11

Approaches to Pesticide Approaches to Pesticide Cumulative Risk Cumulative Risk

Assessment: Policy, Assessment: Policy, Practice, ExperimentationPractice, Experimentation

Anna B. Lowit, Ph.D.Anna B. Lowit, Ph.D.U.S. Environmental Protection AgencyU.S. Environmental Protection Agency

Office of Pesticide ProgramsOffice of Pesticide Programs

Virginia Moser, Ph.D.Virginia Moser, Ph.D.U.S. Environmental Protection AgencyU.S. Environmental Protection AgencyOffice of Research and DevelopmentOffice of Research and Development

22

Outline: Policy & PracticeOutline: Policy & Practice

1.1. Introduction: regulatory context, Introduction: regulatory context, guidance documents, key principlesguidance documents, key principles

2.2. Hazard Assessment: relative potency Hazard Assessment: relative potency factor approach factor approach

3.3. Exposure Assessment: food, water, Exposure Assessment: food, water, residentialresidential

4.4. Cumulative assessment & ‘Track Back’Cumulative assessment & ‘Track Back’

5.5. SummarySummary

33

Introduction Introduction

EPA’s Office of Pesticide Programs is a EPA’s Office of Pesticide Programs is a licensing program regulating pesticide products licensing program regulating pesticide products in the U.S.in the U.S.• Review effects of pesticides on human and ecological Review effects of pesticides on human and ecological

healthhealth

Food Quality Protection Act (FQPA, 1996) Food Quality Protection Act (FQPA, 1996) • Requires EPA to take into account when setting Requires EPA to take into account when setting

pesticide tolerances: pesticide tolerances: ““available evidence concerning the cumulative effects on available evidence concerning the cumulative effects on

infants and children of such residues and other substances infants and children of such residues and other substances that have a that have a common mechanism of toxicitycommon mechanism of toxicity.” .”

44

IntroductionIntroduction

Under FQPA (1996), cumulative risk is Under FQPA (1996), cumulative risk is defined as: defined as: • The risk associated with a group of chemicals The risk associated with a group of chemicals

that are toxic by a common mechanism from that are toxic by a common mechanism from all pathways all pathways

• Multi-chemical & Multi-pathway Multi-chemical & Multi-pathway Food, drinking water, consumer uses Food, drinking water, consumer uses Routes of exposure (oral, dermal, inhalation)Routes of exposure (oral, dermal, inhalation)

55

Introduction: CRA GuidanceIntroduction: CRA Guidance

OPP developed guidance OPP developed guidance document for cumulative risks document for cumulative risks assessments under FQPAassessments under FQPA• Established core principles for Established core principles for

performing cumulative risk performing cumulative risk assessmentsassessments

• Developed tools for calculating Developed tools for calculating multichemical and multipathway multichemical and multipathway risk estimatesrisk estimates

• Not a ‘recipe book’Not a ‘recipe book’

http://www.epa.gov/oppfead1/trac/science/#common

66

Introduction: Key PrinciplesIntroduction: Key Principles Appropriately Integrate Toxicology & Exposure DataAppropriately Integrate Toxicology & Exposure Data

• Time-Frame ConsiderationsTime-Frame Considerations Time to peak effect? Time to recovery?Time to peak effect? Time to recovery? When does the exposure occur? What is the duration of exposure?When does the exposure occur? What is the duration of exposure?

Strive for Realistic & Accurate AssessmentsStrive for Realistic & Accurate Assessments• Use Representative DataUse Representative Data• Avoid Compounding ConservatismsAvoid Compounding Conservatisms

Preserve and Maintain Geographic, Temporal & Preserve and Maintain Geographic, Temporal & Demographic SpecificityDemographic Specificity• Calendar-Base ApproachCalendar-Base Approach

Be Able to “Track Back” Sources of Exposures & Be Able to “Track Back” Sources of Exposures & Perform Sensitivity AnalysesPerform Sensitivity Analyses• Major Risk Contributors Major Risk Contributors

Emphasis of presentation at CRA Workshop

77

Basic Steps in a Pesticide Basic Steps in a Pesticide Cumulative Risk AssessmentCumulative Risk Assessment

Identify common mechanism group (CMG) Identify common mechanism group (CMG) Determine relevant exposure scenarios/pathwaysDetermine relevant exposure scenarios/pathways Identify cumulative assessment group (CAG)Identify cumulative assessment group (CAG) Consider appropriate method(s) & data sourcesConsider appropriate method(s) & data sources Conduct assessmentConduct assessment

• Characterize & select common mechanism endpoint(s), Characterize & select common mechanism endpoint(s), determine chemical potency & select index chemicaldetermine chemical potency & select index chemical

• Convert pesticide residues to equivalents of the index Convert pesticide residues to equivalents of the index chemicalchemical

• Combine/integrate food, water, & residential exposures Combine/integrate food, water, & residential exposures on an internally consistent manner which incorporates on an internally consistent manner which incorporates demographic & temporal-spatial factorsdemographic & temporal-spatial factors

88

Introduction: CMG Guidance Introduction: CMG Guidance

Mechanism of Toxicity-Mechanism of Toxicity--Major steps leading to an -Major steps leading to an adverse health effect following interaction of a pesticide adverse health effect following interaction of a pesticide with biological targets. All steps leading to an effect do with biological targets. All steps leading to an effect do not need to be specifically understoodnot need to be specifically understood

Common Mechanism-Common Mechanism--Two or more pesticide chemicals -Two or more pesticide chemicals that cause a common toxic effect…by the same, or that cause a common toxic effect…by the same, or essentially the same, sequence of major biochemical essentially the same, sequence of major biochemical eventsevents

http://www.epa.gov/fedrgstr/EPA-PEST/1999/February/Day-05/6055.pdf

Group via Common

Mechanism

Pesticides

1010

Common Mechanism of Toxicity?Common Mechanism of Toxicity?

Three general principles to guide common Three general principles to guide common mechanism determinations: mechanism determinations: • Act on the same molecular target at the same Act on the same molecular target at the same

target tissue,target tissue,• Act by the same biochemical mechanism of Act by the same biochemical mechanism of

action, possibly sharing a common toxic action, possibly sharing a common toxic intermediateintermediate

• Cause the same critical toxic effectCause the same critical toxic effect Called the common toxic effect Called the common toxic effect

1111

Common Mechanism of Toxicity?Common Mechanism of Toxicity?

