/amncfmiLas/atLn1am1ettnaLDt.Lleohl00adLLycsP10eoffpDtk1fo2Eo4afopfg52r0nPropstr/gb5t.CidoEmtrmri1/foi-tefalmifmiiutLpLbcrifactcpditalLaoLnacieasLtgrotdtai.(ottaiL/sonDicltomtlooeianDoiunoP.oonlrrLtunesPEnirsfsooteE.o-issobnlcso1ltoulotouabnuet0ltlbttoouitduti0otiltltoootetaeiniflnonp.oeoaffonsg.nioronf/faorpfmaiPtrnuPflaoeFLarFvPimlOttoaoOFoyvAlaOf)AbuoxfmePilAaiinusminFsseamdptOiluopsoermAdrerfvpeeopio1rldppabfuel0aureypti01erenrmae0diwwrotda0eeobdeidrbwtyieoh1goyfbib0fhrtfmybh0riig6in1renbmeimgt0nghmrr0eLaigai1ntnontih0owgngwnoarigltmniinht1toaihghnsg01il 9.2 Standard Calibration Solutions 9.2.1 oLfCt/hMeS1/.M0 pSgc/malLibrfaotritoinficsatatinodnasrdosluatiroenp.repared in methanol via dilution Pitgc 4 til E05-0210 Interim Report #5 Page 48 of65 Page 102 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOO1784 analytical m ethod Method of Analysis for the DeterminatbiyonLCo/fMPeSr/fMluoSrooctanoic Acid (PFOA) in Vegetation 9.2.2 pTrheepafroeldloawsinnegeidseda.typical example: additional concentrations may be S99oo..C22lfuo..F34tnio0c0or0e.0.nt1110.0innAa.0S...f102(000ttei5trul55coea2tsaetdrftieriaeoCnoidnaLna.tlto)lec6vaolCVilb(uo,m25r1m111l1ua..u.00005L00ptmei)soteonanssidtxacnmodnoDacnride(ltu1mt111t1hnsoo0t0000steLoo0i0000r.dna)ttio1o2n5s-mofLsCtLaoDn(n0M0dc000PF.00.e.0..0agE0.i00.n000n0r/020mdt1052a1r55nls55Laat)mirornoaywb-me opurethpabroedttleass 10.0 Batch Set Up 10.1 10.2 ociREnonaentecqhchueeiunbrqentaurmttaarcetlehiianottytonessdaffs(soslacaurmobrfanipcentlolrcedonesltaprenolxaaldtnnsrdlapafciobktretoewdtr)haotit(osotyrvpuyperniordctijraufeeylpcaltyltp.iecrd2oa0cteecsodoruanrnltaerdolsslrsse)pcimkofevouserstrtiwyfiiinefloldclrbutedhateespbaekatcnticlofehiwaesdnt 11.0 Sample Extraction 11.1 1111..32 1111..54 11.6 11.7 ddDtSctfefPCP(fAWhhlofaoiriiaaaeeotmedyellrscrheclacnbrnd.ok-kikpcentatog1i.wrtr3eni2fhnhiazt0abyt0f0ehny,eurydteadm-l5mahmgdssaneeb2chiitLLingnchdoya0retuhee-onphoxdetlo1mmdeefeotdrfrdfisayregLeat1aS.ociitdc2fcfoOnhesLrltS,SenigataolCtroomPsnaozetonkn-EhnoenpmLrNi.neleilttwtartoLHrpSi(uCicsintliobP2laehteaoe.hlteEmrply.aolcesliaenpoelCdpetuaDduirncslnooeraettbdti-sirnnnpsiestmsehtcddheyihseianaaieeolaqsmmaktetrsnopioundeen)oileee.oldxeunodduanSftnsalwcitOtDlSniouhtelaeeiPwnbfaretmllyr.newEwlaiao)cnsisLs.izgrRtockthhicetehlossiheuoepnntltsem2nuho.sa.fbetmlecl1naygaytDpt2isinpshnoSefkroeiwnlafosnnoimrpfntigsn-ntltcyohLseahssohsltsdaiiemaknlas2ka,nmiipge0lpnwlel2earlpsoiednmifltwcgwdteihoeefLzelilnresttaaty-ohithdtosthlrIlekiifubS-ecpfs2ceeuame.tmofnh0mgaoegleer0eouirt-en0uheosmluhtt.an3uhnutnrac02ebpspoeos%eemt,el.ogfs., Page f u f ' E05-0210 Interim Report #5 Page 49 o f65 Page 103 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOOl 131 Exygen Research Method Number VOOO1784 aanalytical m eth o d Method of Analysis for the DeterminatbiyonLoCf/MPeSrf/lMuoSrooctanoic Acid(PFOA) in Vegetation 111111...8190 11.11 1111..1132 11.14 1111..1136 TARtDseeMACSfAhaoxvrheneededmaartapancarddknp-pakaelet5esoyrec2naefi3arzt0tftteiptam-rheuohsr4-tetmegtenseishtaenhhea.eLrapdfsuema-itatsrtsnpeohoemerpha1exesexfplald1.ptdetsapr.r5vlusca8faee0oclecoucdataelttshtfsomugdsikornmnafotLp1niligoautaen-rtrnooes,gpeHidkfcslthobeoePsttlsayhucr1ytLwnaoet-r1parnCoe1i.ordtrdsl1oh0usldav1ppesttmiimooicranyansaoleglgamisgyenmtamhnS2uiuiaLxbneePpstCrm/ie.ilHdnotnese/uLietgMxsoiabcnstrndeooSrowysital/lfashuMcviepcte2mtwvehoo%S.a3nrsni.pi0an.ttsoaabhtmis-rletnCaeachLitcoeopnotercliigrbllopepue(iennescaectatatntssramr.<thit-cfahspui4khdeflg0reeaeoeripmn.Calteuutd)mab.