Document RpnrDJD2DoX0M9O2eLQVa6RyV
s r l in y T i n .F. Extraction of Potassium Perfluorooctanesulfonate from Mallard Serum and Mallard Liver
for Analysis Using HPLC-Electrospray/Mass Spectrometry
DATA REQUIREMENTS Analytical Method Requirements
STUDY DIRECTOR Sean Gallagher
STUDY COMPLETED ON September 06,2001
PERFORMING LABORATORY / TESTING FACILITY Centre Analytical Laboratories, Inc. (Centre) 3048 Research Drive State College, PA 16801 P h o n e :814-231-8032 STUDY SPONSOR
3M Environmental Technology and Safety Services Building 2-3E-09 PO Box 33331
St. Paul, MN 55133-3331 PROJECT
Centre Protocol Number: 00P-023-042 Centre Study Number: 023-042 Total Pages: 104
000-
Centre Study No.: 023-042
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT
Centre Study Number 023*042, entitled "Extraction o f Potassium Perfluorooccanosulfonate from Mallard Serum and Mallard Liver far Analysis Using HPLC-Hectrospray/Mass Spectrometry," conducted for 3M Environmental Technology and Safety Services, was performed in compliance with US EPA TSCA Good Laboratory Practice Standards (40 CFR Part 792) by Centre Analytical laboratories, Inc,
fnilyR. lacker Principal Investigator Centre Analytical Laboratories, Inc.
Sean Gallagher Study Director Wildlife International, Ltd.
_ 3 lfe je i Date
Dote
aideau Spon/or Repnssentath 3MTn vironmental Technology and Safety Services
Centre Analytical Laboratories, Inc.
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Centre Study No.: 023-042
QUALITY ASSURANCE STATEMENT
Centre Analytical Laboratories' Quality Assurance Unit reviewed Centre Study Number 023-042, entitled, "Extraction of Potassium Perfluorooctanesulfonate from Mallard Serum and Mallard Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry". All reviewed phases were inspected for conduct according to Centre Analytical Laboratories' Standard Operating Procedures, the Study Protocol, and all applicable Good Laboratory Practice Standards. All findings were reported to the Study Director and to management.
Phase
1. Protocol Review
2. Extraction, Fortification
3. Raw Data Review
4. Raw Data and Draft Report Review
5. Final Report Review
Date InsDected
Date Reported to Date Reported to
Centre
Study Director and
Manaeement Sponsor Manaeement
11/20/00
11/30/00
12/28/00
11/20/00
11/30/00
12/28/00
12/12/00 3/18-21/00
12/21/00 3/29/01
12/28/00 5/7/01
9/6/01
9/6/01
9/6/01
______________L-C -- < Naomi Lovallo Sr. Quality Assurance Auditor
9/&lo I Date
Centre Analytical Laboratories, Inc.
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Centre Study No.: 023-042
CERTIFICATION OF AUTHENTICITY
This report, for Centre Study Number 023-042, is a true and complete representation of the raw data for the study.
Submitted by;
Centre Analytical Laboratories, Inc. 3048 Research Drive State College, PA 16801 (814)231-8032
Principal Investigator, Centre:
f e "T V - g ^ e / .Deeper Scientist
Centre Analytical Laboratories, Inc.
Date
Centre Analytical Laboratories, Inc. Facility Management: Centre Analytical Laboratories, Inc.
6 'Je f i~ o \ Date
Study Director, Wildlife International:
ju L . I
Sean Gallagher Wildlife International, Ltd,
Centre Analytical Laboratori, Inc.
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Centre Study No.: 023-042
STUDY IDENTIFICATION
Extraction of Potassium Perfluorooctanesulfonate from Mallard Serum and Mallard Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry
CENTRE PROTOCOL NUMBER: 00P-023-042
CENTRE STUDY NUMBER:
023-042
TYPE OF STUDY:
Analytical
SAMPLE MATRIX:
Mallard Serum and Mallard Liver
TEST SUBSTANCE:
Perfluorooctanesulfonate (PFOS)
SPONSOR:
3M Environmental Technology and Safety Services Building 2-3E-09 POBox 33331 St. Paul, MN 55133-3331
STUDY DIRECTOR:
Sean Gallagher Wildlife International, Ltd. 8598 Commerce Drive Easton, MD 21601
SPONSOR REPRESENTATIVE:
Rochelle Robideau 3M Environmental Technology and Safety Services Building 2-3E-09 POBox 33331 St. Paul, MN 55133-3331
TESTING FACILITY:
Centre Analytical Laboratories, Inc. 3048 Research Drive State College, PA 16801
ANALYTICAL PHASE TIMETABLE:
Study Initiation Date: Experimental Start Date: Experimental Termination Date: Study Completion Date:
11/10/00 11/17/00 03/13/01 09/06/01
Centre Analytical Laboratories, Inc.
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GO0 4 3 5
Centre Study No.: 023-042
PROJECT PERSONNEL
The Study Director for this project was Sean Gallagher at Wildlife International, Ltd. The folowing personnel from Centre Analytical Laboratories, Inc., were associated with various phases of the study:
Name Emily Stauffer Dave Bell Melissa Kennedy Whitsel Tiffany Proctor Angela Morgan Michelle Aijmand Lawrence Ord Rickey Keller Karen Risha
Title Scientist Scientist Group/Team Leader Technician Technician Technician Sample Custodian Sample Custodian Scientist
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Centre Study No.: 023-042
TABLE OF CONTENTS
Page TITLE PA G E............................................................................................................................ 1 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT...............................2 QUALITY ASSURANCE STATEMENT............................................................................. 3 CERTIFICATION OF AUTHENTICITY.............................................................................. 4 STUDY IDENTIFICATION.................................................................................................... 5 PROJECT PERSONNEL......................................................................................................... 6 TABLE OF CONTENTS......................................................................................................... 7 LIST OF TABLES....................................................................................................................8 LIST OF FIGURES................................................................................................................. 10 LIST OF APPENDICES.........................................................................................................11 1.0 SUMMARY..................................................................................................................... 12 2.0 OBJECTIVE.................................................................................................................... 12 3.0 INTRODUCTION............................................................................................................12 4.0 TEST SYSTEM...............................................................................................................13 5.0 REFERENCE MATERIAL............................................................................................ 13 6.0 DESCRIPTION OF ANALYTICAL METHOD...........................................................14
6.1 Extraction Procedure.................................................................................................... 14 6.2 Preparation o f Standards and Fortification Solutions................................................. 14 6.3 Chromatography............................................................................................................15 6.4 Instrument Sensitivity................................................................................................... 15 6.5 Description o f Instrument and Operating Conditions................................................. 15 6.6 Quantitation and Example Calculation........................................................................ 16 7.0 EXPERIMENTAL DESIGN.......................................................................................... 18 8.0 RESULTS........................................................................................................................ 18 9.0 CONCLUSIONS..............................................................................................................18 10.0 RETENTION OF DATA AND SAMPLES.................................................................19
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Table I.
LIST OF TABLES Page
Summary o f PFOS in Reagent Blanks............................................................... 21
Table II. Summary of PFOS in Mallard Liver Blanks...................................................... 22
Table III. Summary of PFOS in Mallard Serum Blanks....................................................22
Table IV. Summary o f PFOS Recoveries in Mallard Liver.............................................. 23
Table V. Summary of PFOS Recoveries in Mallard Serum............................................ 24
Table VI. Summary of PFOS in Adult Liver Samples at 0 ppm a.i. (Test Termination 19 W eeks)..................................................................................... 25
Table VII. Summary of PFOS in Adult Liver Samples at 1.8 ppm a.i. (Test Termination 6 W eeks)....................................................................................... 26
Table VIII. Summary of PFOS in Adult Liver Samples at 6.2 ppm a.i. (Test Termination 6 W eeks)....................................................................................... 27
Table IX. Summary o f PFOS in Adult Liver Samples at 17.6 ppm a.i. (Test Termination 19 W eeks)..................................................................................... 28
Table X. Summary of PFOS in Juvenile Liver Samples at 0 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 29
Table XI. Summary of PFOS in Juvenile Liver Samples at 1.8 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 30
Table XII. Summary of PFOS in Juvenile Liver Samples at 6.2 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 31
Table XIII. Summary o f PFOS in Juvenile Liver Samples at 17.6 ppm a.i. (Test
Termination 12 W eeks)..................................................................................... 32
Table XTV. Summary of PFOS in Control Adult Serum Samples...................................... 33
Table XV. Summary of PFOS in Adult Serum Samples at 0 ppm a.i. (Test Termination 19 W eeks)..................................................................................... 34
Table XVI. Summary of PFOS in Adult Serum Samples at 1.8 ppm a.i. (Test Termination 6 W eeks)....................................................................................... 35
Table XVII. Summary of PFOS in Adult Serum Samples at 6.2 ppm a.i. (Test Termination 6 W eeks)....................................................................................... 36
Table XVIII. Summary of PFOS in Adult Serum Samples at 17.6 ppm a.i. (Test Termination 19 W eeks)..................................................................................... 37
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LIST OF TABLES (Continued) Page
Table XIX. Summary o f PFOS in Adult Serum Samples at 17.6 ppm a.i. (6 Week Samples)............................................................................................................. 38
Table XX. Summary of PFOS in Juvenile Serum Samples at 0 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 39
Table XXI. Summary of PFOS in Juvenile Serum Samples at 1.8 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 40
Table XXII. Summary o f PFOS in Juvenile Serum Samples at 6.2 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 41
Table XXIII. Summary o f PFOS in Juvenile Serum Samples at 17.6 ppm a.i. (Test Termination 12 W eeks)..................................................................................... 42
Table XXIV. Summary o f Percent Moistures for Mallard Liver Sam ples....................... 43
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Centre Study No.: 023-042
Figure 1.
LIST OF FIGURES Page
Typical Calibration Curve for PFO S.................................................................46
Figure 2. Chromatogram Representing a 0.1 ng/mL standard for PFO S...................... 47
Figure 3. Chromatogram Representing a 1.0 ng/mL standard for PFO S...................... 48
Figure 4. Chromatogram Representing a Reagent Blank for PFOS (Centre ID Reagent Blank A, Set: 112000A)...................................................................... 49
Figure 5. Chromatogram Representing Control Mallard Serum for PFOS (Centre ID: 0003803 Blank A, Set: 112100B)......................................................................50
Figure 6. Chromatogram Representing Control Mallard Liver for PFOS, (Centre ID: 0003804 Blank A, Set: 112000A).....................................................................51
Figure 7. Chromatogram Representing Control Mallard Serum fortified with 10 ppb o f PFOS (Centre ID: 0003803 Spk A, Set: 112100B)..............................52
Figure 8. Chromatogram Representing Control Mallard Liver Fortified at 1000 ppb with PFOS (Centre ID: 0003804 Spk A, Set: 112100A).................................53
Figure 9. Chromatogram Representing Juvenile Mallard Serum Sample (Centre ID: 0008143, Sponsor ID: 2332, Set: 112700A)..................................................... 54
Figure 10. Chromatogram Representing Juvenile Mallard Liver Sample (Centre ID: 0008108, Sponsor ID: 2337, Set: 112100A)..................................................... 55
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LIST OF APPENDICES
Page
Appendix A Study Protocol 00P-023-042 (Centre Study No. 023-042) and Amendments and Deviation.......................................................................... 56
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Centre Study No.: 023-042
1.0 SUMMARY
Centre Analytical Laboratories, Inc. (Centre) extracted mallard liver samples and mallard serum samples for the determination of perfluorooctanesulfonate (PFOS) in mallard serum and mallard liver according to protocol 00P-023-042 (Appendix A).
The limit of quantitation for mallard liver was 10 ppb and 10 ng/mL for mallard serum. The LOQ for each matrix was determined in a method verification study performed at Centre (Centre Study: 023-044). Residues of PFOS in the mallard liver samples ranged from non-quantifiable levels (a peak was detected at the corresponding analyte retention time but was less than the lowest concentration of the calibration standards (0.0001 pg/mL)) to 22.8 pg/g. These results are reported on a wet weight basis. PFOS in the mallard serum samples ranged from non-quantifiable levels to 218 pg/g.
The average percent recovery standard deviation for PFOS in liver samples was 95% 12%. The average percent recovery standard deviation for PFOS in serum samples was 93% 10%.
2.0 OBJECTIVE
The objective of this study was to determine levels o f perfluorooctanesulfonate (PFOS) in specimens of mallard liver and mallard serum using the analytical methods described in protocol 00P-023-042.
3.0 INTRODUCTION
This report details the results o f the analysis for the determination o f PFOS in mallard liver and mallard serum, using the analytical methods entitled, "Extraction o f Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry" and "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Serum for Analysis Using HPLC-Electrospray/Mass Spectrometry".
The study was initiated on November 10, 2000, when the study director signed Centre protocol number 00P-023-042. The experimental start date was November 17, 2000, and the experimental termination date was March 13,2001.
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Centre Study No.: 023-042
4.0 TEST SYSTEM
The control mallard liver and serum used for the matrix blanks and matrix fortifications was received on May 18,2000 from Wildlife International Ltd., Easton, MD. The control mallard liver was assigned the Centre ID o f 0003804 and the control mallard serum was assigned the Centre ID of 0003803.
Sixty mallard liver samples and sixty mallard serum samples were received at Centre on August 10, 2000. The samples were stored frozen until they were logged in by Centre personnel on August 16, 2000 and stored frozen. An additional twenty mallard liver samples and forty mallard serum samples were received on February 24, 2001 and stored frozen until they were logged in by Centre personnel on March 1,2001.
Sample log-in and chain of custody information can be found in the raw data package associated with this study. Storage records will be kept at Centre Analytical Laboratories, Inc. and a true copy of the storage records can be found in the raw data package associated with this study.