Is there concordance in dose response Is there concordance in dose response and timing between the major steps and and timing between the major steps and the toxic effect?the toxic effect?

Is it biologically/chemically plausible?Is it biologically/chemically plausible? What are strengths & uncertainties of the What are strengths & uncertainties of the

available data?available data? Could there be other an alternative Could there be other an alternative

mechanism(s) of action?mechanism(s) of action?

1212

Relative Potency Factor MethodRelative Potency Factor Method PBPK models would be preferred PBPK models would be preferred

• In vivoIn vivo and and in vitroin vitro pharmacokinetic data not available pharmacokinetic data not available at this timeat this time

• Multi-chemical, multi-pathway models not availableMulti-chemical, multi-pathway models not available Relative toxic potency of each chemical is Relative toxic potency of each chemical is

calculated in comparison to “index chemical” calculated in comparison to “index chemical”

Exposure equivalents of index chemical are Exposure equivalents of index chemical are combined in the cumulative risk assessmentcombined in the cumulative risk assessment

=RPFIndex ChemicalBMD

Chemical nBMD

OP CRA Hazard & Dose Response OP CRA Hazard & Dose Response

• Collaborative effort with EPA-ORD

• Rat data collected from studies at 21 days or longer where inhibition is no longer changing (ie, steady state)

• Use of multiple studies provides robust estimate of pesticide potency & incorporates variability across studies

Relative Potency Factors from OP Cumulative Risk Assessment

1515

NMC CRA Hazard & NMC CRA Hazard & Dose ResponseDose Response

Collaborative effort with ORDCollaborative effort with ORD• Benchmark modeling and dose-response and time course Benchmark modeling and dose-response and time course

laboratory studieslaboratory studies Relative potencies are estimated along with recovery Relative potencies are estimated along with recovery

half lives from acute (single dose) rat dose-time half lives from acute (single dose) rat dose-time response data at or near peakresponse data at or near peak

Dose & Time Course Model UsedDose & Time Course Model Used• Dose-response portion of model is similar to that used for AChE Dose-response portion of model is similar to that used for AChE

inhibition by organophosphatesinhibition by organophosphates• Time course model reflects an exponential decay of inhibitionTime course model reflects an exponential decay of inhibition

Rapid nature of NMC toxicity----Exposure assessment Rapid nature of NMC toxicity----Exposure assessment on single day exposures onlyon single day exposures only

Example: Oxamyl Dose-Time Example: Oxamyl Dose-Time ResponseResponse

1616

Dose of Mixture (mg/kg)

0 2 4 6 8 10 12 14 16

Cho

lines

tera

se a

ctiv

ity (

% c

ontr

ol)

20

30

40

50

60

70

80

90

100

110

120

Expected Values (+ 95% confidence limits)Actual Data (+ sem)

Carbamate Mixture Study:Brain Cholinesterase Activity

Dose-Additive Design

Equipotent Mixture

Control

1717

1818

Exposure Assessment & Exposure Assessment & Probabilistic TechniquesProbabilistic Techniques

Probablistic exposure techniques are routinely Probablistic exposure techniques are routinely applied by OPP for virtually all its pesticide risk applied by OPP for virtually all its pesticide risk assessmentsassessments• More accurate estimate of the entire range of More accurate estimate of the entire range of

exposures and their associated probabilities exposures and their associated probabilities

OPP’s Cumulative Risk Assessments rely on OPP’s Cumulative Risk Assessments rely on probabilistic (Monte-Carlo) techniques to probabilistic (Monte-Carlo) techniques to evaluate exposureevaluate exposure• Food, drinking water, residential uses, multi-pathwayFood, drinking water, residential uses, multi-pathway

1919

Exposure Assessment Exposure Assessment Software & ModelingSoftware & Modeling

Development of probabilistic models that Development of probabilistic models that permit time-based integration of residential, permit time-based integration of residential, food, and water exposures to pesticidesfood, and water exposures to pesticides

• ““Time-Based Integration” = Calendar-based Time-Based Integration” = Calendar-based approachapproach

• Allow probabilistic combining of exposures Allow probabilistic combining of exposures through multiple pathways and routesthrough multiple pathways and routes Single chemical or Multi-chemicalSingle chemical or Multi-chemical Food, Drinking Water, ResidentialFood, Drinking Water, Residential Ingestion, Inhalation, Dermal absorptionIngestion, Inhalation, Dermal absorption

2020

Exposure Assessment Exposure Assessment Software & ModelingSoftware & Modeling

Key concept: Must track potentially Key concept: Must track potentially exposed persons on a daily basis in a way exposed persons on a daily basis in a way that preserves all appropriate linkages and that preserves all appropriate linkages and appropriately allows for co-occurring appropriately allows for co-occurring exposuresexposures• Age, sex, behavior, region, etc.Age, sex, behavior, region, etc.

2121

Exposure Assessment SoftwareExposure Assessment Software OPP has used several software models to OPP has used several software models to

perform its risk assessmentsperform its risk assessments

Presented to FIFRA SAP by OPP along with Presented to FIFRA SAP by OPP along with model development teamsmodel development teams• LifelineLifeline• CARESCARES• DEEM/CalendexDEEM/Calendex• SHEDSSHEDS

All four models All four models • conform to EPA & OPP guidance conform to EPA & OPP guidance • have undergone peer reviewhave undergone peer review• are publicly availableare publicly available

2222

Exposure Assessment Exposure Assessment Software & ModelingSoftware & Modeling

Inputs includeInputs include• Toxicity information (e.g RfD, BMD, NOAEL)Toxicity information (e.g RfD, BMD, NOAEL)• Exposure informationExposure information

ResiduesResidues Food consumption (from USDA’s CSFII)Food consumption (from USDA’s CSFII) Behavior information (e.g., hand to mouth behavior)Behavior information (e.g., hand to mouth behavior)

Output includesOutput includes• Exposure levels (mg/kg bwt/day)Exposure levels (mg/kg bwt/day)• Risk metric (% RfD occupied, Margin of Exposure)Risk metric (% RfD occupied, Margin of Exposure)• Risk “drivers” Risk “drivers”

chemical(s), commodities, or residential uses which chemical(s), commodities, or residential uses which contribute significantly to riskcontribute significantly to risk

