A EPA
EPA/635/R-21 /312b
External Review Draft
www.epa.gov/iris
Toxicological Review of Perfluorohexanoic Acid [CASRN 307-24-4] and
Related Salts
Supplemental Information
February 2022
Integrated Risk Information System
Center for Public Health and Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Washington, DC
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
DISCLAIMER
This document is an external peer review draft for review purposes only. This information
is distributed solely for the purpose of external peer review. It has not been formally disseminated
by EPA. It does not represent and should not be construed to represent any Agency determination
or policy. Mention of trade names or commercial products does not constitute endorsement or
recommendation for use.
This document is a draft for review purposes only and does not constitute Agency policy.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
TABLE OF CONTENTS
TABLE OF CONTENTS iii
APPENDIX A. SYSTEMATIC REVIEW PROTOCOL FOR THE PFAS IRIS ASSESSMENTS A-l
APPENDIX B. BENCHMARK DOSE MODELING RESULTS B-l
B.l. MODELING PROCEDURE FOR CONTINUOUS NONCANCER DATA B-l
B.2. HEMOGLOBIN —FEMALE RATS (Klaunig et al., 2015) B-2
B.3. HEMOGLOBIN —MALE RATS (Chengelis et al., 2009b) B-4
B.4. HEMOGLOBIN —FEMALE RATS (Chengelis et al., 2009b) B-6
B.5. HEMOGLOBIN —MALE RATS (Loveless et al., 2009) B-8
B.6. HEMOGLOBIN —FEMALE RATS (Loveless et al., 2009) B-10
B.7. RED BLOOD CELLS—MALE RATS (Klaunig et al., 2015) B-12
B.8. RED BLOOD CELLS—FEMALE RATS (Klaunig et al., 2015) B-14
B.9. RED BLOOD CELLS—MALE RATS (Chengelis et al., 2009b) B-16
B.10. RED BLOOD CELLS—FEMALE RATS (Chengelis et al., 2009b) B-18
B.ll. RED BLOOD CELLS—MALE RATS (Loveless et al., 2009) B-20
B.12. RED BLOOD CELLS—FEMALE RATS (Loveless et al., 2009) B-22
B.13. HEPATOCELLULAR HYPERTROPHY—MALE RATS (Chengelis et al., 2009b) B-24
B.14. HEPATOCELLULAR HYPERTROPHY—FEMALE RATS (Loveless et al., 2009) B-25
B.15. HEPATOCELLULAR HYPERTROPHY—MALE RATS (Loveless et al., 2009) B-27
B.16. POSTNATAL (Fi) COMBINED RAT BODY WEIGHT ON PND 0 (LOVELESS ET AL., 2009) B-29
B.17. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 2) ON PND 0 (IWAI AND
HOBERMAN, 2014) B-31
B.18. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 1) ON PND 0 (IWAI AND
HOBERMAN, 2014) B-33
B.19. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASES 1 AND 2) ON PND 0
(IWAI AND HOBERMAN, 2014) B-34
B.20. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 2) ON PND 4 (IWAI AND
HOBERMAN, 2014) B-35
B.21. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 1) ON PND 4 (IWAI AND
HOBERMAN, 2014) B-37
B.22. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASES 1 AND 2) ON PND 4
(IWAI AND HOBERMAN, 2014) B-38
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B.23. PERINATAL MORTALITY (PHASE 2) ON PNDs 0-21 (IWAI AND HOBERMAN, 2014) B-40
B.24. PERINATAL MORTALITY (PHASE 1) ON PNDs 0-21 (IWAI AND HOBERMAN, 2014) B-41
B.25. PERINATAL MORTALITY (PHASES 1 AND 2) ON PNDs 0-21 (IWAI AND HOBERMAN,
2014) B-42
APPENDIX C. EVALUATION OF PFHxA ELIMINATION C-l
C.l. EVALUATION OF PFHxA ELIMINATION IN RATS AND MICE C-l
C.1.1. Mice C-2
C.l. 2. Rats C-4
C.2. EVALUATION OF PFHXA ELIMINATION IN HUMANS C-4
APPENDIX D. QUALITY ASSURANCE FOR THE IRIS TOXICOLOGICAL REVIEW OF PFHxA D-l
APPENDIX E. SUMMARY OF PUBLIC COMMENTS AND EPA'S DISPOSITION E-l
REFERENCES 1
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
TABLES
Table B-l. Dose response data for hemoglobin in female rats (Klaunig et al., 2015) B-2
Table B-2. Benchmark dose results for hemoglobin in female rats—constant variance, BMR = 1
standard deviation (Klaunig et al., 2015) B-2
Table B-3. Dose response data for hemoglobin in male rats (Chengelis et al., 2009b) B-4
Table B-4. Benchmark dose results for hemoglobin in male rats—constant variance, BMR = 1
standard deviation (Chengelis et al., 2009b) B-4
Table B-5. Dose response data for hemoglobin in female rats (Chengelis et al., 2009b) B-6
Table B-6. Benchmark dose results for hemoglobin in female rats—nonconstant variance, BMR =
1 standard deviation (Chengelis et al., 2009b) B-6
Table B-7. Dose response data for hemoglobin in male rats (Loveless et al., 2009) B-8
Table B-8. Benchmark dose results for hemoglobin in male rats nonconstant variance, BMR = 1
standard deviation (Loveless et al., 2009) B-8
Table B-9. Dose response data for hemoglobin in female rats (Loveless et al., 2009) B-10
Table B-10. Benchmark dose results for hemoglobin in female rats constant variance, BMR = 1
standard deviation (Benchmark dose results for hemoglobin in female rats
constant variance, BMR = 1 standard deviation (Loveless et al., 2009) B-10
Table B-ll. Dose response data for red blood cells in male rats (Klaunig et al., 2015) B-12
Table B-12. Benchmark dose results for red blood cells in male rats—nonconstant variance, BMR
= 1 standard deviation (Klaunig et al., 2015) B-12
Table B-13. Dose response data for red blood cells in female rats (Klaunig et al., 2015) B-14
Table B-14. Benchmark dose results for red blood cells in female rats—constant variance, BMR
= 1 standard deviation (Klaunig et al., 2015) B-14
Table B-15. Dose response data for red blood cells in male rats (Chengelis et al., 2009b) B-16
Table B-16. Benchmark dose results for red blood cells in male rats—nonconstant variance, BMR
= 1 standard deviation (Chengelis et al., 2009b) B-16
Table B-17. Dose response data for red blood cells in female rats (Chengelis et al., 2009b) B-18
Table B-18. Benchmark dose results for red blood cells in female rats—constant variance, BMR =
1 standard deviation (Chengelis et al., 2009b) B-18
Table B-19. Dose response data for red blood cells in male rats (Loveless et al., 2009) B-20
Table B-20. Benchmark dose results for red blood cells in male rats—nonconstant variance, BMR
= 1 standard deviation (Loveless et al., 2009) B-20
Table B-21. Dose response data for red blood cells in female rats (Loveless et al., 2009) B-22
Table B-22. Benchmark dose results for red blood cells in female rats—constant variance, BMR =
1 standard deviation (Loveless et al., 2009) B-22
Table B-23. Dose response data for hepatocellular hypertrophy in male rats (Chengelis et al.,
2009b) B-24
Table B-24. Dose response data for hepatocellular hypertrophy in female rats (Loveless et al.,
2009) B-25
Table B-25. Benchmark dose results for hepatocellular hypertrophy in female rats—nonconstant
variance, BMR = 10% Extra Risk (Loveless et al., 2009) B-25
Table B-26. Dose response data for hepatocellular hypertrophy in male rats (Loveless et al.,
2009) B-27
Table B-27. Benchmark dose results for hepatocellular hypertrophy in male rats—nonconstant
variance, BMR = 10% Extra Risk (Loveless et al., 2009) B-27
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Table B-28. Dose response data for postnatal (Fl) combined rat body weight on PND 0 (Loveless
et al., 2009) B-29
Table B-29. Benchmark dose results for postnatal (Fl) combined rat body weight on PND
0—nonconstant variance, BMR = 5% relative deviation (Loveless et al2009) B-29
Table B-30. Dose response data for postnatal (Fl) combined mouse body weight (Phase 2) on
PND 0 (Iwai and Hoberman, 2014) B-31
Table B-31. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 2)
on PND 0—constant variance, BMR = 5% relative deviation (Benchmark dose
results for postnatal (Fi) combined mouse body weight (Phase 2) on PND
0—constant variance, BMR = 5% relative deviation (Iwai and Hoberman, 2014) B-31
Table B-32. Dose response data for postnatal (Fl) combined mouse body weight (Phase 1) on
PND 0 (Iwai and Hoberman, 2014) B-33
Table B-33. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 1)
on PND 0—nonconstant variance, BMR = 5% relative deviation (Iwai and
Hoberman, 2014) B-33
Table B-34. Dose response data for postnatal (Fl) combined mouse body weight (Phases 1 and
2) on PND 0 Dose-response data for postnatal (Fi) combined mouse body weight
(Phases 1 and 2) on PND 0 (Iwai and Hoberman, 2014) B-34
Table B-35. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 1
and 2) on PND 0—nonconstant variance, BMR = 5% relative deviation
Benchmark dose results for postnatal (Fi) combined mouse body weight (Phase
1 and 2) on PND 0—nonconstant variance, BMR = 5% relative deviation (Iwai
and Hoberman, 2014) B-34
Table B-36. Dose response data for postnatal (Fl) combined mouse body weight (Phase 2) on
PND 4 Dose-response data for postnatal (Fi) combined mouse body weight
(Phase 2) on PND 4 (Iwai and Hoberman, 2014) B-35
Table B-37. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 2)
on PND 4—constant variance, BMR = 5% relative deviation Benchmark dose
results for postnatal (Fi) combined mouse body weight (Phase 2) on PND
4—constant variance, BMR = 5% relative deviation (Iwai and Hoberman, 2014) B-35
Table B-38. Dose response data for postnatal (Fl) combined mouse body weight (Phase 1) on
PND 4 Dose-response data for postnatal (Fi) combined mouse body weight
(Phase 1) on PND 4 (Iwai and Hoberman, 2014) B-37
Table B-39. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 1)
on PND 4—nonconstant variance, BMR = 5% relative deviation Benchmark dose
results for postnatal (Fi) combined mouse body weight (Phase 1) on PND
4—nonconstant variance, BMR = 5% relative deviation (Iwai and Hoberman,