5.0 REFERENCE MATERIAL
The analytical standard PFOS was received at Centre on June 3, 2000 from 3M Environmental Technology and Services.
The available information for the reference material is listed below. The reference material was stored frozen.
Compound PFOS
Centre Control No.. TCR No.
Purity (% ) Expiration Date
00-023-042
TCR 00017-46
97.9
08/31/01
The molecular structure o f PFOS is given below.
PFOS Chemical Name Molecular weight
o
= =
Perfluorooctanesulfonate 499 (CsFnSCb-)
CaFiyS -- or
O
Note: The neutral molecule and standard form from which PFOS (anion) is derived, is potassium perfluorooctanesulfonate [C8F17SO3K], molecular weight 538.
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6.0 DESCRIPTION OF ANALYTICAL M ETHOD
Analytical methods entitled "Extraction o f Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry" and "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Serum for Analysis Using HPLC-Electrospray/Mass Spectrometry" were used for this study.
6.1 Extraction Procedure
A ~ 0.50 g sample o f liver and a 100 pL aliquot o f the serum was used for the extraction procedure. After fortification o f appropriate samples, the samples were vortexed for ~ 15 seconds. One milliliter o f 0.5 M TBA was added to the liver samples and 0.5 milliliter was added to the serum samples. Two milliliters o f 0.25 M sodium carbonate/sodium bicarbonate was added to the liver samples and 1 milliliter was added to the serum samples. Five milliliters of MTBE was added to both the liver and serum samples. Each sample was placed on wrist-action shaker for ~ 20 min. and then centrifuged for ~ 15 min. Four milliliters o f the organic layer was taken and dried on a nitrogen evaporator and then reconstituted with 1 milliliter o f methanol. Each sample was analyzed by LC/MS/MS electrospray.
6.2 Preparation of Standards and Fortification Solutions
Standard solutions were prepared on September 18, 2000 as specified in Centre Analytical Laboratories' protocol OOP-023-042. An individual stock standard solution of PFOS was prepared at a concentration o f 100 pg/mL by dissolving 10 mg o f the standard (corrected for purity and salt content) in methanol. From this solution, a 1.0 pg/mL fortification standard solution was prepared by taking 1 mL o f the stock and bringing the volume up to 100 mL with methanol.
A 0.1 pg/mL fortification standard was prepared by taking 10 mL o f the 1.0 pg/mL fortification standard and bringing the volume up to 100 mL with methanol. A 0.01 pg/mL standard was prepared by taking 10 mL o f the 0.1 pg/mL standard and bringing to 100 mL with methanol.
A set of standards containing PFOS was prepared by serial dilution o f the 0.1 pg/mL and 0.01 pg/mL solutions in the following manner.
Initial Cone. (pg/mL)1 0.1 0.1 0.1 0.01 0.01 0.01
1of PFOS
Volume (mL) 5 2 1 5 2 1
Diluted to (mL) 100 100 100 100 100 100
Final Cone. (pg/mL)
0.005 0.002 0.001 0.0005 0.0002 0.0001
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Centre Study No.: 023-042
The stock standard solution and all fortification and calibration standard solutions were stored in a refrigerator (4 2C) when not in use. Documentation of standard preparation can be found in the raw data associated with this report.
6.3 Chromatography
Quantification of PFOS was accomplished by LC/MS/MS electrospray. The retention time o f PFOS was ~ 4.4 min. Peaks were detected in the control matrices corresponding to the analyte retention time, but the amounts detected were only significant enough to alter several fortification recoveries and the rest were less than the lowest calibration standard (0.0001 pg/mL).
6.4 Instrument Sensitivity
The smallest standard amount injected during the chromatographic run had a concentration o f 0.0001 pg/mL o f PFOS.
6.5 Description of Instrument and Operating Conditions Instrument: PE SCIEX API 3000 Biomolecular Mass Analyzer
Interface: SCIEX Turboion Spray Liquid Introduction Interface
Computer: Dell UltraScan PI 110
Software: HPLC:
PE SciexAnalyst version 1.1 Windows NT, version 4 H ewlett Packard (HP) Series 1100
HP Quat Pump HP Vacuum Degasser HP Autosampler HP Column Oven
HPLC ColumnrGenesis Cg (Jones Chromatography), 2.1 mm x 50 mm, 4p Column Temp.: 35 C Injection Voi.: 10 pL Mobile Phase (A): 2 mM Ammonium Acetate in ASTM type I water Mobile Phase (B): Methanol Flow Rate: 0.3 mL/min.
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Time % A % B
0 60 40
1.0 0 100
7.0 0 100
7.5 40 60
11.0 40
60
Centre Study No.: 023-042
Ions monitored:
Analvte PFOS
Mode negative
Transition Monitored 499 - 99
Approximate Retention Time ('mini
4.20
Tune File Parameters
Controls IS-Iospray DP-Declustering Potential FP-Focusing Potential EP-Entrance Potential CE-Collision Energy CXP-Collision Cell Exit Potential DF-Deflector CEM-Channel Electron Multiplier
Set -4200.0
-51.0 -230.0 10.0
-70 -5 300.0 2800.0
Gas Flows
Nebulizer Gas Curtain Gas Collision Gas
TIS Temperature
Set
12 13 4 350C
6.6 Quantitation and Example Calculation
Ten microliters o f sample or calibration standard were injected into the LC/MS/MS. The peak area was measured and the standard curve was generated (using 1/x fit weighted linear regression) by Analyst software using six concentrations o f standards. The concentration for mallard liver (pg/g) and mallard serum (pg/mL) was determined from the equations below.
Equation 1 calculated the amount of analyte found (in ng/mL, based on peak area) using the standard curve (linear regression parameters) generated by the Analyst software program. Then Equation 2 calculated the amount o f analyte found in pg/mL (the equations for serum are shown).
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Equation 1:
Analyte found (ng/mL) = (Peak area - intercept^ slope
Equation 2:
Analyte found (pg/mL) = ((anal, found (ne/mLl x FV (mL) x DF x EV (mLY) x 1 pg
(AV (mL) x sample vol. (mL))
1000 ng
Where:
FV = Final Volume
DF = Dilution Factor
EV = Extraction Volume
AV = Aliquot Volume
For samples fortified with known amounts o f PFOS prior to extraction, Equation 3 calculated the percent recovery.
Equation 3: Recovery (%) ~ ((anal, found (pg/mL) - avg. anal, in Ctrl (pg/mL)) x (1000 ng/1 ng)) x 100
amount added (ng/mL)
An example o f a calculation using an actual sample follows:
Mallard serum sample Centre ID 0003803 Spk A (Set: 112100B), fortified at 10
ng/mL with PFOS, where:
peak area
= 6558
intercept
= 515
slope dilution factor
= 7740 =1
ng added (fort level) =
1 ng
avg. amt in controls final volume extraction volume
= =
=
0 (Not quantifiable) 1 mL 5 mL
aliquot volume
= 4 mL
sample weight (volume) = 0.1 mL
From equation I: Analyte found (ng/mL)
-- 16558--5151 7740
From equation 2: Analyte found (pg/mL)
0.8 ng/mL
= (0.8 ng/mL x 1 mL x 1 x 5 mL) x 1 ug
(4mLx0.1mL)
1000 ng
= 0.00976 pg/mL Centre Analytical Laboratories, Inc.
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From equation 3: % Recovery = ((0.00976 pg/mL x - 0 ng/mLl x 1000) x 100 10 ng/mL
= 98%
7.0 EXPERIMENTAL DESIGN
Each set of samples (liver or serum) consisted of one reagent blank, one matrix blank, two matrix blanks fortified at known concentrations, and ~ 20 samples. Each sample was extracted using the appropriate method and then analyzed in duplicate.
8.0 RESULTS
The PFOS found in the reagent blanks are listed in Table I. The PFOS in the mallard liver blanks and mallard serum blanks are given in Tables II & III.
Individual recoveries for PFOS in the liver samples are detailed in Table IV. The average percent recovery standard deviation for PFOS in liver samples was 95% 12%. Individual recoveries for PFOS in the serum samples are detailed in Table V. The average percent recovery relative standard deviation for PFOS in serum samples was 93% 10%.
PFOS in the mallard liver samples ranged from non-quantifiable levels (a peak was detected at the corresponding analyte retention time but was less than the lowest concentration of the calibration standards (0.0001 pg/mL)) to 22.8 pg/g. Individual results are listed in Tables VI-XIII. The results are reported on a wet weight basis.
PFOS in the mallard serum samples ranged from non-quantifiable levels (a peak was detected at the corresponding analyte retention time but was less than the lowest concentration of the calibration standards (0.0001 pg/mL)) to 218 pg/g. Individual results are listed in Tables XIV-XXIII.
The percent moistures for the mallard liver samples are given in Table XXIV.
9 0 CONCLUSIONS
The mallard liver and serum samples were successfully extracted and analyzed according to protocol 00P-023-042.
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10 0 RETENTION OF DATA AND SAMPLES
When the final report is complete, all original paper data generated by Centre Analytical Laboratories, Inc. will be shipped to the sponsor. This does not include facility-specific raw data such as instrument logs, however exact copies o f temperature logs will be submitted. Exact copies o f all raw data, as well as a signed copy o f the final analytical report and all original facility-specific raw data, will be retained in the Centre Analytical Laboratories, Inc. archives for the period of time specified in 40 CFR Part 792. Retained samples o f reference substances are archived by the sponsor.
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TABLES
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Table I. Summary o f PFOS in Reagent Blanks
Sponsor ID
na na na na na na na na na na na na na na na na na na
Centre ID
Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A * Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A* Reagent Blank A Reagent Blank A*
Set Number
111700A 111700A 112000A 112000A 112100A 112100A 112100B 112100B 112700A 112700A 112700B 112700B 030701A 030701A 03070IB 030701B 030801A 030801A
Extraction
Analysis
Date
Date
11/17/00 11/17/00 11/20/00 11/20/00 11/21/00
11/18-19/00 11/18-19/00 11/20-21/00 11/20-21/00 11/22-23/00
11/21/00 11/21/00 11/21/00
11/22-23/00 11/23/00 11/23/00
11/27/00 11/27/00 11/27/00 11/27/00 3/7/01 3/7/01 3/7/01
11/28-29/00 11/28-29/00
11/29/00 11/29/00 3/9/01 3/9/01 3/9-10/01
3/7/01 3/8/01
3/9-10/01 3/11/01
3/8/01
3/11/01
AVERAGE: STANDARD DEVIATION:
Analyte Found (pg/g)
NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ
* Duplicate injection
NQ = Not Quantifiable (A peak was detected at the corresponding analyte retention time but was less than the lowest concentration of the calibration standards (0.0001 pg/mL))
For values recorded as NQ, 0.00005 pg/g (half the value o f the lowest calibration standard, 0.0001 pg/mL) was used to calculate the average and standard deviation.
LOQ = 0.01 pg/g for liver and 0.01 pg/mL for serum
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Table II. Summary of PFOS in Mallard Liver Blanks
Sponsor ID
na na na na na na na na
Centre ID
0003804 Blank A 0003804 Blank A* 0003804 Blank A 0003804 Blank A* 0003804 Blank A 0003804 Blank A* 0003804 Blank A 0003804 Blank A*
Set Number
111700A I11700A 112000A 112000A 112100A 112100A 030801A 030801A
Extraction Analysis
Date
Date
11/17/00 11/17/00 11/20/00 11/20/00 11/21/00 11/21/00 3/8/01
11/18-19/00 11/18-19/00 11/20-21/00 11/20-21/00 11/22-23/00 11/22-23/00
3/11/01
3/8/01
3/11/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (ub/b)
0.000294 0.000300
NQ NQ NQ NQ NQ NQ NQ NQ
Table III. Summary o f PFOS in Mallard Serum Blanks
Sponsor ID
na na na na na na na na na na
Centre
ID
0003803 Blank A 0003803 Blank A* 0003803 Blank A 0003803 Blank A* 0003803 Blank A 0003803 Blank A* 0003803 Blank A 0003803 Blank A* 0003803 Blank A 0003803 Blank A*
Set Number
112100B 112100B 112700A 112700A 112700B 112700B 030701A 030701A 030701B 030701B
Extraction Analysis
Analyte
Date
Date Found (ue/mL)
11/21/00 11/23/00
NQ
11/21/00 11/27/00 11/27/00 11/27/00 11/27/00 3/7/01 3/7/01
11/23/00 11/28-29/00 11/28-29/00
11/29/00 11/29/00 3/9/01 3/9/01
NQ NQ NQ NQ 0.00159 0.00826 0.00825
3/7/01
3/9-10/01
0.00159
3/7/01
3/9-10/01
0.00397
AVERAGE: 0.00239
STANDARD DEVIATION: 0.00333
* Duplicate injection
NQ = Not Quantifiable (A peak was detected at the corresponding analyte retention time but was less than the lowest concentration of the calibration standards (0.0001 pg/mL))
LOQ = 0.01 pg'g for liver and 0.01 pg/mL for serum
For values recorded as NQ, 0.00005 pg/mL (half the value of the lowest calibration standard, 0.0001 pg/mL) was used to calculate the average and standard deviation.
