Document b52X5XEXqNQx5V8DV3X7LGwV3
AR226-1751
STUDY TITLE Extraction of Potassium Perfluorooctanesulfonate from Quail Serum and Quail 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 Phone: 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-023-041 Centre Study Number: 023-041
Total Pages: 102
Centre Study No.: 023*041
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT
Centre Study Number 023-041, entitled "Extraction of Potassium Perfluorooctatiesulionate from Quail Serum and Quail Liver for Analysis Using HFLCElectrospray/Mass Spectrometry," conducted for 3M Environmental Technology and Safety Service, was performed in compliance with US EPA TSCA Good Laboratory Practice Standards (40 CFR Part 792) by Centre Analytical Laboratories, Inc.
Emily HjDecker Principal Investigator Centre Analytical Laboratories, Inc.
Date
Study Director
W ildlife International, Ltd.
Date
R ogjjH kcB A iideau^N . `^fopror Rcpresentativer
3M Environmental Technology and Safety Services
Centre Analytical Laboratories. Inc.
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Centre Study No.: 023-041
QUALITY ASSURANCE STATEMENT
Centre Analytical Laboratories' Quality Assurance Unit reviewed Centre Study Number 023-041, entitled, "Extraction of Potassium Perfluorooctanesulfonate from Quail Serum and Quail Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry". All phases were reviewed 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
Protocol Review
Extraction
Raw Data & Summary Table Review
Raw Data and Draft Report Review
Final Report Review
Date Inspected
Date Reported to Date Reported to
Centre
Study Director and
Management Sponsor Management
12/19/00
12/21/00
12/28/00
12/19/00
12/21/00
12/28/00
2/28,3/1/01
3/19/01
3/23/01
3/23-26/01
4/9/01
5/7/01
9/6/01
9/6/01
9/6/01
Naomi Lovallo Sr. Quality Assurance Auditor
Date
Centre Analytical Laboratories, Inc.
Page 3 of 102
Cenere Study No.: U23-W1
CERTIFICATION OF AUTHENTICITY
This report, for Centre Study Number 023-041, ia 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:
iHLjkiJuc
oilyR. Decker
Scientist Centre Analytical Laboratories, Inc.
Centre Analytical Laboratories, Inc. Facility Management:
Centre Analytical Laboratories, Inc.
StudyDirector, Wildlife International:
ibLJ- .dX L
Sean Gallagher
l
Wildlife International, Ltd.
Sponsor Representative, 3M:
Mio'ot
Date
Centre Analytical Laboratories, Inc.
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Centre Study No.: 023-041
STUDY IDENTIFICATION
Extraction of Potassium Perfluorooctanesulfonate from Quail Serum and Quail Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry
CENTRE PROTOCOL NUMBER: 00P-023-041
CENTRE STUDY NUMBER:
023-041
TYPE OF STUDY:
Analytical
SAMPLE MATRIX:
Quail Serum and Quail Liver
TEST SUBSTANCE:
Perfluorooctanesulfonate (PFOS)
SPONSOR:
3M Environmental Technology and Safety Services Building 2-3E-09 PO Box 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 T echnology 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:
12/08/00 12/18/00 03/08/01 09/06/01
Centre Analytical Laboratories, Inc.
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Centre Study No.: 023-041
PROJECT PERSONNEL
The Study Director for this project was Sean Gallagher at Wildlife International, Ltd. The following personnel from Centre Analytical Laboratories, Inc., were associated with various phases of the study:
Name Emily Stauffer Dave Bell Melissa Kennedy Whitsel John Flaherty Tiffany Proctor Angela Morgan Michelle Arjmand Lawrence Ord Rickey Keller
Shawn Robb
Karen Risha
Title Scientist Scientist Supervisor Operations Manager Technician Technician Technician Sample Custodian Sample Custodian
Sam ple Custodian
Scientist
Centre Analytical Laboratories, Inc.
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Centre Study No.: 023-041
TABLE OF CONTENTS
Page
TITLE PAGE...................................................................................................................... 1 GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT............................. 2
QUALITY ASSURANCE STATEMENT....................................................
3
CERTIFICATION OF AUTHENTICITY.......................................................................... 4 STUDY IDENTIFICATION............................................................................................... 5 PROJECT PERSONNEL.................................................................................................... 6
TABLE OF CONTENTS.................................................................................................... 7
LIST OF TABLES.............................................................................................................. 8
LIST OF FIGURES............................................................................................................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 of Standards and Fortification Solutions...............................................14
6.3 Chromatography.......................................................................................................15
6.4 Instrument Sensitivity............................................................................
15
6.5 Description of Instrument and Operating Conditions............................
15
6.6Quantitation 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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LIST OF TABLES
Table I. Summary of PFOS
Page in Reagent Blanks..................................................21
Table II. Summary of PFOS in Quail Liver Blanks............................................22
Table III. Summary of PFOS in Quail Serum Blanks..........................................22
Table IV. Summary of PFOS Recoveries in Quail Liver......................................23
Table V. Summary of PFOS Recoveries in Quail Serum....................................24
Table VI. Summary of PFOS in Adult Liver Samples at 0 ppm a.i. (Test Termination 19 Weeks).................................................................................25
Table VII. Summary of PFOS in Adult Liver Samples at 1.8ppm a.i. (Test Termination 6Weeks)................................................................................... 26
Table VIII. Summary of PFOS in Adult Liver Samples at 6.2 ppm a.i. (Test Termination 6 Weeks)................................................................................... 27
Table IX. Summary of PFOS in Adult Liver Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)................................................................................. 28
Table X. Summary of PFOS in Juvenile Liver Samples at 0 ppm a.i. (Test Termination 12 Weeks)................................................................................. 29
Table XI. Summary of PFOS in Juvenile Liver Samples at 1.8ppm a.i. (Test Termination 12 Weeks)...............................................................
30
Table XII. Summary of PFOS in Juvenile Liver Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)...............................................................
31
Table XIII. Summary of PFOS in Juvenile Liver Samples at 17.6 ppm a.i. (Test Termination 12 Weeks)................................................................................. 32
Table XIV. Summary of PFOS in Adult Serum Samples at 0 ppm a.i. (Test Termination 19 Weeks)................................................................................. 33
Table XV. Summary of PFOS in Adult Serum Samples at 1.8 ppm a.i. (Test Termination 6 Weeks)................................................................................... 34
Table XVI. Summary of PFOS in Adult Serum Samples at 6.2 ppm a.i. (Test Termination 6Weeks)................................................................................... 35
Table XVII. Summary of PFOS in Adult Serum Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)................................................................................. 36
Table XVIII. Summary of PFOS in Juvenile Serum Samples at 0 ppm a.i. (Test Termination 12 Weeks)................................................................................. 37
Table XIX. Summary of PFOS in Juvenile Serum Samples at 1.8 ppm a.i. (Test Termination 12 Weeks)................................................................................. 38
Table XX. Summary of PFOS in Juvenile Serum Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)................................................................................. 39
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LIST OF TABLES (continued) Page
Table XXI. Summary of PFOS in Juvenile Serum Samples at 17.6 ppm a.i. (Test Termination 12 Weeks).................................................................................40
Table XXII. Summary of Percent Moistures for Quail Liver Samples............................ 41
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LIST OF FIGURES Page
Figure 1. Typical Calibration Curve for PFOS.............................................................. 43
Figure 2. Chromatogram Representing a 0.1 ng/mL standard for PFOS....................... 44
Figure 3. Chromatogram Representing a 1.0 ng/mL standard for PFOS....................... 45
Figure 4. Chromatogram Representing a Reagent Blank for PFOS (Centre ID Reagent Blank A, Set: 122000A )..................................................................46
Figure 5. Chromatogram Representing Control Quail Serum for PFOS (Centre ID: 0011424 Blank A, Set: 122000A ).................................................................47
Figure 6. Chromatogram Representing Control Quail Liver for PFOS, (Centre ID: 0007870 Blank A, Set: 121900A)................................................................. 48
Figure 7. Chromatogram Representing Control Quail Serum Fortified with 500 ng/mL of PFOS (Centre ID: 0011424 Spk A, Set: 122000A)....................... 49
Figure 8. Chromatogram Representing Control Quail Liver Fortified at 10 ppb with PFOS (Centre ID: 0011423 Spk A, Set: 121800A)............................... 50
Figure 9. Chromatogram Representing Juvenile Quail Serum Sample (Centre ID: 0007965, Sponsor ID: 9544, Set: 122000A).................................................. 51
Figure 10. Chromatogram Representing Juvenile Quail Liver Sample (Centre ID: 0007926, Sponsor ID: 9544, Set: 121800B).................................................. 52
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LIST OF APPENDICES
Page
Appendix A Study Protocol 00P-023-041 (Centre Study No. 023-041) and Amendments and Deviations.................................................................... 53
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1.0 SUM M ARY
Centre Analytical Laboratories, Inc. (Centre) extracted quail liver samples and quail serum samples for the determination of perfluorooctanesulfonate (PFOS) according to protocol 00P-023-041 (Appendix A).
The limit of quantation for quail liver was 10 ppb and for quail serum 10 ng/mL. These were determined in the method verification study (Centre study 023-054). PFOS in the quail 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 8.37 pg/g. These results are reported on a wet weight basis. Residues of PFOS in the quail serum samples ranged from 0.00212 pg/mL levels to 263 pg/mL.
The average percent recovery standard deviation for PFOS in serum samples was 108% + 10%. The average percent recovery standard deviation for PFOS in liver samples was 95% 11%.
2.0 OBJECTIVE
The objective of this study was to determine levels of perfluorooctanesulfonate (PFOS) in specimens of quail liver and quail serum using the analytical methods described in protocol 00P-023-041.
3.0 INTRODUCTION
This report details the results of the analysis for the determination of PFOS in quail serum and quail liver, using the analytical methods entitled, "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Serum for Analysis Using HPLC-Electrospray/Mass Spectrometry" and "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry" with the modifications listed in Section 6.
The study was initiated on December 8, 2000, when the study director signed Centre protocol number 00P-023-041. The experimental start date was December 18, 2000, and the experimental termination date was March 8,2001.
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4.0 TEST SYSTEM
The control quail serum and liver used for the matrix blanks and matrix fortifications was received on November 28, 2000 from Wildlife International Ltd., Easton, MD.
The control quail liver was assigned the Centre ID of 0011423 and the control quail serum was assigned the Centre ID of 0011424. Fifty-six quail serum samples and sixty quail liver 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 quail serum samples and twenty quail liver samples were received on February 23, 2001 and stored frozen until they were logged in by Centre personnel on February 26,2001.
Sample login 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 M ATERIAL
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. Stability was not assessed in this study but the reference material was stored according to the certificate of analysis and was used prior to expiration. The solubility of the reference material was not relevant to this study.
Compound
PFOS
Centre Control No. TCR No. Purity (%) Expiration Date
00-023-042
TCR 00017-46
97.9
08/31/01
The molecular structure of PFOS is given below.
PFOS Chemical Name Molecular weight
O
= =
Perfluorooctanesulfonate 499 (C8F17SO3-)
C8F17S -- O'
o Note: rrhe 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 of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Serum for Analysis Using HPLC-Electrospray/Mass Spectrometry" and "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical Compounds From Liver for Analysis Using HPLC-Electrospray/Mass Spectrometry" with the modifications listed below were used for this study.
6.1 Extraction Procedure
A 100 pL aliquot of the serum and ~ 0.50 g sample of liver was used for the extraction procedure. After fortification of appropriate samples, the samples were vortexed for ~ 15 seconds. One mL of 0.5 M TBA was added to the liver samples and 0.5 mL was added to the serum samples. Two mL of 0.25 M sodium carbonate/sodium bicarbonate was added to the liver samples and 1 mL was added to the serum samples. Five mL of MTBE was added to both the serum and liver samples. Each sample was placed on wrist-action shaker for ~ 20 min. and then centrifuged for ~ 15 min. Four mL of the organic layer was taken and dried on an N-Evap and then reconstituted with 1 mL of 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 00P-023-041. An individual stock standard solution of PFOS was prepared at a concentration of 100 pg/mL by dissolving 10 mg of 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 of the stock and bringing the volume up to 100 mL with methanol.
The 0.1 pg/m L fortification standard w as prepared by taking 10 m L o f the 1.0 pg/m L
fortification standard and bringing the volume up to 100 mL with methanol. A 0.01 pg/mL standard was prepared by taking 10 mL of the 0.1 pg/mL standard and bringing to 100 mL with methanol.
A set of standards containing PFOS was prepared by dilution of 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
Centre Analytical Laboratories, Inc.
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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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 analysis using electrospray LC/MS/MS. The retention time of PFOS was ~ 4.4 min. Peaks were detected in the control matrices corresponding to the analyte retention time and some of the amounts detected were significant enough to alter the results reported in this study.
6.4 Instrument Sensitivity
The smallest standard amount injected during the chromatographic run was equivalent to 0.0001 pg/mL of PFOS.
6.5 Description of Instrument and Operating Conditions Mass Spec: PE SCIEX API 3000 Biomolecular Mass Analyzer
Interface:
SCIEX Turboion Spray Liquid Introduction Interface Harvard infusion pump
Computer: Dell UltraScan PI 110
Software:
HPLC:
PE SciexAnalyst 1.1 Windows NT H ew lett Packard (H P) Series 1100
HP Quat Pump HP Vacuum Degasser HP Autosampler HP Column Oven
HPLC ColummGenesis 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 60 40
11.0 60
40
Ions monitored:
Analyte PFOS
Mode negative
Transition Monitored 499 -> 99
Approximate Retention Time (min)
4.40
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 of sample or calibration standard was injected into the LC/MS/MS. The peak area was measured and the standard curve was generated (using 1/x fit weighted linear regression) by Analyst software using six concentrations of standards. The concentration for quail serum and quail liver was determined from the following equations:
Use Equation 1 to calculate 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 use Equation 2 to calculate the amount of analyte found in pg/g (pg/mL for serum samples).
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Equation 1:
Analyte found (ng/mL) = (Peak area - intercept)
slope
Equation 2:
Analyte found (pg/g) = (anal, found (ng/mLt x FV (mL) x DF x EV (mL)) x 1 u.g
(AV (mL) x sample wt. (g))
1000 ng
Where:
FV = Final Volume
DF = Dilution Factor
EV = Extraction Volume
AV = Aliquot Volume
For samples fortified with known amounts of PFOS prior to extraction, use Equation 3 to calculate the percent recovery. Equation 3: Recovery (%) =
(anal, found (ug/g) - avg. analyte in Ctrl (u g/gi) x 1000 ng x 100% amount added (ng/g) x 1 pg
An example of a calculation using an actual sample follows:
Quail liver sample Centre ID 0007870 Spk A (Set: 121900A), fortified at 1000 ng/g with
PFOS.
Where:
peak area
11483
intercept
463
slope
2790
dilution factor
100
ng/g added (fort level) avg. amt in controls final volume
1000 ng/g 0 (Not quantifiable) 1 mL
extraction volume
5 mL
aliquot volume
4 mL
sample weight
0.5 g
From equation 1: Analyte found (ng/mL)
= 141483- 4631 2790
From equation 2: Analyte found (pg/g) =
3.9 ng/mL
(3.9 ng/mL x 1 mL x 100 x 5 mL) x 1 ug
(4 mL x 0.5 g)
1000 ng
= 0.975 pg/mL
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From equation 3: % Recovery = (0.975 iie/mL - 0 ug/mLf x (1000 ng/1 uef x 100% 1000 ng/g
= 98%
7.0 EXPERIM ENTAL 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 quail liver blanks and quail 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% 11%. Individual recoveries for PFOS in the serum samples are detailed in Table V. The average percent recovery + standard deviation for PFOS in serum samples was 108% 10% .