-tte2eehf.t0lehai0ans0kntionoirlatptinhatmodel 12.0 Chromatography 12.1 12.2 12.3 12.4 12.5 evsSUief(cgmbASaILnotoxnxaeenaetoCsntlufclarjtnmgaeleitersenso/ablewiraucMytpneedsrcwrctslaadzpnbalncaiteSerttetneeioetrcehseinid/dddreaourtdMreisgsasealennnsebiynrcfsscaoSbtdyosasdpaaslmrffttsunrmelaoabeoaeiPtsdpdtrtntbamnerxyehdiFlesnrtdoetosea.dwOaeaardrantsfateetma.arseiAiscsmsdnonereheotpttdscnahneax.caouooldmenontuynnenrcsAdrltatsvuautcdrdeeaaceoneeasrnrntsvbstsrladcnedfidyepyeblsabdoeytaohscleraot5litroonoaibicdct-nfcneftudfhh1rasioaexaclioela0ednlrasictenmxbitfgrsnesobastirqeeartarrsnnaatereusadmtoytmatdatgaciftasnoapurtanispbteroetdnlnrttaldeedasitnaeldnths,snerassd.iedauidlstotstnaarisaasasaeonsrjAtmindnrtedntn.idfidudcns.pgliiasatuvntlmeraeirretjMeeEnddndeersuLsgaodsxcsappsan)tatrisotia1.rneldmtosmatnn/reLexncpssdtIaufroethynroenrwseiredrnesnrtaedeecet.xstiaieeaicitbbftngvasdvioh3eesAetaheneen.egaad3l,tdrrnmcnyiiiyanannaesydz(cprnanngnoceaad5lmildtdsrrseuno-erdae.1aepndgcfmss,tq0fltueepioeeiuotpdrorme.xlsvienlliafavteoneurakmasslwstasateeoltcpaevttthranltsteeehheaohraetldlsaeee!seitfl 13.0 Acceptance Criteria 13.1 doCafhu4rgo1hm3teaartmoiogunr.a(mT36hm9eua4sm1t3ush)aormewpuraepspaeerneatnkstotchfoearlrodessaspuoognhfdctesarrtboioontnhaedtiPo3xF6iO9deAa.mauniforno,mwahiplearethnet Page 6 o f ? E05-0210 Interim Report #5 Page SOof 65 Page 104 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V000I784 _________________________ ANALYTICAL METHOD_________________________ Method of Analysis for the DeterminatbioynLoCf/MPeSrf/MiuoSrooctanoic Acid (PFOA) in Vegetation 13.2 13.3 13.4 13.5 13.6 am2asRTAteEHMbmRhtnpn0lhraeeo4nuaeupetr.ntecawyin9isosetr%rrdonhttok9nreeaavccoknbp2tvTlsraoicewdnoeryoedelorrieotitr(inaitsorbnbsit,eRweetchobtoltrsexl,aaatt2aitnafrhicnanilotmtmsntie2iseivkrhoafnaotseu0easaemenncentdn.sPlypood9umrpaF2ststen8ebcanbhlat0uOlset$aoleneaeemr%.ssn)teovlAndtnw.ydfueslaIufnottthaxesnafiihhsmoroctcefctptIeedatainlutbfsileuhblesklececenfddeenercevostoortuettatsbentonuiadnomoltlrtet(nainfas.aterRbfnadusdkorfigrsnlaerdI)csaelaeortf-mtnarnotesetmPmdriflbuapxoepiomFsmebbttaittnlroeOkererttuerabahrranesesAcinrcteetatxtaaseaitdihfrodnheobslacasaisondaujoneltbantupslla.fluslaryritstsltcsOlakseimetiztdomutst-eav.eaiSteulpsnfiobdenieaxntalcdln.etpsdemltssdaiialrpsaotrdlrarrugrebcnieudcdofomsaur,tmstteusnuretotuaehvthabtdietsfxsltneelhee.niietycsdjentatreahzebretncceeetaegoothbcdcdteetptenawyo.chsdnepneuae.edetrtwullshaearirdtsebaihitnbifsibtrenlotbeaell7lgndaietiLm0mim,nlo-Otiekohm1imniatxtQre3snsicuew,c0ta.dlssHu%muit,tttrdhhuIhvttpfaeeghbhioelnndneeeeeaf 14.0 Calculations 14.1 bUgaessneeedtrhaeotenfdopblleyoawtkhienagrMeeaaq)susuaLstiiyonnngxttohsoecfatslwtcaaunrldeaatperrdtohgceruaarmmve:ou(lnint eoafrPrFeOgrAesfsoiounndp(airnamnge/tmerLs), PFOA found (ng/mL) (Peak aresalo-pientercept) 14.2 Ungs/egt(hpepfbo).llowing equation to convert the amount of PFOA found in ng/mL to PFOA found (ppb) = iPFOA founds(anmg/pmleLw) exigfihnta(lgv)olume (mL) x DF) DF " factor by which the final volume was diluted, if necessary. 14.3 Fthoerfsoalmlopwliensgfeoqrtuiafiteiodnwtoithcaklcnuolwatne tahme opuenrctsenotfrePcFoOveAryp. rior to extraction, use Recovery (%) [ total analyte found (ng/g) - analyte found in control (ng/g)] ^ analyteadded (ng/g) Page 7 o l 7 E05-0210 Interim Report #5 Page 51 of65 Page 105 o f 123 I 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 AMNeAthLodYNTuICmAbeLr MV0E0T01H7O85D Method of Analysis for the DMeatemrmmianlaLtiiovnerobfyPLerCfl/uMoSro/MocStanoic Acid (PFOA) in Small Analytical Testing Facility: E3S0txa5yt8egeCRnoeRslleeeasgreeca,hrPcDAhri1v6e801 Approved By: PTaeuchl nCiocanlnLolelayde_r,_LCC-M--S-, E0x^yg4eI_n R_e_se_ar_ch Date Date E05-0210 Interim Report #5 Total Pages: 7 Page 52 of 65 Page 106 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygea Research Method Number VU001785 | A:\ALYT1CAL METHOD Method of Analysis for the DMeatmermmainlaLtiiovnerobfyPeLrCfl/uMorSo/oMcStanoic Acid (PFOA) in Small 1.0 Scope TaScphieidsctmbroyemthHeotirdgichisDtPoeetebrfceotoermrm(apLnlCocye/MedLSif/qoMuriStdh)eiCnihssormolamatlilaotmnogaamrnadmphqayulalincvoteiutrap.tlieodn otof paerftlaunodroemoctaMnoaiscs 2.0 Safety 22..12 pCArolewncasauuyltsitootnhbses.earpvperosapfreialtaebMorSatDoSrybperfaocrteicheas.ndling any chemical for proper safely 3.0 Sample Requirement 33..12 3.3 