Document RjRRywRzEre40D5ya3Y6ZKeOz

3Afl A&2S16- $S7Z 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 Analytical Report Definitive Study: Quantitative Screening of Commercial Lots of Human Serum Obtained in 2004 for Endogenous Fluorochemicals Using Solid Phase Extraction and Liquid Chromatography/Tandem Mass Spectrometry Laboratory Request Num ber EQ5-0442 Testing Laboratory 3M Environmental Health and Safety Operations 3M Environmental Laboratory' _________________ _ _ _____________________________________________ ;_________ _ 3M Environmental Laboratory Project Lead Mark E. Ellefeon Building 2-3E -09 PO Box 33331 Saint Paul, MN 55133-3331 Phone: 651-778-5405 Testing Facility Management W illiam K. Reagen, Ph.D . Building 2-3E-09 PO Box 33331 Saint Paul, MN 55133-3331 Phone: 651-778-6565 C5 c-n CD S ro 5 = a 00 `J p ^ ^ J Requestor Dale Bacon Building 2-3E-09 PO Box 33331 Saint Paul, MN 55133-3331 Phone: 651-778-4736 Page 1 of 19 3JMENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 3M Environmental Laboratory Analytical Report Project Number: EO5-0442 1 Introduction / Summary The following analytical report presents data from com m ercially available normal pooled human serum that were purchased by the 3M Environmental Laboratory and subsequently screened for endogenous fluorochemicals. Thirty-six separate lots o f pooled normal human serum were extracted using a solid phase extraction technique and screened for sixteen different fluorochem icals using liquid chromatography/tandem mass spectrom etry (LC/MS/MS). Serum samples were received from various suppliers between 9/9/04 and 12/2/04. Samples were extracted on 5/11/2005 and analyzed 5/23/2005. Heptafluorobutyric acid (PFBA) was included in the original list o f sixteen target analytes. However, due to instability in the m ethanol extract, analytical data for PFBA w ill not be included in this report. Instead, a separate study w ill be conducted including the re-extraction o f the thirty-six lots o f commercial serum w ith acetonitrile and analysis using a newly developed method that provides isom er resolution. Those results w ill be reported in a separate report. Sample results are summarized in Tables 1 and 2. The average values reported represent samples in which there was detectable analyte. Sample values that were less than the lim it o f quantitation were not included in the average value calculations. Please refer to Section 5 for individual sample results. Page 2 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.05-0442 Tab! 1. Summary of Perfluorinated Acid Results Analyte Mean Median High Low Standard Deviation Number of Samples with Detectable Analyte NFPA PFHA TDHA PFOA (ng/mL) (ng/mL) (ng/mL) (ng/mL) C9 Acid (ng/mL) CIO ACID (ng/mL) C11 C12 ACID ACID (ng/mU {ng/mL) 1.04 43:01tV 0.151'* 2.98 0.813 0.282! 0.185 0.0246 1.05 2.11 0.327 0.378 0.00838 0.0323 0.00633 0.00822 0.118 0.474 0.0232 0.111 321 0.737 5.72 1.97 0.359 0.0950 1.27 . 0.445 0.281 0.59 0.0274 0.137 0.134 0.734 0.0152 0.176 0.0197 0.0833 0.0108 0.0149 38/36 9/33 33/36 36/316 !! 36/36 36/36 36/36 30/36 Table 2. Summary of Other Perfluorinated and Highly Fluorinated Analyte Results Analyte Mean Median High Lew Standard Deviation FBSA (ng/mL) NA1 NA1 0.00542 FOSA (ng/mL) 0.149 0.124 0.446 0.00542 0.0376 NA1 0.0876 PFBS (ng/mL) 0.0165 0.0101 0:0840 0.00545 PFHS PFOS THPFOS THPFDS (ng/mL) (ng/mL) (ng/mL) (ng/mL) 2.55 14.9 NA2 0.0442 2.85 15.5 NA2 0.0386 4.18 27.0 NA2 0.126 0 .6 0 8 2 .0 3 NA2 0.00772 0.0169 1.07 5.94 NA2 0.0227 Number of Samples with Detectable Analyte 1/36 3 6 /3 6 33/36 36/38 36/38 0 /3 6 NA = Not Applicable because this analyte was only detected in one of t ie samples. NA2 = Not Applicable because this analyte was not detected in any of the samples. 