Document aXKg3y4wV92xJJ0Xoj6goE6N
fiR. S.3.6 _ 1017
R -r-- n C::
m. CENTRAL SCIENCE LABORATORY
SCIENCE SERVING AGRICULTURE, FOOD AND THE ENVIRONMENT
( CONTRACT REPORT
STUDY NUMBER HT5602
PERFLUOROOCTANESULFONATE, POTASSIUM SALT (PFOS):
AN ACUTE ORAL TOXICITY STUDY ^ WITH THE HONEY BEE
MAFF
000002
TITLE PAGE
CSL Study Number HT5602
STUDY NUMBER: HT5602 PERFLUOROOCTANESULFONATE, POTASSIUM SALT (PFOS): AN ACUTE ORAL TOXICITY STUDY WITH THE HONEY BEE
REPORT AMENDMENT NO 1
EPPO GUIDELINE 170 OECD GUIDELINE 213
AUTHOR:
Paul Wilkins
STUDY INITIATION DATE: 11 September 2000
STUDY REPORT DATE: 23 March 2001
SUBMITTED TO:
3M Corporation Environmental Laboratory
935 Bush Avenue St Paul, Minnesota 55106
USA
Performing Laboratory:
Environmental R&D Team Environmental Biology Group
Central Science Laboratory Sand Hutton
York Y041 1LZ, UK
Environmental Laboratory Project Number U2723 Page 1 of 5
0C0003
CSL Study Number HT5602
CENTRAL SCIENCE LABORATORY
Study title
Test facility Study number Number of pages:
CONFIDENTIAL REPORT AMENDMENT 1 Perfluorooctanesulfonate, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee Environmental R&D Team, CSL, Sand Hutton, York, UK HT5602 5
Signature /
Study Director P Wilkins Environmental R&D Team, CSL
Pat(dd/mm/yy)
Authorisation of release of report amendment
Signature_____ _______________
Date
Management Representative M H Bew Head Environmental Biology Group, CSL
(dd/mm/yy)
This amendment has been noted by Quality Assurance as correction to the report
Signature.a
Mrs C V Walker QA Auditor Central Science Laboratory
Date a s | c&jol(dd/mm/yyi
Environmental Laboratory Project Number U2723 Page 2 of 5
000004
CSL Study Number HT5602
Reason for amendment: There was a typographical error on pages 7 and 8 of the final report where the Test Item had been mis-spelt as Perfuorooctanesulfonate, Potassium Salt (PFOS). This has been corrected to Perfluorooctanesulfonate, Potassium Salt (PFOS). A formatting error had occurred on page 7, which had obscured the length of exposure. This has been corrected.
Distribution
Sponsor's Representative Sponsor's Monitor Study Director CSL Management Representative QA Manager
Ms R Robideau Mr D Palmer Mr P Wilkins Mr M H Bew Mrs R M Brookes
2 copies 1 copy 1 copy 1 copy 1 copy
Environmental Laboratory Project Number U2723 Page 3 of 5
000005
CSL Study Number HT5602
SPONSOR:
SUMMARY 3M Corporation Environmental Laboratory 935 Bush Avenue St Paul, Minnesota 55106 USA
SPONSOR'S REPRESENTATIVE:
Ms Rochelle Robideau
LOCATION OF STUDY, RAW DATA AND A COPY OF THE FINAL REPORT:
Central Science Laboratory Sand Hutton, York, Y041 1LZ UK
1
CSL STUDY NUMBER:
HT5602
TEST ITEM:
Perfluorooctanesulfonate, Potassium Salt (PFOS)
STUDY:
Perfluorooctanesulfonate, Potassium Salt (PFOS):,An acute oral toxicity study with the honey bee
NOMINAL TEST CONCENTRATIONS: Negative control, Positive control, 4.78, 2.17, 0.991, 0.450 and 0.205 pg a.i./bee
TEST DATES:
Range test: 12-15 September 2000 Definitive test: 21-24 September 2000
LENGTH OF TEST: LENGTH OF EXPOSURE: TEST ORGANISM: SOURCE OF TEST ORGANISMS:
LIFE STAGE OF ORGANISMS:
72 hours
4 hours
HONEY BEE (Apis mellifera L.)
National Bee Unit CSL Sand Hutton, York Y041 1LZ
Adult
ORAL LDso (72 hrs):
STATISTICALLY DETERMINED ORAL NO OBSERVED EFFECT LEVEL (BASED ON MEAN INTAKE):
0.40 pg a.i./bee (95% CL 0.33 - 0.48 pg a.i./bee)
0.21 pg a.i./bee
Environmental Laboratory Project Number U2723 Page 4 of 5
000003
CSL Study Number HT5602
INTRODUCTION
This study was carried out by the Environmental R&D Team, Central Science Laboratory (CSL), for 3M Corporation at the CSL facility in Sand Hutton, York, UK. The tests were conducted from 12lh September to 24th September 2000. The protocol, protocol amendments, raw data, all notes to file and the final report associated with this study will be retained in the CSL GLP archives for a minimum of 20 years after submission of the final report. The test item will be disposed of by CSL within two months after issue of the final report.
OBJECTIVE
The objective of this study was to evaluate the acute oral toxicity of Perfluorooctanesulfonate, Potassium Salt (PFOS) administered to the honey bee (Apis mellifera).
EXPERIMENTAL DESIGN
The oral ingestion method with administration of the test item in aqueous sucrose solution allows for precise exposure over the 4-hour dosing period. This reflects one main potential route of exposure for bees and other non-target: insects: oral intake of contaminated food (pollen, nectar etc.). At the initiation of the test three batches of bees in groups of 10 bees at each dose level received a single dose of the test item dissolved in sucrose. This was removed after 4 hours and replaced with untreated sucrose.
To determine an approximate toxicity a range-finding test was performed. Four doses of the test item separated by a factor of ten were administered to groups of 30 young adult worker honey bees. Five geometrically spaced doses of the test item were then administered to groups of 30 young adult worker honey bees to define the LD50. The toxicity of the test item could not be accurately determined in that trial due to high levels of mortality. Therefore another test (second main test) was carried out with using a lower dose of test item (issued as a protocol amendment). Only the results of the range test and second main test (hereafter referred to as the definitive toxicity test) are reported here. All data are retained on the study file. At initiation of the test, each group of 10 bees was offered 200 pi of the test item in 50% w/v sucrose. After 4 hrs the test feed was removed and the dose taken by each group of bees calculated. Mortality and sub-lethal effects were assessed at 4 hrs then a further 24 and 48 hrs after removal of feeders. In the range test and definitive test an additional assessment was made at a further 72 hrs after removal of the test feeders. All doses of test item are reported as pg a.i. (active ingredient)/bee and have been adjusted for analysed content. At the end of the tests any remaining live bees were killed by freezing and incinerated.