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Table IV. Summary o f PFOS Recoveries in M allard Liver
Sponsor ID
na na na na na na na na na na na na na na na na
Centre
Set
ID Number
0003804 SpkA 111700AD
0003804 Spk A* 111700AD 0003804 SpkB 111700A
0003804 Spk B* 111700A
0003804 Spk A 112000AR 0003804 SpkA* 112000AR
0003804 Spk B 112000A 0003804 SpkB* 112000A
0003804 Spk A 112100A
0003804 Spk A* 112100A 0003804 Spk B 112100AD
0003804 SpkB* 112100AD 0003804 SpkA 030801A
0003804 Spk A* 030801A 0003804 Spk B 030801AD 0003804 Spk B* 030801AD
Extraction Analysis
Amt
Date
Date
Added (ng/g)
11/17/00 11/17/00 11/17/00 11/17/00
11/19/00 11/19/00 11/18-19/00 11/18-19/00
10 10 50 50
11/20/00 11/20/00 11/20/00 11/20/00 11/21/00 11/21/00 11/21/00
11/22/00 11/22/00 11/20-21/00 11/20-21/00 11/22-23/00 11/22-23/00 11/27/00
100 100 4000 4000 1000 1000 20000
11/21/00 3/8/01 3/8/01 3/8/01 3/8/01
11/27/00 3/11/01 3/11/01 3/11-12/01 3/11-12/01
20000 20 20 200 200
AVERAGE: STANDARD DEVIATION:
RELATIVE STANDARD DEVIATION:
% Recovery
79 77 85 84 114 115 106 107 89 88 90 89 92 92 107 101 95 11 12
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 23 of 104
Centre Study No.: 023-042
Table V. Summary of PFOS Recoveries in M allard Serum
Sponsor ID
na na na na na na na na na na na na na na na na na na na na
Centre ID
0003803 Spk A 0003803 Spk A* 0003803 Spk B 0003803 Spk B* 0003803 Spk A 0003803 Spk A* 0003803 Spk B 0003803 Spk B* 0003803 Spk A 0003803 Spk A* 0003803 Spk B 0003803 Spk B* 0003803 Spk A 0003803 Spk A* 0003803 Spk B 0003803 SpkB* 0003803 Spk A2 0003803 Spk A2* 0003803 Spk B2 0003803 Spk B2*
Set Number
112100B 112100B 112100B 112100B 112700A 112700A 112700A 112700A 112700B II2700B 112700B 112700B 030701A 030701A 030701A 030701A 03070IB 03070IB 03070IB 03070IB
* Duplicate injection
Extraction Analysis
Amt
Date
Date Added (ng/mL)
11/21/00 11/21/00 11/21/00 11/21/00 11/27/00 11/27/00 11/27/00 11/27/00
11/23/00 11/23/00 11/23/00 11/23/00 11/28-29/00 11/28-29/00 11/28-29/00 11/28-29/00
10 10 500 500 10 10 10000 10000
11/27/00 11/27/00 11/27/00 11/27/00 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01
11/29/00 11/29/00 11/29/00 11/29/00 3/9/01 3/9/01 3/9/01 3/9/01 3/9-10/01 3/9-10/01 3/9-10/01 3/9-10/01
500 500 100000 100000 100 100 1000 1000 100 100 1000 1000
AVERAGE: STANDARD DEVIATION: RELATIVE STANDARD DEVIATION:
Recovery
98 98 84 86 81 81 91 91 97 97 89 89 79 77 89 93 90 90 118 119 92 11 12
LOQ = 0.01 jtg/g for liver and 0.01 gg/itiL for serum
Centre Analytical Laboratories, Inc.
Page 24 of 104
0005-
Centre Study No.: 023-042
Table VL Summary of PFOS in Adult Liver Samples at 0 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
1961 ^ 1961* 1962 ' 1962* 1963 * 1963* 1964 c 1964* 1965 ^ 1965* 1966 f 1966* 1967 P 1967* 1968 f 1968* 1969 ^ 1969* 1970 1970*
Centre ID
0008006 0008006 0008007 0008007 0008008 0008008 0008009 0008009 0008010 0008010 0008011 0008011 0008012 0008012 0008013 0008013 0008014 0008014 0008015 0008015
Set Number
111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700A 111700ADD 111700ADD 111700AD 111700AD 111700AD 111700AD 111700AD 111700AD
Extraction Date
Analysis Date
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00 11/18-19/00
11/17/00
11/20/00
11/17/00
11/20/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
0.000813 0.000763 0.000509 0.000631 0.000937 0.000887 0.00155 0.00157 0.00110 0.00111 0.00446 0.00432
0.662 0.684 0.238 0.246 0.115 0.106 0.355 0.343 0.138 0.219
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 25 o f 104
000505
Centre Study No.: 023-042
Table VII. Summary of PFOS in Adult Liver Samples at 1.8 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
EOI-0263-21065 ^ E01-0263-21065* E01-0263-21066 ^ EOI-0263-21066* EOI-0263-21067 M EOI-0263-21067* EOI-0263-21068 A ' EOI-0263-21068* EOI-0263-21069 A* EOI-0263-21069* EOI-0263-21075 r EOI-0263-21075* EOI-0263-21076 EOI-0263-21076* EOI-0263-21077 ? EOI-0263-21077* EOI-0263-21078 f E01-0263-21078* EOI-0263-21079 ( ' EOI-0263-21079*
Centre ID
0103870 0103870 0103871 0103871 0103872 0103872 0103873 0103873 0103874 0103874 0103880 0103880 0103881 0103881 0103882 0103882 0103883 0103883 0103884 0103884
Set Number
030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD
Extraction Date
Analysis Date
3/8/01 3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01
3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01
3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01
3/8/01
3/11-12/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (jig/g)
0.359 0.350 0.647 0.631 0.886 0.880 2.11 2.04 0.222 0.232 0.317 0.326 0.219 0.230 0.194 0.197 0.346 0.342 0.0893 0.0888 0.535 0.574
* Duplicate injection
LOQ = 0.01 p.g/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 26 of 104
Centre Study No.: 023-042
Table VIII. Summary of PFOS in Adult Liver Samples at 6.2 ppm a.i. (Test Termination 6 Weeks)
Sponsor
ID
E01-0263-21070 E01-0263-21070* E01-0263-21071 EOI-0263-21071* EOl-0263-21072 EOI-0263-21072* EOI-0263-21073 EOI-0263-21073* EOI-0263-21074 E01-0263-21074* v EOI-0263-21080 ' E01-0263-21080* EOI-0263-21081 EOI-0263-21081* EOI-0263-21082 EOI-0263-21082* EOI-0263-21083 EOI-0263-21083* j EOI-0263-21084 v,, EOI-0263-21084* '
Centre ID
0103875 0103875 0103876 0103876 0103877 0103877 0103878 0103878 0103879 0103879 0103885 0103885 0103886 0103886 0103887 0103887 0103888 0103888 0103889 0103889
Set Number
030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD 030801AD
Extraction
Analysis
Date
Date
3/8/01 3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01
3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01
3/8/01 3/8/01 3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01
3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01 3/8/01
3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01 3/11-12/01
AVERAGE: STANDARD DEVIATION:
Analyte Found (pg/g)
1.04 1.01 0.963 0.907 1.92 1.81 0.791 0.827 1.28 1.34 1.01 1.03 0.0614 0.0578 0.473 0.517 1.66 1.52 0.0862 0.0878 0.920 0.576
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 27 of 104
Centre Study No.: 023-042
Table IX. Summary of PFOS in Adult Liver Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
1991 1991*
1992 f
1992* 1993 v 1993*
1994 r
1994*
1995 a1
1995*
1996 r
1996* 1997 y 1997* 1998 ' 1998* 1999 . 1999* 2000 , 2000*
Centre ID
0008016 0008016 0008017 0008017 0008018 0008018 0008019 0008019 0008020 0008020 0008021 0008021 0008022 0008022 0008023 0008023 0008024 0008024 0008025 0008025
Set Number
111700ADD 111700ADD 111700ADD 111700ADD 111700ADD 111700ADD 111700AD 111700AD 111700AD 111700AD 111700ADD 111700ADD 111700AD 111700AD 111700AD 111700AD 111700ADD 111700ADD 111700AD 111700AD
Extraction Date
Analysis Date
11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00 11/17/00
11/20/00 11/20/00 11/20/00 11/20/00 11/20/00 11/20/00 11/19/00 11/19/00 11/19/00 11/19/00 11/20/00 11/20/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/19/00
11/17/00
11/20/00
11/17/00
11/20/00
11/17/00
11/19/00
11/17/00
11/19/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
15.9 16.4 22.8 21.9 7.02 6.69 2.82 2.93 3.53 3.60 13.2 13.5 5.62 5.58 3.72 3.86 15.5 16.1 5.97 6.10 9.64 6.57
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 28 o f 104
000508
Centre Study No.: 023-042
Table X. Summary of PFOS in Juvenile Liver Samples at 0 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
2301 ^ 2301* 2302 c 2302* 2303 2303* 2305 v ' 2305* 2307 ^ 2307* 2308 M 2308* 2314 2314* 2316 ^ 2316* 2317 2317* 2320 ^ 2320*
Centre ID
0008086 0008086 0008087 0008087 0008088 0008088 0008089 0008089 0008090 0008090 0008091 0008091 0008092 0008092 0008093 0008093 0008094 0008094 0008095 0008095
Set Number
112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A
Extraction Analysis
Date
Date
11/20/00 11/20-21/00
11/20/00 11/20/00
11/20-21/00 11/20-21/00
11/20/00 11/20-21/00
11/20/00 11/20/00 11/20/00 11/20/00 11/20/00 11/20/00
11/20-21/00 11/20-21/00 11/20-21/00 11/20-21/00 11/20-21/00 11/20-21/00
11/20/00 11/20/00 11/20/00 11/20/00 11/20/00
11/20-21/00 11/20-21/00 11/20-21/00 11/20-21/00 11/20-21/00
11/20/00 11/20-21/00
11/20/00 11/20-21/00
11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00
AVERAGE: STANDARD DEVIATION:
Analyte
Found iiiK/g)
NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ NQ
* Duplicate injection
NQ = N ot Quantifiable (A peak was detected at the corresponding analyte retention time but was less than the lowest concentration o f the calibration standards (0.0001 pg/mL))
LOQ = 0.01 pg/g for liver and 0.01 pg/mL for serum
For values recorded as NQ, 0.00005 pg/mL (half the value o f the lowest calibration standard, 0.0001 pg/mL) was used to calculate the average and standard deviation.
Centre Analytical Laboratories, Inc.
Page 29 of 104
000509
Centre Study No.: 023-042
Table XI. Summary of PFOS in Juvenile Liver Samples at 1.8 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
2322 r: 2322* 2323 \ 2323* 2325 2325* 2326 r= 2326* 2328 A' 2328* 2330 Y 2330* 2331 ^ 2331* 2332 f 2332* 2333 fi 2333* 2334 /V\ 2334*
Centre ID
0008096 0008096 0008097 0008097 0008098 0008098 0008099 0008099 0008100 0008100 0008101 0008101 0008102 0008102 0008103 0008103 0008104 0008104 0008105 0008105
Set Number
112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A 112000A
Extraction Date
Analysis Date
11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00 11/20/00 11/20-21/00
AVERAGE: STANDARD DEVIATION:
Analyte Found (ne/e)
0.00454 0.00458 0.00124 0.00129 0.00150 0.00154 0.00236 0.00245 0.00167 0.00165 0.00185 0.00182 0.000719 0.000763 0.000425 0.000430 0.000969 0.000966 0.00356 0.00355 0.00189 0.00126
* Duplicate injection LOQ = 0.01 ng/g for liver and 0.01 (xg/mL for serum
Centre Analytical Laboratories, Inc.
Page 30 of 104
0005. 0
Centre Study No.: 023-042
Table XII. Summary of PFOS in Juvenile Liver Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
2335 2335* 2336 lr 2336* 2337 f 2337* 2338 At 2338* 2339 ? 2339* 2341 ^ 2341* 2342 r 2342* 2343 2343* 2344 f 2344* 2346 ^ 2346*
Centre ID
0008106 0008106 0008107 0008107 0008108 0008108 0008109 0008109 0008110 0008110 0008111 0008111 0008112 0008112 0008113 0008113 0008114 0008114 0008115 0008115
Set Number
112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112100A 112I00A 112100A 112100AD 112100AD
Extraction Analysis
Date
Date
11/21/00 11/21/00 11/21/00
11/22-23/00 11/22-23/00 11/22-23/00
11/21/00 11/22-23/00
11/21/00 11/21/00 11/21/00 11/21/00
11/22-23/00 11/22-23/00 11/22-23/00 11/22-23/00
11/21/00 11/21/00
11/22-23/00 11/22-23/00
11/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/22-23/00 11/22-23/00 11/22-23/00 11/22-23/00 11/22-23/00
11/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/22-23/00 11/22-23/00 11/22-23/00
11/27/00 11/27/00
AVERAGE: STANDARD DEVIATION:
Analyte
Found (pg/g)
0.00197 0.00200 0.00377 0.00378 0.00380 0.00363 0.00555 0.00535 0.00540 0.00518 0.00306 0.00313 0.00312 0.00320 0.00329 0.00332 0.00402 0.00405 0.0141 0.0146 0.00482 0.00341
* Duplicate injection LOQ = 0.01 pg/g for liver and 0.01 jjg/mL for serum
Centre Analytical Laboratories, Inc.
Page 31 of 104
00051
Centre Study No.: 023-042
Table XIII. Summary of PFOS in Juvenile Liver Samples at 17.6 ppm a.i. (Test Termination 12 Weeks)
Sponsor
ID
2347 Ier 2347* 2348 F 2348* 2353 F 2353* 2354 * 2354* 2355 ^ 2355*fC 2357 rft 2357* 2358 P 2358* 2359 p 2359* 2360 ri' 2360* 2363 2363*
Centre
ID
0008116 0008116 0008117 0008117 0008118 0008118 0008119 0008119 0008120 0008120 0008121 0008121 0008122 0008122 0008123 0008123 0008124 0008124 0008125 0008125
Set
Number
112100AD 112100AD 112100AD 112100AD 112100AD 112100AD 112100AD 112100AD 112100A 112100A 112100AD 112100AD 112100AD 112100AD 112100AD 112100AD 112100A 112100A 112100AD 112100AD
Extraction Analysis
Date Date
11/21/00
11/27/00
11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/22-23/00 11/22-23/00 11/27/00 11/27/00 11/27/00
11/21/00 li/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/27/00 11/27/00 11/27/00 11/22-23/00 11/22-23/00 11/27/00
11/21/00
11/27/00
AVERAGE: STANDARD DEVIATION:
Analyte
Found iue/e)
0.0383 0.0370 0.0456 0.0465 0.0234 0.0227 0.0184 0.0189 0.0110 0.0105 0.0616 0.0629 0.0175 0.0189 0.0211 0.0211 0.00623 0.00628 0.0172 0.0191 0.0262 0.0168
* Duplicate injection
LOQ = 0.01 |ag/g for liver and 0.01 fig/mL for serum
Centre Analytical Laboratories, Inc.