3 8 /3 6 Page 3 p f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 2 Reference Substances Table 3. Reference Substances Name N onafluoropentancic acid Synonym or Acronym NFPA (C5 A cid) Standard Source Material Vendor Traceability Number Purity* A ldrich TCR-333 (00017-045) 96.5% Additional Correction Factor (K* salt or H") 0 .9 9 6 Perfluorohexanoic acid PFHA (C 6 A cid) O akw ood Products T C R -673 97.7% 0 .9 9 7 Tridecafluoroheptanoic acid TDHA (C 7 A cid) A ldrich T C R -267 9 8 .2 % 0 .9 9 7 Perfluorooctanoic acid PFOA (C 8 A cid) 3M TCR-123 (99030-030) 95.0% 0 .9 5 8 H cptadecafluorononanoic acid C 9 A cid Oakw ood Products T C R -618 9 8 .0 % 0 .9 9 8 N onadecafluorodecanoic acid CIO A cid Oakwood Products T C R -36 98.01% 0 .9 9 8 Perfluoroundecanoic acid C ll A cid O akw ood Products T C R -619 9 6 .4 % 0 .9 9 8 Perfluorododecanoic acid C l 2 A cid Oakw ood Products TCR-37 (SD 037) 99.65% 0 .9 9 8 Periluorobutanesulfonnm idc FBSA 3M TCR-314 (00017-026) unknow n N one P erfluorooctanesulfon am ide FOSA 3M TC R -280 (99131-038) 98.94% None P erfluorobutane sulfonate PFBS 3M T C R -l 16 (99030-023) 96.7% 0 .8 8 4 4 Pcrfluorohcxane sulfonate PFHS 3M TCR-83 (SE036) 9 8 .6 % 0.911 Perfluoroociane sulfonate PPOS 3M T C R -6 9 6 86.4% 1H, 1 H .2 H .2 H , pcrfluorooctanesulfonic acid THPFOS SynQ uest Labs TCR-343 (00017-055) 9 3 .5 % 1H, 1H, 2H ,2H , perfluorodecan sulfonate THPFDS Paee/3M T C R -6 2 7 9 4 .7 % Perfluorooctanoic acid, isotopically labeled P erflnorooctanc sulfonate, am m onium salt, isotopically labeled PFOA [ U - IJC] PFOS ["O jI Perkin F.lmer RT1 International T C R -744 T C R -756 >97% 100 ug/m L S ta n d ard Reference substance purities have not been cum *ltd for impurities due to shorter chain homulogues which catmot he distinguished by NMR analysis. 0 .9 2 7 0 .9 9 8 0 .9 9 8 None None . NMR results indicate that the PFOS reference substance consists o f 30.8% branched isom er, and the PFOA reference substance consists o f 22.1% branched isomer. These values were used for the determ ination o f the isomer ratios in the serum samples (presented in Table 10). Page 4 o f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO. EOS-0442 3 Sample Preparation The sam ples for this study were prepared as follows: 1. The serum samples were allowed to thaw and warm to room tem perature. 2. Four m illiliters o f serum (or water in the case o f method blanks) w ere added to a 15 mL screw-capped polypropylene centrifuge tube. 3. M atrix spikes and method blanks were prepared by spiking the serum (or w ater) w ith the analytes o f interest. In addition, l3C PFOA and ,802 PFOS were added as surrogates to all serum samples, calibration standards, m atrix blanks, and method blanks at concentrations o f 1.04 and 0.950 ng/m L serum respectively to evaluate extraction method performance. 4. Ten m illiliters o f acetonitrile were added to the serum and the centrifuge tubes were capped and vortex mixed, allowing the serum proteins to precipitate. 5. The tubes were centrifuged at approxim ately 3500 rpm for 20 m inutes to pellet the precipitate. 6. A fter rem oval from the centrifuge, the supernatant was transferred to a bottle containing 70 m L o f water. 