Environmental Laboratory Project Number U2723 Page 5 of 5
OC0007
CSL Study Number HT5602
CENTRAL SCIENCE LABORATORY AMENDMENT TO PROTOCOL
Study title Test facility
Perfluorooctane Sulfonic Acid, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee
Environmental R&D Team, Environmental Biology Group Central Science Laboratory, Sand Hutton, York, Y 041 1LZ
Study number
HT5602
Amendment number: 2
Number of pages: 2
Study Director Paul Wilkins
Signature
Date
(dd/mm/yy)
Management Representative Medwin Bew
Signature
Date i v i - St (dd/mm/yy)
Environmental Laboratory Project Number U2723 page 1 of 2
000008
JJtnAk'L:-
CSL Study Number HT5602
CENTRAL SCIENCE LABORATORY PROTOCOL AMENDMENT
Amendment No 2 to Protocol HT5602
After clarification by the Sponsor, it was requested that the Test Item name be changed from Perfluorooctane Sulfonic Acid, Potassium Salt (PFOS) to Perfluorooctanesulfonate, Potassium Salt (PFOS) and consequently the study title changed from:
Perfluorooctane Sulfonic Acid, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee
to
Perfluorooctanesulfonate, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee.
This has no impact on the Study
Circulation list
Sponsor's Representative Sponsor's Monitor Study Director CSL Management Representative CSL, QA Unit
Ms R Robideau, 3M Mr D Palmer, Wildlife International Mr P Wilkins, CSL Environmental R&D M rM Bew , CSL Mrs R Brookes, CSL
wi
Environmental Laboratory Project Number U2723 page 2 of 2
000009
W ildlife International, Ltd.
ECOTOXICOLOGY & ANALYTICAL TESTING SERVICES
ANALYSIS FORPFOS IN AQUEOUS AND SUCROSE SOLUTIONS USED BY CENTRAL SCIENCE LABORATORIES FOR CONDUCTING HONEY BEE TESTING
Enclosed please find a data summary for our screen for PFOS in aqueous and sucrose solutions employed by Central Science Laboratory in the honeybee studies (Sample IDs HT5601/02 Water and HT5601/02 Sucrose). Included are the results table, calibration curve, and standard/sample ion chromatograms for the screens performed on our triple quadrupole LC/MS/MS system. As can be seen in the data, the samples were less than the limit of quantitation, 0.0500 mg a.i./L (0.000500 mg a.i./L low standard x sample dilution factor (100)). The screen was conducted on October 26, 2000. These results apply to solutions representative of those used in the following studies.
PERFLUOROOCTANESULFONATE, POTASSIUM SALT (PFOS): An Acute Contact Toxicity Study with the Honey Bec (CSL Study Number: HT5601) PERFLUOROOCTANESULFONATE, POTASSIUM SALT (PFOS): An Acute Oral Toxicity Study with the Honey Bee (CSL Study Number: HT5602)
Scientist
MANAGEMENT:
Director, Analytical Chemistry
8598 C om m erce Drive Easton, Maryland 21601 Tel. 410-822-8600 Fax 410-822-0632 E-mail: ecotox@wlldllfeintemational.com European O ffice: Bergkam pw eg 1 7231 CL W am sveld The Netherlands Tel. 31 (0) 575-573048 Fax 31 (0J575-574813 E-mail: aleopold@worldonline.nl
Page 1 o f4
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Page 2 of 4
MacQuan, version 1.6 Printed: Thu, Oct 26, 2000 12:58 Calibration Rie: Cal PFOS 102600 MD Path: Macintosh HD:API3000:PROJECT DATA:454C-101:454C101_102600 MD: Comments: PFOS: 454C-101: Method Development - Screening llsh flake, water, sucrose -{L.C/MS/MS-MRM)
PROS 4 9 9 .0 - > 9 9 .1 No Internal Standard W eighted (1/x)
Intercept = 3 2 3 .1 7 Slope = 8193.32 Correlation Coeff. = 0.99949
Area
c e r tifie d c o p y
OF THE ORIGINAL
p lf, R1WS "
i$2r o ? ,
" ,11 & .
000012
MacQuan, version 1.6
Page 3 of 4
Printed: Thu, Oct 26, 2000 12:59
Calibration Rie: Cal PFOS 102600 MD Path: Macintosh HD:API3000:PROJECT DATA:454C*101:454C101 ,,102600 MD:
Comments: PFOS: 454C-101: Method Development - Screening fish flake, water, sucrose -(LG/MS/MS-MRM)
PFOS_1
STD 0.500 ug a.UL
4675A -011D-6
4.98 in 1 period PROS No Internal Standard Use Area
1: 4.97 MRM, 298 scans
499.0->99.1
Noise Thres.
2.0
Quant Thres. Min. Width M ult Width Base. Width RT Win. (secs) Smooth Expected RT
1.0 12 10 40 10 2 3.70
Area
4 2 3 1 .5 2
Height
3 9 3 .9 0
Start Time End Tim e
3.50 4.17
Integration Width Retention Time
0.67 3.70
Integration Type
A - BB
Thu, Oct 26, 2000 10:34
intensity: 4000 cps
PFOS_2
STD 1.00 ug a.U L
4675A -011D-7
4.98 in 1 period PFOS No Internal Standard Use Area
1: 4.97 MRM, 298 scans
4 9 9 .0 -> 9 9 .1 Noise Thres.
2.0
Quant Thres. Min. Width
1.0 12
Mult. Width
10
Base. Width
40
RT Win. (secs) 1 0
Smooth
2
Expected RT
3.70
Area
8 4 2 1 .3 3
Height
7 4 6 .5 7
Start Time
3.47
End Time
4.14
Integration Width
0.67
Retention Time
3.69
Integration Type
A - BB
Thu, Oct 26, 2000 10:40
intensity: 4000 cps
CERTIFIED COPY OF THE ORIGINAL
A - - w 52
000013
MacQuan, version 1.6
Page 4 of 4
Printed: Thu. Oct 26, 2000 12:59
Calibration R e : Cal PFOS 102600 MD Path: Macintosh HD:API3000:PROJECT DATA:454C-101:4540101,,102600 MD:
Comments: PFOS: 454C-101: Method Development - Screening fish flake, water, sucrose -(LC/MS/MS-MRM)
PFOS_9 4 5 4 C -1 0 1 -
WATER
Thu, Oct 26. 2000 11:22
4.98 in 1 period
pros
No Internal Standard
Use Area
1: 4.97 MRM, 298 scans 4 9 9 .0 -> 9 9 .1
Noise Thres. Quant Thres.
