Document wr7vVKdyRY1oNY97Jb3YDrzpE

YEND-PuOP-373. Coding AR 226 _ 0ST/ FINAL REPORT PROTOCOL 418-014 - `ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS `SPONSOR'S STUDY NUMBER: T-6295.13 FINAL REPORT DATE: 23 Y 1999 001285 PROTOCOL 418-014 ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS SPONSOR'S STUDY NUMBER: T-6295.13 TAOFBCOL NTEE NTS SUBJECT PAGE I. SUMMARY AND CONCLUSION 1 A. Methods 1 B. Results 4 C. Conclusion 6 Il. DESCRIPTION OF TEST PROCEDURES 1 A. Conduct of Study 11 AA. Sponsor 1 A2. Testing Facilty 1-1 A3. Study Number 11 Ad. Sponsor's Study Number 11 AS. Purpose of the Study 1 AS. Study Design 11 A7. Regulatory Compliance 11 ! 001286 SUBJECT A8. Ownership of the Study AS. Study Monitor A10. Altemate Study Monitor A11. Study Director A12. Technical Performance A13. Report Preparation A14. Report Review A.15. Date Protocol Signed A.16. Dates of Technical Performance A.17. Records Maintained B. Test Atticle Information B.1. Description B.2. LovBatch Number B.3. Date Received and Storage Conditions B.4. Special Handing Instructions B.5. Analysis of Activity C. Vehicle Information C.1. Description C2. Lot Numbers C.3. Dates Received and Storage Conditions C4. Special Handling Instructions C5. Analysis of Purity PAGE 2 2 2 2 2 2 2 2 1-3 +3 1-3 3 1-3 1-3 4 14 4 14 14 Ih4 4 4 001287 SUBJECT D. Test Article Preparation and Storage Conditions D.1. Sample Information D.2. Analytical Results E. Test System E.1. Species E.2. Strain E.3. Supplier (Source) E4. Sex E.5. Rationale for Test System E.6. Test System Data E.7. Breeder Male Rat Data E.8. Method of Randomization E.S. System of Identification F. Husbandry F.1. Research Facility Registration F.2. Study Rooms F.3. Housing F.4. Lighting F.5. Sanitization F.6. Feed F.7. Feed Analysis F.8. Water fii PAGE 1-5 1-5 1-5 Im-5 1-5 I-5 1-8 1-6 1-6 6 1-6 1-6 7 n-8 1-8 1-8 1-8 1-8 1-8 1-8 1-9 1-8 001288 susJecT - F.9. Water Analysis F.10. Bedding F.11. Bedding Analysis G. Methods G.1. Dosage Administration G2. Rationale for Dosage Selection G3. Routeof Administration G.4. Rationale for Route of Administration G5. Frequency of Administration G6. Length of Study G7. Method of Study Performance H. Gross Necropsy HA. Fo Generation Rats H.2. F1 Generation Pups I. Statistical Analyses Wl. RESULTS A. Mortality, Clinical and Necropsy Observations B. Body Weights C. ACbosnosluumtpet(igo/ndaVya)launeds Relative (g/kg/day) Feed D. Natural Delivery and Litter Observations D.1. Natural Delivery Observations D2. Cross-Fostering Litter Observations v PAGE 9 9 Io 10 I-10 1110 1-11 111 111 111 1-11 1-15 11-15 1-16 147 1 i-1 1 1-2 n-2 2 1-2 001259 SUBJECT PAGE E. Clinical Observations from Necropsy Observations Birth to Day 21 Postpartum and "3 F. Electron Microscopic Evaluation Day 1 Postpartum Pups of the Liver and Lung from 4 FA. Liver iil F2. Lung 4 REFERENCES Ils APPENDIX A - REPORT FIGURES Figure 1. Matemal Body Weights At Figure 2. Matemal Body Weights - Rats Assigned to Cross Fostering ~~ A-2 APPENDIXB - REPORT TABLES Table 1. Clinical Observations - Summary - Fo Generation Female Rats 81 Table 2. Necropsy Observation-s Summar-y Fo Generation Female Rats B4 Table 3. Body Weights - Precohabitation - Summar-y Fo Generation Female Rats 85 Table 4. Body Weight Changes Fo Generation Female - Precohabitation Rats - Summary - 8-6 Table 5. Matemal Body Weights - Gestation - Summary - Fo Generation Female Rats 87 Table 6. Matemal Body Weight Changes - Gestation - Summary - Fo Generation Female Rats 88 Table 7. Matemal Body Weights - Lactation - Summary - Fo Generation Female Rats 8-9 Table 8. Matemal Body Weight Changes - Lactation - Summary - Fo Generation Female Rats 8-10 v 001290 SUBJECT Table 9. PAGE Absolute Feed Consumption Precohabitation - Summary - Values (g/day) Fo Generation Female Rats ~~ B-11 Table 10. Relative Feed Consumption Values (g/kg/day) Precohabitation -- Summary - Fo Generation Female Rats ~~ 8-12 Table 11. Matemal Absolute Feed Consumption Values (g/day) - Gestation - Summary - Fo Generation Female Rats, B-13 Table 12. GMeastteamtaiolnR-elSautmivmeaFreye-dFCoonGsenuemrpattiioonn VFaelmuaelse(Rga/tksg/day-) B14 Table 13. Matemal Absolute Feed Consumption Values (g/day) - Lactation - Summary - Fo Generation Female Rats B-15 Table 14. Matemal Relative Feed Consumption Values (g/kg/day) - Lactation - Summary - Fo Generation Female Rats B-16 Table 15. Natural Female Delivery Rats Observations - Summary - Fo Generation B17 Table 16. LitterObservations (Naturally F1 Generation Litters Delivered Pups) - Summary - B20 Table 17. Clinical Observations from Summary - F1 Generation Birth to Pups Day 21 Postpartum - B24 Table 18. Necropsy Observations - Summary- F1 Generation Pups B-25 Table 19. Clinical Observations - Individual Data - Fo Generation Female Rats B26 Table 20. Necropsy Observations Female Rats - Individual Data - Fo Generation 8-30 Table 21. Body Weights Fo Generation - Precohabitation Female Rats - Individual Data - 8-33 Table 22. Matemal Body Weights - Presumed Data - Fo Generation Female Rats Gestation - Individual B45 Table 23. Matemal Body Weights - Lactation - Individual Data -- Fo Generation Female Rats 8-53 vi 001291 suBJECT Table 24. Table 25. PAGE DFaetead -CFoonsGuemnpetriaotinoVnaFlueemsal-ePrReactoshabitation - Individual 8:59 Matemal Feed Individual Data Consumption Values - Presumed - Fo Generation Female Rats Gestation - 8-63 Table 26. Matemal Feed Consumption Values - Lactation - Individual Data - Fo Generation Female Rats B-71 Table 27. Natural Delivery, Implantation Sites, and Pup Viability and Sex - Individual Data - Fo Generation Female Rats/ F1 Generation Litters B-74 Table 28. Duration of Gestation and Average Duration of Delivery F(MoinGuetneesr)atPieornPFuepmaPleer LRiatttesr - Individual Data- 877 Table 29. PPousptpBaordtyumWe-iIgnhdtivLiidtutaelr ADvaetara-gFe1s GfernoemrBaitritohntoLiDttaeyrs21 8-80 Table 30. Pup Body Weights from Birth to Day 21 Postpartum Individual Data - F1 Generation Pups B-83 Table 31. Pup Vital Status and Sex from Birth to Day 21 Postpartum Individual Data - F1 Generation Pups B-99 Table 32. Clinical Observations from Birth to Day 21 Postpartum Individual Data - F1 Generation Pups B-102 Table 33. Necropsy Observations - Individual Data - F1 Generation Pups B-104 APPENDIX C- PROTOCOL AND AMENDMENT C-1to C41 APPENDIX D - DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY D-1 APPENDIX E - TEMPERATURE AND RELATIVE HUMIDITY REPORTS E-1t0E-7 APPENDIX F - PATHOLOGY REPORT F-1to F-46 APPENDIX G - STATEMENT OF THE STUDY DIRECTOR vii G-1 001292 SUBJECT APPENDIX H- QUALITY ASSURANCE UNIT FINAL REPORT STATEMENT PAGE H-1to Hs viii 001293 418-014:PAGE I-1 TITLE: ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS ARGUS RESEARCH LABORATORIES, INC. PROTOCOL NUMBER: 418-014 SPONSOR'S STUDY NUMBER: T-6295.13 1 IMMARY AND CONCLUSION A. Methods Seventy-eight were assigned vtiorgtiwn ofedmoaslaegCergir:oCuDpsB(RGrVoAuFp/sPl| aunsd (Sprague-Dawley) rats Il), 42 rats in the control group and 36 rats in the treated group. The rats were administered the test article, PFOS (FC-95), or the vehicle, 0.5% Tween 80, orally (via gavage), once daily beginning 42 days before cohabitation through day 21 postpartum (DL. 21). cThhaendgoessaagnedvgoilvuemneawt aaspp5romxLi/mkagt.elyDotsheagseasmweetriemeadejaucshteddayd.aily for body weight dReaptesnwdeirneg ass on itgheneodrdteorcorfosdse-lfiovsetreyrianngdotrhpehaavramialcaboiklitnyeotficra`tssafmoprlecrcooslsl-efcotsitoenr in g . TTwheentlyit-tferisvewelriteterassspiergndeodsasugcehgtrhoautpf were assigned to our subsets were cross-fostering on DL 1. obtained among the two dosage groups as described in the table below. [C I [ DCosaT|T S Mo-- c0atan1 mTeT |oowovueress1|||7 rI 2 itnoTe ||T UTweoemiseosss ||55a]]] Group. Rats (mg/kg/day)| Reassigned (Dam Treatment) Eight female rats from the control group and two female rats from the treated group were assigned to pharmacokinetic sample collection following delivery. These rats were not cross-fostered. a. Detailed descriptions of all procedures used in the conduct of this study are provided in the appropriate sections of this report and in APPENDIX C (PROTOCOL AND AMENDMENT). 001294 418-014:PAGE 1-2 A. Eo Generation Rats Assigned to Cross-Fostering: poTfrheetmhaeFtosutrgueednydeeralanitvdieorfinoerfscelaminnadil/ceaolrroadbtessaewtrehvrsaetbioeofbnossreeorfvaeenfddfefacoptrspvriooafxbtiihletytaetstlearatsitcltew,iacbeoretaicohnsd,ay Spaocsritfdiocse.ageF.eeBdocdoynsweuimgphttisonwevraelureescworedreedrdeaciolryddeudriwnegemtkahlteyedtlooyscoaonngeaebhipoteaurtriioodn,andadilayt during the gestation period and on DLs 1, 4, 7, 10 and 14. dlhiiteeedr fbseiemzfeaosrleeanirnaditispaluwpeexrvaeimaiebnvilaatltyuiaoattnebodiffrotthr.hedPuliurtapetsrifotonhraotvfiaegbieitslhitetarytiaopnp,elaernegdthstoifllpasrotmuroirtitohn,a bePShtuaaptvsuiswoeartrwebairestxh.armeMicanotreeddemddaulribneghatvheio2r1o-fdathyepdosatmpsarwtausm epvewraielorudae.teeOdxbadsmaeiilrnyveewddhfeomrnavttiethaemlal on DLs 1, 4, 7, 14 and 21 mbJlTaowtoeednmtasyla-mfrpoaultsre.hsoTwuhreeruerbilcnooeolaldencsdtaefmdepcolanelssDawLme2pr2le,ecsveinwatertrihefeucgieonfdle.lreicotrevdefnraomcaDvLas, 21 to 22 from the and samples and the serum were shipped to the Sponsor. The urine and focal All Fo gross generation rats necropsy of the assigned thoracic, The number and distribution of atbodcormoissn-aflosatnedripneglvwiecrveisscaecrraifwicaesd implantation sites were on DL 22 and performed. a (right lateral lobe) from each dam was collected and recorded. A shipped to the fiver section Sponsar alRevastaiso;nsts.haatmApdlibdelsonoowdtesdreaelmicpveelnretrawialfiustgeecrdowlaelnrecdetessdaecrprruiifmoircsetadomospanlcerDisfGiwce2er5viaantdheexianfmeirinoerdvefonregross fsSahpsiotnppspuoerpd.wtoaAsthlefiovSeurpnosdnescdtoeirao.dn (Draigmhtslwaitetrhalnolosbuer)vfivrionmgepaucphsdwaeemreswhsaiaspcrprieefditcateiodnteahdfetaanrdthe , missing or presumed cannibalized, A:2. F1 Generation Pups Assigned to Cross-Fostering: eDvaaylu1atoefdlafcotravtiiaobni(ltpyosattplaeratsutmt)wwicaesedaecfhindeadyaosf tthhee Pups present in each liter were counted once each day of birth. postpartum day. Each fitter was period. The wveesiigohrtasedweforretrheecpourpdesdoonnceDeLasc1h day during (birth), 4, 7, the 14 postpartuPmhypseircioadl. siPgnusp and 21 and at sacrifice were body dlPeeusapidsonosfnoauDnndLdsfdoe2ra0tdhe4o,rcatshuaecsreliufoinfcgedsdeaabntedhcatouhrestehlieovefmromwroeriribebuunpndrdeicstoeynrdwvietiroen.exFaomrianlel dpfuoprsgfroousnsd ed. Pups with gross 001295 418-014:PAGE 1-3 tesions found on 10 21, the lungs, DLs liver 2 to and g4rowsesrelepsrieosnserwveerdeapnrdesfeorrvpeudp.s found dead on DLs 5 TfOhenmeaDllLuen4pg,uscparsnasdpse-trfhoeitsletirev,reerwdwheFer1reegpeprnoesessrieabrtlvieeo.dn.lPiuTtpehrses wwlieutnrhgesgrcuolslsedletsoiofnivsewmearlseparnedsefrivveed. afrcohm tdhoesfaigres tgernoucuplolendepaucphs dfaryom(ceoantcrholdopsooalgeonglryoouanpnDd(iLtehs1)ep. eficTvtehirevweelruonfegIsctoaleln)edcfttroharem wliivtehrowuetreevarleutaatiinoend.. Remaining culled pups were sacrificed and discanded slOeenslieoOcntLs.ed21fG,orraolclsospllulepecsstiiwoonnesroewfepsroaecorlriefeditcaseidanemvdpi.aledSseicx(aplpeiitrtteairttsieporen)raofsnudbseextamwienreerdafnodrogmrolsys ttshiaeemrpS)lpaoennsdswoterhr.eescaomlplelcetsedweviraetcheentirnifefruigoerd.veTnahecasvearufmrosmabemlapoclohdesapunwpde,rleipvoesorh.liepBdple(adppedtro A-3. Eo Generation Rats Assigned to Pharmacokinetic SampleCollection: ddVeialamibvsielrtayyssoabinsgdenrecvidianttioicoaclnrsoosbwsse-efrrovesattpeierorinnfsgo,.rmbeoddyaswepirgehvtiso,usfleyeddecsocnrsibuemdptfioronFovagleuneesraatnidon iPnhaGrrmoaucposkiI naentdicIl,saremsppleectsivweelry,e oconllDeLct1e4d.frMoimlkesigahmtpalneds approximately two to six hours postdosage on DL 14. two Fo generation wers colleated rats vvsiehtsiscpewpreeadrwetaossathipeefriSfcpoeordnmseaodnr.d. aFgorllooswsinngeccroomppslyetoifotnheofthmoirlakcisTcah,mepalbsdeaomcmpoillnleeascltwiaoennr,defpeemiavilce creenctorridfedu.gedBalnododthseamspelreusTmhwweearnseuscmhoblielperpceatdenddtovditiahsettrhiSebpuoitnnifsoenorrioo.fr viAmepnlliaavnectraasvteiacotnainosnditaesndwas omfileka-csehcrdeatminglegfltasniddse fornolym)twheeraexiclloalrlye,cttheodraacnidc,shaibpdpoemdintaolthaendSpiongnusionarl the regions Ad. E1Co4lGleencetiroant:ion PupsAssignedtoPharmacokineticSample fpVoiesartbefirloiirntgmy.eadnOadnsclpDirnLeivc1ai4lo,uosbllisyteerdrvseasatcisrosinibsge,ndebfdoodrtoyF1twheegiegpnhhetarsramtaaincodonklpiiunteeprtsobassesrivganteidontso ewrearsee. were gach psuacpr,ifpiocoeldeadn(dpebrloiotedr)s,amcepnltersifwuegrede collected via the ic sample collontion inferior vena cava from and the serum was shipped o the. Sponsor. The to the Sponsor. liver from each pup was collected, pooled (per liter), andshipped 001296 418-014:PAGE I4 B. Results aAlnldFnoegcernoeprsaytioobnsefrevmaatlieonrsatwsesruervciovnesdidtoersecdheudnurleeladtseadcrtiofitchee. adAlmlinaidsvterrastieonclionfical PFOS. pBroedcyohwaebiigthattigoaniansndwegreestarteidouncepedriinodtsh,e a1s.6commg/pkagr/eddaytodtohseacgoentgrrolougproduurpivnagltuhees bMoadtyerwneailghbtodgyaiwnesifgohrttghaeiennstiwreerleacvtaartiiaobnlepetrhirooduwgehrouetgtehneerlaalcltyatcioonmppaerriaobdleand raemdouncgedthine tfhoeur1c.r6omsgs-/fkogs/tdearyeddsousbasgeetsg.roAubpsobleugtiennbiondgyawtetihgehtesndweorfethselightly precohabitation period and continuing through the gestation and lactation periods. Absolute and administered relative the 1.6 feed consumption mg/kg/day dosage values were generally reduced for rats of PFOS during the precohabitation, gestation and lactation periods, as compared to the control group values. gTrhoeupd.uraAtsioonbsoefrgveestdaitnioanptreenvdieoudstloy cbeonrdeudcutceeddsitnutdhye(A1.r6gumsg/Rkegs/edaarychdosage pLraebiomrpatloarniteast,ioInncl.o,ssP;rotthoecoalve4r1a8-g0e08n)u,mbtheirs orfediumcptliaonntawtaisonassistoecsipaetreddawimthwmaisnimal oslfidgehtlliyverreydupceerdp,urpes(uinltmiingnuitneas)sltiegnhtdleydrteodubceedreddeulcievedriend tlhitete1r .s6izmeg./kTgh/edadyuration dosage dosage group. group The live was also litter size before slightly reduced. cross-fostering The gestation in the 1.6 index was mg/kg/day 100% in each dosage group; all pregnant rats delivered live offspring. aPrutpiclmeo-rttraelaitteydwdaasmsgre(astuebsstetinC)p.upPsufprommortteasltitayrtwiacsle-atlrseoatiendcdraeamssedfoisntperuepds bfyrotmest rteesstulatrotfclteh-etsreeaitnedcrdeaasmess fionsptuerpedmorbtyalviethyi,clteh-etrvieaabtielditydainmdsic(essu(bnseutmbA)e.r oAfsliave npuupmsbeornsDoLf s4urpvrievciunlglpinugp/snupmebrelritotferpaunpdslcivreoslsit-tferosstiezreesdatDLw1e)igwheirneg rweedruecerde,duacnedd torneaDtLed4dparmescufloisntgerinedthbeysetesttwoarstiucblsee-ttsr.eatPedupdamomrsta(lsiutybsientpuD)pswafsrocmovmephaicrlaeb-le to that in pups (subset 8). from vehicle-treated These observations dams fostered indicate that in by vehicle-treated utero exposure to dams the test article is responsible maternal milk for did pup not mortality, adversely and that potential affect the viability exposure to of the pups. the test article via Pup body weights awnedreD)r.edauscceodmopnarDeLd 1toinpbuopthbosduybsweetisghotfsdianmtsheadsmuibnsiesttseroefddaPmFsOSad(msiunbissettesreCd mtihlekvaefhtiecrleDL(1subrseedtusceAdapnudpBb).odPyowteeintgihatls,exrpeogasrudrleestso tohfeinteusttearrotiecxlepovsiaurmeattoemthael 001297 418-014:PAGE 1-5 test article subsets D or vehicle. (pups from After DL 1, pup body vehicle-treated dams weights fostered were somewhat reduced in by test article-treated dams) and A (pups compared to from test the pups article-treated dams from vehicle-treated fostered by dams cross vehicle-treated dams), fostered to vehicle- as treated dams (subset B). After DL 1, pup in subset C (pups from test article-treated body weight reductions were greatest dams fostered by test article-treated dams). Tvewhoiclliet-tterrseaitnesdudbsaemts)Aa(npduposnferloimttetresitn asrutbiscelte-tCre(aptuedpsdafrmosm ctreostssa-rftoisctlee-rterdeattoed dpuapmsmocrrtaolsist-yfowsatsergerdewaittehstteisnttahretsicelet-wtoresautebdsedtasm.sT)hhearde pwuapssntohamtidlikdinnotthenurse; stomach of necropsied 57.1%, 100% in subsets A, and 87.0% of the pups C and D, respectively. that were found dead and mEivaclruoastcioopnyorfetvheealleivderisnocfrepauspesd tnhuatmbdeierdsoorfwpeerreoxsiascroimfeiscewditohninDhLep1abtyoceylteecstroofn Qpuuapnstiftraotmiodnaomfspetrreoaxtiesdomwietsh i1n.d6icmagt/ekdgs/ldiaghytlPyFmOoSr,e ctohmanpatrwiecdewtihteh ncounmtrboelrs.of peroxisomes in treated pups. were not discemible between Differences control and in cellular membranes or mitochondria treated pups in the samples examined. mEivaclruoastcioopnyorfetvheealleudngTsyopfeplul ppseuthmaotcdyietdesorcwoentraeinsiancrgifliacmeedlloanr bDoLdi1ewsiitnh beoltechtron control Type Il and 1.6 mg/kg/day pneumocytes and dosage lamellar group bodies pups, with an in lungs from increased number of treated pups. Although ssiognmiefilcaanmteldliafrfemraetnecreiawla(ssunrofatcetdanbte)twwaesenidtehneticfoinatbrloelwigtrhoiunpalavnedol1a.r6lmugmi/nkag,/dnaoy dosage group. 