7. Th sam ples were passed through a preconditioned SPE cartridge and eluted w ith 2 mL o f methanol. Q uantitation was accomplished using high performance liquid chromatography tandem mass spectrom etry. The following param eters ware used: L iquid C h rom atograph: Hewlett-Packard* Series 1100 Liquid Chromatograph system Analytical column: Keystone BetasilTM Cu 2.1xl00tmn, 5pm particle size Column tem perature: 30 "C Stop Time: 20.0 m inutes Flow rate: 300 pL/min Injection volume: 5 pL M obile phase components: Solvent A: 2.0 mM ammonium acetate in ASTM Type I w ater Solvent B: HPLC Grade M ethanol Solvent Gradient: Urns 0.00 1.00 14.50 15.50 16.50 20.00 %5 20% 20% 90% 90% 20% 20% Page 5 o f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 M ass S pectrom eter: MDX Sciex API 4000 Q-Trap Mass Spectrom eter D etector Software: AnalystTM Version 1.4 Ionization Mode: Electrospray Negative Temperature: 450 C G SI: 35 GS2: 45 Entrance Potential: -10 Analysis Type: M ultiple Reaction M onitoring (MRM) Table 4. Acquisition Parameters ComDOund NFPA(C5 Acid) PFHA (Ce Acid) TDHA(CrAcid) PFOA (CsAdd) Ce Add C10Aeid Cn Acid C12Acid FBSA FOSA PFBS PFHS PFOS THPFOS THPFDS PFOA [1,2-iaC] PFOS T o d Q1 Mass (amu) 262.9 313.0 362.9 413.0 463.0 512.9 563.0 613.0 297.9 497.9 298.9 398.9 498.9 426.9 526.9 414.9 5034) Q3 Mass tamul 219.0 268.7,118.9 3 1 8 .7 ,1 6 8 .8 ,1 1 8 .9 3 6 8 .9 ,2 1 9 .0 .1 6 9 9 4 1 8 .7 ,1 6 8 .9 ,2 1 8 .9 4 6 8 .8 ,2 1 8 .9 ,2 6 9 .1 518.7,268.9,218.8 568.7.168.9.318.7 780 77.9 9 8 .9 ,7 9 .9 9 8 0 ,8 0 .0 80 .0 ,9 8 .9 ,1 3 0 9 4 0 6 .8 .8 0 .9 5 0 a7 .8 1 .0 369.8 103.0,34.0 Dsclusterina Potential -30 -45 -35 -45 -45 -35 -45 -50 -55 -90 -85 -95 -120 -70 -70 -40 -100 W hen m ultiple transitions were monitored, the total ion current (TIC) was used to quantify each analyte, thereby providing greater sensitivity than individual Q3 ion m easurem ents. 4 Analysis 4.1 Calibration 4.1,1 Calibration Curve An extracted calibration curve was formed by spiking rabbit serum at prescribed levels and extracting the curve in the same manna- as the samples. Stock solutions were individually prepared from neat fluorochemicals and diluted in methanol. Samples were quantified versus a 15-point extracted calibration curve. Page 6 of 19 3U ENVIRONMENTAL LABORATORY PROJECT NO.E05 0442 The regression analysis was performed using a quadratic curve fit weighted 1/x. Calibration standards were within 25% (30% for the Lower Limit o f Quantitation (LLOQ)) with a coefficient o f detennination X).990. Low-level calibration standards which were not within 30% were not included in the calibration curve. Calibration standards that were not greater than 2x the method blanks were not included in the calibration curve, except whore noted. 4.1.2 Continuing Calibration Verification (CCV) A Continuing Calibration Verification (CCV) was run at least once every 10 samples, bracketing the samples with the calibration curve. The CCV recovery was within 25% for all analytes. 4.13 System Suitability Five system suitability standards were analyzed before the initial calibration curve and after the last CCV. These samples had area counts with a %RSD o f <5% and a retention time %RSD o f2% when evaluated independently except for the following instances: The system suitability samples for each o f the perfluorinated acids and FOSA failed. In each case title %RSD o fthe retention times was w ell within tolerance limits, but the area counts varied by as much as 20.3%. The failure o f every perfluorinated acid tested suggests the presence o f a systematic effect on that class o f analytes. This should be kept in mind when evaluating the data presented in this report. 4.1A Lower Lim it of Q uantitation (LLOQ) The LLOQ for each analyte was equal to the nominal concentration o f the lowest standard in the calibration curve that met accuracy requirements o f 70% -130% and that had at least twice as many area counts as the method blanks. 