2.0 1.0
Min. Width Mult. Width
12 10
Base. Width RT Win. (secs) Smooth Expected RT
40 10 2 3.70
Area
0.0
Height
0.0
Start Time
End Time
0.00 0.00
Integration Width
0.00
Retention Time Integration Type
0.00
intensity: 4000 cps
PFOSJIO 4 5 4 C -1 0 1 -
SUCROSE
Thu, Oct 26, 2000 11:28
4.98 in 1 period
pros
No Internal Standard
Use Area
1: 4.97 MRM, 298 scans
499.0->99.1 Noise Thres.
2.0
Quant Thres. Min. Width
1.5 12
M ult Width
10
Base. Width
40
RT Win. (secs) Smooth
10 2
Expected RT
3.70
Area
3 8 9 .0 6
Height
41.96
Start Time
3.50
End Time
3.77
integration Width
0.27
Retention Time
3.59
Integration Type
A* - BB
intensity: 4000 cps
CERTIFIED COPY OF THE ORIGINAL
iNliTlALS
Jz
KS HSV RECEIVED
C;A A U K T
000014
CSL Study Number HT5602 TITLE PAGE
<H
STUDY NUMBER: HT5602 PERFLUOROOCTANESULFONATE, POTASSIUM SALT (PFOS): AN ACUTE ORAL TOXICITY STUDY WITH THE HONEY BEE
REPORT
EPPO GUIDELINE 170 OECD GUIDELINE 213
AUTHOR:
Paul Wilkins
STUDY INITIATION DATE: 11 September 2000
STUDY REPORT DATE: 23 March 2001
SUBMITTED TO:
3M Corporation Environmental Laboratory
935 Bush Avenue St Paul, Minnesota 55106
USA
Performing Laboratory:
Environmental R&D Team Environmental Biology Group
Central Science Laboratory Sand Hutton
York Y041 1LZ, UK
Report No 1 of 6
Environmental Laboratory Project Number U2723 Page 1 of 25
000015
CSL Study Number HT5602
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT
Study number: HT5602
Report title: Perfluorooctanesulfonate, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee
I, the undersigned, declare that the objectives laid down in the protocol were achieved and that the data generated are valid. As stated in the protocol no samples of dosing solutions were collected for chemical analysis. The report fully and accurately reflects the procedures used and the raw data generated in the above study.
This study was conducted in accordance with the UK GLP Regulations SI 1999 (No. 3016). These regulations are in accordance with the Organisation for Economic Co-operation and Development (OECD) Principles of Good Laboratory Practice (1997 ENV/MC/CHEM (98) 17). It is my understanding that the UK and OECD regulations meet the following standard:
Japan Ministry of Agriculture, Forestry and Fisheries, 59 NohSan, Notification No. 3850, Agricultural Production Bureau, (Tokyo, 1984).
The test item was not characterized in accordance with full GLP compliance; however, the characterization was performed according to 3M Standard Operating Procedures and Methods, and all raw data are being maintained in the 3M archives. The Sponsor has stated that the test item is being recharacterized in accordance with GLP. No samples of the dosing solutions were collected for chemical analysis.
Signed: '
$ ______ Date:.
Mr P Wilkins Study Director Environmental R&D Team, CSL
a / _(dd/mm/yy)
Environmental Laboratory Project Number U2723 Page 2 of 25
000016
CSL Study Number HT5602 QUALITY ASSURANCE STATEMENTS Quality assurance inspections of this study were made on the following dates:
<&
Date Activity
08/09/00
Verification of study plan
Range test 11/09/00 12/09/00
14/09/00
Dispensing (test item and toxic reference) Preparation of dilutions Dosing bees Assessments Assessments
Main test 18/09/00
Dosing bees
20/09/00
Assessments
Definitive test
20/09/00
Dispensing (test item and toxic reference)
21/09/00
Preparation of dilutions
Dosing bees
Assessments
Weighing feeders
22/09/00
Assessments
Date reported 08/09/00
15/09/00 15/09/00
15/09/00
21/09/00 21/09/00
25/09/00 25/09/00
25/09/00
Signed O A cL.fr*....
Date..?^.(,?>?.<&..|.J.l (dd/mm/yy)
Mrs C V Walker QA Auditor Central Science Laboratory
This report has been audited by the Quality Assurance Unit of the Central Science Laboratory.
Signed.
.... .................................... D a te..^ .l..fe k |.fc .L . (dd/mm/yy)
Mrs C V Walker QA Auditor Central Science Laboratory
Environmental Laboratory Project Number U2723 Page 3 of 25
O O O tti-7
aft CSL Study Number HT5602
3ft REPORT APPROVAL
SPONSOR: 3M Corporation
i TITLE: Perfluorooctanesulfonate, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee
CSL STUDY NUMBER: HT5602
STUDY DIRECTOR:
- -.aft
DATE X h \ o Z \ Q \ __ (dd/mm/yy) Mr P Wilkins Environmental R&D Team, CSL
Approval for Report issue MANAGEMENT:
_________li/Lth ^ A________
DATE______2-3 * V Cr\______ (dd/mm/yy) Mr M H Bew Head Environmental Biology Group, CSL
Environmental Laboratory Project Number U2723 Page 4 of 25
000018
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CSL Study Number HT5602 TABLE OF CONTENTS
TITLE PAGE.............................................................................................................................................1
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT..................................................2
QUALITY ASSURANCE STATEMENTS............................................................................................. 3
REPORT APPROVAL.............................................................................................................................. 4
TABLE OF CONTENTS.......................................................................................................................... 5
SUMMARY.............................................................................................................................................. 7
INTRODUCTION..................................................................................................................................... 8
OBJECTIVE.............................................................................................................................................. 8
EXPERIMENTAL DESIGN..................................................................................................................... 8
MATERIALS AND METHODS.............................................................................................................. 9
Test item...................................................................................................................................................10
Toxic reference.........................................................................................................................................10
Conditions during the tests.......................................................................................................................10
Dose preparation.................................................................................................................................... 11
Dose administration............................................................................................................................... 11
Observations.............................................................................................................................................11
Analysis of data...............................................................
