v»EPA
United States
Environmental Protection
Agency
Screening the Vast
PFAS Landscape:
In Vitro Toxicokinetic Testing and
LC-MS/MS Analysis
Marci Smeltz
U. S. Environmental
Office of Research
Presentation for Waters Southeast Mass Spectrometry Users Meeting
Tuesday, July 21, 2020
The views expressed in this presentation are
those of the author and do not necessarily reflect
the views or policies of the U.S. EPA
-------
vvEPA
United States
Environmental Protection
Agency
EPA Office of Research and Development
¦ Office of Research and Development (ORD) is the scientific research
arm of the EPA
¦ Research is conducted by ORD's four Centers
¦ 10 facilities across the USA
¦ Research is conducted by a combination of Federal scientists, contract
researchers, and postdoctoral, graduate student, and post-
baccalaureate trainees Research Triangle Park, NC
1
Athens, GA
-------
AEPA
United States
Environmental Protection
Agency
What are PFAS?
PFAS = per- and polyfluoroalkyl
substances
Man-made chemicals used in industry
and consumer products worldwide
since the 1950s
Repel water, resist heat, and protect
surfaces
F F F F F F F o
I I l l I l I 4-
F—C—C —C—C—C—C—C—C
l l I l l l I ^nL1
F F F F F F F OH
F F F F F F F F 0
l l l I l l l l ll
F-C-C-C-C-C-C-C-C-S-OH
I l l I l l l l ll
FFFF FFFFO
PFOA
PFOS
NOTHING STICKS TO
F F F
^ f T ,o
F-C-C-C-O-C-C/
i i 1 i hh
F F F F 0H
GenX
NH,
Elftuw!
Oliaei et al 2013, Environ Sci Pollut Res
2
-------
• Bioaccumulative
¦ Accumulate over time
¦ Absorption > elimination
3
vvEPA
United States
Environmental Protection
Agency
CDC 2017, Fourth Report on Human Exposure
to Environmental Chemicals
Why are PFAS receiv ng lots of attention?
• Widespread occurrence
¦ PFAS in 97% of American
population
¦ Even in arctic polar bears
-•-PFOS
-•-PFOA
H PFHxS
-•-PFNA
2000 2004 2006 2008 2010 2012 2014
• Persistence
¦ Carbon-fluorine bonds are
some of the strongest
¦ Little degradation in
environment
Gonzalez-Gaya et al 2019, Environ. Sci.: Processes Impacts
-------
vvEPA
United States
Environmental Protection
Agency
How many PFAS exist?
¦ More than 4700 PFAS recognized by OECD
-------
v»EPA
United States
Environmental Protection
Agency
How many PFAS exist?
More than 4700 PFAS recognized by OECD
As industry continues to invent, the number will increase
PFAS-Like Properties
List Details
Description: List consists of all [
substructure filter is designed to
j 1.1.lUlllLlllii imuari
Number of Chemicals: 6648
{rtv.ny*n«.nt#l Prot^C
AQ+TKy
-------
AEPA
United States
Environmental Protection
Agency
What's the CompTox Chemicals
Dashboard?
https://comptox.epa.gov/dashboard
One-stop-shop for chemical, toxicological, and exposure information
Almost 900,000 chemicals inventoried
&EPA
United Slates
Environmental Protection Home Advanced Se«di Botch Search late '
Agoncy
875 Thousand Chemicals
Product/Use Categories Assay/Gene
~ Identifier substring search
See what people are saying, read the dashboard comments!
Ctte the Dashboard Publication dick here
Latest News
Read more news
New list of dioxins and dioxin-like compounds added
December 18th, 2019 at 7 50:44 Ml
Dioxins and dioxin-like compounds (DLCs) are a well known class of compounds that are highly toxic envwonmentaJ persistent organic pollutants
-------
v»EPA
United States
Environmental Protection
Agency
What is the EPA doing about PFAS?
EPA PFAS Action Plan (2019)
¦ Assist states, tribes, and communities address
PFAS with short-term solutions and long-term
strategies to address PFAS
PFAS-Related Challenges
¦ Developing/validating laboratory analytical
methods for measuring PFAS
Assessing PFAS chemical toxicity
Developing standard toxicity values for PFAS
chemicals
Characterizing potential human exposure
pathways
Managing PFAS containing materials and waste
Testing drinking water treatment technologies
Identifying site remediation technologies
SEPA i
EPA's Per- and Polyfluoroalkyl
Substances (PFAS) Action Plan
-------
v»EPA
United States
Environmental Protection
Agency
Which PFAS are we interested in?