4 2 B lanks 4.2.1 M ethod Blanks Method blanks consisted o f aliquots of Milli-Q water extracted in the same m anna as the samples. All o f tile method blanks in tins study had area counts < 50% o f the LLOQ for their respective analytes with the following exceptions: PFHA: Two out o fthree method blanks was greater than 50% o f the LLOQ; none were above the LLOQ. C7 Acid: One o f three method blanks had area counts greater than 50% o f the LLOQ; none w o e above the LLOQ. C l 1 Acid: One o f three method blanks had area counts greater than 50% o f the LLOQ; none w o e above the LLOQ FOSA: Two out o f three method blanks were greater than 50% o f the LLOQ; none w o e above the LLOQ. Page 7 o f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 PFBS: Two method blanks had area counts greater than 50% o f the LLOQ; one showed anomalous contamination and was above the LLOQ. 42 2 M atrix Blanks Matrix blanks consisted o f aliquots o f blank rabbit serum extracted in the same manner as the samples. Three matrix blanks were prepared and analyzed with the samples. Matrix blanks containing endogenous levels o f analyte that were above the LLOQ are detailed below. Table 5. Endogenous Analyte in Rabbit Serum Sample ID TDHA C11 ACID PFBS THPFOS THPFDS LLOQ (ng/mL) 0.00504 0.00992 0.00506 0.00520 0.00503 MxBIk 050511-1 0.0065 0X>196 0.00941 0.0260* 0.0146 MxBIk 050511-2 0.00252* 0.0138 0.00729 0.0260* 0.0131 MxBIk 050511-3 0.0102 0.0169 0.00774 0.0249 0.0137 Average 0.00641 0.0168 0.00815 0.0256 0.0138 ` Please refer to the iw te to file included in the raw data describing the calculation o f the endogenous levels. The average endogenous values were used to correct the matrix spike response. 4 2 3 Solvent Blanks Methanol blanks were analyzed throughout the analytical run. Analyte response was less than Vi the LLOQ in all cases. Surrogate PFOA [1,2-13C] and PFOS [180z] were added as surrogates to all serum samples, calibration standards, m atrix blanks, and method blanks to evaluate the extraction m ethod perform ance. A ll PFOA [1,2-l3C] surrogate recoveries w ere w ithin 30% , but two of the PFOS [18Oi] recoveries were outside 30%, extending the surrogate recovery range to +/-34%. Specific surrogate recovery data is presented below. Page 8 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 Table 6. Surrogate Recovery Sample ID PFOA [1,2-13C] PFOS [1*OJ Recovery, % Recovery, % M xB lk_050511-1 M xBlk_050511-2 M dB (c_050S11-1 M dBlk_050511-2 LS_Q5Q5115-1 L S _ 0505115-2 L S _ 0505115-3 H S _05 0 5 1 1 S -1 HS 0505115-2 H S_0505115-3 T N -A -6840 T N -A -6643 T N -A -6847 T N -A -6934 T N -A -6935 T N -A -6944 T N -A -6959 T N -A -6960 T N -A -6967 T N -A -6973 T N -A -6975 T N -A -6977 TN-A-6981 110% 102% 80% 99% 114% 109% 98% 85% 120% 90% 80% 93% 78% 102% 89% 80% 85% 78% 91% 76% 84% 82% 83% m% 98% 112% 103% 103% 94% 98% 95% 98% 88% 87% 82% 79% 72% 82% 79% 80% 66% 85% 76% 82% 73% 67% Sample ID PFOA [t,2 -1*C] Recovery, % PFOS f sOd Recovery, % TN-A-6992 TN-A-6994 TN-A-6995 TN-A-6997 TN-A-7000 T N -A -7006 TN-A-6938 TN-A-S939 TN-A-6941 TN-A-6950 TM-A-6953 TN-A-8955 TN-A-6962 TN-A-6964 TN-A-6966 TN-A-8970 TN-A-8972 TN-A-6978 TN-A-3979 TN-A-S982 TN-A-6984 TN-A-6966 TN-A-6988 94% 87% 83% 84% 78% 91% 92% 78% 88% 96% 78% 85% 84% 89% 83% 84% 73% 74% 92% 113% 86% 81% 85% 83% 77% 74% 75% 72% 84% 76% 88% 82% 85% 81% 81% 85% 86% 75% 84% 75% 80% 78% 87% 86% 83% 83% 4 .4 Laboratory Control Spikes (LCS) Six Laboratory Control Spikes (LCS) were prepared in rabbit serum and analyzed w ith the samples. Results were corrected for endogenous levels o f analyte in the m atrix (where appropriate), and are presented below. Page 9 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 Table 7. Matrix Spike Recoveries