2014) B-37
Table B-40. Dose response data for postnatal (Fl) combined mouse body weight (Phase 1) on
PND 4 Dose-response data for postnatal (Fi) combined mouse body weight
(Phase 1) on PND 4 (Iwai and Hoberman, 2014) B-38
Table B-41. Benchmark dose results for postnatal (Fl) combined mouse body weight (Phase 1
and 2) on PND 4—nonconstant variance, BMR = 5% relative deviation
Benchmark dose results for postnatal (Fi) combined mouse body weight (Phase
1 and 2) on PND 4—nonconstant variance, BMR = 5% relative deviation (Iwai
and Hoberman, 2014) B-38
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Table B-42. Nested Model summary for perinatal mortality (Phase 2) on PNDs 0-21, BMR = 1%
extra risk Nested Model summary for perinatal mortality (Phase 2) on PNDs 0-
21, BMR = 1% extra risk (Iwai and Hoberman, 2014) B-40
Table B-43. Nested model summary for perinatal mortality (Phase 1) on PNDs 0-21, BMR = 1%
extra risk Nested model summary for perinatal mortality (Phase 1) on PNDs 0-
21, BMR = 1% extra risk (Iwai and Hoberman, 2014) B-41
Table B-44. Nested model summary for perinatal mortality (Phase 1 and 2) on PNDs 0-21, BMR =
1% extra risk Nested model summary for perinatal mortality (Phase 1 and 2) on
PNDs 0-21, BMR = 1% extra risk (Iwai and Hoberman, 2014) B-42
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
FIGURES
Figure B-l. Dose response curve for the Linear model fit to hemoglobin in female rats (Klaunig et
al., 2015) B-3
Figure B-2. Dose response curve for the Polynomial Degree 3 model fit to hemoglobin in male
rats (Chengelis et al., 2009b) B-5
Figure B-3. Dose response data for hemoglobin in female rats (Chengelis et al., 2009b) B-7
Figure B-4. Dose response data for hemoglobin in male rats (Loveless et al., 2009) B-9
Figure B-5. Dose response curve for the Polynomial Degree 3 model fit to hemoglobin in female
rats (Loveless et al., 2009) B-ll
Figure B-6. Dose response data hemoglobin in male rats (Klaunig et al., 2015) B-13
Figure B-7. Dose response curve for the Linear model fit to hemoglobin in female rats (Klaunig et
al., 2015) B-15
Figure B-8. Dose response data for red blood cells in male rats (Chengelis et al., 2009b) B-17
Figure B-9. Dose response curve for the Exponential 5 model fit to red blood cells in female rats
(Chengelis et al., 2009b) B-19
Figure B-10. Dose response curve for the Linear model fit to red blood cells in male rats
(Loveless et al., 2009) B-21
Figure B-ll. Dose response curve for the Linear model fit to red blood cells in female rats
(Loveless et al., 2009) B-23
Figure B-12. Dose response data for hepatocellular hypertrophy in male rats (Chengelis et al.,
2009b) B-24
Figure B-13. Dose response curve for the Multistage Degree 3 model fit to hepatocellular
hypertrophy in female rats (Loveless et al., 2009) B-26
Figure B-14. Dose response curve for the Multistage Degree 1 model fit to hepatocellular
hypertrophy in male rats (Loveless et al., 2009) B-28
Figure B-15. Dose response curve for the Hill model fit to postnatal (Fl) combined rat body
weight on PND 0 (Loveless et al., 2009) B-30
Figure B-16. Dose response curve for the Polynomial Degree 3 model fit to postnatal (Fl)
combined rat body weight (Phase 2) on PND 0 (Iwai and Hoberman, 2014) B-32
Figure B-17. Dose response curve for the Polynomial model (poly 3) fit to postnatal (Fl)
combined rat body weight (Phase 2) on PND 4 Dose-response curve for the
Polynomial model (poly 3) fit to postnatal (Fi) combined rat body weight (Phase
2) on PND 4 (Iwai and Hoberman, 2014) B-36
Figure B-18. Dose response curve for the Exponential 5 model fit to postnatal (Fl) combined
mouse body weight (Phases 1 and 2) on PND 4 Dose-response curve for the
Exponential 5 model fit to postnatal (Fi) combined mouse body weight (Phases
1 and 2) on PND 4 (Iwai and Hoberman, 2014) B-39
Figure B-19. Dose response curve for the Nested NCTR model fit to perinatal mortality (Phase 2)
on PND 0-21 (Iwai and Hoberman, 2014) B-40
Figure C-l. Fits of population pharmacokinetic model to data for male (top row) and female
(remaining rows) mice following 2-350 mg/kg oral exposure PFHxA C-3
Figure C-2. Fits of human PFHxA data from ski-wax technician blood samples C-5
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
ABBREVIATIONS AND ACRONYMS
ADME
absorption, distribution, metabolism,
i.v.
intravenous
and excretion
LDH
lactate dehydrogenase
AFFF
aqueous film-forming foam
LLOQ
lower limit of quantitation
A:G
albumin:globulin ratio
LOQ
limit of quantitation
AIC
Akaike's information criterion
LOAEL
lowest-observed-adverse-effect level
ALP
alkaline phosphatase
LOD
limit of detection
ALT
alanine aminotransferase
LOEC
lowest observed effect concentration
APTT
activated partial thromboplastin time
MCH
mean cell hemoglobin
AST
aspartate aminotransferase
MCHC
mean cell hemoglobin concentration
atm
atmosphere
MCV
mean corpuscular volume
ATSDR
Agency for Toxic Substances and
MOA
mode of action
Disease Registry
MW
molecular weight
AUC
area under the curve
NCTR
National Center for Toxicological
BMD
benchmark dose
Research
BMDL
benchmark dose lower confidence limit
NOAEL
no-observed-adverse-effect level
BMDS
Benchmark Dose Software
NPL
National Priorities List
BMR
benchmark response
NTP
National Toxicology Program
BUN
blood urea nitrogen
ORD
Office of Research and Development
BW
body weight
OECD
Organisation for Economic
Cmax
maximum concentration
Co-operation and Development
CAR
constitutive androstane receptor
OSF
oral slope factor
CASRN
Chemical Abstracts Service registry
osRfD
organ/system-specific oral reference
number
dose
CBC
complete blood count
PBPK
physiologically based pharmacokinetic
CHO
Chinese hamster ovary (cell line cells)
PC
partition coefficient
CI
confidence interval
PECO
populations, exposures, comparators,
CL
clearance
and outcomes
CLa
clearance in animals
PFAA
perfluoroalkyl acids
CLh
clearance in humans
PFAS
per- and polyfluoroalkyl substances
CPHEA
Center for Public Health and
PFBA
perfluorobutanoic acid
Environmental Assessment
PFBS
perfluorobutane sulfonate
CPN
chronic progressive nephropathy
PFCA
perfluorinated carboxylic acid
DAF
dosimetric adjustment factor
PFDA
perfluorodecanoic acid
DNA
deoxyribonucleic acid
PFHxA
perfluorohexanoic acid
DTXSID
DSSTox substance identifier
PFHxS
perfluorohexane sulfonate
EPA
Environmental Protection Agency
PFNA
perfluorononanoic acid
FTOH
fluorotelomer alcohol
PFOA
perfluorooctanoic acid
GD
gestation day
PFOS
perfluorooctane sulfonate
GGT
y-glutamyl transferase
PK
pharmacokinetic
HAWC
Health Assessment Workplace
PND
postnatal day
Collaborative
POD
point of departure
HCT
hematocrit
PODhed
human equivalent dose POD
HED
human equivalent dose
PPAR
peroxisome proliferated activated
HERO
Health and Environmental Research
receptor
Online
PQAPP
programmatic quality assurance
HGB
hemoglobin
project plan
HSA
human serum albumin
PT
prothrombin time
IQR
interquartile range
QA
quality assurance
IRIS
Integrated Risk Information System
QAPP
quality assurance project plan
ISI
Influential Scientific Information
QMP
quality management plan
IUR
inhalation unit risk
RBC
red blood cells
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
RfC reference concentration
RfD oral reference dose
RNA ribonucleic acid
ROS reactive oxygen species
RXR retinoid X receptor
SD standard deviation
TP total protein
TRI Toxics Release Inventory
TSCATS Toxic Substances Control Act Test
Submissions
TSH thyroid stimulating hormone
UF uncertainty factor
UFa interspecies uncertainty factor
UFc composite uncertainty factor
UFd evidence base deficiencies uncertainty
factor
UFh human variation uncertainty factor
UFl LOAEL to NOAEL uncertainty factor
UFs subchronic to chronic uncertainty
factor
Vd volume of distribution
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
APPENDIX A. SYSTEMATIC REVIEW PROTOCOL FOR
THE PFAS IRIS ASSESSMENTS
1 A single systematic review protocol was used to guide the development of five separate IRIS
2 PFAS [per- and polyfluoroalkyl substances] assessments (i.e., perfluorobutanoic acid [PFBA],
3 perfluorohexanoic acid [PFHxA], perfluorohexane sulfonate [PFHxS], perfluorononanoic acid
4 [PFNA], and perfluorodecanoic acid [PFDA]). This "Systematic Review Protocol for the PFAS IRIS
5 Assessments" was released for public comment and subsequently updated. The updated protocol
6 and prior revisions can be found at the following location:
7 http://cfpub.epa.gov/ncea/iris drafts/recordisplay.cfm?deid=345065
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
APPENDIX B. BENCHMARK DOSE MODELING
RESULTS
As discussed in the body of the report (Section 5), the endpoints selected for benchmark
dose (BMD) modeling were hepatocellular hypertrophy from Chengelis etal. (2009a) and Loveless
etal. (20091: hemoglobin and red blood cells from Chengelis etal. (2009a). Loveless etal. (20091.
and Klaunigetal. f20151: postnatal body weight decreases from Loveless etal. f20091 and Iwai and
Hoberman f20141: and perinatal mortality from Iwai and Hoberman f20141. The animal doses in
the study were used in the BMD modeling and then converted to human equivalent doses (HEDs)
using the ratio of animal-to-human serum half-lives. For endpoints with successful BMD model fit,
the modeling results are presented in this Appendix.