001298 Cc. Conclusion 418-014:PAGE 1-6 Trehdeuc1.e6d mfge/ekdgc/odnasyudmopstaiogne voaflPuFesOSforctahueseFdo rgeednuecrtaitoinosn idnabmosd,yawseiogcchutrgraeidn iannad previous multigeneration study with the test article (Argus Research Laboratories, Inc., Protocol 418-008). As previously observed, the s1t.6udmyga/lksgo/draeydudcoesdatghee odfurPaFtiOoSnsadofmigneissttaetrioend atondthpearFtourgiteinoenr,actaiuonseddams in this preimplantation loss and reductions in litter size and pup growth. Cross-fostering of pups from test article treated dams and vehicle treated dams revealed that the effects on pup mortality that occurred in thepreviously conducted study (418-008), can be attributed to in utero exposure to the test article, and that potential exposure to the test article via maternal milk did not adversely affect the viability of the pups. Exposure to the test article via both in utero exposure and matemal milk caused reductions in pup growth. |J Mildred S. Christian, Ph.D., Fello, ATS Executive Director of Research 23 -T1)P Date Athlon Alan M. Hoberman, Ph.D., DABT Director of Research 4Jon 25 ayfhond G. York, PD, JorsT Associate Director of Reearch and Study Director rps 7 Date u-99 Date 001799 I. DESCRIPTION OF TEST PROCEDURES A. Conduct of Study: 418-014:PAGE II-1 A. Sponsor: 3M Corporate 55144-1000 Toxicology, 3M Center Building 220-2E-02, St. Paul, Minnesota A2. Testing Facility: Argus Research Laboratories, Pennsylvania 19044-1297 Inc., 905 Sheehy Drive, Building A, Horsham, A3. Study Number: 418-014 A4. Sponsor's Study Number: T-6295.13 AS. Purpose of the Study: The purpose of following PFOS this study treatment was to evaluate survival of of C:CDBR VAF/PIus F1 Fo generation generation pups female rats cdruorsisn-gfopsrteemraetdindgu,rignegsttahteiolnacatnatdiolnacpteartiioond.toF1difgfeenreerntaitaitoenbpeutpwseewnereeffects on Pups exposed maternal milk. tSoetlheectteesdtpahratircmlaecionkuitneertoicansdamppulpessewxeproesecdolltoecttheed test article from via Fo generation female rats and F1 generation pups. AS. Study Design: The requirements of as the basis of study the U.S. design. Food and Drug Administration (FDA) were used A7. Reaulatory Compliance: The study was conducted in regulations of the U.S. Food compliance with Good Laboratory and Drug Administration (FDA), Practice (GLP) the Japanese Ministry (EEC). of Health and Welfare (MHW) There were no deviations from and the GthLePEurergoupleatainonEsctohnaotmaifcfeCctoemdmtuhneity quality or integrity of the study. Quality Assurance Unit findings derived from the 001200 418-014:PAGE II-2 inspections provided to during the the Study conduct Director aonfdthtihsesTteusdtyiangreFadcoilciutmy eMnatneadgeamnednhta.ve been AB. Ownership of the Study: TheSponsor owns the study. tissues are the property of the All raw data, Sponsor. analyses, reports and preserved AS. Study Monitor: Marvin T. Case, D.V.M., Ph.D. A10. Alternate Study Monitor: Andrew M. Seacat, Ph.D. A1. Study Director: Raymond G. York, Ph.D., DABT (Associate Director of Research) A12. Technical Performance: MJaorhgnarF.etBaMm.etMtar,tiBn.S(.R(eDsieraecrtcohrAosfsLoacbioartaet)ory Operations) Corrie L. Pabst (Quality Control Associate) A.13. Report Preparation: JRoayAmnonnFdraGz.eeY,orMk.,SP.h(.SD.t,udDyACBooTrdinator) Susan Karen K. G. PBarrakdesrh,aAw.,A.B.(SR.ep(oDrattaAdMmainniasgtreamteorn)t Specialist) A.14. Report Review: MAilladnreMd. SH.oCbherrismtaina,n,PPhh.D..D,.DFAelBlTow,(DAirTeScto(rExoefcRuetsievearDcihr)ector of Research) A.15. Date Protocol Signed: 23 October 1998 001301 A.16. Dates of Technical Performance: 418-014:PAGE 11-3 Rat Arrival Dosage Period (42 days before cohabitation 270CT 98 until DL*21) Cohabitation Period 02 NOV 98 - 28 JAN 99 13 DEC 98 PM - 19 DEC 98 AM Day 0 of Delivery Presumed Period Gestation (DG 0) 14 DEC 98 - 19 DEC 98 04 JAN 99 - 03 JAN 99 DL DL 1 4 Culling Culling (Control Pool) (Cross-Fostered Litters) 04 JAN 9 - 09 JAN 99 08 JAN 99 - 11 JAN 99 DG 25 Sacrifice DL 14 Pharmacokinetic Group Sacrifice 08 JAN 99 - 12 JAN 99 17 JAN 99- 22 JAN 99 F1 Generation Pups Sacrifice (DL 21) Fo Generation Dams Sacrifice (DL 22) 25 JAN 99 - 28 JAN 99 26 JAN 99 - 29 JAN 99 A17. Records Maintained: The original report, raw vehicle components are data and retained reserve samples of the in the archives of Argus bulk test article and Research Laboratories, Inc. one Any year preserved tissues are retained in after the mailing of the draft final the archives of the Testing Facilty report, after which the Sponsor will for decide their discarded at final the disposition. All Testing Facility. unused prepared Unused bulk test test article formulations article was retumed to were. the Study Monitor. B. TestAticle Information: B.1. Description: PFOS (FC-95) - a light-colored powder B.2. Lot/Batch Number: 217 (Expiration date: May 2000) B.3. Date Received and Storage Conditions: The test article temperature. was received on 21 October 1998, and stored at room a. DLis used as an abbreviation for day of lactation or day postpartum. 001302 B.4. Special Handling Instructions: 418-014:PAGE 14 Standard respirator safety precautions (use of and safety goggles) were protective clothing, gloves, dust-mist taken when handling the bulk test article and prepared formulations. B.5. Analysis of Activity: Information article is on regarding the identity, file with the Sponsor. strength, composition and purity of the test C. Vehicle Information: CAA. Description: 0.5% (RO. Tween 80 in Reverse Deionized Water) Osmosis Membrane Processed Deionized Water C.2. LNoutmbers: Tween 80 - MO3HOS, L06662 and M2947 C.3. Dates Received and Storage Conditions: The Tween 1 September 81099w8a(slotreLcOe8i6v6ed2)onan3dD1e7ceSempbteerm1be9r81(9l9ot8M(Olo3tHMO2S9)4,77), from RJ..TO.. Bdaekieorn,izPehdilwiapstbeurrgis, aNvaeiwlaJbelresfery,omaandcownatsinsutooursedsoautrrcoeoamt ttehmepTeersattiunrge.FacTihliety and is maintained at room temperature. C.4. Special Handling Instructions: rSetsapnidraatrodr,ssaaffeettyypgroegcgaulteisoonrs s(aufseetyofglpraostseecstiavnedclaotfhaicneg-,shgileolvde)s,wedruset-tmaiksetn when handling the vehicle. C5. Analysis of Purity: Neither the Sponsor nor contaminants likely to be the Study Director was present in the vehicle aware of any potential that would interfere with the results of this study. 001203 D. Test Article Preparation and Storage Conditions: Suspensions were prepared daily at the Testing Facility at concentrations of 0 and 0.32 mg/mL. Records are maintained in the raw data to document how the test article formulations were prepared. Prepared suspensions were stored at room temperature. D.1. Sample Information: ri =mem Tr oT[ oefee e J=eT eTo]] Er pe ay Lot LoseE2 smi | 280CT98 hd ethos 25NOV 98. D.2. Analytical Results: Data verifying the stability of the test article in the vehicle for 48 hours under the conditions of administration are on file with the Sponsor. Homogeneity of the prepared formulations is on file with the Sponsor. Results of the concentration analyses have not been received. E Test System: EA. Species: Rat E.2. Strain: Cri:CDBR VAF/Plus (Sprague-Dawley) 001304 E3. Supplier (Source): 418-014:PAGE Il-6 Charles River Laboratories, Inc., Portgage, Michigan Ed. Sex: Female (Male rats of the and were not considered same source and strain were part of the Test System). used only as breeders E.5. Rationale for Test System: TSyhsetCerm:bCeOcauBsRe:VA1F)/tPhlisusstra(iSnporfagrautew-Daaswulseye)d rianttwhaesresperloedcutcetdivaesatnhde Test TdeesvteilnogpmFeacnitlailtyt"o;xicaintyds3t)udtihees;te2s)t ahritsitcolreiciaslpdhaatramaacnadloegxipcearlileynaccetievxiesitnatthtehe species and strain. E6. TestSystem Data: Number of Rats Approximate Date of Birth Approximate Age at Arrival Weight () on the Day after Arrival Weight (g) at Study Assignment 86 23 AUG 98 66 days 179-226, 200 - 244 E7. Breeder Male Rat Data: Number of Rats Approximate Date of Birth Approximate Age at Arrival WWeeiigghhtt ((gg)) oatnCtohheabDiatyatAifotner Arrival 110 13 JAN 98 77 days 317-352 514-866 E.8. Method of Randomization: Ubapsoins oafrrcivoaml,pumtaelre-gaennderfaetmeadlerarantdsowmeruneitas.ssiAgftneerdatcocliinmdaitviioduna,l7h8ouvsirignign foenmathlee wraetisghwtesreresceolredcetdeddufrorinsgtuadcycloimnattihoen.basTihseoffepmhyasliecarlataspwpeeraeraanscseigannedd body to dosage groups (42 generated (iwnetihgehtc-oonrtdreorlegdr)oruapndaonmdiz3a6tiinonthperotcreedautreedsg.roup) based on computer- Within each cohabitation dosage group, consecutive with breeder male rats, one order male was used to assign rat per female rat. female rats Rats were to assigned to cross-fostering or pharmacokinetic collection depending on the order 0061305 418-014:PAGE II-7 of delivery the control and the availability of rats for cross-fostering. Eight female rats from group and two female rats from the treated group were designated for pharmacokinetic sample collection following delivery; these rats were not cross- fostered. Twenty-five litters per dosage group were cross-fostered on DL 1. The litters were assigned such that four subsets were obtained among the two dosage groups, as described in the table below. Subset assignments were made based on order of deliveries. esac|WNiostar| Doge | onrcea ReToe | cane GT roup n Rats TT Tot(meg/kngicdaey)|| owes| Re6 assigned1| (DatmoTrmeeatsment|| NAumb]er | Uwewss[5 oo |1 w6e T 15 |owT teesmsss ||5co] On DL 4, cross-fostered litters were culled to five male and five female pups per litter, where possible, using a table of random units. E.9. System of Ident ition: E.9.a. Fo Generation Rats: Rats were permanently identified using Monel self-piercing ear tag (Gey Band and Tag Co., identification Inc., No. numbers uMpSoPnTas2s0i1g01n)m.entMatloethreatTsewsetrineggFiavceinlituyn'isqbureeepdeerrmamnaelnetrat population. Female rats were assigned temporary numbers at receipt and given unique permanent identification numbers when assigned to the study. Cage tags were marked with the study number, permanent rat number, sex, test article identification and dosage level. E.9.b. F1 Generation Pups: Pups were not individually identified during lactation; all parameters were evaluated in terms of the litter. toms 1 and 2 a. See APPENDIX D (DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY), b. See APPENDIX D, item 3. 001306 Ff Husbandry: 418-014:PAGE II-8 F.1. Research Facility Registration: eUtSsDeqA. Registration No. 23-R-099 under the Animal Welfare Act, 7 U.S.C. 2131 F.2. Study Rooms: The study rooms were maintained under conditions of positive airflow relative to a hallway and independently supplied with a minimum of ten changes per hour of 100% fresh air that had been passed through 99.97% HEPA filters. Room temperature and humidity were monitored constantly throughout the study. Room temperature was targeted at 64F to 79F (18C to 26C); room humidity was targeted at 30% to 70%. See APPENDIX E (TEMPERATURE AND RELATIVE HUMIDITY REPORTS). F.3. Housing: Fo generation rats were individually housed in stainless steel, wire-bottomed ccoahgaebsi,taetxicoenptpedruiordi,ngetahcehcpoahiarboiftartaitosnwaansdhpoousstepdaritnutmhepemraioldes.ratD'surciangge.the Beginning no later than DG 20, Fo generation female rats were individually housed in nesting boxes, except during the collection interval for urine andfecal samples. During this collection interval (DLs 21 to 22), the female rats were housed individually in metabolism cages. Each dam and assigned litter were housed in a common nesting box during the postpartum period. All cage sizes and housing conditions were in compliance with the Guidefor the Care and Use of Laboratory Animals. F.4. Lighting: An automatically-controlled fluorescent light cycle was maintained at 12-hours light:12-hours dark, with each dark period beginning at 1900 hours EST. F.5. Sanitization: Cage pan liners were changed approximately three times each week. Cages were changed approximately every other week. Bedding was changed as often as necessary to keep the rats dry and clean. F.6. Feed: Rats were given ad libitum access to Certified Rodent Diet #5002 (PMI Nutrition Intemational, St. Louis, Missouri) in individual feeders. 001307 418-014:PAGE I1-9 F.7. Eeed Analysis: Alenvaellysseexsceweedrienrgoutthienemlayxpiemrfuomrmceodncbenyttrhaetifoenedlismiutpsplfioerrc.erNtioficedonfteaemdionrants at deviations analyses. fCroopmieesxpoefcttheedrensuutrlittsioonfatlhreefqeueidreamneanltyssweseraeredeatveaicltaebdlebyintthheeseraw data. cNoeintthaemrintahentSsploinkseloyrtonohratvheebSeteundyprDeisreencttoirnwtahes faeweadrethaotf waonuylpdotheanvtiealinterfered with the results of this study. F8. Water: Lmoecmablrwaanteer(tRh.aOt.hwaadtebre)ewnapsraovcaeislsaebldebtyo pthaessraatgseatdhriobuigthumafrreovmerasneaoustmoomsaitisc watering access system and/or individual the processed water as a bacteriostat. water bottles. Chlorine was added to F.9. Water Analysis: The processed water is analyzed twice contamination (Lancaster Laboratories, annually for possible chemical Lancaster, Pennsylvania) and monthly fPoernnpsoyslsivbalneiab)a.cteCroipailecsonotfatmhienartesiuolnts(AonfaltyhteicwaalteLrabaonraaltoyrsieess,arIenca.,vaCihlaalbfloentin, the raw data. cNoeintthaemrintahentSsploinkesloyrtonohratvheebSeteundyprDeisreencttoirnwtahes waawtaerretohfatawnoyuplodtehnativael interfered with the results of this study. F.10. Bedding: Bed 0'cobs was Groups, Maumee, used as Ohio), the nesting material (Andersons' Industrial Products F.11. Bedding Analysis: cNoeintthaemrintahentSsploinkesloyrtonohravtheebSeteundyprDeisreencttoirnwtahse baewdadriengoftahantywpooutlendtihaalve interfered with the results of this study. Analyses for conducted annually and documented in the raw data. possible contamination are 001308 G. Methods: G.1. Dosage Administration: 418-014:PAGE I-10 Tee [ rom T e--me [TE] E=ET CrToTmmTo]een] =Elem e [[E =rE rErrEE eS=EeerEeE TeerErE ESREEEERsssa EEEEaREnES ENEE E l E Ey G.2. Rationale for Dosage Selection: Dosages were selected on the basis ofa previous two-generation study conducted with the test article (Argus Research Laboratories, Inc., Protocol 418-008). In this study the Fo generation matemal and paternal no- ohbisgehrevrabdloes-aegffeesctc-aluevseeld(rNeOduEcLt)ioonfsPiFn ObSodwyawsei0g.h1tmgga/iknga/ndadyr(e0d.u4cmegd/fkege/dday and consumption values). The Fo generation reproductive NOEL was greater than 3.2 mg/kg/day; no effects growth on in mating, fertility or the F1 generation estrous cycling occurred. The NOEL for viability offspring was 0.4 mg/kg/day (1.6 mg/kg/day and and higher dosages caused preimplantation loss and reductions in litter size, pup viability, growth and survival). 001309 418-014:PAGE 11-11 The (0.4 mF'g1/gkegn/edraaytidoonsamagteercnaaulseadndrepdautcetmiaonlsNiOn EboLdoyfwPeiFgOhSt was gain 0.1 and mg/kg/day reduced feed dcoonssaugmepotfio0n.4vmalgu/eksg)/.daTyh;enoF1efgfeencetrsaotniomnarteipnrgodourcfteirtvielitNyOocEcLurwraeds.grTehaeteNrOtEhaLn a (fo0r.4vimagbi/lkitgyladnayd dgorsowatgheicnatuhseeFd2stgielnlbeirrathtsioanndofrfesdpurcitngiownassin0.li1ttemrgs/ikzge,/dpauyp viability, growth and survival). G3. Routeof Administration: Oral (gavage) G.4. Rationale for Route of Administration: aTdhmeinoirsatlra(tgiaovnagine)threoudteevewlaospmseenltecatledanfdorruesperobdeucctaiuvsee:tox1i)cothliosgywastsudtihees;roauntde of 2) it is one of the possible routes of human exposure. G.5. Frequency of Administration: G.5.a. Fo Generation Female Rats: Female 42 days rats were given the test article or the prior to cohabitation through DL 21. vehicle once daily beginning Dosages were adjusted daily for body weight changes and given at approximately the same time each day. G.5.b. F1 Generation Pups: pFo1ssgiebnleyraetxipoonspeudptso wtehreetensottadrtiircelcetldyurgiinvgenmathteertneaslt agretsitclaet,iobnut(imnauytehraoveexpboeseunre) or via matemal milk during the lactation period. G6. Lengthof Study: . Approximately 3 months G.7. Method of Study Performance: G.7.a. Cohabitation and Cross-Fostering Procedures: Twenty-five cohabitation rats for assigned to the rats assigned to ctohnetrtorleagtreodugprwoeupreancdohtahbeitceodhaobniteatdiaoyn prior to period consisted of a maximum of six days*. The cohabitation period for the remaining a. See APPENDIX D, item 4. 001310 418-014:PAGE I-12 rmaatsxiinmtuhme ocfonftirvoeldagyrso.upDaunrdintghecorhaatbsitaastsiiognn,ebdottoh tchoenttrroelaatenddgPrFoOuSp-twraesataed groups were co-housed with untreated male breeders, one male rat per one female rat. Female rats with spermatozoa observed in a smear of the vaginal contents and/or a copulatory plug observed DG 0 and assigned to individual housing. in situ were considered to be at Several dams assigned to the control group that delivered on the first day of pnaeretudreidtiofonroccrcousrsr-efodswteerriengalpluorwpeodsetso.deTlhiivserparnodvirdeetdaian ptoheoilronfatcuornatlropludpasmusntil available for immediate cross-fostering of pups from PFOS-treated preventing these pups from nursing from their PFOS-treated dams. dams, thus. Srteamrotivnegdwiitmhmetdhieadteelliyve(raisessoofonthaesdpaamrtsurointiotnhewasseccoonmdpldeayt,eda)llfpruopmsthweeirre rgersopuepctoirvae dPFaOmSs-tarnedatpeldacdeadmw.ithWheietnhera adnaomthearssdiagmneadstsoigthneedcotnotrtohlegcroonturpolwas not available, then a dam from the control pool was used. The litter from that control pool dam was then sacrificed via decapitation. This cross-fostering procedure resulted in four groups of 12 or 13 dams/pups, i.e., Control/PFOS (Subset A), Control/Control (Subset B), PFOS/PFOS (Subset C) and PFOS/Control (Subset D). This procedure allowed distinctions to be made between PFOS treatment. prenatal and postnatal effects of the continuous matemal G.7.b. Fo Generation Rats Assigned to Cross-Fostering: The Fo generation female rats were observed for viability at least of the study and for general appearance at least once during the twice each acclimation day period. The rats were test article, abortions, also examined for clinical observations of premature deliveries and/or deaths before effects and of the `approximately one hour postdosage. wBeoidgyhtwseiwgehrtes rweecroerdreedcodradileyddautrilneagstthoendcoesdaugreinpgertihoedaaccnldimaattsiaocnripfeicrei.od.FeBeoddy consumption values were recorded `weekly to cohabitation, daily during at least once during the the presumed gestation acclimation period and period, on DLs 1, 4e,x7p,ec1t0edantdhat14t.heFpeuepdscboengsaunmpttoicoonnwsausmenomtattaebumlaaltefdeeadf.ter DL 14, when it was The female rats were continually evaluated (24 hours per day) for clinical observations during parturition [performed under red light conditions during the (daDrGk 0petroiothde(ttiimmee etahechfirpsut ppuwpaswaosbsdeerlivveedrewd)a,slreencgothrdoefd)pa,rtduuirtaitoinon(otifmgeeosftation 001311 418-014:PAGE Il-13 observation N-1 pups in of last pup each litter), mliitnteurssitzheest(idmeefionfeodbasesravlaltpiuopnsofdetlhievfeirrestd)puapnddipvuipded by veixaabimliintyataitobnirotfh.thPeulpitsterthfaotr eviiatbhielritaypwpeeraereedxasmtiilnlebodmfoorr vtihtaatl dsiteadtubseaftorbiertihn.itiTalhe lungs were considered removed stillbom; paunpdsiwmimtherlsuendgsinthwaattefrl.oatPedupwserweitcholnusnigdsertehadtlsiavnekbowmeraend to whhaveen dtiheedpsuhporstlwyerafeteerxbaimrtihn.edMadtuerminagl tbheeha2v1i-odrayofptohsetpdaarmtsumwpaesrieovda.luOabtseedrdvaeidly meaxtpeercntaeld bmeahtaevrinoarlwbaesharveicoorrwdeerdeornecDoLrsde1d,,4,if 7a,nd14wahnedn2p1r.esDenetv,iaotnioanlsl oftrhoemr days during the postpartum period. cFoollgeecntieornatoifounrifneemaalnedrfaetcsalwesraemphloeussefdroimndDiLvisdu2a1lltyoi2n2m. etFaoblolloiwisnmgctahgee2s4f-ohrour cicoel,lescttoiroendinftreorzvealn, (s-a7m0pCleosr wbeerloewc)oallnedctsehdiipnpteodcteonttrhiefuSgpeotnusboesr,foprlaancaeldysoins.dry mOanteDmLa2l2,rabtlsovoida sthaempilnfeesri(orapvpernoaxciamvaatealn4yd mtrLanesafcehr)rewdeirnetocsolelreucmtesdefpraormattohre