A n aly te NFPA S a m p le ID Nominal Observed R ecovery, % Cone., ng/mL Cone., ng/mL LS. .0505115-1 LS 0505115-2 LS 0505115-3 HS 050511S-1 HS 0505115-2 HS 0505115-3 0.306 0.306 0 .3 0 9 3.06 3.06 3 .0 6 0 .5 1 7 0 .3 7 3 0 ,3 3 6 2.89 3.48 3.55 169% 122% 109% 94% 114% 116% % RSD 23% 11% PFHA LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0.303 0.303 0.303 3.03 3.03 3.03 0 .6 9 9 0.281 0.224 3.43 2.79 3.09 231% 93% 74% 113% 92% 102% 65% 10% Page 10 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 Table 7. Matrix Spike Recoveries (continued) PFOA LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 ' HS 0505115-2 H S_0505115-3 0 .2 9 7 0 .2 9 7 0.297 2 .9 7 2 .9 7 Z97 0.235 0.129 0.136 2 .4 6 Z82 2.7 C9 Acid LS 0505115-1 LS 0505115-2 LS 0505115-3 H S. 0505115-1 HS 0505115-2 H S _0505115-3 0.300 0.300 0.300 3 .0 0 3 .0 0 3 .0 0 0.285 0 .2 3 7 0.31 2 .1 4 . 3.03 3 .0 4 C 10A C ID LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 H S_0505115-3 0.311 0.311 0.311 3.11 3.11 3.11 0.291 0.323 0 .3 2 3 3.30 3.16 3.06 C12 ACID LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0 .3 0 0 .3 0 0 .3 0 2 .9 9 2 .9 9 2 .9 9 0.323 0.371 0.341 2.91 3.65 3.56 FBSA LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0.300 0.300 0.300 3 .0 0 3 .0 0 3 .0 0 0.305 0*354 0 .3 1 8 3.00 3.25 3.51 79% 43% 4B% 83% 95% 91% 95% 79% 103% 71% 101% 101% 94% 101% 101% 105% 102% 98% 105% 124% 114% 97% 122% 119% 102% 118% 105% 100% 108% 117% 38% 6.9% 13% 19% 5.9% 3.8% 7.0% 12% 7.8% 7.8% Page 11 of 19 Table 7. Matrix Spike Recoveries (continued) 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 A n aly te FOSA PFHS PFOS S a m p le ID US 0505115-1 LS 0505115-2 LS 0505115-3 H S _0505115-1 HS .0505115-2 HS 0505115-3 US 0505115-1 US 0506115-2 US 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 N om inal C o n e ., ng/m L O b se rv e d C o n e ., R e co v e ry , % ng/m L 0.299 0.299 0.299 2.99 2.99 2.99 0.370 0.274 0.251 2 .7 9 3 .4 5 2 .3 6 124% 92% 84% 93% 115% 79% 0.315 0.315 0.315 3.15 3.15 3.15 0.305 0.302 0.317 3 .1 2 2 .8 5 2 .9 0 97% 96% 101% 99% 90% 92% Q C 050202-001 Q C 050202-002 Q C 050202003 C 050202-004 Q C 050 2 0 2 -0 0 5 Q C 050202-006 0.308 0.308 0.308 3.08 3.08 3.08 0.195 0.160 0.230 3 .0 7 3 .0 8 2 .5 2 63% 52% 75% 100% 100% 82% % RSD . 21% 19% 2.6% 4.9% 18% 11% Page 12 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 Tabid 7. Matrix Spike Recoveries (continued and corrected for endogenous target analytes) A nalyte TDHA S am p le ID LS 0505115-1 LS 0505115-2 LS 0505115-3 H S 0505115-1 HS 0505115-2 HS 0505115-3 N om inal C o n e., np/m L 0 .3 0 2 0.302 0 .3 0 2 3.02 3.02 3.02 O bserved C one., ng/m L E ndogenous Level in M atrix, ng/m L O bserved C o n e., C orrected fo r E ndogenous L evel, ng/m L R ecovery, % 0.392 0.354 0.334 2.93 2.84 . 2.74 . 0.00641 0.00641 0.00641 0.00641 0.00641 0.00641 0.39 128% 0.35 115% 0.33 108% 2.9 97% 2.8 94% 2.7 91% % RSD 8% 3% C11 ACID LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0 .2 9 7 0 .2 9 7 0 .2 9 7 2.97 2.97 2.97 PFBS LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0 .3 0 4 0 .3 0 4 0.304 3.04 3 .0 4 3 .0 4 0.355 0.292 0.310 2 .8 9 2 .8 2 2 .6 9 10.8 0.314 0.342 2 .9 7 3 .1 0 3 .1 4 0.0168 0.0168 0.0168 0 .0 1 6 8 0 .0 1 6 8 0 .0 1 6 8 0.00815 0 .0 0 8 1 5 0 .0 0 8 1 5 0 .0 0 8 1 5 0 .0 0 8 1 5 0 .0 0 8 1 5 ! . 