psaSfaSSAounnraagtoobadmm)ljdlle,eiyppmclatpslehltsieer.aasteostv.ncSicoesoenhAgnlsoloo.soletupopceflertrrdtSinoneaoecasbnnatteideenlssplspahyraimrofo,nnormcgiodpcefzloeidetespsghunslfneeraeoneercdrsescietzeeboiwexsserttshaiwfraalogealnrirectybehtsiineaaooednmsnxvsrud.tepypfrretflanhieicccciseitegifie.soihienanntdPmt. ltafipoarnmPcloelezoatahcelcutleahneonnewtstshboastameeoffmrosupfaarrpslgoimelnezecdgpesanualreipbsnnslot(saaicn-ulnmlopentdppfsitomllisaieoreiextndthiinhe.adorinessaf 4.0 Reagents and Standards 44444.....53412 AAPMWecmeraeftmtthloueaonronn-iroHtoiruloiPm-lceLtHa-ACnPHcoLegiPtcCraLaAtdgCeecr-aigddrAae-.dCSe.iSg.mRae-aAglednritcGhrade 3.0 Instrument and Equipment 3.1 5533....5234 5AAAsp1o05LnldmavmhceleiovyLLgnnithtciddnsceiieasseptclppqoetbooeurcafssdioolpaaatlbrbpolnmleleceeadcaetppntwcaroocaanlleiwyyptdhppaeldbrrimaqooaleupptvMaiyyoadllrfafeeoisranncrsebeehpalSrdeeccopaeeimvennnkcogttartirltnriituofofotmuugem0gggre.aeeer0pati0ettnthuui0rjobbe0(nceeLc1asstCao..pgnr/a.MdbcalqSepu/MaaobnSfltei)t.paoutifmoinnp.jiencgtinugp 5-t2o0(21 555...876 2DI2ims5p-LomcsLlaebLalerDHmPPEicLrnCoaprvripioaewlts-kmi(t5.o0u-t1h0b0outLtl,es1.00-200uL). Page 2 of ** E05-0210 Interim Report #5 Page 53 of65 Page 107 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V000i 78S 1 anaLyticaT m ethod Method ofAnalysis for the DMeatmermmainlaLtiiovnerobfyPLerCfl/uMorSo/oMcStanoic Acid (PFOA) in Small 5555555.......1111191132540 TAWDWSCiPeuisrasnEtsiptosuterotvperi-ssaifmapauccSbeigutzetlieuteoepemrcnps.Paisa(mpph1laeeaa0btkVnt0leeie-afsr1oc..o0l6fd0s.0cpcpin(Lnlgian)ngdtC115108m-1SL0P0pBopclLya)pr,trrwoidpigtyhelsed.niseptousbaebsleatti3p0s0. 0 rpm. 6.0 Chromatographic System 6.1 6666....2345 TMM(AGPenr/oomaNabbdlp:iiyilleee8teinr2caPPta5thhPul0aarrC5eoss-ee:og0lr((5u3aAB2m0m)1)n:3C:::0F)Ml2uoemtphMhaansAoelmRmP o(KnieuymstoAnceetSacteieinntiWfica)t,e2r.1 mm x 50 mm, 5p Tim2208e120....(0.0050min) %2666255555A %3337755555B F(mloLw00000/.....3m3333Riantte 666...867 RIQnuujenacnTttiiiomtanteiV:ono-:l2umP3eeam:kin1A5urtepeasL.-(ceaxntebrneailnsctraenadseadrdtocaalsibmrauticohnacsu5rv0ep. L). TophteimabizoevtehceoHndPiLtiConssyasrteemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 MPFoOdAe:. Electrospray Negative MRM mode, monitoring 413 -->369 m/z for The above conditions are intended as a guide and may be changed in order to optimize the MSMS system. Page ' o f 7 E05-0210 Interim Report #5 Page 54 of65 Page 108 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001785 | AN.-U.YT1CALMETHOD ' Method ofAnalysis for the DMeatemrmmianlaLtiiovnerobfyPeLrCfl/uMorSo/oMcStanoic Acid (PFOA) in Small 8.0 Preparation of Solutions 8.1 Mobile Phase 8.1.1 a2mmmMonaiummmaocneituamte taoce1t0a0te0imnLwoaftewraitserp.repared by adding 0.154 g of Alternate volumes may be prepared. 9.0 Standard Preparation 9.1 9999S....t1111a....n4132darTmafAmaomiAPdrnphrfo1L0eLSpeae1ma2t.ptrh1on.o5oos10aaattcfx2frhmonplpket5ityechogmh/gLtkFamle/di/eamcmoiLaasLt1na1rettnLDL.lteo00laidLycsfP0offipDtk1ofEc4oafpog2raPnrpstrg/b5ttCiodEmtriif/o-iefolamifmuitLcpbrniactcdtalLaoLnaieasStrotdti.(ottaisLooncilotloeoalnoiDunu.oolnrrtutsePnirfsisooteoE.oi~osnlconltult1ubanuett0btooitdiot0iltoooaetnfnlpneoaffosg.ioornf/farpfmaiPrnuPlaeFLarvFilmOtotoOoyvAlafuA)obxmlePaiiumsinFsesmdptOuoopermAdrerfeeopil1dpapfbu0areyit10erenre0diwwodta0obdeirwtbiyehg1oyfib0htrfmhr0bii6ginernmmeigtnghmreLaagi1tnontih0ongonwarlgtnmih1atoinsh0gtIl 9.2 Standard Calibration Solutions 9.2.1 9.2.2 pToLrhCfetep/hMaefroSe0ldl/.1oMawpsSignnc/gemaelidLisberfdaao.trtiytoipfniiccsaatatlinoednxaasrmodlspulateiro:enap.dredpitaiorendalincomnectehnatnraotliovnias dilution may be S99o..oC22flou..F34ntoicore211511ntni00Aan0..S.f000(t000tietrnlcoea2tgaetdrt/ierimeoCnodnaLna.tlto)lec6vaolCVilbu(o,m25r1m111lua..u.000L000ptemi)soteonanssidtxacnmodnoDacnride(ltuttm1t1h1nsooot000steLoooi000r.dna)ttio1o2n5s-mofLsCtLaoDnn(ndcPFge0a00251Ei/n.r.n....m125000dtarnsLlaat)miroronawy -bmeopurethpabroetdtleass Page 4 o f ? E05-0210 Interim Report #5 Page 55 of65 Page 109 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001785 _______________________ ANALYTICAL METHOD_______________________ Method ofAnalysis for the DMeatemrmmianlaLtiiovnerobfyPLerCfl/uMorSo/oMcStanoic Acid (PFOA) in Small 10.0 Batch