PFOS in the quail 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 8.37 pg/g. Individual results are listed in Tables VI-XIII. The results are reported on a wet weight basis.
PFOS in the quail serum samples ranged from 0.00212 pg/mL levels to 263 pg/mL. Individual results are listed in Tables XIV-XXI.
The average percent moistures for the quail liver samples are given in Table XXII.
9.0 CONCLUSIONS
The quail liver and serum samples were successfully extracted and analyzed according to protocol 00P-023-041.
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10.0 RETENTION OF DATA AND SAM PLES
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 of temperature logs will be submitted. Exact copies of all raw data, as well as a signed copy of the final analytical report and all original facility-specific raw data, will be retained in the Centre Analytical Laboratories, Inc. archives for the period of time specified in 40 CFR Part 792. Retained samples of reference substances are archived by the sponsor.
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TABLES
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Table I. Summary of PFOS in Reagent Blanks
Sponsor ID
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*
* Duplicate injection
Set Number
121800A 121800A 121800B 121800B 121900A 121900A 121900B 121900B 122000A 122000A 122000B 122000B 030501A 030501A 030601A 030601A
Extraction Analysis
Analyte
Date
Date Found (pg/mL)
12/18/00
12/19/00
NQ
12/18/00 12/18/00
12/19/00 12/20/00
NQ NQ
12/18/00 12/19/00 12/19/00
12/20/00 12/21/00 12/21/00
NQ NQ NQ
12/19/00 12/21-22/00
NQ
12/19/00 12/21-22/00
NQ
12/20/00 12/20/00
12/22/00 12/22/00
0.000200 0.000200
12/20/00 12/20/00 3/5/01 3/5/01 3/6/01
12/22-23/00 12/22-23/00
3/6-7/01 3/6-7/01 3/7/01
NQ NQ ND ND NQ
3/6/01
3/7/01
NQ
AVERAGE:
NQ
STANDARD DEVIATION:
NQ
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)) ND = Not Detected
For values recorded as ND,zero was used to calculate the average and standard deviation. For values recorded as NQ, 0.00005 pg/mL (half the value of the lowest calibration
standard, 0.0001 pg/m L) w as 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 Quail Liver Blanks
Sponsor ID
na na na Na 454 454 456 456
Centre ID
0011423 Blank A 0011423 Blank A* 0011423 Blank A 0011423 Blank A* 0007870 Blank A 0007870 Blank A* 0007872 Blank A 0007872 Blank A*
Set Number
121800A 121800A 121800B 121800B 121900A 121900A 030601A 030601A
Extraction Analysis
Date
Date
12/18/00
12/19/00
12/18/00
12/19/00
12/18/00
12/20/00
12/18/00
12/20/00
12/19/00
12/21/00
12/19/00
12/21/00
3/6/01
3/7/01
3/6/01
3/7/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
NQ NQ 0.00113 0.00109 NQ NQ 0.00120 0.00110 0.000590 0.000578
Table III. Sum m ary o f PFOS in Quail Serum Blanks
Sponsor ID
na na na na na na na na
Centre ID
0011424 Blank A 0011424 Blank A* 0011424 Blank A 0011424 Blank A* 0011424 Blank A 0011424 Blank A* 0011424 Blank A 0011424 Blank A*
Set Number
121900B 121900B 122000A 122000A 122000B 122000B 030501A 030501A
Extraction Analysis
Analyte
Date
Date Found (pg/mL)
12/19/00 12/21-22/00
12/19/00 12/21-22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00 12/22-23/00
12/20/00 12/22-23/00
3/5/01
3/6-7/01
3/5/01
3/6-7/01
AVERAGE:
STANDARD DEVIATION:
0.0136 0.0144 0.0138 0.0151 0.0127 0.0109 0.00555 0.00559 0.0115 0.00384
* 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 of 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 IV. Summary of PFOS Recoveries in Quail Liver
Sponsor ID
na na na na na na na na 454 454 454 454 456 456 456 456
Centre ID
0011423 Spk A 0011423 Spk A* 0011423 SpkB 0011423 SpkB* 0011423 Spk A 0011423 Spk A* 0011423 SpkB 0011423 SpkB* 0007870 Spk A 0007870 Spk A* 0007870 SpkB 0007870 Spk B* 0007872 Spk A 0007872 Spk A* 0007872 Spk B 0007872 Spk B*
Set Number
121800A 121800A 121800A 121800A 121800B 121800B 121900A 121900A 121900A 121900A 121900A 121900A 030601A 030601A 030601A 030601A
Extraction Analysis
Amt
Date
Date
Added (ng/g)
12/18/00 12/19/00
10
12/18/00 12/19/00
10
12/18/00 12/18/00
12/19/00 12/19/00
2000 2000
12/18/00 12/18/00
12/20/00 12/20/00
1000 1000
12/18/00 12/20-21/00 12/18/00 12/20-21/00
20000 20000
12/19/00 12/19/00 12/19/00 12/19/00 3/6/01
12/21/00 12/21/00 12/21/00 12/21/00 3/7/01
1000 1000 20000 20000 20
3/6/01 3/6/01
3/7/01 3/7/01
20 200
3/6/01
3/7/01
200
AVERAGE: STANDARD DEVIATION:
RELATIVE STANDARD DEVIATION:
% Recovery
102 104 101 103 113 110 80 80 99 103 85 84 93 86 82 84 94 11 12
* Duplicate injection
LOQ = 0.01 |tg/g for liver and 0.01 |tg/mL for serum
Centre Analytical Laboratories, Inc.
Page 23 o f 102
Centre Study No.: 023-041
Table V. Summary of PFOS Recoveries in Quail Serum
Sponsor ID
na na na na na na na na na na na na na na na na
Centre ID
0011424 Spk A 0011424 Spk A* 0011424 Spk B 0011424 SpkB* 0011424 Spk A 0011424 Spk A* 0011424 SpkB 0011424 SpkB* 0011424 Spk A 0011424 Spk A* 0011424 SpkB 0011424 SpkB* 0011424 Spk A 0011424 Spk A* 0011424 SpkB 0011424 SpkB*
Set Number
121900BD 121900BD 121900BD 121900BD 122000A 122000A 122000A 122000A 122000B 122000B 122000B 122000B 030501A 030501A 030501A 030501A
Extraction Analysis
Amt
Date
Date
Added (ng/mL)
12/19/00 12/19/00 12/19/00 12/19/00 12/20/00 12/20/00
12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00
12/22/00 12/22/00
500 500 100000 100000 500 500
12/20/00 12/20/00 12/20/00 12/20/00 12/20/00 12/20/00 3/5/01 3/5/01 3/5/01 3/5/01
12/22/00 12/22/00 12/22-23/00 12/22-23/00 12/22-23/00 12/22-23/00 3/6-7/01 3/6-7/01 3/6-7/01 3/6-7/01
100000 100000
500 500 100000 100000 100 100 1000 1000
AVERAGE STANDARD DEVIATION RELATIVE STANDARD DEVIATION
% Recovery
103 106 90 93 106 103 97 103 108 110 113 98 122 122 107 115 106 9 9
* Duplicate injection
LOQ = 0.01 (ig/g for liver and 0.01 (ig/mL for serum
Centre Analytical Laboratories, Inc.
Page 24 o f 102
Centre Study No.: 023-041
Table VI. Summary of PFOS in Adult Liver Samples at 0 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
451 451* 452 452* 453 453* 454 454* 455 455* 456 456* 457 457* 458 458* 459 459* 460 460*
Centre ID
0007867 0007867 0007868 0007868 0007869 0007869 0007870 0007870 0007871 0007871 0007872 0007872 0007873 0007873 0007874 0007874 0007875 0007875 0007876 0007876
Set Number
121800AD 121800AD 121800A 121800A 121800AD 121800AD 121800A 121800A 121800AD 121800AD 121800A 121800A 121800AD 121800AD 121800A 121800A 121800AD 121800AD 121800A 121800A
Extraction Date
Analysis Date
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/19/00
12/18/00
12/19/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/19/00
12/18/00
12/19/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/19/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/19/00
12/18/00
12/19/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/19/00
12/18/00
12/19/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
0.215 0.215 NQ NQ 0.303 0.304 NQ NQ 0.114 0.111 NQ NQ 0.0762 0.0739 0.0103 0.0104 0.236 0.235 0.00117 0.00117 0.0953 0.113
* Duplicate injection
N Q = N ot Quantifiable (A peak w as detected at the corresponding analyte retention tim e
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 of 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 25 o f 102
Centre Study No.: 023-041
Table VII. Summary of PFOS in Adult Liver Samples at 1.8 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
E01-0259-20780 EOI-0259-20780* EOI-0259-20781 EOI-0259-20781* EOI-0259-20782 EOI-0259-20782* EOI-0259-20783 EOI-0259-20783* E 0 1-0259-20784 E01-0259-20784* EOI-0259-20790 EOI-0259-20790* EOI-0259-20791 EOI-0259-20791* EOI-0259-20792 E01-0259-20792* EOI-0259-20793 EOI-0259-20793* EOI-0259-20794 EOI-0259-20794*
Centre ID
0102997 0102997 0102998 0102998 0102999 0102999 0103000 0103000 0103001 0103001 0103007 0103007 0103008 0103008 0103009 0103009 0103010 0103010 0103011 0103011
Set Number
030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601 AD 030601AD 030601AD 030601AD 030601AD 030601A 030601A 030601 AD 030601AD 030601AD 030601AD
Extraction Date
Analysis Date
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/7/01
3/6/01
3/7/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
0.800 0.722 0.149 0.134 0.341 0.311 0.455 0.422 0.0988 0.0843 0.210 0.210 0.177 0.191 0.0124 0.0121 0.0381 0.0394 0.0629 0.0720 0.227 0.224
* Duplicate injection LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 26 o f 102
Centre Study No.: 023-041
Table VIII. Summary of PFOS in Adult Liver Samples at 6.2 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
E01-0259-20785 E01-0259-20785* E01-0259-20786 EOI-0259-2078 6* EOI-0259-20787 EOI-0259-20787* EOI-0259-20788 E01-0259-20788* E 0 1-0259-20789 E01-0259-20789* EOI-0259-20795 EOI-0259-20795* EOI-0259-20796 EOI-0259-20796* EOI-0259-20797 E01-0259-20797* EOI-0259-20798 EOI-0259-2079 8* EOI-0259-20799 EOI-0259-20799*
Centre ID
0103002 0103002 0103003 0103003 0103004 0103004 0103005 0103005 0103006 0103006 0103012 0103012 0103013 0103013 0103014 0103014 0103015 0103015 0103016 0103016
* Duplicate injection
Set Number
030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601AD 030601 AD
Extraction Date
Analysis Date
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
3/6/01
3/8/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
1.15 1.20 1.18 1.19 1.43 1.42 1.60 1.51 1.06 1.17 0.216 0.208 0.115 0.0934 0.0801 0.0797 0.187 0.218 0.0901 0.0954 0.715 0.606
LOQ = 0.01 |tg/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 27 o f 102
Centre Study No.: 023-041
Table IX. Summary of PFOS in Adult Liver Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
481 481* 482 482* 483 483* 484 484* 485 485* 486 486* 487 487* 488 488* 489 489* 490 490*
Centre ID
0007877 0007877 0007878 0007878 0007879 0007879 0007880 0007880 0007881 0007881 0007882 0007882 0007883 0007883 0007884 0007884 0007885 0007885 0007886 0007886
Set Number
121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD 121800AD
Extraction Date
Analysis Date
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
1.05 1.05 4.10 4.17 8.37 8.30 1.41 1.43 1.54 1.51 0.415 0.392 1.17 1.15 0.511 0.509 2.42 2.48 0.351 0.360 2.13 2.39
* Duplicate injection
LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 28 o f 102
Centre Study No.: 023-041
Table X. Summary of PFOS in Juvenile Liver Samples at 0 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9503 9503* 9506 9506* 9508 9508* 9512 9512* 9515 9515* 9518 9518* 9520 9520* 9522 9522* 9525 9525* 9526 9526*
Centre ID
0007907 0007907 0007908 0007908 0007909 0007909 0007910 0007910 0007911 0007911 0007912 0007912 0007913 0007913 0007914 0007914 0007915 0007915 0007916 0007916
Set Number
121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B
Extraction Date
Analysis Date
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
NQ NQ NQ NQ NQ NQ NQ NQ 0.000785 0.000755 NQ NQ 0.000734 0.000672 0.000718 0.000656 NQ NQ NQ NQ 0.000251 0.000316
* Duplicate injection
N Q = N ot Quantifiable (A peak w as detected at the corresponding analyte retention tim e
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 of 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 29 o f 102
Centre Study No.: 023-041
Table XI. Summary of PFOS in Juvenile Liver Samples at 1.8 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9528 9528* 9529 9529* 9530 9530* 9531 9531* 9533 9533* 9535 9535* 9540 9540* 9541 9541* 9543 9543* 9544 9544*
Centre ID
0007917 0007917 0007918 0007918 0007919 0007919 0007920 0007920 0007921 0007921 0007922 0007922 0007923 0007923 0007924 0007924 0007925 0007925 0007926 0007926
Set Number
121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B 121800B
Extraction Date
Analysis Date
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
12/18/00
12/20/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (ug/g)
0.0130 0.0128 0.00295 0.00295 0.00904 0.00876 0.00417 0.00435 0.0119 0.0117 0.0113 0.0114 0.00635 0.00623 0.00305 0.00297 0.0126 0.0126 0.00514 0.00521 0.00792 0.00393
* Duplicate injection LOQ = 0.01 |ig /g for liver and 0.01 fxg/mL for serum
Centre Analytical Laboratories, Inc.