Page 32 o f 104
Centre Study No.: 023-042
Table XIV. Summary o f PFOS in Control Adult Serum Samples
Sponsor
ID
E01-0263-21005 E01-0263-21005* E01-0263-21006 ^ E01-0263-21006* E01-0263-21007^ E01-0263-21007* E01-0263-21008 ^ E01-0263-21008* E01-0263-21009 a E01-0263-21009* E01-0263-21023 EOI-0263-21025* EOI-0263-21026 EOl-0263-21026* EOl-0263-21027^ EOI-0263-21027* E01-0263-21028 <' E01-0263-21028* E01-0263-21029 Y E01-0263-21029*
Centre ID
0103810 0103810 0103811 0103811 0103812 0103812 0103813 0103813 0103814 0103814 0103830 0103830 0103831 0103831 0103832 0103832 0103833 0103833 0103834 0103834
Set Number
030701A 030701A 030701A 030701A 030701A 030701A 030701A 030701A 030701A 030701A 030701B 030701B 03070IB 030701B 030701B 030701B 030701B 030701B 030701B 030701B
Extraction Date
Analysis Date
Analyte Found (ue/mL)
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
3/7/01
3/9-10/01
AVERAGE:
STANDARD DEVIATION:
0.0463 0.0455 0.00828 0.00928 0.00954 0.0103 0.00319 \ 0.00452 ^ 0.00423v'n 0.00616 ^ 0.00285 0.00207
NQ NQ NQ NQ 0.00428 0.00265 NQ NQ 0.00797 0.0134
* Duplicate injection
NQ = Not Quantifiable (A peak was detected at the corresponding analyte retention time but w as less than the low est concentration o f the calibration standards (0.0001 (ig/m L))
For values recorded as NQ, 0.00005 pg/mL (half the value o f the lowest calibration standard, 0.0001 pg/mL) was used to calculate the average and standard deviation.
LOQ = 0.01 pg/g for liver and 0.01 pg/mL for serum
Centre Analytical Laboratories, Inc.
Page 33 o f 104
Centre Study No.: 023-042
Table XV. Summary of PFOS in Adult Serum Samples at 0 ppm a.i. (Test Termination 19 Weeks)
Sponsor
ID
1961 M 1961* 1962 F 1962* 1963 1963* 1964 1964* 1965 ^ 1965* 1966 1966* 1967 /A 1967* 1968 f5 1968* 1969 ^ 1969* 1970 f ' 1970*
Centre ID
0008026 0008026 0008027 0008027 0008028 0008028 0008029 0008029 0008030 0008030 0008031 0008031 0008032 0008032 0008033 0008033 0008034 0008034 0008035 0008035
Set Number
112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100B 112100BD 112100BD 112100BD 112100BD 112100BD 112100BD 112100BD 112100BD
Extraction Analysb
Analyte
Date
Date Found (its/mL)
11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/23/00 11/27-28/00 11/27-28/00
0.00605 0.00594 0.00494 0.00501 0.00844 0.00825 0.00798 0.00761 0.0104 0.0103 0.0358 0.0357
6.11 6.02
11/21/00 11/21/00 11/21/00 11/21/00 11/21/00 11/21/00
11/27-28/00 11/27-28/00 11/27-28/00 11/27-28/00 11/27-28/00 11/27-28/00
3.80 3.81 0.843 0.849 1.41 1.38
AVERAGE: STANDARD DEVIATION:
1.22 2.03
* Duplicate injection LOQ " 0.01 |ag/g for liver and 0.01 ng/m L for serum
Centre Analytical Laboratories, Inc.
Page 34 o f 104
Centre Study No.: 023-042
Table XVI. Summary of PFOS in Adult Serum Samples at 1.8 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
E01-0263-21010 *\ E01-0263-21010* E01-0263-21011 *1 E01-0263-21011* EO1-0263-21012/* E01-0263-21012* E01-0263-21013 t* E01-0263-21013* E01-0263-21014 1 E01-0263-21014* EOI-0263-21030 C EOI-0263-21030* E01-0263-21031 ^ E01-0263-21031* E01-0263-21032 E01-0263-21032* E01-0263-21033 p E01-0263-21033* E01-0263-21034 r E01-0263-21034*
Centre ED
0103815 0103815 0103816 0103816 0103817 0103817 0103818 0103818 0103819 0103819 0103835 0103835 0103836 0103836 0103837 0103837 0103838 0103838 0103839 0103839
Set Number
030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD
Extraction Analysis
Analyte
Date
Date
Found (uE/mL)
3/7/01 3/7/01 3/7/01 3/7/01
3/10/01 3/10/01 3/10/01 3/10/01
12.8 12.1 18.2 20.1
3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01
3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01 3/10-11/01
22.5 24.0 20.4 20.4 10.6 10.6 0.870 0.854 6.70 6.43 0.443 0.468 8.88 8.38 1.68 1.74
AVERAGE: STANDARD DEVIATION:
10.4 8.15
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 (jg/mL for serum
Centre Analytical Laboratories, Inc.
Page 35 o f 104
Centre Study No.: 023-042
Table XVII. Summary of PFOS in Adult Serum Samples at 6.2 ppm a.i. (Test Termination 6 Weeks)
Sponsor
ID
E01-0263-21015 ^ E01-0263-21015* E01-0263-21016 E01-0263-21016* E01-0263-21017 ^ E01-0263-21017* E01-0263-21018 *1 E01-0263-21018* E01-0263-21019 M E01-0263-21019* E01-0263-21035 Ie E01-0263-21035* E01-0263-21036 r E01-0263-21036* E01-0263-21037 PE01-0263-21037* E01-0263-21038 fE01-0263-21038* E01-0263-21039 ? E01-0263-21039*
Centre
ID
0103820 0103820 0103821 0103821 0103822 0103822 0103823 0103823 0103824 0103824 0103840 0103840 0103841 0103841 0103842 0103842 0103843 0103843 0103844 0103844
Set
Number
030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701 AD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD
Extraction Analysis
Analyte
Date Date Found (ue/mL)
3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01 3/7/01
3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10/01 3/10-11/01 3/10-11/01 3/10-11/01
47.1 44.7 40.4 35.5 45.5 44.4 23.5 24.7 20.3 20.8 n .r 17.3 2.83
3/7/01 3/7/01 3/7/01
3/10-11/01 3/10-11/01 3/10-11/01
2.87 3.48 4.12
3/7/01
3/10-11/01
61.9
3/7/01 3/7/01 3/7/01
3/10-11/01 3/10-11/01 3/10-11/01 AVERAGE:
60.3 3.69 2.79 26.2
STANDARD DEVIATION:
20.0
* Duplicate injection
LOQ = 0.01 fig/g for liver and 0.01 jig/m L for serum
Centre Analytical Laboratories, Inc.
Page 36 o f 104
Centre Study No.: 023-042
Table XVIII. Summary of PFOS in Adult Serum Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)
Sponsor
ID
1991 M 1991* 1992 F 1992* 1993 M 1993* 1994 F 1994* 1995/1 1995* 1996 P 1996* 1997 1997* 1998 P 1998* 1999 *4 1999* 2000 2000*
Centre ID
0008036 0008036 0008037 0008037 0008038 0008038 0008039 0008039 0008040 0008040 0008041 0008041 0008042 0008042 0008043 0008043 0008044 0008044 0008045 0008045
* Duplicate injection
Set
Number
112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD 112100BDD
Extraction Analysis
Analyte
Date
Date
Found fue/mL)
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
11/21/00
11/28/00
AVERAGE:
STANDARD DEVIATION:
41.0 41.1 47.0 49.5 199 198 93.9 93.0 182 180 218 217 214 212 156 156 156 161 200 205 151 65.0
LOQ = 0.01 jig/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 37 o f 104
Centre Study No.: 023-042
Table XIX. Summary of PFOS in Adult Serum Samples at 17.6 ppm a.i. (6 Week Samples)
Sponsor ID
E01-0263-21020 ^ E01-0263-21020* E01-0263-21021 ^ E01-0263-21021* E01-0263-21022 E01-0263-21022* E01-0263-21023 M E01-0263-21023* E01-0263-21024 S ' E01-0263-21024* E01-0263-21040 C E01-0263-21040* E01-0263-21041 p E01-0263-21041* E01-0263-21042 p E01-0263-21042* E01-0263-21043 P E01-0263-21043* E01-0263-21044 f E01-0263-21044*
Centre ID
0103825 0103825 0103826 0103826 0103827 0103827 0103828 0103828 0103829 0103829 0103845 0103845 0103846 0103846 0103847 0103847 0103848 0103848 0103849 0103849
Set Number
030801AD 030801AD 030701AD 030701AD 030701AD 030701AD 030701AD 030701AD 030801AD 030801AD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD 030701BD
* Duplicate injection
Extraction
Analysis
Date Date
3/7/01
3/11-12/01
3/7/01
3/11-12/01
3/7/01
3/10/01
3/7/01
3/10/01
3/7/01 3/7/01
3/10/01 3/10/01
3/7/01 3/7/01 3/7/01
3/10/01 3/10/01 3/11-12/01
3/7/01
3/11-12/01
3/7/01
3/10-11/01
3/7/01 3/7/01
3/10-11/01 3/10-11/01
3/7/01 3/7/01
3/10-11/01 3/10-11/01
3/7/01
3/10-11/01
3/7/01
3/10-11/01
3/7/01
3/10-11/01
3/7/01 3/7/01
3/10-11/01 3/10-11/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (ufi/mL)
18.6 18.1 152 151 156 157 137 138 42.2 39.1 55.9 62.1 49.8 43.2 15.8 17.7 18.5 17.3 14.1 14.4 65.9 57.5
LOQ = 0.01 ng/g for liver and 0.01 |tg/mL for serum
/
Centre Analytical Laboratories, Inc.
Page 38 o f 104
Centre Study No.: 023-042
Table XX. Summary of PFOS in Juvenile Serum Samples at 0 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
2301 C 2301* 2302 ? 2302*i 2303 2303* 2305 /** 2305* 2307/h 2307* 2308 ^ 2308* 2314 2314* 2316 /V 2316* 2317 r 2317* 2320 r 2320*
Centre ID
0008126 0008126 0008127 0008127 0008128 0008128 0008129 0008129 0008130 0008130 0008131 0008131 0008132 0008132 0008133 0008133 0008134 0008134 0008135 0008135
Set Number
112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A
Extraction Date
Analysis Date
Analyte Found (ug/mL)
11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00
AVERAGE: STANDARD DEVIATION:
NQ NQ NQ NQ 0.00458 0.00453 NQ NQ 0.00278 0.00272 NQ NQ NQ NQ 0.00381 0.00379 0.00626 0.00601 0.00141 0.00153 0.00190 0.00222
* Duplicate injection
NQ = N ot Q uantifiable (A peak w as detected at the corresponding analyte retention tim e but was less than the lowest concentration o f the calibration standards (0.0001 pg/mL))
For values recorded as NQ, 0.00005 pg/mL (half the value o f the lowest calibration standard, 0.0001 pg/mL) was used to calculate the average and standard deviation.
LOQ = 0.01 pg/g for liver and 0.01 pg/mL for serum
Centre Analytical Laboratories, Inc.
Page 39 o f 104
Centre Study No.: 023-042
Table XXI. Summary of PFOS in Juvenile Serum Samples at 1.8 ppm a.i. (Test Termination 12 Weeks)
Sponsor
ID
2322 ^ 2322* 2323 M 2323* 2325 f 2325* 2326 r5 2326* 2328/*4 2328* 2330 2330* 2331 Al 2331* 2332 ** 2332* 2333 F52333* 2334 ^ 2334*-
Centre ID
0008136 0008136 0008137 0008137 0008138 0008138 0008139 0008139 0008140 0008140 0008141 0008141 0008142 0008142 0008143 0008143 0008144 0008144 0008145 0008145
Set Number
112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700D 112700D 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A 112700A
Extraction Date
Analysis Date
Analyte Found (ue/mL)
11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00 11/27/00 11/28-29/00
AVERAGE: STANDARD DEVIATION:
0.0464 0.0472 0.0274 0.0281 0.0497 0.0487 0.0160 0.0156 0.249 0.250 0.00788 0.00747 0.0101 0.00998 0.0113 0.0110 0.0339 0.0339 0.0554 0.0555 0.0507 0.0701
* Duplicate injection LOQ = 0.01 ng/g for liver and 0.01 ig/mL for serum
Centre Analytical Laboratories, Inc.