PFAS Screening Library creation: PFAS Landscape
¦ Maximize read-across
¦ Capture structural diversity
1220 PFAS currently in TSCA inventory
Brief Communication
A Chemical Category-Based Prioritization Approach for Selecting 75 Per- and
Polyfluoroalkyl Substances (PFAS) For Tiered Toxicity and Toxicokinetic Testing
(trace I'allewicz.' Ann M. Richard,' Antony J. William*,' ('hriuopher M. drulke,' Herder Sam*,' Ja\on Iximhert,'
Pamela l>. Xoye*/ Michael J. DeVila.' Ranald S. Hints,* Mark Strynar.* Annette (iui\eppi-EUt,* and Knurl/ S. Thomas'
Environmental Health Perspectives
014501-1
127(1) January 2019
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vvEPA
United States
Environmental Protection
Agency
Which PFAS are we interested in?
¦Initially, 75 PFAS selected from the PFAS Landscape, but the PFAS of
interest now expands to near 200 unique structures and the Landscape
up to 430 unique PFAS
9
EPA interest; Value for read-across
In vivo data
Data lacking; capturing structural diversity
Workflow
Step:
1: EPA
PFAS WG 2 3
4: EPA-PFAS
5: PFAS-Landscape Categories
31 vV
iljit
Final 75 PFAS Selection
If 0 oO ~I ¦¦¦¦ I ¦¦ aa a ¦¦¦¦¦ ¦ aa
10
5
0
25
20
15
10
l.llll.
N EPA PFAS WG 31 ~ EPA-PFAS ¦ PFAS Undscape
PFAS-Landscape
1
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j
...11I1I.
1..I.iiiii...
Ill
I !l0,o jL.iilil.l I1..I.11
P op
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vvEPA
United States
Environmental Protection
Agency
Which PFAS are beiing evaluated?
OH
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vvEPA
United States
Environmental Protection
Agency
How are we examining these PFAS?
¦A range of targeted and tiered high-throughput toxicity assays to serve
as guide for potential human health risk
¦ New approach methodologies (NAMs) used
¦ Alternative test methods and strategies to reduce, refine, and/or replace
mammalian animals
¦ In vitro tests/assays, in chemico assays, in silico algorithms
¦ Endpoints for PFAS work
¦ Hepatotoxicity
¦ Immunotoxicity
¦ Developmental toxicity
¦ Mitochondrial toxicity
¦ In vitro toxicokinetic assays
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oE
United States
Environmental Protection
Agency
What are in vitro toxicokinetic assays?
Toxicokinetics: the study of how a substance gets into the body and
what happens to it in the body
¦ Can be used to look at how chemicals move throughout the body and
lead to harmful effects
¦ Often viewed as a function of dose over time
Kinetic data can inform...
§
¦ Bioavailability (degree of a substance to enter I
circulation when introduced to body)
¦ Bioaccumulation potential (absorption » excretion)
¦ Metabolite formation (transformation of original chemical ^
new entity; can lead to bioactivation or detoxification)
12
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vvEPA
United States
Environmental Protection
Agency
What are in vitro toxicokinetic assays?
¦ Toxicokinetics: the study of how a substance gets into the body and
what happens to it in the body
1
H
a
a
Human
Hepatocytes
^ I
u
u
Human
Plasma
\
H
Hepatic
^ ¦ UMIUIMlj
Plasma Protein
Binding
1
J
r
In Vitro
In Vivo
Extrapolation
~\
I
1
i
i
j
1
1
Steady State
Blood
Concentrations
13
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AEPA
United States
Environmental Protection
Agency
What is plasma protein binding?
Assay to assess the free
(unbound) fraction of chemical to
proteins within the blood
¦
¦ Unbound molecules permeate through
eel! membranes to reach 'target'
¦ Determine by equilibrium dialysis,
ultrafiltration, and/or ultracentrifugation
Ultracentrifugation assay used
¦ Human plasma (10-donor pool, mixed
sex) centrifuged to separate aqueous
fraction from albumin, lipoproteins, and
fatty acids
¦ Mixtures of up to 4 PFAS (10 \iW\) were
included with each plasma sample, run
in triplicate
Lower free drug
concentration
Add
plasma
protein pi
u
oU
D ° O
°
O O °
o O
0
J v
o° fo
|o oo
o
0
o
0
fo
O ° '
o
-Drug
- Plasma
protein
High protein
binding
Low protein
binding
o o
o
¦O
Kieltyka et al 2016, J Pharm Sci
©o°®
©°©
° °
Higher free drug
concentration
14
-------
oE
United States
Environmental Protection
Agency
What is plasma protein binding?