2323

Exposure Assessment Exposure Assessment Software & ModelingSoftware & Modeling

Use data from well-known surveys to Use data from well-known surveys to generate and evaluate specific daily generate and evaluate specific daily exposures for individualsexposures for individuals• Use available databases to address each Use available databases to address each

component of simulationcomponent of simulation• Incorporates seasonal and other aspectsIncorporates seasonal and other aspects

2424

Populations Groups AssessedPopulations Groups Assessed

Separate assessments were based on survey Separate assessments were based on survey information on the following age groups: information on the following age groups: • Infants <1Infants <1• Children 1 - 2 years oldChildren 1 - 2 years old• Children 3 - 5 years oldChildren 3 - 5 years old• Children 6 - 12 years oldChildren 6 - 12 years old• Youths 13 - 19 years oldYouths 13 - 19 years old• Adults 20 - 49 years oldAdults 20 - 49 years old• Adults 50+ years oldAdults 50+ years old• Females 13 - 49Females 13 - 49

2525

Regions AssessedRegions Assessed

2626

Software Inputs: Software Inputs: CSFII 1994-96/1998 Food CSFII 1994-96/1998 Food

Consumption SurveyConsumption Survey Nationally Representative/Statistically-BasedNationally Representative/Statistically-Based

• Intakes of individuals residing in 50 states and D.C. Intakes of individuals residing in 50 states and D.C. • 21,662 individual participants interviewed over the period 21,662 individual participants interviewed over the period

1998 Supplemental Children’s Survey 1998 Supplemental Children’s Survey • ~5000 children ~5000 children • birth through 9 years old birth through 9 years old • integrated into 1994-96 CSFIIintegrated into 1994-96 CSFII

Consisted of:Consisted of:• 2 non-consecutive days using in-person 24 hour recalls (ca. 2 non-consecutive days using in-person 24 hour recalls (ca.

3-10 days apart)3-10 days apart) Covers all seasons of year and all days of weekCovers all seasons of year and all days of week

2727

USDA Pesticide Data Program USDA Pesticide Data Program (PDP) Residue Data(PDP) Residue Data

Statistically-reliable sampling procedure designed to be Statistically-reliable sampling procedure designed to be representative of US food supply representative of US food supply • Approximately 600 samples per commodity per yearApproximately 600 samples per commodity per year

Samples collected at terminal markets and distribution Samples collected at terminal markets and distribution centerscenters• Samples prepared as if for consumptionSamples prepared as if for consumption

PDP has tested more than 50 different commodities and PDP has tested more than 50 different commodities and more than 300 pesticides/metabolitesmore than 300 pesticides/metabolites• Fresh/frozen/canned fruits & vegetables, fruit juices, milk, grains, Fresh/frozen/canned fruits & vegetables, fruit juices, milk, grains,

meat/poultry/pork, corn syrup, etc. meat/poultry/pork, corn syrup, etc. • Emphasis on children’s foodsEmphasis on children’s foods

Reliable analytical methods with low limits of detectionReliable analytical methods with low limits of detection

2828

““Track Back” in Food ExposureTrack Back” in Food Exposure

0%

5%

10%

15%

20%

25%

30%

Pot

ato

Str

awbe

rry

Pea

ch

Wat

erm

elon

Can

talo

upe

Cuc

umbe

r

Spi

nach

Nec

tarin

e

Pea

r

Lettu

ce, h

ead

Gra

pe

App

le

Bea

n, s

nap

Lettu

ce, l

eaf

Cab

bage

Ora

nge

Che

rry

Thiodicarb

Pirimicarb

Oxamyl

Methomyl

Methiocarb

Formetanate hydrochloride

Carbofuran

Carbaryl

Aldicarb

Sum of H X F

Food

PARENTPESTICIDE

2929

Cumulative DW AssessmentCumulative DW Assessment

Regional level screenRegional level screen Watershed-based modeling for surface Watershed-based modeling for surface

water sourceswater sources Shallow ground water for private wellsShallow ground water for private wells ““Typical” usage patternsTypical” usage patterns Daily distribution over multiple yearsDaily distribution over multiple years Estimates compared with, calibrated Estimates compared with, calibrated

against monitoringagainst monitoring

3030

For DW, Each Regional Location For DW, Each Regional Location Reflects …Reflects …

Geographic area with high potential for Geographic area with high potential for combined (cumulative) exposurecombined (cumulative) exposure• Influenced by both use and relative toxicitiesInfluenced by both use and relative toxicities

Location-specific conditionsLocation-specific conditions• environmental data (soil/site, weather, crops)environmental data (soil/site, weather, crops)• Major crop-pesticide combinations within that Major crop-pesticide combinations within that

areaarea Vulnerable drinking water sources within Vulnerable drinking water sources within

the regionthe region

3131

Residential Exposure Residential Exposure AssessmentAssessment

Extensive use of survey data and other pesticide use Extensive use of survey data and other pesticide use informationinformation

Use of distributions for residues and behavior/activity Use of distributions for residues and behavior/activity elementselements

• Hand-to-mouth activitiesHand-to-mouth activities

• Choreographed adult activities/Non-scripted playChoreographed adult activities/Non-scripted play

• Transfer Coefficients/Dislodgeable Foliar ResidueTransfer Coefficients/Dislodgeable Foliar Residue Use of a calendar based model to address the temporal Use of a calendar based model to address the temporal

use of residential usesuse of residential uses Region-specific analysesRegion-specific analyses

3232

Residential Exposure Residential Exposure AssessmentAssessment

Assessment performed for the following Assessment performed for the following uses:uses:• Indoor UsesIndoor Uses• Pet UsesPet Uses• Home Lawn and GardenHome Lawn and Garden• Golf CourseGolf Course• Public Health UsesPublic Health Uses

Cumulative MOEs for Children 1-2 Region A Seven Day Rolling Average Analysis

1

10

100

1000

10000

100000

1000000

1 10 19 28 37 46 55 64 73 82 91 100

109

118

127

136

145

154

163

172

181

190

199

208

217

226

235

244

253

262

271

280

289

298

307

316

325

334

343

352

361

Julian Days

Food MOE PRZM-EXAMS Water MOE Total MOE Inhalation MOE Dermal MOE Oral (non-dietary) MOE