Page 30 o f 102
Centre Study No.: 023-041
Table XII. Summary of PFOS in Juvenile Liver Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9548 9548* 9549 9549* 9551 9551* 9552 9552* 9555 9555* 9559 9559* 9561 9561* 9562 9562* 9563 9563* 9564 9564*
Centre ID
0007927 0007927 0007928 0007928 0007929 0007929 0007930 0007930 0007931 0007931 0007932 0007932 0007933 0007933 0007934 0007934 0007935 0007935 0007936 0007936
Set Number
121900A 121900A 121900A 121900A 121900A 121900A 121900A 121900A 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900A 121900A 121900AD 121900AD 121900A 121900A
Extraction
Analysis
Date
Date
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/21/00
12/19/00
12/21/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
0.00303 0.00305 0.0100 0.0105 0.0123 0.0125 0.00316 0.00302 0.0410 0.0408 0.0340 0.0341 0.0446 0.0445 0.0116 0.0121 0.0193 0.0197 0.00184 0.00201 0.0182 0.0156
* Duplicate injection LOQ = 0.01 p.g/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 31 o f 102
Centre Study No.: 023-041
Table XIII. Summary of PFOS in Juvenile Liver Samples at 17.6 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9567 9567* 9570 9570* 9575 9575* 9579 9579* 9580 9580* 9581 9581* 9582 9582* 9583 9583* 9584 9584* 9588 9588*
Centre ID
0007937 0007937 0007938 0007938 0007939 0007939 0007940 0007940 0007941 0007941 0007942 0007942 0007943 0007943 0007944 0007944 0007945 0007945 0007946 0007946
Set Number
121900A 121900A 121900AD 121900AD 121900A 121900A 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD 121900AD
Extraction Date
Analysis Date
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/21/00
12/19/00
12/21/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
12/19/00
12/23/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/g)
0.00904 0.00912 0.0424 0.0429 0.00629 0.00623 0.0880 0.0898 0.0461 0.0441 0.0492 0.0451 0.0747 0.0733 0.156 0.132 0.0983 0.101 0.0350 0.0370 0.0593 0.0417
* Duplicate injection LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 32 o f 102
Centre Study No.: 023-041
Table XIV. Summary of PFOS in Adult Serum Samples at 0 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
451 451* 452 452* 453 453* 454 454* 455 455* 456 456* 457 457* 458 458* 459 459*
Centre ID
0007887 0007887 0007888 0007888 0007889 0007889 0007890 0007890 0007891 0007891 0007892 0007892 0007893 0007893 0007894 0007894 0007895 0007895
Set Number
121900BD 121900BD 121900B 121900B 121900BD 121900BD 121900B 121900B 121900B 121900B 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD
Extraction
Analysis
Date
Date
12/19/00
12/23-24/00
12/19/00 12/19/00
12/23-24/00 12/21-22/00
12/19/00
12/21-22/00
12/19/00 12/19/00 12/19/00
12/23-24/00 12/23-24/00 12/21-22/00
12/19/00 12/19/00 12/19/00 12/19/00 12/19/00
12/21-22/00 12/21-22/00 12/21-22/00 12/23-24/00 12/23-24/00
12/19/00 12/19/00 12/19/00 12/19/00
12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00
12/19/00
12/23-24/00
12/19/00
12/23-24/00
AVERAGE: STANDARD DEVIATION:
Analyte Found (pg/mL)
11.6 11.8 0.0102 0.00909 9.04 10.4 0.00274 0.00212 0.0125 0.0123 6.32 6.46 6.58 6.35 0.105 0.102 8.75 8.50 4.78 4.64
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 (j.g/mL for serum
Centre Analytical Laboratories, Inc.
Page 33 o f 102
Centre Study No.: 023-041
Table XV. Summary of PFOS in Adult Serum Samples at 1.8 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
EOI-0259-20752 EOI-0259-20752* E01-0259-20753 EOI-0259-20753* EOI-0259-20754 EOI-0259-20754* EOI -0259-20755 E01-0259-20755* EOI-0259-20756 EOI-0259-20756* EOI-0259-20762 EOI-0259-20762* EOI-0259-20763 EOI-0259-20763* EOI-0259-20764 EOI-0259-20764* EOI-0259-20765 EOI-0259-20765* E 0 1-0259-20766 EOI-0259-20766*
Centre ID
0102969 0102969 0102970 0102970 0102971 0102971 0102972 0102972 0102973 0102973 0102979 0102979 0102980 0102980 0102981 0102981 0102982 0102982 0102983 0102983
Set Number
030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD
Extraction Date
Analysis Date
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/mL)
10.9 10.9 7.92 8.01 16.4 16.0 35.8 33.1 24.3 23.7 1.50 1.89 5.01 4.98 1.57 1.84 0.134 0.117 10.5 9.79 11.2 10.7
* Duplicate injection LOQ = 0.01 |ag/g for liver and 0.01 (J.g/mL for serum
Centre Analytical Laboratories, Ine.
Page 34 o f 102
Centre Study No.: 023-041
Table XVI. Summary of PFOS in Adult Serum Samples at 6.2 ppm a.i. (Test Termination 6 Weeks)
Sponsor ID
E01-0259-2075 7 E01-0259-20757* E01-0259-2075 8 E01-0259-20758* E01-0259-20759 EOI-0259-20759* EOI-0259-20760 EOl-0259-20760* EOI-0259-20761 EOI-0259-20761* EOI-0259-20767 EOI-0259-20767* EOI-0259-20768 EOI-0259-20768* EOI-0259-20769 E 0 1-0259-20769* EOI-0259-20770 EOI-0259-20770* EOI-0259-20771 EOI-0259-20771*
Centre ID
0102974 0102974 0102975 0102975 0102976 0102976 0102977 0102977 0102978 0102978 0102984 0102984 0102985 0102985 0102986 0102986 0102987 0102987 0102988 0102988
Set Number
030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD 030501AD
Extraction
Analysis
Date
Date
3/5/01 3/5/01 3/5/01 3/5/01 3/5/01 3/5/01 3/5/01
3/7-8/01 3/7-8/01 3/7-8/01 3/7-8/01 3/7-8/01 3/7-8/01 3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
3/5/01
3/7-8/01
AVERAGE:
STANDARD DEVIATION:
Analyte Found (ug/mL)
52.2 56.1 102 94.7 80.4 84.6 69.2 62.9 84.6 86.7 4.57 4.07 2.72 2.61 4.02 3.87 3.24 3.31 2.10 1.70 40.3 39.7
* Duplicate injection LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 35 o f 102
Centre Study No.: 023-041
Table XVIL Summary of PFOS in Adult Serum Samples at 17.6 ppm a.i. (Test Termination 19 Weeks)
Sponsor ID
481 481* 482 482* 484 484* 485 485* 487 487* 488 488* 489 489* 490 490*
Centre ID
0007896 0007896 0007897 0007897 0007898 0007898 0007899 0007899 0007900 0007900 0007901 0007901 0007902 0007902 0007903 0007903
Set Number
121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 121900BD 030501AD 030501AD 121900BD 121900BD
Extraction
Analysis
Date
Date
12/19/00 12/19/00 12/19/00 12/19/00 12/19/00 12/19/00 12/19/00
12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00 12/23-24/00
12/19/00
12/23-24/00
12/19/00
12/23-24/00
12/19/00
12/23-24/00
12/19/00
12/23-24/00
12/19/00
12/23-24/00
3/5/01
3/7-8/01
3/5/01
3/7-8/01
12/19/00
12/23-24/00
12/19/00
12/23-24/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (ug/mL)
260 263 43.1 42.4 118 106 179 172 133 127 15.8 16.8 232 228 6.87 6.80 122 93.0
* Duplicate injection
LOQ = 0.01 |ig/g for liver and 0.01 (ig/mL for serum
Centre Analytical Laboratories, Inc.
Page 36 o f 102
Centre Study No.: 023-041
Table XVIII. Summary of PFOS in Juvenile Serum Samples at 0 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9503 9503* 9506 9506* 9508 9508* 9512 9512* 9515 9515* 9518 9518* 9520 9520* 9525 9525* 9526 9526*
Centre ID
0007947 0007947 0007948 0007948 0007949 0007949 0007950 0007950 0007951 0007951 0007952 0007952 0007953 0007953 0007954 0007954 0007955 0007955
Set Number
122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A
Extraction
Analysis
Date
Date
12/20/00 12/20/00 12/20/00 12/20/00
12/22/00 12/22/00 12/22/00 12/22/00
12/20/00 12/20/00
12/22/00 12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/mL)
0.00486 0.00385 0.00718 0.00754 0.00365 0.00319 0.00431 0.00457 0.00789 0.00528 0.00378 0.00481 0.00367 0.00395 0.00552 0.00438 0.00354 0.00388 0.00477 0.00142
* Duplicate injection
LOQ = 0.01 ng/g for liver and 0.01 ng/mL for serum
Centre Analytical Laboratories, Inc.
Page 37 o f 102
Centre Study No.: 023-041
Table XIX. Summary of PFOS in Juvenile Serum Samples at 1.8 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9528 9528* 9529 9529* 9530 9530* 9531 9531* 9533 9533* 9535 9535* 9540 9540* 9541 9541* 9543 9543* 9544 9544*
Centre ID
0007956 0007956 0007957 0007957 0007958 0007958 0007959 0007959 0007960 0007960 0007961 0007961 0007962 0007962 0007963 0007963 0007964 0007964 0007965 0007965
Set Number
122000D 122000D 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A 122000A
Extraction Date
Analysis Date
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
12/20/00
12/22/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/mL)
0.0762 0.0670 0.0273 0.0284 0.00785 0.00824 0.0480 0.0476 0.0488 0.0494 0.0416 0.0413 0.0192 0.0189 0.0321 0.0334 0.00969 0.00983 0.0158 0.0166 0.0324 0.0198
* Duplicate injection LOQ = 0.01 |ig/g for liver and 0.01 [ig/mL for serum
Centre Analytical Laboratories, Ine.
Page 38 o f 102
Centre Study No.: 023-041
Table XX. Summary of PFOS in Juvenile Serum Samples at 6.2 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9548 9548* 9549 9549* 9551 9551* 9552 9552* 9555 9555* 9559 9559* 9561 9561* 9562 9562* 9563 9563* 9564 9564*
Centre ID
0007966 0007966 0007967 0007967 0007968 0007968 0007969 0007969 0007970 0007970 0007971 0007971 0007972 0007972 0007973 0007973 0007974 0007974 0007975 0007975
Set Number
122000D 122000D 122000D 122000D 122000D 122000D 122000B 122000B 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D
Extraction
Analysis
Date
Date
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/22-23/00
12/20/00
12/22-23/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/mL)
0.137 0.160 0.382 0.390 0.222 0.208 0.0542 0.0535 0.243 0.218 0.161 0.168 0.185 0.171 0.231 0.229 0.259 0.254 0.171 0.173 0.203 0.0838
* Duplicate injection LOQ = 0.01 p g/g for liver and 0.01 (xg/mL for serum
Centre Analytical Laboratories, Inc.
Page 39 o f 102
Centre Study No.: 023-041
Table XXI. Summary of PFOS in Juvenile Serum Samples at 17.6 ppm a.i. (Test Termination 12 Weeks)
Sponsor ID
9567 9567* 9570 9570* 9575 9575* 9579 9579* 9580 9580* 9581 9581* 9582 9582* 9583 9583* 9584 9584* 9588 9588*
Centre ID
0007976 0007976 0007977 0007977 0007978 0007978 0007979 0007979 0007980 0007980 0007981 0007981 0007982 0007982 0007983 0007983 0007984 0007984 0007985 0007985
Set Number
122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D 122000D
Extraction
Analysis
Date
Date
12/20/00 12/20/00
12/24/00 12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
12/20/00
12/24/00
AVERAGE:
STANDARD DEVIATION:
Analyte Found (pg/mL)
0.329 0.355 0.483 0.512 0.241 0.252 0.406 0.424 0.287 0.254 0.295 0.285 0.459 0.473 0.521 0.524 0.553 0.564 0.532 0.509 0.413 0.114
* Duplicate injection LOQ = 0.01 pg/g for liver and 0.01 pg/m L for serum
Centre Analytical Laboratories, Inc.
Page 40 o f 102
Centre Study No.: 023-041
Table XXII. Summary o f Percent M oistures for Quail Liver Samples
Sponsor ID
451 452 453 454 455 456 457 458 460 482 484 486 488 490 9506 9508 9512 9518 9520 9522 9526 9530 9540 9543 9544 9548 9551 9552 9555
Centre ID
0007867 0007868 0007869 0007870 0007871 0007872 0007873 0007874 0007876 0007878 0007880 0007882 0007884 0007886 0007908 0007909 0007910 0007912 0007913 0007914 0007916 0007919 0007923 0007925 0007926 0007927 0007929 0007930 0007931
% Moisture
Sponsor ID
66.7 64.4 68.4 65.6 64.1 71.8 59.5 71.2 69.0 55.0 60.6 72.5 61.5 65.1 70.7 72.1 67.4 73.2 73.5 68.3 67.9 70.0 66.7 57.1 55.8 59.4 81.3 53.5 65.0
9559 9561 9562 9563 9564 9570 9579 9584 9588 E01-0259-20780 EOI-0259-20781 EOI-0259-20782 EOI-0259-20783 EOI-0259-20784 EOI-0259-20785 EOI-0259-20786 EOI-0259-20787 EOI-0259-20788 EOI-0259-20789 EOI-0259-20790 EOI-0259-20791 EOI-0259-20792 EOI-0259-20793 EOI-0259-20794 EOI-0259-20795 EOI-0259-20796 EOI-0259-20797 EOI-0259-20798 EOI-0259-20799
Centre ID
0007932 0007933 0007934 0007935 0007936 0007938 0007940 0007945 0007946 0102997 0102998 0102999 0103000 0103001 0103002 0103003 0103004 0103005 0103006 0103007 0103008 0103009 0103010 0103011 0103012 0103013 0103014 0103015 0103016
% Moisture
78.3 80.6 57.8 63.4 34.6 85.2 88.7 59.6 65.2 70.3 57.8 70.6 67.0 67.7 64.5 72.3 67.6 68.0 68.0 69.3 70.9 67.0 65.3 74.5 65.7 71.7 72.8 71.3 72.1
Centre Analytical Laboratories, Inc
Page 41 o f 102
Centre Study No.: 023-041
FIGURES
Centre Analytical Laboratories, Inc.
Page 42 o f 102
Centre Study No.: 023-041
Figure 1. Typical Calibration Curve for PFOS
<_o : in
Ioo
r
\<o
i** I* t\ *
TT
Concentration, ng/mL
1O
L
1CM
CO CM
K
! CM
sol
: CM
CM
Io |CM
L< r*
! 00
| "d
1
1 !'M*
ro
! CM rd
M ' M*
o o q o q q o o- o q
0) m
0) 0) 0) CO CM *
0) o0) doo doo doo o> GO N
doo
CO
dooto
doo
doo doo - doo
CO CM v
d
sjunoo `eajy
Centre Analytical Laboratories, Inc.
Page 43 o f 102
Centre Study No.: 023-041
Figure 2. Chrom atogram Representing a 0.1 ng/m L standard for PFOS
jg sfcatasB a
081
Sample Type: Concentration: Acq. Data: Acq. Tima:
QC 0.100
12/22/00 10:37:10 AH
Modified: Bunching factoc: Noise Threshold: Area Threshold: Num. Smooths: RT Window: Expected RT: Sep. Width: Sep. Height: Exp. Peak Ratio: Exp. Adj, Ratio: Exp. Vai. Ratio: Use Relative RT:
Yes 1 1.91 9.55 1 30.0 4.51 0.20 0.01 S.00 4.00 3.00 No
min
$
c
233 . 220
i 200
180j
isoi
40 ;
120 i
too -
Int. Type: Retention Time: Area:
Height: Start Time: End Time:
Manual 4.49 min
1039.0 counts 2.18e+002
4.40 4.59 in
50, 0 4 40-
20 J
oU
I,'4 * 1 5.12 5.4 5.90 * ? J .5.53 ,, .7.45
J
3.5 4.0 4.5 Time, min
Centre Analytical Laboratories, Inc.
Page 44 o f 102
Centre Study No.: 023-041
Figure 3. Chromatogram Representing a 1.0 ng/mL standard for PFOS
Sample Type:
Concentration: Aeq. Oat: Acq. Time:
Standard 1.000
12/22/00 11:15:0 AM
1161 1100-
Modified: Bunchinq Tactor: Noise Threshold: Area Threshold: Num. Smooths: RT Window: xpected RT: Sep. width: Sep. Height: Cxp. Peak Ratio: exp. AdJ. Ratio: exp. Val. Ratio: Ose Relative R7:
Yes 1000:
1
1.91 9.35
600
1 30.0
sec
800
4.51 min
0.20
700-
0.01 5.00
4.00 3.00 No
it 1
600 ; ? 1 .500 -
Int. Type: Retention Time: Area: Helqht: Start Time: end Time:
Manual
4.50 min 468.6 counts 1.13e*00:
4.40
4.61 min
1400-
300 -j
200-i
100 \
ol
0.S f.O 1.5 2.0 * 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5
Tim, min
Centre Analytical Laboratories, Inc.