B.l. MODELING PROCEDURE FOR CONTINUOUS NONCANCER DATA
BMD modeling of continuous noncancer data was conducted using EPA's Benchmark Dose
Software (BMDS, Version 3.2). For these data, the Exponential, Hill, Polynomial, and Power models
available within the software are fit using a benchmark response (BMR) of 1 standard deviation
(SD) when no toxicological information was available to determine an adverse level of response.
When toxicological information was available, the BMR was based on relative deviation, as outlined
in the Benchmark Dose Technical Guidance (U.S. EPA. 20121. An adequate fit is judged on the basis
of ax2 goodness-of-fit p-value (p > 0.1), scaled residuals at the data point (except the control)
closest to the predefined BMR (absolute value <2.0), and visual inspection of the model fit In
addition to these three criteria for judging adequacy of model fit, a determination is made as to
whether the variance across dose groups is homogeneous. If a homogeneous variance model is
deemed appropriate on the basis of the statistical testprovided by BMDS (i.e., Test 2), the final BMD
results are estimated from a homogeneous variance model. If the test for homogeneity of variance
is rejected (p < 0.05), the model is run again while modeling the variance as a power function of the
mean to account for this nonhomogeneous variance. If this nonhomogeneous variance model does
not adequately fit the data (i.e., Test 3; p < 0.05), the data set is considered unsuitable for BMD
modeling. In cases where a model with # parameters = # dose groups was fit to the data set and all
parameters were estimated and no p-value was calculated, that model was not considered for
estimation of a point of departure (POD). Among all models providing adequate fit, the benchmark
dose lower confidence limit (BMDL) from the model with the lowest Akaike's information criterion
(AIC) was selected as a potential POD when BMDL estimates differed by less than threefold. When
BMDL estimates differed by greater than threefold, the model with the lowest BMDL was selected
to account for model uncertainty.
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1 For relative liver weight, a BMR equal to 10% increase from the control mean was used
2 based on a biological consideration. For continuous developmental toxicity data, a BMR equal to
3 0.5 SD was used. The use of 1 SD for the BMR for continuous endpoints is based the observation
4 that shifting the distribution of the control group by 1 SD results in ~10% of animals falling beyond
5 an adversity cutoff defined at the ~1.5 percentile in the control group fCrump. 19951. This roughly
6 approximates the 10% extra risk commonly used as the BMR for dichotomous endpoints. Thus, the
7 use of 0.5 SD for continuous developmental toxicity endpoints roughly approximates the extra risk
8 of 5% commonly used for dichotomous developmental toxicity endpoints.
B.2. HEMOGLOBIN-FEMALE RATS fKLAUNIG ET AL.. 20151
Table B-l. Dose response data for hemoglobin in female rats (Klaunig et al..
2015.)
Dose (mg/kg-day)
Number of
animals
Mean (g/dL)
Standard deviation
0
10
15.5
0.97
5
10
15.7
0.73
30
9
15.5
0.79
200
20
14.7
0.91
Table B-2. Benchmark dose results for hemoglobin in female rats—constant
variance, BMR = 1 standard deviation (Klaunig et al.. 2015)
Models
p-Values
Test 2
p-values
AIC
1 SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
0.83237671
0.7551
127.5023
182.1091
120.47632
-0.016328458
Exponential 3
0.57454386
0.7551
129.4505
189.9502
120.87504
0.002065941
Exponential 4
0.83237684
0.7551
127.5023
182.0793
120.47647
-0.015845878
Exponential 5
0.57331833
0.7551
129.4525
188.501
120.85884
-0.000961421
Hill
NA
0.7551
131.4228
42.56095
31.718073
0.000755846
Polynomial (Poly 3)
0.56681655
0.7551
129.4634
191.4936
123.01116
0.00113966
Polynomial (Poly 2)
0.56681165
0.7551
129.4634
191.4757
123.0163
0.001201719
Power
0.57425021
0.7551
129.451
190.1218
123.04966
0.002120702
Linear
0.83402366
0.7551
127.4983
182.7286
122.7699
-0.014122961
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Linear Model with BMR of 1 Std. Dev. for the BMD
and 0.95 Lower Confidence Limit for the BMDL
18
16
14
12
10
o
—©
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.3. HEMOGLOBIN-MALE RATS fCHENGELIS ET AL.. 2009B1
Table B-3. Dose response data for hemoglobin in male rats (Chengelis et al..
2009b)
Dose (mg/kg-day)
Number of
animals
Mean (g/dL)
Standard deviation
0
10
15.6
0.51
10
10
15.4
0.58
50
10
15.4
0.65
200
10
14.3
1.08
Table B-4. Benchmark dose results for hemoglobin in male rats—constant
variance, BMR = 1 standard deviation ((Chengelis et al.. 2009bl
Models
p-Values
Test 2
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
0.704934
0.064112
91.82015
110.5213
77.18423
0.615129749
Exponential 3
0.539535
0.064112
93.49724
142.4976
78.66252
-0.004155158
Exponential 4
0.704934
0.064112
91.82015
110.5301
77.18593
0.615097154
Exponential 5
0.539535
0.064112
93.49724
142.4988
78.6632
-0.004163015
Hill
NA
0.064112
95.52553
58.25582
51.28692
1.3298 x 10 6
Polynomial (Poly 3)
0.853527
0.064112
91.4376
151.5179
81.3468
-0.001390585
Polynomial (Poly 2)
0.558344
0.064112
93.46342
143.161
81.22378
-0.006077488
Power
0.539814
0.064112
93.49673
141.0967
81.06874
-0.007648406
Linear
0.719614
0.064112
91.77892
112.5099
79.80677
0.5847594
This document is a draft for review purposes only and does not constitute Agency policy.
B-4 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Polynomial Degree 3 Model with BMR of 1 Std. Dev.
for the BMD and 0.95 Lower Confidence Limit for the BMDL
18
16
Q t» m
14
12
10
o
$
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.4. HEMOGLOBIN-FEMALE RATS fCHENGELIS ET AL.. 2009B1
Table B-5. Dose response data for hemoglobin in female rats (Chengelis et al..
2009b)
Dose (mg/kg-day)
Number of
animals
Mean (g/dL)
Standard deviation
0
10
15.6
0.46
10
10
15.8
1.4
50
10
15.2
0.85
200
10
14.6
0.83
Table B-6. Benchmark dose results for hemoglobin in female
rats—nonconstant variance, BMR = 1 standard deviation (Chengelis et al..
2009b)
Models
p-Values
Test 3
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
0.2289302
0.0118
113.344
177.8625
106.4881
0.107636338
Exponential 3
0.2289313
0.0118
113.344
177.8107
106.4883
0.107765953
Exponential 4
0.0996002
0.0118
115.1073
145.8206
37.82113
0.036336773
Exponential 5
0.165197
0.0118
114.3214
53.45159
25.38329
0.084452285
Hill
NA
0.0118
116.3216
68.93813
40.08308
0.084913389
Polynomial (Poly 3)
0.2265515
0.0118
113.3649
179.1794
109.5823
0.105758473
Polynomial (Poly 2)
0.2265515
0.0118
113.3649
179.1817
110.5758
0.105711892
Power
0.2265515
0.0118
113.3649
179.174
109.6196
0.105830655
Linear
0.2265515
0.0118
113.3649
179.1809
110.1348
0.10575497
Both constant and nonconstant variance models failed to model the variance. Therefore, this data set is not
amendable for BMD modeling.
This document is a draft for review purposes only and does not constitute Agency policy.
B-6 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Hemoglogin (HGB)
13.5
0 20 40 60 80 100 120 140 160 180 200
Dose
Figure B-3. Dose response data for hemoglobin in female rats (Chengelis et al..
2009b).
This document is a draft for review purposes only and does not constitute Agency policy.
B-7 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.5. HEMOGLOBIN-MALE RATS fLOVELESS ET AL.. 20091
Table B-7. Dose response data for hemoglobin in male rats (Loveless et al..
2009)
Dose (mg/kg-day)
Number of
animals
Mean (g/dL)
Standard deviation
0
10
15.4
0.5
20
10
15.5
0.41
100
10
4.5
0.7
500
10
9.9
2.8
Table B-8. Benchmark dose results for hemoglobin in male rats nonconstant
variance, BMR = 1 standard deviation (Loveless etal.. 2009)
Models
p-Values
Test 3
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
<0.0001
<0.0001
199.3758
9.377807
7.193218
-2.377990291
Exponential 3
<0.0001
<0.0001
239.5418
855.7401
0
0.802984088
Exponential 4
<0.0001
<0.0001
190.0784
6.631732
3.474481
-0.818335738
Exponential 5
0.071088
<0.0001
138.3961
70.68336
21.21839
-2.570261966
Hill
0.07109
<0.0001
138.3961
38.98926
21.0774
0.362134233
Polynomial (Poly 3)
<0.0001
<0.0001
239.7107
891.7542
383.4838
0.626839397
Polynomial (Poly 2)
<0.0001
<0.0001
239.7107
891.7546
383.3929
0.626839015
Power
<0.0001
<0.0001
239.7107
891.7544
383.3928
0.626839886
Both constant and nonconstant variance models failed to model the variance. Therefore, this data set is not
amendable for BMD modeling.
This document is a draft for review purposes only and does not constitute Agency policy.
B-8 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Hemoglobin (HGB)
18
0
0 50 100 150 200 250 300 350 400 450 500
Dose
Figure B-4. Dose response data for hemoglobin in male rats (Loveless etal..
2009V
This document is a draft for review purposes only and does not constitute Agency policy.