Day of the Year

Example of Time based exposure Example of Time based exposure profile: Organophosphatesprofile: Organophosphates

Children 1-27-day

Inhalation + Total

Food

Water

Oral (non-dietary)

Dermal

MO

Es

3333

3434

Public Participation ProcessPublic Participation Process Numerous Public Technical Briefings on methods Numerous Public Technical Briefings on methods

and approaches for cumulative risk assessment and approaches for cumulative risk assessment and resultsand results

FIFRA Science Advisory Panel meetings on FIFRA Science Advisory Panel meetings on methods and approachesmethods and approaches• More than 20More than 20

Preliminary assessment –public comment and Preliminary assessment –public comment and Science Advisory Panel meetings Science Advisory Panel meetings

Revised assessment(s)–public commentRevised assessment(s)–public comment Website dedicated to cumulative risk assessment Website dedicated to cumulative risk assessment

http://www.epa.gov/pesticides/cumulative/

3535

Pesticide Cumulative RisksPesticide Cumulative Risks

Organophosphates (OP) Organophosphates (OP) NN-methyl carbamates -methyl carbamates Triazines Triazines ChloroacetanilidesChloroacetanilides

3636

Pesticide Cumulative RisksPesticide Cumulative Risks

Pyrethroids—Work has only just begunPyrethroids—Work has only just begun• Draft common mechanism grouping Draft common mechanism grouping

reviewed & supported by SAP, June 2009reviewed & supported by SAP, June 2009• OPP & ORD developing PBPK models for OPP & ORD developing PBPK models for

use in the pyrethroid cumulative risk use in the pyrethroid cumulative risk assessmentassessment

• Linkage between probabilistic exposure Linkage between probabilistic exposure assessment (SHEDS) and PBPK modelsassessment (SHEDS) and PBPK models

3737

Key PrinciplesKey Principles Appropriately Integrate Toxicology & Exposure DataAppropriately Integrate Toxicology & Exposure Data

• Time-Frame ConsiderationsTime-Frame Considerations Time to peak effect? Time to recovery?Time to peak effect? Time to recovery? When does the exposure occur? What is the duration of exposure?When does the exposure occur? What is the duration of exposure?

Strive for Realistic & Accurate AssessmentsStrive for Realistic & Accurate Assessments• Use Representative DataUse Representative Data• Avoid Compounding ConservatismsAvoid Compounding Conservatisms

Preserve and Maintain Geographic, Temporal & Preserve and Maintain Geographic, Temporal & Demographic SpecificityDemographic Specificity• Calendar-Base ApproachCalendar-Base Approach

Be Able to “Track Back” Sources of Exposures & Be Able to “Track Back” Sources of Exposures & Perform Sensitivity AnalysesPerform Sensitivity Analyses• Major Risk Contributors Major Risk Contributors

3838

Thank You!Thank You!

Approaches to Pesticide Approaches to Pesticide Cumulative Risk Assessment: Cumulative Risk Assessment:

Policy, Practice, Policy, Practice, ExperimentationExperimentation

Ginger Moser, Ph.D., D.A.B.T.Ginger Moser, Ph.D., D.A.B.T.TAD/NHEERL/ORD/US EPATAD/NHEERL/ORD/US EPA

moser.ginger@epa.govJuly 14, 2009July 14, 2009

3939

AcknowledgementsAcknowledgements

Statistical expertise: Virginia Statistical expertise: Virginia Commonwealth University Commonwealth University • Drs. Chris Gennings, Hans Carter, Jr.Drs. Chris Gennings, Hans Carter, Jr.• Graduate students including but not limited to Graduate students including but not limited to

Michelle Casey, Adam HammMichelle Casey, Adam Hamm

Technical collaborations: US EPATechnical collaborations: US EPA• Drs. Dave Herr, Stephanie Padilla, Anna Drs. Dave Herr, Stephanie Padilla, Anna

Lowit, Jane Ellen SimmonsLowit, Jane Ellen Simmons• Pam Phillips, Kathy McDaniel, RenPam Phillips, Kathy McDaniel, Renéée e

MarshallMarshall4040

BackgroundBackground

Humans are exposed to multiple Humans are exposed to multiple chemicalschemicals

Effects of chemical mixtures may not be Effects of chemical mixtures may not be adequately predicted by studying adequately predicted by studying individual chemicalsindividual chemicals

Component-based mixtures risk Component-based mixtures risk assessment is aided by experimental assessment is aided by experimental design combining:design combining:• exposure evaluationsexposure evaluations• quantitative chemical informationquantitative chemical information• appropriate statistical analyses appropriate statistical analyses 4141

Theories of AdditivityTheories of Additivity

TerminologyTerminology• Zero interaction = additivityZero interaction = additivity• Synergy, antagonism = response greater, less Synergy, antagonism = response greater, less

than predicted under additivitythan predicted under additivity

Dose additivity = chemicals interacting as if Dose additivity = chemicals interacting as if they were dilutions of one anotherthey were dilutions of one another• Does not require same shape of dose-responseDoes not require same shape of dose-response• Does not require common mechanism of actionDoes not require common mechanism of action• Combinations of sub-threshold doses may be Combinations of sub-threshold doses may be

activeactiveBerenbaum, J. Theor. Biol. 114:413-431,1985 4242

Isobolographic ApproachIsobolographic Approach

Classic method of describing dose-additivityClassic method of describing dose-additivity• Isobols of equi-effective dosesIsobols of equi-effective doses• Requires multiple dose-response determinations with Requires multiple dose-response determinations with

different dose combinations of each chemicaldifferent dose combinations of each chemical• Data intensiveData intensive

Isobolographic Analysis

Chemical A Dose

0 2 4 6 8

Che

mic

al B

Dos

e

0

5

10

15

20

A ED50

B ED50

ED50 isobol

Isobolographic Analysis

Chemical A Dose

0 2 4 6 8

Che

mic

al B

Dos

e

0

5

10

15

20

ED50

ED20ED10

4343

Ray ApproachRay Approach Dose-response along ray of mixture with fixed proportions Dose-response along ray of mixture with fixed proportions

of componentsof components Uses individual chemical dose-response curves plus Uses individual chemical dose-response curves plus

mixture curvemixture curve Inferences limited to mixing ratio testedInferences limited to mixing ratio testedIsobol = curve fitted to Isobol = curve fitted to points with fixed points with fixed responseresponseRay = curve fitted to Ray = curve fitted to points with fixed ratiospoints with fixed ratios

