Document a4w2EdaO4YqynR2x0QZVQkqMN

3M ENVIRONMENTAL LABORATORY Protocol-AnalyticalStudy QuantitativeAnalysisofPerfluorooctaneSulfonicAcid PotassiumSalt (PFOS, T-6295)inCynomolgus Monkeys FollowingAdministrationofa 4- Week CapsuleToxicityStudy 3M ET & SS StudyNumber: AMDT-041598.1 In-VivoStudyReferenceNumber: Covance6329-222 TestSubstance:PerfluorooctaSnuelfoniAccid PotassiumSalt(PotassiuPmerfluorooctane SulfonateC),F17SO3-K'(PFOS,T-6295) Name and AddressofSponsor: 3M ToxicologyServices Building220-2E-02,3M Center St.Paul,MN 55144-1000 Name and AddressofBiological-PhaFsaecility: Covance LaboratorieIsnc. P.O.Box 7545 Madison,WI 53707-7545 Name and AddressofAnalyticaTlestingFacility: 3M EnvirorunentTaelchnologyand Services 935 Bush Avenue St.Paul,MN 55106 DateofApprovalby Sponsor: 4/21/98 ProposedAnalyticaIlnitiatiDoante: 7/15/98 ProposedAnalyticaClompletionDate: 7/15/99 Author:Glenn Langenburg Ki@idyDwctCEe5@@/ Pr'm'cipAanlalyticaIlnvestigator Date 1.0 PURPOSE This studyisdesignedto quantitativedleyterminetheextentofabsorptionofPFOS and various metabolitesincynomolgus monkeys orallyadministeredPFOS. 2.0 REGULATORY COMPLIANCE Thisstudywillbe conductedinaccordancewiththeEPA Good LaboratoryPracticeregulations 40 CFR part792 and applicablceurrentEnvironmentalTesting& SafetyServices(ET&SS) SOP's and methods (seesections8.0 and 9.Ofor specifiScOP's and methodsrelatedtothis protocoo. 3.0 TEST MATERIALS 3.1 Test,Control, and Reference Substances and Matrices 3.1.1 AnalyticalReference Substance: PerfluorooctaneSulfonicAcid Potassium Salt (C8F,7SO3-K@ (PFOS, T-6295)) 3.1.2 AnalyticalReference Substance Matrix: Cynomolgus monkey serum and liver tissue 3.1.3 AnalyticalControl Substance Matrix: Cynomolgus monkey serum and liver tissue(Controltissue1,05346). 3.2 Source ofMaterials 3.2.1 AnalyticalReference Substance: 3M ICP/PCP Division 3.2.2 AnalyticalReference Substance Matrix: Covance ResearchProducts(The sourceof themonkeys willbe documented intheraw data) 3.2.3 AnalyticalControl Substance Matrix: Covance ResearchProducts.The tissue was from a controlanimal,105346. 3.3 Number ofTest and Control Samples 3.3.1 Test samples Number Dose Level Group & Route PFOS Collections' Sex (mWWday) 1 2 Male Orallyby 0 0 Blood predose(-6daysfrom dosing),and (Control) 2 Female gelcapsules on Days 2, 7,14,and29. 2 3 Male Orallyby 0.02 (low-dose) 3 Female gelcapsules 0 Livertissuoen Day 29. 0 Bloodpredose(-6daysfromdosing)a,nd on Days 2,7 14,and29. 0 -Livertissuoe'n Day 29. 3 1Male Orallyby 2.0 0 Bloodpredose(-6daysfromdosing)a,nd (high-dose)1Female gelcapsules on Days 2,7,14,and29 1 1 0 Livertissuoen Day 29. 1-Bloodcollectefdromanimalswillbe centrifugbeydCovance,serumharvesteda,ndserumshippedto3M ET&SS foranalysis 3.4 Identificationof Test and Control Samples. Test samples willbe identifiedby Covance atthetime ofcollection. 2 3.5 PuritayndIdentiotfyMaterialCsh.aracterizaotftihoentesmtateriailstshe responsibiliotfytheSponsor. 3.6 StabilitoyfReferenceMateriaL Testingforthestabiliotfythetestmaterialunderthe conditionsof administratioinstheresponsibiliotfytheSponsor. 3.7 Storage Conditions for Test Materials. Analyticalreferencesubstancewillbe storedat ambienttemperature.Analyticalreferencmeatriceswillbe storedator below approximately-20*C,untilextractionA.llextractwsillbe storednominallyat4'C until analyzed.Testand Controlsampleswillbe receivedaccordingtoAMDT-S-10. 3.8 Dispositionof Specimens. Biologicaltissueasnd fluidswillbe retainedfortherequired time periodas perGLP regulatio4n0 CFR part792.195forstudieslongerthan 28 days. 3.9 SafetyPrecautions.RefertotheMaterialSafetyData Sheetsforthesubstancesused for associatesdafetyinformationE.xercisecautionwhen handlingknivesforcuttingthe sample matrices.Safetyglassesp,rotectivcelothingand appropriatheand protection shouldalwaysbe wom when working intheEnvironmentalLaboratory. 