Page 40 o f 104
Centre Study No.: 023-042
Table XXH. Summary of PFOS in Juvenile Serum Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)
Sponsor
ID
2335 r 2335* 2336 f 2336* 2337 f 2337* 2338 ivA 2338*' 2339 2339* 2341 2341* 2342 * 2342* 2343 PH 2343* 2344 f 2344* 2346 r 2346*
Centre ID
0008146 0008146 0008147 0008147 0008148 0008148 0008149 0008149 0008150 0008150 0008151 0008151 0008152 0008152 0008153 0008153 0008154 0008154 0008155 0008155
Set
Number
112700B 112700B 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700B 112700B 112700D 112700D 112700D 112700D 112700B 112700B
Extraction Analysis
Analyte
Date Date Found Iiie/mL)
11/27/00 11/27/00 11/27/00 11/27/00
11/29/00 11/29/00 11/29-30/00 11/29-30/00
0.0159 0.0162 0.118 0.124
11/27/00 11/27/00 11/27/00 11/27/00 11/27/00
11/29-30/00 11/29-30/00 11/29-30/00 11/29-30/00 11/29-30/00
0.181 0.183 0.0584 0.0592 0.0889
11/27/00 11/27/00
11/29-30/00 11/29-30/00
0.0840 0.0556
11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00 11/27/00
11/29-30/00 11/29/00 11/29/00
11/29-30/00 11/29-30/00 11/29-30/00 11/29-30/00
11/29/00 11/29/00
0.0558 0.0380 0.0383 0.0700 0.0717 0.0676 0.0688 0.Q371 0.0373
AVERAGE: STANDARD DEVIATION:
0.0734 0.0466
* Duplicate injection
LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 41 o f 104
Centre Study No.: 023-042
Table XXIII. Summary of PFOS in Juvenile Serum Samples at 17.6 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
2347 F 2347* 2348 ir 2348* 2353 F 2353* 2354 2354* 2355
2355* 2357 AO 2357* 2358 ? 2358* 2359 F 2359* 2360 2360* 2363 A 2363*
Centre ID
0008156 0008156 0008157 0008157 0008158 0008158 0008159 0008159 0008160 0008160 0008161 0008161 0008162 0008162 0008163 0008163 0008164 0008164 0008165 0008165
Set
Number
112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D 112700D
Extraction Analysis
Analyte
Date
Date
Found (ue/mL)
11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00 11/27/00 11/29-30/00
AVERAGE: STANDARD DEVIATION:
0.490 0.496 0.150 0.156 0.894 0.908 0.591 0.621 0.399 0.384 0.547 0.547 0.512 0.517 0.402 0.404 0.521 0.514 0.439 0.434 0.496 0.184
* Duplicate injection LOQ = 0.01 |i.g/g for liver and 0.01 |xg/mL for serum
Centre Analytical Laboratories, Inc.
Page 42 o f 104
Centre Study No.: 023-042
Table XXIV. Summary of Percent Moistures for Mallard Liver
Samples
Sponsor
Centre
Average
ID ID % Moisture
1961
0008006
69.7
1962
0008007
73.4
1963
0008008
75.7
1964 0008009 69.2
1965
0008010
71.9
1966 1967 1968 1969 1970 1991 1992 1993 1994 1995 1996 1997 1998 1999 2000 2301 2302 2303 2305 2307 2308 2314 2316 2317 2320 2322 2323 2325 2326 2328 2330 2331 2332 2333 2334 2335
0008011 0008012 0008013 0008014 0008015 0008016 0008017 0008018 0008019 0008020 0008021 0008022 0008023 0008024 0008025 0008086 0008087 0008088 0008089 0008090 0008091 0008092 0008093 0008094 0008095 0008096 0008097 0008098 0008099 0008100 0008101 0008102 0008103 0008104 0008105 0008106
75.0 72.8 70.4 69.5 72.9 75.3 71.8 71.9 71.8 71.5 72.4 70.5 71.3 69.8 72.0 76.6 77.3 72.7 72.5 72.3 72.0 75.0 69.6 71.8 71.6 71.6 72.5 73.2 72.6 72.0 68.9 71.4 71.7 69.8 68.1 71.7
Centre Analytical Laboratories, Inc.
Page 43 o f 104
Centre Study No.: 023-042
Table X X IV (cont'). Summary o f Percent M oistures for M allard Liver
Samples
Sponsor
Centre
Average
ID__________ ED______% Moisture
2336 2337 2338 2339 2341 2342 2343 2344 2346 2347 2348 2353 2354 2355 2357 2358 2359 2360 2363 E01-0263-21065 E01-0263-21066 E01-0263-21067 E01-0263-21068 E01-0263-21069 E01-0263-21070 E01-0263-21071
EO1-0263-21072
E01-0263-21073 EO1-0263-21074 E01-0263-21075 E01-0263-21076 E01-0263-21077 E01-0263-21078 E01-0263-21079 EO1-0263-21080 EO1-0263-21081 EO1-0263-21082 EO1-0263-21083 EO1-0263-21084
0008107 0008108 0008109 0008110 0008111 0008112 0008113 0008114 0008115 0008116 0008117 0008118 0008119 0008120 0008121 0008122 0008123 0008124 0008125 0103870 0103871 0103872 0103873 0103874 0103875 0103876
0103*77
0103878 0103879 0103880 0103881 0103882 0103883 0103884 0103885 0103886 0103887 0103888 0103889
69.7 69.7 69.8 70.6 72.6 78.2 70.9 69.2 70.9 69.7 70.3 73.2 68.5 67.8 74.5 72.0 73.6 69.0 74.1 73.8 81.0 82.4 75.8 78.2 77.4 77.5
74.8
77.5 82.7 71.7 77.9 77.5 76.0 63.8 78.2 73.6 76.7 81.4 72.3
Centre Analytical Laboratories, Inc.
Page 44 o f 104
ooo
/
Centre Study No.: 023-042
FIGURES
Centre Analytical Laboratories, Inc.
Page 45 o f 104
Centre Study No.: 023-042
Figure 1. Typical Calibration Curve for PFOS
q ri
!m I i
q Tf
to to
;^T :^ I # ?
q to
in I cm" t' i
finnrpntratinn pg/mL
{C'if
Centre Analytical Laboratories, Inc.
Page 46 o f 104
Centre Study No.: 023-042
Figure 2. Chromatogram Representing a 0.1 ng/mL standard for PFOS
gamp
| Sample Index: ( Sample Type:
! Concentration: I Acq. Date: ; Acq. Time:
Standard
0.100
11/22/00 12:11:52 PM
; Modified: I Bunching Factor: ! Noiae Threshold: I Area Threshold: i Num. Smooths: I FT Window:
Expected FT: Sep. Width: Sep. Height: Exp. Peak Ratio: Exp. AcTj. Ratio: Exp. Val. Ratio: Use Relative RT:
Y1es
2.82
12410..008
4.52 0.20 0.01 5.00
4.00
3.00
No
sec min
Int. Type: Retention Time: Area:
Manual 4.55
1548.4
min counts
`* "'
l, . J '* , /' `
_,v*,' ! /*' '
4.54
.. .
Centre Analytical Laboratories, Inc.
Page 47 of 104
Centre Study No.: 023-042
Figure 3. Chromatogram Representing a 1.0 ng/mL standard for PFOS
spi
5 Comment____
| Sample Index: Sample Type:
Concentration: ' Acq. Date: ; Acq. Time:
Sta1n.0d0a0rd
11/ 22/00 12:49:51
"ritBr-irn-fsf^l-:r ^ 2A`^
2695 2500
Modified: : Bunchinq Factor:
' Noise Threshold: Area Threshold: ! Num. Smooths: AT Window: | Expected RT: ] Sep. Width; * Sep. Height: | Exp. Peak Ratio:
Exp. Adj. Ratio: 1 Exp. Val. Ratio: | Use Relative RT:
Y1es
2,82
114.08
30.0
4,52 0.20 0.01 5.00
4.00
3.00 Mo
sec xin
2000
& 1500 I
1000
600
rlnt. Type: * Retention Time:
Area:
Manual 4.54
8355.5
dn counts
4.54
34 Tim, min
S
Centre Analytical Laboratories, Inc.
Page 48 o f 104
*"0528
Centre Study No.: 023-042
Figure 4. Chromatogram Representing a Reagent Blank for PFOS (Centre ID Reagent Blank A, Set: 112000A)
Semple Index: Sample Type:
Concentration: Aeq. Date: Acq. Time:
8
Bo.l-aoonok
11/20/00 05:21:16 PM
Modified:
Yes
Bunching Factor: 1
Hois Threshold: Ax.a Threshold: Nun. Smooths:
2.82 14.Q8
1*
8a
RT Window:
30.0 sec 2?
Expected RT:
4.52 in 1
Sap. Width:
0.20
Sep. Height:
0.01
--
Exp. peak Ratio: S..00
Exp. Adj. Ratio: 4.00
Exp. val. Ratio: 3.00
Use Relative RT: No
InC. Type: Retention Tima: Area:
Manual 4.57
432.0
min counts
139 . 120
ioo j
801Ii i 80.
! 40I
201
0-
1. 2
i. -iK :v
4.66
4.47f/^ ,^5.,00
8.9l* 7.22*
i
Time, min
T
Centre Analytical Laboratories, Inc.
Page 49 o f 104
000529
Centre Study No.: 023-042
Figure 5. Chromatogram Representing Control Mallard Serum for PFOS (Centre ID: 0003803 Blank A, Set: 112100B)
Sample Index: Sample Type: Concentration: Acq. Date: Acq. Time:
10 ' Blank 0.000 11/23/00 03:38:52 AM
,
200
4.53
'Modified: Bunching Factor:
Ya 1
150
Noise Threshold: 2.82
Area Threshold: Nun. Smooths:
14.08 1
&
*
RT Window: Expected RT: Sep. Width: Sep. Height:
30.0
4.52
S0.20 0.01
sec min
ec
100
i
Exp. Peak Ratio: 5.00
Exp. Adj. Ratio: 4.00
so
Exp. Val. Ratio: 3.00
4.66
Use Relative RT: Ho
Int. Type:
Manual
Retention Time:
4.54
min
4,45
VK5.07 . 5 M
6.4+ # J7
770
'V- >_/-'/--A-J.V/*' **v.
0-
123
4
67
Area:
612.8 counts
Time, min
Centre Analytical Laboratories, Inc.
Page 50 o f 104 G O ! <r*
Centre Study No.: 023-042
Figure 6. Chromatogram Representing Control Mallard Liver for PFOS, (Centre ED: 0003804 Blank A, Set: 112000A)
cffimMtSBSowoS?
I Sample Index:
10
; Sanpl Typ: 1 Concentration:
*Blank
0 000
1 Aeq. Oat: Acq. Tim:
11/20/00 05:46:39 PH
Modified: Bunching Factor: Moil Threshold: Area Threshold: Hum. Smooths: RT Window: Expected RT: Sep. Width: Sep. Height: Exp. Peak Ratio: Exp. Adj. Ratio: Exp. Val. Ratio: Us Relativ RT:
ya i 2.82
14.08 1
30.0
4.52 0.20 0.01 5.00
4.00 3.00 No
IK in
1 |
Int. Typ: Retention Time: Area:
Manual 4.56 in
529.8 counts
227 200 | 150 100 50
4.50
4.43
4.66
I- .5.00 S ''0
6.60
7.58
Tim, min
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Centre Study No.: 023-042
Figure 7. Chromatogram Representing Control Mallard Serum fortified with 10 ppb of PFOS (Centre ID: 0003803 Spk A, Set: 112100B)
Sample Index: Sample Type:
Concentration: Acq. Date: Acq. Time:
QC . 0.800 11/23/00 04:04:15 AM
Modified:
Yes
Bunching Factor: 1
Noiae Threshold: 2.82
Area Threshold: ' 14.08
Num, Smooths:
1
8
RT Window:
30.0 sec
Expected RT:
4.52 min c
Sep. Width: Sep. Height:
0.20 0.01
j J5
Exp. Peek Ratio: 5.00
Exp. Adj. Ratio: 4.00
Exp. Val. Ratio: 3.00
Use Relative RT: No
Int. Type:
Manual
Retention Time:
4.53
Area:
\ 6557.7
tin counts
21! 2000
1500 1000
500,
4.53
4 Time, min
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Figure 8. Chromatogram Representing Control Mallard Liver Fortified at 1000 ppb with PFOS (Centre ID: 0003804 Spk A, Set: 112100A)
Sample Index: Sample Type:
Concentration: Acq. Date: Acq. Tine:
12 QC
1.500 11/22/00 02:31:00 PM
Modified: Bunching Factor; Noise Threshold: Area Threshold: Nun. Smooth*: RT Window:
Expected RT: Sep. Width: Sep. Heiqht: Exp. Peak Ratio; Exp. Ad}. Ratio: Exp. Val. Ratio: Use Relative RT:
Yes i 2.12 .14.08 1 30.0
4.52 0.20 0.01 5.00 4.00 3.0Q No
sec min
# I *
Int. Typo:
Manual
Retention Tine:
4.-54
Area:
10563.0
min counts
3251 3000 2500 2000 1500 1000 .500
4.53
Tim, min
57
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Figure 9.