Ray Design

Chemical A Dose

0 2 4 6 8

Che

mic

al B

Dos

e

0

5

10

15

20

ED50

ED20ED10

1:5 ray

2:1 ray

4444

Advantages of Ray DesignsAdvantages of Ray Designs

Useful for any number of chemicalsUseful for any number of chemicals Economical and efficient design to test for Economical and efficient design to test for

interactionsinteractions Provides statistical test of additivityProvides statistical test of additivity Mixture of study can be tailored to address Mixture of study can be tailored to address

experimental question(s)experimental question(s) Hypothesis-testing as well as -generatingHypothesis-testing as well as -generating

4545

General Methodology for Additivity General Methodology for Additivity Analysis using Ray DesignsAnalysis using Ray Designs

Dose-response model is fit to single chemical dataDose-response model is fit to single chemical data Additivity model (predicted) along fixed ray is Additivity model (predicted) along fixed ray is

generated based on single-chemical data and generated based on single-chemical data and mixing ratio of each chemical mixing ratio of each chemical

Dose-response model (observed) is fit to Dose-response model (observed) is fit to experimental mixture dataexperimental mixture data

Fitted models (predicted vs observed) are tested for Fitted models (predicted vs observed) are tested for departure from additivity, departure from additivity, e.g.e.g., , • Equality of parameters for experimental and additivity Equality of parameters for experimental and additivity

modelmodel• Experimental model fits within confidence limits of Experimental model fits within confidence limits of

predicted modelpredicted model• Equality of statistically derived thresholdsEquality of statistically derived thresholds

4646

Considerations Using Ray DesignsConsiderations Using Ray Designs Adequately characterize shape of individual Adequately characterize shape of individual

and mixture dose responseand mixture dose response Dose-response characteristicsDose-response characteristics

• Maximal responsesMaximal responses• Slopes Slopes

Focus on chemical selections, Focus on chemical selections, combinations, and mixing ratios of combinations, and mixing ratios of interestinterest

Also important: dose-rate, sequence and Also important: dose-rate, sequence and route of administrationroute of administration

4747

Mixture of 5 Organophosphorus Mixture of 5 Organophosphorus PesticidesPesticides

Why OPs? Why OPs? • Widely used pesticides, still Widely used pesticides, still • Potential for human exposure to multiple OPs through Potential for human exposure to multiple OPs through

use on foods and other commercial crops, pets, use on foods and other commercial crops, pets, garden, homegarden, home

• Common mode of action (inhibition of Common mode of action (inhibition of acetylcholinesterase)acetylcholinesterase)

• Epidemiological studies implicate OPs for Epidemiological studies implicate OPs for neurological adverse effects not predicted by neurological adverse effects not predicted by individual chemicalsindividual chemicals

Why 5 OPs?Why 5 OPs?• Monitoring data show 99% of food products have Monitoring data show 99% of food products have 5 5

pesticide residues (USDA Pesticide Data Program)pesticide residues (USDA Pesticide Data Program)4848

Mixture of 5 Organophosphorus Mixture of 5 Organophosphorus PesticidesPesticides

Which OPs?Which OPs?• Relevance based on potential human exposures, Relevance based on potential human exposures,

usage patterns, food residuesusage patterns, food residues• Overlapping geographical usageOverlapping geographical usage• Chlorpyrifos, diazinon, malathion, acephate, Chlorpyrifos, diazinon, malathion, acephate,

dimethoatedimethoate These were among top 10 OPs in use in USThese were among top 10 OPs in use in US

What ratios?What ratios?• Proportions based on predicted dietary Proportions based on predicted dietary

exposures estimated by Dietary Exposure exposures estimated by Dietary Exposure Estimate Model (DEEMEstimate Model (DEEMTMTM))

4949

Environmentally Relevant Environmentally Relevant Proportions (Ratios)Proportions (Ratios)

Dose ratios Dose ratios 0.031 (chloryprifos) 0.002 (diazinon)0.031 (chloryprifos) 0.002 (diazinon) 0.825 (malathion) 0.04 (acephate)0.825 (malathion) 0.04 (acephate) 0.102 (dimethoate)0.102 (dimethoate)

Chlorpyrifos DEEM values

Acute Population Adjusted Dose – based on RfD5050

http://water.usgs.gov/nawqa/pnsp/usage/maps/

2002 Pesticide 2002 Pesticide Usage MapsUsage Maps

5151

Mixture of Organophosphorus Mixture of Organophosphorus PesticidesPesticides

Would we expect dose-additivity?Would we expect dose-additivity?• Default assumption, but…Default assumption, but…• Old literature (50’s, 60’s) shows non-additive Old literature (50’s, 60’s) shows non-additive

interactions in about half of binary OP interactions in about half of binary OP combinationscombinations

• Well-known OP interactions with malathion Well-known OP interactions with malathion due to inhibition of detoxifying enzymesdue to inhibition of detoxifying enzymes

• Several potential kinetic sites for interactionsSeveral potential kinetic sites for interactions• Recent data suggest non-additivity dependent Recent data suggest non-additivity dependent

on sequence of administrationon sequence of administration5252

ApproachApproach Use multiple endpoints to fully characterize Use multiple endpoints to fully characterize

interactionsinteractions• BrainBrain and blood ChE inhibition, and blood ChE inhibition, motor activitymotor activity, ,

gait score, tail pinch responsegait score, tail pinch response Evaluate influence of malathion in the Evaluate influence of malathion in the

mixture by removing it (reduced ray)mixture by removing it (reduced ray) Acute oral dosing, tested at 4 hr (time of Acute oral dosing, tested at 4 hr (time of

peak effect), male peak effect), male

Long-Evans rats, Long-Evans rats,

n=10/dosen=10/dose

Adult, PND17Adult, PND175353

Brain CholinesteraseBrain Cholinesterase

Malathion had no effect up to 500 mg/kgMalathion had no effect up to 500 mg/kg

ACEPHATE

Dose (mg/kg)

0 3 10 30 60 120P

erce

nt C

ontr

ol C

hE A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

| | | | | |

CHLORPYRIFOS

Dose (mg/kg)