Page 45 o f 102
Centre Study No.: 023-041
Figure 4. Chromatogram Representing a Reagent Blank for PFOS (Centre ID Reagent Blank A, Set: 122000A)
Smpr~Rlimim22QOOA^OOB~.rSimpi
Sample Index: 1 Sample Type: Concentracin: Acq. Date: , Acq. Time:
8 Blank 0.000 12/22/00 12:05:48 PM
1 Modified: ' Bunching Factor: i Noise Threshold: ' Area Threshold:
Hum. Smooths: ' RT Window: ' Expected RT: ' Sep. Width: ; Sep. Height: . Exp. Peak Ratio: Exp. Adj. Ratio:
Exp. Val. Ratio: Use Relative RT:
Yes 1 1.91 9.55 1 30.0 4.51 0.20 0.01 5.00 4.00 3.00 No
sec min
490
450I
4001
350 i
-- 300 1
a- j 250-
s# -- 200
' Int. Type: Retention Time:
i Area: ' Height: i Start Time: ' End Time:
Manual 4.50 min
2299.8 counts 4.<0e*002 cps
4.39 min 4.7 min
ISO*
too :
50
0-
'v 5.0! 5 3e 5.8 j 28 8.8 8.85 7.3! 7-58 7.71
" 77 f i f s r S S 4.0 .5 5.0 5.5 8.0 6.5 7.0 7.5
Time, min
'
-
Centre Analytical Laboratories, Inc.
Page 46 o f 102
Centre Study No.: 023-041
Figure 5. Chromatogram Representing Control Quail Serum for PFOS (Centre ID: 0011424 Blank A, Set: 122000A)
Sample Index: Sample Type: Concentration: Acq. DaCe: Acq. Time:
10 Blank 0.000 12/22/00 12:31:00 PM
Modified:
Bunching Factor: Noise Threshold: Area Threshold: Hum. Smooths: RT Window:
Expected RT: Sep. Width: Sep. Height: Exp. Peak Ratio: Exp. AdJ. Ratio: Exp. Val. Ratio: Use Relative RT:
Yes 1 1.91 9.55 1 30.0
4. SI 0.20 0.01 $.00 4.00 3.00 No
sec min ,
I
Int. Type: Retention Time:
Ares: Height: Start Time: End Time:
Manual 4.SO min
4770.8 counts l.03%+003 cps
4.39 min 4.43 min
1083 1000 :
800
700
600
500
400 {
300-
200-
1
100-
i
0-
A.n
t.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5
Tim*, min
-
6.5 7.0 7.5
Centre Analytical Laboratories, Inc.
Page 47 o f 102
Centre Study No.: 023-041
Figure 6. Chromatogram Representing Control Quail Liver for PFOS, (Centre ID: 0007870 Blank A, Set: 121900A)
. Sample Index: | Sample Type: < Concentration: j Acq. Date:
Acq. Time:
52
Blank 0.000 12/21/00 11:29:44 AM
i Modified:
Yes
: Bunching Factor: 1
I Noise Threshold:
1 Area Threshold: 1 Hum. Smooths: j RT Window:
1.91 9.55 1 30.0
sec
: Expected RT:
4.51 in
| Sep. Width:
0.20
1 Sep. Height: ! Exp. Peak Ratio:
0.01 5.00
1
! Exp. Adj. Ratio: Exp. Val. Ratio:
4.00 3.00
$
I
! Use Relative RT: Ho
c
j Int. Type:
1 Retention Time: | Area:
j Height:
: Start Time:
} End Time:
Manual 4.48
482.8 counts l.oie+oo;
4.37 in 4.61 in
108 100 90-
70 1
60 < 50, 40 . 50 20,0 0-
14.57 .. . . . . *,J
3.5 4.0 4.3 Tim, mm
,5.oe
517 5.57
Centre Analytical Laboratories, Inc.
Page 48 o f 102
Centre Study No.: 023-041
Figure 7.
Chromatogram Representing Control Quail Serum Fortified with 500 ng/mL o f PFOS (Centre ID: 0011424 Spk A, Set: 122000A)
| Sample Index: Sample Type:
! Concentration: ; Acq. Date: ' Acq. Time:
12
QC 2.000 12/22/00 12:56:17 PM
Modified: Bunching Factor: Noise Threshold: Area Threshold: Num. Smooths: AT Window: Expected AT: Sep. Width: Sep. Height: Exp. Peak Aatio: Exp. Adj. Aatio: Exp. Val. Aatio:
Use Aelative AT:
Yes 1
1.91 9.55 1 30.0 4.51 0.20 0.01 5.00 4.00 3.00 No
.
&
I
Int. Type: Aetention Time: Area: Height: Start Time: End Time:
Manual
4.49 7570.S counts 1.79e+003 cps
4.40 4.62 min
1000
1400 !
1200-
1000 ;
800i
600, *
400 :
200 i 1
0-
1.5 2.0 2 5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 Time, min
7.0. 7.5
Centre Analytical Laboratories, Inc,
Page 49 o f 102
Centre StudyNo.: 023-041
Figure 8.
Chromatogram Representing Control Quail Liver Fortified at 10 ppb with PFOS (Centre ID: 0011423 Spk A, Set: 121800A)
Spl Type ! Concentration: ! Aeq. Saco: ; Aeq. Tiaa:
4.000 12/19/00 0 2 :0 1:40 PH
] IMltUdl | Bunching factor:
i Hois Threshold: I Aroa Threshold: | Mu m . Smooths: ; AT Window: , txpeetod AT: ; lap. Width: i lop. Netqht: tap. Peak Ratio: ' Cop. Ad}. Ratio: I Cap. Val. Aatio: ; Use Aolativo AT:
1.91
19 .1$
30.0
4 .SI 0.20
05..0010
4.00 3.00
No
toe min
; Int. Typo: ' Aotoneion Tina: ! Aroa: ' Koiqhe: ! Itart Time: ! end t i n s :
4 .SO 17214.
S.3le*003 4.43 4.41 min
t 3000
I
^2000
6 0 OS 7.0 - 7 9
Centre Analytical Laboratories, Inc.
Page 50 o f 102
Centre Study No.: 023-041
Figure 9.
Chromatogram Representing Juvenile Quail Serum Sample (Centre ID: 0007965, Sponsor ID: 9544, Set: 122000A)
i Sample Index: ! Sample Type: I Concentration:
1 Acq. Date: | Acq. Time:
57
Unknown N/A
12/22/00 10:29:14 PM
! Modified: ! Bunching Factor:
* Noise Threshold: Area Threshold:
\ Num. Smooths: RT Window: | Expected RT:
' Sep. Width: j Sep. Height: j Exp. Peak Ratio: ' Exp. Adj. Ratio: 1 Exp. Val. Ratio: | Use Relative RT:
Yes l 1.91
9.55 1 30.0 4.51 0.20 0.01 5.00 4.00 3.00 No
min
m C
; Int. Type: Retention Time:
Area: : Height:
Start Time: : End Time:
Manual
4.SO min 5149.2 counts 1.19e*003 cps
4.40 min 4.59 min
1200 > 11001000 00 00 ; 700 600500 < 400 | 300 200100 ;
0-
3.5 4.0
4.S
______Time, min______
Centre Analytical Laboratories, Inc,
Page 51 o f 102
Centre Study No.: 023-041
Figure 10. Chromatogram Representing Juvenile Quail Liver Sample (Centre ID: 0007926, Sponsor ID: 9544, Set: 121800B)
1 Sample Index: 1 Sample Type: j Concentracin: j Acq. Date: | Acq. Time:
59 Unknown
N/A 12/20/00 .01:33:11 PM
2101 ; 2000 -j
| Moditied:
Yes
I Bunching Factor: - 1
I Noiae Threshold: 1.91
1 Area Threshold:
1 Hum. Smooths: j RT Window:
9.55
l
30.0
sec
Expected RT: 1 Sep. Width:
; Sep. Height-: i Exp. Peak Ratio: 'I Exp. Adj. Ratio:
4.51 0.20 0.01 5.00 4.00
min
m
5
Exp. Val. Ratio:
! Use Relative RT:
3.00 No
%c
Int. Type:
1 Retention Time:
Area:
> Height:
. Start Time:
i End Time:
Manual
4.47 min
7435.8 eounts
2.06e*003 cps
4.38 4.58
ma inn
1000 j
1600 I
1400 -i
i
t200 !
1000 !
800
600 [
400 -
200
0-
" ^ 5 ----- ^
T 2d) s 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5
- Tim, min ,,
.
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Centre Study No.: 023-041
APPENDIX A
Study Protocol 00P-023-041
(Centre Study No. 023-041) and
Amendments and Deviations
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.m .
Centre Study No.: 023-041
' Centre Protocol No. 00P-023-041
STUDY PROTOCOL
STUDY TITLE
EXTRACTION OF POTASSIUM PERFLUOROOCTANESULFONATE FROM
QUAIL SERUM AND QUAIL LIVER FOR ANALYSIS USING HPLC- '
ELECTROSPRAY/MASS SPECTROMETRY
SPONSOR
3M Environmental Technology and Safety Services
.
Building 2-3E-09
'
. PO Box 33331
St. Paul, 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 NUMBER : 00P-023-041
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Centre Study No.: 023-041
Centre Protocol No. OOP-023-041
Title: EXTRACTION OF POTASSIUM PERFLUOROOCTANESULFONATE FROM QUAIL SERUM AND QUAIL LIVER FOR ANALYSIS USING HPLC. . . ELECTROSPRAY/MASS SPECTROMETRY
TABLE OF CONTENTS ______ _________ ;__________________________ __________ ;______________ Page TABLE OF CONTENTS .............. ................................................................................ 2
1. PURPOSE..................................
3
2. TEST MATERIALS...... .....................................
3
3. SPONSOR......... ...................................................................... ...............................4
4. TESTING FACEJIY AND PERFORMING LABORATORY....................;......4
5. PROPOSED EXPERIMENTAL TIME-FRAME................
4
6. JUSTIFICATION FOR THE SELECTION OF THE TEST SYSTEM................. 4
7. . SAMPLE PROCESSING, STORAGE AND IDENTIFICATION
...:. 5
8. ANALYTICAL METHOD ............................................................................. 5
9. EXPERIMENTAL DESIGN....................!...................... :..................................... 10
10. PROTOCOL AMENDMENTS AND DEVIATIONS........................................... 10
11. RECORDS.............. ..........:................................................ ............................ ......11
12. QUALITY ASSURANCE......... !................... ................ ...........'...................... 12
13. DATA AND REPORT ...... ........;..............*....................................................12 14. ARCHIVE STATEMENT...................................................................................... 13
15. REFERENCES___ .'............................ .... ........................................................13
16. PROTOCOL APPROVAL....................................................... i............................14
APPENDIX: ANALYTICAL METHODS...................................... ................................. 15
I
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Centre Study No.: 023-041
f
.
Centre Protocol No. OOP-023-041
Study Title:
i
EXTRACTION OF POTASSIUM PERFLUOROOCTANESULFONATE FROM
QUAIL SERUM AND QUAIL LIVER FOR ANALYSIS USING HPLC-
ELECTROSPRAY/MSS SPECTROMETRY
1. PURPOSE
The purpose of this study is to analyze quail serum samples and quail liver samples for residues of perfluorooctanesulfonate (PFOS) using methods entitled, ' "Extraction Of Pbtassium 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 use '
'
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:
' Perfluorooctanesulfonate
1-Octanesulfonic acid.l, 1,2,2,3,3,4,4,5,5,6,6t7,7,8,8,8-
heptadecafluoro-, potassium salt
.
CAS Number
2795-39-3
'
Molecular Weight: 499 (CjFnSOj- )
O
II .
C8F 17S-- O
II
o
Note: The neutral molecule and standard form that PFOS (anion) is derived from is perfluorooctanesulfonate potassium salt [CgFnSOjK], molecular weight 538.
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Centre Protocol No. OOP-023-041
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 the raw data package.
All standards/test substances and any prepared solutions must be identified with a unique label or number on the container or cross-referenced to the container.
HAZARD INFORMATION
A current MSDS for the chemical(s) used in this study will be maintained at the
testing facility.
3. SPONSOR
3M Environmental Technology and Safety Services
Building 2-3E-09
PO Box 33331
' St. Paul, MN 55133-3331
.
STUDY DIRECTOR: Rochelle Robideau TELEPHONE: 651-778-7065 FAX NUMBER: 651-778-6176
.
. .
. .
-
"
.
4. TESTING FACILITY AND PERFORMING LABORATORY
Centre Analytical Laboratories, Inc. (Centre)
3048 Research Drive State College, PA 16801
-
PRINCIPAL INVESTIGATOR:
Emily R. Stauffer, Centre TELEPHONE: 814-231-8032 FAX NUMBER: 814-231-1580
5. PROPOSED EXPERIMENTAL TIME-FRAME
Experimental Start Date Experimental Termination Date Report Issued
Dec. 11,2000 Dec. 15,2000 Dec. 30,2000
..
6. JUSTIFICATION FOR THE SELECTION OF THE TEST SYSTEM '
Quail serum samples arid liver samples collected by Wildlife International, Ltd. will be used as the test systems for this study. Quails represent wild bird populations and they are an EPA recommended species because they do well in a laboratory environment Quail habitats include bushy pastures, grassy roadsides,
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Centre Study No.: 023-041
Centre Protocol No. OOP-023-041
farmlands! and open woodlands, and they are considered land 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 fed PFOS.
'
-
A complete description of the test system along with all of die in-life parameters is documented in the sponsor's protocol no. 454/01060Q/QP/SUB454, Project No. 434-104 that is associated with this analytical protocol.
7. SAMPLE PROCESSING, 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 prior to extraction; however, they will be homogenized by laying the samples on a flat surface and . mashing them by hand as 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.substanceduring 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 container or cross-referenced to the container.
8. ANALYTICAL METHOD
The serum samples will be analyzed according to the analytical method titled "Extraction Of Potassium Perfluorooctanesulfonate Or Other Buorochemical Compounds ' From Serum For Analysis Using HPLG-Electrospray/Mass Spectrometry" (3M method number ETS-8-4.1) and the control liver samples will be analyzed according to the method titled "Extraction Of Potassium Perfluorooctanesulfonate Or. Other.Fluprochemical Compounds From Liver For * Analysis Using HPLC-Electrospray/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, and calibration standards will be prepared as follows:
I
Stock Solution Prepare a stock solution of PFOS at 100 |ig/mL by weighing out 10.0 mg of analytical standard (corrected for 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-mL LDPE bottles) in a refrigerator at 2C to 6C for a . maximum period of 6 months from the date of preparation.
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Centre Study No.: 023-041
(entre Protocol No. 00P-023-041
Fortification Solutions a. 1.0 ug/mL 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.
b. 0.1 ug/mL Fortification Solution - Pipette 10.0 mL of the 1.0 pg/ml
fortification solution into a 100-mL volumetric flask and bring up to
r" vluitrwtthUHSthsnoI......
'
c. 0.01 Ug/mL Fortification 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
date of preparation.
-
Calibration Standards Prepare six LC/MS/MS calibration standards in methaiiol via dilution of the 0.1 pg/mL and the 0.01 pg/mL fortification solutions.
This is a typical example; additional concentrations may be prepared as
needed.
' .