Chromatogram Representing Juvenile Mallard Serum Sample (Centre ID: 0008143, Sponsor ID: 2332, Set: 112700A)
slllllSIj i l l
SM plO In d M t
is
io a p lo Type:
C oncontxatioa: Acq. D lM i Acq. T U a:
Onkaova
/A
11/29/00 0 4 :0 7 :5 1
M odiflad>
itching re c to r:
M oiao T h ro e h o ld : X rta T hroahold: No*, f o o t ha: RT W indow: E ap o c ta d RT: Sop. W idth: Sop. B oightt Cap. foak R atio : Cap. A d). M tio : Cap. V ii! R atio:
Oao Ra l a C lvo RT:
Yat 1
2 .1 2 1 4 .0
1 2 0 .0
4 .SI 0 .2 0 0 .0 1 5 .0 0
4 .0 0 2 .0 0 (to
t a t . Typo:
R o to a tle n Tiaw : A zoa:
holgfce: S ta r t T ina: Cnd T ijao:
M anual 4 .3 0
4 * 7 7 .3
1.41*0003 4 .4 0
4 .5 1
j1400
11tM2100O00.-,IJ
TOO MO MO
4.0 Tim, mn
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Centre Study No.: 023-042
Figure 10. Chromatogram Representing Juvenile Mallard Liver Sample (Centre ID: 0008108, Sponsor ID: 2337, Set: 112100A)
8_______ Sample index: Sample Type:
Concentration: Acq. Date: Acq. Time:
21
Unknown
N/A 11/22/00 04:25:14 PM
j Modified;
I Bunching Tactor: ; Noise Threshold: | Area Threshold: | Hum. Smooths: 1 RT Window: 1 Expected RT: i Sep. Width; ' Sep. Height:
: Exp. Peak Ratio: j Exp. Adj. Ratio: ( Exp. Val. Ratio: i Use Relative RT:
Y1es
2.82
114.08
30.0 4.52 0.20 0.01 5.00
4.00 3.00
No
sec in
I Int. Type:
Manual
1 Retention Time:
4.54
i Area:
11166.9
min counts
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APPENDIX A
Study Protocol 00P-023-042
(Centre Study No. 023-042) and
Amendments and Deviation
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STUDY PROTOCOL
STUDY TITLE EXTRACTION OFPOTASSIUM PERFLUOROOCTANESULFONATE FROM MALLARD SERUM AND MALLARD LIVERFOR ANALYSIS USING HPLC-
ELECTROSPRAY/MASS SPECTROMETRY
SPONSOR 3M Environmental Technology and Safety Services
Building 2-3E-Q9 PO Box 33331
SuPhul, MN 55133-3331
DATA REQUIREMENTS Analytical Method Requirements
TESTING LABORATORY Centre Analytical Laboratories. Inc. (Centre)
3048 Research Drive State College, PA 16801
Phone 814-231-8032
PROTOCOL IDENTIFICATION NUMBERt OOP-023-042
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Centre Protocol No. OOP-023-042
Title: EXTRACTION OFPOTASSIUM PERFLUOROOCTANESULFONATEFROM MALLARD SERUM AND MALLARD LIVERFOR ANALYSIS USING HPLCELECTROSPRAY/MASS SPECTROMETRY
TABLE OF CONTENTS
. _________ __________________________ ____ __ ?BSE.
TABLE QF CONTENTS.............................................................................................. 2
1. PURPOSE....................................... ...........................................................-- 3 2. TEST MATERIALS.......................................................................................... 3
3. SPONSOR.........................- ...................................................... ......... ..............4 4. TESTING FACILITY AND PERFORMING LABORATORY.................. 4 5. PROPOSEDEXPERIMENTALTIME-FRAME...................... .................. . 4
6. JUSTIFICATION FOR THE SELECTION OFTHE TEST SYSTEM................ 4
7. SAMPLEPROCESSING, STORAGE AND IDENTIFICATION....................... 5
8. ANALYTICAL METHOD................................................................................. 5
9. EXPERIMENTALDESIGN.......................................
10'
10. PROTOCOL AMENDMENTS AND DEVIATIONS..........................................10
11. RECORDS................................................................................................. 11 12. QUALITY ASSURANCE................................................................................11
13. DATA AND REPORT...................................................................................... 12
14. ARCHIVESTATEMENT................................................................................ 12
15. REFERENCES..................................................................................................13
16. PROTOCOLAPPROVAL................................................................................ 14
APPENDIX: ANALYTICAL METHODS................................................................... 15
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Studv Title:
EXTRACTION OF POTASSIUM PERFLUOROOCTANESULFONATEFROM
MALLARD SERUM AND MALLARD LIVER FORANALYSIS USING HPLCELECTROSPRAY/MASS SPECTROMETRY
1. PURPOSE
The purpose of this study is to analyze mallard serum samples and mallard liver samples for residues of perfluorooctanesulfonate (PFOS) using methods entitled, '"Extraction Of Potassium Perfluorooctanesulfonate Or Other Fluorochemical Compounds From Serum For Analysis Using HPLC-Electrospray/Mass Spectrometry" (3M method number ETS-8-4.1) and "Extraction Of Potassium Perfluorooctanesulfonate Or Other Fluorochemical Compounds From Liver For Analysis Using HPLC-Electrospray/Mass Spectrometry" (3M method number ETS-8-6.0) with'the modifications listed in Section 8.
The Quality Assurance Unit of Centre Analytical Laboratories, Inc., will audit the study for compliance with EPA TSCA Good Laboratory Practice standards 40 CFR Part 792(1).
2. TEST MATERIALS The following analytical standards will be used:
Test Material PFOS
TCR Number TCR00017-46
Purity (%) 97.9
Expiration Date 08/31/01
Chemical name and structure of the compound is presented below.
PFOS Chemical Name: IUPAC Name:
CAS Number Molecular Weight:
Perfluorooctanesulfonate
1 -Octanesulfonic acicU.1,2,2,3,3,4,4,5,5,6,6,7,7,8,8,8heptadecafluoro-, potassium salt 2795-39-3'
499 (CbFnSOT)
O
C8F 17 S -- 0
O
Note: The neutral molecule and standard form that PFOS (anion) is derived from is perfluorooctanesulfonate potassium salt [QF17SO3K], molecular weight 538.
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A record of test and reference substance receipt, storage conditions, and a record of use will be maintained at Centre Analytical Laboratories, Inc. Forms documenting chain-of-custody and shipping records for tracking of the test substances will be included as part of therawdatapackage.
All standards/test substances and any preparedsolutions must be identified with a unique label ornumberon thecontainerorcross-referenced to the container.
HAZARD INFORMATION
A current MSDS for the chcmical(s) used in this study will be testing facility.
at ^
3. SPONSOR
3MEnvironmental Technology and SafetyServices Building 2-3E-09 POBox 33331 St. Paul, MN55133-3331
STUDY DIRECTOR: Rochelle Robideau TELEPHONE: 651-778-7065 FAXNUMBER: 651-778-6176
4. TESTING FACILITYAND PERFORMINGLABORATORY
Centre Analytical Laboratories, Inc. (Centre) 3048 ResearchDrive State College, PA 16801
PRINCIPALINVESTIGATOR: EmilyR. Stauffer, Centre TELEPHONE: 814-231-8032 FAXNUMBER: 814-231-1580
5. PROPOSED EXPERIMENTAL TIME-FRAME
Experimental Start Date Experimental TerminationDate Report Issuet)
Nov. 6,2000 Nov. 24,2000 Dec. 30,2000
6. JUSTIFICATIONFORTHESELECTION OFTHE TEST SYSTEM
Mallard serumsamples and liver samples collected by Wildlife International, Ltd. will be used as the test systems for this study. Mallards represent wild* bird populations and they are an EPA recommended species because they do well in a laboratory environment. Mallard habitats include ponds, lakes and marshes, and
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they are considered aquatic grazers. The use of these matrices for the study was to determine if PFOS was deposited in the liver and serum of mating adults that were fedPFOS.
7. SAMPLEPROCESSING,STORAGE AND IDENTIFICATION
No processing will be required for the serum samples. Due to the small amount of sample available, the liver samples will not be homogenized priorto extraction; however, they will be homogenized by lying the samples on a flat surface and mashingthemby handas best as possible.
Each sample will be assigned a unique sample identification number at Centre, which will be used for tracking and identification of the samples. The samples will be stored in a temperature'monitored freezer, maintained at < -10 C, except when removed for extraction and analysis as described in the method. The samples will be kept isolated from the test substance during storage.
Sample receipt and storage location and conditions during the study will be documented. All samples and any resulting sample extracts will be identified
with a unique label or sample number. Such identification will be either on the containerorcross-referenced tothe container.
8. ANALYTICALMETHOD
_
Th serum samples will be analyzed according to the analytical method titled "Extraction Of Potassium Perfluorooctanesulfonate Or Other Fluorochentical Compounds From Serum For Analysis Using HPLC-Electrospray/Mass Spectrometry" (3M methodnumberETS-8-4.1) andthe control liver samples will b analyzed according to the method titled "Extraction Of Potassium Perfluorooctanesulfonate Or Other Fluorochemical Compounds Rom liver For Analysis Using HPLC-Bectrospray/Mass Spectrometry" (3M method number ETS-8-6.0). The following modifications will be applied to each method:
1. There will be no surrogate standard used in the extraction procedure. The stock, fortification, andcalibration standards will be preparedas follows:
Stock Solution
, , . ...
,,,
Prepare a stock solution of PFOS at 100 pgfmL by wetghing out 10.0 mg of
analytical standard(correctedfor percent salt and percent purity). Adjust final
volume to 100 mL with methanol in a 100-mL volumetric flask. Store this
stock solution (in 125-mLLDPE bottles) in a refrigerator at 2C to 6C for a
maximumperiodof6 months fromthe dateof preparation.
Fortification Solutions
,,, T .L
,T
a l n i.o/mt.'Fortification Solution - Pipette 1.0 mL of the 100 pg/mL stock
solution into a 100 mL volumetric flask. Bring up to volume with
methanol.
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b n i iiy/mT. Fortification Solution - Pipette 10.0 mL of the 1.0 pg/ml
fortification solution into a 100-mL volumetric flask and bring up to volumewith methanol.
c. nni no/mT. Friificfltion Solution - Pipette 10.0 mL of the 0.1 pg/mL
fortification solution into a 100-mL volumetric flask and bring up to volume with methanol
Store all fortification standard solutions (in 125-mL LDPE bottles) in a
refrigerator at 2C to 6C for a maximum period of 6 months from the dateof preparation.
Calibration Standards
. . . . . . ... . ,
Prepare six LC/MS/MS calibration standards m methanol via dilution of
the 0.1 pghnLandthe0.01 pg/mLfortification solutions.
This is a typical example; additional concentrations may be prepared as needed.
Initial Cone. (pe/mL)
0.1 0.1 0.1 0.01 0.01 0.01
Volume (mL) 5. 2. 1 5. 2 1
Diluted to (mLl 100 100 100 100 100 100
Final Cone. (pp/mL'i
0.005 0.002 0.001 0.0005 0.0002 0.0001
Store all calibration standard solutions (in 125-mL LDPE bottles) in a
refrigerator at 2C to 6C for a maximum period of 6 months from the date ofpreparation.
2. The liver samples will not be processed to make the homogenate described in the method Section 12.2 to 12.12. Due to limited sample available, samples will be mashedon a flat surface byhandpriorto extraction.
3. The sample weight for liver will be 0.50 g ( 0.05) and the sample volume for serum will be 100 pL The amount of 0.5 MTBA added to the serum sample
will be 0.5 mL and the amount of 0.25 M sodium carbonate/sodium bicarbonate buffer added will be 1.0 mL Samples will be placed on a wristaction shaker for - 20 min. at level 10 and then centrifuged for - 15 min @
-3000 rpm. The samples will onlybe filtered with 0.2 pm nylon mesh filter if necessary. Samples will be reconstituted in a final volume of. 1 mL of methanol for both matrices.
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4. The samples will be analyzed under the following conditions: LC/MS/MS System and Operating Conditions (Turbolonspray)
Mass Spec: P E S OE X API 3000 Biomolecular Mass Analyzer
Interface:. SCIEXTutbolon SprayLiquid Introduction Interface Harvardinfusion pump
Computer PowerMacintosh 03
Software: PE SciexAnalyst 1.1 Windows NT
HPLC:
Hewlett Packard (HP) Series 1100 HP Quat Pump HP Vacuum Degasser HP Autosampler HPColumn Oven
HPLCColumn:Genesis C* (Jones Chromatography), 2.1 mmx 50 mm, 4p. ColumnTemp.: 35 C Injection Voi.: 10 pL Mobile Phase (A): 2 mM AmmoniumAcetate in ASTM type I water Mobile Phase (B): Methanol How Rate: 0.3 mL/min.
Time %B
0 60 40
1.0 0 100
7.0 0 100
7.5 40 60
11.0 40
60
It may be necessary to adjust the HPLC gradient in order to optimize instrument performance. Columns with different dimensions (e.g., 2.1 nun x 30 mm) and columns from different manufacturers (Keystone Betasil C|* etc.) can be used, provided equivalent chromatography is obtained.
Ions monitored:
Analvte PFOS
Mode negative
Transition M9nilorgd 499 > 99
Approximate Retention Time (min'
4,20
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On a day-torday basis, the retention times may vary slightly depending on the batch of mobile phase, etc.
Example Tune File Parameters
The following values are provided as an example. Actual values may vary from instrument to instrument Also, these values maybe changedfromtimeto time in orderto optimize forgreatest sensitivity.
The mass spectrometer is tuned using a 0.5pg/mL PFOS solution, prepared via dilution of the stock solution in methanol. The solution is infused (using a "T"
connector) at 10 pL/min into a 0.2 mL/min stream of mobile phase consisting of 40% methanol and 60% 2 mM ammonium acetate. The analytes are initially tuned for the parent ion and then tuned for the product ion. Once the instrument is tuned, the optimized parameters are savedas a "tunefile". This tunefile is then used during routine analysis. The tuning procedure may be repeated as necessary to ensure optimal sensitivity.
Controls IS-Iospray
OR-Orifice RNG-Focus Ring
QO-Quad0 Rod Offset IQl-Interquadllens
ST-Stubbies ROl-Quadl Rod Offset IQ2-Interquad21ens
R02-Quad 2 rodoffset ST3-Stubbies
R03- Quad3 rodoffset DF-CEMDeflection Plate CEM-Channel Electron Multiplier
Set -4000.0 -61.0 -270.0
10.0 9.3 15.0 9.3 20.0 84.0 100.0 86.0 300.0
2800.0
Gas Rows
Nebulizer Gas Curtain Gas Collision Gas TIS Temperature
Set
12 13 4 350C
Calibration Procedures
a. Inject the same volume (between 10 to 20 pL) of each calibration standard(preparedin MeOH) into the LC/MS/MS.
b. Use linear, 1/x weighted standardcurves for quantification. Linear standard curves are generated for each set by linear regression using the appropriate software system. Any calibration standards
falling outside 30%, based on its calculated concentration, must
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be excludedfromthe calibrationcurve. However, the total number of calibration standards thatmaybe excludedmustnotexceed30% of the total numberof standardsinjected.
c. The correlation coefficient (r) for calibration curves generated mustbe 20.9925 (1^0.985). Ifcalibrationresults fall outside these limits, then appropriate steps should be taken to adjust instrument operation, and the relevantset of samples must be reanalyzed.