012 5 10 20 25 30 50Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100

||| |||

DIAZINON

Dose (mg/kg)

05 25 50 75 125

150

250P

erce

nt C

ontr

ol C

hE A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

||||||

DIMETHOATE

Dose (mg/kg)

0 5 10 25 50 75Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100

| | | | | |

Moser et al., Tox. Sci. 86:101-115, 2005

5454

Brain CholinesteraseBrain CholinesteraseMixture DataMixture Data

BRAIN CHOLINESTERASE: FULL RAY

Total Mixture Dose (mg/kg)

010 55 100

200

300

450

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100Predicted additivityExperimental mixture

The full and reduced rays showed synergism, and were not different from each otherThe full and reduced rays showed synergism, and were not different from each other1.5 to 2.1-fold shift in ED20 or ED50; 6 to 19-fold shift in thresholds1.5 to 2.1-fold shift in ED20 or ED50; 6 to 19-fold shift in thresholds

BRAIN CHOLINESTERASE: REDUCED RAY

Total Mixture Dose (mg/kg)

0.01.

89.

617

.535

.052

.578

.8P

erce

nt C

ontr

ol C

hE A

ctiv

ity (

X±S

EM

)0

20

40

60

80

100

Predicted additivityExperimental mixture

Moser et al., Tox. Sci. 86:101-115, 2005

5555

Motor ActivityMotor ActivityACEPHATE

Dose (mg/kg)

0 3 10 30 60 120

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

| | | | | |

CHLORPYRIFOS

Dose (mg/kg)

012 5 10 20 25 30 50

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

| | | | | |

DIAZINON

Dose (mg/kg)

05 25 50 75 125

150

250

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

||||||

DIMETHOATE

Dose (mg/kg)

0 5 10 25 50 75

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

| | | | | |

Malathion had no effect up to 500 mg/kgMoser et al., Tox. Sci. 86:101-115, 2005 5656

Motor ActivityMotor ActivityMixture DataMixture Data

MOTOR ACTIVITY: FULL RAY

Total Mixture Dose (mg/kg)

010 55 100

200

300

450

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

Predicted additivityExperimental mixture

MOTOR ACTIVITY: REDUCED RAY

Total Mixture Dose (mg/kg)

0.01.

89.

817

.535

.052

.578

.8

Act

ivity

Cou

nts

(X±S

EM

)

0

40

80

120

160

200

240

Predicted additivityExperimental mixture

The full and reduced rays showed synergism, and were not different from each otherThe full and reduced rays showed synergism, and were not different from each other1.2 to 1.7-fold shift in ED20 or ED50; >3-fold shift in thresholds1.2 to 1.7-fold shift in ED20 or ED50; >3-fold shift in thresholds

Moser et al., Tox. Sci. 86:101-115, 2005

5757

Adult Mixture SummaryAdult Mixture SummaryEndpointEndpoint Full Ray*Full Ray* Reduced Reduced

Ray*Ray*Full vs. Full vs. Reduced**Reduced**

ED20/50 ED20/50 DifferenceDifference

Brain ChEBrain ChE YesYes YesYes NoNo 1.5-2.1X1.5-2.1X

6-19X threshold 6-19X threshold shiftshift

Blood ChEBlood ChE YesYes YesYes YesYes 1.2-1.9X1.2-1.9X

Motor ActivityMotor Activity YesYes YesYes NoNo 1.2-1.7X1.2-1.7X

>3X threshold >3X threshold shiftshift

Gait ScoreGait Score YesYes NoNo YesYes 1.6-1.7X1.6-1.7X

Tail Pinch Tail Pinch ResponseResponse

NoNo NoNo -- -- ----

* significantly different from additivity, greater-than-additive (synergism)* significantly different from additivity, greater-than-additive (synergism)** significant difference between full and reduced rays** significant difference between full and reduced rays

Moser et al., Tox. Sci. 86:101-115, 2005 5858

PND17 Brain CholinesterasePND17 Brain CholinesteraseAcephate

Dose (mg/kg)

0 3 10 30 60 120

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100 Chlorpyrifos

Dose (mg/kg)

0 1 2 5 10 20%

Co

ntr

ol C

hE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100 Diazinon

Dose (mg/kg)

0 2 6 15 30 6025

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

Dimethoate

Dose (mg/kg)

0 5 10 25 50 75

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100 Malathion

Dose (mg/kg)

010 50 100 250 500125

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

Moser et al., Tox. Sci. 92:235-245, 2006 5959

Brain CholinesteraseBrain CholinesterasePND17 Mixture DataPND17 Mixture Data

BRAIN CHOLINESTERASE: FULL RAY

Total Mixture Dose (mg/kg)

0 10 20 40 60 100 165

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

Experimental mixturePredicted additivity

BRAIN CHOLINESTERASE: REDUCED RAY

Total Mixture Dose (mg/kg)

0.0 1.8 3.5 7.0 10.5 17.5 28.9

% C

on

tro

l Ch

E A

ctiv

ity (

X±S

EM

)

0

20

40

60

80

100

Experimental mixturePredicted additivity

The full and reduced rays showed synergism, and they were different from each otherThe full and reduced rays showed synergism, and they were different from each other1.3 to 2-fold difference in ED20 or ED501.3 to 2-fold difference in ED20 or ED50

Moser et al., Tox. Sci. 92:235-245, 2006 6060

PND17 Mixture SummaryPND17 Mixture Summary

EndpointEndpoint Full Ray*Full Ray* Reduced Ray*Reduced Ray* Full vs. Full vs. Reduced**Reduced**

ED20/50 ED20/50 DifferenceDifference

Brain ChEBrain ChE YesYes YesYes YesYes 1.5-2.1X1.5-2.1X

Blood ChEBlood ChE YesYes YesYes YesYes 1.7-2.3X1.7-2.3X

Motor ActivityMotor Activity YesYes NoNo YesYes 1.3-2.6X1.3-2.6X

Gait ScoreGait Score YesYes YesYes YesYes 2.2-3X2.2-3X

Tail Pinch Tail Pinch ResponseResponse

YesYes NoNo YesYes 3.5X3.5X

* significantly different from additivity, greater-than-additive (synergism)* significantly different from additivity, greater-than-additive (synergism)** significant difference between full and reduced rays** significant difference between full and reduced rays