12
RESULTS AND DISCUSSION.............................................................................................................. 12
Range test.........................................................................
12
Definitive test...........................................................................................................................................12
CONCLUSIONS......................................................................................................................................13
Table 1. Oral LDsoof PFOS from definitive test data............................................................................ 13
Table 2. Oral toxicity of dimethoate toxic reference from definitive test data......................................13 Environmental Laboratory Project Number U2723
Page 5 of 25
OC0019
CSL Study Number HT5602
REFERENCES......................................................................................................................................... 13 PRINCIPAL WORKERS......................................................................................................................... 14 DISTRIBUTION OF REPORT............................................................................................................... 14 APPENDIX 1 (TABLES A1-A8)........................................................................................................... 15 DETAILED ORAL TOXICITY TEST RESULTS.................................................................................15 APPENDIX 2 ........................................................................................................................................... 23 INTERIM CERTIFICATE OF ANALYSIS FOR TEST SUBSTANCE...............................................23
Environmental Laboratory Project Number U2723 Page 6 of 25
000020
I
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SPONSOR:
SPONSOR'S REPRESENTATIVE: LOCATION OF STUDY, RAW DATA AND A COPY OF THE FINAL REPORT:
CSL Study Number HT5602
SUMMARY 3M Corporation Environmental Laboratory 935 Bush Avenue St Paul, Minnesota 55106 USA
Ms Rochelle Robideau
Central Science Laboratory Sand Hutton, York, Y041 1LZ UK
CSL STUDY NUMBER:
HT5602
TEST ITEM:
Perfuorooctanesulfonate, Potassium Salt (PFOS)
STUDY:
Perfuorooctanesulfonate, Potassium Salt (PFOS): An acute oral toxicity study with the honey bee
NOMINAL TEST CONCENTRATIONS: Negative control, Positive control, 4.78, 2.17, 0.991, 0.450 and 0.205 pg a.i./bee
TEST DATES:
Range test: 12-15 September 2000 Definitive test: 21-24 September 2000
LENGTH OF TEST:
72 hours
TEST ORGANISM: SOURCE OF TEST ORGANISMS:
LIFE STAGE OF ORGANISMS:
HONEY BEE (Apis mellifera L.)
National Bee Unit CSL Sand Hutton, York Y041 1LZ
Adult
ORAL LD50(72 hrs):
STATISTICALLY DETERMINED ORAL NO OBSERVED EFFECT LEVEL (BASED ON MEAN INTAKE):
0.40 pg a.i./bee (95% CL 0.33 - 0.48 pg a.i./bee)
0.21 pg a.i./bee
Page 7 of 25
000021
*
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CSL Study Number HT5602 INTRODUCTION
it ^
This study was carried out by the Environmental R&D Team, Central Science Laboratory (CSL), for 3M Corporation at the CSL facility in Sand Hutton, York, UK. The tests were conducted from 12th September to 24IhSeptember 2000. The protocol, protocol amendments, raw data, all notes to file and the final report associated with this study will be retained in the CSL GLP archives for a minimum of 20 years after submission of the final report. The test item will be disposed of by CSL within two months after issue of the final report.
OBJECTIVE
The objective of this study was to evaluate the acute oral toxicity of Perfuorooctanesulfonate, Potassium Salt (PFOS) administered to the honey bee (Apis mellifera).
EXPERIMENTAL DESIGN
The oral ingestion method with administration of the test item in aqueous sucrose solution allows for precise exposure over the 4-hour dosing period. This reflects one main potential route of exposure for bees and other non-target insects: oral intake of contaminated food (pollen, nectar etc.). At the initiation of the test three batches of bees in groups of 10 bees at each dose level received a single dose of the test item dissolved in sucrose. This was removed after 4 hours and replaced with untreated sucrose.
To determine an approximate toxicity a range-finding test was performed. Four doses of the test item separated by a factor of ten were administered to groups of 30 young adult worker honey bees. Five geometrically spaced doses of the test item were then administered to groups of 30 young adult
worker honey bees to define the LD50. The toxicity of the test item could not be accurately determined in that trial due to high levels of mortality. Therefore another test (second main test) was carried out with using a lower dose of test item (issued as a protocol amendment). Only the results of the range test and second main test (hereafter referred to as the definitive toxicity test) are reported
here. All data are retained on the study file. At initiation of the test, each group of 10 bees was offered 200 pi of the test item in 50% w/v sucrose. After 4 hrs the test feed was removed and the dose taken by each group of bees calculated. Mortality and sub-lethal effects were assessed at 4 hrs then a further 24 and 48 hrs after removal of feeders. In the range test and definitive test an additional assessment was made at a further 72 hrs after removal of the test feeders. All doses of test item are reported as pg a.i. (active ingredient)/bee and have been adjusted for analysed content. At the end of the tests any remaining live bees were killed by freezing and incinerated.
Environmental Laboratory Project Number U2723 Page 8 of 25
000022
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CSL Study Number HT5602
The nominal doses administered are shown below.
Range finding oral test dose levels
Dose group
PFOS
(pg a.iybee)
1 103
2 10.3
3 1.03
4 0.103
Control 1
50% w/v sucrose
containing 5% acetone
Control 2
50% w/v sucrose
Dimethoate (pg ai/bee)
0.60 0.30 0.15 0.075 50% w/v sucrose containing Triton X-100 N/A
Definitive test dose levels
Dose group
PFOS
(pg a.iTbee)
1 4.78
2 2.17
3 0.991
4 0.450
5 0.205
Control 1
50% w/v sucrose
containing 5% acetone
Control 2
50% w/v sucrose
Dimethoate (pg ai/bee)
0.60 0.30 0.15 0.075
-
50% w/v sucrose containing
Triton X-100 N/A
Estimates of LD50values were calculated for the main oral test. The no observed effect levels were determined by a statistical evaluation of the mortality data.
Negative controls, solvent controls (5% acetone) and positive control groups (dimethoate as BASF 40 formulation nominally containing 400 g/1) were maintained concurrently to check that the bees were reacting normally to the test item during the studies.
MATERIALS AND METHODS
The methods, species used and route of administration described in this protocol are based upon pro cedures specified in EPPO Guideline 170, Guideline on Test Methods fo r Evaluating the Side-Effects o f Plant Protection Products on Honey Bees; and the OECD Test Guideline 213, Honeybees, Acute Oral Toxicity Test. In order to control bias, bees were impartially distributed to treatment and control groups. No other potential sources of bias are expected to affect the results of the study. The study was considered valid with: a) control mortality of 10% or less; and b) the calculated dimethoate LD50 fell within the OECD standard, i.e. 0.10 - 0.35 pg ai/bee.