treufbreisgearfatteerdrceemnotrviaflugfe.romTmheetasbeorluimsmwacsagtinrga.nsfTerhreedsianmtpolpeoslywperroepyslpeunne tinubaes lcoalbleelcetdiownitthimtehpoeinstt.udyAlnlusmabmeprl,esratweidreentiifmimceadtiioant,edlaytferoofzecnololnectdiroyn,icse,tusdtyordeady and frozen (-70C or below) and shipped to the Sponsorforanalysis. G.7.c. F1 Generation Pups Assigned to Cross-Fostering: Day first 1 of lactation day on which (postpartum) was all pups in a litter defined as the day of birth were individually weighed. and was also the pEearciohd.littDerewaadspeuvpasluoabtseedrvfeordvaitabtilhietsyeattilmeeasstwtewriecereeamcohveddayfroofmthteheponsesttpianrgtubmox. T(ihneclpuduipnsg pgrreossesnetxitnerenaaclhphliytsteircwaleraenocmoaulniteesd) ownecree eraeccohrddaeyd.foPrhtyhseicpaulpssigonnsce each DLs 1 day during (birth), 4, the postpartum period. Pup 7, 14 and 21 and at sacrifice. body weights were recorded on On DL female 4p,ucprsospse-rfolisttteer,rewdheF1regepnoessriabtlieo.n litters were culled to five male and five 001312 418-014:PAGE Il-14 G.7.d. Fo Generation Rats Assigned to Pharmacokinetic Sample Collection: dViealbiivleirtyy oabnsderclviantiicaolnsobwseerrveatpieornfso,rmbeodd,yawseipgrhetvsi,oufseleyddecsocnrsiubmepdtifoornFvoalgueenseraantdion dams assigned to cross-fostering. Blood, rats in milk and Groups | liver and samples were II, respectively, cdoelsleicgtneadtefdrofmoreipghhatrmaancdoktiwnoeFtiocgseanmerpalteion `caolplpercotxiiomnatpeelrydtowsoatgoesgirxohuopurosn DL 14. Milk samples postdosage on DL 14. were collected Each dam was removed from dam the was niensjteicntgedbionxtraanvdenionudsilviyduwailtlhy housed one unit foofroaxpyptroocxiinm(a2t0elUySfPouurnhiotusr/sm.L, The Lot aNpupmrboexirma5t2e0l6y4,fievexpmiirnatuitoensdbaetfeorJeulym,il2k0s0a0,mpmlaensuf(aatctleuarsetd1b0y0 OLsboprern)sample) were collected. The (-70C or below) and samples shipped were immediately to the Sponsor. frozen on dry ice, stored frozen Following collected milk from sample collection, blood the matemal rats via the isnafmerpiloersve(napapcroaxviamaatnedlytr4anmsfLereraecdhi)ntwoere Tsehreusmesreupmarwaatsortrtaunbsefse.rrTehdeinstoamppolleysprwoepyrleenseputnubiensalraebferliegderwaittehd tcheentsrtiufudgye. number, rat All samples wideernetifiimcmateidoina,tdealtyeoffroczoellneoctnidorny, study day ice, stored and collection frozen (-70C timepoint. or below) and shipped to the Sponsor. aFnoldlotwhienmgilcko-llseecctrieotninogfgmlilakndasndfrbolmootdhesaaxmiplllaersy,, taholriavceircs,ecatbidoonm(irniaghlt alantderianlgluoibnea)l regions of each dam (left side only) below) and shipped to the Sponsor. were collected, stored frozen (70C or G.7.e. 1 Generation Collection: Pups Assigned to Pharmacokinetic Sample pViearbfiloirtmyeadndasclpirneivcailouosblsyerdveastciroinbse,dbfoodr yF1wegiegnhetraantodnlipttuepr sobassesrivganteidontso wcerrosesfostering. bCeafposreaannddlaafbteelredrettuebnteisonweorfepowoeliegdhepdup(csoammbpilneesdfwoerisguhbt,setqoutehnetnuesaereisnt 0.001 g) pharmacokinetic analyses. DBlLoo1d4,spaomoplleeds(wpeerrelitctoerl)leacntdedtrvainastfheerriendfeirnitoor sveernuamcasvepaarfartoomretaucbehs.pupThoen samples were polypropylene spun in a refrigerated tubes labeled with the csetnutrdiyfunguem.beTrh,epsuepriudmenwtiafsicattriaonns,fedrarteed into of 001313 418-014:PAGE Il-15 collection, frozen on study day and dry ice, stored fcroolzleenct(i-on70tiCmeoproibnetl.owA)llasnadmpshliepspweedrteo immediately the Sponsor for analysis. The liver from each below) and shipped pup was collected, to the Sponsor for pooled (per analysis. liter), stored frozen (-70C or H. Gross Necropsy': gAlrlosFso ngeecnerroaptsiyoonfrtahtes wtheorreacsiacc,raifbidcoemdibnyalcaarnbdonpedlivoixcivdiescaesrpahywxiaastipoenr,foarnmdeda. eGvraolsusatiloens.ionRsewpreerseenrteattaiinveed pihnonteougtrraalphbsufoffermeadte1m0a%l fgorromsaslilnefsoironpsosasrieblaevafiultaubrlee in the raw data. HA. Fo Generation Rats: iTmhpelafnetmaatlioenrsaittsewsewreeresarcercifoircdeedd.onADlLiv2e2r.seTcthieonnu(rmibghetrlaatnedraldilsotbrei)buftrioonm oefach dam was collected, for analysis. stored frozen (-70C or below) and shipped to the Sponsor pFhoallromwaicnogkicnoemtpilcetcioollnecotfimoinlwkesraempsalcericfoiclelde.ctiBonlooond DL 14, female rats and liver samples assigned were to sciotlelsecwtaesd arsecporredveido.usly described. The number and distribution of implantation lReastisontsh.atUdtiedrinwoterdeelsitvaeirnaedliwttietrhwe1r0e%saacmrmifoinceiduomn sDulGfid2e5tao ncdonefxiarmmitnheedabfosregnrcoess of implantation sites. mDiasmsisngwiotrh pnroessuurmvievdincganpnuipbsalwiezerde.sAacrbilfoioceddsaafmteprlethe(alpapsrtopxuimpawtealsyf4oumnLd)dweaasd, collected, separator prior to sacrifice, via tubes. The samples the inferior vena cava and were spun in a refrigerated transferred centrifuge. into serum The serum iwdaenstitfriacnatsifoenr,reddatientoofacoplolleyctpiroonp,ylsetnuedytdubaey laanbdelceodllweictthiosnttuidmyepnouimnbt.er,Threatsamples were immediately frozen on dry ice, and stored frozen (-70C or below) and a. Atable of random units was group female rat and one F1 used to select one generation control Fo generation control group male and female tpoupprforvoidmewchoincthroallltiissssuueessfeorxpaomtiennteidalatconmepcarroaptsiyvewehriestoreptaatihnoeldo,giicnaolrder evaluations. 001314 418-014:PAGE II-16 dshaimppweadstocotlhleecStpedo,nsstoorrfeodrfarnoazleynsi(s-.70ACliovrerbseelcotwi)onan(rdigshthilpatpeerdaltlootbhee) Sfproomnseoarch for analysis. H2. F1 Generation Pups: Pups that died before vital status at birth, as epxraemviionuastliyondeosfcrtihbeeldi.tterPufoprspwuipthvigabriolsitsy were evaluated lesions were for preserved preserved in in Bouin's Bouin's solution solution, for possible future and the liver was evaluation. preserved in The lungs were neutral buffered 10% formalin for possible future evaluation. pPuuppss cfurlolmeddoanmsDLas4siwgenreedstaocrtihfeicceodntvrioaldpeocoapliatantdiocnu.lleTdheonluDnLgs1 aanndd tahlleoltihveerr lwiettrere) aconldletchteedfifrsrtotmenthpeufpisrstttheatn wceulrleedfopuunpdsdferaodm t(hneo caounttorloylsipso)oflr(oimrreeascpehctive of ldiovserasgweegrreorueptaoinneDdLin1.neTuthreallubnugfsfewreerde1r0e%tafionremdalininBofuorinp'ossssioblluetifount,uarend the evaluation. evaluation. Remaining culled pups were sacrificed and discarded without Pups found lesions and fdoeratdheorcasuacsreifoifceddeabtehcaoursteheofmomroriibbuunnddictonydwiteiroen. examined for For all pups gross. found dead on DLs 2 was preserved to in 4, the lungs were preserved in neutral buffered 10% formalin, Bouin's Solution, and the liver for possible future evaluation. Pups with gross lesions found for possible future evaluation. on DLs For all 2 to pups 4 were found preserved in dead on DLs Bouin's 5 to 21, solution the lungs, liver and possible gross future lesions were evaluation, preserved in neutral buffered 10% formalin for rOenspDecLti1v4,e dliattmerss wasesriegsnaecdritfoictehde bpyhacramrabcoonkidnieotxiicdesaasmpphlyexicaotliloenc.tioInndaivniddutahle,ir pooled samples described. The (per pups litter) of blood and liver were collected were examined for gross lesions. as previously lOensioDnLs.21G,raollsspulpessiwonesrewesraecrirfeitcaeidnevdiaindenceauptirtaaltbiuonffaenredd e1x0am%ifnoerdmalfionr. grSoisxs litters bpleorosdubasnedtlwiveerr.e Brlaonoddomslaympselleesctweedreforcoclollelcetcetdiovniaoftpheooilnefedrisoarmvpelneasc(apvear lfirttoemr) of seaamcphlpeusp,weproeolsepdun(peinralitrteerf)riagnedrattreadncsefnetrrriefdugien.to Tsheerusmerseupmawraatsortrtaunbsefs.erreTdheto cpoolllyepctrioopny,lsetnuedtyudbaesy alanbdelceodllweictthiotnhteimsetpuodiyntn.umAblelrs,amraptleidsenwteifriecatiimomne,ddiaatteeloyf frozen on analysis. dry ice, stored frozen (-70C or below) and shipped to the Sponsor for 001315 418-014:PAGE Il-17 All pups selected for continued examinedforgross lesions. observation were sacrificed via decapitation and lL Statistical Analyses: Averages and appropriate. percentages were calculated. Litter values were used where 001316 Wm. RESULTS 418-014:PAGE Ill-1 A. Mortality, Clinical and and 2; Individual Data Necropsy Observations - Tables 19 and 20 (Summaries -Table1s All rats survived to scheduled sacrifice. Allalctaadtvioenrspeerciloidniscawleorbesecrovnastiidoenrsedduurnirneglattheedptroectohheabtietsattairotni,clegebsetcataiuosneatnhde sienceindeinncreastswienrtehenoltabdoorsaatgorey-deenvpiernodnemnetnta.ndTohrestheeoobbsseerrvvaattiioonnssianrcelucdoemdmonly cmihsrsoimnogr,hibnroorktehneaa,nds/coarlymitasial,liagbnreadsiionnciosonrst,helohceaaldiz,edneaclko,petaciilaanodn/tohrefohreealdi,mb, ldiemhbysd,ranteicokn,anpder/ionreablachke,michar,obmroodkaecnryoorrsrwhoelal,enscfaobreopnawthdeighiet aadndorptraoilt,ruding xiphoid. gTahleacotnolcyelnee)cirnoposnyeocbosnetrrovlatgiroonuwpadsaamnaisnsgiuginnaeldmtao spsha(rpmraceoskuimneedtitco bseamaple collection on lactation day 14 (DL 14) B. Body Weights individual Data (Figures 1 -- Tables and 2; Summaries 21 through 23] - Tables 3 through 8; Btootdhyewceoingtrhotlggarionuspduvrailnuegst,hebepgriencnoihnagbiotnatdiaoynspe1r5iotdo 2w2eroef rtheeduscteuddya(sDcSosmp15ared wtoei2g)hatnodncDonStsin3u6intgo t4h3r;otuhgeh bDoSdsy 2w9eitgoht36l.osRsawtassingrbeoatthedroisn atghee g1.r6oumpgs/klgos/tday gdeonseargalelgyroocupc.urrAeldthoonudghaycsal4c2ultaot4ed3 foofr tahesesvteund-ydaanydiwnteerrveala,stshoeciwaetiegdhtwiltohsses. tmhoevaenmiemnalt rofootmhewmhaelne tbhreeerdatesrwreatrseicntoohatbhietaendiomnaldrayoo4m2.anAdsmaatriensugltaoctfivtihtyesien cwhearnegreesd,ubcoeddyawnedigahbtsoglauitnesbfoordythweeiengthitrse wpererceohslaibgihttaltyiroendpuecreidodon(DDSS143to(9463).6% of control) in the 1.6 mg/kg/day dosage group. dMoastaegmealgrbooudpyiwnetihgehtfirgsatitnwsocwoenteiknsueodf ttohebegesrteadtuicoendpienritohde {1d.a6ymsg0/ktgo/1d4ayof gestation, DGs entire gestation 0 to 14). As period (DGs a 0 result, to 20) body weight gains were reduced for in the 1.6 mg/kg/day dosage group. the Matemal body weights throughout gestation, tended to be reduced in the 1.6 mg/kg/day dosage group `Raeddmiunciesdtemreadte1m.6almgb/okdgy/dwaeyigPhFtsOSperdsuirsitnegdtihnetlhaecttawtoiosnupbesreitosd of rats (subsets C and D), 001317 418-014:PAGE Ill-2 2asndcoB)m.paLraectdattiootnhebordatyswienitghhetssuhbasdettsheademxipneicstteedreddetghreeeveohficvalreia(bsiulbitsyetbsutAwere csuobmspeatrsabCleanbdetDwe(teenstsaurbtsiceltes-tAreaatnedd Bgr(ovuephsi)c.leMcaotntermoallgrbooudpys)weaingdhtbgeatiwnesenfor the entire lactation subsets. period (DLs 1 to 22) were essentially comparable among the four C. Absolute (g/day) and (Summaries - Tables Relative 9 through (a/ka/day) Feed 14; Individual Consumption Data - Tables Values 24 through26) Arbesdoulcuetdef(ogr/rdaatys)aadnmdinriesltaetriveed(tgh/ekg1/.d6aym)g/fkeged/dcaoyndsoumspatgieonofvPalFuOeSs wdeurrienggetnheerally precohabitation, values. gestation and lactation periods, as compared to the control group D. Natural and 16; DIenldiivveirduyaalnDdatLaitt- eTraObblesser2v7attihornosug(hSu3m1)maries - Tables 15 D.1. Natural Delivery Observations `The duration of gestation tended group, both when the duration of to be reduced gestation was in the 1.6 calculated mg/kg/day dosage in whole days and to tahsesonceiaarteesdt wtietnhthmionfiamadlayp.reAimsplparnetvaitoiuosnlyloosbss;etrhveeda,vetrhiasgerenduucmtbioenr woaf s idemlpilvaenrteadtilointtesritseizsep.erThdeamduwraatsiosnliogfhtdleylirveedruycepde,r rpeuspul(tiinngmiinnuatessl)ighttelnydreeddutocebde froesdtuerciendg iinntthhee 11..66 mmgg//kkgg//ddaayyddoossaaggeeggrrooupu.p wTahseallisvoe lsiltitgerhtsliyzreebdeufcoerde, carnoss- observation related to total litter size and associated with stilbirths. The gestation live offspring. index was 100% in each dosage group; all pregnant rats delivered D.2. Cross-FosteredLitter Observations After cross-fostering subsets. on DL 1, live litter sizes were comparable among the four fPousptemroerdtablyittyeswtaasrtgircleea-ttersetatiendsduabmsse)t;C1(9p.u2p%s offrtohmetepsutpasrtwiecrlee-tfroeuantedd ddeaamdsor presumed cannibalized on DLs 2 to 4. subset A (pups from test article-treated Pup mortality was dams fostered by also increased vehicle-treated in dams) on As DLs 2 to 4; 9.0% a result of these of these pups were found increases in pup mortality, dead or presumed cannibalized. the viability indices (number of 001318 418-014:PAGE 111-3 lrievdeupcuepd,s aonndDnLu4mbperrescuolflsiurnvgi/vninugmbpoeufrpspuppesr lcirtoesrsa-fnodstleivreedlitotner DsiLz1e)s waetrweeighing were reduced on DL 4 preculling (pups from vehicle-treated dams in these fostered two subsets. Pup mortality in subset by test article-treated dams) was D vceohmipcaler-atbrleeatteod tdhaamtsi)n.suTbhseesteBo(bpsueprsvaftrioomnsveihnidcilcea-tteretahattedinduatmesrofeoxstpeorseudrebyto the taretsitclaertviicalemiastreemsapolnsmiilbkledifdornpotupadmvoertrasleiltyy,afafnedctthtahte potential exposure to viability of the pups. the test aPdumpinbiosdtyerweedigPhFtfOiSte(rswuebsreetsreAduacneddCo)n, aDsLc1ominpabortehdstuobspeutpsboofdpyuwpesigfhrtosm idnatmhse subsets of pups from Potential exposure to dams administered the vehicle (subsets B the test article via matemal milk after DL1 and D). reduced pup DboLdy1,wpeuipghbtso,dyrewgeairgdhltesssweofreinsuotmeerowheaxptorseudruecteodtihnestuebssteatrsticDle(oprupveshifcrloem. After varethiiccllee--ttrreeaatteeddddaammssffoosstteerreeddbbyyvteehsitclaer-ttircelae-tterdeadtaemdsd),amass)caonmdpAar(epduptos tfhreomputpesst fDrLom1,vpehuipclbeo-dtryeawteeidghdtamresdufcotsitoenrsedwebryeveghriecaltee-sttreinatseudbdseatmsC (subset B). (pups from After test article-treated dams fostered by test article-treated dams). SpuepxsraotnioDsLan4dptohsetelualcitnagt)iowneirnedecxom(pnaurmabbelreofamivoengputphse ofnourDLcr2os1s/-nfuomstbeerredof live subsets E. OClbisneircavlatOibosnesrv(aStuimomnasrfireosm-BTiarbthletso 1D7ayan2d1 1P8o;sItnpdairvtiduumalanDdatNae-cropsy Tables 32 and 33 tTrweoatelidttderasmisn )suabnsdetonAe(lpitutpersinfrsoumbsteestt Car(tipculpes-tfreraotmedtedsat marstifcolset-etrreeadtebdydvaehmiscle- wfoasstegrreedatbeystteisnt tahrteiscelet-wtoreastuebdsedtasm.s)Thheardepwuapss ntohatmidlikdinnotthenusrtseo;mapcuhpomfor5t7a.l1it%y, 100% Cand and 87.0% of the D, respectively. pups that were found dead and necropsied in subsets A, Aclolnostihdeerrecdlinuincralelaantdednteoctrhoepstyesotbasretricvlaetiboencsauisnet:he1F)1thgeenienrcaitdieonncepsuwpesrweenroet odbosseargvea-tdieopnesnidnecnltu;deodr b2l)atchketiopbosfertvaial,tiloanboorcecdurbrreedatihninogn,lydaornke ilnitctoerl.or,Clminiiscsailng hindpaw pelvis at digit and necropsy bent tail. One on DL 21. pup in subset A had slight dilation of the renal 001319 418-014:PAGE lll4 F. Electron Microscopic Evaluation ofLiver and Lung from Day 1 Postpartum Pups (APPENDIX G FA. Liver Uhtetpraatstorcuyctteursalolfy,putphesrferwoemrdeaimnscrteraesaetdednwuimtbhe1r.s6 omfg/pkegr/odxaiysoPmFeOsSw,ithcionmpared `nwuitmhbceorntoroflpse.roQxuainstoimteastiionntorfeapteerdopxuipsso.meDsififndeirceantceeds silnigchetlllyulmaorrmeemtbharnantweisceorthe `mitochondria were samples examined. not discemible between control and treated pups in the F2. Lung Tanydpe1.Il6pmnge/ukmgo/cdyatyedsocsoangteaignrionugplapmueplsl;arhobwoedvieers,wethreereidaepntpiefiaerdedintbootbhe caonntrol tirnecarteeadsepdupnsu.mbAletrhoofugThypseomIIepnlaemuemlolcarytmeatserainadl (lsaumreflalcatrabnto)diweassinidleuntnigfsiafbrloem within group alveolar and 1.6 lumina, no mg/kg/day significant difference dosage group. was noted between the control 001320 REFERENCES 418-014:PAGE IIl-5 1. U.S. Food and Harmonisation; DGruuigdeAldimnieniosntrdaettieocnti(o1n99o4f).toxiIcnitteymtaotiroenparlodCuocntfieonrefnocr e on medicinal products. No. 183. Federal Register, September 22, 1994, Vol. 59, 2. RUe.gSu.lFatoioodnsa;ndFinDarluRgulAed.min2i1stCraFtRionP.artGo56o.d Laboratory Practice 3. JPraapcatniceeseStMainndisatrrdyfoofrHSeaafletthyaSntdudWieelsfoanreDr(1u9g9s7,).MHGWooOdrLdaibnoarnacteory Number21, March 26, 1997. 4, E1u9r8o9poenanthEecaocncoempitcanCcoembmyuntihetyEu(r19o8p9e)a.nCEocuonnciolmidcecCisoimomnuonnit2y8 oJfulayn lOaEboCrDatodreyciprsaicotni/cer.ecOofmimceinadlaJtoiuornnaolnocfotmhpelEiaunrcoepewaitnhCporminmcuinpilteisoefs:good Legislation. 32 (No. L 315; 28 October): 1-17. 5. Christian, M.S. and Mutagenicity Tests. VEonyvtierko,nPm.eEn.ta(1l98P2r)o.tecItnioVnivAogeRnecpyr,odWuacsthiivnegatnodn, D.C. National Technical Information Service, Springfield, VA 22161 U.S. Department of Commerce, 6. nCharlitsrteixaonn,eM.(SP.ro(c1e9e8d4i).ngsReopfrNoadlutcrteixvoenetoSxiycmiptyoasinudmt,erNateowloYgoyrekvaAlcuaatdieomnsy of of Sciences, November 7, 1983), J. Clin. Psychiat. 45(3):7-10. 7. CLoanntgr,olP.DLa.t(a19i8n8)t.he EChmabrrlyeos aRinvderFeCtra:lCDDeveBlRopRmaetn.taClhaTroxliecsitRyiv(eTerratology) ALrabgoursatRoersieesa,rcInhc.L,aWbiorlamtionrgiteosn,, IMncA.) 01887-0630. (Data base provided by 8. UInssetitoufteLaobfoLraabtoorraytAonriymaAlnsi.malNaRteisonoaulrcAecsad(1e9m9y6).PreGsusi,dWeafsohritnhgetoCna,reD.aCn.d 9. Salewski, E. (1964). Implantationsstellen Farbemethode zum am Uterus der Rate. makroskopischen Nachweis von Arch. Pathol. Exp. Pharmakol. 247:367. 001321 APPENDIX A REPORT FIGURES . 001322 MATERNAL BODY WEIGHTS Figur1e | i jh. e" ! _Let -- ; o MTTTTUITETETTR acs hov yu a! 3 &2 z 3 DAYOF STUDY DAYOF GESTATION > MATERNAL BODY WEIGHTS RATS ASSIGNED TO CROSS-FOSTERING - Figur2e [EE 7 || a. nl --- = nd g: m 1 - ~~pd _ oo o re a _. Zao ~ lw" rg Brees - | TS . sto 1 ' 1Bor nmoli . 28 REEEEEREEEEC.. e--r~a-s amen =o svmmeron I sunseron 3 APPENDIX B REPORT TABLES 001325 nn esse soos ea wns enn warn, goa fe se os wt ve ommmncnonmer "a oe o ala he 11 Gy aber"or rte assigned from Group 1 5 5 28 R3 8 2 a an rosie comes eo rn [r-- wo we reas erin vs oe skiimn rosdibLe CHORE+ (DATS wars)eanorWsEisvencheue. 28 8 3 - g 3 &3 5 Shem, cobb oe EN vars ue ue ve p---- ue we ue EE NE ER SR Er Se Movi ins 20 sr 2 ora aoSaeOr ksAi A A1RTatsbresncemonrion 5 1VE6MIwCaL/eaT/dSuy)dosJargoeuh9Srohuep dcarmnsstcoronsts.erfeodstweirtehd wwietht1s.6antiotm/uga/dary odosssigeergerronup litters. g 288 Ry we . oe :2 : 3 g 28 52 8 T a Ey -- i :8 2 5 g8 z 8 3g g EE g2 8 es x oo & 38 2  i3 2 SELE E E S n SRt So TEpL at sgeee acnee 2 : & a 2 / + 7 ISTE S i Son e i ipsiseFoe ocee g8&8 222Z 2E& 3: TABLE 10 (PGE 1): RELATIVE FEED CONSINPTION VALUES (G/W/ONT) - PRECOHNITATION - SUMSUAAY cr omvenaTion roo RTS g 23ii{ 8g :z g ii: 3 ta 2 tr, tt 1 en os SA no ama oe : g3o 832 8 i2 OE ariesSousOtoe See toesEometes wih 1.8SYVBIdOosNe grep Liters. Beit aesfof dee Sohhad Stars ch TeremtSrorefomenee bE RT EE IE art et, ss et ti 8 52 & . 