Assay to assess the free
(unbound) fraction of chemical to
proteins within the blood
¦ fu
¦ Unbound molecules permeate through
cell membranes to reach 'target'
¦ Determine by equilibrium dialysis,
ultrafiltration, and/or ultracentrifugation
Ultracentrifugation assay used
¦ Human plasma (10-donor pool, mixed
sex) centrifuged to separate aqueous
fraction from albumin, lipoproteins, and
fatty acids
¦ Mixtures of up to 4 PFAS (10 |iM) were
included with each plasma sample, run
in triplicate
1 hf, 37*C, 150 rpm
(AF); fr»»
Fu = [AF] /[T5]
T5 SAMPLE
15
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AEPA
United States
Environmental Protection
Agency
What is in vitro hepatic clearance?
Oxidation
(Cytochrome P450"s)
o
Metabolite
Polar
Species
Ding
Q
Conjugation
Conjugation
{Glucuronidation etc.)
Stable Adducts
Non-polar
Species
Renal Elimination
(Urine)
Biliary Elimination
(Stool)
Liver is the major site of drug
metabolism in body (hepatic)
Hepatic clearance (CLhepatic) is
measure of the rate of
elimination of a chemical from
the liver
Models to study metabolism:
¦ Human liver microsomes
¦ Recombinantly expressed
enzymes
¦ Hepatocytes contain full
complement of hepatic drug
metabolizing enzymes
16
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oE
United States
Environmental Protection
Agency
What is in vitro hepatic clearance?
Hepatocyte
£ rt
\ I /
I H
For the PFAS work...
¦ Substrate depletion approach
using primary human
hepatocytes (50-donor pool,
mixed sex) at 1 nM PFAS
concentration
¦ Time course: 0, 15, 30, 60, 90,
120, and 240 min with non-linear
regression fit
¦ Work completed by collaborator
at National Toxicology Program
(NIEHS) [David Crizer]
17
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vvEPA
United States
Environmental Protection
Agency
How do we analyze these assay samples?
¦ Both assays require concentration determination of parent PFAS
¦ EPA has a range of analytical capabilities (single quads, triple quads,
high resolution mass spec)
18
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vvEPA
United States
Environmental Protection
Agency
Which PFAS of interest are LC-able?
-------
v»EPA
United States
Environmental Protection
Agency
What guided our LC-MS/MS method
development journey?
oEFft
METHOD 533: DETERMINATION OF PER- AND
POLYFLUOROALKYL SUBSTANCES IN DRINKING WATER BY
ISOTOPE DILUTION ANION EXCHANGE SOLID PHASE
EXTRACTION AND LIQUID CHROMATOGRAPHY/TANDEM
MASS SPECTROMETRY
[APPLICATION note]
Waters
THt Kiua or what's fossimjl*
An Alternative Ionization Technique for Perfluorinated Alkyl Substance
(PFAS) Analysis: Evaluating UniSpray for Water and Soil Samples
Kiift Organtini. Stuart Oeftfle. and Ken Rosnack
METHOD 537,1
EPA Document #; EPA/600/R-20;'006
DETERMINATION OF SELECTED PER- AND
POI YFLI ORINATED AI.KVL SUBSTANCES IN DRINKING
WATER BY SOLID PHASE EXTRACTION AND LIQUID
CHROMATOGRAPHY/TANDEM MASS SPECTROMETRY
(LC/MS/MS)
[technology brief]
Ultra Low-Level Detection of Perfluoroalkyl Substances (PFASs)
Using the PFC Analysis Kit
Lauren Mullin and Jennifer Buroess
Antlybul ml BtwuJyOc* Ommaay (20)$ 41VJ507-J5»
RESEARCH PAPtR
ICS > 13 > 13.060 > 13.060.50
ISO 21675:2019
Water quality — Determination of perfluoroalkyl and
polyfluoroalkyl substances (PFAS) in water — Method
using solid phase extraction and liquid chromatography-
tandem mass spectrometry (LC-MS/MS)
A single analytical method for the determination of 53
legacy and emerging per- and polyfluoroalkyl substances (PFAS)
in aqueous matrices
Timothy L. Co-ggan' ¦ Tarun Anumol1 • James Pyke1 • Shimeta1 ¦ Bradley O. Clarice1
mime
nence&Tecflnology
5. Cite Thb: £nvWon Ut firctaot 2019, SJ, 4717-47J7
Identification of Per- and Polyfluoroalkyl Substances in the Cape
Fear River by High Resolution Mass Spectrometry and Nontargeted
Screening
James McCord ° and Mark Strgur'
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v»EPA
United States
Environmental Protection
Agency
What LC-MS/MS settings were used?