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CSL Study Number HT5602
Test item
The PFOS (IUPAC Name 1-Octanesulfonic acid, 1,1,2,2,3,3,4,4,5,5,6,6,7,7,8,8,8-heptadecafluoro-potassium salt, CAS Number 2795-39-3, white powder, lot 217) used in this study was supplied by Wildlife International Ltd. on behalf of 3M Corporation. Each sample of the material tested in this study was uniquely labelled with the Test Item number NBU105. The test item was stored, as recommended by the sponsor, at 16-20 C prior to use.
PFOS was dispersed in acetone (analytical grade) before use due to a low solubility in water (approximately 500 mg/1). A dispersion test was carried out, before the toxicity tests were performed, in which PFOS was dispersed in acetone at the approximate concentration required to deliver the highest doses. The homogeneity of the mixture was assessed after 2 hours. In this report "solution" is used to include material which may be suspended or dispersed. Homogeneity of such solutions of the PFOS were checked visually during dispersion tests before the day of the test and immediately before use. All solutions were re-mixed prior to use. Solutions of the test doses were homogenous for the purpose of administration. The test item formed a clear solution on mixing at the highest dose, after two hours there was a slight sediment at room temperature at a concentration of 86 |ig a.i./pl PFOS (the approximate concentration required to prepare the highest diluted dose in the test).
Toxic reference
Dimethoate toxic standard (NBU90), BASF 40 lot 37M 9020106 (a blue liquid EC formulation, 37.4% w/w ai, 400 g/1 nominal concentration), was purchased from UAP York on 14/06/00 and was stored at 0-11C (on one occasion the maximum temperature recorded was 14C - this was after the door to the refrigerator had been opened) suitable to maintain stability for 2 years according to information supplied by the manufacturer. Dose and LD50calculations are for corrected dimethoate concentration based on data supplied by BASF.
The honey bee (Apis mellifera) is useful in evaluating the potential hazards of agricultural chemicals to nontarget insects since it is an important pollinator of various agricultural crops. There is also a substantial data base on the effect of agrochemicals upon bees with which to categorize potential hazards.
Worker honey bees (Apis mellifera) were obtained from colonies belonging to the CSL National Bee Unit. Bees from the colony 32 were used throughout the tests. Bees were examined prior to the start of the test and shown to be free of acarie, nosema and amoeba. The colony from which the bees were taken had not been treated with a varroacide within the last 4 weeks. Worker bees were collected from the hive by using a small amount of smoke, gently shaking them from the combs and transferring them (40-50 per cage) into cylindrical mesh cages. In the laboratory the mesh cages were placed into the incubator (25 + 2C, 65 + 5% relative humidity) to starve the bees for 1.5 to 2 hrs before the test.
Conditions during the tests
Test bees were housed in test chambers which are clean, well ventilated, inverted petri dishes, measuring approximately 9 cm in diameter. A small inverted petri dish (approximately 3 ml) to contain the sucrose solution was affixed in each chamber. Each test chamber contained 10 worker bees and was identified by study number, dosage group and replicate. After dosing, test cages were kept in darkness at 25 2C and 65% 5% humidity, except during observations. Temperature and relative humidity within the environmental chamber were recorded continuously during the test.
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CSL Study Number HT5602
Dose preparation
Doses of the test item were made by preparing a solution of 103 pg a.i./pl (range-finding test) and
47.8 pg a.i./pl (definitive test) PFOS and making a series of dilutions from this. Doses were prepared
I
in acetone and then further diluted in 50% w/v sucrose for dosing. In the range test and definitive test 0.5 ml of each acetone solution was then diluted to 10 ml with 50% w/v sucrose giving a final acetone
concentration of 5%. Although this concentration of acetone was greater than recommended in the
OECD guidelines (1%) it was required to ensure the highest concentrations remained in solution for
the oral test doses. One set of control bees was treated with the same level of acetone in 50% w/v
sucrose (0.5 ml acetone in 10 ml sucrose) and a second set of control bees was treated with 50% w/v
sucrose.
Stock solutions of the positive control, dimethoate, were prepared in deionised water containing 1 g/1 Triton X-100. Stock solutions of 2.80 pg ai/pl (range-finding test) and 3.00 pg ai/pl (definitive test) dimethoate in deionised water containing 1 g/1 Triton X-100 were prepared and a series of dilutions made in 1 g/1 Triton X-100. This series of dilutions (0.5 ml) were then further diluted to 10 ml with 50% w/v sucrose. Control bees were dosed with 0.5 ml 1 g/1 Triton X-100 in deionised water diluted to 10 ml with 50% w/v sucrose. The tests were carried out in parallel with those for PFOS.
Samples of the sucrose solution used in the study were analysed by Wildlife International Ltd. and no PFOS background levels were found.
Dose administration
The bees were anaesthetised with carbon dioxide immediately before dosing and gently tipped out onto filter paper and counted into the petri dish cage (drones were discarded). Each group of 10 bees was offered 0.2 ml of a given concentration (or controls as above), the dose being measured into a small, pre-weighed, glass feeder within the cage using a variable volume pipette. This is equivalent to 20 pl/bee. At each dose for the test item and dimethoate and for each control there were 3 replicate cages of 10 bees. Doses were administered within 2 hours of preparation.
After 4 hours the glass feeders were removed and weighed and the sucrose feeders filled with approximately 3 ml 50% w/v aqueous sucrose so that bees had continuous access to sucrose for the remainder of the study. The dose consumed was determined by comparison of the weight of the dose remaining in the glass feeders with the weight of a known volume of the test solutions.
Observations
Mortality and any bees knocked down, i.e. alive but immobile, or stumbling were assessed when the feeders were removed and 24, 48 and 72 hours after removal of the test feeders (i.e. 4, 28, 52 and 76 hours after the start of the test).
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CSL Study Number HT5602
Analysis of data
Analysis of the data comprised plotting probit mortality recorded after 24, 48 and 72 hours against the logarithm of dose (CSL Probit 1 package). A least - squares regression (Finney 1971) was fitted to these. Toxicity was expressed as LD50 in pg a.i. per bee with the 95% confidence limits, NOEL and slope of the response curve. The NOELs (no observed effect levels) were estimated using Student's t-test (p<0.05).