3 a Seine Garon mtu ecu aes Wi 2Re mines o vena. BE Voicn ie Souinht$IaSIeT a Son fots eres winsH eviehaarrr Slimeeem"are istre. BE I enn EE 11 phSELES SITET ser day 10 of backanion, materna ee commepeion sain i 2g82g &Z?32 A i BD BR een - EEREREa emer sno0 me ton + < 8 3 :g TUE wm mm FE mn - . & 8 ? a  ET Le EE EEe h en : g 82a i3 Z Bio OE WH mE ma ma ven lin he on ie FRE EE tn co art cn ge se 5bE ISIE CR TL erTnbee rs e . 1.6 ma/ks/day dosage group damscross:fostered vith 1.4 wa/ha/day dosage group litters. 2 g 3 3 8 7 a28 8 Em b---- ne mbt 4 3 +Sn E BnoE s E tn fgEa iro oeoe g 3 8: 3 8 i women mn waime enamn wei atl re Jr E Fi o g OA J g cn ERELIT 5 BB 8B VIL3.CG6 ImmaaD//kkgSgidOdbNayyId)dosB aoge ar gSi rrhoouve ppCd? dreaea omssncrre oomsnesse..rffoosntcearereehdd S with 1.e 6 ma/kadb /edtaytedi powosese grouMpctereli.tters. vith VERLCA control aver Titers > ~oEo or I a eee, | 3 EEyah[r= om BCrEanny UE ILaka frEe gerowaiver B RATTAE N Sn en of SS ws pce cs g 5 8 2 @ Hw. ETT p------ cbte obhe de ge TT on rewnc on xvor uivringwim onsen a rm onil on uull $uvl u2v FBEES S I SE IS A LEA BT EE hare 2 2 3 & 8 32 2 uo To a i i a FE HE Ses Tie eo te BE s i 25 g | | Bogen HES gH gen REESE. saan OA 0 vstPips : FEISTY re] apists " ST os 2i i IEEE 8 or Sr apinsises eV POWER. resrisany 8g lon contienadat necropey. hipaa sonatas g torts aris 2 8 g o8 33& 1&a] LE e b FE O E IE LLY HI S re R we 38 a8 zt g g i g g  BS 3 g 8  5 i ii i 2 2 2 3 8 2 8 g 3g 3: 2 FFmaa. g:: 3:f BU a FE + ATER g s8 3# g8 8E HE EF ce i 5g 3ii a 8 | momER BEBE ESL RL LR BS esa th < 8 3 - or i : g I EE mnt. i s i : z g3 z: t | 2 ii east hamo essen cher ony on dekd tun wan presen geass on doy 35 of gestion gg i i 2 ETT EREI es sa en 1 tr 1 ion g 8< :32 3& g g 58g4 ii Hane 2 5 i >& i hy 4: :g: g 3 & g 3 '2 8g 3 B | i EEERELEEED 3 8 Ta eh A. es 32ob MLLE I n R e & ~~ - -- we 8 2 ' 2 * | treo oe ; A BI EE Sl Rly ewe sree ens | 3 LJ | ]| g EPA St i penss ee 8 3 : i: | | | i -- ----LIL 2 < g | ] . BESET 5 g 83 & -* CERETEEES 3o E 25a? " D pr ho a SaAn o ) o g3 g ia ; & | ERRE E ve g .52 23i& ~~ i. :: | : 32 2 32 2 FR g <. Ez 8 3S | 22 2 2& 8 ~ EE g R3N 82 i | | ; | me i Sy 3 & i i i - oe ; - * : ny - Bernese fred noowE won moon ER B RT nnpn as XCLBED Pho AVERAGES) en n - or - a :3 l :ig ~ | g 8 R & 3 | BOE EER Iparr ni o " _ & TB ro ge rt resr 1 Le 8g er enoloto ae tron Fem sr enlie eI -g &3 3e LTO oS Z ob IODA gR RRR do sronfe I Aes sreee en : : 2 | se g 38288 2#3:3 & | | ER 3 & done re ener St re ow ee. | a ri on i on FE Bip BE mihi vse BEB BRELL LLL S . LICUerwarenot cross: fostered;velues axchuded from group sverspen. TIT g 22g 3E3 3 jg i] ua LiE ioo 8 s as eh ised between tiv of che cee 2nd ne pp, J oy pope. BE aanSe hm Soe SHEN Se detegoes Hrsers: EPRI I SRR Sm pretest sue averages. :3 |i 2 3 | jit i i i Rg TEER Cu Caleuiated 0 tha arent tenth of 4 dr Fo dE or B0b3 TI E REIaE dt he si tt fnTh pai 0 2 u beeen ron fonared, partially cibtseervsede . xif grea average. - 3ia 3 H REe 1 BTd R se sis n ot ce e fe a-- kp, s-- tot 101 ron SE i ' s 3 G8 ] Sore Cotecrisi - oo + miei veaieay errs i ------ Ee 3 oHLL LE mn AE 2 od HE HEN BL a aaa BE a &EEL FE s Wie pais x 1 some i oF Fab Welsms oF Sa a EL 3 | Eoand ani RnERE EERE 8 2 ud eios Le ator cans0 Ww ove rans eneSyver as AT B 8B E5 hTEeSii te ensn co tonne vn 1 nat cnn tg ce 3 LI STS I NI rySeiad ee dite. -~ - 3 o4 = g : j ~~ BoRBIGEMEHEHEERCT- : 3b Se A SO ee o3 . Vehicle control group dams crofostered vith vehicle control 1iccers. BodHGHEEOaEnEnaEnal, i 3 UimEEEEaay LE &  CEE EIEEE EE 8 mmm i i EO iniasy sow Soe dan omaripa ELT 1Sr sects pt 2 LRSE Tn en Sa pa Cd z 8 ATR ERR 5 $2000 0 hn ph An el Xr - B Simm E MM LrC E EER T _-- 2 2 Boone HELD SHS Ra RAR dH z Q o BEw T i A Isonoa i cC ont, wT e ssaotconse 83 EFE HEg ISEfe p me E ErA 8 gE onninnbsdiniionna., ; RE ER teescross{Ioven ith 1.423/3 Sori Sroup. Ete 5 8 REEENEESLn i Bn ryEen eeeoh, AA SoyBp er 2 By TSII I EA BOAR ARERR REAR 3 ViRE Euaa 8 BIEVER RI co nn moms comin BEN Fh SedEaTors TG oar i em reat . &  hmm ga bn MHI ILI IRSA SL B REE OR EE NEHEOEH E BRE g: [J a p---- i 3BIETEEEeE CEST ee BR eek TOLLDMING "SECOND LETTER" IWOICATES THE DAY FOSTFAKTUN THE EVENT OCCURRED. % I : :& 8 8 I ---- + once rare em on g 8 33 IS oS 8 Cs g L8 @& J vr nit. EC LCA A o idl, EE W tl ow ie g 3 ii BB FL EE r hie entedE fec2 otoupaverse g HAI EAH HTTAIR IE R a g E Ee Iue co me &BbFi LEE;EnIR 3: g 3 & 2 2 zg K Is 3 Temeotekseeup TERE suescron EE rn es ET aLvs pnv sa Js 2 23 8 32 a =3~ g 2% 8g 8 4 i SE 28 23 | ~~ i ee & 3 7 ne . vos co. S Q E _i-- mReR i Emwieentmesn J SG 3 a: a g 3 2 & 8 8 a - +; TpiHERES i iipe Er : 2 ne & i 3 8 a < ] g 32 28 B + APPENDIX C PROTOCOL AND AMENDMENT 001434 418-014:PAGE C-1 PRIMEDICA AroseBRToegesae nohayLSLaAaeponreTStrein.Sdennessg em PROTOCOL 418-014 STUDY TITLE: PURPOSE: SPONSOR'S STUDY NUMBER: T-6295.13 Oral (Gavage) Cross-Fostering Study of PFOS in Rats "The purposeofthis study is to evaluate pup survival of F1 rats following PFOS treatment of Cr:CDBR VAF/Plus female rats during premating, gestation and lactation. F1 pups will be cross-fostered during the lactation period to differentiate via maternal milk the differences for in utero exposure and exposure through matemal milk.Selected pharmacokinetic samples will be collectedfrom Fo generation females and F1 generation pups. TESTING FACILITY: STUDY DIRECTOR: Argus Research Laboratories, Inc. 905 Sheehy Drive, Building A Horsham, Pennsylvania 19044-1297 Telephone: (215) 443-8710 Telefax: (215) 443-8587 Raymond G. York, Ph.D., DABT Associate DiroefcRet seo arr ch SPONSOR: 3M Corporate Toxicology 3M Center, Building 220-2E-02 St. Paul, Minnesota 55144-1000 STUDY MONITOR: TMearlveipnhTo.neCase(,51D).V7.3M.5,-3P1h8.D0. Telefax: (651) 733-1773 ALTERNATE `STUDY MONITOR: Andrew M. Seacat, Ph.D. Telephone: (651) 575-3161 Telefax: (851) 733-1773 001436 418-014:PAGE C2 Protocal 41P8a.g0e142 REGULATORY CITATIONS: U.S. Food Guideline oannddeDtercutgioAndmoifntiosxtircaittyiotno (r1e9p9r4o)d.uctIinotnemfaotrimoendailcCionnalfeprreodnuccetso.n HFaerdmeornailsation; Register, September 22, 1994, Vol. 59, No. 183. U.S. Food and Drug Administration. Good Laboratory Practice Regulations; Final Rule. 21 CFR Part 58. Japanese Tor Safety Ministry Studies of on Health Drugs, aMnHd WWelOfradriena(n1c99e7N).uGmboeord2L1a,boMraartcohry26P,ra1c9t97i.ce Standard EaImcuecrneodppatetaainoncneEocbnoynctoohmmeiplcEiuCaronocmpemeuawnintihEtcpyori(nn1oc9mi8pi9l)ce.sCooCfomugmnouconidiltldyaeobcoifrsaaitnoonrOoyEnprC2aDc8tiJdcueelc.yis1Oif9of8ric9/iraoelncJotomh-uenal of the European Communities: Legislation. 32 (No. L 315; 28 October): 1-17. REGULATORY COMPLIANCI This study will be conducted in compliance with the Good Laboratory Practice (GLP) regulations cited above. All changes Director and otrhreeSvipsoinosnosr,ofdtahtisedpraontodcomlaisnhtalalinbeed documented, signed with the protocol. by the Study Tanhde wQiulalliintsypAesctsucrriatinccalepUhnaiste(sQAofU)thweilsltauuddyitinthaeccporortdoacnolc,etwhiethratwhedSattaanadnadrdthOeperreapotritn,g Procedures of Argus Research Laboratories, Inc. TachceurfaitnaellyrerpeofrltecwtislltihnecrluadwedaatsataotbetmaeinntedsidgunreidngbtyhtehpeeSrtfuodrymaDnicreecotfortthheatsttuhdeyraenpodrtthat aslilgnaipfpilciacnatbdleeviGaLtPionrsegfurloatmiGonLsPwerergeulfaotliloonwsedocicnutrh,eecaocnhduwicltl of be the study. described Should in detail, together with how the deviation might affect the quality or integrity of the study. SCHEMATIC OF STUDY DESIGN AND STUDY SCHEDULE: See ATTACHMENT 1 to the protocol. 00143 418-014:PAGE C-3 Protocol 418.014 Page 3 R AND LE: Identification: TestAric. Name: Physical Description: Lot/Batch Number: Specific Gravity: Purity: Expiration Date: PFOS (Synonym: FC-95). Light-colored powder. 217. ~0.6. 98.9%. May 2000. Information on the identity, composition, strength and purity of the test article is onfile `with the Sponsor. Vehicte: the raw data. 0.5% Tween 80 in Reverse Osmosis Membrane Processed Deionized Water (R.O. Deionized Water). Supplier and lot identification of Tween 80 to be documented in * conducted. Neither the Sponsor nor the Study Director is aware of any potential contaminants likely to be present in the vehicle components that would interfere with the results of this study. Therefore, no analyses other than those mentioned in this protocol will be: `Safety Precautions: Gloves, mask, appropriate eye protection and a uniform/lab coat are to be wom during formulation preparation and administration. The Material Safety Data Sheet (MSDS) is. attached to the protocol (ATTACHMENT 2). Storage: Bulk Test Article: Vehicle Components: Prepared Vehicle: Prepared Formulations: `Room temperature. `Room temperature. `Room temperature. Room temperature. 001438 418-014PAGE C4 Protocol 41P8a.g01e4.+ JAlullitaenstGaurltbiicnlseksih,iMpmaenntasgteoorfthFeoTremsutliantgioFnasc,ilattythsehopruedviboeusaldydcrietsedseadddtroetshse aanttdention of telephone number. c`aSrhtiopnmsensthsouslhdoubled include labeled aipnpfroorpmraitaitoenlyc.onTcehrenirnecgipsiteonrtasgheocuolnddibteionnostiafinedd shipping in advance of shipment. FORMULATION: Freqouf Preepnaractioyn: tFhoermsutlabaitliitoynsof(tshuespteesntsiaortnisc)lewiilnltbhee pvreheipcalreefdorda4i8lyhaotutrhseuTnedsetrintgheFaccointdyi.tioDnastoafverifying administration are on file with the Sponsor. Detailed preparation procedures are attached to this protocol (ATTACHMENT 3). Adjustment for Purity: The test article will be considered 100% pure for the purposeof dosage calculations. Testing Facility Reserve Samples: TthheecSopuorsnesoorf wil this reserve study. a sample (1 The Testing g) of each Faciity will lot of the reserve bulk test a sample a(r5timcLle)uosfeedacduhrilontgof the vehicle components used during the course of this study. Samples will be stored under the previously cited conditions. ANALYSES: `Samples additional to those described below may be taken if deemed necessary during the course of the study. Bulk Test Article Sampling: No analysesofthe Information on the bulk test article will be stability of the bulk test caortnidculecties dondufrilienwgitthhethceouSrpsoensoofrt.his study. Analyses of Prepared Formulations: HHoowmeovgeern,eirteycoarndds wsitlalbbiletymaoifnptraeipnaerdetdofdoromcuulmaetinotnshoiswotnhefilteeswtitahrttichlee Sponsor. formulations wer, prepared. 0143 418-014:PAGE C-5 Protocal 41P8a.g0e1s4 `Concentration ofTest Article Formulations: Concentration of study. Duplicate tshaempplreepsa(r2edmLforemauclha)tiwoilnlsbweiltl abkeenvefrirfoimedthdeurfiirnsgt the and course of this last preparation orenmtahienidnagysparmepplaeresdw.illObneerseatmaipnleed oaftetahechTseesttiwnigllFbaceilsthyipapsedbafcorkuapnaslyasmipsl;etsh.e Backup `samples will be stored frozen (-70C or below) and discarded at the Testing Facility upon request of the Sponsor. `ShippingInstructions: Samples to be analyzed will be shipped (frozen on dry ice) to: Kris 3M J. Hansen, Ph.D. Environmental Technology and Safety Services. 9Bu3i5ldBiungsh2-A3v6e-n0u8e St. Paul, Minnesota 55133-3331 Telephone: (612) 778-6018 Telefax: (612) 778-6176 Therecipientwill be notified in advance of `sample shipment. DISPOSITION: Prepared formulations will be discarded at the Testing Facility. All remaining bulk test article will be retumed to the Study Monitor at the previously cited address upon completion of all work with the test article. TEST SYSTEM: `Species/Strain and Reason for Selection: The Cr:CDBR VAF/Plus (Sprague-Dawley) rat was selected as the Test System because: studies; 2) 1) this strain of historical data rat was used in and experience tehxiestreaptrtohduecTteisvteiangndFadcielvietyloTMp"m; eanntdal3t)otxihceittyest article is pharmacologically active in the species and strain. 001430 418-014:PAGE C-6 [---- Page 6 Number: IPnoiptiuallaptoipounlasteiloenctaecdclfiomrastteudd:y: 528n64dvmia2rtg4ienmdftfehememaaleleaetraretasdt.sg(r3o0upi)n.the control group Population selected for pharmacokinetic sample collection (day 14 postpartum): 12 mated female rats (six per dosage group). BWode y iganhdAt ge: Female rats will which time they be will ordered to have be expected to body be at weights least 60 dofay2s0o0fgagteo .22A5ctguealacbhodayt receipt, weights at will be recorded the day after receipt and will be documented in the raw data. The weight range will be included in the final report. Sex: Female be used rats will only as bberegeidveernstahnedteasrtearntoitclce.onsMiadleererdatospafrtthofetsheamTeesstoSuyrscteema.nd strain will Soures: Charles River Laboratories, Inc. The rats will be shipped in filtered cartons by air freight andor truck from Charles River Laboratories, Inc., to the Testing Facility. Identification: Fo Generation: a Rats are peeormaMnePnTtly20i1d0en1t)i.fieMdaulsainrgatsMoanreelgivseenlfu-npiieqruceinpgeeramrantaegnst(dGeentyfiBcaantdioannd Tag nBumbaersrupson aasssdiagnnemdettneotmptohreaTreystniunmgbFearcsiltayt'escbariepetdaenrdmagliveenratunpoipquuleatpieonr.manent identification numbers when assigned to the study. F1 Generation: Pups will not be individually identified during lactation; all parameters will beevaluated in terms of the litter. co1448 . 418-014:PAGE C-7 Price 41r8.0e14 ANIMAL HUSBANDRY: All cage sizes and housing conditions are in compliance with the Guide for the Care and Use of Laboratory Animals'. Housing: Fo Generation Rats/F1 Generation Litters: Fo generation rats wil be individually housed in stainless steel, wire-bottomed cages, except during the cohabitation and postpartum periods. During cohabitation, each pair of rats will Beat duis calocton intervals or urine and fecal samples. During these presumed be housed in gestation, Fo the male rat's cage. Beginning no later than day 20 generation female rats will be individually housed in of nesting postpartum period. cEoalclehctdiaonmiantnedrvaalsss,igthneedfelmitatelrewirlaltsbewilhlobuesehdouinseadcionmdimvoidnuanlleystiinnmgebtoaxbodluirsinmgcatghees. Nesting Material Bedding material (bed-'cobs) will be provided. Bedding wfiolrbpeocshsanigecodntaasmoifntaetnioans nareececsosnadruyctteodkeaennputahlelyaaninmdaldsocruymeanntdecdleiannt.he raw data. Room Alr, Temperature and Humidity: "The animal room is independently supplied with at least ten changes per hour of 100% fresh air that has been Room temperature will passed through 99.97% HEPA filters (Airo Clean room). be maintained at 64F (18C) to 79F (26C) and monitored constantly. Room humidity will also be `monitored constantly and maintained at 30% to 70%. Light: An automatically controlled 12-hour light: 12-hour dark fluorescent light cycle will be maintained. Each dark period will begin at 1900 hoursEST. Diet: Rats will be given Certified Rodent Diet #5002 (PMI Nutrition Intemational) available ad libitum from individual feeders. 00144) 418014:PAGE C8 Protocol 41P8a.g0e1s4 Water: `atWhnartaoeuurtgohwimlaaltrbieecveawrvaastieelroaibsnlmgeoaascidcsleimsbseitmsubymsrftarenomem. ibAneldfliovwriaedtuueaslrebw.ioltltClhbeiesofrairtnotemacwaihlelldobcteaolatsdhodeuercdcaegteoasnthdoerpfarsosmed {phraonce1s.s2epdpwmatcehrloarsinae baatcttehreiotsitmaeto;fparnoacleyssisse.d wWaatteerriisseaxnpaelcytzeeddtmoocnotnhtlayinfonropomsosribele bacterial contamination and twice annually for possible chemical contamination. Contaminants: N{we0oiutblhedeprirtnehtseeernfSetpreionnwtishtoehrtcnehroetrirfteihseeudlSdtisteuto,dfytthhDieisrdesrctitunodkryi.nigsTawhweaarrefetooorrefet,harennyonpaeonsttaeilnnytgisaemlsactooetrnhiteaarlmtiahtnaalnnetvteshloslsitkehelayt routinely performed by the feed supplier or those mentioned in this protocol will be conducted. RANDOMIZATION, COHABITATION AND CROSS-FOSTERING PROCEDURES: cUopmopnutaerirv-agle,nmeraalteeadnrdafnedmoamleunriattss. wiAlfltebreaacscsliimganteidont,o7i8ndviivrigdiunalfehmoaulsienrgatosnwitlhebebasis of dScouenrltierncogtleadgcrcfoloirumpsattaiunoddny.3o6nThitnheetfhbeeamstairlseeaotrfeadptshgywrsioilulcpabl)eabaapsspseeidagrnoaenndccteoomadpnoudstaebrgo-edgygernwoeeuripagsthet(ds42(rewicenoitrghdheetd ordered) randomization procedures and treated with either the vehicle or the test article for42 days prior to cohabitation. Wctoihhteahcbioinnttearatoiclohngrdwooiustphagwbierllegebrdoeeurcpo,mhacalobenistreeacdtus,otinoveneedoamryadplerreiowrrialtltpobeecrofuhesamebaidtlateotiaroasnts.ifogTrnhrefaetrsmaaatlssesaisrgasntisegdtnoetdo ttohe trartesatweidthgrsopuepr.maTthoezocaohoabbsiteartvieodn ipneraiosdmewiallrcoofntshiestvoagfainamlacxoinmteunmtsofafnidv/eordaaysc.opuFleamtaolrey plug observed in situwill be considered to be at day 0of presumed gestation and assigned to individual housing. fMoaltloewds:feomfatlheer4at2sf(etmhaolsee wriattsh acsosnifginremeddtoevtihdeecnocnetoroflmagrtoiunpg,)3w0illmbaeteadssfiegmnaeldearsats will basesaisgsniegdnteodtthoetthreeactoendtrgorlogurpo,u2p4fomractreodssf-e{omsatleerirnagtspwuirllpobseesa;ssoifgtnheed3t6o ftheematlreeartaetds gdroosuapgefogrrcoruospsw-iflosbteerdiengsipgunraptoesdesf.orApnharadmdaictoikoinnaletsiicx msaamtpeldefecomlalleectriaotns(farsodmeescarcihbed breelmoawi)nifnogllfoewmianglederlaitvserwyi,thbuctontfhiersmeerdatesviwidlelnncoetobfemcartoisnsg-ftohsatterweed.re Faisnsaillgyn,eadntyo the control group prior to cohabitation will be assigned to the control pool for cross-fostering 001443 418-014:PAGE C-9 esos Page 9 purposes. Any remaining female rats with confirmed evidence of mating that were waistshioguntedfutrothtehreetvraelautaetdiognroatuptphreidoirstcorectoihoanboiftatthieon`SwtiuldlybDeirseacctroirfiacnedd atnhde dSitsucdayrMdoenditor. Day 1 of lactation (postpartum) is. defined as the day of birth and is also the first day on wafhtiecrhalalllpuppuspsininaaliltittetreraraeredeilnidivveirdeudalalnydwegirgohoemded(pbuyp body weights the dam). will berecorded Tpwhaerlmvaecdokaimnsetaincdstahmepirlerecsoplelcetcitvioenl(itstiexrsdwailmlsbeansdelleitctteersd paenrddaosssaiggenegdrotuop)t.heThese mr |ee r [ox] pUopsttpoa2r4tulmit.terTshpeerlidttoesrsawgiell gbreouapsswiilglnebde srueacshstihgantefdofuorrscurbossest-fsoasrteeroibngtaoinneddayam1ong the two dosage groups as described in the table below. L Group| FemaleRas pda) | CT Tovewa| To Toman] gred |_(Dam Traatme = [| me [1] 5 |ww 1o} Detailed procedures for reassignment of litters for cross-fostering are attached to this protocol (ATTACHMENT 4). On day 1 postparturn, pups from dams assigned to the control pool will be culled, sacrificed via decapitation and handled as described in ATTACHMENT 4. On day 4 postpartum, cross-fostered litters. will be possible. Pups not selected culled to five male and five female for continued evaluation will be pups per litter, where itation; the lungs (saved in `Bouin's solution) and the liver (saved in sacrificed via decap neutral buffered 10% formalin) will be collected from the first ten culled pups from each dosage grofuorpp(oismseisbpleectfiuvteuroefhliitstteor)padtehtoelrogmiicnaeldetvoalbueataitond.a4Ryempaoisntipnagrtcuumllaenddpups will bpreessaecrrviefdiced via decapitation and discarded without necropsy evaluation. 