Sample Analysis Inject the same volume used for the calibration standards (between 10 to 20 pL> of each sample, fortification, control, etc. into the LC/MS/MS. Each sample must be analyzedin duplicate.
a. Standards corresponding to at least six concentrations must be included in ananalytical set.
b. Inject an entire set of standards (six) at the beginning of the run and inject standards interspersed about every 5-10 samples. All sample injections mustbe bracketedbystandardinjections.
c. Each set of samples analyzed (not to exceed 25) must include at least one reagent control (extraction solvents only), at least one matrix control, and two matrix control samples fortified at known concentrations andcarriedthroughthe procedureto verifyrecovery.
d. The concentration of each sample, fortification, control, etc. is determined from the standard curve based on the peak area of the analyte in all standards injected during a set The standardresponses must bracket responses of the residue found in the sample set If necessary, dilute the samples andre-analyze to give a response within the standard curve range.
e. Fortification recoveries within 80 to 120% are acceptable for fortifications at the LOQ level. Recoveries between 80 to 120% are acceptable for fortifications at levels greater than the LOQ. Failure to meet these criteria requires an investigation of cause and a full reanalysisofthe affected samples.
f Simples in which no peaks are detected at the corresponding analyte retention times will be reported as ND (not detected). Samples in which pr?Ware detected at the corresponding analyte retention times but areless than 3:1 signaknoise will be reported as NQ (not quantifiable).
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g. Background levels of analyte found in control/blank samples that correspond to values below the LOQ, but are still quantifiable, will be used to correct fortification recoveries.
h. If samples are not loaded on the instrument to be analyzed the day they are extracted, samples must be stored refrigerated at approximately 2C to 6C until analysis and analyzedpreferablywithin aweek.
9. EXPERIMENTALDESIGN
Samples obtained by Wildlife International Ltd. have been shipped to Centre Analytical Laboratories, Inc. for analysis.
All samples will be analyzed according to methods, "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Serum for Analysis Using HPLC-EIectrospray/Mass Spectrometry" and "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Liver for Analysis Using HPLC-EIectrospray/Mass Spectrometry" with the modifications detailed in Section 8.
Methods to control bias will include assay of untreated control samples, fortification of untreated control samples to obtain recovery data, and replicate analysis offortified samples to provide an indication ofreproducibility.
The average recovery and relative standard deviation of the fortified samples will be Also, the average residue found and standard deviation for each matrix will be calculated.
10. PROTOCOL AMENDMENTS AND DEVIATIONS
Any deviations from the protocol or from the analytical method as provided will be documented and repottedpromptly to the Study Director. Planned changes to this protocol or to the analytical method will be made in writing as an amendment and approved by the Principal Investigator and the Study Director. Any amendments or deviations will be appended to this protocol and included in the final report
1 Pm.rvni .mmdments: Planned changes to the approved protocol shall be documented by amendments that clearly describe the change, justification for the change, andimpacton the study. Amendments will be signedanddatedby the Principal Investigator and Study Director. Copies of amendments will be sent to the qualityassurance unit.
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2. Protocol deviations: Protocol deviations, which are one time and unplanned deviations fromthe protocol, shall be documented in the studyrecords, noting the nature of the deviation, potential effect or impact on the study, and corrective action if required. Protocol deviations are signed and dated by the Principal Investigator andStudyDirectorandreviewedby QAU.
11. RECORDS
i
Records to be maintained include, but are not limited the following (as appropriate): 1. Sample trackingsheet(s) 2. Sample receiptrecords, storage history, andchains of custody 3. History andpreparation of standards (stock, fortification, calibration) 4. Descriptionof anymodifications tothe method 5. Instrumentrunsheets, bench-sheets or logs 6. Analytical datatables 7. All chromatographic and instrumental conditions 8. Sampleextraction andanalysis dates 9. A comple t listing of studypersonnel, signatures andinitials 10. Chronological presentationof all studycorrespondence 11. Anyotherdatanecessaryforthereconstructionof the study
All chromatograms will containthefollowing:
a. Sample identification, date, arrow or other indication of the area of interest,andinjection numbercotresponding to the run.
b. Additionally, fortifications will include the fortification level of the analyte.
c. Analytical standard chromatograms will additionally include the concentration (e.g., pg/ml, ngftriL, ppb, ppt, etc.).
Each data set will contain information on temperatures, flow rales, column parameters, gases, instrument parameters, and instrument type, etc. The Centre study number will also be recorded on the fust chromatogram of each daily analytical run.
12. QUALITYASSURANCE
Centre QA Unit will review the protocol, audit the study conduct and the study documentation (including raw data and final report) to ascertain that all QA/GLP procedures are adhered to. Centre QA Unit will inspect the study at intervals adequate to assure compliance to GLP's, and will report the findings of the audits to thePrincipal Investigator, CentreManagement, andthe StudyDirector.
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13. DATA AND REPORT
Centre Protocol No. OOP-023-042
1. AHraw data and the original signed protocol will be maintained in the study file. This data includes the protocol amendments, protocol deviations, laboratory notebooks, analytical standard solution preparation, sample chain of custody sheets, sample work sheets, chromatograms, calibration, curves, and any other appropriate data generated. Raw data not used will be stored in thestudyfile. Thereason forexclusion will be documented.
2. A final report will be issued by Centre, approved and signed by the Study Director and sufficient for submission to EPA. The report contents should include, but not limitedto:
1. Objectives andprocedures stated inthe protocol 2. Analytical andstatistical methods used 3. Test materials identified by name, lot, purity, and othercharacteristics 4. Nameof testing facility andstudy initiation andcompletion dates 5. Statementprepared andsigned by the quality assurance unit 6.Tables containing all applicable data 7. AHchromatographic andinstrumental conditions 8. A complete listing of Centre studypersonnel
Centre will send a copy of the draft report to the Study Director, who will return the repot with comments. Centre Analytical will make revisions andfinalize the report with the approval of the Study Director.- The Centre QA unit will conduct an audit of the final draft report and data files to assure accuracy and GLP compliance. The final report must be approved by the Study Director prior'to signature. A statement of QAU inspections and a statement of GLP compliance will be included in the final report.
Any corrections or additions to the final report shall be in the form of an
amendment by the Principal Investigator. The amendment shall clearly identify
the part of the final report that is being corrected and the justification for correction. The amendment shall be signed and dated by the Principal Investigatorandthe StudyDirector.
REPORTDISTRIBUTION: Original: StudyDirector Copy: Centre Archives
14. ARCHIVE STATEMENT
Study records to be maintained: Records to be maintained for the study include all raw data, observations recorded during the conduct of the study, documentation, chromatograms, specimen tracking information, and study related correspondence.
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This includes a description of equipment used during the conduct of the study. All characterization data andany shipping records shall be retained. Document archives: Upon completion of the study, the study records, protocol and amendments, and the final report and amendments shall be retained in the 3M document archives. If it is necessary to substitute acopy for an original record, it will be certified as an exact copy. Centre will retain facility-related original and a copy of the final report andrawdatapackage.
15. REFERENCES 1 U.S. Environmental Protection Agency. Toxic Substances Control Act (TSCA). Good LaboratoryPractice Standards. Final Rule, 40 CFR Part 792.
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16. PROTOCOL APPROVAL This protocol was audited by the Quality Assurance Unit of Centre Analytical Laboratories, Inc. Quality Assurance Reviewer, Centre
Quality Assurance Auditor
----------------------
Principal Investigator, Centre
W '/
Facility Management, Centre
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APPENDIX; ANALYTICAL METHODS A. "Extraction of PotassiumPerfluorooctanesulfonate orOtherFluorochemical Compounds From Serum for Analysis Using HPLC-Electrospray/Mass Spectrometry" B. "Extraction of Potassium Perfluorooctanesulfonate or OtherFluorochemical Compounds FromLiver for Analysis Using HPLC-Electrospray/Mass Spectrometry"
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BEST COPY AVAILABLE
Centre Study No.: 023-042 CentreProtocol No. OOP-023-042
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.* JJ PFOSAA(3MSpecialtyChemicalDivision),molecularweight 5J5. ...fS-4 HtFOSE-OH(3MSpecialty ChemicalDivision), molecularweight 570
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s a iiS ^ P P ro ^ J M te 'sp ild iii amoonts for standard] a id spikes
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r M D L /L Q Q v ain es Tor r a b b it si
Compound MDL
~r-L in e a r C alib ratio n B an g e (L C R ) | " -- ----------- -
- (PP*))*-
A pproxim ate concentrations to be nsed fo r preparino th.
Standard C alibration C n m . - r E
c
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Ki ^ S o id
Prepared range LCR. from \o f standards l `. ciirve . (ppbjfng/m L)' \:-.(PPb) j .
v-j (a ffai.) `
% Recovery .r-.Range .'
.; ,RSD - Range .
..1;'\ -
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':i:'&-$k' 0ib)~(ngfalj' t (ng/mL) C
V - .,i r . } ; f { * .`L*.'*
MT/N'T- -r^!
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Rabbit Snni : 'r; ' fiy 7
: Preparedrange rffofstandards *
f^O ^ L V
LCR from % Recovery [' cunt ,/; U. Range,.-:!
(ppb) C; .(ogAriL) ?,
RSD .Range
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^ ,7 7 - n o J 0 s-' - - i .
S ' W i ^ 9 .7 i^ $ S $ 0 . 9 X 'V.1-4.W :
ffigbcmye5' S p?
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Compound: PKJSKAB& kK Preparedfange . VLCRfrom:
Rabbit Serum ; p c tiS & i j& .Vcrve
/(bWngnL)" *" ^ 1f'^ r* j ^OngftSL) S
^ $ & ~ 4
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'' ,?
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.Prepord range . ..LOtfromV Recovery. f ' f standards:'; `.V;!"curve 7;>^ YfRjinge'^m
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-'9 7 -io'^\/rVf; -....tV a J f i5.
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' .fK1:5.'.Nitrogen vpiator,Organomtion
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- " >
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JhfiO ^Q ^in te t" Store & a 1
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.^ ;;lfl Pat^ amalvsis and Calculations
.
'v -iL
" ' " if-:1 "
3*^4t.CinirmculatneithLaOymeiLagaehdqenostistyfolKftiblnepleiavmerBh.o*.m^og,-e-n*a-*te.b>y'reconimg..ea.c.h..mass of
V _ /.-iS^tiiAveraedoHitvfm e/m H^Averaee massima) of the aliarots '
:;; V;VT^3iF. 7j';stiarikdit^mf an^S^te^W^ l^o^irMfi%| ^*tqPiFloOnS:,>^^'irol'f;t/e?f-f;l.n-oyr.o'-iie-jm-. ic. alsin(libnition ' -
'! -i \u*-.
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^uLofStandardxConcentt^onfue/niLl^Kiial Concentration (jl/E o r^ - > c|.i . -A ' ^ . mg Liver/ rmLhmogetfite*.;`.,;-,.ofPFOSinLiver.
`
i '-\'^^:;i.';& .i w.vK**ie-wfe.r`Tto 1L 2 f-or -il*-,ev i
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3-
14.0 Methoo Pertormanc:
*
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B EST CO PY AVAILABLE CentreProtocol No. OOP-023-D42
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V .'V-'t>'
Centre Protocol No. OOP-023-042
r
LOQC o m p o u n d J M D L '
L inear C alibration Range (LCR)
"
-- ---------
(ppb) A p p rir io ia te co ncentrations to b e used fo r p rtp a ria the
S ta n d a r d C alib ra tio n C u rv e
,
*
. U ^ S * U t 4 S . v '26-9.- 3 0 p p b - : 1 2 0 0 p p b : . . - ''>
A \ , - i ; . pi 3_5Q l*. 11.1 12 ppb -.1 2 0 0 p jS -TM .- '
>J r i O i A A S " ' 1^ 24.6 / V O . 3 0 p p b li 1200ppb
v.
~ -- ---------. :-- -- ------------
. ---------------------
c i r u S i i i J H JV< 108 V 3 4 5 60 p p b V 900 p p b * :.
; ----------
M ^ o - 5 a y : J a ; S 2 J . - . .2 6 2 , 6 0 p p b - 1 2 0 0 ppb . .. < r f u s h i V < i t ( i ; : j 3 : 9 : c u m ; 3 0 p p b - 1200 p p b .' . :. . ;
-- --- .
~r.-- ;-- --
e V V -.--o r-u .
. ----------
Refer Eni n n
-A \rr^ ^ ..j.;
9 n .1/1U 1,-1.
r '
C `-
1.-^Prepared - Range of. .*.^C R rfin, Ringe o f^ i jjL ^ 'ilro ^
v75- Liver,*', ;'-`rM gc'of.. . avenge
ivcqpijcfl ]/ffiowstd ;
/matrix^ 4 stand*; pb) (aiw j (ppb) (staL) - .t.l'v- .J.
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C'.&cSZm
;M "i . ; c a m *
<**) ( a r tii
. s s b m
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6.19-1237
^T -iiob -- /.-A iL t^ 5 W ? S 0
, 60- | 3M. l ^ i S o S
C o m p o a o d : P F S A -*. \"..*.rv v {j Vi*'.