0 .3 3 8 0 .2 7 5 0 .2 9 3 2 .8 7 2 .8 0 2 .6 7 10.8* 0 .3 0 6 0.334 2 .9 6 3 .0 9 3 .1 3 114% 93% 99% 97% 94% 90% 3550%* 101% 110% 97% 102% 103% 11% 3.6% 62% 2.9% THPFOS LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0 .3 1 2 0 .3 1 2 0 .3 1 2 3 .1 2 3 .1 2 3 .1 2 NR NR 0.589 2 .9 4 3 .0 6 2 .4 4 0.0256 0.0256 0.0256 0.0256 0.0256 0.0256 NR NR NR NR NR 0.56 181% 2.9 93% 3.0 97% 12% 2.4 77% THPFDS LS 0505115-1 LS 0505115-2 LS 0505115-3 HS 0505115-1 HS 0505115-2 HS 0505115-3 0 .3 0 2 0 .3 0 2 0 .3 0 2 3.02 3.02 3.02 0.330 0.306 0.302 2 .9 4 2 .8 5 2 .9 4 0.0138 0.0138 0.0138 0.0138 0.0138 0.0138 0 .3 1 6 0 .2 9 2 0.2B8 2.93 2.84 2.93 105% 97% i 95% 97% 94% 97% 5.1% 1.8% *This value can be eliminated as an outlier at the 99% confidence level and will be excluded in all further calculations. NR Not Reported because measurable amounts o f target analyte were not observed. Page 13 o f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-Q442 4.5 Accuracy The accuracy for each analyte reported was calculated by taking the average o f the matrix spike recoveries for that analyte. The results are presented in Table 8. Table 8. Method Accuracy for individual Analytes Analyte NFPA PFHA TDHA PFOA C9 Acid C10 Acid C11 Acid C12 Acid % Accuracy 21% 17% 5% 27% B% 1% 2% 14% Analyte FBSA FOSA PFBS PFHS PFOS mpFos THPFDS % Accuracy 8% 2% 3% 4% 21% 12% 3% 4.6 Sample Related Comments The extraction and analysis o f the samples was performed utilizing a non-validated m ethod. The m atrix spike results, PFOA [1,2- C] and PFOS [l8Oz] surrogate recoveries, and the calibration curve should be considered when evaluating the accuracy and precision o f the sample results. 5 Individual Sample Data Tables 9 and 10 summarize individual sample data. Table 9. Perfluorinated Acid Results Sample ID NFPA (ng/mL) PFHA (ng/mL) TDHA (ng/mL) PFOA C9 Acid C10 ACID C11 ACID (ng/mL) (ng/mL) (ng/mL) (ng/mL) C12 ACID (ng/mL) LLOQ (ng/mL) 0.102 0.00505 0.00504 0.0991 0.0150 0.0259 0.00992 0.00997 TNA 6340 0.445 < LLOQ < LLOQ 3.89 0.656 0.198 0 .0 9 6 8 0 .0 2 8 2 TNA5843 1.51 < LLOQ 0.0752 0.509 0.127 0.0768 0.0259 0.0212 TNA 6847 1.56 0.0133 0.118 3.55 0.756 TNA 6934 0.866 0.00633 0.0232 5.72 1.19 TNA 6935 1.04 0.00838 0.0964 3.48 1.17 0.309 0.450 0.551 0.141 0.188 0 .3 8 4 0.0207 0.0264 0.0833 TNA 6944 1.43 < LLOQ 0.290 5.67 1.18 0.584 TNA 6959 0.795 0.00964 0.195 2.52 1.96 0.193 TNA 6960 0.932 0.00889 0.210 2.91 0.611 0.300 TNA 6987 0.450 < LLOQ 0.270 3.66 0.531 0.209 TNA 6973 0.680 0.00652 0.352 3.60 0.940 0.391 TNA 6975 0.682 0.00779 0.398 4.02 0.906 0 .4 0 2 The uncertainty o f these results (based on isotnpically labeled surrogate recovery) is +/- 34%. 0 .1 9 6 0.734 0.121 0.109 0 .1 2 4 0.149 0.0465 0.0370 0.0190 0 .0 1 9 8 0 .0 3 2 8 0 .0 2 9 5 Page 14 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.EOS-0442 Tabla 9. Perfluorinated Acid Results (continued) Sample ID NFPA In g /m L i PFHA (ng/mL) TDHA (ng/mL) PFOA 9 Acid (ng/mL) (ntfmL) C10ACID (ng/mL) C11ACID (ng/mL) C12 ACID (ng/mL) LLOQ (ng/mL) 0.102 0.00505 0.00504 0.0991 0.0150 0.0259 0.00992 0.00997 TNA6977* TNA 6981 1-23 0.327 <LLOQ 0.0442 < LLOQ 0.0555 0.406 2.26 0.0950 0.423 0.0374 0.147 0.0152 0.0708 < LLOQ < LLOQ TN A 6992 1.09 <LLOQ 0.239 3.32 0.842 0 .3 1 9 0 .1 5 8 0 .0 1 2 4 TNA 0994 1.75 0.00694 0.0750 0.770 0 .2 0 6 0.0830 0.142 0.0223 TNA 6995 1.06 < LLOQ 0.198 2 .0 0 1,41 0.169 0 .6 8 8 0.0147 TM A 8997 1.32 < U-OQ 0.179 2 .3 3 1.97 04202 0.637 0.0185 TN A 7000 1.08 < LLOQ 0.174 3.92 0.717 0.332 0 .1 1 2 0.0240 TNA7006 0.818 < U O Q 0.166 3 .3 2 0.686 0.306 0.0946 0 .0 1 2 8 TNA 6938 1.09 < u o q 0.474 4 .3 6 1.20 0.421 0 .1 6 5 0 .0 1 5 0 TNA 6939 0 .6 1 5 0.0323 0.0826 2.01 1.09 0.344 0.28 0.0302 TNA 6941 1.02 < LLOQ 0.0608 4 .0 7 1.61 0.599 0 .4 2 4 0.0527 TNA 6950 2.11 <LLOQ 0.0925 3.20 0.792 04229 0 .1 1 6 0.0115 TNA 6953 1.27 < LLOQ 0.0442 3 .0 8 0.785 0.249 0.0753 0.0150 TNA 6955 1.26 < U .O Q 0.111 2420 0.603 0.341 0 .1 9 6 0 .0 3 0 9 TNA 6962 1.09 < LLOQ < LLOQ 3.51 0.690 0.311 0.111 0 .0 1 8 3 