001443 418-014:PAGE C-10 Protoca 4P1a8ge.01%0 ADMINISTRATION: RR outee andasfoo r Chon ice: aTdhmeinoirsatlr(agtiaovnagien )threoudteevwelaospmseelnetcatledanfdorruesperobduecctaiuvsee:to1x)itchoilosgwy asstudtihees;roaunted of 2) itis one of the possible routes of human exposure. Meat ndFhreqo uend cy: FoGenerationFemale Rats: pFreimoraltoe croahtasbwiitllatbieongitvhernoutghhedtaesyt2a1rtipcolsetorptarhteumv.ehDicolseaognecsewdilalilbyebaedgjiunsntiendgd4a2ildyafyors body weight changes and given at approximately the same time each day. 1 Generation Pups: Rationale for Dosage Selection: eFx1pgoesneedrattoitohneptuepsts awritlilcnleotdubreindgiremcattlyemgiavlegnetshteattieosnt (airntiucltee,robuetxmpoasyurbee) poorsvsiiabmlaytemal `milk during the lactation period. Dosages were selected on the basis of a previous study conducted with the test article. (Argus Research Laboratories, Inc., Protocol 418-008). Dosage Levels, Concentrations and Volumes: wz | mp p[Sym|pCy pt [= A] apssuen | e etc cred 00% rss r asse e rwiloefdc otnooesclos htosson ripacHto rcdo 14 ESTS, ANALYSES AND MEASUREMENTS -Fo GENERATION: Viability: All Periods: At least twice daily. 001444 418-014:PAGE C-11 Protocel P41a8o01141 ClO inicb al serv and/a orGt enei ralAo ppen arans ce: Acclimation Period: At least once. Dosage Period: Twice daily. Prior to administration and once `approximately one hour:postdosage. Matemal Behavior: Days 1, 4, 7, 14 and 21 postpartum.Observed abnormal behavior will be recorded daily. Clinical observations may be recorded more frequently than cited above, if deemed appropriate by the Study Director. and/or Study Monitor. Body Weights: Acclimation Period: Dosage Period: At least once. Daily. Sacrifice: "Terminal weight. Feed Consumption Values (recorded and tabulated): Acclimation Period: At least once. Dosage Period: Weekly to cohabitation. Daily duringpresumed gestation and days 1, 4,7, 10 and 14postpartum. Feed consumption will not be tabulated after day 14 postpartum, when it is expected that pups will begin to consume matemal feed. Feed consumption values may be recorded more frequently if it is necessary to replenish the feed. These intervals will not be tabulated. MatingPerformance: Mating will be evaluated daily during the cohabitation period and confirmed by observation of spermatozoa in a smear of the vaginal contents and/or a copulatory plug observed in situ. 00144% . 418-014:PAGE C-12 Protocol 4P1a8g.e01142 Natural Delivery: Female rats will be evaluated for: Clinical Observations During Parturition [to be performed under red light conditions during the dark period (time each pup is observed will be recorded)] Dobusreartvieodn)o.f Gestation (day 0 of presumed gestation to the time the first pup is Length of Parturition (time of observation of last pup minus the time of observationofthe first pup divided by N-1 pups in each litter). Litter Size (defined as all pups delivered). Pup Viability at Birth. Pharmacokinetic Sample Collection: Caps and labeled tubes will be weighed (combined weight, to the nearest .001 gram) before and after retention of pooled pup samples for subsequent use in pharmacokinetic analyses. These weights will be documented in the raw data, and copies of these weights will be included with the packing list priotro shipment. Blood, Milk and Liver Samples- Fo Generation Rats: ofsample caliectons (blood and milk) wil be recorded in the raw data. Blood, milk and liver samples will be collected from six matemal rats designated for pharmacokinetic sample collection per dosage group on day 14 postpartum. The time Milk samples will be collected approximately two to six hours postdosage on day 14 pfoorstappaprrtouxmi.matEealcyhfoduarmhowuilrls.beTrheemdovaemdwiflrlobmethienjneectsetdinigntbroaxveannodusilnydiwviitdhuaolnlye huoniutsoefd oxytocin approximately five minutes before milk samples (at least 100 pL per sample) are collected. The samples will be immediately frozen on dry ice and `maintained frozen (-70C or below) until shipment to the Sponsor for analysis. Following milk sample collection, blood samples (approximately 4 mL each) will be collected from separator tube the matemal rats s. The samples w via il the inferior be spun in vena a refr cava igerat and transferred in ed centrifuge. The to serum serum will be transferred into polypropylene tubes labeled with the study number, animal identification, date of collection, study day and collection timepoint. All samples will be immediately frozen on dry ice and maintained frozen (-70C or below) until shipment to the Sponsor for analysis. 00144 418-014:PAGE C-13 Protocol 4P1a8ge01143 FTolllsoewcirnegticnolgleogcnltliayon)ndwosiflflmrbioelmkctaohnleldeacxbtilelodao,rdyf,rsotahzmoeprnlaecasinc,d,aastbiodvreoermdis(ne-act7li0oannCd(oirignhbgteulilonaatwle)rarulentgliiolobsnehs)ioapfnmdeeantcthhe Gtoatmhe(lSoptosnisdoer for analysis. `Blood and Liver Samples - F1 Generation Pups: pBolsotopdasrtaumwmpi,llepbsoeowslilepldubn(epiecnroalllireteecfrtr)eidagnevridaattetrdhaencseifnnetfrrerriiefoudrgievn.etonTashecerausvmearsfuermpoamwrilealtaobcrehtpturubaepnsso.fnerTrdheaedyi1nf4o apomlpylperospylene tubes Pellastion, study day laanbdelceodllweictthiotnhteismetpuodiyntn.umAblelrs,amanpilmeaslwiildlenbteifiicmamteiodni,atdealtye forfozen (-70C or below) unti shipment to the Sponsor for on Gy ice and maintained frozen analysis. Tbehleowv)eruntfirlosmheiapmcehnptutpo will the Sbpeocnoslloerctfeodr,anpaoloylseids(.per ter), frozen and stored (70C or `Shipping Instructions: AISlnalamslpailmeepdslwewisitlhwibltelhebsehsiampmappienldteasoinannefddrsofzerenontzeotnno(dK-rr7yi0sicJCe. ovHriaabneoslveeonrw,n)iPughnh.ttiDl.m,saihalit.ptmhAeenpptarcfekovriinoagnuasllliyystsciwisit,ledbe address. Both the recipient and the Study Monitor will be notified in advance of sample shipment. rine and Fecal Sample Collection: oFfougfeinneeraantnditoefnrevcfaaellm,asslaaemmprplalteessswfiwrlilolbmebdeahycoosulsl2ee1dctteiondd2ii2nvtipodoucsaeltnlptyarriintfuummge.etatFbuobolellsio,swmpilncgaactgehedeso2fn4o-rdhrcoyoulirlceectainond Sotlolrcetdiofrnozen (-70C or below) until shipmentfor analysis. `Shipping Instructions: I`AlnSlaalmsupadlmeepdslwewisitlhwibltlehebsehsiampmappielndteasoinnaenfddrofszreeonnzteotnno(dK-rr7yi0sicJC.e oHvriaabneoslveeonrw,n)iPughnh.ttiDl.m,asihla.itptmAheenpptarcfekovriinoagunaslllsyytsciwisit.ledbe adress. Both the recipient and the Study Monitor will be tified in advance of sample shipment. METHOD OF SACRIFICE - Fo GENERATION RATS: Rats will be sacrificed by carbon dioxide asphyxiation. 001443 418-014 PAGE C-14 Protocol 4P1a8ge01144 NEC- FR oGEO NERP ATIOS NRAY TS: hvGarilacushastalileosniiso(nsasutewasiblleebxoeafmrrieantaneiddnoeamdt niunneicntersuotwpirslalylbwbeiulflufsbeeerderdetto1as0ien%leedfc,otriomnanoleridnceofrnotrtropopolrsgosrvioibudleep fcruoattnutfrrreoolm Yspsescuiefiscaflolryacniytepdobsesliobwl,e halilsototphaetrhtoilsosguiecsalwielvlableuadtiisocnasrdoefdg.ross lesions). Unless RD atseNl otiveri aLn ittger: ERaxtasmitnhaetddfoorngortodseslilveesrioansl.iteUrtweriil wbiellsbaecrsitfaiciendedonwidthay1205%oafmpmroensiuumemdsuglefsitdaettiooncaonndfi the absence of implantation sites. `Scheduled Sacrifice - Rats Assigned to Pharmacokinetic Sample Collection: Onedeadysae1mt4r(peeotfsitsnpigadregtluoamnnl,yd)msiwlfikrloalmnbetdhcbeollaolxoeidlcltaserdaym,aptslheodsre,ascciarci,lbiaveebdrdpsoremecivtniiaooulns(laryni,gdhftirnolgazuteeinrnaaallnlrdoebsgeti)oornaesnddof (e7h0narCmoarcobkeilnoawt)iucntsitahmsephtilhpeomrceaocnlitlce,tcotaitbohnde,omFSiponoagnelsnoaernradftoirpeoalnnvafilceymvsaiissl.ceeraFatolswliwoliwlibnebgepteshraefcrocirofmmiepcdle.de,tiTaohnnde 2 gross Humber naencdrdoipsstyriobfution of implantation sites will be recorded. Scheduled Sacrifice - Day 22 Postpartum: Orantsdfaoyll2o2winpgosrtepmaorvtaulm,frbolomomdestaamboplliessmwiclalgbinegc.ollected from the Fo generation female BThleooidnfseraimorrpelvfeersniga(eracapatpveradocaxenindmtratitfreualgn4eys.fmeTrLheedesaicenhtro)uswmiewlriulblmebsceeopltlareracanttseofdrefrtrrueobdmesit.nhteoTmphoaeltysepamrmaoplplyerlasetnsweivlilabe `bSapenudnscoilnlaalbecetlieodnwtiitmheptohients.tuAdlyl nsuammbpelre,s awinlilmbael iidmemnetdifiiactaetiloyn,frdoazteenoofncodlrlyecitcieona,nsdtudy day aintained frozen (-70C or below) until shipment to the Sponsor for analysis. Tahnde fpeelmvailcevirsatcserwaiAlwlitlllihvebenresbepecetrsifaoocnrrim(feridicg.ehdt,Tlhaatneedrnalaumlgobrbeoesr)safnrneodcmdrieosaptcsrihybuodtfaitomhnewoiftlhliombrpealcaicnco,tlalaetcbitodenodm,siinteasl ill be frozen arencdorsdteodr.ed (-70C of below) until shipment to the Sponsor for analysis. 001449 418-014PAGE C-15 Protocol 4P1a8g-e01145 Dams with No Surviving Pups: Dams with no surviving pups will be sacrificedafter the last pup is found dead, missing or presumed cannibalized. PrSTimaoamrtptelmoealswiarlclartibfveiicaes,pthuaenbiilnnofoeardiroserfarvmiepgnleareact(eaadpvpacreaonxtnridimfatutrgaeenl.syfTe4rhrmeeLds)iewnritluolmsbeweirlcluomblleseectptraaerndsafftreoorrrmteutdbhieenst.o aThe oFPoonlllydarpsytrioiopcnye,lasentnueddtymuadbiaenytlaaaibnnedeldecdoflwrlioetzchetnitoh(ne-t7si0tmueCpdoyoifnntub.meblTeohrw,e) usannatiimlmpasllheiiwdipelnlmtibefenitcotiatmitmohened,iSdapatoteenlsyooffrrofzoren analysis. (lAi4v7ger0rossCescotnieobcnerl(oorpiwgs)hytuonlftaittlehrseahltilhpoombreaenc)itfc,rtooamtbhedeaoSmciphnodanlasomarnwdiflolrpebatenvaiclcoylvsliiessc.cteerPda,oswfitrlploazbreetnupamenrddfaostrtamoefroder.dthAese dams will be excluded from summary tables. Rats Found Dead or Moribund: SGRehalstiesvretvrhayattwiiodlniebisoermeaaxrdaeem.siancTerhidfeifcoreradttshbweeiclclaabuuesseeexoofafmmdioenraeitdbhuofnordrmgcorornoidsbistuilnoedns,icooannbsdo.irttAiioolnnivooenrr pstrehceetmdiaoatnyur(tfehieght FSaathteierpawmlielllnotbbeteo)rtfehcreoormSdpeedoa.ncshAodbroafrmotrewadinlalfleybtseiussc.eosl_laePcnrtdeeodgr,ndafernlocizyveensrtaeadntdupssuatpnosrdewduiltl(eb-rie7n0eeCxcoaonmrtiebnneetldsowto)of utfnheteimlale `exatmemnotnpioussmibsluel.fidUetetroicoofnfaiprpmartehnetlaybsneonncpereogfniamnptlarnattastwiiolnl be stained sites". with 10% `Shipping Instructions: AI`lnSlcalsmupadlmeepdslewwiisltlwhibltlehebsehsiampmappielndetasfiranonezdednfsroeonnztedntroy(Ki7ic0iesvJCi.aoHoravnbeesmleiongw,h)tPuhmn.atDii.l.,shaAitptpmhaeecnkptirnfeogvriioasuntsawlliyyslciibst.eed `address, Both the recipient and the Study Monitor will be notified in advance of sample shipment. TESTS. ANALYSES AND MEASUREMENTS - F1 GENERATION: Viability: Postpartum Period: dLaiitltye.rs Twihllebpeupobssienrevaecdhfolritdteeawidllpbuepscoautnlteeadstotnwciece daily. 001448 418-014:PAGE C-16 Protocol 4Pa1g8e0164 linical ervatic ar r ce: Postpartum Period: Once daily. Clinical observations may be recorded more frequently than cited above,if deemed appropriate by the Study Director and/or the Study Monitor. Body Weights: Postpartum Period: Days 1 (birth), 4, 7, 14 and 21 postpartum. Sacrifice: "Terminal weight. METHOD OF SACRIFICE = F1 GENERATION RATS: As previously cited for Fo generation rats. decapitation. NECROPSY - F1 GENERATION RATS: Culled pups will be sacrificed via Gross lesions will be retained in neutral buffered 10% formalin for possible units `will be used to select one control group future rat of each evaluation (a table of random all tissues examined at necropsy will be retained, in order to provide of gross lesions). Unless. sex from which control tissues for any possible histopathological evaluations `specifically cited below, all other tissues will be discarded. `Pups Found Dead on Day 1 Postpartum: As described previously, caps and and labeled tubes will be weighed (combined weights, after retention of pooled pup samples. These to the nearest weights will .001 gram) before be documented in the raw data, and copies.of these weights will be included with the packing list prior to shipment. Be bin. The lungs wil be removed and immersed in water. Pups with lungs that Pups that die before examination of the litter for pup viability will be evaluatedfor vital Sink will be identified as stillborn; pups with lungs that float will be identified as lesions will be preserved livebor, in Bouin's and to have died shortly after birth. Pups with gross `solution for possible future evaluation. "The lungs will be preserved in Bouin's solution evaluation. and the liver will be preserved in neutral `buffered 10% formalin for possible future 001458 . 418-014:PAGE C-17 Protocal 4Pa1g8e01147 ups Found Dead or Moribund. to 21 Postpartum: Pups found dead or sacrificed because of moribundity will be use of death or the `moribund condition. examined for gross Pups with gross lesions lfeveoasulinuodantsoinaonnd;dagyfrosro2stshteolec4saipoonsstopfarptuupmswfilolunbde opnredsaeyrsve5d tion 2B1oupions'stpsaorltuutimonwilflorbpeopssriebsleervfeutdurien neutral buffered 10% formalin. For all pups found dead on days er will 2 to 4 postpartum, the lungs will be preserved in neutral buffered be preserved 10% formalin in for Bploouusnisgnis'baslensdfoulttuhuteiroelnievvaeanrldwuialttlhieboen.l.ipvrFeosreralvlepduipnsnefuoturnadl dbeufafderoend d1a0y%s f5otroma2l1inpofosrtppaorstsuibml,etfhueture evaluation. Pups NotSelectedfor ContinuedObservation -Days 1and4 Postpartum: Adllap4yuppsoscutlplaerdtuomn d(aallyo1thpeorsctuplalretdupmup(sp)upwsillfrboemsdacarmisficaesdsviiganeddectaopitthaeticoonn.troTlhpeoollu)ngasnd and the liver wil be collected from the first ten pups culled day (control pool only on day (iespective of litter) from 1 postpartum); the lungs will each dosage group on each bfoermraeltianinfeodr ipnosBsoiubilne'sf solut uture ieovna,luaantdiotnh.e Rlievmerasinwiilnl gbecurlelteadinpeudpsinwnilelutbrealsabcurfiffeirceedd 10% and discarded without evaluation. Scheduled Sacrifice - Litters Assianed to Pharmacokinatic Sample Collection: On day 14 postpartum, liters assigned to the pharmacokinetic sample collection willbe saancdrilfiivceerdwiwliltchotlhleeicrterdesapsecdteisvcerdibaemds.preIvnidoiuvisdluya.l,"Tphoeolpeudpssawimlpllbeese(xpaermilintetedr) foofr bglroososd. lesions. Scheduled Sacrifice- Litters Assigned to Sacrifice on Day 21 Postpartum: On day 21 postpartum, six litters per subset will be randomly selected for collection of pooled samples (per litter) of blood and liver. Blood samples will be collected via the inferior vena cava from each pup, pooled um separator tubes. The `samples will be spun in a (per lrietfterri)gaernadtetdracnesnfterrirfeudgei.ntTo hseerserum will be transferred into polypropylene dy day tubes labeled andcollection with the study number, amples animal identification, date of collection, stu will be immediately frozen on dry ice and maintained frozen timepoint. (-70C or All s below) until shipment to the Sponsor for analysis. 001452 418-014:PAGE C-18 Protocol 4P1a8g.e01148 Tpohoelepdup(spewrilllitbeer),exfaromzienneadnfdorstgorroesds (le7s0ionCs.orTbheelolwiv)erunftriolmsheiapcmhenptuptowitlhlebSepcoonlsleocrtefdo,r analysis. All other remaining pups will be sacrificed and discarded without further evaluation. `Shipping Instructions: `AlSlasmpalmepslewisllwiblel bsehimpapiendtafirnoezdenfroonzednry(i-c7e0viCaoorvbeemliogwh)tumntaiill.shAippmaecnktinfogrlaisntalwylslisb.e iadndcrleusdse.d wiBtohththtehesarmecpilpeisenatnadndsetnhtetSotKurdiys MJ.onHiatnosrewni,llPbhe.Dn.o,tiaftietdhienpardevviaonucseloyfcistaedmple shipment. STATISTICAL EVALUATION: Aapvperroapgrieastea.ndAdpdeirtcieonnatlagpersocwieldlubreescaalncdul/aotreda.nalLyitsteesr vmaalyuebsewiplerbfeorumseedd,iwfhdeereemed appropriate. DATA ACQUISITION, VERIFICATION AND STORAGE: DDiarteactwoilrlabned/hoarnadp-paronpdrlioartecommapuntaegre-rmeecnotrdpeedr.soRneneclorwditshwiinll2b1edraeyvsiaefwteedr gbeynetrhaetiSotnu.dyAll obreigbinoaulnrdecaonrddsinwidlelxbede.stAorceodpiynotfhaellarrcahwivdeastaofwitlhlebTeesstuipnpgliFeadcitloittyh.eASllpoornisgoinralupdaotna will yreeqauresaftt.erPmraeisleirngveodf ttihsesdureasftwiflilnbalersetpoorrte,daatfttehrewhTiecshtitnigmeFatchileitSypaotnnsoorcwhiallrgbeefcoornotnaected to determine the disposition of these materials. 001453 418-014:PAGE C-19 Protocol 4P1a8ge01149 RECTO BOEMR AINTDAINSED: Protocol and Amendments. Test Article, Vehicle and/or Reagent Receipt, Preparation and Use. Animal Acquisition. Randomization Schedules. Mating History. Treatment (if prescribed by Staff Veterinarian). `General Comments. Clinical Observations and/or General Appearance. Tissue and Sample Collection, Processing and Shipment. Cap and Labeled Tube Weights. Body Weights. Feed Consumption Values. NLiatteurraOlbsDeelrivvaetriyonOsb.servations. GOrrogsasn NWeecirgohptssy(IOf brseqeurivraetdi).ons. Photographs (if required). Study Maintenance (room and environmental records). Feed, Water and Bedding Analyses. Packing and/or Shipment Lists. KEY PERSONNEL: Executive Director of Research: Mildred S. Christian, Ph.D., Fellow, ATS Director of Research: Alan M. Hoberman, Ph.D., DABT AssociateDirector of Research and Study Director: Raymond G. York, Ph.D.,DABT Director of Laboratory Operations: John F. Bamett, B.S. Manager Manager of of ASntiumdaylCoOopredriantaitoinosn:anVdalMeerimebeA.r,ShIanrsptietru,tiMonSal.Animal Care and Use Committee: Dena C. Lebo, V.M.D. Manager of Consultant, VReetgeurliantaorryyPCaotmhpollioagnyc:e:W.BRarabyaBrraoJw.nP,atDt.eVr.sMo.n,,PBh..DA.., ACVP 001454 418-014:PAGE C-20 Protocal 4P1a8g.e01240 il ORT: A comprehensive draft final report be finalized following consultation will with bteheprSeppoansroerd.onThcoemprelpeotritownilolfitnhcelusdteudtyheand wil following: Summary and Conclusion. Experimental Design and Method. Evaluationof Appendices: Test Results. Figures, Summary and Individual Tables Summarizing the Above GDaLtPa,CPormoptloicoalncaendStAastseomceinatt,edReApmoertnsdmofenSutpspoarntdinDgevDiaattiao(nisf,apSptruodpyriDaitree)ctaonrd's QAU Statement. i NAL ANIMAL Ts : ITnhsetitpurtoiconeadluArneismdaelsCcrairbeedanidn tUhsisepCroomtomciotltehea.veAbllepernorceevdiuerweesddbeysctrhiebeTdesitnitnhgisFapcriolttoyc'osl thatinvolve discomfort, dsitsutdryesasnoirmaplasinwitlo tbhee caonnidmualcst.ed in a manner to avoidorminimize The Sponsor's signature necessity for conducting below documents this study and the the fact fact that that this information concerning is not an unnecessarily the dpurpolciecdautirveesswteurdey mavaayilabbeleobftoarimneeedtfirnogmtthheesStaptoendsopru.rpoNsoeaslotfemtahteivsteud(iyn. vitro) 001454 418-014:PAGE C-21 Protocol 4P1a8g.e01214 REFERENCES: 1. CThersitsst.iaEn,nvMi.rSo.nmaenndtaVloyPtreokt,ecPt.iEo.n(A1g9e8n2c).y,IWnaVsihviongRetporno,dDu.cCt.ivNeatainodnaMluTteacghenniiccailty Information Service, U.S. Department of Commerce, Springfield, VA 22161. 