Set Up 10.1 10.2 ocRiEnonaenetcqhchueeiunbrqentaurmttaracetelhiianottytonesdsfafs(soslacaurmobrfanipcentlorlcedonesltarpeonlxaladtnnsrdlapafciobkterotewdtr)haot(itosotyrvpupyreniordctijraufeelypcaltyltp.iecrd2oa0tceecsodoruanrntlaredlosslrsse)pcimkofevouserstrtiwyfiiinefloldcrlbutedhateespbaekatcnticlofehiwae.sndt 11.0 Sample Extraction 11.1 111111...234 11111t...S76 11.8 1111..190 11.11 AEDmCLTCAmdfAHWnIoSieloroneedodLesluaeealnncapmadmdtnoiddoaltedsogwsyreSLwonuihsftifdztwhgftraeewmiapeu,eodeber1ttiodgrahenntsLlebshgeltearei1dyaepzyomtcttrmepolheo~sh.efamp5rpuef2enpscaotelpmhasLetlcepntCreaemmeuesyidlLmrfbtuoisnlouipLuodeennaopaslnfgnsmgeSiatlfcieonaeenatPtuHooprnhgfttdcEiisnnultePnle-iemedebmn3nttLilcioehnfege0asttaCio.ritcetanx0rooaihartmftdswrn0arwuatSoaiainpesgdaOmrssefdholptesgpilSeeanllpmumerOC.)trkaaleus.ueLlefyummbfs~v(CNosfei1pLSiLnioor2zozoo(CPlgr-eeflgtulddEl5e~i/ry,emnMrai2apfocstavc6mohe0rptpSlaaoradoi/or/imvm2pnts~MtilsfroaseyuaaLii1pbcmlnStdl1lbuee)to)lm0ugl.ee.Lsnmestebr.miaeefnn.DyceobsoaLuce1lroDtof9tpee.neeunnna2ite.ssrwstoplecairmsuftii.aeipufwaLnlriugeetgdgre)gttehi.teesceti1tlston-uut0alutoiabuoncbtmfemetteiol.aoLsianvnn(gaermdfusrroahnermIadattSddihukafdraaeyyymntre3.obaLdSosl 12.0 Chromatography 12.1 12.2 12.3 12.4 esgUAiLaSmtsiInhnanainetCstntjuemmajnaheeee/nsleeerceMcppytdsrtnrntlllbzaepaeietSdtetdneeicrsher/sddra.eMoedeiessasnsteafesrs.fScaodtAe,oaalmftsumrescsPtstodptvahaayehFleafnemleesediOnarcdtsbyepma.aaiArmblnalnar3eoiedltab.c-tauigles1roonyniearcA0nrntntttaure.inaseonorsitbasvcannfitSmpyavealgeytolssneaapilttn,dbicinalncofdaerhndaoeaarsifdanndalrtsra(irgtssdotraaeqoerdsnntnemtuos.dagtmasiucasarmenrtceurstettadtousislnts,tesusabfditaesntbooeaiastbtelnortmlednofotfuioh.pwinifsvnremltieeceecnadarlufgLaoussilsnripierdt1bsnbdceemt/reoryxdpaasfonatroewrainderdroctecdetsanisteibcfthlegdaiioleteesbhanetntdrtswodbaciatnfnaesaeetriggadsnneiodntmaidntaonnrjrneapdnfcecdtflspiuvioetsnaoeiretlrogvamnalirdonnnyeksaalwtsd)eonyaS.iavdnrar-teeadhlhrIalatlneaeess)tl Page 5 of 7 E05-0210 Interim Report #5 Page 56 of 65 Page 110 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001785 _______________________ ANALYTICAL METHOD_______________________ Method ofAnalysis for the DMeatemrmmianlaLtiiovnerobfyPLerCfl/uMoSro/MocStanoic Acid (PFOA) in Small 12.5 Sevsaoxemfcrtsewpuelsadercesratesalsiynbpsdrotaaenrtmdsioe.rnessshptoaounnlddsaersndoschtooenuxclcdeenbetedraftsuitoratnnhdeuarsriddnilgureMtesdpaosasnnLsdeysrn.exan3Aa.3lnyyz(oesdra.meqpulievsaltehnatt) 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.5 13.6 RTasaAEHmwRbmteodMC1htpn0nlhear3af4eonahuauep.rettuen0acr9wyn4risioesrt%rgrdn%rooth9ont1kraeehaaccmvnkoptb23nvlrTaoitwceodnrstyeodeaesleroriaeetoih(tritrnsariodbnmbittsoR,eweeotcihob.trtlegsolxo,uau!aattiaatnrrhinncalinlo.fttmamntdseieiirvkh(maofsonts0eue3Taaesaennncetdb.nsl6ypPom9homdumrea9s2tsF8teenebcapnbhat0uul5Osteoaaelale4neemr%s.se)mvntelvonA1dt.twnysafdueal3usufItnotltaxseniiashhuhf)smoctcrefcathtpIoeaedaraitntmlofsbieetwlhulbsekeececuenpfuedeecrnsdetovoolrtruateatdtsotneenupainodmlotblspr(tinntfaaa.stbaeRerbfyeanrdusdkofinergeaasl)InrdaesclatnkeormftttrnansrtohesttdmmPrifelbuoatppoeieosm-FchebmbttatlinefrooeOeeketrteuxarahabrarrsneencstnAlirrteeatrataoxdtseaeiadihafrnodhsobsslalaccasaisoysanujponuelbattsnutpllsoeafluogarylsstirttsdtclnaslskmhiftez1ietdomut.tdsetae-t0cvaioutpelesfobisndaneiaenartnxlcAn.dreltdsemtlsdgbdtsaiiolaisartoonrdle/rorarucregginnyutdeotndcmfouasehr,mtstustenaurreeodtmtuemvthaathtbideitsfsxlnteoelPhaneie.3ietiycsjdtxetannFtr6erzahebertineiOc9ecdeexategootbhdccdeAetetpaitnoeway.c.hsdsnfmenpaeupe.detearuwltrhlsiuarerniadteksebaiithnbi-sifietforbeneotlebralenl7xlogndoietaimL,m0timm,unlro-tiOwaetkoimhmn1isatxctarQes3hinsutecidw,cii0tpa.dlsolsHeuu%mieat,ttntrdhhruhIt7vteptaeehgbfhihoe0nlnnnideeeee.saetf 14.0 Calculations 14.1 bgUaessneeedtrhaeotenfdopblleyoawtkhienagrMeeaaq)susuasLtiiynongnxttohsoecfatslwtcaaunrldeaatperrdtohgceruaarmvme:ou(lnint eoafrPrFeOgrAesfsoiounndp(airnamnge/tmerLs), PFOA found (ng/mL) " (Peak aresalo-pinetercept! x