Initial Cone. (pg/mL) 0.1 0.1 0.1 0.01 0.01 0.01
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
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 of preparation. -
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 mashed on a flat surface by hand prior to 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 M TBA 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 only be filtered with 0.2 pm nylon mesh filter if
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Centre StudyNo.: 023-041
1
Centre Protocol No. 00P-023-041
necessary. Samples will be reconstituted in a final volume of 1 mL of methanol for both matrices.
4. The samples will be analyzed under the following conditions: LC/MS/MS System and Operating Conditions (Turbolonspray)
Mass Spec: PE SCIEX API 3000 Biomolecular Mass Analyzer
- Interface: SCIEX Turboion Spray Liquid Introduction Interface ' Harvard infusion pump
Computer Power Macintosh G3
Software:
PE SciexAnaiyst LI Windows NT
' .
.HPLC: .
Hewlett Packard (HP) Series 1100 EBPQuat Pump
_ HP Vacuum Degasser HP Autosampler
` HP Column Oven .
HPLC Column:Genesis Cg (Jones Chromatography), 2.1 nun x 50 mm, 4p
Column Temp.: 35 C .
. Injection Vol.: 10 pL
Mobile Phase (A): '2 mM Ammonium Acetate in ASTM type I water
Mobile Phase (B): Methanol
Row Rate: 0.3 mL/min.
.
. Time 3 k A SLS.
0 6 0 . .40
1.0 0
100
7.0 0 : ' 100
7.5 4 0 . 60
11.0 40 . 60
.
.
I-
It may be necessary to adjust the HPLC gradient in order to optimize
instrument performance. Columns with different dimensions (e.g., 2.1
mm x 30 mm) and columns from different manufacturers (Keystone
Betasil Cis etc.) can be used, provided equivalent chromatography is
obtained.
`
Ions monitored:
.
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Centre Protocol No. OOP-023-041
Analvte PFOS
I Approximate
Mode lramilip.O.Monitpred Retention Time (mini
negative
499 -> 99 '
4.20
On a day-to-day 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 may be changed from time to time in
order to optimize for greatest sensitivity.
.
The mass spectrometer is tuned using a 0.5pg/mL PFOS solution, prepared via
dilution of the stock solution in methanol. -1116 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 saved as a "tune file". This tune file is then
used during routine analysis. The tuning procedure may be repeated as necessary
to ensure optimal sensitivity.
'
Controls
.
. Set '
. ' IS-Iospray
-4000.0
OR-Orifice
-
-61.0
- RNG-Focus Ring `
- -270.0
QO-Quad0 Rod Offset
- 10.0
IQ1- Interquad 1 lens
9.3
ST-Stubbies
-15.0
ROl-Quadl Rod Offset
. 9.3
IQ2-lhter quad 2 lens
20.0
R02-Quad 2 rod offset
-84.0
ST3-Stubbies'
100.0
R03- Quad 3 rod offset
86.0
. DF-CEMDeflection Plate
300.0
CEM-Channel Electron Multiplier
2800.0
` Gas Flows :
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 (prepared in MeOH).into the LC/MS/MS.
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Centre Study No.: 023-041
. Centre Protocol No. OOP-023-041
b. Use linear, 1/x weighted standard curves 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 be excluded from the calibration curve. However, the total number of calibration standards that may be excluded must not exceed 30% of the total number of standards injected.
c. The correlation coefficient (r) for calibration curves generated
must be 0.9925 (r^0.985). If calibration results fall outside these limits, then appropriate steps should be taken to adjust instrument operation, and the relevant set 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 analyzed in duplicate.
-'
a. Standards corresponding to at least six concentrations must be included in an analytical 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 must be bracketed by standard injections. *
'
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 and carried through the procedure to verify recovery.
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 standard responses
must bracket responses of the residue found in the sample set -If
necessary, dilute the samples and re-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 reanalysis of the affected samples. -
f. Samples in which no peaks are detected at the corresponding analyte retention times will be reported as ND (not detected). Samples in which peaks are detected at the corresponding analyte retention times
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Centre Protocol No. OOP-023-041
but are less than 3:1 signalmoise will be reported as NQ (not quantifiable).
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 analyzed preferably within a week.
9. EXPERIMENTAL DESIGN
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-Electrospray/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 of fortified samples to provide an indication of reproducibility. .
The average recovery and relative standard deviation of the fortified samples will
be calculated. Also, the average residue found and standard deviation for each
matrix will be calculated.
-
Moisture determination will be performed On all liver samples according to
Centre SOP CAL-10B/11 with the following modifications:
.
1. The sample weight will be - 0.5 g.
.
2. The drying time will be - 12 hours.
10. PROTOCOL AMENDMENTS AND DEVIATIONS ,
Any deviations from the protocol or from the analytical method as provided will
be documented and reported promptly 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.
.
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"
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CentreProtocol No. OOP-023-041
1. Protocol amendments: Planned changes to the approved protocol shall be documented by amendments that clearly describe rite change, justification for the change, and impact on the study. Amendments will be signed and dated by the Principal Investigator and Study Director. Copies of amendments will be sent to the quality assurance unit.
2. Protocol deviations: Protocol deviations, which a one time and unplanned deviations from the protocol, shall be documented in the study records, 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 and Study Director and reviewed by QAU.
11. RECORDS
Records to be maintained include,-but are not limited the following (as
appropriate):
1. Sample tracking sheet(s)
-
2. Sample receipt records, storage history, and chains of custody
3. History and preparation of standards (stock, fortification, Calibration)
4. Description of any modifications to the method
5. Instrument run sheets, bench-sheets or logs '
.
6. Analytical data tables
' .-
7. All chromatographic and instrumental conditions
8. Sample extraction and analysis dates
9. A complete listing of study personnel, signatures and initials
10. Chronological presentation of all study correspondence
11. Any other data necessary for the reconstruction of the study
All chromatograms will contain the following:
'
a. Sample identification, date, arrow or other indication of the area of interest, and injection number corresponding 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, ng/mL, ppb, ppt, etc.).
Each data set will contain information on temperatures,, flow rates, column
parameters, gases, instrument parameters, and instrument type, etc. The Centre
study number will also be recorded on the first chromatogram of each' daily
analytical run.
'
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entre Study No.: 023-041
Centre Protocol No. 00P-023-041.
12. QUALITY ASSURANCE
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 the.Principal Investigator, Centre Management, and the Study Director.
`
t
13. DATA AND REPORT
1. All raw data and the original sighed protocolwill be maintained in the study file. This data includes the protocol' amendments, protocol deviations, laboratory notebooks, analytical standard solution preparation, sample chain o f custody sheets, sample work sheets, chromatograms, calibration curves, and any other appropriate data generated. Raw. data not used will be stored in the study file. The reason for exclusion 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 limited to:
'
1. Objectives and procedures stated in the protocol .
2. Analytical and statistical methods used
3. Test materials identified by name, lot, purity, and other characteristics
4. Name of testing facility and study initiation and completion dates
. 3. Statement prepared and signed by the qualify assurance unit
6. Tables containing all applicable data
_
7. All chromatographic and instrumental conditions
5. A complete listing of Centre study personnel '
.
Centre will send a copy of the draft report to the Study Director, who will return - the report with com m ents. Centre Analytical will make revisions and finalize 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 GLPcompliance. 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
Investigator and the Study Director.
.
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REPORT DISTRIBUTION: Original: Study Director
' Copy: Centre Archives
'-
14. ARCHIVE STATEMENT
i
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. This includes a description of equipment used during the conduct of the study. All characterization data and any 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 a copy for an original record, it will
be certified as an exact copy. Centre will retain facility-related originals and a copy
of the final report and raw data package.
15. REFERENCES
.
1 U.S. Environmental Protection Agency. Toxic Substances Control Act (TSCA).
. Good Laboratory Practice Standards. Final Rule, 40 CFR Part 792.
'
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Centre Study No.: 023-041
Centre Protocol No. OOP-023-041
16. PROTOCOL APPROVAL This protocol was audited by the Quality Assurance Unit of Centre Analytical Laboratories, Inc. Quality Assurance Reviewer, Centre
''UtLjtou: IW Naomi Lovallo, Phi).
Quality Assurance Auditor
/iT
Principal Investigator, Centre
, /.In ha Date
Emily Stauffer Facility Management, Centre
Date
Study Director, 3M
. Date
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Centre Study No.: 023-041
Centre Protocol No. OOP-023-041 .
APPENDIX: ANALYTICAL METHODS
A. "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical
Compounds From Serum for Analysis Using HPLC-Electrdspray/Mass
Spectrometry"
'
B. "Extraction of Potassium Perfluorooctanesulfonate or Other Fluorochemical
Compounds From liv er for Analysis Using HPLC-Electrospray/Mass
Spectrometry"
`
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Centre StudyNo.: 023-041
"'CentreProtacol-Nor00P-O23-041
\ V; '" "
VV.' ' . . OROther
i . i ' 1. '. ^ : '
. "-.'J -'
AdoptionDate: 03/01/99
Revision Date: 1/3 7/99 :
. ... Approved B y .;:
i " V . . > ' ' f ' ` .. :.'v _ >.J,. ,: -. ; ^ p ite ; -------- :----;--j -
__ -- ' ': t l / k, - ;
- ^ c h m o a l R e v w w e r <
\V
. '-
U S co tt ifip A w r / noM y v
1.1 - Scope: This methodis forthe e*u<KuwuiputHiuB.pan
wotherfluorocheimcalcorapoundsfromscnmi.
/ ,. .
..
1-2 Applicable compounds: Fluorochemical surfactants or other fluorinatcd compounds '
^ the
ra<`*H)V"K ' monkey, and human scrum dr otherfluids as designated in
O:
,v'r :
. W&W ;-*
ei$-s-u -' ;..
f *V:.f. v ;r#* .yt'.B.mwtwcdn--onjotftWrrOuSo.fitreoarSaaeenniiwca-.j^. .* ''. ' . *
P*e 1of14
, .
A j o y h $ S v !' ;
Centre Analytical Laboratories, Inc. . - ............... < - ............. Page 16 of 43
Centre Analytical Laboratories, Inc.
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Page 69 of 102
Centre StudyNo.: 023-041
-Centr ProtocolNo;-0eP=Q23*e<tt-
t* rnesesampiegetraets are anaiyzeaiouowingmemoaiiia-a-a.i or other a p p r a t i a t e '
fflCthrvi
'V't'-V^
: - *; -* < :
LD P g fw fn ffla
' '' V-
\ V"-'. 'if-. .
3.1 PFOS: pekluoroocoeulfonStefanionofpotassium salt) C^F17SOj' - ; '
3 J PFO S^perflooioot^esulfonyLm deCiF^O jN H j'.f y
33 PF0SA *rtfe^w *Jbri!^
*
. . ....
'
3.7 Surrogate standard: lH-lH-2H-2Hpaiuorooctane s kf omcaci d: ^. : ' . ' . .
4.0 W AH M ff^ Awp t^ im o ws 4.1 Health and'rafety warnings . 4.1.1 " UseimirnmlnmeanriniH/esneciallvlaboratory-coats, mm
;
5.1 There areno interferences known at this time.
6.0 EoriffMgwT 'T 6.1
acceptable.
6.1.1
Vortex mixer, VWR, Vortex Genie 2 .
'
o
' 6.1.2 Centrifuge,Mistral IQ00 or IEC ' .'
' 6.1.3 Shaker, Eberbach or V W R ..
.
.. . :
Centre Analytical Laboratori, Inc. Centre Analytical Laboratories, Inc,
' Page 17 of 43 Page 70 o f 102
Centre StudyNo.: 023-041
(^ntre-ProtoeeWo,-G0P-023-044--
... .
Oxford Dispci a e ^ j ^ lO-OniL '^ a ...:-, >'..: ..
73 Syringes,'capableo fnmasuringd jiL to50 jtLV:?j S?- ..." ' ' .
.7-10 . Graduatedp ip e te ^ .'K ^ ;.' 4 - - V f
: " " , "V
7.11 Syringes, disponbleplasric.^cc'jjW -. :; . >tS'-
. ' :
S-odium h*y:dii,a>dse*(Nria^0H-'),J.T B akeror equival.e'vn.~tyTL*v/ *1->4 &r;: : J
TctrabutyIaminommiihydrogen sul&tc(TBA), Kodak or equivalent & ' ' A
\ ;
;; *
Sodiumcarbonate(Na,CO)j; J.T.Bakerorequivalent :
Sodium bicaiitonr ^ a H C ^ ,
equivalent ; .
\'
Methy-T-Butl Ether.OWim- iiso-W.Wf-gtt.asVs:d'.*istilleid-orHPLTC 'gr*ad.e/' * . ,.r *-* *.:'\X*: *"
Methanol, Omoisolv, glass distilled orH PLC grade A/ . ' ..
Serum orblood, ihnen from su p p lie r'
...
Fluorochemical standards : .
.'
o 8^.1 PFOS (3M Specialty Chemical Division), molecularweight 538 8.9.2 PFOSA (3M Specialty Chemical Division), molecularweight 499
;v.->*/.;:V;V- : \-r-*.*.*fV*\v>'V...".'i.r'S..'i...';.*..,.H_..x_mJ_ca-'o"aETofTPWFO~-~S4J"f'iam.Swan"as;:''%\.'V. . *
.y..**
P a p J o f 14
CentreAnalytical Laboratories, Inc.
Page 18 of 43
Centre Analytical Laboratories, Inc.
Page 71 o f 102
/Cntre Study N o.: 023-041 Ciffi'Protocol No:0QP-Q234)4i------ ---------
Y', '? 8-103.1 TBXrjuire achtikpriom iachuse'toensirrcpH-T
..' ^.neededusi ng
^ '
8.11
1.10ppm EtFOSE-OH.)
8.11J- Weigh approximately 100 mg ofPF05 into a'100 mL v h m e t 'fla ^ d l^ r fi^ '
. the actual weight ; ..
.Vi .V.
'
8.11.4 Brin to'voltme with methanol
1
!
approximately SO ppm.
o 8-11.6 Dilute working standard 1with methanol for aworking standard 2 solution of
approx. 5.0 ppm. ;
. ' :v...
Centre Analytical Laboratories, be. Centre Analytical Laboratories, Inc,
. Page 19 of 43 Page 72 o f 102
Centre Study No.: 023-041 -- ...... CentreProtoeeHfe-QQP-023-641------ -
Centre Analytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
Page 20 of 43 Page 73 o f 102
Centre.Study No.: 023-041
CentreProtocol No. 00PO234541^---
. - l 1-t- Tw '^aliquot^othg'iq^ropri^ volne.iaveaa nialrixhl9rit-'. - . ;
- 'vi'*'.;-Typically usa t^sSiiidnl ntitraticiii'and.spiking amounts listed in Tbel '
.*< :; .Vit the end ofth u m tia n ^ l ik V in duplicte,twcwtandard carves, for a total of . . eighteenstandar3s,rt^m p blanla, aid two method blanks. - ...
Referto validation!---- S * * i i i m i i i i / '
"-
t; i...*
ETW-4.1
,j
(,
v-.`>v ^.'t.'-T
\
Exrncanioooa
ofPPFFOS
fro.m..Serua*m
.:
':
{_.. ..
** 1/* *!:
Centre Analytical Laboratories, he
Centre Analytical Laboratories, Inc.
Page6 of 14
$ 5 0 ^ ' '
Page 21 of 43
Page 74 o f 102
Cntre Study No.: 023-041
mm*
'CeatreProtocol Nor0oe^e3*04h
'y: ; Approximate spUcingafflounts for standards and spikes
$ r. v. t ^ t t i n g L m L^ matrix
'
^-.Working standard/,;:
' ^(Srrox. conc.)'^'* . - i i *
'*A!v>papnraolx'y.*tef*iVmna*ml'caotnr.iex,*o',,*f " :r,Vi B lank-/';,.