B-9 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.6. HEMOGLOBIN-FEMALE RATS fLOVELESS ET AL.. 20091
Table B-9. Dose response data for hemoglobin in female rats (Loveless et al..
2009)
Dose (mg/kg-day)
Number of
animals
Mean (g/dL)
Standard deviation
0
10
15.6
0.7
20
10
15.8
0.8
100
10
15.6
0.4
500
9
13.3
0.9
Table B-10. Benchmark dose results for hemoglobin in female rats constant
variance, BMR = 1 standard deviation (Loveless et al.. 2009)
Models
p-Values
Test 2
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
0.214488
0.107799
89.79631
134.0618
104.1801
1.219441036
Exponential 3
0.50507
0.107799
89.16158
264.9174
126.3561
-0.026708969
Exponential 4
0.214489
0.107799
89.7963
134.0137
104.1803
1.219907658
Exponential 5
0.505122
0.107799
89.16147
266.3836
126.3586
-0.034100913
Hill
NA
0.107799
91.14613
115.7416
102.1374
-1.06893E-06
Polynomial (Poly 3)
0.800193
0.107799
87.16312
268.4412
127.6129
-0.023130227
Polynomial (Poly 2)
0.800179
0.107799
87.16315
267.1194
127.8182
-0.018215642
Power
0.452941
0.107799
89.2806
372.7895
126.1226
0.000358346
Linear
0.259194
0.107799
89.41767
141.5272
111.6505
1.119316772
This document is a draft for review purposes only and does not constitute Agency policy.
B-10 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Polynomial Degree 3 Model with BMR of 1 Std. Dev.
for the BMD and 0.95 Lower Confidence Limit for the BMDL
18
16
m(D a
14
12
10
o
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.7. RED BLOOD CELLS-MALE RATS fKLAUNIG ET AL.. 20151
Table B-ll. Dose response data for red blood cells in male rats (Klaunig et al..
2015.)
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
9.2
0.17
2.5
10
8.8
1.52
15
9
8.66
0.92
100
19
8.8
1
Table B-12. Benchmark dose results for red blood cells in male
rats—nonconstant variance, BMR = 1 standard deviation (Klaunig etal.. 20151
Models
p-Values
Test 3
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
<0.0001
0.863381
143.4547
808.9905
155.542
0.078851
Exponential 3
<0.0001
0.863381
143.4551
832.8003
104.7724
0.070665
Exponential 4
<0.0001
0.863381
144.0729
-9999
0
-9999
Exponential 5
<0.0001
0.863381
145.454
865.033
103.3948
0.080901
Hill
NA
0.863381
121.2797
-9999
0
-9999
Polynomial (Poly 3)
<0.0001
0.863381
143.4552
774.4171
120.0894
0.076434
Polynomial (Poly 2)
<0.0001
0.863381
143.4552
774.9496
132.5465
0.076318
Power
<0.0001
0.863381
143.4552
775.0219
106.6368
0.076271
Linear
<0.0001
0.863381
143.4552
775.3403
153.4792
0.076151
The only model provides adequate fit is the Hill model, but the Hill model failed to estimate BMDL.
Both constant and nonconstant variance models failed to model the variance data.
This document is a draft for review purposes only and does not constitute Agency policy.
B-12 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Red Blood Cell (RBC)
12
10
8 —
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.8. RED BLOOD CELLS-FEMALE RATS fKLAUNIG ET AL.. 20151
Table B-13. Dose response data for red blood cells in female rats (Klaunig et
al.. 201 SI
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
8.14
0.52
5
10
8.23
0.58
30
9
8.12
0.37
200
20
7.48
0.68
Table B-14. Benchmark dose results for red blood cells in female
rats—constant variance, BMR = 1 standard deviation (Klaunig et al.. 20151
Models
p-Values
Test 2
p-values
AIC
1SD
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential 2
0.896578
0.204474
88.37423
153.772
105.8225
-0.0212
Exponential 3
0.702156
0.204474
90.30213
168.4422
106.2748
0.001301
Exponential 4
0.896578
0.204474
88.37423
153.7727
105.8231
-0.02121
Exponential 5
NA
0.204474
92.30211
168.1679
30.68698
0.001229
Hill
NA
0.204474
92.28508
40.32119
31.54188
0.000759
Polynomial (Poly 3)
0.690261
0.204474
90.3147
175.8228
109.4569
0.000354
Polynomial (Poly 2)
0.69227
0.204474
90.31254
173.1861
109.4699
0.000719
Power
0.701613
0.204474
90.3027
169.0362
109.5006
0.000962
Linear
0.900552
0.204474
88.36539
155.595
109.1493
-0.01798
This document is a draft for review purposes only and does not constitute Agency policy.
B-14 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Linear Model with BMR of 1 Std. Dev. for the BMD
and 0.95 Lower Confidence Limit for the BMDL
10
9
Estimated Probability
Response at BMD
O Data
BMD
BMDL
0 50 100 150 200
Dose
Figure B-7. Dose response curve for the Linear model fit to hemoglobin in
female rats (Klaunig et al.. 2015).
This document is a draft for review purposes only and does not constitute Agency policy.
B-15 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.9. RED BLOOD CELLS-MALE RATS fCHENGELIS ET AL.. 2009B1
Table B-15. Dose response data for red blood cells in male rats (Chengelis et
aL_2009b)
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
8.89
0.32
10
10
8.84
0.281
50
10
8.88
0.69
200
10
8.17
0.593
Table B-16. Benchmark dose results for red blood cells in male
rats—nonconstant variance, BMR = 1 standard deviation (Chengelis etal..
2009b)
Models
p-Values
Test 3
p-values
AIC
1SD
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.211487
0.046614
60.49036
113.155
66.64235
0.93824
Exponential 3
0.211488
0.046614
60.49034
113.324
66.64082
0.936908
Exponential 4
0.112482
0.046614
61.90217
63.88893
16.43649
1.512764
Exponential 5
0.123707
0.046614
61.75292
51.86424
17.01699
1.668664
Hill
NA
0.046614
61.38555
49.50692
16.01355
1.522475
Polynomial (Poly 3)
0.208929
0.046614
60.51469
115.2832
69.56043
0.914633
Polynomial (Poly 2)
0.208929
0.046614
60.51469
115.2939
69.56068
0.914397
Power
0.208929
0.046614
60.51469
115.2866
69.56292
0.914574
Linear
0.208929
0.046614
60.51469
115.2954
69.55948
0.914492
Both constant and nonconstant variance models failed to model the variance data.
This document is a draft for review purposes only and does not constitute Agency policy.
B-16 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Figure B-8. Dose response data for red blood cells in male rats (Chengelis et
aL_2009b).
This document is a draft for review purposes only and does not constitute Agency policy.
B-17 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.10. RED BLOOD CELLS-FEMALE RATS fCHENGELIS ET AL.. 2009B1
Table B-17. Dose response data for red blood cells in female rats (Chengelis et
aL_2009b)
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
8.62
0.338
10
10
8.53
0.696
50
10
8.32
0.491
200
10
7.93
0.43
Table B-18. Benchmark dose results for red blood cells in female
rats—constant variance, BMR = 1 standard deviation (Chengelis et al.. 2009bl
Models
p-Values
Test 2
p-values
AIC
1SD
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.819031
0.13452
61.22185
145.9541
94.47522
0.13169
Exponential 3
0.819031
0.13452
61.22185
145.9541
94.47455
0.13169
Exponential 4
0.828537
0.13452
62.86949
112.0384
27.37312
-0.13653
Exponential 5
0.527493
0.13452
63.2218
145.9511
16.32358
0.131694
Hill
NA
0.13452
64.90674
95.16729
22.04822
0.034663
Polynomial Degree 3
0.805881
0.13452
61.25422
148.2376
97.83829
0.128637
Polynomial Degree 2
0.805881
0.13452
61.25422
148.2285
97.83846
0.128826
Power
0.805881
0.13452
61.25422
148.2268
97.80444
0.128858
Linear
0.805881
0.13452
61.25422
148.2178
97.81736
0.129033
This document is a draft for review purposes only and does not constitute Agency policy.
B-18 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Exponential Degree 5 Model with BMR of 1 Std. Dev.
for the BMD and 0.95 Lower Confidence Limit for the BMDL
10
9 ~ i
ill
0
/
Si 6
Estimated Probability
c
° s
Response at BMD
Q. ^
tn
CD A
o
O Data
9
BMD
z
1
BMDL
±
n
u
0
50 100 150 200
Dose
Figure B-9. Dose response curve for the Exponential 5 model fit to red blood
cells in female rats (Chengelis etal.. 2009b).
This document is a draft for review purposes only and does not constitute Agency policy.
B-19 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.ll. RED BLOOD CELLS-MALE RATS fLOVELESS ET AL.. 20091
Table B-19. Dose response data for red blood cells in male rats (Loveless et al..
2009)
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
8.89
0.36
20
10
8.95
0.34
100
10
8.46
0.41
500
10
6.09
1.27
Table B-20. Benchmark dose results for red blood cells in male
rats—nonconstant variance, BMR = 1 standard deviation (Loveless et al.. 20091
Models
p-Values
Test 3
p-values
AIC
1SD
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.281567
0.991476
64.79171
52.64163
38.9282
0.746143
Exponential 3
0.376218
0.991476
65.03997
78.0673
43.76706
-0.23387
Exponential 4
0.281572
0.991476
64.79167
52.63495
38.92813
0.744847
Exponential 5
NA
0.991476
66.45382
91.34257
46.77432
-0.01779
Hill
NA
0.991476
66.44302
97.70618
94.33382
-0.01642
Polynomial (Poly 3)
0.291705
0.991476
65.36868
73.55976
45.76816
-0.24307
Polynomial (Poly 2)
0.291695
0.991476
65.36872
73.60792
45.76059
-0.24304
Power
0.341547
0.991476
65.16156
77.54244
46.28623
-0.27696
Linear
0.445951
0.991476
63.87203
59.08585
44.57007
0.743535
This document is a draft for review purposes only and does not constitute Agency policy.