The LD5o was used to classify the toxicity of the test item according to the ICBB (1985). The categories used were:
Highly toxic: Moderately toxic: Slightly toxic: Virtually non-toxic:
less than 1pg ai/bee 1-10 pg ai/bee 10-100 pg ai/bee greater than 100 pg ai/bee
Data for the dimethoate 24-, 48- and 72-hour mortality results were analysed using the CSL probit program (Probit 1, version 4).
RESULTS AND DISCUSSION
The results of the oral tests are summarised in Tables 1 and 2 and detailed results are listed in Appendix 1. Further details are given below. The oral toxicity of the test item resulted in a 72 hr LD50 value for the honeybee of 0.40 pg a.i./bee.
Ranee test
Mortality was observed at all doses from mean intake 1.0 pg a.i./bee with a steep dose response between a mean intake of 1.0 and 10 pg a.i./bee. Sublethal effects were observed as knockdown and stumbling at 4 hrs at the two highest dose levels with these individuals dying by 24 hrs.
Definitive test
There was significant mortality at all doses above a mean intake of 0.21 pg a.i./bee with a steep dose response between a mean intake of 0.45 and 2.2 pg a.i./bee. The 72 hr LD50 was calculated as 0.40 pg a.i./bee (95% confidence limits 0.33 - 0.48 pg a.i./bee). The NOEL was 0. 21 pg a.i./bee at 72 hours based on mean intake data (Student's t-test p <0.05). Sublethal effects were seen observed as knockdown at the highest dose level at 4 hrs with these individuals dying after 24 hours.
The 24, 48 and 72 -hour contact LD50 values for dimethoate in the definitive test are shown in Table 2. Mortality of bees exposed to the toxic reference in the tests allowed calculation of an oral LD50 of 0.12 pg ai dimethoate/bee at 24-hours and 0.11 pg ai dimethoate/bee at 48 and 72-hours. These results show a toxicity level within the ranges reported by the OECD guidelines showing the bees were reacting normally in this test.
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CSL Study Number HT5602
CONCLUSIONS
Exposure of honeybees to PFOS by oral dosing resulted in a steep dose response curve with significant mortality above a mean intake of 0.21 pg a.i./bee and a 72 hr LD50 calculated as 0.40 pg a.i./bee. The NOEL based on statistical analysis of the data (Student's t test p<0.05) was 0.21 pg a.i./bee at 72 hrs.
These results demonstrate the oral toxicity of PFOS as 0.40 pg a.i./bee and according to the ICBB (1985) is classified as highly toxic by ingestion.
Table 1. Oral LDsnof PFOS from definitive test data
Time Hrs 24
Oi/l Q -J
(pg a.iibee) 0.72
95% Confidence Interval for LD50
0.60 - 0.85
48 0.46 0.32 - 0.55
72 0.40 0.33-0.48
Estimate of slope of response line 5.0 4.7 4.8
Table 2. Oral toxicity of dimethoate toxic reference from definitive test data
Time Hrs
LD5o (pg a.iybee)
95% Confidence Interval for LDS0
Estimate of slope of response line
24
0.12 0 .1 0 -0 .1 4
6.4
48
0.11 0.096-0.13
7.3
72
0.11 0.096 - 0.13
8.1
REFERENCES
European and Mediterranean Plant Protection Organization. 1992. Guideline on Test Methods for Evaluating the Side-Effects o f Plant Protection Products on Honey Bees. EPPO Bulletin, 22, 203-215.
Finney, D. J. 1971. Statistical Methods in Biological Assay, 2nd ed., Griffin Press, London.
Internationa] Commission for Bee Botany (ICBB). 1985. Recommendations fo r harmonization o f methods fortesting hazard o f pesticides to honeybees. Third Symposium on harmonization of methods for testing hazard of pesticides to bees, England.
Organization for Economic Cooperation and Development. 1997. Guideline 213 Honeybees, Acute Oral Toxicity Test.
Environmental Laboratory Project Number U2723 Page 13 of 25
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CSL Study Number HT5602
PRINCIPAL WORKERS
STUDY MANAGEMENT
Study Director CSL Management Representative
Mr P Wilkins Mr M H Bew
OPERATIONAL SUPERVISORS
Principal Scientist Scientist & Test Item Controller
Scientist & Deputy Test Item Controller Scientist Scientist Scientist
Dr H M Thompson Miss U B Pietsch
Miss S C Green Mr A H Battersby Dr R J Spinks Mr M J Kelly
QUALITY ASSURANCE
QA Manager QA Auditor
Mrs R M Brookes Mrs C V Walker
DISTRIBUTION OF REPORT
Sponsor's Representative Sponsor's Monitor Study Director CSL Management Representative QA Manager
Ms R Robideau Mr D Palmer Mr P Wilkins Mr M H Bew Mrs R M Brookes
2 copies 1 copy 1 copy 1 copy 1 copy
Environmental Laboratory Project Number U2723
.
CSL Study Number HT5602
APPENDIX 1 (TABLES A1-A8) DETAILED ORAL TOXICITY TEST RESULTS
Table A l. Results of range-finding oral dosing tests with PFOS - Mortality
Nominal Actual dose2 dose1 (pg a.i./bee)
(jig a.iVbee)
Cage No.
Cumulative number dead (n=10)
4 hrs
24 hrs 48 hrs 72 hrs
103 96 46 5 10 10 10 * 103 94 47 5 10 10 10
103 101 48 7 10 10 10
1
10.3 10 49 0 10 10 10 10.3 10 50 0 10 10 10 10.3 10 51 0 10 10 10
1.03 1.0 52 0 0 2 3 1.03 1.0 53 0 1 1 2 1.03 1.0 54 0 0 0 4
1
] 0.103 0.10 55 0 0 0 0
J 0.103 0.10 56 0 0 0 0 0.103 0.10 57 0 0 0 0
O3 0 58 0 0 0 0 O3 0 59 0 0 0 0 ! O3 0 60 0 0 0 0
04 0 61 0 0
04 0 62 0 0
04
0
63 0
0
1based on nominal consumption of test solution of 20jd/bee based on amount of test solution consumed (average of 10 bees)
3acetone dosed 4undosed
0 0 0
0 0 0
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CSL Study Number HT5602
Table A2. Results of range-finding oral dosing tests with PFOS - Sub-lethal effects
Nominal Actual dose2 dose1 (pg a.i^bee)
(Hg a.i./bee)
Cage No.