TNA 6964 1.28 U O Q 0.259 4.00 0.933 0.386 0.117 0 .0 1 7 6 TNA 6966 0.665 < LLOQ < LLOQ 2.81 0 .5 6 2 0.249 0.0916 0 .0 1 9 5 TNA 6970 0.781 < u o q 0 .1 5 9 4 .1 7 0.917 0.322 0 ,1 4 8 0 .0 1 1 9 TNA 6972 1.19 < a o o 0.123 3.79 0.761 0257 0 .1 2 6 0.0192 TNA 6978* 1.36 < a o Q 0.0481 0,358 0.114 0.0274 0.0152 < LLOQ TNA 6979 1.11 < a o Q 0.128 3.21 0.568 0.304 0 .1 5 6 0.0161 TNA 6982 0.751 < LLOQ 0.0660 2 .1 6 0.653 0.173 0.0547 < LLOQ TNA 6984 0.899 U Q 0.0636 2 .1 9 0.558 04261 0.164 0 .0 1 0 8 TNA 6986 0.500 < LLOQ 0.0549 2.31 0.460 0.179 0.0394 < LLOQ TNA 6988 0.916 < LLOQ 0.0443 1.93 0.610 0.239 0.164 < LLOQ M ean 1.04 0.0111 0.151 2 9 8 0.813 0.282 0 .1 8 5 0 .0 2 4 6 M edian H igh 1.05 0.00838 0.118 3.21 0.737 2.11 0.0323 0.474 5.72 1.97 0.281 0.599 0.134 0,734 0.0197 0 .0 8 3 3 Low 0.327 0.00633 0X232 0.359 0 .0 9 5 0.0274 0.0152 0 .0 1 0 8 S tan d ard D eviation 0.378 0.00822 a m 1.27 0.445 0.137 0 .1 7 6 0 .0 1 4 9 # OF HITS 36 9 33 36 36 36 36 30 TN-A-6977 and -6078 ate from the name lot ofpooled serum. The uncertainty o f these results (based on isotopically labeled surrogate recovery) is +/- 34%. Page 15 o f 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 Table 9. Sample Results for Additional Perfluorinated and Highly Fluorinated Analytes Sample ID FBSA FOSA PFBS PFHS PFOS THPFOS THPFDS (ng/mL) (ng/mL) <ng/mL) (ng/mL) (iig/mL) (ng/mL) (ng/mL) LLOQ (ng/mL) 0.00501 0.00498 0.00506 0.0158 0.103 0.00520 0.00503 TNA 6840 < LLOQ 0.356 0.00795 3.07 2 1 .9 TNA 6843 TNA 6847 < LLOQ < LLOQ 0.124 0.0694 0.00854 0.00854 0.606 1.62 2 .7 8 2 2 .4 TNA 6934 < LLOQ 0.314 0.0112 1.95 27.0 TNA 6935 < LLOQ 0.133 0.0101 1.75 14.5 TNA 6944 < LLOQ 0.0786 0.00969 3.66 16.9 TNA 6959 < LLOQ 0.232 0.00979 2 .8 5 12.4 TNA 6960 < LLOQ 0.166 0.0107 3.27 14.6 TNA 6967 0.00542 0.446 0.0238 3.04 19.7 TNA 8973 <LLOQ 0.156 0.0172 3.94 16.9 TNA 6975 LLOQ 0.127 0.00858 3.97 18.0 TNA 6977* < LLOQ 0.118 0.0840 0.662 2.03 TNA 6981 < LLOQ 0.122 < LLOQ 1.17 125 TNA 6992 < LLOQ 0.159 0.0103 3.66 16.6 TNA 6994 TNA 6995 < LLOQ 0/116 0.0211 1.37 2 .7 8 < LLOQ 0.248 0.0160 2 .8 8 11.6 TNA 6997 <LL0Q 0.224 0.00942 2.92 12.0 TNA 7000 < LLOQ 0.0906 0.0148 3.97 1 7 .5 TNA 7005 < LLOQ 0.202 0.00989 3.43 19.2 TNA 6938 < LLOQ 0.0446 0.00711 238 16.3 TNA 6939 < LLOQ 0.0967 0.0202 1.65 6.43 TNA 6941 < LLOQ 0.130 0.00991 290 2 1 .8 TNA 6950 < LLOQ 0.0378 0.00545 284 14.9 TNA 6953 < LLOQ 0.0459 0.00804 4.08 2 1 .9 TNA 6955 <LLOQ 0.0981 0.00588 273 9.82 TNA 6962 < LLOQ 0.162 0 .0 1 2 2 3.64 17.8 TNA 6964 TNA 6968 < LLOQ < LLOQ 0.187 0.135 0.0218 0.00802 4.18 3.10 20.1 15.2 TNA 6970 < LLOQ 0.238 0.00737 3.22 18.3 TNA 6972 < LLOQ 0.124 0.0557 282 14.4 TNA 6978* < LLOQ 0.109 0.00577 0.640 2.21 TNA 6979 < LLOQ 0.121 0.0127 1.97 19.0 TNA 6982 < LLOQ 0.0903 0.0144 1.53 14.7 TNA 6984 < LLOQ 0.108 < LLOQ 1.32 14.2 TNA 6986 < LLOQ 0.0712 0.0567 1.57 14.1 The uncertainly o f these results (baseil un isutopically labeled surrogate recovery) is +/- 34%. < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ sLLOQ < LLOQ < LLOQ <LLOQ <LLOQ < LLOQ < LLOQ < LLOQ <LLOQ < LLOQ <LLOQ <L10Q < LLOQ < LLOQ < LLOQ < LLOQ <L10Q <LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ < LLOQ 0 .0 5 6 2 0.106 0.126 0 .0 0 7 7 2 . 