ACQUITY UPLC l-Class FTN
Equipped with Waters RFC Kit
CORTECS T3 2.7 jjM 3.0x100 mm
Column Temp: 55°C
Flow Rate: 0.6 mL/min
Run Time: 6.5 min
Mobile Phase A: 95:5 water:
acetonitrile with 2.5 mM ammonium
acetate
Mobile Phase B: 5:95 water:
acetonitrile with 2.5 mM ammonium
acetate
Xevo TQ-S Micro
MRM transitions determined
Acquisition Polarity: ESI+ and ESI-
Capillary Voltage: 0.4 kV
Source Temperature: 150°C
Desolvation Temperature: 500°C
Desolvation Gas Flow: 1000 L/hr
Cone Gas Flow: 150 L/hr
19 mass-labelled PFAS (Wellington
Laboratories, MPFAC-24ES) was
included for quantitation
21
f&l "..."
1
WELLINGTON
LABORATORIES
Standards for Environmental Testing and Research
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v»EPA
United States
Environmental Protection
Agency
How's the method working?
B
0 F F F F
11 I I I I
HO — $—I \-
O F F F F
DTXSID30170109 DTXSID5030030 DTXSID8031865 DTXSID30382104 DTXSID90382620
PFBS PFOA CI-PFNA 7:3 FTCA
OFFFFFFF
HQ—S— \—
OFFFFFFFF
DTXSID3031864
PFOS
I [
F F
DTXSID50375114
PFPE-6
H
O F F F F F F
DTXSID50469320
PFHxSA
DTXSID3059921
PFTeDA
DTXSID7027831
NMeFOSE
0.80 1.00 1 20 1 40
2.00 220 2.40 2.60 2 80 3.00 320 3.40
S20 540 5.60
¦ Mixture of PFAS run at 100 ppb
Most have estimated LOQ < 50 ppt
¦ Most PFAS were analyzed in ESI
negative; others were monitored as
acetate adducts, fragmented in-source
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vvEPA
United States
Environmental Protection
Agency
What about DMSO stock quality?
¦ DMSO is a common solvent used for assays
¦ RADAR = MRM (MS/MS) + MS full scan
¦ Monitor for any interferences and impurities
¦ Application Note: 720005033EN
¦ Created scoring system for quality of stocks
23
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vvEPA
United States
Environmental Protection
Agency
What about DMSO stock quality
¦ DMSO is a common solvent used for assays
¦ RADAR = MRM (MS/MS) + MS full scan
¦ Monitor for any interferences and impurities
¦ Application Note: 720005033EN
¦ Created scoring system for quality of stocks
EM-MS full Sun
J
DTXSID3031862
Perfluorohexanoic acid
/
. .. .. ..
24
DTXSID70880215
Perfluoro-2-methyl-3-
oxahexanoic acid
ESI- MS Full Scan
MRM
T /
I tnm
mr
-ir,:
S —i S =_
>>>,» >» ,k|" j
i n i
Nil I!
«J" 133 j*j» i , i ill /iii ir*"I till
'r >»
¦ n
Liberatore et al 2020,
Environ Sci Techno! Lett
. | , > If' . f. ft ' 1 ' 'T
iwitojSTjw j.-o ' j» »io * " »to 3 to ' »to ' i« "
1 oo ' i to ' i to ' i to ' 1 to' 1 to t» ' < ' i to " i to ' ?to ' J to J» ' ? » ' ?to ?m ' 2« ' in
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vvEPA
United States
Environmental Protection
Agency
What did we find from the plasma protein
binding assay?
50 LC-able PFAS have determined fraction unbound data
Fu I binding to plasma proteins t
25
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v»EPA
United States
Environmental Protection
Agency
What did we find from the plasma protein
binding assay?