2. Cnharlitsrteixaon,neM(.PSr.o(c1e9e8d4i)n.gsReofprNoadlutcrteixvoenetoSxiycmiptyoasnidumt,erNateowloYgoyrekvaAlcuaatdieomnsyooff Sciences, November 7, 1983), J. Clin. Psychiat. 45(9):7-10. 3. LCaonntgr,olP.DLa.t(a19i8n8t)h.e EChmabrrlyeos aRinvderFeCtrail:DCeDveBlBoRpRmaetn.taClhaTrolxiecsitRyiv(eTrerLaatboolroagtyo)ries, Inc., Wilmington, MA 01887-0630. (Data base provided by Argus Research Laboratories, Inc.) 4. LInasbtoirtuatteoorfyLAanbiomraaltso.ryNaAtniiomnaall RAecsaoduermcyesPr(1e9s9s6,)W.asGhuiindgetfoonr,tDh.eCC.are and Use of 5. SImaplleawnstkait,ioEn.ss(t1e9l6l4e)n. aFmarUbteemreusthdoedreRaztutem. maArkcrho.skPoatphioslc.hEexnp.NaPchhawremiaskovlo.n 247:367. 001458 PROTOCOLAPPROVAL: FOR THE TESTING FACILITY OQ fp -- ADliraenctMo.r HoofbReersmeaanr,chPh.D., DABT A ond G. York! ., DABT Associate Director of Research `Study Director Leen boiler Dena C. Lebo, V.M.D. Member, Institutional Animal Care and Use Committee FOR THE SPONSOR Don: 7%, Marvin T. Case, D.V.M., Ph.D. `Study Monitor 418-014:PAGE C-22 ProtocolP4a1g8e.0214 2 outlE Date 23-Oct-98 Date _@aDt Date 270.f 2g Date 00145 418-014:PAGE C-23 ATTACHMENT 1 `SCHEMATIC OF STUDY DESIGN AND STUDY SCHEDULE 001458 ATTACHMENT 1 418-014:PAGE C-24 ProtocPoalg1e8.100124 STUDYSCHEMATIC `CROSS-FOSTERING STUDY* Startof Fende Rats Premating Period (0 Cohabtzion | Presumed Period Gestaion Gla Ped Endo Schedued Sacre! Postpartum Lactfon Peo ME a. b. c. d. Dosage Period. Fproortoacdodli.tional details see "Tests, Analyses and Measurements section of the coFn1tgiennueirnagtiunotnilpduapys w2i1llpboestcpraorstsu-mf.ostered beginning on day 1 postpartum and F1 generation pups will be sacrificed on day 21 postpartum. Fo generation dams will be sacrificed on day 22 postpartum. Samples for analysis of pharmacokinetics will be collected from six dams and their respective litters on day 14 postpartum. These litters will not be cross-fostered and will be sacrificed following sample collection. 0145 418-014:PAGE C-25 ATTACHMENT 1 ProtocPolag4e128001(42 SCHEDULE" . 270CT98 Animals Arive - Acclimation Begins. 02NOV 88 Dosage Period - Female Rats (42 dayspriorto cohabitation until day 21 postpartum). 13DEC981P4 MD-E1C99D8EC98 AM 19 DEC 98 Cohabitation Period. . LFiarsstt PPoossssiibbllee DDaayy 00 ooff PPrreessuummeedd GGeessttaattiioonn.. 04 JAN 99 13 JAN 99 First Possible Delivery (Day 21 of presumed gestation). Last Possible Delivery (Day 25 of presumed gestation). 04 JAN 99 12 JAN 99 First Possible Day 1 Postpartum Culing - `Control Pool Only. Last Possible Day 1 Postpartum Culling - Control Pool Only. 08 JAN 99 16 JAN 99 First Possible Day 4 Postpartum Culling - Cross-Fostered Liters. Last Possible Day 4 Postpartum Culling - Cross-Fostered Litters. 08 JAN 99 13 JAN 99 First Possible Day 25 of Presumed Gestation Female Sacrifice. Last Possible Day 25 of Presumed Gestation Female Sacrifice. 17 JAN 99 26 JAN 99 First Possible Day 14 Postpartum Pharmacokinetic Sacrifice. Last Possible Day 14 Postpartum Pharmacokinetic Sacrifice. 24 JAN 99 - 02 FEB 99 Scheduled Sacrifice - F1 Generation Pups (Day 21 postpartum). 25 JAN 99-03 FEB 99 `Scheduled Sacrifice - Fo Generation Dams. (Day 22 postpartum). 08 JUN 99 Draft Final Report. a. The study initiation date is the day the Study Director signs the protocol. 001.468 418-014:PAGE C-26 ATTACHMENT 2 MATERIAL SAFETY DATA SHEET 001468 . 418-014:PAGE C-27 wTaATTEARISAHLEETSEY 3 cantar Ssiae-1000 St. Paul, Minnesota 1-800-364-3577 or (612) 737-8501 (24 hours) Copyright, 1958, Minnesota Hining and Manufacturing Cospany. AiBILnLfLeoirrtmiogasPhtetirdsonaprvfreoosirevorivtdneheded.1pthuarsCtpoosoppsiyesidngofinapnrfdou/ploelrrlwdyiotwhuntlionlaoidzciihnnaggngoeSfsMthupinrsloedsusc: ts 20 DrEeiiroerhienrsagtrcehedesscweoinpttyhitnshoerobttiahneitneenodtriirognionmoaflSH1e,3arranenisdnsglad oprrofoitthertuhiesreeon. TOhIAVFIDoSEIbOWNw:F:LiwOnRDCHEBMrIanCdALSFluorochesical Surfactant 10 NoaMBmiEyR0/eU3s..ePa7.s 0000-.551111335.50.90008042-71 3oe8..00a2t0r7--s0e1r0e4.-15 0o00.-s51111d3s5.-0029a1r515--28 Z2O5P5FE0oRESDoE:rCEJSat,nruMaaornyvse2mb,ero 19089,8 i1987e DocumENT: 10-3796-8 i innedien Shs wo. Percent TPoPoTOTTAAASSSSeSTITiUomMn PPPEEERRRFFFLLLUUUOGGNRNOOOAAALLLKKYYYLLL SSSUUULLLFFFOOOMMMATTTEEE.................. 2s2e78794i5..2s30e9---e34033382 -8-- 786 PoTTAASSSSTTiiNm PPEERRFFLLUUGGRNOOAALLOYLL SSUULLFFOOMWTTEE........... usaerrez.zseen.s 21 oa 2 piso DATA VVBaOirIooLnnINe GpaPEOsIsNuw TR:e:..e .11.2.I.1r vI1evI1e. 1eees NWMIAAA ESSvoPacEpuCosIRtFaLITICITcOYNmRwIAwTaET:E1R.:110110111111011111l1 GWaalistkanDe6 Haters PERCENTVOLATILE: ........c.c.. 0%(Bulk) VMISCeOScITYp:oint o1c1 I eaI ns NWI(pD0.1% Aqueous) APP[ElAaRnAtNGcEolAorDed,ODOfRr:ee flowing powder. Ae bbreviations: NID - Wot De utersined WIA - Not Applie cable Gh - Approxiemeaeteenley 00146 418-014:PAGE C28 MJSaDnSu:aryFC2-99,5 F19L9U8ORAD Brand Fluorochesical Surfactant Pace 2 "37 FIRE AD EXPLOSION HAZARD DATA FFFLLAAAMASPHSIAPLBOLEIENTLL:II.NM.II.TT.SSco-cLoUEzELoLTo:n.1e..e.1e.1n.1e.0n.0s. NNOIRAE MIA ATOTGATTIONTENPERATURE: |... NIA EXMTaItNeGrU,ISHCIaNrbGonMEDdIiAo:xide, Ory chemical, Foss SHPoEeCIrALtvfieuvFelsIlR'EpprrabFosaItsnGeodHscrTetIiNavGroeroPucRnplOdroCetEsahDrsiUmunRsrgE,e,S:Wdeaisinascnltdudainbndrgelaehtgehsli,mnegtf,aacpepsaerlmafats-ukc,so,ntaanbidunnekde,r cost rotictive covering for exposed aress of the head. UNUSUAALaaFrIdREouAsNDDeEcXoPmLpOSoIsOiNtiHoAnZAsReDcSt:ion for products of combustion. a REACTIVITY 0ATA STABILITY: Stable INNCoOMtPAsTpIpBlIiLcIaTbYle.- WATERIALS/CONDITIONS TO AVOID: HAZARDOUS POLYMERIZATION: Hazardous polyserization Will not occur. MAaiZAunRoDorOinUdSeM,oDnEToCowOxiMidPceOSIVaTnadpIoOrNCsa,PrRbOGoDnaUsCeDTsiSo:xoirdeP,arOtixciudleasteosf.Sulfur, Hydrogen SL EWIROWENTAL INFORMATION SPoGIhLnsLpeorRuvEneSdPOpNorSreEc:waautteiornstofraovmoidotdhuesrtisnegc.tioCnAsU.TIONV!acuAusv,acuuusms eran ignition source. Clean up residue with water. eclteasnwereepcionugld Place in an approved metal container. Seal the container. REDCoOMMnEoNtDEreDlDeIaSsePOSTAoL:Waterways processes that could result ionr aseqwueart.ic cDooncnoetntruasetioinnsprgordeuacttesr torhan i$n7d7u0storfiatlhe orlocwoesmtmeErcCiSa0l ofracLiClSiOtyconicnentthreatpiroens.enceInocfinaeractoembiunstianble Amlatteerrniaatli.ve:CombDuisstpioosne pofroWdauscttes wpirlolducitncliundea HfFa.ciliDtiyspopsearlsitted to 001468 Abbreviations: NID - Not Determined WIA - Not Applicable CA - Approximately 418.014PAGE C29 JJaOnSu:aryFC2-95,5 P19L8U8ORAD Brand Fluorochesical Surfactant mee 8 =. EWIROWENTAL INFORMATION (continued) accept chemical waste. ENOEVIrREOMrEe1NraTia)AsotLarnistDcAiTmsAgFh:/i{lsL;hepL4oC8nS-iOvs,r-maFcaErtCoShDoe,haidrDuaMspi)nhenncoiuda(PmMiga/sglen,aphaR=laeSisOnbopawgr/o1Ts;reoluCstOs(D)S==.a30l80s4og/l, S7a: 80020 = Nal. REVVGoOUlLCaAtTLiOelRsseY HOIrzNgbFaOn&RiMcAETxIeCOmoNpm:tpouSnodlsv:entWsI:A.WA. Soirncse retglauploastsiio.ns Uv.aSr.y, EPcAonsWualztardaopupslicMaabsltes Nruemgbuelrat=ionNsonOer a(uMtGhTorUi.St.iss EPA vazardous) TThSias,pErIoNdEuCcSt,coCOmSpLl,iesALCWSi,thMItThIe acnhdemiKcoraela.registration requirements of ETPiCnReA WtaAgZaAnRdD: CLNAoSS:PRESSURE: No REACTIVITY: Wo ACUTE: Yes CHRONIC: Yes 6. SkaESTED FIRST AID Tr To evFeSoacnOotkmeIeastrt:`iGyatfliunsshedeiyaetes mwietdhicallargaetteanmtoiuonnt.s of water for at least 15 SKoITNmamtoCsdomiuizTnAtaGetTl:aydflculsohthisnkgi.n wIifthirrliatragteioanmoupnetrssisotfs,watcearl.l Raempohvyesician. Mash Contaminated clothing before reuse. IN2AiLaoAnlTaaIjOnsNay:sipstyoanpstoncsonotcicnuuer,, rcealmlovea ppheyrssiocniatno. fresh air. If IFDrSWiAnLkLOtWuEoD:lasses of Water. Call a physician. 7. PRECAUTIONARY INFORMATION Tm v`eAvPoRiOdTEsCyTeIOcNo:ntact. Wear vented gogoles. co1a64 Abbreviations: W/O - Not Determined NIA - Not Applicable CA - Approximately 418-014:PAGE C-30 JWSaDnSu:aryFC2-99,5 F19L9U8ORAD Brand Fluorochesical Surfactant "7. PRECAUTIONARY INFORMATION (continued) PAGE 4 SKVINoiPdROsTkEiCnTIOcNo:ntact. Aivarial. A pair of Wgelaovresappmradoeprfiraotme gtlhoevefsollWohweninhgansdatleirnigslt(hsi)s are pBFeoervcasorominenangld,epdrc:oovteercbatulitloysnl. irtuebPmbresort.e4sctiUnvseeecesgosnaearrmeyonrttsomopr(rsoetvohefenrttthhesaknfionlgllcooowvneitsan)cgt:shouhledad bpeolymsatdehylofeneei/tphoelryvoifnytlhiedenfsollcohwlionrgidmeat(eSrairaalnse:x). REvCUeOsnMetMiEwlNiaDttEheDdsVpEpaNrrTeoaIp.LrAiTaItPOerNo:vliodcealsuexfhfaiucsitentvevnetniltaitliaotni.on tUsoemainintaaiwnell: "Taaisnsoitonasdeqbuealtoew, ruesceomampepnrdoepdrieaxtpeosurreespilrisaittosr.y prIofteecxthiaouns.t ventilation RE`SAPvIoRiAdTObRrYeatPhRiOnTgECToIfONd:ust. Select one of the Pespirators based on airborne concentration ofofllcoownitnagminNaInOtSsH aanpdproivned hacaclo?rdmaansckesWuiptphliOedSKAairregruelsaptiiroantso:r, fhualllf--fsaacsek dduusstt aanndd mmiisstt rreessppiirraattoorr,, full face supplied air respirator. PREDVoENnTaItONsaOt,F AdCrCinIkDENorTALsmoIkNeGESwThIeOnN:using areas thoroughly With soap and water. thaisshprhoadnudcst.afWtaesrhhaenxdploisnegd and before eating. RECKOeMeMpENcDoEnDtaSiTnOeRrAGdEr:y. Keep container closed When not in use. FINREonfAlNaDsaEzXbPlLeO.SION AVOIDANCE: OTNcHooEnRtsamProRikEinCnaAgtU:iToISnOmNooAfkRiYtnhgIeNwFhtORoiMblAaeTcIcuOosNi:nagnd/tohrissmporkoeducatndcalneadrestoulttheinformation of the hazardous decomposition products mentioned in section 4 of this hSOS. WIS MAZARD RATINGS: HPEEARLSTOHN:AL2PROFTLEACMTMIAOBNI:LITXY:(S0ee pRErAeCcTaIuVtIiToYn:s, 0section 7.) EXPOSURE LINITS INGREDIENT PPOOTTAASSSSIIUUMM PPEERRFFLLUUGGRROOAALLKKYYLL SSUULLFFOONNAATTEE...... PPOOTTAASSSSIIUUMM PPEERRFFLLUUCORROOAALLKKYYLL SSUULLFFOONNAATTEE...... VALUE UNIT 00..11 MMGG//MM33 00..11 HMWGG//MK33- TYPE AUTH SKIN TM hA =HYY 00146 TTWHAA M 34 YY Abbreviations: N/D - Not Determined N/A - Not Applicable CA - Approximately 418-014:PAGE C-31 MSDS: FC-95 FLUORAD Brand Fluorochemical Surfactant PAGE S January 29, 1998 [Ep -- wareozEn Weve wr TRE Am sn P+OXTAIISNSTTIWNOTOaAPTEIRC OFNL:UORLOiAsLo tKeYdL sSuUbnLssFotONatyAnheTce.Ee.so.vs.eiirnaudlimlc0a.sbt1vkepdoaiswruiortWoehGr/nbK'eyYS' otrhu,endessroTurHteASaKnepIoaNurstriecfrueolruatretloyY, eeHe etance. enicies san s1iwr sein Soaorpeion: SSouEncE0oF EXePOSvURE eLIsAITExDApToAs:ure Guidelines 8. HEALTH HAZARD DATA eveOTcoTnm,cr:iicatrion: signa/aysptons can include redoess, swelling, saA w coAmTo:ssationir(asteerrporoelmonwgedsweotrlirnegpseataendd Scoenntiacnt.): enaytbeeabrsosrbed through the skin and persist in the body for an -- A: etal 17 inhaled. pay be absorbed by innalation and persist in the body for an extended Single overexposure, above recommended guidelines, may cause: o Jeitatione (upper resEpniaratwoarsy)t:, sisgonvss/hsiyosgptaonsdsscnaeneziinngc.lude rLL-- : mot a Likely route of exposure To this product. fIlriinegsastseayirtesult from a single swallowdng of a sederate quantity of nay be naratul 1 allowed. moTeAtGeEpNeTEiIeTiYt:y assays indicets the product is not mutagenic. co1466 Sobravistionss Nib CNet outerained WIA | Wot Applicwsie GA - Approvisstely 418-014:PAGE C-32 MJEaDnSu:aryFC2-99,5 1F9L9U8ORAD Brand Fluorochesical Surfactant PAGE & "6. HEALTH WAZARD DATA (continued) REMPoRtODUtCeTrIaVtEo/gDeEnViEcLOiPnHEtNhTeALratTOXsItNSo:ral doses below asternally toxic Levels. OTTHhEiRsHEpArLoTdHucHtAZ4sARDnotINkFnOoRwMnATItOoN:Contain any substances regulated under California Proposition 65. A Product Toxicity Summary Sheet is available. SECTION CHANGE OATES HEADING SECTION CHANGED SINCE Movesber 05, 1997 ISSUE Abbreviations: ND - Not Determined NIA - Not Applicable CA - Approximately Tbehecoirnrfeocrtmat45ionOfinthethdiasteMatiesrsuieadl. Saf3etyWAKDEaStaNSOhWeAeRtRA(NMTSIDESS), isEXPbReElSiSeEvDedORto WIENRPCLIHEADN,TABIINLCILTUYDINOGR,FIBTUNTESNSOTFOLRIMAITPEADRTTOI,CULAANRY PIUNRPPLOISEED OWRARCROAUNRTSYEOFOF uPhEeRtFhORoMrANtChEe O3RM UpSrAoGdEucOtFisTRAfDiEt.forUsearpairstirceuslpaornsipbulreposfeoranddetseuriatianbilneg for Cuasnera'fsfemcetthotdheofuseuseandorapappplliiccaattiioonn.of Gaiv3enprtohdeucvta,riseotymeofoffWahcitcohrsartehat tuhneiquuseelry weivtahliunatethetheuse3rM'sprkondouwcltedtgoedeatnedracionnetrowlh,ethiterisitesissefnittialfotrhaat particular purpose and suitable for user's ssthod of use or application. D3ueptroovitdheesreimnoftoermaptoisosnibiilniteyletchtartonieclecftorroaniacs atrasnesrfveircematyo ihtasvecursetsoumletresd. irinenpfsrorerrsmoearntsti,aotniooombnitssasiianosendstoorriotmsalt4ceorGma4pttliaeobtnaessneesisnmaytorhinsoatcciunbrefaocrsyss.atciuorInrn,enatd3dMiatswiaoktneh,se no inforaation in the NSDS available directly from 3. 00146 418-014:PAGE C-33 ATTACHMENT 3 TEST ARTICLE PREPARATION PROCEDURE 001463 418-014:PAGE C-34 ATTACHMENT 3 Protocol 418.014 Version:418:014 (2P2a0gCeT91o8l2) TEST ARTICLE PREPARATION PROCEDURE Test Article: PFOS. Vehicle: 0.5% Tween 80 in R.0. Deionized Water. A. Purpose: The purpose of this procedure is of dosage suspensions of PFOS to provide a method for and the vehicle for oral the preparation administration to rats on Argus Study 418-014. 8. General Information: 1. All suspension containers wil be labeled and color coded. Each label wil Snpuemcbiefry,thceonpcreonttorcaotlionnu,mbdeors,agteestleavretli,clpereipdeanrtaitfiicoantidoant,eA,regxupisrabtaitocnhdate and storage conditions. 2. XSu_spenDsaiiolnys wil be prepared: Weekly _For__daysofuse 3. VehiclDeaiwilly be prepared: X_ Weekly ~~ _For__daysofuse 4. Suspensions will be prepared at a final dosage volume of mUkg. 5. SXa_fety:Gloves, lab coat, goggles or safety glasses and faceshield X__ HDuasltf--MFiascte RReessppiirraattoorr -- Full-Face Respirator/Positive Pressure Hood Tyvek SuitApron 6. Dosage solutions adjusted for Free base and % Pury. -- Yes X_ No (Calculations based on 100%) __ FreeBase __ Purity 7. Sampling requirements: Cited in protocol. 8. Storage: Cited in protocol. 001469 418-014:PAGE C-35 ATTACHMENT 3 Version; 418.P1ro4toc2o2lG4C18T.80194 Fagezorz TEST ARTICLE PREPARATION PROCEDURE NOTE: One the final volumes are achieved, stir bars are to be added to the containers; mixing should occur during sampling and/or administration. C. Preparation of Vehicle: 1. Add the required amount of R.O. deionized water to an appropriately labeled container. Heat the water to 50C, +5C, add the required `amount of Tween 80 and mix until uniform. (See TEST ARTICLE `CALCULATIONS for exact quantities.) D. Test Article Suspension Preparation: 1 To prepare the suspension for the high dosage group, add the required amountoftest article (See TEST ARTICLE CALCULATIONS) into an Vaephpircolper,iaatdedlyasiszierdb,alraabenldedhecaotnttahienemri.xtQurSe atod8t0otCh,e 2r5eqCui,reind aamwoautenrtbwaitthh for approximately 30 minutes or until the test article dissolves. 2. `Once the test article has dissolved, remove the suspension from the water bath and spin the suspension while it cools. (Be sure there is a visible vortex, this will achieve the desired emulsion. This may be prepared the day before use.) fl org Written by: -- Approved le Date: 23-ccr9 Clarification: 3// No ___Ybs (See attached clarification form.) ntialDate : ChoidX.pBhooupn? 001499 418-014:PAGE C-36 ATTACHMENT 4 'CROSS-FOSTERING PROCEDURE co147g 418-014:PAGE C-37 ATTACHMENT & CROSS-FOSTERING PROCEDURE ProtocPoalg4e181.001114 The cohabitation period will begin one day earlier for the rats assigned to the control group. Dams assigned to the control group that mate on the first day wil be allowed to deliver and will retain their natural pups until needed for cross-fostering purposes. This will provide a pool of control dams available for immediate cross-fostering to pups from PFOS-treated dams, thus preventing these pups from nursing from their PFOS-treated dams On the remaining days of cohabitation, both control and PFOS-treated groups will be co-housed with male breeders, one male rat to one female rat. Starting with this set of deliveries, all pups will be removed immediately (as soon as parturition is complete) from their respective dams and placed with either another dam assigned to the control gthreonupa odraamPfFrOoSm-ttrheeactoendtrdoalmp.oolIfwaildabem uassesd.ignTehdetfoitttherefrcoonmtrotlhagtrcoounptriosl npootolavdaialmablweil,l then be sacrificed via decapitation. `The first ten pups from the control pool (irrespective of iter) determined to be at day 1 postpartum wil be decapitated, and the lungs (saved in Bouin's solution) and the liver (fustauvreedhiinstnoepuattrhaollobguifcfaelreedva1lu0a%tiofno.rmaTlihne) rwielmabienicnogllpeucptesdfarnodm tphreesceornvterodlfporooplosdsaimblsewill be sacrificed via decapitation and discarded without necropsy evaluation. This cross-fostering procedure wil result in four groups of 12 dams/pups, i... CPoFnOtrSo/llPCFoOntSro(lSu(bSsuebtseDt).A),ThCiosnptrrooclePduFrOeSw(ilSluablsleotwBd)i,stPiFnOctSi/oCnosnttorboel m(SaudbesebteCt)weaennd prenatal and postnatal effects of the continuous maternal PFOS treatment. 00147). 418-014:PAGE C-38 ~ PRIME]DICA ArpSsnResearchDLiaboraBtoreiesa, t os SR PROTOCOL 418-014 Oral (Gavage) Cross-Fostering Study of PFOS in Rats `SPONSOR'S STUDY NUMBER: T-6295.13 Amendment 1 -- 4 January 1999 v CIEL uae Pagar Do ASTER 5 PRT The first ten pups from the control pool (irrespective of litter) and the first ten pups that are found dead (no autolysis present) per dosage group determined to be at day 1 postpartum will be decapitated at approximately 1 to 3 hours after birth, and the lungs and the liver wil be retained. Several (minimum of two) iSag oh Aa EAI FE EELe L. A procedure provided by the Sponsor is documented in the raw data (stored under refrigeration in scintillation vials) (see attached)] for future evaluation. The a ron buffered 10% formalin, respectively, for possible future evaluation. The ee sma and discarded without necropsy evaluation. The liver sections in McDowell-Trump's Fixative will be shipped (on cold packs), for evaluation by electron microscopy, to: JPeatahnonleogdyeAWsasrodciates Intemational 4915 D Prospectus Drive Durham, North Carolina 27713 Telephone: (919) 544-5257 Telefax: (919) 544-3218 The remaining samples will be shipped (ambient conditions) to the Kris J. Hansen, Ph.D., at the address cited in the protocol for histopathological `and biochemical evaluation. 