DF x aliquot factor DAlFiq-uofatcftaocrtobry-w1h0ich the final volume was diluted, if necessary. Page 6 o f 7 E05-0210 Interim Report #5 Page 57 of65 Page 111 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number VOOOI7B5 _______ ________________ANALV11CAL METHOD__________________________ j Method of Analysis for the DMeatmermmainlaLtiiovnerobfyPeLrCfl/uMorSo/oMcStanoic Acid (PFOA) in Small 14.2 Fthoer fsoalmlopwleisngfoerqtiufaietidonwtiothcaklncouwlanteatmheopuenrtcseonft rPeFcOovAeryp.rior to extraction, use Recovery (%) = [total analyte found (ng/mL) - analyte found in control (ng/mL)] analyteadded (ng/mL) 14.3 Unsge/gth(eppfbo)l.lowing equation to convert the amount of PFOA found in ng/mL to PFOA found (ppb) = fPFOA fousnadm(pnlge/mwLei)gxhtf(inga)l volume (mLH E05-0210 Interim Report #5 Page 7 o f 7 Page 58 of 65 Page 112 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 ANALYTICAL METHOD Method Number: VOOOI786 Method of Analysis for theMDeatmermmailnSateirounmofbPyeLrfClu/MorSo/oMctSanoic Acid (PFOA) in Small Analytical Testing Facility: SE30txaSyt8egeRCneoRslleeeagsreeca,hrPcDAhri1v6e801 Approved By: .VPTaeuchl nCiocnalnLolelayder, LC-MS, Exygen Research _D_a_te_IQ lW o 't / JVoihcneJPF/7rlae?hsiedretynt, Operations, Exygen Research Date E05-0210 Interim Report #5 Total Pages: 7 Page 59 oj 65 Page 113 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001786 | ANALYTICAL METHOD Method of Analysis for theMDeatmermmailnaSteirounmobfPyeLrfClu/MorSo/oMctSanoic Acid (PFOA) in Small 1.0 Scope TaScphieidsctmrboyemthHeotirdgichisDtPoeetebrcfeotoermrm(apLnlCocye/MedLSif/qMouriStdh)eiCnihssormolamatlilaotmnogaamrnadmphqayulasnceotriuutamptli.eodn otof paerftlaunodreomoctaMnoaiscs 2.0 Safety 22..21 pACrolewncsaauuyltstiootnhbses.earpvperosapfreialtaebMorSatDorSybperfaocrteicheas.ndling any chemical for proper safety 3.0 Sample Requirement 33..12 3.3 pNsSAaraotommljseeppaacllmeste.tspc1mloemullpseLtrcototociofeatnselslspoitnrwsogaecmidesdptnuoleerceeofsdomewrdpeilxlfelottrberalecsyteistrophuneam.cwifstiaoemdropilonemst.hteeHmsoapwmeerpavltieunrrg,efpbroleazfnoernfeosuresrteuh.mis 4.0 Reagents and Standards 44444.....15324 APMWAemcearefttmhletuoaroonn-nrHoiiotulroPm-iclLetHaA-CnPHcoLgePicCrtLaaAdCtgeecr-aigAddrae-.dCeS.Sig.mRae-aAglednritcGhrade 5.0 Instrument and Equipment 5.1 555555......356724 As5ADA1pIoS20Lnil5dsvammhpc-elemioovyLLgnnstiLhticaddnscebiieLasseplctlpepDqoetbooeurmPcafdssioEolpaaiactlrbpbolnnmreellocaeeadcapertppnrtiwcaorpoocaanweleliwyyptdth-sppaemldbrrm(iaoqo5aloeup0ptvuMa-iyyoatd1hlrlfafeeoiOsarbnncrsOeboehepaulStredecctLoplaeeie,mvnenkns1ocg.tta0irtrltnriuti0ofooftm-uugem20gggre0.aeeer00pati0ettnuthuui0rLjobbe0()nceeLc1.sastCao..pgnr/a.MdbcalqSepu/aMaobnSlftei)t.paotuifmoinnp.jiencgtinugp 5-t2o(112) 555555......1118911320 S2AVDWPuoimasErtptotLeeovprxcssaiepalcSerbeu.aetlutrepemHpsPiPm(ap1kLea0tCVnt0ei-afsv1oc.i0al6d0l.0kcicpt.(Llgan) dtC1108-1S0P0EpcLa)r,trwidigthesd.isposable tips. Page 2 ul`1 E05-0210 Interim Report #5 Page 60 of65 Page 114 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V000 i786 AlS/nATlCAL METHOD Method of Analysis for theMDeatmermmailnSateirounmobfyPeLrCilu/MorSo/oMctSanoic Acid (PFOA) in Small 55..1154 WCernistrt-ifaucgtieoncasphaabkleer.of spinning 15 mL polypropylene tubes at 3000 rpm. 6.0 Chromatographic System 6.1 6666....5234 MTMG(APerno/omaNabbdlp:iiyilleeet8enir2caPPta5thhPul0aarrC5osse-ee:go0rl((5u3aBA2mm0)1)n:3C:::0F) l2MuomeptMhhaasnAeomlRmP o(KnieuymstoAnceetSacteieinntiWfica)t,e2r.1 mm x 50 mm. 5p Tim2208el20...O.f.0050min) %6622655555A %3733755555B FfmloLw00000/.....m33333Riantel 666...876 RQInuujeancnTttiiiomtanteiVo: no-:lu2mP3eeam:kin1Au5rtepeasL.