B-20 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Linear Model with BMR of 1 Std. Dev. for the BMD
and 0.95 Lower Confidence Limit for the BMDL
100
200 300
Dose
400
^^Estimated Probability
^^Response at BMD
O Data
BMD
BMDL
500
Figure B-10. Dose response curve for the Linear model fit to red blood cells in
male rats (Loveless etal.. 2009).
This document is a draft for review purposes only and does not constitute Agency policy.
B-21 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.12. RED BLOOD CELLS-FEMALE RATS fLOVELESS ET AL.. 20091
Table B-21. Dose response data for red blood cells in female rats (Loveless et
al.. 20091
Dose (mg/kg-day)
Number of
animals
Mean
(million/pL)
Standard deviation
0
10
8.34
0.43
20
10
8.53
0.52
100
10
8.32
0.27
500
9
6.85
0.63
Table B-22. Benchmark dose results for red blood cells in female
rats—constant variance, BMR = 1 standard deviation (Loveless et al.. 20091
Models
p-Values
Test 2
p-values
AIC
1SD
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.21884
0.087567
57.58768
133.0328
102.9324
1.002642
Exponential 3
0.331861
0.087567
57.49047
238.0109
116.9504
-0.08346
Exponential 4
0.21884
0.087567
57.58768
133.0037
102.9322
1.002848
Exponential 5
0.331861
0.087567
57.49047
238.0095
116.9523
-0.08345
Hill
NA
0.087567
59.42671
113.2878
101.18
-2.4E-06
Polynomial Degree 3
0.320732
0.087567
57.53481
261.7164
122.0763
-0.18275
Polynomial Degree 2
0.320735
0.087567
57.5348
261.8718
122.0761
-0.1845
Power
0.330478
0.087567
57.49587
243.0686
122.3971
-0.09028
Linear
0.268591
0.087567
57.17798
142.5548
112.3638
0.87655
This document is a draft for review purposes only and does not constitute Agency policy.
B-22 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
1 n
Frequentist Linear Model with BMR of 1 Std. Dev. for the BMD
and 0.95 Lower Confidence Limit for the BMDL
-LU
Q
8 ®
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.13. HEPATOCELLULAR HYPERTROPHY-MALE RATS fCHENGELIS ET
AL.. 2009B1
Table B-23. Dose response data for hepatocellular hypertrophy in male rats
(Chengelis etal.. 2009b)
Dose (mg/kg-day)
Number of
animals
Incidence
Percentage of incidence
0
10
0
0
10
10
0
0
50
10
0
0
200
10
7
70
This data set is not considered appropriate for BMD modeling because there is a single dose group showing a high
incidence response (70%) in contrast to no response in all other groups.
Hepatocellular Hypertrophy, Centrilobular
Figure B-12. Dose response data for hepatocellular hypertrophy in male rats
(Chengelis etal.. 2009b).
This document is a draft for review purposes only and does not constitute Agency policy.
B-24 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.14. HEPATOCELLULAR HYPERTROPHY-FEMALE RATS fLOVELESS ET
AL.. 20091
Table B-24. Dose response data for hepatocellular hypertrophy in female rats
(Loveless et al.. 2009)
Dose (mg/kg-day)
Number of
animals
Incidence
Percentage of incidence
0
10
0
0
20
10
0
0
100
n
0
0
500
10
5
50
Table B-25. Benchmark dose results for hepatocellular hypertrophy in female
rats—nonconstant variance, BMR = 10% Extra Risk (Loveless et al.. 2009)
10% Extra Risk
Scaled
residual
for dose
Models
p-Values
AIC
BMD
BMDL
group
near BMD
Dichotomous Hill
l
17.86294
405.8782
97.0233
1.22E-07
Gamma
0.98176
19.86399
319.4271
97.38679
0.000597
Log-Logistic
1
17.86294
442.544
96.1137
-1.2E-09
Multistage Degree 3
0.996008
15.98505
267.9741
96.31573
-0.24575
Multistage Degree 2
0.956334
16.4752
201.2327
82.64841
-0.53848
Multistage Degree 1
0.610724
18.92693
104.2041
53.56868
-1.08184
Weibull
1
17.86294
449.1777
97.95169
-1.1E-08
Logistic
1
15.86296
438.966
209.1724
4.68E-07
Log-Probit
1
17.86294
402.6258
93.01619
7.1E-07
Probit
1
17.86294
411.492
185.6167
1.31E-09
This document is a draft for review purposes only and does not constitute Agency policy.
B-25 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Multistage Degree 3 Model with BMR of 10% Extra
Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
i
0.9
Dose
Figure B-13. Dose response curve for the Multistage Degree 3 model fit to
hepatocellular hypertrophy in female rats (Loveless et al.. 2009).
This document is a draft for review purposes only and does not constitute Agency policy.
B-26 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.15. HEPATOCELLULAR HYPERTROPHY-MALE RATS fLOVELESS ET AL..
20091
Table B-26. Dose response data for hepatocellular hypertrophy in male rats
(Loveless et al.. 2009)
Dose (mg/kg-day)
Number of
animals
Incidence
Percentage of incidence
0
10
0
0
20
10
0
0
100
10
4
40
500
10
10
100
Table B-27. Benchmark dose results for hepatocellular hypertrophy in male
rats—nonconstant variance, BMR = 10% Extra Risk (Loveless et al.. 2009)
10% Extra Risk
Scaled
residual
for dose
Models
p-Values
AIC
BMD
BMDL
group
near BMD
Dichotomous Hill
l
17.46023
85.47371
28.3855
-1.1E-06
Gamma
0.999944
17.46046
70.57884
20.71965
0.00025
Log-Logistic
1
15.46023
85.49796
28.38513
2.91E-07
Multistage Degree 3
0.997823
15.54154
59.28867
16.83509
-0.20131
Multistage Degree 2
0.902071
17.8587
46.58058
16.60448
-0.44287
Multistage Degree 1
0.391117
20.9779
18.16542
10.6581
-1.10904
Weibull
0.987025
17.51174
62.10697
19.73504
0.025832
Logistic
0.999997
17.46024
89.81641
41.88635
4.78E-05
Log-Probit
1
17.46023
78.71963
26.71976
-7.5E-12
Probit
0.999765
15.47853
71.58692
37.71366
0.012573
This document is a draft for review purposes only and does not constitute Agency policy.
B-27 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Multistage Degree 1 Model with BMR of 10% Extra
Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
i ©
0 100 200 300 400 500
Dose
Figure B-14. Dose response curve for the Multistage Degree 1 model fit to
hepatocellular hypertrophy in male rats (Loveless et al.. 2009).
This document is a draft for review purposes only and does not constitute Agency policy.
B-28 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.16. POSTNATAL (Fi) COMBINED RAT BODY WEIGHT ON PND 0
fLOVELESS ET AL.. 20091
Table B-28. Dose response data for postnatal (Fl) combined rat body weight
on PND 0 (Loveless et al.. 2009)
Dose (mg/kg-day)
Number of
animals
Mean (g)
Standard deviation
0
20
7.1
0.9
20
20
6.8
0.6
100
20
6.3
0.4
500
20
5.8
0.4
Table B-29. Benchmark dose results for postnatal (Fl) combined rat body
weight on PND 0—nonconstant variance, BMR = 5% relative deviation
(Loveless et al.. 2009)
Models
p-Values
Test 3
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.000613
0.257697
150.4828
154.17
126.6598
-2.10808
Exponential 3
0.000613
0.257697
150.4828
154.2311
126.6606
-2.10618
Exponential 4
0.417875
0.257697
138.3442
28.86879
18.04413
-0.30628
Exponential 5
0.417869
0.257697
138.3442
28.89287
18.02549
-0.3069
Hill
0.721731
0.257697
137.8148
20.37779
10.61916
0.013748
Polynomial (Poly 3)
0.000461
0.257697
151.0527
164.7639
137.82
-2.14368
Polynomial (Poly 2)
0.000461
0.257697
151.0527
164.763
137.8213
-2.144
Power
0.000461
0.257697
151.0527
164.7277
137.8381
-2.14471
Linear
0.000461
0.257697
151.0527
164.7482
137.8256
-2.14516
This document is a draft for review purposes only and does not constitute Agency policy.
B-29 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Hill Model with BMR of 0.05 Rel. Dev. for the BMD
and 0.95 Lower Confidence Limit for the BMDL
^ 5
°4
LO
DC 3
^^Estimated Probability
^^Response at BMD
O Data
BMD
BMDL
100
200 300
Dose
400
500
Figure B-15. Dose response curve for the Hill model fit to postnatal (Fl)
combined rat body weight on PND 0 (Loveless et al.. 20091.
This document is a draft for review purposes only and does not constitute Agency policy.
B-30 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.17. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 2) ON
PND 0 flWAI AND HOBERMAN. 20141
Table B-30. Dose response data for postnatal (Fl) combined mouse body
weight (Phase 2) on PND 0 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
20
1.562
0.120
7
17
1.561
0.119
35
19
1.579
0.115
175
20
1.447
0.180
Table B-31. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 2) on PND 0—constant variance, BMR = 5% relative deviation
(Iwai and Hoberman. 2014)
Models
p-Values
Test 2
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential
0.5476652
0.11356
-83.22986065
110.1988
72.6152
-0.18098
ExponentiaB
0.6427413
0.11356
-82.21886799
162.9802
78.154
-0.00108
Exponential
0.5476662
0.11356
-83.22986423
110.2315
72.6152
-0.18210
Exponential
0.6427936
0.11356
-82.21893566
163.6378
78.15859
-0.00024
Hill
NA
0.11356
-80.21900716
80.26504
36.86639
0.37613
Polynomial (Poly 3)
0.971011
0.11356
-86.19475647
151.5619
80.06441
-0.00309
Polynomial (Poly 2)
0.8402282
0.11356
-84.08587922
140.6661
79.398
-0.018554
Power
0.6428503
0.11356
-82.21900901
172.0405
121.5756
2.26045E-
05
Linear
0.5601161
0.11356
-83.27482038
111.6004
75.16344
-0.16700
This document is a draft for review purposes only and does not constitute Agency policy.
B-31 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Polynomial Degree 3 Model with BMR of 0.05 Rel.
Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
(D
to
c
o
w o
(D
* o
0,
0,
W
•$>
20
40
60
80 100
Dose
120
140
160
^^Estimated Probability
^^Response at BMD
O Data
BMD
BMDL
Figure B-16. Dose response curve for the Polynomial Degree 3 model fit to
postnatal (Fl) combined rat body weight (Phase 2) on PND 0 (Iwai and
Hoberman. 20141. X-axis is dose (mg/kg-day) and y-axis is mean body weight (g).
This document is a draft for review purposes only and does not constitute Agency policy.
B-32 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.18. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 1) ON
PND 0 flWAI AND HOBERMAN. 20141
Table B-32. Dose response data for postnatal (Fl) combined mouse body
weight (Phase 1) on PND 0 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
19
1.597
0.166
100
19
1.484
0.100
350
19
1.365
0.237
500
13
1.396
0.187
Table B-33. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 1) on PND 0—nonconstant variance, BMR = 5% relative
deviation (Iwai and Hoberman. 2014)
Models
p-Values
Test 3
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential 2
0.1454254
0.01314
-38.99028302
153.0166
106.9649
-0.9649
Exponential 3
0.1454276
0.01314
-38.99031401
152.9732
106.9641
-0.9649
Exponential 4
0.0502847
0.01314
-37.01452559
152.2536
22.05564
-0.9633
Exponential 5
NA
0.01314
-38.08812965
101.2731
78.25327
-0.6831
Hill
NA
0.01314
-38.08803429
100.2818
93.46723
-0.6814
Polynomial (Poly 3)
0.1237777
0.01314
-38.66793064
163.1923
116.9646
-0.9923
Polynomial (Poly 2)
0.1237777
0.01314
-38.66793064
163.1927
116.9612
-0.9923
Power
0.1237777
0.01314
-38.66793064
163.1924
116.9832
-0.9923
Linear
0.1237777
0.01314
-38.66793064
163.1923
117.1098
-0.9923
Both constant and nonconstant models failed to model the variance data.
This document is a draft for review purposes only and does not constitute Agency policy.
B-33 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.19. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASES 1 AND
2) ON PND 0 flWAI AND HOBERMAN. 20141
Table B-34. Dose response data for postnatal (Fl) combined mouse body
weight (Phases 1 and 2) on PND 0 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
27
1.577
0.154
7
17
1.561
0.119
35
19
1.579
0.115
175
20
1.447
0.180
100
19
1.484
0.100
350
19
1.365
0.237
500
13
1.396
0.187
Table B-35. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 1 and 2) on PND 0—nonconstant variance, BMR = 5% relative
deviation (Iwai and Hoberman. 2014)
Models
p-Values
Test 3
p-values
AIC
5% relative deviation
Scaled residual
for dose group
near BMD
BMD
BMDL
Exponential
<0.0001
<0.0001
-113.7083011
143.1894
105.9785
-0.7569
Exponential
<0.0001
<0.0001
-113.7082872
143.218
105.9774
-0.7574
Exponential
<0.0001
<0.0001
-111.765811
142.2114
45.41936
-0.7464
Exponential
<0.0001
<0.0001
-114.9537952
124.9707
78.92966
-1.0491
Hill
<0.0001
<0.0001
-114.8291753
120.1336
86.39634
-0.9939
Polynomial (Poly 6)
<0.0001
<0.0001
-113.1738765
151.9313
116.5059
-0.8482
Polynomial (Poly 5)
<0.0001
<0.0001
-113.1738878
151.8383
114.2464
-0.8469
Polynomial (Poly 4)
<0.0001
<0.0001
-113.1738762
151.9337
114.3852
-0.8482
Polynomial (Poly 3)
<0.0001
<0.0001
-113.1738765
151.9313
114.3477
-0.8482
Polynomial (Poly 2)
<0.0001
<0.0001
-113.1738765
151.9313
114.3472
-0.8482
Power
<0.0001
<0.0001
-113.1738821
151.8842
114.494
-0.8473
Linear
<0.0001
<0.0001
-113.1738823
151.8741
114.3477
-0.8474
3oth constant and nonconstant models failec
to model the variance data.
This document is a draft for review purposes only and does not constitute Agency policy.
B-34 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.20. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 2) ON
PND 4 flWAI AND HOBERMAN. 20141
Table B-36. Dose response data for postnatal (Fl) combined mouse body
weight (Phase 2) on PND 4 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
20
2.844
0.307
7
16
2.850
0.320
35
19
2.976
0.335
175
20
2.726
0.442
Table B-37. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 2) on PND 4—constant variance, BMR = 5% relative deviation
(Iwai and Hoberman. 2014)
Models
p-Values
Test 2
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential
0.2719642
0.3216
62.91273812
169.116
79.86226
-0.259556406
ExponentiaB
0.1915765
0.3216
64.01402032
171.7121
88.6223
0.000775748
Exponential
0.2719642
0.3216
62.91273812
169.1154
79.86194
-0.259556232
Exponential
0.1915772
0.3216
64.01401491
171.7578
88.62246
0.000965695
Hill
0.191581
0.3216
64.01398562
90.59589
37.60928
1.049750693
Polynomial (poly 3)
0.6280973
0.3216
60.04847961
167.4876
89.7897
-0.008262428
Polynomial (poly 2)
0.3908438
0.3216
62.1874628
164.7988
88.29103
-0.042810195
Power
0.1915812
0.3216
64.01398451
174.0668
106.2633
4.3382E-06
Linear
0.2746896
0.3216
62.89279543
168.0092
81.96061
-0.247433913
This document is a draft for review purposes only and does not constitute Agency policy.
B-35 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
3.5
3
Frequentist Polynomial Degree 3 Model with BMR of 0.05 Rel.
Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
2.5
9
-4>
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.21. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASE 1) ON
PND 4 flWAI AND HOBERMAN. 20141
Table B-38. Dose response data for postnatal (Fl) combined mouse body
weight (Phase 1) on PND 4 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
18
2.966
0.460
100
19
2.771
0.248
350
17
2.256
0.650
500
11
2.382
0.482
Table B-39. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 1) on PND 4—nonconstant variance, BMR = 5% relative
deviation (Iwai and Hoberman. 2014)
Models
p-Values
Test 3
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential
0.0805287
0.01040
90.85546713
84.46765
60.74916
-0.0400
ExponentiaB
0.0805258
0.01040
90.85553976
84.40685
60.76206
-0.0404
Exponential
0.0358662
0.01040
92.22063699
59.93938
28.42237
0.3237
Exponential
NA
0.01040
90.99580479
113.1456
55.45824
-0.6168
Hill
NA
0.01040
90.99580314
102.7811
95.12445
-0.6171
Polynomial (poly 3)
0.064938
0.01040
91.28582701
94.9049
70.88712
-0.1219
Polynomial (poly 2)
0.064938
0.01040
91.28582701
94.90514
70.88731
-0.1219
Power
0.064938
0.01040
91.28582692
94.90368
70.88898
-0.1219
Linear
0.064938
0.01040
91.28582698
94.90395
70.88785
-0.1220
Both constant and nonconstant models failed to model the variance data.
This document is a draft for review purposes only and does not constitute Agency policy.
B-37 DRAFT-DO NOT CITE OR QUOTE
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.22. POSTNATAL (Fi) COMBINED MOUSE BODY WEIGHT (PHASES 1 AND
2) ON PND 4 flWAI AND HOBERMAN. 20141
Table B-40. Dose response data for postnatal (Fl) combined mouse body
weight (Phase 1) on PND 4 (Iwai and Hoberman. 2014)
Dose (mg/kg-day)
Number of litters
Mean (g)
Standard deviation
0
38
2.902
0.387
7
16
2.85
0.320
35
19
2.976
0.335
175
20
2.726
0.442
100
19
2.771
0.248
350
17
2.256
0.650
500
11
2.382
0.482
Table B-41. Benchmark dose results for postnatal (Fl) combined mouse body
weight (Phase 1 and 2) on PND 4—nonconstant variance, BMR = 5% relative
deviation (Iwai and Hoberman. 2014)
Models
p-Values
Test 3
p-values
AIC
5% relative deviation
Scaled
residual
for dose
group
near BMD
BMD
BMDL
Exponential
0.1128908
0.2000
147.7837189
96.33303
71.43861
-0.0192
ExponentiaB
0.0802807
0.2000
149.2060003
120.9626
73.66277
-0.3003
Exponential
0.1128924
0.2000
147.78368
96.31896
71.43858
-0.0189
Exponential
0.4079141
0.2000
145.7743154
155.2243
103.1196
0.4065
Hill
0.3365496
0.2000
146.2589902
165.6672
139.2494
0.3173
Polynomial (poly 6)
0.110707
0.2000
147.8372495
103.2643
78.89672
-0.0983
Polynomial (poly 5)
0.110707
0.2000
147.8372495
103.2643
79.3315
-0.0983
Polynomial (poly 4)
0.110707
0.2000
147.8372495
103.2643
78.87018
-0.0983
Polynomial (poly 3)
0.110707
0.2000
147.8372495
103.2643
78.84946
-0.0983
Polynomial (poly 2)
0.110707
0.2000
147.8372495
103.2643
78.84795
-0.0983
Power
0.0672452
0.2000
149.6433094
118.3717
79.54891
-0.2558
Linear
0.110707
0.2000
147.8372495
103.2643
78.84954
-0.0983
This document is a draft for review purposes only and does not constitute Agency policy.
B-38 DRAFT-DO NOT CITE OR QUOTE
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Frequentist Exponential Degree 5 Model with BMR of 0.05 Rel.
Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
3.5
3
2.5
-------
Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.23. PERINATAL MORTALITY (PHASE 2) ON PND 0-21 flWAI AND
HOBERMAN. 20141
Table B-42. Nested Model summary for perinatal mortality (Phase 2) on PND
0-21, BMR = 1% extra risk (Iwai and Hoberman. 2014)
Model type
Litter-specific
covariate
Intralitter
correlation
Goodness
of fit p-
value
AIC
BMD
BMDL
Nlogistic
Yes
Yes
0.223
145.10
150.6
24.50
Yes
No
0.0037
158.29
157.6
39.26
No
Yes
0.2113
141.20
151.4
24.77
No
No
0.003
154.50
158.0
39.03
NLogistic model with intralitter correlation is selected as the best model based the lowest AIC value.