Liver ' matrix '
Preparai range of ;
j standards (ppb) (itali
6Range of
;*avenge';*
UrLCLJpoeT .. Range o f- ^ ^ tg R - fig te i i towsld
, :^ t = o i . g L C .
w t.:
^'ebrve *
(ppb) (estai) 3ppb)rfdf< (ppb) (ctaL) <(ibt?n7rnul
Rabbit *. .'1 9 r 37. ' 12,1200
'b 000
H Prepared :11:Range of : rtXCKJhioi.V,.| JRasgeei ^^LCKdfiTCri'
: liver* ;nutr*v
'^rangeir; standards**
..avenge -^aw CuryeAj
^ 'crve (peli)(BttaU .r(w>b)(-V
'j;,"T7J ' amre u '
.... 'tiViza >i2fj^..ryo>3M
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- .-i
'
1 I*.
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'A
O
Ion Pair Standard Carves - Tissue
^riijgafric*:
Standard number: Equipment number . final solvent and TO: Blank Uver/identifier:
1 7:
F C m i i t f d a ^ r o t -. - . - . ... ........
SuTogatestdapproz^OOjspm: v r..-'*> V : : V
X
Aetna! nceatratioiu of standards a the FC mix - -* ~
t '"v!-.":-i * -X-rX:.
`
p r o s / PFOSA.
tFOSS PFOSEA. . M556 .
; au -
AJJ -
Stdcccc Sidcone; ^Sticaae? *SuJ cone Stdcon. Std cobfi - Stdcoae ` Ami jpked Density
nj/mL: ntaiL-'
ugfSN ngAnL- : vi/mL ` ttg/xnL mL` * ' ' `8
0300- 0300 :0 3 0 0 -' 0 3 0 0 1 . 0300 0300
: 0.002 . 0.167
0300 : 0300 - 0300 . *0300/ 0300 0300
0304 - 0.167
0300 0300 ; . 0300 , '0300^- .0300 0300
- 0300c - 0300 v ;-0300.. . 0300 'J V0300 . , . 0300 : oioo - 0300 -.. 0300 0300Hi 0 3 0 0 '. 5.0300
0.010 - 0.167
. 0.020 " 0.167
` 0.040
0.167
r 5.00 T '.: '5.00 > --V5.0O * 590 -: 'w 5:00,S ''5.00
0.0t0 -. 0.167
J.00.' -100 .:. 5.00.-: 5.00 - s s f > y - 5.00
' 0.020
0.167
3.00 * 5.00 : ' .5.00 . . 'T.OO-- . -5.00. --5.003k v. . 0.030 0.167 '
S O J O i: *; 50.0 . ;:*50.0 : 50.0 Vr:50.0\: '- J O .O 'A ' u*.' - '
0.004 .4-.. .- 0.167
C alculated concentrations of standards in the samp I e j n a l m ^ \ * - -
V -/::'.:-
PFOS * .PFQSA PFOSAA EtFQSE PFOSEA V.MSKJ
; Fimi _ !*Hnl
Final cone
, Tmali v'Raal./
;':FmaY m
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r t- - ng/j '
sis - n ^ t " B g /t':-.
m
5.99. 599 - 599 12.0 - 12.0 . . ito
599 . 599 ' : 599 i 12.0 12-0 . 12.0 v -*.W
10?. tv Z 0.005
293 ... 29.9 - v 299 . 299 299 v 2 9 3 . - " - ` A /S an y p U ik A ' & r t + y
599 .
- 599 . <-! 599 . , *593 - v v :
.PBJ'dioc s
: 120 ! 120..: :;.I20
t-120 .-120 .H T. 120 >
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299 . 299 : .. 299 -299 * '299 v, ' 2 9 9 - - -
3v"4^''
5 9 9 .- -599 i 599 - 599 " 599 - 599
V*-" '`.v
89a i-" 198 Wt : .. 898 - * 898v- v* 898 ;*
119.. - 1198 .1 1 9 1
119S r. .1198 - 1198 ^
' S A ` i y f ^ ^ S b^ V b'& flr
't
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PROTOCOL AMENDMENT
Page 1of 1
Centre Study Number. 023-042
Amendment Number 1 Effective Date: 11/21/00
Centre Protocol Number. 00P-023-042
DESCRIPTION QF AMENDED SECTION I. 6: Justification for the Selection of the Test System
AMENDED TO
1. Add the following:
A complete description of the test system along with all of thein-life parameters is
documented in the sponsor's protocol no, 454/010600/MP/SUB454, Project No. 454-
105 that is associated with this analytical protocol.
________
RATIONALE
l. At therequestof the sponsor, adescription of the in-life parameters forthe test system was not to beincluded in this protocol that covers the analytical phase for the analysis of mallardliver andserum.
IMPACT ON THE STUDY
1. No negative impact on the study.
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to-
Date I lA '/ t / o r )
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0 9 / 0 6 / 0 1 _ 1 6 : 0 9 FA_T 4 1 0 J 2 2 0 6 3 2
WILDLIFE INTERNATIONAL
SEP. 6.2031 3*2901 DOTCEN HESRRCH
@003
NO,5 9 5
P .3
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PROTOCOL AMENDMENT
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Centre Study Number 023-042
AmendmentNumber. 2 EffectiveDate: 11/29/00
Centre Protocol Number: DOP-Q23-042
DESCRIPTIONOPAijiagma dECDON 1. 9; ExperimentalDesign
AMENDEDTO
1. Addthefollowing:
Moisture detenmnaticawill beperfotmodonalllivertangles accordingtoAOAC OfficialMethod950.46SectionB<(a), withdiefollowingmodifications: . 1. Thesampleweightwill be - 0.5 g. 2. Thedryingtin will be - 12-14boon.
RATIONALE
TM-- "
At therequestofthesponsor,moisturedctcnninaHbnawill be donsfareveryliver
sanqde. Due tolimitedsamplesize, thesample'weightusedwill be -0.5 insteadof
-10 g. Thedryingrimerequiredforsuchasmall samplesize docs nothavetobe 16-18
bouts,so thesamples willbe driedfor-12-14 houts
IMPACT ON THE STUDY
1, Nonegativeimpactontis study.
igatorSignature
Date
SponsorRepresentative (StudyDdector))St ignatare
QU i Date
CentreQAUReview ffr? */t/>'a/ ________ ____ r FefhimJ*vrM-iv 12.1988/1
KOT&-. TV AMdrihHAKT W45 L-TuLD H ^/Cefsi T>Wt oilA JM l. w*5 COST
fYvrCe Iptte
^ljo\
r e c e iv e d T ire: s e p . 6 .
4 :0 6 pm
Centre Analytical Laboratories, Inc.
3
PRINT T ift SEP. 6. 4:08Pn
Page 101 of 104
Centre Study No.: 023-042
. 0 9 / 0 . 8 / 0 1 _ 1 8 : 0 3 FAT 4 1 0 g 2 2 0 8 3 2
1 WILDLIFE INTERNATIONAL
SEP, S .2 0 0 1 3 : 29PM
EXYOEN RESEARCH
@004 NO.595 P.4
, M.a,___fto
Centre Analytical Laboratories, Inc.
Rytote (turnu p
PROTOCOL AMENDMENT ; Amenrtmrnl-NmnbeK 2
Ef&ggveEte: 03/22
PagI of 1
CerneStndyNumber:023-042
CentraProtocol Number: 00P-Q23-frt2
D B S g U P T T O If O F A M E N D ]
1. $8: SamplaAnaljr(f) 2- g 9: Brpegnwntal Petipa 3. } S: LC/M SM S System m d Opeating Owdte'oat(TnrbcJbajjray)
AMPN-nm-np
""
1. Change Uselast salience to read the Mowing;
Sanle a whichpeek ins dejected atihe em aponang nalyte iBtEntan dines batate less than the loweat concentration ofthe eaEhrattan ittnditrd* (0.0001 pg/mL)win be reponed as
NQ (not quantifiable). 2. Add the following:
The following eqnadcnj willbe usedforqnanritatinn of the samples:
1-Analyte FbrindCngAnL)=,(peakarea-intercept)/slope
XAiulyte Found (ps/s) a fTP1-0*fn"" Hft"*3 IP W i) ^ Iff 8TT1' fcnLl * TQDOno AVCinL)*ample w tf c jx ip g
WhereFVa final volume, DF= ditaian factor, EV= extraction voten*-, AV = aliquot v o te *
3 -R c c o v e ry (9 6 ) = /a n a l, f a r n d lg t a U r - i r e . m i t r e ia a r i fu e /m L lt t Ip o n fl, T tn n g .
emonnt added(BgtmL)x 1 pg 3.Chinge the campearhoed to Dell tOmSan PU10
SAHtWALB
---- -- --
1-3. Clarify bowthe samplesare qosatitared andreportedend the totrmnans usai to acquire
dat.
IM P A C T O K
1-3. No negati impactan the ffldy-
ipal te W rig a to r Signature
__ P'
Stndy Director Signature
'
Date
Dam CentreQAUB*Sew
ix 199S/l
rel' M '& w bfr JtJ i'X iu o 0 ^ J o i Xufc otZ&KAL WA8 U r 5 6 6 flftjrtt (va OA-TTiis jlr.I* fjilU M
RECEIVED TIME SEP. 6. 4:06PM
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Centre Study No.: 023-042
3048 Research Drive, State College, PA 16801.
Phone: (814)231-8032, Facsimile: (814)231-1253
C entre S tudy N um ber: 023-042
PRO TO CO L AMENDMENT Amendm ent N um ber 4
D ate o f O ccurrence: 08/24/01 '
Page 1of 1
C entre P rotocol N u m b e r OQP-023-042
1. 3 : S P O N S O R
D ESCR IPTIO N O F A M EN D ED SECTIO N
AMENDED TO
1. Change the study director from Rochelle Robideau at 3M Environmental Technology and Safety Services to Sean Gallagher at Wildlife International, LTD, 8598 Commerce Drive, Easton, MD 21601, phone: 410-822-8600, fax: 410-822-0632.
2. Change Rochelle Robideau to the Sponsor Representative.
R A TIO N A LE
-2 . B y re q u e st o f R o ch e lle R o b id e au (fa x d a te d 08/24/01 in c lu d e d in c o rre sp o n d en c e s e c tio n o f the raw d ata associated w ith this study)
IM PA C T O N TH E STU D Y 1-2. N o n e g ativ e im p a c t on th e study.
tZ M
S in g F acility M anagem ent Signature
W -, Incom ing S tudy D irector Signature
D ate
' L
D ate
C entreQ A U R eview
L 6 1
30 0 1
February 12.1998/1
Centre Analytical Laboratories, Inc.
Page 103 of 104
Centre Study No.: 023-042
Ms o e / O l -- 15x08 Fa i 410 822 06J2
WILDLIFE international
SEP. 6 .2 0 0 1 3:29PM D frtZ H RESEARCH
@002 NO.5 35 P . 2
Centre Analytical Laboratories. Inc.
3048 H e " * Tvh . Bate C a l l t r x . PA
PROTOCOL DEVIATION
D eriiiim N um ber I
D a te o f O c e a n s * : (1)imS/00(2 ) 11/19/00, (?)1 1 /2 3 /0 0 (4 ) W29-3OV0(S)3 /lQ d lI
C a m e Study N um ber 023-042
C en W fto to c o l N u m b e r OOP-Q23-042
d e sc r ip t io n o f d e v ia t io n
A^eptedL { 8 S e m p le A s s a y s * (e ) P a g e 9 :
ib c o v o m S i 7 9 *
Spk A a u d its duplicate iyoctinn.ni S et 1U700AD.
fo r Centra lam pto 0003804
i s * C iin..
f ____ l i m
P a a es 8-9: Included a {laudani in A s caEfcrstion carve fla t fell 11I700A D (C091M O-5).
3 . 4 8 ^ H - b r Hn n P ro ce d u re s (b ) P l g w 8 -9 : B tc iu d e d a
to r n th e fib ratio n curve that feU
v rirk ^ fc o fh s
Set1 X 2 1 0 (C 091800-3).
4. 5 g CaEbrsikin P to ced aea (b) Pages 8-9: B d a d id ilO rn d irt ta {to calitaticin cn rv o that fell
oarideiSO ^ itsc a lc o li oeocoiwaai SK 112700D (C091800-6),
5.{ 8 S a m p le A n a ly sis ( ) P i g e 9* A c c e p te d rtW V erfM o f 7 9 % s a d 7 7 * f e r C e n t a m p l e 0 0 0 3 8 0 3
S pk A A d il d u p licate m je& m io Sot O307O1A.
&&L ib lei
t-f. ftntocal deviation
ACTIONS TAKEN
.
la - m miment IcnnxL SOImvidTM. **
ScccstcdB y/D ;
IM PACI ON THB STUDY
...
14. NonegaflveiinpaaQa n ^ y bKKniitteovraDiTCi^rfchetpiiEiEcovcricjfcrrfv. jmriv
&H w ithin dm eccepoblo range o f 80S>-I20%.
'
2.4. N a n e g ativ e in a c t on the study b ecam e tb curva w a s etili w ithin accep tab le E tx a r
wpAsmente.
3. No negative impact on tiestudy becatratae d e s u n ta of standardscadmiai faro tb curve
w as 3ess th a n 2 0% o f tba t r a i num ber o f standards injected.
'csTigutar Signature
jltj l
D ate
. P' J^giIW JfV
etudyDirector S itu an e
f
9 Wof
D ate
k. Cefc. &u>fulfil
<lX lo*LL,
Due
CAL QAU R eview .
-fy*4 sA'ol Mmurr12.19M7
feT& T t i
-agufcticN 4(0(d1
OSJ&VAL MrS U&T eb
fHi?w2r cftS
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G005S4