Number knockdown (K) or stumbling (S)
4 hrs
24 hrs 48 hrs 72 hrs
103
96
46 4K
0
0
0
103 94 47 1S,4K 0 0 0
103 101 48 1S.2K 0 0 0
10.3
10
49 2K
0
0
0
10.3 10 50 1S,3K 0 0 0
10.3
10
51 2K
0
0
0
1.03 1.0 52 0 0 0 IK 1.03 1.0 53 0 0 0 0 1.03 1.0 54 0 0 IK 0
0.103 0.10 55 0 0 0 0 0.103 0.10 56 0 0 0 0 0.103 0.10 57 0 0 0 0
o3 0 58 0 0 0 0 o3 0 59 0 0 0 0 o3 0 60 0 0 0 0
04 0 61 0 0
04
0
62 0
0
o4
0
63 0
0
1based on nominal consumption of test solution of 20jil/bee based on amount of test solution consumed (average of 10 bees) acetone dosed
4undosed
0 0 0
0 0 0
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CSL Study Number HT5602
Table A3. Results of range-finding oral dosing tests with dimethoate - Mortality
Nominal Actual dose12
dose1
(gg a.iTbee)
(pg a.i7bee)
Cage No.
Cumulative number dead (n=10)
4 hrs
24 hrs
48 hrs
72 hrs
0.60 0.56 31 3 10 10 10 0.60 0.54 32 0 10 10 10 0.60 0.60 33 0 10 10 10
0.30 0.25 34 2 10 10 10 0.30 0.30 35 0 10 10 10 0.30 0.30 36 1 10 10 10
0.15 0.15 37 0 8 8 8 0.15 0.15 38 0 4 5 5 0.15 0.15 39 0 5 7 7
0.075 0.075 40 0 0 0 0
0.075
0.074
41
0
0
0
0
0.075
0.074
42
0
1
1
2
0 0 43 0 0 0 0 0 0 44 0 0 0 0 0 0 45 0 0 0 0
1based on nominal consumption of test solution of 20pl/bee 2based on amount of test solution consumed (average of 10 bees)
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CSL Study Number HT5602
Table A4. Results of range-finding oral dosing tests with dimethoate - Sub-lethal effect^
Nominal Actual dose2 dose1 (ug a.iibee)
(fig a.iibee)
Cage No.
Number knockdown (K) or stumbling (S)
4 hrs
24 hrs
48 hrs
72 hrs
0.60 0.56 31 2K 0 0 0 0.60 0.54 32 2K 0 0 0 0.60 0.60 33 7K 0 0 0
0.30 0.25 34 IK 0 0 0 0.30 0.30 35 0 0 0 0 0.30 0.30 36 0 0 0 0
0.15 0.15 37 0 0 0 0 0.15 0.15 38 0 0 0 0 0.15 0.15 39 0 0 0 0
0.075
0.075
40
0
0
0
0
0.075
0.074
41
0
0
0
0
0.075
0.074
42
0
0
0
0
0 0 43 0 0 0 0 0 0 44 0 0 0 0 0 0 45 0 0 0 0
1based on nominal consumption of test solution of 20pl/bee based on amount of test solution consumed (average of 10 bees)
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CSL Study Number HT5602
Table AS. Results of definitive oral dosing tests with PFOS - Mortality
Nominal Actual dose12 dose1 (pg a.iTbee)
(fig a.i^bee)
Cage No.
Cumulative number dead (n=10)
4 hrs
24 hrs
48 hrs
72 hrs
4.78 4.8 316 3 10 10 10 4.78 4.8 317 3 10 10 10 4.78 4.8 318 3 10 10 10
I
2.17 2.2 319 1 10 10 10 2.17 2.2 320 0 10 10 10 2.17 2.2 321 1 10 10 10
0.991 0.991 0.991
0.96 322 0 0.96 323 0 0.96 324 0
7 10 10 8 9 10 6 99
0.450 0.45 325 0
14
5
I--TI
0.450 0.45 326 0 3 7 7 0.450 0.45 327 0 2 4 6
--1t
0.205 0.21 328 0 0 0 1 0.205 0.21 329 0 0 0 0
I 0.205 0.21 330 0 0 2 2
O3 0 331 0 1 1 1 O3 0 332 0 0 0 0 O3 0 333 0 0 0 0
04 0 334 0 0 0 0 04 0 335 0 0 0 0 04 0 336 0 0 0 0
1based on nominal consumption of test solution of 20pl/bee based on amount of test solution consumed (average of 10 bees) acetone dosed
4undosed
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9 m
i*
j3
I*
\w
i *
1 I
--
I 1
I 3
CSL Study Number HT5602
Table A6. Results of definitive oral dosing tests with PFOS - Sub-lethal effects
Nominal Actual dose2 dose1 (fig a.iTbee)
(fig a.i./bee)
Cage No.
Number knockdown (K) or stumbling (S)
4 hrs
24 hrs
48 hrs
72 hrs
4.78 4.8 316 IK 0 0 0 4.78 4.8 317 IK 0 0 0 4.78 4.8 318 IK 0 0 0
2.17 2.2 319 0 0 0 0 2.17 2.2 320 0 0 0 0 2.17 2.2 321 0 0 0 0
0.991 0.96 322 0 0 0 0 0.991 0.96 323 0 0 0 0 0.991 0.96 324 0 0 0 0
0.450 0.450 0.450
0.45 325 0 0.45 326 0 0.45 327 0
IS 0 0 0 00 0 IK IK
0.205
0.21 328 0
0
0
0
0.205
0.21 329 0
0
0
0
0.205
0.21 330 0
0
0
0
03
0
331 0
0
0
0
O3 0 332 0 0 0 0
O3 0 333 0 0 0 0
O4
0
334 0
0
00
O4
0
335 0
0
00
O4
0
336 0
0
0
0
1 based on nominal consumption of test solution of 20fil/bee based on amount of test solution consumed (average of 10 bees)
3 acetone dosed 4 undosed
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CSL Study Number HT5602
Table A7. Results of definitive oral dosing tests with dimethoate - Mortality
Nominal Actual dose2 Cage dose1 (fig a.iybee) No.