C
o
Q.
tn
(D
CC
Frequentist Nested Logistic Model with BMR of 0.01 Std. Dev. for
the BMD and 0.95 Lower Confidence Limit for the BMDL
0.14
0.12
0.1
0.08
0.06
0.04
0.02
0
20
40
60
80 100
Dose
120
140
160
^^Estimated Probability
^^Response at BMD
O Data
BMD
BMDL
Figure B-19. Dose response curve for the Nested NCTR model fit to perinatal
mortality (Phase 2) on PND 0-21 (Iwai and Hoberman. 2014). X-axis is dose
(mg/kg-day) andy-axis is mortality.
This document is a draft for review purposes only and does not constitute Agency policy.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.24. PERINATAL MORTALITY (PHASE 1) ON PND 0-21 flWAI AND
HOBERMAN. 20141
Table B-43. Nested model summary for perinatal mortality (Phase 1) on PND
0-21, BMR = 1% extra risk (Iwai and Hoberman. 2014)
Model type
Litter-specific
covariate
Intralitter
correlation
Goodness
of fit p-
value
AIC
BMD
BMDL
Nlogistic
Yes
Yes
0.053
356.23
206.1
105.8
Yes
No
<0.0001
478.37
238.9
177.2
No
Yes
0.0593
353.33
201.7
98.61
No
No
<0.0001
477.04
233.1
162.7
Because none of the models provided adequate fit (goodness of fit p-value > 0.1) to Phases 1 data, no BMDL could
be estimated.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
B.25. PERINATAL MORTALITY (PHASES 1 AND 2) ON PND 0-21 flWAI
AND HOBERMAN. 20141
Table B-44. Nested model summary for perinatal mortality (Phase 1 and 2) on
PND 0-21, BMR = 1% extra risk (Iwai and Hoberman. 2014)
Model type
Litter-specific
covariate
Intralitter
correlation
Goodness
of fit p-
value
AIC
BMD
BMDL
Nlogistic
Yes
Yes
0.024
495.44
150.9
85.15
Yes
No
<0.0001
632.14
199.7
138.2
No
Yes
0.029
491.80
147.7
83.59
No
No
<0.0001
629.52
195.2
134.0
Because none of the models provided adequate fit (goodness of fit p-value > 0.1) to Phases 1 data, no BMDL could
be estimated.
This document is a draft for review purposes only and does not constitute Agency policy.
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APPENDIX C. EVALUATION OF PFHXA
ELIMINATION
C.l. EVALUATION OF PFHXA ELIMINATION IN RATS AND MICE
Pharmacokinetic parameters were estimated separately for male and female rats and mice
using a hierarchical, Bayesian framework to allow for the partial pooling of time-course
concentration data across multiple studies. Data extracted from the studies described above were
fit to the following model formulation, which describes the absorption (when necessary),
distribution, and elimination phase of PFHxA through a two-compartment pharmacokinetic model:
Ct = absflagii(-Ai - BOe-boW* + + B^ (C-l)
Here, / represents the ith compartment for PFHxA measurement (e.g., plasma, liver, kidney).
Ai and £?, represent the ratio of chemical mass going to each empirical compartment, normalized by
the central compartment volume, resulting in units of PFHxA concentration. For PFHxA
concentrations measured in the plasma (i.e., central compartment) following intravenous (i.v.)
exposure, absfiag,i\s set to zero to remove the absorption term.
Conventionally, each compartment with pharmacokinetic data is fit independently to
equation C-l and tissue-specific half-lives for each species and sex are derived from the estimated p,
i.e., ti/2,1 = ln(2)//?;. However, when a compound is in the elimination phase, (3 should be constant
across all tissues. To determine this PFHxA-specific /3 and use the time-course concentration data
from every study across multiple compartments, a partial pooling of data in a hierarchical Bayesian
framework assumes that, although /?, differs for each tissue, they are all sampled from a common
group distribution. Following completion of the Markov-chain Monte Carlo, the top-level posterior
distribution of /3 is used to determine the median PFHxA half-life, with uncertainty, for each
species/sex. The remaining study-level coefficients are used to estimate the additional
pharmacokinetic values, for example, area under the curve (AUCmf), clearance (CL), volume of
distribution (Vdp).
Along with the half-life analysis, a separate distribution of CL = dose/AUCinf and Vdp = CL/(3
is generated for each experiment (study/route/dose/sex), where AUCmf is obtained by integrating
equation (C-l) from time = 0 to in infinity, to yield
AUCinfi=^ + ^-abSfla3'i(Ai+Bi) (C-2)
lnf'1 at ft kabSjl 1 J
Median and 5th and 95th percentiles of the distributions for t^i, CLt and Vdp are then pooled across
each study/route/dose to calculate the species- and sex-dependent values.
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C.l.l. Mice
Data for male and female mice were obtained from Gannon etal. f20111 who evaluated the
pharmacokinetics after single oral doses of 2 and 100 mg/kg. Original data files were provided by
Shawn Gannon, The Chemours Company, Wilmington, Delaware (personnel communication).
Although the data for the 2 mg/kg dose appeared appropriately censored below the dose-specific
limit of quantification (LOQ), the 100 mg/kg data appeared to reach a plateau just above the
corresponding LOQ (~0.25 [ig/g plasma), in a concentration range for which clearance after the
2 mg/kg dose was quite rapid. EPA interpreted this result as indicating an interfering background
signal. For this reason, only data with measured concentration >0.5 |ig/g plasma were used for the
100 mg/kg dose. The resulting statistics for the elimination half-lives (90% confidence interval)
are 2.8 hours (1.0-7.0 hours) and 6.7 hours (2.2-16 hours) for females and males, respectively.
Female mouse data were from Daikin Industries (2010). who exposed groups of mice to 35,
175, or 350 mg/kg PFHxA by oral gavage and measured serum concentration at time-points up to
24 h. Because three separate mice were analyzed at each time point, means and standard
deviations were calculated and used for statistical modeling. Data at the first time points with
concentrations below the lower limit of quantification (LLOQ) were assigned a value of LLOQ/V2
for the purpose of computing the means. Specifically, for the 24-h time point, two of three animals
in the 175 and 350 mg/kg dose groups had results
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
Gannon Male 1
2 mg/kg Gavage dose
Gannon Male 2
100 mg/kg Gavage dose
5 10 15 20 25
time [hrs]
Gannon Female 1
2 mg/kg Gavage dose
5 10 15 20 25
time [hrs]
Gannon Female 2
100 mg/kg Gavage dose
5 10 15 20 25
time [hrs]
Daikin Female 1
35 mg/kg Gavage dose
5 10 15 20 25
time [hrs]
Daikin Female 2
175 mg/kg Gavage dose
iO2!
[A
1 102i
\\\
E
cn
g
E, 10° i
u
-——
u
8 10-1]
v s
c
o
u
(O
E 10-2,
ro
£
(O
jC
Pi
M
O
Q.
O
i—1
1 ! 1 1
5 10 15 20 25
time [hrs]
Daikin Female 3
350 mg/kg Gavage dose
5 10 15 20 25
time [hrs]
—- 90% C.I.
— median
\ in vivo data
5 10 15 20 25
time [hrs]
Figure C-l. Fits of population pharmacokinetic model to data for male (top
row) and female (remaining rows) mice following 2-350 mg/kg oral exposure
PFHxA.
Source: Data from Gannon et al. (2011) and Daikin Industries (2010).
This document is a draft for review purposes only and does not constitute Agency policy,
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C.1.2. Rats
PFHxA the following PK data for male and female rats were evaluated:
• Chengelis etal. (2009a): male and female Sprague-Dawley rats exposed once by
intravenous injection (i.v.; 10 mg/kg) or by single-day or Day 25 of repeated gavage (50,
150, or 300 mg/kg). (i.v. data for males and females and oral data for males are provided in
published tables. Oral data for females were obtained by digitizing the plot of single-day
exposure data. The 25-day female rat data, however, were not digitized or used because the
digitization process has some uncertainty; reported dose-specific half-lives for females
were quite similar for the single- and 25-day studies, and results for males were similar
with and without the 25-day data.)
• Dzierlenga et al. (2019): male and female Sprague-Dawley rats exposed by i.v. (40 mg/kg)
or by gavage (40, 80, or 160 mg/kg; data from National Toxicology Program website).
• Gannon etal. (2011): male and female Sprague-Dawley rats exposed by gavage (2 or
100 mg/kg; data from study authors).
• Iwabuchi etal. (2017): male Wistar rats exposed by gavage (0.1 mg/kg; data from published
tables or digitized from figures).
The resulting statistics for the elimination half-lives, clearance values, and volumes of
distribution (with 90% confidence intervals) are listed in Table 5-3 (Section 5.2.1, Approach for
Animal-Human Extrapolation ofPFHxA Dosimetry).
Data for human PFHxA analysis were extracted from Nilsson etal. (2013) where PHFxA
concentrations were measured in the blood of ski wax technicians exposed to PFAS compounds
over the course of multiple ski seasons. Because timing of the initial PFHxA exposure and the
resulting absorption kinetics are unknown for this population, we fit a one-compartment infusion
pharmacokinetic model to the reported time-course data:
Here, / represents the ith ski wax technician and tinp represents the time at which exposure
to PFHxA ends. All other model parameters are the same as described above for the rat and mouse
fits. Briefly, this model assumes a constant exposure to PFHxA throughout the ski season when
time is less than t,„/. Once t,„/is reached, PFHxA is eliminated under a first order elimination
assumption.