-(ceaxntebrneailnsctraenadsaerddtocaalsibmrauticohnacsu5rv0ep. L). oTphteimabizoevtehceoHndPiLtiConssyastreemin.tended as a guide and may be changed in order to 7.0 MS/MS System 7.1 PMFoOdAe:. Electrospray Negative MRM mode, monitoring 413 -* 369 m /z for TophteimaibzoevtehecMonSdMitiSonssystaerme .intended as a guide and may be changed in order to 8.0 Preparation of Solutions 8.1 M obile Phase 8.1.1 a2mmmMonaiummmaocneituamte taoce1t0a0te0imnLwoaftewraitserp.repared by adding 0.154 g of Alternate volumes may be prepared. Page 3 of 7 E05-0210 Interim Report #5 Page 61 of 65 Page 115 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001786 _______________________ ANALYTICAL METHOD_______________________ Method ofAnalysis for theMDeatmermmailnSateirounmobf PyeLrfClu/MorSo/oMctSanoic Acid (PFOA) in Small 9.0 Standard Preparation 9.1 9999S....t1111a....n1324darTmaafAAmmioPdnrphrfo1LeLSpe0aem1a2tptr.hn.o5o1oso10aaattcf2fxrmhonplkpet5ityechogmh/gLtakFmle/i/demacmoiLaas1Ltna1itetnLDL.tloe00alidLycfsP0fofipDtk1ofcEoa4fopg2raPnrsptrg/b5ttodiCEtmrii/fo-ifeolamifmuitLcpbrinactcdtalLaLonaieasStrottdi.o(tatiosLonciloltoeolanuoiDnuo.olnrrtutsePinrsifoosteo.Eoi-socnlon1ltultubanue0tttbooidti0otitloooatenfnlpneoaffogs.oironf/farfpmiaPrnuPlaeFLarFvlitmOtoOooyvAlaAf)uboxmPleaiiusmiFnsesmdpOtuopoermAdrerfeeoipl1dpafpbu0areyti01reenre0diwwodt0aobedirwbtiyehg1oyfibh0trmfh0briingi6ernmmgeitnghmreLagai1tnontih0ongonwarlgtnmih1toaihngs0Itl 9.2 Standard Calibration Solutions 9.2.1 9.2.2 poTLrfhCetep/hMaefroSe0ld/.l1MoawpsSignnc/egmaelidLisberfdaao.trtiytoipfniiccsaatatlinoednxaasrmdoslpualteiro:enap.dredpitaiorendalincomnectehnatnraotliovnias dilution may be Concentration Final o f Fortification Volume Diluted to Concentration Solution fng/mL) (mL) (m L) (ng/mL) 100 5.0 100 5.0 100 2.0 100 2.0 100 1.0 100 1.0 5.0 10 100 0.5 2.0 10 100 0.2 9.2.3 9.2.4 1naSA.0tetloe2tedrereCndaa.tltolec6vaolCilbu,rm1ua0pteisotonanssidtxacnmodnoacnrdet1hsn0st0i.rnati1o2n5s-mofLstLaDndP0aE.r1dnsamrroawy -bme opurethpabroetdtleass 10.0 Batch Set Up 10.1 10.2 ocREinonaenetcqhchueeiunbrqnetaurmttaracetelhiianottytonesdsaffs(soslacaurmobrfanipcentlorlcedonesltarpeonlxaaldtnnsrdlapafciobktertoewdtr)haoti(tosotyrvpupyerniordctijraufeeylcpaltyltp.iecrd2oa0tceecsodoruanrnltaerdlosslrss)epcimkofevouserstrtiwyfiiineflolcdlrbutedhaetespbaekatcnticlofehwiaesndt Page 4 of? E05-0210 Interim Report #5 Page 62 of65 Page 116 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: P0001131 Exygen Research Method Number V0001786 | analytical m ethod ! Method ofAnalysis fortheMDeatmenmnailnaSteirounmobf yPeLrCflu/MorSo/oMctSanoic Acid (PFOA) in Small 11.0 Sample Extraction 11.1 111111...234 111111...567 11.8 1111..190 11.11 ECLmAACDmdfTMAVaIoSsiloroenedoLdsalueaalnncapmnrdydtaondtatloedoesewrysbwLSniesufitxfztefwhteadeirwampueodbeefermttiogaohentslLdsbhel1tearree,1yaeyomat-tcmtrmprh-osh1s.efeSmr2peuuLenpopatlprmmhLtlcpletCereoeaumeesyiidLrcbffnotnlunosLoueeedusnaosanfganseStmitlfecioemenpiaPatt.HdoprnhfgettcEpinultnePle-ameeldeb3denLnilcienfeeias0ttdaCio.nritctannr0oihorgtmtfdmtwrn0arouaooianepsigdarsfnxadholpetigpSeeanltSmOerCw.thrkaaOusuileeey(mbsfs~mvl(Cosfeil1LSLoo)nar2oL.(CPmlrg-flguddEl5i/-p,empnMNri2aoscolac6m0eeplotpSlaydotir/sov/m2mepnstMisiorfsezruaLiitoplcm2ednSbthue)p)o0gula.am.Lseystetbv.mael.eDyaecabsoLinel"olDlfotpene.aen2naitberscrCsonpllciameusetfaaianiuapwtpLnrlftetgeedorge)rtttei.riivtfescgeuuio1lttohnu-ugs0llattueiabeuolnc.tmmyemetttioua.eLoanbsnngearmdousrsahnfe(aIdatfSsddhukoedraaenrynmturie.3fomdLy5l 12.0 Chromatography 12.1 12.2 12.3 12.4 tsigUAamiesvISLhnnananieetCsntjeutmmjanraheeeessn/lecereepcMpuytdsrtnrntllslzaebpaeetiSdtetdiencerscher/sdda.reaoMedeiesssanlstafeisesr.focaSbtdAe,aaomlrftsumreascsPttsopdtvthaaayehiFleafnemloeesedinOacrdstnbypema.aaiArbmlnlasranSeioetdlbtaaa.tc-ugiesrl1onnonyiaercA0nndrtntttuare.niaaseonorrsiatvsbcanndfitSmpvealaygeytsoslcneapaitlton,dbaiicnlconnfedanrhdaoaeascrifdannldrastea(ritgsrnsdtoaraeqoerdtsnntntmreuos.adatmgsaitcuamsaenrricturseoetttaduotissnlnt,tsesusafbditeasunobtoaeaitsbstleonrtmliedfoonntfiuohpwi.ignfnrvselmtieecceeMandlarufLgauossilasnripdeirstbs1nbdceesmtr/eoryLdxpasfaonatyroewrainredndroctecdtexanssitebciflthegdiaiole3betesahnetn.tdrosw3tdbacatifnaenaeets(irgsndanegioodntmidantrannrojrneapendcefcdtfqlisuvipsoetnaourietelrgovmaniliradvonnnyekasaalwtds)enoyl5a.iaevdn-rrante1abdherIatll0naecesas)tl 12.5 