"k ^ r. OiSOOppm>tv. * . -V'vlOSCWi . 0.005 ppm
S iA 0.500 ppm 'V -..,i:.20
-- . -t 0.01O p p m =
S1VT5.00 ppm V I v 5 r'vi-
0.025 ppm .:>
SPA*,15,QQppm-.n*rv 3v.-,.'-I0 ,W;
0.050ppm:i : - ;
-': ; :ir5:00ppm f-V'-i
20 r-S.r,- ' .v.0.100 ppm
i.:s^ '5 0 .0 ppmv x.i
5 .-.-. A
0.250p p m . . .
>* '5 0 ,0 ppm A
10 V. -.- ` 0.500ppm .
SO.Oppm'.- . . ; 15 0.750ppm .
v 50.0 ppm 4&*~ V .'20- ,>. -1.00 ppm'.
)\ ;<**t
12.0 Puno^^ime
________ _______________ .
.
. ..
12.1 Obtain frozensamples and aliow^o th aififroom ianperature or in a lukewarm V ' '.
'
l i j . Vortex mix for 15 seconds, flien tranaaT.O mL orpther appropriatevnlnrn p a- ' ''*
1 -- 1 A e o n unused samplesti^fteezwaftCTartncbon^omitshave M a i lanovw l'Y.f'Si;' ' " .' 12.4 Record^thpinitial voiuinebn the e j * it io V w o ^ '^ ^ ^ V ./-^\ . '. .;S y '{ ' 12;S. LabeltbtiibewithUM'S^^^iuimb^.'samapIeD/dauan(lttiahtiiiltialsl S t t a h i ^ ' ^ " '*.. .
O
12 .10 To eachample, add 1 bicarbonatebuffer. - .**", . '
V ;*v ^ -
12.11 Using ffi Oxford Dispenser, add 5 m Lm e^l>tert-butyle^.''> :?''' . i
12.12 Cap eachsample and put on theshaker ata setting o f300 rpm, for20 minutes. " .' .'- , '
11.13 Centrifuge for 20 to 25 minutes at a setting o f3500 rpm,' oruntil layers are web ''
separated.-.-
' . '
'
:*4fV
Centre Analytical Laboratories, be. Centre Analytical Laboratories, Inc.
Page 22 of 43 Page 75 o f 102
Centre Study No.: 023-041 CentreProtocol NorOOP-023-041------
f *a '
mLof ytapaaraxconcepnanon-oi-wanuarumyfinfrf
.
- '''.-niT n f f TMT-nfgirimgggStaildardHHinitialmatrix volume(mT) ^
-i-. ,
rJ. - = `
.
. i.
.f '
. .
- -* ' '-**V'
..j
' *
...
.*
: for specificMDLand limit ofromtitatiori (LOQ) valncsfsce A ttachment B and O . V :
U*
# *"-- *.
4I guMMif n c M i
4 4Villi W u t l u i ^ h rtf f f l u m l M *
*
; J. :"?. :
/ r-i >*__' -p 4 ^ ;- $fc
1 Referto"sectiw5iofETS-8-5.rfem efl^'peifcnnanreeritaii..'
:
O
'l5.0 PoLLtmowPREvpmnwandWasteManagement---- ' ' ~ ; L/ . - '
15.1 Simpfe vmmH iipnwt in hiohamd container. flammable solventwaste dispngjvt . '
high BTU containers, and used jlasi pipettewastei disposed in broken glass containers ...
. located in thelaboratoiy. .
; v' y.l i
' ' : .- ' ,
Centre Analytical Laboratories, Inc.. Centre Analytical Laboratories, Inc.
Page 23 of 43 Page 76 o f 102
Centre Study No.: 023-041
Centre-Protocol-No.-Q0P-()23-Q44-
_
-FAl)l-M-3.1 f "Analysis ofScrum or w m * n.TM
.w ..
HPLC-Electrospray'iass Spectrometry" _ ... ;
im nr
VtA_A m c m p a c ii J im & f ^ ^ " '
' ' '
19.1. ETS-8-5.1, "Analysis ofSeram or OtferFluid Extracts for Fluorochemicais using
- ' ' i^HPLC-ElectrosprayMasSpe^metry" '^
. ;.
.
; ; ; 20.0 Rgvisiiws'J =
: v -~- '
;
- ..fflBfit
f v-r- 1
: % ; > ,. Revisioa
>: /. . Ppm,Pntg1nng1^ : .. . - r . -p
' -1
Section 1221 Changed toinclude sample orageat room tenerature. ' "04/02/99.
Section 12.13 Added th e '& a ta ^ e d '"!^ / % v ^ \ X
0
,< .
Centre Analytical Laboratories, Inc.
Centre Analytical Laboratories, Inc.
Page 24 of 43
Page 77 of 102
Centre Study No.: 023-041
Centre ProtbTOpNorOO?-Q23-Q4-l-
Is T:
Y*;.i4.**
.Study #
M in C-Bak#
. j-.,.. S a R o p t s S t d f n ->.
><:
/S.W W D iy
: v.;. #
F s
DateSjptked/Aaalyst ^ *<rf*V-5kl
v \ .'V J - :- A . fi'C C V
./w V -'iV /rV
. S-'. 'V '.is M S " -.' -- yf'.SJiY V.
^ 3 V i ' -. M SD <
T V .-.-.a- ..; , . . .
.pprotO Jfm : ichial
VY*. fi! `ft ?. `t
t t p / n iS r i a j
#v
/j a?**?' '' v-j>< - *.'vJh'A 'i T ^ '. tt;
if.: F C M x V .7 '^ C o n n a t p p ratJO p ^'
*etali*nom '# T t t -
.* ;V" ', ' v y
. ; V i ' 1
" - - .. . - ViVV - . . .
. . ..-. I- .
*
.
'Vol.*'' 'A-:--.
.t^e^nr'x *
- v - ' -- . -V v ,;- /V .
^ ' v p a - y ^ y .- r 5 :4 r-^.y f v.
-- ''V
' - - 'l - J B T V 4
. -? v . L
<*-; S .-4 ---
..
< .- -V
_ V
-
.-- \ a r
-
: V -V * /. * \ ; , W
*.' . ', - - \
** V
.r.V V V W
7 v h r-r~
> -.stjC'-
v .
' 4- ' -U v .*
* t H * -
-, ;
-w'-. t..
^ --i
.V'J"
."V*i '4 ?.`fV.V t
\^
1. **-*'v . .. /S * t.'
. V . -C ..-. -
'.. r- W -, . . .* '<
- ...
'.'ft'* - .... `7 * *it .
* / r:
; -;
: - .
'** .
"V ~
*.v> . - e ^ s r
v , ;,* <*
.. \
%*. *. V.7*V'L Vi *. '
*... . ' .* / -
.
V - r f '- 'J ii.- .
.W -:
.
X ; ..v .' 7
. .
.
v` - i
- : .- .. 'O P '- ^ W > ' V ! . ' V - . - a j -
. .........
. v ' - s t s s r r . - r K - S - . - ' * .t .. T. '
.. >
.-- . . i-.
`NS j h ' - a
W-- >
moant^oay- ~
Ji n iM E x tru ltM Method -
1
V o rm isI ... .
r r r r v'< r.-v .> : - : . . . . .
A ^7* ....
Pw IU M S S " - . . . . . erf:. <A^ *v*- Vohime
^ y r n L 'n ..
T-. . . ...
. r i p a t a i a L o T U M TB *-nH 10. r f t - . :'.. -
025n p c o o l w i . o r NCCh/0_UM NaHCOt h d ftr - :
M i * .. - Std.# - j --**- w * *':*!'*' ' M,'T
--w
' " - f l " -i-.
.
*<.# .TN*A- ***'
t J - . X .
Shaltcraced; -
viCoioiftm w M d: \ .
io o r-
.m n w w d m |.i|iaM| Bf MoMie |w g
. . * .-v : *!
: -A
' .
rwaoNiBooaiEwBoninnB drrnet*
Add mcthiaol
. v *
' mL
T em ocnnre
. TN-A ** ' \ - r
.y
_Yortex30ce.
-
. s ' Coat CaLVmfcatioas usedsamematrixts forstd carve.
v j . . , , **. * . *
Centre Analytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
Page 25 of 43 Page 78 o f 102
Centre Study No.: 023-041
'"" CentreProtocol No.D0P-023-044--
Compound ; M D L *: LO Q . LineTar C.a.lVibra/tion*Ra'ng.e (LCR.) - /. ..- ... * ~
v!5pp> ) | (PPb)' Approxunateconcentratioiu to be nsed for preparing the
Standard Calibration Curve '''.i'.'//..]
.,
PF,OS
1.74 v 5.55? 5 ppb1000 p p b ' j . . . - ....' * ?... -.. . , . -,
PF.O SAX-b , U l ' v 4.79 ; 5 ppb -1000 ppb ~ > if.
.
PEOSAA3 - 3.46 ,v .20.5 ? 5ppb.-1000ppb
`
.
EtFOSElOH v ' t U i 36.2, 5ppbr 1000ppb `f :.f. M556^T-V<;1-6.03S 19.2 \ 5w *rlO O O ppb^.|-- ;
5 - . ' ,. .
PFOSEA':??ri l r^ .7 J S ; 18J2; 5ppb-1000ppb 'f"i..
.. -'.v . . . ., . .,.
tserum '
. ; . * .*
. 9jMI
^
: . Please seeLOQ Summary'inftMDrErtuSinETS
t - '
5.0-V-I forfartherinformation. ... *
o
i ' AtnehmiattB:MDL/LOQSangtuy ' ETSw-S-4i.1 .:(|
... . . . ......
Pha*gee1l1looffiU4
' -i W 1 * ?.:V."*- v /
V J*' IWiiitMefPFOSa-- S w `
:
*V J.'; '/ *.i !''* ^ *r,** `j1'-'V'
Centre Analytical Laboratories, Inc.
Page 26 of 43
Centre Analytical Laboratories, Inc.
Page 79 o f 102
wmmmmmmm
r
Centre Study No.: 023-041
"entre-Protocol-No.-00P-Q23*Q41..
. Preparedrange - "LCRfrom-; -% Recovery :-;.R SD >; j.;- Rabbit Serum w ofstandards is "^3'iaye S-- f Range":.':' ^.1.Range
fob)'(ng/mL) '
-V .
v?(ngAnL) '
- *V
FuflR&ge*-:; r^VMslm :? .^.S^97S* :tjo .\u X :-
`94
s i SM04 $ - ' -jO^
H iglicijrie/'l.
:i`b9.v7tJ--,S,'r7S*,.5 .';i'5-06-w::. i *.: .Sij& iiss-
Vf
Preparedtarige.^ ..LCRfrom;. % Recovery RabbitSerum : "ofstavSsJ f J.^i'eutve.jw .a 'R ange :
:(ppb) (ngkaL)'"
IfeaK
..R S D .-r Range
Full.Range '' Low Curie.i ',*
/ j M ; : i*
mm*S.n-lW ;ff
High curve -: . 244- 97 ' ;'.j
Jmff -.t* Mpv-: ijja*'*-b ij'i
.>* *
.*. *4.'
i K . v ; y ; ftsn -im f
^
2^
/S.'lb-M /
'-* v' ;
it*
Compound: PFOSAA- V Preparedrange
Rabbit Sum .'.ofstandards ; (ppb) (ngftnL)
..-.LaCcRufr'rvoems.;;-;* .' (ng/mL) i
^Recovery
Full Range ' ! ,0991-974 ; .'*>24.7 -fj74`?'
Low Curve V
^"9.74V 247 v
S l-lll ;'iV
:-4 .is-i^ p
i-V 'V
High curve ' '.^ -9 7 4 77 : . 9 M - n i? y f S -in v ?
1/X w i 0991-974 ; .' 74 - 974/;;
:-.4.n-23i4J^>*;
o
Centre Analytical Laboratories, Inc.
Centre Analytical Laboratories, Inc.
Page 27 of 43 Page 80 o f 102
Centre Study No.: 023-041
-Gentre-ProteeGlt'Or0P-023*0AL.
^Preparedrange v :>LCRfiom'-: % Recovery ^ R SD .,.-
;RabbitSerum S ' ofstandards.
\? Range, 2-:Range
!
i
%
S
8
ffe ftg fa )
f?(p b )^ (tig/mL) -f
1 j-.y. ':;:...V...
't**** "r '*//
:
t
MVl Lg*&.&1 7? : ***'j
?i?,*4#-9i'-.a9.7 ii 77410 . ; X i j S j . ' :
.<97407
MV .1..-.2*.J%X-
K g h p S ^ lj $W M M
$ * $ & & ? /j!W409 ..: i'njPu.s.yj
`y '.-it. i/ x - y z g f
% $ jf- 9 7 S ? ^ 4 1 1 T;* .'ii.i-2n :-
' __
* .-.'a Compottlidi P
F
Q
^ S
E
^
g
ijA
^
V-,.*f.vv:"*',.*.
%'
*
s'i
.Prepared range- LCRfiom %RecoTery - RSD .
Rtbbit.S7.c^zuxn
ofstancards^ (ppb) .(ngftE U
Range
Y/*rRange*l1
-7 '- < * `*.*2 1 ^ ^^ng/mL)7;,`
Y.-v.V'fji-:
FuHRange 'vf ^ 0 5 9 3 -9 7 ^
/ ' K lO i - ' C*0*.1-1' S.2*-.;
Low Curve ;i ;^f4J3 - 248 9 .7 j@ il
. Highjanve '2 .4 9 3 -7 7 6 ??:
iA0.9>4 ~;.
i/x
.7^0993-9 W ^ i m i - M t 6<_t
aa.iv ...i..i.?
Compound:M556 >;
.Prepared range.- ,'LCRroni ;%Reeyery, 1-RSD
Rabbit Scran ..Yofitandards^-
i-R an g e^H
(ppb) (ng/mL); y% fe>b)^-
(ng/mL) *
' ' s tte t
FuflRange, . 0993 791` 7724.S-976. '.."8406 . -. '`-4.4-2--W17.9' .
Low Curve';'
9.76 - 974 ~ ioo-m j :; 'i9 5 - ii2 ^ ;
High curve '..;
97.6 - 97 ' Sl-H l.' ' ; : 5.il:?;74 ?
i ;;> v - `0993 - 976 ir- : 9.76- 97 ` :. .97410.
4.77-199 '
->:.* i
o
Centre Analytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
Page 28 of 43 ' Page 81 o f 102
Centre Study No.: 023-041
-kntfe-PretocolNo.-00EJ)23Q4]___
w .ttQ S A M S FOSAAv EtFOSE PFOSEA V M S. taf.AM
AO "
SldcooeJ SUoocTr. StdcoaS\Sld cno'-J.Sld concvx' Sslcac' o Anft ' 7,..' fin i n i
niftnL ':
-"^ugAnL^i. ttsAnL ' pjtedniL 'mL V.
0300 0300. 5.00
0307 > 0332,2* 3*0301 * r 0.521 0301
0.010 : - 1.015 ~
0307 0332tf MOSOIS J.0321.-. 0301 . 0.020 ' 1X25 '
5 .0 7 . - 5J2-S 3575.01. t '*521.;:. .- 52)1 . 0.005 . 1.010 `
5.00.. 5.07 5J2 3 5.01 .>521 . 5.01 .0X10
1.015 :
5.00. . 5X7. 532 V.Ol.' M & * ' . 5.01 0.020 - .- L025 50.0 : 50.1 . 532 . - 50.1; jt V S2.1-' y. 50.1 . 0.005 . 1.010
50.0 1 50.1 . 532 . 50.11* *`;52.\ .?, k--50.1... 0.010 - ' 1X15 .