01. 473 418-014:PAGE C-39 . ReaforsChoangne: rorolifsimtasre TcohnetiSnpuoendseovralhuaastiroenquoefsttheedrtehaastotnhsesfoerardeddituicoendalpsuapmspulrevsivable orbesteairnveeddfionr previous studies. 2. Cross-Fostering Procedure (Paragraph 4 of Attachment 4 to the protocl): oThoe fourth paragraphofthe Cross-Fostering procedure has been changed to This cross-fostering procedure will result in four groups of 12 dams/pups, i.e., Control/Control (Subset B), Control/PFOS (Subset A), PFOS/Control (Subset D) and PFOS/PFOS (Subset C). This Procedure will allow distinctions tPoRbOeSmvaedaetmbeenttween prenatal and postnatal effect of the continuous maternal Rea forsCho angn e: This corrects the protocol. Ge - Reh OFTens Alan M. Hoberman, Ph.D., DABT Director of Research Date A eo 25: nd G. York, Associate Director of Study Director . DABT Date Research and Deng Clube Dena C. Lebo, V.M.D. ons Date Chairperson, Institutional Animal Care and Use Committee MainsTae 04902 ManvinT. Case, DV.M., PhD. Date Study Monitor oo01474 418-014:PAGE C40 Protacal 418-014 Version: 412.P9a0gDee1c1a34f82) FIXATIVE PREPARATION PROCEDURE Fixative: McDowell-Trump's Fixative A Purpose: TofheMcpDuorwpeolsle-Torfutmhpis'sprFoicxeatdiuvreefiosr ttoispsruoevisdaempalmeestfhoordstfuodryth4e18p-r0e1p4a.ration B. General Information: 1. Al solution containers will be labeled. Each label will specify the protocol number, fixative identification, preparation date, expiration date and storage conditions. 2. Fixative will be prepared: _ Daily _ Weeky _x Once 3. SXa_fety:Gloves, lab coat, goggles or safety glasses and face shield XC Dust-Mist Respirator X Half-Face Respirator _ Full-Face Respirator/Positive Pressure Hood Tyvek SuiApron X_ Other: Chemical Fume Hood 4. Dosage suspensions adjusted for % Activity/Purity or Correction Factor: -- Yes X_ No (Calculations based on 100%) 5. Sampling requirements: Cited in protocol. 7. Storage: Room temperature col. 475 418-014:PAGE C41 Protocol 418014 Version: 418(-F29a0Deg1co948r) FIXATIVE PREPARATION PROCEDURE c. Preparation of Fixative: McDowell-Trump's Fixative 1 Add 860 mLs of R.O. Deionized water into an appropriately labeled container. 2 Add 100 mLs of 37% Formaldehyde to the container and stir until uniform. 3. Aundidfo4rm0, mLs of Gluteraldehyde to the container and stir until 4. Add 11.6gof Sodium Phosphate Monobasic to the container and stir until uniform. 5. Add 2.70gofSodium Hydroxide to the vessel and stir until uniform. [NOTE: The final solution should have a pHofapproximately 7.3.] Written By: (Jlel20i:[h0opns Approvw ed lohe Date: RINE Clarification: Se __ Yes [see attached clarification form] tniiatsDate: eLeughiXK.ropueit 2/1ofes 001478 APPENDIX D DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY c0147 418-014:PAGE D-1 DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY 1. Two extra female from the treated group (Group I) were used for pharmacokinetic sample collection on day 14 of lactation (DL 14), 22 January 1989. These two female rats in tum received all collections and were sacrificed on DL 14. This deviation did not adversely affect the outcome of the study because the pharmacokinetic samples were not analyzed. 2. Eight female rats rather than six female rats from the control group (Group I) were used for pharmacokinetic sample 21 January 1999 and 22 January 1999. collection The two on DL 14, 17 extra female January 1999, rats received all sample collections and were sacrificed on DL 14. This deviation did not adversely affect the outcome of the study because the pharmacokinetic samples were not analyzed. 3. A total of 25 litters from the control (Group I) and treated (Group Il) groups `were cross-fostered rather than 24 liters. This deviation did not adversely affect the outcome of the to each dosage group. study because only one additional litter was added 4. Twenty-five female rats rather than all of the rats in the control group (Group I) were placed into cohabitation on 13 December 1998. The cohabitation deviation did period for these rats not adversely affect lasted six days the outcome of rather than five days. the study because it This provided sufficient mated rats for cross-fostering. All deviations are documented in the raw data. LA fs 23-3) R: York, PRD. DABT Date Associate Director(of Resegrch and Study Directo APPENDIX E TEMPERATURE AND RELATIVE HUMIDITY REPORTS 1579 418-014:PAGE E-1 ARGUS Temperature and Relative Humidity Report Location: Room 04 Protocol Number: 418-014 Range of Dates: 27-Oct-1998 13:15 to 23-Nov-1998 15:49 STpaercgieetsR:aRnagle: TToottaall NNuummbbeerr ooff HDoauyrss:: Total Number of Data Points: TSempFeraTtSurFe | Rela3t0i%vetoHu7m0i%dity 652051 6520851 634 634 Mean (50): MMeadxiiamnu:m: Minimum: NNumubemroobffPPaeaiinnrttss HInigRhan(%g)e: (%): Number of Points Low (4): 1 @19| 2 80 e7a6d4 65722 665 28% 64 (1000)|| 633 (90)9 oon 1 2 Report Generated: 04-Feb-1999 at 17:26 COMMENTS: REVIEWED BY: 0. Nod... ATE: 2.897 Cumulative by Location (v04.01.97 Co%47- (neo 418-014:PAGE E-2 ARGUS Temperature and Relative Humidity Report Location: Room 27 Protocol Number: 418-014 Range of Dates: 23-Nov-1998 15:49 to 31-Dec-1998 14:10 STahrogtets:Range: TTToootttaaall NNNuuummoomobbfffbRDebaeoayursrnP:eanrt: er Temperature | ae Relative Humidity srw5ds0on srt5gh0n `Mean ( SD): NMMeaisxntioimnmu:am:: 709 1aT2se6 (0.7) 571 83a2 (6.1) Number of pains NumberofPoints Number of Points Low G41 in Range High (%): (%): EI813 0 + (100.0) (0.0) A913 0 (100.0) (0.0 Report Generated:04Fe1099 37:28 comments: -- REVIEWED BY: an Tid. DATE: 379 , CumbyLocation(vO40197 0014399 148/ 418-014:PAGE E-3 ARGUS Temperature and Relative Humidity Report Location: Room 35-37 Protocol Number: 418-014 Range of Dates: 31-Dec-1998 14:10 to 18-Jan-1999 08:45 STpaercgieetsR:aRnagte: TTToootttaaalll NNNuuummbbmeerroboofffeDHDaoartuyarss:P:oints: Temperature 64F 10 79F 2a21r4 | Relative Humidity 30% to 70% or24 Moan (2 5D): MMMiaenxdiiimamnuu:mm:: Number of Points in Range (4): NumberofPoints High (%): Numob fPoeinr ts Low (%): se un| sa er E778H34 ss5e0s wr wo| a7 mp 0 .0) 0 0.0 o .0) 10 @3 Report Generated: 04-Feb-1999 at 17:30 commeNTs: REVIEWED BY: th. Mad. Date: 2.8917 Cumulative Sv assiin (NBO GT 0014589 418-014:PAGE E4 ARGUS TemperatureLoacnadtiRoenl:atRioveomHu2m7idity Report Protocol Number: 418-014 Range of Dates: 18-Jan-1999 08:45 to 29-Jan-1999 09:57 TSpaercgieetsR:aRnagte: TToottaall NNuummoobbffDHeoeauyrrsrs:: Total Number of Data Points: TFem0per7at9ure | Rela3t0i%vetoHu7m0i%dity 21520 21520 26 265 Mean (50): MMeadxiiamnu:m: Minimum: NNuummbbeerr ooff PPooiinnttss iHnigRhan(4g)e: (4): Number of Points Low (4): 04 @on | 3 (83 70240 67044 651 31 26 (0000)|| 215 049 oo 9 | o o Report Generated: 04-Feb-1999 at 17:33 COMMENTS: REVIEWEDBY: Mm. ad. DATE: 2.691 Corilatrea ARE OR DST 0014 3 418-014:PAGE E-5 ---------- ARGUS-------- Relative LHoucmaitdiiotny:DReovioamti0o4ns Report Protocol Number: 418-014 Range of Dates: 27-Oct-1998 13:15 to 23-Nov-1998 15:49 SHpuemciideist:y rTaatrget Range: O3NDowa-t1e998 T1i4m00e R2H8.61 30% to 70% Date Time RH. H=Value out of range - High L =Value out of range - Low R.H. = Relative Humidity (%) Report Generated: 13-May-1999 at 10:45 LL These deviations did not adversely affect the outcomeo interpretation of the study. "The following deviation(s) impacted on the outcome otfh study as described: Study Director: Z = : ; __ Deviations by Location (v04.01.97) Date: (3 124-99 001 83 418-014:PAGE E-6 ARGUS Relative Humidity Deviations Report Location: Room 35-37 Protocol Number: 413.014 Range of Dates: 31-Dec-1998 14:10 to 18-Jan-1999 08:45 SHpuemciideist:y rTaatrget Range: saDantes jsainnieew OOs6llianniisessee OGesualniioene OOeeJvaanniisee9s Osvanise T2i10m0e RsHsl. 220000 SSiOLL 0o0t0o00 SS1LL 00230000 5S2al 00650000 SSeSlL 0600 Sel 30% to 70% Date Time RH. H=Value out ofRraHng=e-RHeilgathiv_eLHum=iVdailtuye(4o)utofrange - Low Report Generated: 13-May-1999 a 10:50 / These deviations dd nt adversely affect the outcome or interpretation of the sudy. "The following deviations) impacted onthe outcome of the study as described: Study Director: ra : f = Dexiolions by tocaervGs 1:67 Date: 13M FD corp 1185 418-014:PAGE E-7 ARGUS Relative Humidity Deviations Report Location: Room 27 Protocol Number: 418-014 Range of Dates: 18-Jan-1999 08:45 to 29-Jan-1999 09:57 Humidity Target Range: Species: rat 30% to 70% 24D-Jaatn-e1999 Ti06m0e0 R7H0.4M Date Time RH. H=Valus out of range - High L = Value out of range - Low R.H. = Relative Humidity (%) Report Generated: 13-May-1999 at 10:54 + These deviations did not adversely afec the outcome or interpretation of the study. The following deviation(s) impacted an the outcome of the study as described Study Director: Z 2 = Date: (M899 Deviations by Location (v04.01.87) 001484 APPENDIX F PATHOLOGY REPORT 00145 P77 --- 418-014:PAGE F-1 PAaCthooloomgfSycpiAenscaespopnchciatayotnsesiamIsnttoenarnCoartpoiroa nal == --_-- PATHOLOGY REPORT (ANCILLARY STUDY) ELECTRON MICROSCOPIC EVALUATION OF LIVER AND LUNG IN CRL:CDBR VAF/PLUS RATS) ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS CLIENT STUDY NUMBER 418-014 PAISTUDY NUMBER EM 99.46 Prepared by: James B. Nold, D.VM. Ph.D, Diplomate, A.C.V.P. Pathology Associates International 4915-D Prospectus Drive Durham, NC 27713 April 5,1999 PREPARED FOR 3M CORPORATE TOXICOLOGY, 3M CENTER,ST. PAMNU 551L 44-1, 000 001483 --4c9e-- 1r5 0 Pro-- spectus-- Ore + D-- urham,-- Nori Car-- olina. 2-- 7713 + (e9-- 1e9) s544.-- 5257 (-- 91e 9) 544r e .321Fe Ae 8X Pathology Narrative Transmission Electron Micrograph Interpretation Forms Transmission Electron Micrographs Quality Assurance Statement 418-014:PAGE F-2 Section 1 n m v 001489 418-014:PAGE F-3 L Pathology Narrative 001499 418-014:PAGE F-4 PrimeticsAnAcrigulsayStPuuiyoNloegy41R8a.p0e1r4 PATHOLOGY REPORT (ANCILLARY STUDY) ELECTRON MICROSCOPIC EVALUATION OF LIVER AND LUNG IN CRL:CDBR VAF/PLUS RATS) ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS CLIENT STUDY NUMBER 418-014 PAI STUDY NUMBER EM 99.46 SUMMMARY nUelornaattralucrtatrapluyp,litvheerrserweormedianncrseaessetdendumwbietrhs1.o6fmpge/rokrgi/sdoaryme(shiwgiht-hdionshe)epPatFoOcSytes of tchoempnaurmebderwiotfhpceornotxriolsso.meQsuainnttirteaattieodnaonfimpaelrso. iDsiofmfeesreinncdeiscaitnecdelslluilgahrtlmyemmobrreantheasn otrwice `mitochondria were not discernible between control and treated animals in the samples. examined. Type II pneumocytes containing lamellar bodies were identified in both ctoynpterIoTlspnacnudmhoicgyht-edsosaendanliammaellsl;arhobwoedvieers,inthleurnegsapfpreoamretdretaotebdesaanmpilnecsr.easAeldthnouumgbhesroomfe. ldiafmfeelrleanrcmeawtearsianlot(esudrfbaecttwaente)nwcaosntriodelnatinfdiaebaletweidthsianmpallevse.olar lumina, no significant PROCEDURES "fTrhoemobsjeelcetcievde doafyt-h1ispsoustdpyarwtausmtroaepxuapmsinien abny oerlaelct(rgoanvmaigec)rocsrcoospey-ftoheselrinvgersstaunddy olufngs PFOS. In addition to a general qualiative evaluation or differences between control and shturerepfaatatecodtcayantnteislm,aalmmsee,lmltbahrebauonldteireaassntidrnucmttihuteroauclhneognn.ddrpioailntcshainncgleusdeind hqeupaanttoitcayttieosn, oafndptehreoxqiusaonmteitsawtiiotnhionf Ftesot atrhteicclreososr-fthoestveerhiincglsetuodnyc.e6d6aifleymbaelgeinCnrinCgD4E2BdaRysVpArFi/oPtoLUcoShabriatastiwoenrewigtihvaenmtahlee through day 21 postpartum according to Text Table 1. Grow | `NuFmebmerleofhMaasted PmFOaS gDonsage ae PFOS Concentration| mi Dosage Volume. LT 30s6 |oQenao | 0 | -- 5 | CSo 1apmoT separomraa mr.aTyhese soefdparma1 seamnadctohRniesispwe0ecrsesSOHIeS| rasedweFreosscfBoiacesm dafodllowin1 gramep5 lce cToelalri] n 8 " co1494 418-014:PAGE F-5 ensssERT a `The first ten pupsfromthe control pool (irrespective of liter) and the first 10 pups that were found dead (no autolysis present) from the treated group determined to be at day | liver were collected. Samples of lung and liver were thin-sliced and placed in McDowell- `Trump fixative (McDowell EM, 1976), and submitted to this laboratory for electron YS submitted samples were identified by dam number only. The five digit number is the dam number; the suffix 1 or 2 denotes the pup number from that dam's litter. [ Cowol| Omgi Highg Dow(d omps hwamy)| Lh examination. Thick (1u) sections were cut, stained with toluidine blue, and examined to select representative areas for further electron microscopy processing. Thin sections RESULTS AND DISCUSSION = Individual interpretations of light and electron micrographs A for each animal are shown in , characterized by discontinuous plasma membranes, organelles, dilatationofendoplasmic reticulum and lossof detailofcytoplasmic the nuclear envelope, and dilatation ofmitochondrial cristae. Fixation, however, was still considered adequate to assess for 001459, 2 > the presence of peroxisomes in the hepatocytes and lamellar bodies in the lung and to `make general comparisons between control and treated samples. LIVER a a LW dilated cristae, both artifacts due to suboptimal fixation. Dilatation of endoplasmic: TLaIndh,. the average numberofperoxisomes counted per hepatocyte photograph was 7 peroxisomes per cell (Text Table 3). Fetal and neonatal rat liver is a hematopoietic PO. TT [Mean Conoor= 770 [MemHighDose= 161] was higher in the high-dose Tn animals compared with controls (Text Table 3). Otherwise, LUNG Control (0 mg/kg/day). Lung samples were atelectatic and not inflated with fixative. alveolar and airway spaces. Figures 6-8 illustrate some normal cellularity and ultrastructural pulmonary anatomy. Type II pneumocytes (PN2) were infrequently preumocytweesre. identified by the presence of lamellar bodies, which are cytoplasmic - 001493 418-014:PAGE F-7 PrmedicsAnAcrigloaySuPtayvaNoo.gy41R5e0p1o4r vacuoles containing electron-dense laminated membrane-like profiles. Lamellar bodies represent the material secreted into the alveolar spaces as surfactant. Surfactant or lamellar material within airway spaces was very rarely identified in control lungs. The tthyipnemIaprngeiunmoofcycytteop(lPaNs1m). isTychpaeraIctpearcisutmioccalyltyesfladtitfefnereednatinadtelifnreosmthteypaelvIeoplaareusmpoaccyetaess.a MreapnreysepnrteaumwoacnystietsonapspteagaerebdettwobeeencPubNo2isdaalndwiPtNho1ust,deisspteicnicatlilvye slaimneclltahresbeodliuensg,sawnedrmeay neelosnatraelpraensdenhtaedd tneortmbineaelnbfruolnlycheixoplaarndeepidtwhietlhiuami.r. Additionally. some cuboidal epihelial Type I pncumocytesand lamellar bodies were infrequently identified in these control wluenrgesaalnsdo pnroesreelnitawbliethqiunanctaiptialtliaorny clouumlidnba.eassessed of them. Some lamellar-like bodies eHsisgenht-iDaollsyes(i1m.i6lamrgt/okgc/ondtaryo)l.saUmlptlreasst.ruTchtuerrael appaptehaorleodg.yohfotwheveerh,igtho-bdeosienlcurneagssewdasn.umbers opfutryppoeseIsIwpanseuemaosiceyrttehsaannidn claomnetlrloalrsabomdpileess;.atThleeascto,lildaepnsteidfyniantgutreheomf mfoorstilolufsttrhaetiavleveoli waintdhisnepctaapimlaladreydleufmiinndavaenqduoanntliytaatsimoanlilmpnruacmtbiecarl.werSeomseeenlawmiethlianrablovedoileis. wFeirgeuriedsen9d-f1i2ed illustrate some representative lung fields from th high-dose animals. CONCLUSION (Ulctornatsrtorluoctur1a.l6emvga/lukagt/idoanyo(fhliigvhe-rdaosned)lPunFgOsSofidreantpifuipesd firncormedasaemdsnturmebaeterdswoifth 0 peroxisomes within hepatocytes of high-dose rat pup livers. Quantitation of peroxisomes. iDnidfifcearteendcessliignhtcleyllmuolarremtehmabnrtawniceestohremniutmobcheorndofiipaerwoexriesnoomtesdiisnctermeiabtleed abneitmwalese.n cortrol and treated animals in the samples examined. In lung samples the presence of type II preumocytes and lamellar bodies was evaluated. Type II peumocytes containing. lamellar bodies were identified in both controls and high-dose animals; however, there appeared to be an increased numberoftype II pneumocytes and lamellarbodies in lungs fexrtormacterlelautleadr saanmdplweist.hinSocampeilllaarmyellluamrinbao.dieAsltahnodulgahmeslolmaer mlyaemleilnl-alrimkaetfeirgiuarlessuwrefracetant) wcoanstriodlenatnidfiarbelsetewditshaimnpallevse.olar lumina, no significant difference was noted between SIGNATURE OF AUTHOR Jersnuit James B. Nold, D.V.M., Ph.D., Diplomate, A.C.V.P. " bpd S 1449 Date 01494 418-014:PAGE F-8 PrimedicaAnAcriglluasrySuPdatyhoNloo.gy$1R8e.p0o1r REFERENCES MeDowell, EM and Trump, BF: Histologic fixative for routine diagnostic light and electron microscopy. Arch. Pathol. Lab. Med. 100:405414, 1976. 1s 01494 418-014:PAGE F-9 IL Transmission Electron Micrograph Interpretation Forms' ! iFnoarrmcshiavreedtsytpuodgryafplhei.cally corrected versions of signed/dated raw data sheets maintained 001498 418-014:PAGE F-10 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No. 418-014; EM.99.46 AnimalNo: 189041 Treatment Group: _0 mg/kg Control Sex: _anponplicable Species/Strain:fCoirgCeDsOBRVAFPIus. Tissue: __Liver Block No(s).: _99.46: 715683, Z15684, Photo/Negative No(s)._Z1S686.88 Sigaificant Lesions (check one): _____ Yes _X_ No Interpreting Pathologist (signature and date): Features: _ ZI3683:- Hepatocyte, several erythrocytes in sinusoid, and nucleus of probable endothelial cell on right ide of photograph. Six (6) peroxisomes evident in large central hepatocyte. Dilated blebs in RER are pseudo-peroxisomes. Abundant fine granular material, glycogen, in cytosol. Dense mitochondria with some granular deposits and dilated cristae. Poor definitionofendothelial cell. 215684: Several hepatocytic nuclei present, butpoorcytoplasmic margin definition. Some swelling of mitochondria. Dilated blebs in RER. Bile canaliculus in lower left comer. Two (2) peroxisomes in central cll. In adjacent two cells only single peroxisomes are clearly evident. Z15686: Single hepatocyte surrounded by erythrocytes. No clear definition of endothelial cells. Distended cytosol with glycogen-like granular particles. Abundant mitochondria and RER. Mitochondria have some granular densities and dilated cristae. Dilated blebs in RER. Ten (10) peroxisomes evident. 215687: Single hepatocyte. Granulocytic cell (hematopoietic)presentin bottom left comer. Distended cytosol with granular material. Dense mitochondria with some `granular deposits. Two (2) peroxisomes evident in central hepatocyte. 