0.0457 0 .0 4 8 0.0311 0.0365 0.0425 0.0353 0.0366 0.0413 0.0351 0.0417 0 .0 2 4 9 0 .0 2 8 0.0273 0 .0 3 4 0.0358 0 .0 2 5 9 0 .0 3 4 9 0 .0 2 7 9 0.0753 0.0273 0.0287 0.0345 0.0461 0.0281 0.0466 0.0319 0 .0 4 8 5 0 .0 7 9 9 0.0581 0.0529 0.0459 Page 16 of 19 Mkkfo 3M ENVIRONMENTAL LABORA TORY PROJECT NO.EOS-0442 Table 9. Sample Results for Additional Perfluorinated and Highly Fluorinated Analytes (continued) Sample ID FBSA FOSA PFBS PFHS PFOS THPFOS THPFDS (ng/mL) (ng/mL) (ng/mL) (ng/mL) (ng/mL) (ng/mL) (ng/mL) LLOQ (ng/mL) 0.00501 0.00498 0.00506 0.0158 0.103 0.00520 0.00503 TNA 6988 M ean M edian H igh Low < LLOQ NA NA 0.00542 0.00542 0.101 0 .1 4 9 0.124 0.446 0.0378 < LLOQ 0.01645 0.0101 0.084 0.0055 1.45 2 .5 5 2 .8 5 4 .1 8 0 .6 0 6 15.7 < l-LOQ 0.0586 14.9 NA 0.0442 15.5 NA 0.0366 27 NA 0.126 2.03 NA 0.00772 S ta n d ard D eviation NA 0.0876 0.0169 1.07 5.94 # OF HITS 1 36 33 36 36 ' TN-A-6977 and --6978 are from the same tot o f pooled serum. th e uncertainty o f these results (based on isotopicslly labeled surrogate recovery) is +/- 34%. NA 0.0227 0 36 Table 10. Isomer Ratios for Individual Samples Sample ID TNA6840 TNA6843 TNA6847 TNA 6934 TNA 6935 TNA6944 TNA 6959 TNA 6960 TNA 6967 TNA 6973 TNA 6975 TNA 6977** TNA 6961 TNA 6992 TNA 6994 TNA 6995 TNA 6997 TNA 7000 TNA 7006 TNA 6938 TNA 6939 TNA 6941 TNA 6950 TNA 6953 Sample Origin California M issouri P e n n sy lv a n ia P e n n sy lv a n ia New York M ichigan V irginia V irginia M aryland M ontreal, Q uebec M ontreal, Q uebec M assachusetts Texas C alifornia California California California C alifo rn ia C alifornia New York Naw York New York M ichigan M ichigan PFOA% Branched isomer <LLOQ* <LLOQ* 3.2% 1.3% 6.8% 2.7% 3.9% Z3% <LLOQ* 2.3% 2.3% <LLOQ* <LLOQ* <LLOQ* 3.1% 4.2% 3.9% 2.2% LLOQ* 2.3% 1,1% 0.7% 1.6% 0.8% PFOS % Branched Isomer 37% 35% 42% 46% 34% 33% 37% 36% 40% 39% 40% 35% 40% 36% 38% 39% 39% 40% 39% 40% 33% 35% 37% 39% Page 17 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 TNA 5955 TNA6962 TNA 6964 TNA 6986 TNA 0970 TNA6972 TNA 6978*' TNA 6979 TNA 6982 TNA 6984 TNA 6986 TNA 6988 M ichigan Virginia Virginia V irginia M aryland M aryland M assachusetts M assachusetts Texas Texas Texas Texas 1.6% <LLOQ* <LLOQ*. <LLOQ* <LLOQ* 2.1% <LLOQ* 3.5% 0.8% <LLOQ* <LLOQ* <LLOQ* 34% 39% 39% 37% 38% 36% 36% 30% 41% 30% 41% 33% A verage: 2.5% " * 37% S tan d ard D eviation 1.4%*" 3.3% *The branched isomer in the sample was < LLOQ **TN-A-6977 and -6978 are i'ram the same lot of pouted serum. ***The average and standard deviation represent only those samples with isomer values above the LLOQ. 6 Data/Sample Retention All original raw data and the analytical report have been archived at the 3M Environm ental Laboratory. The test materials and analytical reference standard reserve samples, as w ell as the samples pertaining to this project are archived at the 3M Environmental Laboratory. Reserve samples, digital copies o f original data and all original paper data w ill be retained in the archives o f 3M Environmental Laboratory for a period o f at least 10 years following report signing. 7 C onclusion Under the conditions o f the study, endogenous fluorochemicals were found to be present in all samples. NFPA, PFOA, G> Acid, Cio Acid, Ci i Acid, FOSA, PFHS, PFOS and THPFDS were detected in every lot o f serum. TDHA and PFBS were detected in all but three samples, and the C u Acid was detected in 30 o f 36 samples. Conversely, PFHA was only detected in 9 o f 36 samples, and FBSA was only detected in 1 o f 36 samples. THPFOS was not detected in any o f the samples using the present methodology. PFOA [1,2-13C] surrogate recoveries w ere all w ithin 30% while PFOS [,s02] surrogate recoveries were all within +/- 34%. M ethod accuracy for individual analytes ranged from +/-1% to +/- 27% w hile overall method uncertainty (based on isotopically labeled surrogate recovery) was +/- 34%. Page 18 of 19 3M ENVIRONMENTAL LABORATORY PROJECT NO.E05-0442 The 3M Environmental Laboratoiy's Quality Assurance U nit has audited the data and report for this project. Quality Assurance Representative D a te Page 19 o f 19