. 50.0- .50.1 532 :S 0 .i ' ei52.1J -> 50.1. >0X15 i- . 1.020 V
50.0 50.1
532 .- 50.1 T x m bt "v 50,1 -, 0.020 . 1X25 ; '
Calculated concentration ofstandard! in thesamplejnatrix
*'' '
rros rrosA . rrosAA r ^ W Q S t p _ ? O S E A y ^ M S S {J r. Serrent
'fi "V `
Finiconc ' Fhdcne. Finalcane' Finalcooc.<;Tmai
^coDc/f Anrtipikid
safari. safari. .' * l/mL-"-r. tcAnL'X*'ni/niL ng/nL~j 7nt/m L ' v*fl.VtL
3 100/ 524..'. *454w 5.01 .* *5.131 s ion- 0X05 t'i.;:'.
.724. '
9J9 25.1 .
10.4 *2J
9.7 *. . 243 .'
: 9.93 v25J
.< \
.VlOa.'v 25X
J d S r:-
'Stthrente
*. . '*"v'1 -:/ ' ..
J 50.0 > 52.4 - 49.4 . 50.1 . . 5U Finaleonep
./." * '
97. - 919 . - 104 : 97J V v99J V 102 .1Bfa
*l^**#- ' .
24 1 251 . 493 500 . . 735 ' .'146
.. 263 524 7t2
-24 -.49
-
252 V .501 v v 74
. 23 ;i 500 y 513 7 .*
'-f;? .-r JsjLv-.'
975." " t a
103 97 993 1017 /y--*
. J . -
.. jy". .. . ` ''. V'>*1.* *:
Validated rant- appro
concentration ^ ' ? -TVv.,it- .*.-r-r-./ w
V
Serra
a. pros rrosA ... PFOSAA . ; EtTOSMH rrosx . VOMMi--
Rjbbil .. Boom
/5X0*1000 1. 5X0-1000 1-. 5.00-1000 !1 5X0-1000 . Enemaonly. Uaetahwfocrabbit -.V -.*
1
5.00-1000 ,,1" 3X0-1000 ' v..-. .
Rat . Esntaoty. Us*loesforrabbit -. i . ' i . Jt,
taonitey&Planm Estamaonly. Usevalue*forrabbit
- * ... * s&
Human Estmesonly. Usevaluesforrabbit
CentrAnalytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
; .i Page 29 of 43
Page 82 o f 102
Centre Study No.: 023-041 CentreProtocol-No. 0QP-02-3-041-------!-----
l.l Scope: This method is.................... other fluorochemical confounds from1liver.
1.2 AppUcableCompounds: Fluorocbezziical surfactants or otherihionnated compounds.
I CentrAnalytical Laboratories, Inc.
Centre Analytical Laboratories, Inc,
Page 30 of 43 Page 83 o f 102
Centre Study No.: 023-041 entre*etoe^-No.-00P-023-04J.
..
. . . ___ _ .__.... .
' ;*V<.-i*&-:'>?-.*'v''/w ... ... -
3.5 PFOSEA: perfluororfsmsuifopyl etfiylamde'c ^ O ^ fOE jC H jJH :>; :
;,-3; 6
'3.7 . Surrogate standard: lH-lH-2H-2Hpcrluorooctanc^il&w^^cid
]
V , - *' _]' '
M WIM^AIWOBTIM^'- : ~X ' ' '"4.1 Healt.h a..n.d^Safe<ty^W atir*nin"gs:*! -,
?^ -I' .
.5.1 There are no nterferences known at this time. :-
M Bouiwhent' ' "
"
5.1 The following eqmpment is'used whileperforming this method. Equivalent
acceptable.
'
" ,,
. 5.1.1 Ultra-TurmT25 Grinder forgrinding versamples ; . ` .
o 1.2 Vortex mixer, VWR, Vertex Genie 2 .
' '5.1.3 Centrifuge, Mistral lOOOorOEC
.
... . -
5 .1 .4 Shaker,EberbachorVWR .
''
:
'
y^ ' ' . * *B.
ETS4 4 .0
i ; V ". Page2ofM '
.., ' v.*j fj*ExtnctioDofFFOSomLiver; ?*vt ' V c . j . '
^ t .*-r ' *1/,'.,.|J.b
t* ^. . .. sf
J *i ..* ."*-*" -,^
Centre Analytical Laboratories, Inc.
Page 31 of 43
Centre Analytical Laboratories, Inc. .
Page 84 o f 102
Centre Study No.: 023-041 -Ointra Protocol No fjQPjm -nai
- 7.5 ; 7.6 7.7 7.8
' 7 5 Labels'. .
7.10 OuxuforraduDispisepnesonrs-JiU^^KM>lUjOjUjrjmi -77..1111 SSywrMingMesf, .e.n.a.bLIle. oiifmeasunngSC pm TLfto. 5Cl0 7.12 Graduatedpipettes -
7.13 Syringes,disposabIepIastic,'3cc
. - . v . X ' .
~s -,
'('i & & r - Z -
y - V-:-}''. '. >.,
O
8.7 Methanol,OmnisoIv.'giassdistilJedorIffLCgrade
8.S Liver,frozenfiomsupplier' ' 8.9 Dry ice 6 om supplier
r -
; - V ; * - / 'w v ' V ` "t .
8 .10 Flnorochemicalstandards
' 8.10 .1 PFOS (3M Specially Chemical Division), molecularweight 538
'
V.. . .
-.Vi.'v;`
ETS4-8.0
.VX
BaactiMOfrFOStiom'Vr'^,4-*~;^'>
-
Pi
.
jii-3.o
f
H
* .* ; V ,:V
i' /.'
1 f.
Centre Analytical Laboratories, Inc.
Page 32 of 43
Centre Analytical Laboratories, Inc.
Page 85 o f 102
Centre Study No.: 023-041
. - -------- ------ :------------------------- -------------- CentreProtocol-Norl)0P-Q23-041.
( . v.^
^^ ; 8. J . 1 - B requires fc h ^ p n o rt^ iin se t^ g ssn re pH - 10. Adjust as *
' needed using 1 N NaOH soliibon.',
V-
'
V ir. . S . ''S-'V" v-:i*w jT o '1
-.-!i'\W r ^^y26Jg fsp iu acait(M iate (N C O Jan d 21.0go fsod iu m
McafoaidetNaHOCO jeta* 1 L volumetric flask andbripg'tb.yolmewith vtni;.-'
QI% ^ e r :A`Starrre>ina lLN:aigoe e- bottle!.'-. ' -. . .S7.-JH:.' Vy .. .' - ;
Standardspreparation.'^.> : :`j *''}
\
-jtp-h, '
'
^ P r e p a r e PFOS. staidaids fbr the standard curve.
^\ *
'
othoflucrochermcalstandards,asqjpropriaie.'Mnltcoiiponir ;\ `
. y.m%fluorochcmical standards are acceptable (for example, one working sta n d a^ *^ -.
j i j ^ l .1 0 p p^m Et .F..OS E -*OryH . )` '
v-
>.113 Weigh approximately 100 mg o fPFOS into a 100 m L volumetric flask and record.' ,^I`>
V; , ' t h e K t u a l we i g h t . ' `V-
~ ' ' .
'. v . S g V * ''" '
i.114 Bring to volume with methanol for a stock standard ofapproximately 100ppm^ ,
' ... (jig/mL). . . . .
'
.115 Dilute the stock solution with methanol tor a working standard 1 solution of - `
;!. approximately 50ppm. . :
.
/ ;
.
ETS-W.0
Page4 of 14
ytical Laboratories, Inc.
Page 33 of 43
Centre Analytical Laboratories, Inc.
Page 86 o f 102
Centre Study No.: 023-041 Canre1PrecoctdNo.-e0PG234)4---------- --
* s . * , ;--- ^-- *~x
><>>' *
' ;. the accuracyoftheextraction/vi. V '*
.* , `"(r-- --r**^ to dotCQQlQC * "
Centre Analytical Laboratories, he. Centre Analytical Laboratories, Inc,
Page 34 of 43' Page 87 o f 102
Centre Study No.: 023-041
I
Centre-Protocol No.-00P-023-04-l------- - -U- -
O'
.JV \ '? / '
-EIS4-6.0 V-/ -;. -f': .'.. --yExttaoiooofPFOS b a a tin c V.,'
Centre Analytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
'I '^ eio ru .
' ^"Cv'T -V .
r-.
Page 35 of 43
Page 88 o f 102
Centre Study No.: 023-41
- Centre Protocol No:-00P-023-04-1-
- ApproximateSpikingAmontsforCalibrationStandards#
^Working Standard :-
ph: i: Approx.^finalcore, of
83-.(Approx. Cone.) *-
frTPFSinW^'
v':V i- \*!>'*.* .i-'-v.-BIank-X'^./
S ^fiiSOppjntv,.- v2vV.v Jsi.v 0.005ppm i"
5^(150O\^t? . ' 4 *7-; /.'iu' 0.010ppm v
vV:'04fttSn A
10 .:0.025ppm . .
. - !.,0i0'nDw^r>; -. : 20
0.050ppm.
-ji 0.50ffljn'gp*? V*.40.*.-t. 0.100ppm : '': "5.0ppm-Sr,- a =:.-io --y '' 0^50ppm
5.0pprit-.\^{ii / $20..-v ; . OJOOppm. .'
:'- 5.0ppm-.. ^ 3 0 ^ . -
0.750ppm - .
' SOppm'^^.:'';.' X'S7*Vk- KvV" 1.00ppm
................... !?:-.V...'* . . . ^
:.i : 13.0'PRorm inm ' - V.v;r
?? '?xir~': :i-.'
- *'
. ;.. - .
... ` . 1^.2 Cut approximately 1 g ofliver using * dissecting scalKl'ns partoftheprocedure is best ' "
. ' " . ' v. T' '". perforated quickly, not allowing the Ivct to
I r ' -.`t-.-
.. '. .
'. ' " 12 J Weighthe santole directly into"afaredplastic sanm ulevuE*^' '
. " ' . ' .
' '; : *3.6 ^ 4 5 m L * o f in te r to xa m p u le v ia L " ..
iV '^ v ^ .V - '
i:-y- :;V
:; 12.7 Grind thesample! Put thegrinderprobe in thesimple and grind foro u t 2tri3mke^jprV ' '
...... "'until thesample is homogeneous./'. . :j
. .
' . 12.8 Rujse^ejTObe'in to 'th is^ le w th 2J i k i `w ^ `u sm g a 'p $ ^ V l''^ :1! .
. . 12.9 Take thegrinderapart ariddean itwith methanol aftereach sample. Refer to A M D T -E P -^ ^ *^ *
-22.:
. v-;r . .
..t hr
a*'. Uw
12.10 Cap thesample and vertex for 15 seconds. Label thesampule vial with thestudy number, .''
' weight, liverID, dateand analyst initials.'' .. V
. ' '
O
- -V , --
v;
. ." I':V\
EIS44D . T-'.-r ' ., 'v..; ;:.:vir'.:Exiraeti.o-n-o-f;P-F--OSfci1mLiw.-7..;^',....... v. .1 iaP ' H M ,.
.
" ; *''i
.1 .-... v
- ..... v-. v.. -..'1* .'... . f
"...*>' f o i *
`.at
Centre Analytical Laboratories, Inc.
Page 36 of 43
Centre Analytical Laboratories, Inc.
Page 89 o f 102
Centre Study No.: 023-041 ~Centre!ProtocoWorOQP-923-04-1---
O
15.1 Sample wasteis disposed in biohazard containers, flammable solvent waste ;sVt;^ T'cj ^
high BTU containers, and used glass pipettewaste is disposed in broken'glass containers '
located in thelaboratory.; ' , '
r
T 'l .
'-
Centre Analytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
Page 37 of 43
Page 90 o f ioi
*
r
Centre Study No.: 023-041
..
.
--1-- -----------------------;-- L--------- "Centro Pioiui ol-Nor0Q?-Q33-044-
reagentis atpH 10. If not, adjust accordingly. '
'" To eachsunpic^d?Km L o2m TBA and 2 m Lofthe0l25 M sodium caiV m , j :
*
-T ^-.
.
^ OxfordDispenser, add5 ml)mcthyU^-butyl ether.^; '. "
\
-
12.19ap achambleand put onflfshidcarat scttinzof30 rpm. r M m m i^ I '
.
. :'*
'.~?:;&`
;Vt\
>TV*V"i1` .*-.: i
..
( :.. 1^ vCcotnftje for'V20--to'W25 mwiniritcrKXsctting df35(Xlipm#"orimt layers arewwee"ll *epea. . .
as m 12.10. "..
p le on theanalvtical m tioffm 'evl^^^im C^drv. ^jproriirm>|y j `3 ' '.
..
to'cji CCTbi&^'tube^sing grduated^Mp^i^^ v ` -
:\ *
12J 5 V w tg mix for 30 seconda^.;;;;-'?
' - ! '' J ?--?.*.'. v .-':
^ f f .; '
2 6 A tta c h > f l ? n m wlftw m f l i f i l t f tw % 1 M 'n m W a n d tra n e fW H ia --
t . l * . '
~
--
ai uovui wiui inenuoy numoer, animai numoer ana gender, Sampletimepoint,
. m- '.oaax,'finalsolvent, extractiondatc,andanalyses)performing the'gttrSio -St -. \ . 1 ' !
12J 8 Cap'inditore extracts'atroom temperature or it 3 jpnanately4 *C until a iS r a s 'i^ K '
!
; u iS a u ^ ^ j u ^
.. ; orindndemstuifybinder,aRirojiriate. " . '
":' "
>.' . .. >/ '
. - . ^ r " " v
o
Paie ( of 14
i Vv .
.Centre Analytical laboratories, Inc.
Page 38 of 43
Centre Analytical Laboratories, Inc.
Page 91 o f 102
Centre Study No.: 023-041
~! CentreProfocol No. OOP-023^041-
* :v *:V'.- v* -V/ V ' : v-: iT ^ A ^ ch m e a t B, MDL/LQ values od summary . v ; !i .** l;
%*. ^^a^tf^Cainwatjnstandardcalculationandconcmtrationr a ft
* ' .
i . *'
.
: . l&o
0 <; 'VV>~
; 18.1 The v^da^^jit'ssoaated with this method is ETS-8-i.O & 7.0-V-l.
' . 18.2 AMDT-EP-22,J^^S^amtenanceofUltra-TuiraxT-25" " .'?''
. lis
or Othff ic o n ic Huorodhanical Surfactants i&om
Liver for AnalysisUnng.HFC-Hectrospray/Mass Spectrometry^ -:. .
' -'
*
' lg.O
* * i . *...
'c.*;>.**-'. .> ; > . >.''A - ".*Lfrk'JrElj^fTfi^yrX* ^* ` :
=.;* J r . V.r^`` g2r
*. *
'
o.
E IS + tO .'
1 . . * Centre Analytical Laboratories, Inc.
Centre Analytical Laboratories, Inc,
.
'. .
1
......................
Page 39 of 43 '
Page 92 o f 102
Centre Study No.: 023-041 '"CentreProtocol No. 00P-Q23-Q41------- ---------
Centre Analytical Laboratories, Inc.