C.2. EVALUATION OF PFHXA ELIMINATION IN HUMANS
(C-3)
This document is a draft for review purposes only and does not constitute Agency policy.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
1 Similar to the methods described for the rat and mouse, /?, for each ski wax technician is
2 sampled hierarchically from a population distribution while all other parameters in the model are
3 fit only to the individual technician. Finally to use limit of detection (LOD) data reported in this
4 study we implemented a left-censored likelihood function in the Bayesian inference model for
5 samples reported below the LOD (<0.05 ng/mL). This ensured that the likelihood function for these
6 data were sampled only from a probability distribution with an upper bound at the LOD.
7 Results for each ski wax technician are shown below following sampling of the technician-
8 specific posterior distributions. Technician half-lives (90% credible interval) are presented in the
9 panel for each technician with the population half-life determined to be 13.78 (5.51-32.86) days.
10 Technicians 1-8 represent data from the 2007-08 ski season, when samples were taken late enough
11 in the spring to allow quantification of post-exposure clearance.
Population half-life (days): 11.45 (6.06 - 21.21)
half-life (days): 8.75 (3.21 - 17.06)
101
10-J
8 lo-3
S 10"a
10"11
ZZZlZLZ^
• -fechl
A "fechl-cens
2 4 6 8 10
Months since Sept
half-life (days): 13.67 (6.87 - 25.77)
S 10-2
E 10-3
- —
• -fech2
A "fech2-cens
2 4 6 8 10
Months since Sept
half-life (days): 8.63 (2.98 - 16.74)
10"1
0 10"4
t-
©
ri 10-7
2 10-1O
10"13
-*
• "fech3
A "fech3-cens
2 4 6 8 10
Months since Sept
half-life (days): 9.88 (3.61 - 19.69)
• "fech4
A "fech4-cens
2 4 6 8 10
Months since Sept
half-life (days): 14.04 (5.10 - 59.02)
101
10"3
£ 10"3
10"'
• "fech5
~ "fech5-cens
4 6 8 10
Months since Sept
half-life (days): 12.84 (5.10 - 31.40)
• "fech6
~ Tech6-cens
half-life (days): 12.60 (4.36 - 42.52)
4 6 8 10
Months since Sept
10"1
j 10"3
• "fech7
~ "fech7-cens
half-life (days): 13.56 (5.49 - 33.06)
4 6 8 10
Months since Sept
M
• "fech8
A Tech8-cens
4 6 8 10
Months since Sept
Figure C-2. Fits of human PFHxA data from ski-wax technician blood samples.
Blue circles represent data above LOD while black triangles are data samples
reported at the LOD (<0.05 ng/mL). 90% credible intervals are illustrated with the
light blue bands and dashed lines.
This document is a draft for review purposes only and does not constitute Agency policy.
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APPENDIX D. QUALITY ASSURANCE FOR THE IRIS
TOXICOLOGICAL REVIEW OF PFHXA
This assessment is prepared under the auspices of the U.S. Environmental Protection
Agency's (EPA's) Integrated Risk Information System (IRIS) Program. The IRIS Program is housed
within the Office of Research and Development (ORD) in the Center for Public Health and
Environmental Assessment (CPHEA). EPA has an agency-wide quality assurance (QA) policy
outlined in the EPA Quality Manual for Environmental Programs (see CIO 2105-P-01.11 and follows
the specifications outlined in EPA Order CIO 2105.1.
As required by CIO 2105.1, ORD maintains a Quality Management Program, which is
documented in an internal Quality Management Plan (QMP). The latest version was developed in
2013 using Guidance for Developing Quality Systems for Environmental Programs fOA/G-11. A
National Center for Environmental Assessment (NCEA)/CPHEA-specific QMP also was developed in
2013 as an appendix to the ORD QMP. Quality assurance for products developed within CPHEA is
managed under the ORD QMP and applicable appendices.
The IRIS Toxicological Review ofPFHxA is designated as Influential Scientific Information
(ISI) and is classified as QA Category A. Category A designations require reporting of all critical QA
activities, including audits. The development of IRIS assessments is done through a seven-step
process. Documentation of this process is available on the IRIS website:
https://www.epa.gOv/iris/basic-information-about-integrated-risk-information-system#process.
Specific management of PFAS assessments is documented in a Programmatic Quality
Assurance Project Plan (PQAPP). APQAPP is developed using the EPA Guidance for Quality
Assurance Project Plans fOA/G-51. and the latest approved version is dated October 2020. All PFAS
assessments follow the PFAS PQAPP, and all assessment leads and team members are required to
receive QA training on the PFAS PQAPP. During assessment development, additional QAPPs may be
applied for quality assurance management. They include:
Title
Document number
Date
Program Quality Assurance Project
Plan (PQAPP) for PFAS Assessments
L-CPAD-0031652-QP-1-3
October 2020
An Umbrella Quality Assurance
Project Plan (QAPP) for Dosimetry
and Mechanism-Based Models
(PBPK)
L-CPAD-0032188-QP-1-2
December 2020
This document is a draft for review purposes only and does not constitute Agency policy.
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Title
Document number
Date
Quality Assurance Project Plan
(QAPP) for Enhancements to
Benchmark Dose Software (BMDS)
L- H E EAD-0032189-QP-1-2
September 2020
ICF-General Support of CPHEA
Human Health Assessment Activities
QAPP
L-CPAD-0031961-QP-1-2
April 2021
1 During assessment development, this project undergoes quality audits during assessment
2 development including:
Date
Type of audit
Major findings
Actions taken
August 2020
QA audit
None
None
July 2021
QA audit
None
None
3 During Step 3 and Step 6 of the IRIS process, the IRIS toxicological review is subjected to
4 external reviews by other federal agency partners, including the Executive Offices of the White
5 House. Comments during these IRIS process steps are available in the docket [insert chemical
6 docket number—make sure the comments are in the docket] on http: //www.regulations.gov.
This document is a draft for review purposes only and does not constitute Agency policy.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
APPENDIX E. SUMMARY OF PUBLIC COMMENTS
AND EPA'S DISPOSITION
This document is a draft for review purposes only and does not constitute Agency policy.
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Supplemental Information for the Toxicological Review ofPFHxA and Related Salts
REFERENCES
Chengelis. CP: Kirkpatrick. IB: Myers. NR: Shinohara. M: Stetson. PL: Sved. DW. (2009a).
Comparison of the toxicokinetic behavior of perfluorohexanoic acid (PFHxA) and
nonafluorobutane-1-sulfonic acid (PFBS) in cynomolgus monkeys and rats. Reproductive
Toxicology 27: 400-406. http: / /dx. doi. o r g/10.1016 /i. r epr otox. 2 0 0 9.01.013
Chengelis. CP: Kirkpatrick. IB: Radovskv. A: Shinohara. M. (2009b). A 90-day repeated dose oral
(gavage) toxicity study of perfluorohexanoic acid (PFHxA) in rats (with functional
observational battery and motor activity determinations). Reproductive Toxicology 27:
342-351. http://dx.doi.Org/10.1016/i.reprotox.2009.01.006
Crump. KS. (1995). Calculation of benchmark doses from continuous data. Risk Analysis 15: 79-89.
http://dx.doi.Org/10.llll/i.1539-6924.1995.tb00095.x
Daikin Industries (Daikin Industries Limited). (2010). Oral (gavage) acute pharmacokinetic study of
PFH ammonium salt (ammonium salt of perflourinated hexanoic acid) in mice. Osaka, Japan.
Dzierlenga. AL: Robinson. VG: Waidvanatha. S: Devito. MI: Eifrid. MA: Gibbs. ST. etal. (2019).
Toxicokinetics of perfluorohexanoic acid (PFHxA), perfluorooctanoic acid (PFOA) and
perfluorodecanoic acid (PFDA) in male and female Hsd:Sprague dawley SD rats following
intravenous or gavage administration. Xenobiotica 50: 1-11.
http://dx.doi.Org/10.1080/00498254.2019.1683776
Gannon. SA: Tohnson. T: Nabb. PL: Serex. TL: Buck. RC: Loveless. SE. (2011). Absorption,
distribution, metabolism, and excretion of [l-14C]-perfluorohexanoate ([14C]-PFHx) in rats
and mice. Toxicology 283: 55-62. http://dx.doi.Org/10.1016/j.tox.2011.02.004
Iwabuchi. K: Senzaki. N: Mazawa. D: Sato. I: Hara. M: Ueda. F. etal. (2017). Tissue toxicokinetics of
perfluoro compounds with single and chronic low doses in male rats. Journal of
Toxicological Sciences 42: 301-317. http://dx.doi.org/10.2131/its.42.301
Iwai. H: Hoberman. AM. (2014). Oral (gavage) combined developmental and perinatal/postnatal
reproduction toxicity study of ammonium salt of perfluorinated hexanoic acid in mice.
International Journal of Toxicology 33: 219-237.
http: //dx.doi.org/10.1177/1091581814529449
Klaunig. IE: Shinohara. M: Iwai. H: Chengelis. CP: Kirkpatrick. TB: Wang. Z. etal. (2015). Evaluation
of the chronic toxicity and carcinogenicity of perfluorohexanoic acid (PFHxA) in Sprague-
Dawley rats. Toxicologic Pathology 43: 209-220.
http://dx.doi.org/10.1177/0192623314530532
Loveless. SE: Slezak. B: Serex. T: Lewis. 1: Mukerii. P: O'Connor. TC. etal. (2009). Toxicological
evaluation of sodium perfluorohexanoate. Toxicology 264: 32-44.
http://dx.doi.Org/10.1016/i.tox.2009.07.011
Nilsson. H: Karrman. A: Rotander. A: van Bavel. B: Lindstrom. G: Westberg. H. (2013). Professional
ski waxers' exposure to PFAS and aerosol concentrations in gas phase and different particle
size fractions. Environmental Science: Processes & Impacts 15: 814-822.
http: / /dx. doi. or g/10.10 3 9 /c3 em3 0 73 9e
This document is a draft for review purposes only and does not constitute Agency policy.
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U.S. EPA (U.S. Environmental Protection Agency). (2012). Benchmark dose technical guidance.
(EPA/100/R-12/001). Washington, DC: U.S. Environmental Protection Agency, Risk
Assessment Forum, https: //www.epa.gov/risk/benchmark-dose-technical-guidance
This document is a draft for review purposes only and does not constitute Agency policy.
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