seSxoacfmtewpeadlerestrsaeynsspdtoaernmds.eresshpoounlsdesnsohtoeuxldcebeedfustrathnedradrdilurteesdpoannsderse.anAalnyyzedsa.mples that Page 5 d 7 E05-0210 Interim Report If5 Page 63 oj 65 Page 117 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOO1131 Exygen Research Method Number VOOO1786 | ANALYTICAL METHOD Method ofAnalysis for theMDeatmermmailnSateirounmobfPyeLrfClu/MorSo/oMctSanoic Acid (PFOA) in Small 13.0 Acceptance Criteria 13.1 13.2 13.3 13.4 13.3 13.6 TRHmasaAbtewEMRodsC1htpnan0la3hrfaeoeahnu4aempetr.u0ntceraw4rni9yisoterrgr%r%odonhtp19okraneehmavaccnolk32pntvetrTlaoewidonsrtceyadesolerariemeotorhitrt(isainrdbmtboiste,Rwneeoitcuh.lotrgosexlotuu,aa2tisatnaarrnhcnin.llofttamtimdste2ieinvkbr(mhfostaoeu03Tsaeaeseaenncndtbn.6slpyhom9momuoreP9as2stet8enebbcapbnFhtu0ul5sataeotle4leeaemOsr%s.ae)mnvtevol1ntt.niwsydaAufenal3suIunftotlaxsenehsifsuih)mortcacfcedhtpoaIaedratamitnloifsbewttehublakseceucpefnunedeeedrcosltoolrratuetdesdtaentounpavndomoltspbt(ttnaafina.ebatherRbefyrnaduskdlofineeearsnalrd)IcesalatnkottmftnesanrrthogstPmmderfinlbuotpepoeiorF-schtmebtbaeieflntirooOekeertrtaeaxraabhreranseetncltAirrneeteaortxdaaesesaaiidfarrodnseoashslalaccsssoiyptnauujnoltbtattpuslhlsoefmlagouslryaitsttlasdcntshkalsifmetziutdonmut.dtsettacveaiseoultpssfinoadbentaeirnta1lcrAldn.etdstbemlssbddi0iaoialatoseondroelrrrugrcenniytudtennordfosuhmaeremsgtutesnaurertomd-u/hmteavteabteiimtsfxtlnsaoePxhlenei3eiiytctjdsxteLtnaFt6rerarzhebeirtieOna,c9cedteexegochtobdccedAteteattipawteo.yh.chedssfnmenpauepde.eadetrtulnrlusih.eanrairdteksebahinitbi-eifsifotareenlortebae7lnoxlgondietiLn0,mmmtim,ulroe-Otiweathm1ionwmisatxcatQ3erhsinitcuedw,i0cti.pdllsbosHeuu%eait,tltntrdrhuahIh7tvettfeneghahhiho0neidnnnkeeeeeasft 14.0 Calculations 14.1 gUbeasnseeedtrhaeotenfdoplbleyoawtkhienagrMeeaaq)susuaLstiiynonngxttohsoecfatslwtcaaunrldeaatperrdtohgceruaarmmve:ou(lnint eoafrPrFeOgrAesfsoiounndp(airnamnge/tmerLs). PFOA found (ng/mL) /Peak aresalo-pientercept) x DF x aliquot factor DAlFiq=uofatcftaocrtobry=w2h0ich the final volume was diluted, if necessary. 14.2 Fthoerfsoallmowpleinsgfeoqrtuifaietidonwtoithcaklcnuolwatne tahme opuenrctsenot frePcFoOveAryp. rior to extraction, use Recovery (%) = [total analyte found (ng/mL) - analyte found in control (ng/mL)] ^ _______________________ analyteadded (ng/mL)____________________________ Page 6 o f 7 E05-0210 Interim Report #5 Page 64 of65 Page 118 o f 123 3M Environmental Laboratory Project No. E05-0210 Exygen Protocol Number: POOOl 131 Exygen Research Method Number VOOOI786 | ANa L yTICALMETHOD ~~! Method of Analysis for theMDeatmermmailnSateirounmobfyPeLrfClu/MorSo/oMctSanoic Acid (PFOA) in Small 14.3 Uppsbe.the following equation to convert the amount of PFOA found in ng/mL to PFOA found (ppb) " 1PFOA fosuanmdptlnegv/moluLmt xe f(minLal)volume tmLll E05-0210 Interim Report #5 Page ? of'? Page 65 of65 Page 119 o f 123 3M Environmental Laboratory Project No. E05-0210 Protocol Exygen P0001131; 3M Study Number E05-0210 Amendment 3 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 PROTOCOL AMENDMENT NO. 3 Amendment Date: May 16, 2005 Performing Laboratory 3M Environmental, Health, and Safety Operations 3M Environmental Laboratory 935 Bush Avenue St. Paul, MN 55106 Laboratory Project Identification E05-0210 E05-0210 E05-0210 Interim Report #5 GLP Protocol Amendment #3 Page 1 of 4 Page 120 o f 123 3M Environmental Laboratory Project No. E05-0210 Protocol Exygen P0001131; 3M Study Number E05-0210 Amendment 3 This amendment modifies the following portion(s) of the protocol: P ro tocol rea ds: T e s t in g F a c il it y Exygen Research 3058 Research Drive State College, PA 15801 Phone: (814)272-1039 A m en d to r ea d : T e s t in g F a c h j v e s Exygen Research 3058 Research Drive State College, PA 15801 Phone: (814)272-1039 3M Environmental Laboratory Building 2-3E-09 935 Bush Avenue St. Paul, MN 55106 R eason: Addition of 3M Environmental Laboratory as a testing facility allows selected water samples to be sent to the 3M facility for analysis of PFBS, PFHS, and PFOS using the most current version of ETS 