O-3-i
0.2-
0.1-
0.0-
¦£
J> x6a
^
^
/ / / / y s/ /
'¦ -0 .aO ^ ^ jJC-® *°
f f f / ¦*" " ¦•*
,0^ «/ ^ ^ JT *
^ / / / ^ / /
^ /
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v»EPA
United States
Environmental Protection
Agency
What did we find from the plasma protein
binding assay?
OFFFFFFFFFF
II
0=S-
O FFFFFFFFFF
Na
PFAS
Avg Fu
LogK0w
Perfluorohexanesulfonic acid
0.0008
3.47
Potassium perfluorooctanesulfonate
0.0040
3.39
Perfluorobutanesulfonic acid
0.0128
2.57
Sodium perfluorodecanesuifonate
0.0546
1.83
LogK,
ow
0=S OH
II
o
* This is based on Dashboard
Average Predicted LogKow
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vvEPA
United States
Environmental Protection
Agency
Any observations from the hepatic
clearance assay?
¦ More than 20 LC-able PFAS assessed
1. In vitro hepatic clearance scree
¦ 0 and 4 hr time points for active and inactive hepatocytes
¦ Compared time ratios to examine for clearance potential
28
Legacy PFAS are stable
Perfluorooctanoic acid (DTXSID8031865)
¦5-
Active Hepatocytes Inactive Hepatocytes
Perfluorooctanesulfonic acid (DTXSID3031864)
12t
Active Hepatocytes Inactive Hepatocytes
Sulfonic acid PFAS are stable, too
Perfluorooctanesulfonic acid (DTXSID3031864)
1.2-1
Active Hepatocytes Inactive Hepatocytes
Perfluorohexanesulfonic acid (DTXSID7040150)
I I I
OH
II
•
•
I I I
rH 0
0
3=
O
Active Hepatocytes Inactive Hepatocytes
Perfluorobutanesulfonic acid (DTXSID5030030)
20n n F F F F
OH F F F
Active Hepatocytes Inactive Hepatocytes
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vvEPA
United States
Environmental Protection
Agency
Any observations from the hepatic
clearance assay?
¦ More than 20 LC-able PFAS assessed
1. In vitro hepatic clearance scree
¦ 0 and 4 hr time points for active and inactive hepatocytes
¦ Compared time ratios to examine for clearance potential
Polyfluoroalkyl substances are LESS stable than
corresponding perfluoroalkyl substances
Perfluorooctanoic acid (DTXSID8031865)
2H, 2H, 3H, 3H-Perf I uo rooctanoic acid (DTXSID20874028)
-------
oE
United States
Environmental Protection
Agency
Any observations from the hepatic
clearance assay?
More than 20 LC-able PFAS assessed
2.
CUD
C
LO
03
CD
i_
u
c
O)
¦4—'
rc
O)
u
c
03
i_
03
(D
Metabolic stability time course
0, 0.25, 0.50, 1, 1.5, 2, 4 hr time points
Non-linear fit to determine half-life (T1/2)
Compound Name
Half-life (min)
Clearance (iiL/min/million cells)
Perfluorobutanoic acid
44769343
1.55E-05
Potassium perfluorohexanesulfonate
21340366
3.25E-05
Perfluorohexanoic acid
237257
2.92E-03
Ammonium perfluorooctanoate
88735
7.81 E-03
Pntassii im nprfli inrnhi itanpsi ilfnnatp
Perfluorononanoic acid
1155
6.00E-01
Perfluorooctanesultonic acid
PerfluoroM-methoxvbutanoicI acid
346.5
2.00E+00
2H,2H,3H,3H-Perfluorooctanoic acid | 101.4 | 6.83E+00
N-Ethylperfluorooctanesultonamide
57
1.22E+01
3-(Perfluoro-2-butyl)propane-1,2-diol
35.87
1.93E+01
Perfluoro-3,6,9-trioxatridecanoic acid
29.71
2.33E+01
Nonafluoropentanamide
25.45
2.72E+01
3,3-Bis(trifluoromethyl)-2-propenoic acid
19.77
3.51 E+01
4:2 Fluorotelomer sulfonic acid
17.5
3.96E+01
Octafluoroadipamide
12.8
5.41 E+01
Perfluoropentanamide
10.63
6.52E+01
N-Methylperfluorooctanesulfonamide
10.17
6.81 E+01
2,2,3,3,4,4-Hexafluorobutanoic acid
4.209
1.65E+02
Perfluorooctanesulfonamide
2.789
2.48E+02
Perfluorononanoic acid (DTXSID8031863)
2H, 2H, 3H, 3H-Perfluorooctanoic acid (DTXSID20874028)
1.50t f
0.00
Time (hr)
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vvEPA
United States
Environmental Protection
Agency
What is IVIVE?