4.0 EXPERIMENTAL - Overview Animals were administeregdelcapsulesorallyd,aily(7 days/week)for28 days.Dose levelsof PFOS inthecapsulevaried(0,0.02,and 2.0mg/kg) between groups(controll,ow-dose,and high-d,osreespectivelyB)l.ood sampleswere collecteadtregularinterval(ssee3.3.1).Liver tissuesampleswere collecteadtthetimeof sacrific(eDay 29). Seraand livertissuewere shippedto 3M ET&SS fortheanalyticaplortionofthisstudy.See Covance Study #CHW 6329- 222 forinformationregardingthein-lifpeortionof thestudy. The seraand livertissuewillbe extractevdiaan ionpairingmethod,or othermethod as appropriatea,nd thenanalyzedviaElectrospraIyonization/MasSspectrometry(Tandem Mass SpectrometryE)SI/MS(MS), or otherappropriatteechniquef,orPFOS and otherfluorochemical metabolites. 5.0 EXPERIMENTAL- AnalyticalMethods Livertissueand serawillbe targetedforanalysisM.ethods areavailableforextractioannd analysiosf fluorochemicalfsrom seraand livertissuevariousanimalmatrices(seeattachments). The methods willbe validatefdorcynomolgus monkey matricespriortoor concurrentlwyith sample analysis. 6.0 DATA ANALYSIS Datawillbe reportedasthemilligrams(mg) ofPFOS or metaboliteperunitof tissueo,r asa concentratioonf fluoridpeertissueo,r inotherclearldyefinedunits,asappropriateQ.uantitation 3 willbe performedusingcalibratiocnurves.These curveswillbe determinedusinglinear regressioannalysis.Statistiucsed,atthediscretioonfthePrincipaIlnvestigatomra,y include averagesand standarddeviationosfconcentrationfsordifferendtose groups.Ifnecessary, simplestatistictaelstssuch as Student'st-tesmtay be appliedtodeterminestatisticdailfference. No finalreportwillbe preparedforthisstudy.A datapackagewillbe providedtoCovance for inclusioinntheirfinalreportforCHW 6329-222. 7.0 MAINTENANCE OF RAw DATA AND RiEcoRDs 7.1 The followingraw dataand recordswillbe retainedinthestudyfolderinthearchives accordingtoAMDT-S-8. 7.1.1 Approved protocoland amendments 7.1.2 Study correspondence 7.1.3 Shippingrecords 7.1.4 Raw data 7.1.5 Approved finalreport(originaslignedcopy) 7.1.6 Electronicopiesofdata 7.2 Supportingrecordstobe retainedseparatelfyrom thestudyfolderinthearchivesaccording toAMDT-S-8 willincludeb,utnotnecessarilbye limitedto,thefollowing: 7.2.1 Trainingrecords 7.2.2 Calibratiornecords 7.2.3 Instrumentmaintenancelogs 7.2.4 StandardOperatingProcedures,Equipment Procedures,and Methods 7.2.5 Appropriatespecimens 8.0 REFERENCES 8.1 8.2 8.3 8.4 8.5 8.6 8.7 8.8 8.9 8.10 8.11 8.12 8.13 8.14 AMDT-S-1, Chemical Tracking AMDT-S-2, Solutionsand StandardsMaking AMDT-S-3, PersonnelTraining AMDT-S-4, GeneralLab DocumentationSystems AMDT-S-5, GLP-relatedDocumentationSystems ANOT-S-6, GeneralLab Records ANMT-S-7, GLP-relatedRecords AMDT-S-8, Archives AMDT-S-9, GLP Program and Responsibilities AMDT-S- 10,Sample TrackingSystem AMDT-S- 12,AnalyticaMlethod Validation AMDT-S- 15,LaboratoryPracticeasnd Data Management AMDT-S-16, QualityAssuranceUnit AMDT-S-17, AnalyticaElquipment SystemsDocumentation 4 8.15 AMDT-S-18, RoutineCalibratioCnhecks ofBalances 8.16 AMDT-S-20, DailyCalibratioCnhecks ofAutomaticPipettors 8.17 AMDT-S-21, Annual CalibratioCnhecks ofAutomaticPipettors 9.0 ATTACHMENTS 9.1FACT-M-1.0 Extractioonf PFOS or OtherAnionicFluorochemicalSurfactantfsrom Liver Using AnalysisUsing HPLC ElectrospraMyass Spectrometry 9.2FACT-M-2.0 Analysisof FluorochemicalisnLiverExtractsusingHPLC Electrospray/Mass Spectrometry 9.3FACT-M-3.0 Extractioonf PFOS or OtherAnionicFluorochemicalSurfactantfsrom Serum Using AnalysisUsing HPLC ElectrospraMyass Spectrometry 9.4FACT-M-4.0 Analysisof FluorochemicalsinSerum ExtractsusingBPLC Electrospray/Mass Spectrometry 5