8-154 "Determination of Perfluorinated Acids, Alcohols, Amides, and Sulfonates in Water by Solid Phase Extractions and High Performance Liquid Chromatography/Mass Spectrometry". The following modifications to ETS 8-154.1 will be incorporated into the study: (1) The sample volume extracted and the final SPE elution volume may be adjusted, at the analyst's discretion, to better achieve the detection limits specified in study objectives. As written, ETS 8 154.1 calls for a 40 mL extraction volume with a 5 mL elution volume which results in an eight fold sample concentration. The actual volumes used and the resulting overall concentration factor will be given in the final report. Associated method quality control samples will demonstrate that the volume modifications do not impact method accuracy and precision. (2) A solvent (unextracted) calibration curve may be analyzed with the extracted calibration curve to ascertain extraction efficiency. Samples will be analyzed using the extracted curve unless otherwise stated in the final report. Data from the solvent calibration curve will not be included or discussed in the final report unless it is used to strengthen experimental observations/explanations. (3) ETS 8-154.1 makes no mention of sample surrogate spikes; however, all samples will be spiked with at least one of the following radiolabeled surrogates, PFOA [1,2 13C] (perfluorooctanoic acid), PFOS [1802] and/or PFNA [1,2 13C] (perfluorononanoic acid - C9).The concentration of the surrogate spike may vary depending on the collection event, but typical samples are spiked with a nominal concentration of 0.05 ng/mL. Surrogate spike recoveries will be documented and discussed in the final report. Surrogate recoveries between 10025% will be deemed as meeting method criteria for accuracy. (4) Several method blanks may be prepared and analyzed to better determine a "representative" area count value for the method blank. The method blank area count is instrumental in determining the method limit of quantitation and understanding the average and range of area counts possible is critical. If a method blank value is used as a "zero point" in the final calibration curve, then a brief explanation in the final report or In the raw data as a Note to File must be E05-0210 GLP Protocol Amendment #3 Page 2 of 4 E05-0210 Interim Report #5 Page 121 o f 123 3M Environmental Laboratory Project No. E05-0210 Protocol Exygen P0001131; 3M Study Number E05-0210 Amendment 3 included to describe why that specific method blank was selected as the zero point (closest to the average value, most conservative (largest) value, etc.) (5) The overall analytical uncertainty for each target (non-surrogate) analyte will be estimated using both the method accuracy and precision. The method accuracy and precision will be calculated using data from laboratory contort spike (LCS) samples prepared and analyzed with the sample set(s). Replicate LCSs must be prepared each day samples and/or other quality control samples are prepared. A sample calculation of how the analytical uncertainty was determined will be provided in the final report and/or raw data. A new revision of ETS 8-154 may incorporate all or several of the modifications listed above. If a new version of ETS 8-154 is issued during the course of this study, sample results must meet the new method requirements regardless if they are listed above. 3M Environmental Laboratory management will approve all documented deviations to the 3M Environmental Laboratory quality system (SOPs, methods, etc.) that occur during the course of this investigation. E05-0210 E05-0210 Interim Report #5 GLP Protocol Amendment #3 Page 3 of 4 Page 122 o f 123 3M Environmental Laboratory Project No. E05-0210 Protocol Exygen P0001131; 3M Study Number E05-0210 Amendment 3 Amendment Approval William K. Reenen, Ph.D. 3M Environmental Laboratory Manager Date Jaisimha Kesari, Weston Solutions, Study Director im m fate E05-0210 E05-0210 Interim Report #5 GLP Protocol Amendment #3 Page 4 of 4 Page 123 o f 123