Moser et al., Tox. Sci. 92:235-245, 2006 6161

Summary of OP MixturesSummary of OP Mixtures

Greater-than-additive interactions (Greater-than-additive interactions (i.e.i.e., , synergism) detected with both mixtures at synergism) detected with both mixtures at both agesboth ages

• Interactions depended on endpointInteractions depended on endpoint• Significant differences at low end of dose-Significant differences at low end of dose-

response (threshold)response (threshold)• Comparing the reduced to the full ray indicated Comparing the reduced to the full ray indicated

an influence of malathion on most endpointsan influence of malathion on most endpoints Degree of influence depended on endpointDegree of influence depended on endpoint

6262

Mixture of 7Mixture of 7 N N-Methyl Carbamate -Methyl Carbamate PesticidesPesticides

Why carbamates?Why carbamates?• Broad agricultural and residential usesBroad agricultural and residential uses• Exposures from food residues, drinking water, Exposures from food residues, drinking water,

and home – dermal, inhalation, oraland home – dermal, inhalation, oral• Common mode of action (inhibition of Common mode of action (inhibition of

acetylcholinesterase)acetylcholinesterase) Why 7 carbamates?Why 7 carbamates?

• Of 10 carbamates being regulated, these 7 Of 10 carbamates being regulated, these 7 had high contribution to cumulative risk had high contribution to cumulative risk assessmentassessment

6363

Mixture of 7 Mixture of 7 NN-Methyl Carbamate -Methyl Carbamate PesticidesPesticides

Which carbamates?Which carbamates?• High usage and contribution to cumulative risk High usage and contribution to cumulative risk

assessmentassessment• Carbaryl, carbofuran, formetanate HCl, Carbaryl, carbofuran, formetanate HCl,

methiocarb, methomyl, oxamyl, propoxurmethiocarb, methomyl, oxamyl, propoxur What ratios?What ratios?

• Proportions based on relative potencies using Proportions based on relative potencies using BMD10 (10% brain ChE inhibition)BMD10 (10% brain ChE inhibition)

• Proportions based on California database of Proportions based on California database of tonnage sold in 2005tonnage sold in 2005 Surrogate for total (aggregate) exposuresSurrogate for total (aggregate) exposures

6464

Relative PotenciesRelative Potencies

Ald

icar

bFo

rmet

anat

eC

arbo

fura

nO

xam

ylTh

iodi

carb

Met

hom

ylM

ethi

ocar

bC

arba

ryl

Pro

poxu

rP

irim

icar

b

RP

Fs

(95%

co

nfi

den

ce li

mit

s)

0.01

0.1

1

10

N-Methyl Carbamate Cumulative Risk Assessment, 2007

6565

California DatabaseCalifornia Database

http://www.cdpr.ca.gov/docs/pur/purmain.htm 6666

Carbamate ProportionsCarbamate Proportions

Relative Potency Relative Potency Factor MixtureFactor Mixture

CarbarylCarbaryl .42.42 PropoxurPropoxur .29.29 MethiocarbMethiocarb .20.20 MethomylMethomyl .05.05 FormetanateFormetanate .02.02 Oxamyl Oxamyl .01.01 CarbofuranCarbofuran .01.01

CA Environmental CA Environmental MixtureMixture

MethomylMethomyl .41.41 CarbarylCarbaryl .39.39 Oxamyl Oxamyl .13.13 CarbofuranCarbofuran .04.04 Formetanate Formetanate .03.03 MethiocarbMethiocarb .003.003 PropoxurPropoxur .002.002

6767

ApproachApproach Use several endpoints to characterize Use several endpoints to characterize

interactionsinteractions• BrainBrain and RBC ChE inhibition, and RBC ChE inhibition, motor activitymotor activity

Evaluate influence of mixing ratiosEvaluate influence of mixing ratios Acute oral dosing, tested at 40 min (time of Acute oral dosing, tested at 40 min (time of

peak effect), male Long-Evans rats,peak effect), male Long-Evans rats,

n=10/dosen=10/dose

Adult, (PND17)Adult, (PND17)

6868

Brain CholinesteraseBrain Cholinesterase

PROPOXUR

Dose (mg/kg)

0.00.

31.

03.

010

.020

.0

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100

CARBARYL

Dose (mg/kg)

0.0

3.0

7.5

15.0

30.0

50.0

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100METHIOCARB

Dose (mg/kg)

0.00.

52.

05.

012

.025

.0

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100

METHOMYL

Dose (mg/kg)

0.0

0.1

0.3

0.6

1.3

2.5

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100

6969

Motor ActivityMotor Activity

PROPOXUR

Dose (mg/kg)

0.00.

31.

03.

010

.020

.0

Act

ivity

Cou

nts

(X±S

EM

0

0

50

100

150

200

CARBARYL

Dose (mg/kg)

0.0

3.0

7.5

15.0

30.0

50.0

Act

ivity

Cou

nts

(X±S

EM

)

0

50

100

150

200

METHIOCARB

Dose (mg/kg)

0.00.

52.

05.

012

.025

.0

Act

ivity

Cou

nts

(X±S

EM

)

0

50

100

150

200

METHOMYL

Dose (mg/kg)

0.00

0.10

0.25

0.60

1.25

2.50

Act

ivity

Cou

nts

(X±S

EM

)

0

50

100

150

200

7070

Brain CholinesteraseBrain CholinesteraseRPF Carbamate MixtureRPF Carbamate Mixture

N-Methyl Carbamate Cumulative Risk Assessment, 2007

Confidence limits analysis suggests additivity7171

Brain CholinesteraseBrain CholinesteraseRPF Carbamate MixtureRPF Carbamate Mixture

Test of additivity not rejected (p=0.066)Test of additivity not rejected (p=0.066)

BRAIN CHOLINESTERASE: RELATIVE POTENCY MIXTURE

Dose (mg/kg)

0.00.

31.

33.

78.

615

.9

Per

cent

Con

tro

l ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100 Predicted additivityExperimental mixture

7272

Motor ActivityMotor ActivityRPF Carbamate MixtureRPF Carbamate Mixture

Test of additivity not rejectedTest of additivity not rejected

MOTOR ACTIVITY: RELATIVE POTENCY MIXTURE

Mixture dose (mg/kg)

0.00.