Page 93 o f 102
; Centre Study No.: 023-041
Centre Protocol No. 00P-O23-O41----
curves m each ofjthese matrices were extracted andanalyzed with the rabbit livercurves to "
detemune ^ v i c n ^ ^ R ^ S ^ iii'the rat.teyine, andmonkey liver'curves were equivalentto ">erabbitresponses; th&SorcijiieirM D L`and LOQ will be assumed to be equivalentto thnV. values as determined for thSrabbiciiier. .Ki.iv.:.5 ' * j V - . *.. -v.u ..
Refif
a ,.H.
.: Uvcr matrix-
. Prepared.'
nnge of.
standards Mi) fncfriL)
Rangeof lXQEfrb; Rangeof .-SytCKfirn-
average . fiayccgcyg: jfSnrstd ^ ft ||rij3 ij8 curve f .'.curvev .-EocunffesH
fppb) (W M^TOtfftLF Coot) (w jviiSjS5oi
; Ranged .ffiJWJtfinS^
i g k jd t$ |2 g g g ^ v- cave >. B (rntai)
Rabbit .19-1237 12-1200 W ^ m iS 'a W , 3 0 S ^ ? M
CorapoundtPFO SA *.
V* ;rf;**:- . . `
:: -v ' ' - i & i - - .
' Liver matrix
Prepared : . rangeof:*
standards-'
Raneeof f>;Ifcftomt* ..i*acvuerrvag*ee..M-4iifgrar*.v8eWcu`rv>evs;
R m geof^gfffS ^E : lo w ] td .': 5 ^ ^ ^
.Raageot : wcgL-finn?; highad " S S ir a i - i
.* cove
B carre
(poi) (astat) ufttfofH (eali) M d )
^W iwW j 'g j& v ft5 y
Rabbit ." 419-1237 12-1200 li^ O O ^ : 12-300
Comnontid : PFOSA .-v 'fc '.'i -
1.
Liver matrix
Rabbit
Prepared range of: . standards' JmMieeMJ
416-1232
Rangeo f. " LCR-from.'- Raneeof v^LOkfiad*^
-avenge ^.ave curve..: lowml
;'eurre
carve -'cfenfveSw curve
frMfneW.) "(iwk)WwU- 0* froM)
-0*6)
-
12-1200 ;:^ 3 o|1 mo^ 30- 900 ^a-'6o.0rvi.9-s0..-0','t;--
;sL ,c
o
Attachment B:MDL/LOQ Value]
i ; ; ETS4-40
' . ; ; -! - . ExtractionofPFOSfromLiver
. .Page 12of1
Centre Analytical Laboratories., Inc. Centre Analytical Laboratories,.Inc.
Page 41 of 43 Page 94 o f 102
Centre Study No.; 023-041 CentreProtocol N6.00P-023-041-----
'.Prepared.'
^ *^Uver : m rieof
i5 S S S rmatrix V
v\
' standards . tab) fato.)
.Rangeof IXRfrom. .Range of ... LCRfiemT
average
curve .^.lowrad v !*" low itd^
V` curve C c m '.: (p>b) (t/inU'taS) (tthA.) tab) (nartnU bob)
Ss M r : -- Rabbif1?" ,A6.(7-123..5 31-900 .*31 Tm * ;/W A v Y / N ftiY !
* Compoc nd rp p oSE
-- liver ' matrix'
Rabbit
u fiN a M i. 'S S i f
CPP*)taw i.17
^V/RaavnergaegoefVA^r<atLvWOteilafryiwsewna>^t
. R asp of . :-LCR.om,
bw Jtd s S $ t 0
j ,
^ v
'^ S .ta g
f h
llO lS o adV ^sa
n ia
" s;
; - curve tab) (ae/mL)
caive:|j -tabMiiSSiLj-
4^earve ^
.Ti--M fagftnL)
NTA . v*t$0fes8 s * ; ? # * : * * *
Compomid :M 56 N
. Y ; V -:Y : j $ S ' : :
Prepared liver range o f. matrix! standards
tab).(ftnH
.-.Rangeof - 'LRrfffan--,1
evc" s y g g F ^ cnrvf ;
Y k > w rtd .> ji|V K ^ ^ h g b a d .& f i ^ - : ;
V curve .`'.eitfVfegil tab) (atbnU a'taiVfmrtliC!:
.
ve. ..S ic s^ S V ;
Rabbit 6.17-1235 31-1200 :.|p |p 00H
O .t
Attabom C: StandardCakahtiou
ETS-K.0
;.
,C. ExtractionofPFOS fromLiver
CentreAnalytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
fnUofu Page 42 of 43 Page 95 o f 102
Oapppp
Centre Study No'.: 023-041
Centre Protocol No. OOP-023-041! ; \J
"T ; / '/ '-O .. f'-Standard n n m b e n ''..'- ;*
^ P j i t t ic ' -.*
? C> - - Equipmentnumber:' ' - ;. . -/r Fiinnaili jolrveehkt and TON: !
.
r . .. '' oBilaanukiBivneirn/Iudaeunitiuliienn
Metiod/rerision: \ .
. *.- ;
* -*^ : v * " > ' * * * X. :
Calcnlated coacentraflons ofstandards In the sample.m a frix y \
PFOS7 FFOSA. Final J ivFinal ' one/.,- cane s
" ' X 539 -539
'125 29.9 " .29.9
PFOSAA Fini cow*
: 539 T O O , S3-
EtFOSE. PFOSEA '.Finals V Final ;
conc-V cow :
' tit V 539 539 125) 1Z0 S 3 . 2)3
M55d r` Rnn?
'
Stdcpnc
.am iate Stdcoac
cow ^rilgg ' ngfaL .
s'*
jCI m
539
4> ~ Too
125) / v . S 3 - ,'x ScrrvnSi
- An Ain't '
piked mL
0505
595 S9S 599
593 .593
593
120 120 V 120 120 120 120
*R%lta"lC**'
299 299 - 299 299 299 599 599 599 . 599 599 599 :
ojoo.-i. t e '
!98 . 898
.898
898
898
898
1198
5198
w n - 1198
1198
1198 |
i. N
\ ;
V *+ *
Rabbit Bovine
Rat ` Monkey
approximate concentrations
pros
FF05A
PFOSAA
5-1000a * I 5.1000ppb I 5-1000H*
M o ia hr. we rabbit valiuoecs.
^
Egnratq only, we rabbit vahia.
Eaw atg only, we rabbit valua.
I EffiSgOH I
I 5-1OOPppb [
" rOAA
5-1000m
' PFOSEA b-------r , , .
TM
Aittdm unt G Standard
ETS-8-4.0 Eanacdsa ofPFOS finn Liver
F*ge U of 14
CentreAnalytical Laboratories, Inc. Centre Analytical Laboratories, Inc.
Page 43 of 43 Page 96 o f 102
SEP. 6.2001 !1:39AM ENVIRONMENTAL LAB 2 3E 09
Centre StudyNo.: 023-041
NO. 5647 P.2
304* Koeirch Drive, State College, PA 16*01.
Phoaei (14)231.8032,Fiatatile! (814)23M253
PROTOCOL AMENDMENT
Page 1 of 1
Centre Study Number 023-041
Amendment Number 1 Effective Date: 12/18/00
Centre Protocol Number 00P-023-041
DESCRIPTION OF AMENDED SECTION 1. $ 9: Experimental Design
AMENDED TO 1. Change the last paragraph to read as follows:
Moisture determination will be performed on all liver samples according to AOAC Official Method 950.46 Section B. (a), with the following modifications: 1. The sample weight will be - 0.5 g. 2. The (faying time will be - 12-14 hours.
RATIONALE
1. At the request of the sponsor, moisture determinations will be done for every liver sample. Due to limited Sample size, the sample weight used will be -0.5 g instead of -2 g. The drying time required for such a small sample size does not have to be 16-18 hours, so the samples will be dried for -12-14 hours
IMPACT ON THE STUDY 1. No negadve impact on the study.
W-A f o / A
rincipaJhvestigator Signature
2p :
sentativeYStudy Director) Signature
Date
Centre QAU Review 4/4L /i4z(<r
__________________
February 12.1998/1
RECEIVED T IIC SEP. 6. 1 :32PM
Centre Analytical Laboratories, Inc.
Page 97 o f 102
SEP. 6.2001 11:39AM ENVIRONMENTAL LAB 2 3i 09
Centre StudyNo.: 023-041
HO. 5647 P. 5
Centre Analytical Lab o rato rie s, Inc.
JOSgRtstarch Drive. gate CoBeee.PA 1O0.
Phone: (814)2314032. Pacatati f14tt31-Un
PROTOCOL AMENDMENT
Amendment Number 2
Effective Date: 03/28/0j
Page 1 of 1
Centre StudyNumber. 023-041
Centre Protocol Number 00P-O23-Q41
DESCRIPTIONOP AMENDED SECTION
1. 1 8: SampleAnalysis (f) 2. i 9: ExperimentalDesign 3. i 8: LMS/MS System and Operating Conditions (Turbolbnspray)
AMENDEDTO
1. Change thelast sentence to read the following: Samples in which peaks aredetected at the corresponding analyte retention times hut areless thanthe lowest concentration of the calibration standards (0.0001 pg/mL) will be reported as
NQ (not quantifiable). 2. Add the following:
Thefollowing equations will be used for quantitation of the samples:
1. AnalyteFound(ng/mL) (peakarea - intercept) / slope
2. Anaiyte Found (pg/g)* /nalvte found Cne/mDx EVfmLl xDF.x EV (mLl x 1 ug AV(mL)x sample wL(g)x 1000 ng
WhereFV final volume, DF dilution factor, EV extraction volume. AV aliquot volume
3.Recovery(%) /anal, found fug/mP - *VIT*"alvte"etri fag/mLOx 1000 ng x 100 amount added (ng/mL) x 1 fig
3.Change the computerlisted to Dell UltraScanFI110
RATIQm S
1-3. Clarifyhow the samples are quantitated andreported and file instruments used to acquire
data.
IMPACT ONTHE STUDY 1-3. No negative impact on the study.
FmicipalWti>tator Signature
1
U>Q l Date
RECEIVED TIME SEP. 6 . 1 :32PM
Centre Analytical Laboratories, Inc.
Centre QAUReview__lho/rf
_ ,_
Febmary 12.1998/1
Page 98 o f 102
Centre StudyNo.: 023-041
Centre A nalytical L a b o ra to rie s, Inc.
3048 Research Drive, S tate College, PA 16801.
Phone: (814)231-8032, Facsim ile: (814)231-1253
Centre StudyNumber 023-041
PROTOCOL AMENDMENT Amendment Number 1
Date of Occurrence: 08/24/01
Page 1of 1
Centre Protocol Number OQP-023-041
DESCRIPTIONOF AMENDED SECTION
1. 3: SPONSOR
ED 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.
RATIONALE
1-2. By request of Rochelle Robideau (fax dated 08/24/01 included in correspondence section of die rawdata associated with this study)
IMPACTON THE STUDY
1-2. No negative impact on the study.
Jerong Facility Management Signature
Date
y ftASLMa
Incomliinnegr SQtfuiifdtays DH iiMreActtAoPr SCiigonn aa thuiPre* *
o\ Date
Centre QAU Review
^ I boIOt February 12,1998/1
Centre Analytical Laboratories, Inc.
Page 99 o f 102
SEP. 6.2001 11:40AM ENVIRONMENTAL LAB 2 3E 09
Centre Study No.: 023-041
HO. 5647 P. 8
Centre Analytical Laboratories, Inc.
3048 ResearchDrive, State College,? A 15*01.
Phone: (*14) 231-8032,Facsimile (814)23M253
PROTOCOL DEVIATION
Deviation Number 1
Date of Occurrence: (1) 12/21/00
Centre Study Number 023-041
ContraProtocol Number OOP-023-041
DESCRIPTIONOFDEVIATION
1. $ Amendment #1: Did notperformmoisture determination on the following sample* became there waano (ample available afterextraction:
Centre Sample ID 0007875 0007879 0007881 0007883 0007885 0007907 0007911 0007915 0007917
0007918
C lim tID
459 483 485 487 - 489 ' 9503 9515 9525 9528 9529
Centre Strnnle ID
0007920 0007921 0007922 0007924 0007928 0007937 0007941 0007942 ' 0007943 0007944
Client ID 9531 9533 9535 9541
95 9567 9580 9581 9582 9583
1. Protocol deviation issued. Recorded By/Date:
ACTIONS TAKEN I amendment letncd. SQPrevieion. etc...
^ S tS u -fla / S j \6jf^\
IM PA C T ON TH E STU D Y
1. N o results far the moisture determination will be available for these samples.
RECEIVED TIME SEP. 6 . l= 32P n
Centre Analytical Laboratories, Inc.
February IZ 1998/2
Page 100 o f 102
SEP. 6.2001 11 :40AM ENVIRONMENTAL LAB 2 3E 09
Centre Study No.: 023-041
NO. 5647 P. 7
Centre A nalytical Laboratories, Inc.
3048ResearchDrive,StateCoOegs,PA16801. Phone.'(814)2314032,Facsimile: (814)231-1253
PROTOCOL DEVIATION
. DeviationNumber 2
Date of Occurrence: (1) 12/21/00
Centre Study Number 023-041
CentreProtocol Number OOP-023-041
1 D ESC R IPTIO N OF D EV IA T IO N
1. Amendment#1: Weighed leu than- 0.5 g for the moisture determination on the following sample because it was all of the sample available afterextraction:
Csatr* Sample IP 0007913 0007867 0007880 0007927 0007936 0007938
nient TO
9520 451 484 9548 - 9564 - 9570
W eishtfri
0.34 0.30 0.33 0.32 0.26 0.27
1. Protocol deviation issued.
ACTIONS TAKEN !< m nibnem limed. SOP revision, ate..,
I
i
i
IM PACT O N TH E ST U D Y
I. No negative impactbecause moisture was still calculated and the results did not vary significantly fromthe moisture results for samples (hit weighed -0.5g.
i I
j
RECEIVED TIME SEP. 6 . 1=32PM
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Pebmsry I X 1M8/2
Page 101 o f 102
SEP. 6.2001 11 :40AM ENVIRONMENTAL LAB 2 3E 09
Centre Study No.: 023-041
NO. 5647 P. 6
304S Research Oltre, State College,PA 1SM1.
Phones (814) 331-8032, Facsimile: (814)231-1253
PROTOCOL DEVIATION Deviation Number 3
Date of Occurrence: (1) 12/21/00, (2) 12/21/00 and 3/12/01
. CentreStudyNumber 023-041
Centre Protocol Number OOP-023-041
DESCRIPTIONOFDEVIATION
1. $ Amendment #1: Did notperformmoitturedetermination on the following samples because there was no sample available afterextraction:
Centre SemnleTD 0007877 0007939 0011423
Client ID 481 9575 na
2. i Amendment#1: The oven temperaturefor the moisture determinations was 110C
ACTIONSTAKEN 1. .mendmi-n, l d - SOP risimi, etc...
1-2. Protocol deviation issupdr-N
Recorded By/Date:
A 3^Z2jfli
IMPACTONTHESTUDY
1. Noresults for the moisture determination will be available for these samples. 2. Nonegative impart
ly A u / CpAci^al^estigaior Signature
% In r Date
S t^ ic ^ /S p o n s o r R ^ r e C t S tature CALOAUReview
S /v ' '
M l. li* !* ,
RECEIVED TIME SEP. 6 . 1:32PM
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Page 102 o f 102