L (fig a.iTbee)
i * Cumulative number dead (n=10)
4 hrs
24 hrs 48 hrs
72 hrs
0.60 0.57 301 8 10 10 10
0.60
0.58 302 2
10 10 10
0.60
0.60 303 5
10 10 10
L
0.30
0.30 304 4
10 10 10
0.30
0.30 305 2
10 10 10
I
0.30
0.30 306 5
10 10
10
0.15 0.15 307 1 9 10 10 0.15 0.15 308 0 7 9 9 0.15 0.15 309 2 6 6 7
0.075
0.074
310
0
2
2
2
0.075
0.075
311
0
1
1
1
0.075
0.074
312
0
1
1
1
0 0 313 0 0 0 1 0 0 314 0 0 0 0 0 0 315 0 1 1 1
1 based on nominal consumption of test solution of 20p.l/bee based on amount of test solution consumed (average of 10 bees)
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CSL Study Number HT5602
Table A8. Results of definitive oral dosing tests with dimethoate - Sub-lethal effects
Nominal Actual dose2 dose' (pg a.iJbee)
(pg a.i./bee)1
Cage No.
Number knockdown (K) or stumbling (S)
4 hrs
24 hrs
48 hrs
72 hrs
0.60
0.57 301 2K 0
0
0
0.60 0.58 302 6K 0 0 0
0.60
0.60 303 5K 0
0
0
0.30
0.30 304 3K 0
0
0
0.30
0.30 305 3K 0
0
0
0.30
0.30 306 2K 0
0
0
0.15 0.15 307 0 0 0 0 0.15 0.15 308 0 0 0 0 0.15 0.15 309 0 0 IK 0
0.075 0.075 0.075
0.074 310 IK
0.075
311
0
0.074 312 0
0 0 0
0 0 0
0 0 0
0 0 313 0 0 0 0 0 0 314 0 0 0 0 0 0 315 0 0 0 0
1 based on nominal consumption of test solution of 20pl/bee based on amount of test solution consumed (average of 10 bees)
Environmental Laboratory Project Number U2723 Page 22 of 25
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+ ! #
:* rm
:m
I
;*
\I
>
j*
im
CSL Study Number HT5602
APPENDIX 2 INTERIM CERTIFICATE OF ANALYSIS FOR TEST SUBSTANCE
INTERIM CERTIFICATE OFANALYSIS
Revision 1(9/7/00)
Centre Analytical Laboratories COA Reference #: 023-0ISA 3M Product: PFOS,Lot217 Reference #: SD-018
Teat Name
Purity1
Appearance
Identification NMR
Metals (ICP/MS) 1. Calcium 2. Magnesium 3. Sodium 4. Potassium1 5. Nickel 6. Iron 7. Manganese
Total % Impurity (NMR) Total % Impurity (LC/MS) Total % Impurity (GC/MS) Related Compounds POAA Residual Solvents (TGA) Purity by DSC Inorganic Anions (IC)
1. Chloride 2. Fluoride 3. Bromide 4. Nitrate 5. Nitrite 6. Phosphate 7. Sulfate'* Organic Acids 5 (IC) 1. TFA 2. PFPA 3. HFBA 4. NFPA
Elemental Analysis6: l. Carbon 2. Hydrogen 3. Nitrogen 4. Sulfur 5. Fluorine
Specifications White Crystalline: Powder
Result 86.9%
Conforms
Positive
1. 0.005 wtVwt.% 2. 0.001 wt./wt.% 3. 1.439 wt./wt.% 4. 6.849 wt./wt.% 5. <0.001 wt./wt.% 6. 0.005 wt./wt.% 7. <0.001 wt./wt.%
1.93 wt./wt.% 8.41 wt./wt.%
None Detected
;'il t
1. Theoretical Value =17.8% 2. Theoretical Value = 0% 3. Theoretical Value = 0% 4. Theoretical Value = 5.95% 5. Theoretical Value = 60%
0.33 wt./wt.% None Detected Not Applicable3
1. <0.015 wt./wt.% 2. 0.59 wt./wt.% 3. <0.040 wt/w t.% 4. <0.009 wt./wt.% 5. <0.006 wt./wt.% 6. <0.007 wt./wt.% 7. 8.76 wt7wt.%
1. <0.1 w t/w t.% 2. <0.1wt./wt.% 3. 0.10 w t/w t.% 4. 0.28 wt./wt.%
1. 12.48 w t/w t.% 2. 0.244 w t/w t.% 3. 1.74 w t/w t.% 4. 8.84 w t/w t.% 5. 54.1 w t/w t.%
COA023-018A
Page 1 of 3
Environmental Laboratory Project Number U2723
Page 23 of 25 .
GG0037
CSL Study Number HT5602 #
m
INTERIM CERTIFICATE OF ANALYSIS
Centre Analytical Laboratories COA Reference #: 023-018A
Date of Last Analysis: 08/31/00
Expiration Date: 08/31/01
I r*
Storage Conditions: Frozen <-10C
[ Re-assessment Date: 08/31/01
i
`Purity = 100% - (sumof metal impurities, 1.45% +LC/MS impurities, 8.41%+Inorganic
I
Fluoride, 0.59%+NMR impurities, 1.93%+organic acid impurities, 0.38%+POAA, 0.33%)
Total impurity from all tests = 13.09%
Purity = 100% - 13.09% = 86.9%
1Potassiumis expected in this salt form and is therefore not considered an impurity.
3Purity by DSC is generally not applicable to materials of low purity. No endothermwas observed for this sample.
4Sulfur inthe sample appears to be converted to SO4 and hence detected using the inorganic anionmethod conditions. The anion result agrees well with the sulfur determination in the elemental analysis, lending confidence to this interpretation. Based onthe results, the SO4 is not considered animpurity.
5TFA HFBA NFPA
PFPA
Trifluoroacetic acid Heptafluorobutyric acid Nonofluoropentanoic acid
Pentafluoropropanoic acid
theoretical value calculations based onthe empirical formula, CgFnSOj'K* (MW=538)
This work was conducted under EPA Good Laboratory Practice Standards (40 CFR 160). /
COA023-018A
Environmental Laboratory Project Number U2723 Page 24 o f 25
G CCOai
CSL Study Number HT5602
INTERIM CERTIFICATE OFANALYSIS
Centre Analytical Laboratories COA Reference #: 023-018A
LC/MS Purity Profile:
Impurity C4 C5 C6
C7
Total
wt./wt. % 1.22 1.33 4.72
1.14
8.41
Note: The C4 and C6 values were calculated using the C4 and C6 standard calibration curves, respectively. The C5 value was calculated using the average response factors fromthe C4 and C6 standard curves. Likewise, the C7 value was calculated using the average response factors from the C6 and C8 standard curves.
Prepared By: ___________________ ______
___ ;
David S. Bell
Date
Scientist, Centre Analytical Laboratories
Reviewed B y :____________________ [______
___
John Flaherty
Date
Laboratory Manager, Centre Analytical Laboratories
Environmental Laboratory Project Number U2723 Page 25 of 25
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