¦ In vitro-in vivo extrapolation = IVIVE
¦ Model approach that allows in vitro data to be extrapolated to estimate
corresponding in vivo effects
¦ Start at tissue/biomarker level -> estimate external exposure
¦ Steady-state concentration (Css)
¦ Concentration of compound in body that stays consistent
¦ This takes into account plasma protein binding and hepatic clearance data
31
Forward Dosimetry
Starting at external exposure, working inward to estimate systemic/tissue level dose
Exposure
Exposure
routes
Activity and
environmental
concentration
distributions
Inhalation
=F
0
Ingestion
Dermal
Exposure Prediction Evaluation
(ExpoCast tools: e.g., SHEDS-HT, HEM)
Internal Dose
Pharmacokinetic
modeling
king
SI
Tissue metric
distributions
J±L
gfEK,
| Vidn
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AEPA
United States
Environmental Protection
Agency
What is IVIVE?
i
Z2
Human
Hepatocytes
Uny
-
t
Steady State
Blood
Concentrations
Human
Plasma
Plasma Protein
Binding
Wetmore etal 2012, Toxicol Sci;
Wetmore2015, Toxicology
32
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v»EPA
United States
Environmental Protection
Agency
What is IVIVE?
In Vitro - In Vivo
Extrapolation
[Conc]ss =
Dose Rate * Body Weight
CL
¦WholeBody
Steady State
Blood
Concentrations
CL
R
CL
H
Assumptions
Exposure at 1 ng/kg/day
Linear kinetics
100% oral bioavailability
CI_R= Fy * GFR
where GFR ~ 6.7 L/hr
Fu * QL * CL
Int
CLh =
QL + Fu * CL|nt
CLjnt = HPGL * Vl * CLjnvitro
where Ql ~ 90 L/hr
Wetmore etal 2012, Toxicol Sci;
Wetmore 2015, Toxicology
where HPGL * 137 million cells/g
VL ~ 1820 g
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v»EPA
United States
Environmental Protection
Agency
What did IVIVE show with PFAS data?
Tr'
\
%
Hepatocytes
9
0
UWji
Plasma
Hepatic Clearance
-------
v»EPA
United States
Environmental Protection
Agency
What is the big picture of this PFAS
toxicity effort?
Plasma
Hepatic Clearance
-------
vvEPA
United States
Environmental Protection
Agency
Summary of findings
¦ Experimental in vitrotoxicokinetic data (Fu and Clhepatic) are being
measured on over 120 PFAS for use in IVIVE modeling
¦ Multi-residual LC-MS/MS method developed to analyze more than 60
unique PFAS
¦ Plasma protein binding data indicate high binding rates, with 75%
exhibiting Fu values from 0.001 - 0.05
¦ Assuming an external exposure of 1 ^g/kg/day, Css predictions ranged
from 0.16-895 pM, with a median value of 23.29
¦ These Css estimates eventually will be combined with other high-
throughput screening data to help identify PFAS risk to humans
¦ Continuing data generation for additional PFAS and toxicokinetic
assays for bioavailability, metabolite identification, and renal reuptake
36
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v»EPA
United States
Environmental Protection
Agency
Acknowledgements
ORD-CCTE
Lucas Albrecht
Mike DeVito
Annette Guiseppi-EIie
Josh Harrill
Keith Houck
Mike Hughes
Richard Judson
Jen Korol-Bexell
Anna Kreutz
Stephanie Padilla
Grace Patlewicz
Matthew Phillips
Ann Richard
Tim Shafer
Adam Swank
¦ Rusty Thomas
¦ John Wambaugh
¦ Barbara Wetmore
¦ Antony Williams
ORD-CEMM
¦ Scott Clifton
¦ Matt Henderson
¦ James McCord
¦ Larry McMillan
¦ Mark Strynar
NTP
¦ David Crizer
Waters
¦ Aurelie Marcotte
¦ Kari Organtini
NTP
National Toxicology Program
U.S. Department of Health and Human Services
smeltz.marci@epa.gov
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