31.

33.

78.

615

.9

Act

ivity

Cou

nts

(X±S

EM

)

0

50

100

150

200

250Predicted additivityExperimental mixture

7373

Brain CholinesteraseBrain CholinesteraseCA Carbamate MixtureCA Carbamate Mixture

BRAIN CHOLINESTERASE: CALIFORNIA ENVIRONMENTAL MIXTURE

Dose (mg/kg)

0.00.

10.

61.

42.

86.

0

Per

cent

Con

trol

ChE

Act

ivity

(X

±SE

M)

0

20

40

60

80

100 Predicted additivityExperimental mixture

Preliminary statistical analyses reveal non-additivity (synergy)Preliminary statistical analyses reveal non-additivity (synergy) 7474

Motor ActivityMotor ActivityCA Carbamate MixtureCA Carbamate Mixture

MOTOR ACTIVITY: CALIFORNIA ENVIRONMENTAL MIXTURE

Dose (mg/kg)

0.00.

10.

61.

42.

86.

0

Act

ivity

Co

un

ts (

X±S

EM

)

0

50

100

150

200

Predicted additivityExperimental mixture

Preliminary statistical analyses reveal non-additivity (synergy)Preliminary statistical analyses reveal non-additivity (synergy) 7575

Summary of 7-Carbamate MixturesSummary of 7-Carbamate Mixtures

Additivity (zero interaction) with mixture Additivity (zero interaction) with mixture ratios based on relative potency factorsratios based on relative potency factors

Greater-than-additive interactions (Greater-than-additive interactions (e.g.e.g., , synergy) with mixture ratios based on synergy) with mixture ratios based on amounts sold in Californiaamounts sold in California

Differences in outcome based on mixing Differences in outcome based on mixing ratios of componentsratios of components

7676

Other Studies, Similar MethodologyOther Studies, Similar Methodology 4 DBPs producing hepatotoxicity (mice)4 DBPs producing hepatotoxicity (mice)11

• Ratio based on average seasonal proportion of 35 water treatment Ratio based on average seasonal proportion of 35 water treatment facilitiesfacilities

• No departure from additivityNo departure from additivity 18 PHAHs decreasing serum T4 (rats)18 PHAHs decreasing serum T4 (rats)22

• Ratio based on average concentrations found in human breast milk Ratio based on average concentrations found in human breast milk and food sourcesand food sources

• Concentrations in range of human body burdensConcentrations in range of human body burdens• Synergy emerged as dose increasedSynergy emerged as dose increased

6 synthetic estrogens producing estrogenic actions (6 synthetic estrogens producing estrogenic actions (in vitroin vitro ER-ER-αα reporter gene and reporter gene and in vivoin vivo uterotrophic assays) uterotrophic assays)33

• With and without phytoestrogensWith and without phytoestrogens• Ratios based on relative potenciesRatios based on relative potencies• Interactions depended on concentrations and components of mixture Interactions depended on concentrations and components of mixture

1 Gennings et al., J Agr Biol Environ Stat, 2:198-211, 19972 Crofton et al., Env Health Perspec 113:1549-1554, 20053 Charles et al., Tox Appl Pharm 218:280-288, 2007 7777

Considerations for Environmentally Considerations for Environmentally Relevant Mixture ResearchRelevant Mixture Research

Appropriate experimental design and statistical Appropriate experimental design and statistical analysesanalyses• Specify dose- or response-additivity hypotheses, design Specify dose- or response-additivity hypotheses, design

and analyze experiment appropriatelyand analyze experiment appropriately

Strategically select specific exposure scenariosStrategically select specific exposure scenarios• Potentially worrisome chemicals, Potentially worrisome chemicals, e.g.e.g., high-use, , high-use,

environmentally persistent environmentally persistent • Rational mixing ratios, Rational mixing ratios, e.g.e.g., reflecting potential or known , reflecting potential or known

human exposurehuman exposure• Site-specific combinations and ratiosSite-specific combinations and ratios

Use of fixed-ratio ray designs can provide efficient Use of fixed-ratio ray designs can provide efficient and focused research of mixturesand focused research of mixtures 7878

Cadillac of Mixture AssessmentsCadillac of Mixture Assessments

Quantitative component-based mixtures risk Quantitative component-based mixtures risk assessment that includes:assessment that includes:• Exposure scenarios reflective of human exposuresExposure scenarios reflective of human exposures• Environmental relevance in composition of mixtureEnvironmental relevance in composition of mixture• Defined dose-response data addressing common Defined dose-response data addressing common

toxic pathwaytoxic pathway• Experimental data on actual mixturesExperimental data on actual mixtures• Evaluation of additional influences, Evaluation of additional influences, e.g.e.g., age, gender, , age, gender,

etc.etc.• Biologically based modeling (Biologically based modeling (e.g.e.g., PBPK) to describe , PBPK) to describe

interactionsinteractions

7979

Volkswagen of Mixture AssessmentsVolkswagen of Mixture Assessments

Less data-intensive approaches for Less data-intensive approaches for (partially) defined mixtures(partially) defined mixtures

Given exposure data and some measure Given exposure data and some measure of acceptable level (of acceptable level (e.g.e.g., RfD/C), RfD/C)• Hazard quotient (HQ) or index (HI)Hazard quotient (HQ) or index (HI)• Target organ toxicity dose (TTD)Target organ toxicity dose (TTD)• Toxicity equivalence factor (TEF)Toxicity equivalence factor (TEF)

Given only compositionGiven only composition• Analysis of sufficiently similar mixturesAnalysis of sufficiently similar mixtures

8080

ChallengesChallenges Determination of key events of componentsDetermination of key events of components

• Target organ and toxicityTarget organ and toxicity No complete dose-response data of componentsNo complete dose-response data of components

• Some statistical methods address thisSome statistical methods address this Not all components are identifiedNot all components are identified

• Evaluate data for similar mixtureEvaluate data for similar mixture• Evaluate known partial mixture with and without Evaluate known partial mixture with and without

undefined fractionundefined fraction Exposure and responseExposure and response

• ChronicityChronicity• TimingTiming• Aggregated routesAggregated routes• Dose-dependent transitionsDose-dependent transitions

These are research areas being proposed in NHEERL

8181

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US Environmental Protection Agency, Research Triangle Park, NC 8282

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