215688: Hepatocyte. Erythvoid precursor present in lower lef sinusoid. Dilated blebs in RER. Cytosol expanded with fine granular material, probably glycogen. Three (3) peroxisomes present. Conclusions: Normal hepatocytes. Plasma membranes poorly defined. D2e3n/s5e=m4i.t6opcehronodxriisaomweisthpesrohmeepagtroacnyutlearcdviedpeonsLi.ts and dilated cristae. Suboptimal fixation. co149 418-014:PAGE F-11 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.: 418-014:EM:99.46 AnimalNo:_1_ 89041 Treatment Group:_0mg/kgControl Sex: __non applicable. Species/Strain:RCAa.CDOBRVAFRIus Tissue: Lung Block No(s).: 99.462 715689 Photo/Negative Nofs).:_Z15693 Significant Lesions (check one): Yes X No Interpreting Pathologist (signatureanddate): Faleaamteulrleasr:-lZik1e56b8o9dy:. CSaepviellraarlytwyipteh1tphrneeeuemaodcoyttheelsia(lPNceIl)l pnruecsleein.tIsnurlruomuendninogf ccaappiillllaarryy.is 2N1o56p9r0e:umoTcyyptee2lpanmeelulmaorcbyotdeie(sPpNr2e)sepnrte.sen, containing multiple lamellar bodies (<7). Bnuecnleeia.thPtNh1e PtNo 2riigshtasciadpeilolfarpyhoctoongtarianpihngwiatnhemroyrtherotcryatnes,lupcleanttelcey,toapnldasemn.dothelial cell 2ro1u5n6d91n:uclCeoulsl.apOstehderalcveelolslalresssepctlae.arlOyndetfoipniesd,abPutNapwpietahrftloabteenprdicmayrtiolpylaPsNm1sanidn large collapsed 215692: septa. No Collapsed pncumocyte lamellarbodies evident alveolar septa. On top is a thin-layered PN1, and in the bottom left ilamlealrlgaerr pdreobfriilseaonfdatPwNoI.leuCceonctyrteasl.lyAalpspoeasrosmteo lbiepiad cdaepbirlilsariy plruemseenntcionnttahiencianpgilsloarmye lumen. Endothelial cell cytoplasm is evident, but no endothelial cell nucleus. No pneumocyte lamellar bodies 215693: Collapsed alveolar identified. septa. Attop is a profile ofa PN in the alveolar space. Bareenepartohbaibtliys aentdhoitnh-ellaiyaelrceedllPNnLuc,leain.dAtto tbhoettleofmt sliefdteisoafnpheortyothirsoacyPtNeL..JuSsutbtjoactehnetringuhctleiis a collapsed alveoli and thin-line PN1. No pneumocyte lamellar bodies identified. Conclusions: Normal lung tissue. Fixation adequate. Ouly one PN2 identified. o0149g 418-014:PAGE F-12 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Srudy No: 418-014:EM:9946 Animal No.: __18904:2 Treatment Group: _0ma/ke/Control Sex: _non applicable Specie/Strain:ECUa.COPAR VAFRIu Tissue: iver Block No(s): 99.21456946- 3 Photo/Negative No(s).;_Z15698 Sigaificant Lesions (check one): _____ Yes __X No Interpreting Pathologist (sigaature and date): fFeoautnudreeds:blebZsIo3f69R4E;R.HeCpyattoopclyatsem. fMuulltoifpglreamnuiltaorchgolnydcroigae,n.soPmoeorREenRdoatnhedlimaulltciepllle wdeiftihnistoimoneignrsainnuulsaoriddse.posNiutcsleaantdeddielartyetdhrcorcisyttaee.inTbhorteteom(3l)epfe.roMxiitsoocmheosndirdieantaifpipeeda.r dense 2Ge1p5o6s9i5ts: anHdedpialtaotceydtcer.istDaiel.atCeydtoRpElRa.smMiucltmiaprlgeindsenasnedmeintdootchheolnidarlicaewllisthpgoroarnluyladerfined. 2S1i5x6(966):perHoexpiastoocmyetse.ideMniafniyed.dilated biebs of RER. Dense mitochondria with granular `Tdweepolsviets(a1n2d) dpielraotxeidscormisetsaei.deTntoifpielde.f hepatocyte has swollen, less dense, mitochondria. W21h5i6c9h7:areSdeivfefricaulltpotoordliyf-ffeirxeentdihaetepaftrocoymtepse.roxMiasnomyesli.ghFtiavned(d5)arpkedriolxaitseodmbelsebisdeonftiRfEieRd,in s2i1n5g6l9e8:moreH-ecpiartcoucmystcer.ibLeadrhgeepactloecayrtea.reas in cell contain fine granular material consistent with glycogen. Five (5) peroxisomes evident. Conclusions: Normal hepatocytes. Suboptimal fixation. 31/5 = 6.2 peroxisomes per cell. 0149 418-014:PAGE F-13 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.:_418-014;EM-99.46 Animal No: 189042 `Treatment Group: _0mg/kgControl Sex: __non applicable. `Species/Strain:RCUa.CDBBR VAFPIus Tissue: Lung Block No(s).: _991.56949 -64 Photo/Negative No(s):_Z15704 Significant Lesions (check one): Yes _X No Interpreting Pathologist (signature and date): Features: Z15699: Alveolus containing several cuboidal pneumocyie cells. The lower clealrlgeapcpuebaorisdatol cceolnltsadion anoctocuopnlteaismnaalnlylalmaemlellalarrbbooddiieess,,pbruotbaabplpyeaarPtNo2.be pTnheeuomtohceyrttewso which haven't flattened out into PN1s or are terminal bronchiolar epithelia. At the left andboriomare capillaries lined by endothelial cells. The flattened PN1 at the bottom contains a segment of luminal surface with some dense electron-dense material. Some 2di1l5a7t0ed0:criAsltvaeeoilnamristpoaccheonwdirtiha.capillary/erythrocyte at left and pneumocytes around top alanmdelrilgahrt.bodTioesp.lefPt Npneautmroigchyttie csolingthatilnysmtohrreeee"leecmtprroyn"dveancsueo.les, possibly extruded 215701: Capillary Matulltoiwpelre rPiNghItswliitnhinegndaoltvheeolliaarl scpeasce.d. No lamellar bodies or PN2 evident. 2be15o7u0t2s:ideCetlhle ctayppiellcaornytaainndinegndcoetnhterlailallacmeelllaar tboopdylefnt,otbuctlecaorlmyprdiesscseerdnibbleet.weIetnaptpheears to p21n5e7u0m3o;cytAelvoenolruisghatnadndlilnionwgercelclesn.trLalarcgelel.PNT1hiaspipseaanrsartoeaocofnctoalilnaapssiendglaelvleaomlealrlasrepbtao.dy ainlvceeonltaerrsoefptpahoitnobgortatpoh.m cCeanptielrlaarnydarnigdhte.ndDoitlhaetleiducmriasttlaeftionfspohmoetomgirtaopchh.ondCroilal.apsed e2m1p5t7y04vacAulovleeoclounstawiintihnlginsionmgepnleaumemlolcayrtmeast.eriCaell. iDnelnosweerrcleelfltactocmenetrecrolnetfaticnosntpaairntsiaaly- couple clear vacuoles. although this cel is lined by a flattened PNL. `Conclusions: Normal lang and alveoli. A few lamellar bodies identified. 01500 418-014:PAGE F-14 `TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 418-014:EM:9946 `Animal No.: 8912-] Treaument Group: _0mg/kg, Control Sex: _nonapplicable SpeciedSirain;C.CDBRR VAFRud. Rat `Tissue: Liver Block No(s): __99.46:5 Z15773- Photo/Negative No(s).:_Z15777 Significant Lesions (check one): Yes _X No Interpreting Pathologist (signature and date): ______ Features: Z15773: Hepatocyte. Some segments of hematopoietic cells are present in ptisheedrioulxpaiptsieoornmreaisnghdctobculnoetmbeebdri..ngEonfdtohteheRliEaRl caenllddseotmaielsdialraetiinodnisotfintchte,nauncdlefairxaetnivoenliospepo.orS.ixTh(e6)re ZZbLl15e5b77b77i45n;:g oHHfeeRppEaatRtoo.ccyyFttoeeurswiu(tr4hr)ocpuaennrdaoelxdiicbusyloumsseaosmcetohuhenebtmoeadtt.oopmoisentdicscienlulss.oidMaalrekreydthdriolcayttieonatantodp pZrieIgrSho7t.x7i6Es:nodmHoeetshpeacltoiuoancltyetcdee.lliss poorly-defined and fixation poorly-fixed and disrupted. is suboptimal. Eleven (11) Marked dilation and blebbing of the RER. Mitochondria are denseand contain some granular deposits and dilated cristae. ZS1e5v7e7n7(:7)Hpeepraotxoicsyotmeesapcpoeuanrtsedsi.milar to those described above. Iti poorly fixed and RER andnuclear envelope are dilated. Seventeen (17) peroxisomes counted. Conclusions: Suboptimal fixation. Nine (9) peroxisomes/hepatocyte (average o5f cells) 001508 418-014:PAGE F-15 `TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 413-014;EM-9945 AnimalNo: 189121 Treatment Group: 0me/ke,Conrol Sex: __non applicable Species/Strain: CALCDOBRVAFPus Rat Tissue Lug Block No(s): __9971.547668 :6 Photo/Negative No(s).:_Z15772 Significant Lesions (checkone): _____ Yes __X No Interpreting Pathologist (signature and date): Features: Z15768: A rowof cuboidal epithelial cells, probablybronchiolar epithelium. `2bTo1hd5eic7ee6sn9,t:sruaAglggasemsiatnli,lveserecoveeralallPfcNco2un.btoaiiSdnaoslmseeepviedtrihaseltleieanmldpectdeylclrsv,iascctoauneosliiensstmewinthtoicwchihotnmhdabrryioanh.cahvieolbaerenepliatmheellliaurm. `The two central cells contain a few lamellar bodies and are probably PN2s. The smaller ddZaaLrrSkk7e7lr0in:cielnlAglacvpeeploelacurossntwtaioitnbhse lainbiansgalceclellsl.and underlying four lamellar bodies and is c3olPlNa2p.seTdhaelvoetohlearr tsewpotal.inTinhgeceslmlasllare ZPINSlTs7.1:Areaseems to beaterminal bronchiolejunctioning with an alveolar area with a sfqlauteieezneedinPNb1et(wleoewnertwrioghctu)b.oiCdeanlteeprtitohpeliisalacPeNs.2 fight one contains a lamellar body in is lumea. wAitththfivreiglhatmealnlda rl ebtodairees.c a pIit lalaprpieeas.r s Ttohe ZI5772: Two large cuboidal epithelial cellsor pneurmocytesand a PN1 lining cell (bottom). Some dilated cristae in mitochondria noted. `Conclusions: Appears normal. A few random PN2s with lamellar bodies noted. 01500 418-014:PAGE F-16 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.: 418-014; EM:99.46 Animal No.: __18915:. Treatment Group: _Q mg/kg, Control Sexi _nonapplicable Species/Strain:CR.aCoDBBRVAEPIs Tissue: Liver Block Nofs).: _99.46.7 Photo/Negative No(s).2.1_5Z71653767 Significant Lesions (check one): _____ Yes _X__ No Interpreting Pathologist (signaure and date): `Faebautnudraenst:gl2y1c5o7g6e3n:-likHeempaatetroiacly.itaePlpahsomtaogmreampbhrhaansesa laanrdgeenadroetahoeflciayltoceplllsasamrefiploloerdlwyi-th adenfdidnieldatdeudectroisptaoeo.rTfhiexrateioins.diMliattioocnhoofntdhreiRaEarRe,desohsoewianngd acsonrtoauinndsbolmeebsdoennscerodsesp-osescittison. ZA15bi7l6e4c:anHaelipcautloucsytie,prseismeinlatrin10lZo1w5er76l3ef.t. FFiovuert(5e)enpe(r1o4x)ipseormoexsicsooumnetsedc.ounted. Z15765: sinusoids. HEeipgahttoc(y8t)e.perPooxoirsofmixeasticoonunctleeadr.ly evidenced by loss of endothelial cell lining g2r1a5n7u6l6e:s, pSrioxb(a6b)lpyemroosxtilsyomgelsyccooguennt,edb.utMaluscohcoonfsitshteecntytwoiptlhapsomocronftixaaitnisonf.ine dispersed EZ1l5e7v6e7n:(1H1e)ppaetroocxyitseosmiemsilcaorunotetdh.ose described above. Many dilated blebs of RER. 8`.C8o/nccelluls(ifoinvse:coSunutbeodp)t.imal fixationofUssue. Number of peroxisomes counted: 001503 418-014:PAGE F-17 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.: "418-014; EM-09.46 Animal No.: 18915: Treatment Gr0oma/ukg,pCont:rol Sex: __non applicable Species/Strain:RCCia: DOVABFPR Is Tissue: ___Lung Block No(s): 99.46: Photo/Negative Nofs).Z:I_L55775682- Significant Lesions (check one): Yes __X No Interpreting Pathologist (signature and date): ______ eFleeactturroens-:deZnIs5e7c3y8t:opCloaslmliacpsmeadraglivneocloanrtasiepntian.gAaifreswpoarcgeaantelbloeRsiaonmdIsa ilanregde bdiylaattehdinbleb. Tluwmoinaca.piLllaamreilelsairntbhoedciesowlithlinaapraepeaucsmoonectyadtienssoormefreleamienltlhaer saitrruscptaucreess farreeenwoittehviindetnhte. 2le1f5t 7a5p9p:earAsltvoecoolnutsaainndaselpatmae.llmaorsbtolydycowliltahpisnetdh. eenPdNoItshelliinaelaicrelslpcaycteso.plaCsamp.illSamrayllatertop dZa1r5k76n0u:cleAurseaion fthceolcleantpesredapspeepaira,s wtiotbhaeiarnsupcalceeustooefat.PfNT.he space is lined by thin margin loufmPeNn.L AStuobjpacreingthttiostahicselslp,apcreiobsabalcyapaiPllNa2ry,cwohnitcahincionngtaailnasrgaeslianmgelelllaarmbeloldayr wbiotdhyininthiets 2c1yt5o7p6l1a:sm.PhTothoegcrlaepahrccoelnltaiintnshesceevnertaelrcaappiplelaarrsiteosbinethaepnceolulmaopcsyetde.septa. The large lumen catyttohpelbaostm.tAomirconstpaaicnesissolmineedpablye-fsattatineinnegdlPiNpi!d mcaytteorpilaalsmi.ntNeormliaxmeedllwairthboedndioetshneolitaeldcienll p2n1e5u7m6o2c:ytCeesn.traTlhidsarpkrocfeillle aipnpleuamrsentombayeabPeNt2ubwuilatrh saufrefawctlaanmtel(llaamrebloldariebsodiny)i.ts clyutmoipnlaa.sdmo.rsaFllattoteannedd tPoNt1hecryitgohtpolfatsmhilsinPeNs2t,hecornetmaaiinndseormofetlhameealilrasrpamactee.riaTlw. oAcafpoiclulsaroyf lamellar material is in the airway lumen. `COothnecrlulsaimeolnlsa:r Nliokremsatlrucftetuarlesluwaegr.eOfrneeewPiNt2hiwnascaipdielnltairfyieldumwiintah,saomaenslianmdgellleafrobcoudsieosf. lamellar material was present in airway lumen. 001504 418-014:PAGE F-18 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 418-014:EM.99.46 Animal No.: _18915.2 `Treatment Group: Omg/kg,Control Sex: __non applicable. Species/Strain:RCaUs.CDORRVAFPlu Tissue: __ Liver Block No(s).:92195.7543-6:0 Photo/Negative Nofs).:_Z15757 Siguificant Lesions (check one): ___Yes _X No Interpreting Pathologist (signamwre and date: Fdielaattuartieosn:s nZoItSe7d3i3n;nucHleepaartcoacvyetleopwei.thAnbuumnedraonutsgdrialnautleadrbglleybcsoogfeRnEinRcyprteospelnats.m.Some M(9i)topcehroonxdirsioameasppceoaurntdeedn.se with some granular densities and some dilated cristae. Nine 2ce1l5ls75a4re: pHoeoprlaytodceyftienesdurdruoeutnodepdoobryfeirxyatthiorno.cytHeespaatnodcyhteemactoonptoaiientsimcacenllys.diElnadtoedthseelgimalents 2Ta1nw5do7b5(l52e:)bspoLearfroRgxEeisRcoeamnnetrsdalcnohuuecnpltaeetador.ceyntveelwoipteh.abFuinvdea(n5t)gpleyrcooxgeins.omeBslecbobunitnegd.of RER abundant. ZI5756: `stcaoimnpilnegt.e Field cell. Some Cweiltlhssheavsernaulmheerpoautoscdyitleast,edhobwleevbesro,foRnElRy,onseomatetodparckeaantedrswoimteh nluicghlteus dilated cristaenotedin mitochondria. Four (4) peroxisomes cousted. is a Tcohuentcienlgl.at lower right appearstohave more peroxisomes, but is an incomplete cell for TZhaw1se57ld5ve7en:s(e1S2me)ivtpeoercarhlooxhniedsproaimtaoecswyicttoehusn,gtraealdnt.uhloaurgdhecnesnitteiersceanldwniuthmenruoculseudsiliatoendlbylceobsmpolfeRteERc.el. It f`iCxoantcilouns.ions: Average number of peroxisomes = 6.4 (5 cells counted). Suboptimal 01504 418-014:PAGE F-19 `TRANSMISSION ELECTRON MICROGRAPHINTERPRETATION Study No: 418-014 EM.99.46 AnimalNo: 189152 `Treatment Group:_0mg/kg,control Sex: non applicable SpeciesSrain:CA.CDRBRVARFiz Rat Tissue: __Lung Block No(s).: __99.46Z-110.5748 Photo/Negative No(s).:_Z15752 `Significant Lesions (check one): _______ Yes --X No Interpreting Pathologist (signature and date): Features: Z15748: Two alveoli with intervening septum and capillary. PNIs line the `septum, and there is alarge cell antached to the right. This cell has minimal cytoplasmic Zcoar1pg5ia0ln8leal9rl:yeslA,ulmmvieegnoh,lutbsbu,etliaisnlinnyogmpcpherolescs,yetnaet.nincSpoonllemauepmsloeadcmyseleelpsaar. maLtthieinsriipnahgloptisorgeprroaempsohee.ntytweitchoinnt the ains a slianmgellellaambeoldliaersbwoidtsh cinytioscpyltaospolfaspmrojbuastblbyenPeAaZt:h the ruclevs. At conve 1p are several bZoIdSiTe5s0.; Cuboidal epithoeflpriobaublmy bronchiole, Centrallyis a PN2 wilatmelhlar ZLSTSL: Alveolus and lining PMLs. At lower right is a capillary with a lamellar body in lthaemellulmaenboodfietsh.e capillary. Attopleft,in collapsed septal area, is a PM2 with several ic2sa1pa5si7lil5na2gr:ileesAclaetvleceownlituteshrotanhnrdeleefltovwawecirutohilgleihsnti.wnhgipcnheuspmpoecaytteoshisavfreeemeofllaasmelsltanrctbuodrieess.in AhtetmopTroigiht Ccaopnicllluarsyiolnusmi:naS.everal PN2and lamellar bodies identified. Some lamellar bodies were in 01508 418-014:PAGE F-20 `TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 418-014;EM-9946 Animal No.: 18952 Treatment Group: 1.6my/ks,high-dose Sex: __non applicable Species/Strain: CACDRRRVAFPlu Bat Tissue: _ Liver Block No(s). _99.46-15 Photo/Negative No(s).Z_1Z51752722-6 Significant Lesions (check one): __X Yes ______ No Interpreting Pathologist (signature and-- date):------------ Features: Z15722; Hepatocyte. Cytoplasm contains abundant ine gramular material, galnydcosgoemn.e dSiloamteed dcirilsattaeed.blTewbesontfyR(E2R0.) peMriotxoicshoomnedsriiadeanptpifeiaerd.deAndsjeawcietnhtgernadnoutlhaelridaelpocseilts 2is15po7o2r3l:y-Sdiemfiilnaerd t(opoZo1r57fi2x2a.tioNn)u.merous round dilated biebsof RER. Bile canliculus 2p1r5es7e2n4t: inSbiontutsooimdocfopnhtoationgirnagpshe.veTraelne(r1y0th)rpoceyrtoexsi.soSmeegsmceonutnotefd.hematopoietic `pgoroarnluylofciyxteid.c ceTlwl oinitnocpomlepflt.etEendhoetphaetloicayltecselonanridghlti.piAd-fcoenwtapienriongxipseormiseisnuasroeindoatlecde,lbuatre sot counted, 215725: Hepatocyte. Bile canalculus at top: sinusoid in lower right of photograph. Numerous dilated blebs of RER. Fifteen (15) peroxisomes counted. Z15726: Hepatocyte. Bile canaliculi at top right and left. Numerous, variably-sized dilated blebs of RER. Nineteen (19) peroxisomes counted. (C4onccelllusscioounnst:ed)I.ncrSeuabsoepdtinmuamlbefrixsatoifonp.eroxisomes. Average 16 peroxisomes/hepatocyie co1s0y 418-014:PAGE F-21 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No:418-014:EM-9946 AnimalNo: 1852.1 Treatment Group:16me/ke,high-dose: Sex: _nonapplicable Species/Strain:CiLCDRBRVAFPIu Tissue: __Lung Block No(s): __99.46:16 Photo/Negative No(s).2_1Z517517721 Significant Lesions (check one): __X Yes ____ No Interpreting Pathologist (signatvre and date): FheaavteuarePsN:Z1b5e7t1w7e;enAltvheeoml.usHowwitehvelrin,inagPcNel1ls.maTrwgionlsatriglesPepNaIrsataersetphreePseNa2t afnrdomaptpheear 10 sinusoid. appear to bSeeveexrtraalcellalmuellarl.ar bTohdeiePsNaIreonprtehseelnetftacsosnotciaaitnesd awbiutnhdtahnetPiNnZe, garnadnusloarmmeatoefrtihaelm 2in15i7s1c8y:toCpollalsam.psceodnsairsetaenotfwailtvehogllayrcsoegpetan.. Two central cells contain lamellar bodies, `tnhorreema(l3)puinlmoonne,arayndarscehivteenct(u7r)eiinstuhnecoltehaerr.becBaoutsheceolflsthaepcpoelalrapsebdesPtaNte2. , although, tZhLeS7ce1n9u:ioTlewsoacreellassswoictihataebdunwidtahntcicleian.trCieolaetsraalplpyeaarrettowroepcruebsoeindtablrcoenlclsh.iolOanrecelclosn,taainnds amsuslodcpilaete(d5w)iltahmetlhlealrefbtodcieels,.buTthiist mcaelyl abpepeexatrrsacteollbuelaar.PNB2o.tAhlcaelrlgsecolnatmaeilnlaarbubnoddayntis granular Z15720: mAaltveeroilau,scoannsdisltiennitngwicetlhls.glTycwoogecne.ls contain lamellar bodies, six in one and ZiIgShTt2Li:n thAelovtehoelru.s,Sfoimveellianminegllcaerllmsa,taenrdiaalcaapppielalrasrytionbetoepxruiagchetlolfulpahrotograph. This is ploasmseilbllayr baojduinecst,iaonndwiatphpaeatretromibnealPNb2rso,nchiole. Twoofthe pneumocytes contain multiple S`Coomnecleuxstiroancse:lluTlhaerrleamaeplpleaarrmatebreiailncnroetaesde.d numbers ofPN2s containing lamellar bodies. 001508 418-014:PAGE F-22 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION StudyNoi 418-014EM-9946 Animal No.18952-2 Treatment Group: 1.6mg/kg,high-dose Sex: __non applicable Species/Strain:CA:CDOBR VAFRI Rat Tissue: __ Liver Block No(s): __99.46-17 Photo/Negative Nos):2_1Z5I7S127-16 Significant Lesions (check one): __X Yes _____ No Interpreting Pathologist (signature and date): ____ Features: Z15712. Hepatocyie. Poorly-defined sinusoids 22d endothelium at top Fight `fsaionnmdeebgorgatrntaunolumalraorfmapdtheeoprtoiosagilrtascpoahnn.sdisdTtiehlneatthewediptcahrtigostlcayyect,oegSceyont.moeMpildtaioslcamhtaeopdnpdberlaierabssavopofpleRuaEmrRid.neonusEseiwgaihnttdh(h8a)abvuendant pZeIrSoTiLs:omeHsepiadteonctiyftiee.d. Hematopoietic cell right. Bile canaiculus preseat in lower right. Abundant fine granular material. glycogen, in cytoplasm. Appeas to be increased numbers of peroxisomes. Twenty-seven (27) peroxisomes present. Z15714: Hepatocyte. Many dilated blebsofRER and abundant glycogen particles in cytoplasm. Fourteen (14) peroxisomes identified inthiscell. cpZyeItrSooTpx1liSass:omm.eHseMpiiadtteooncctyhitfoein.eddriiMnaatwnhiiytshdcisllloatmeedgrbalnebusloarfRdeEpRosiatnsdaanbdunddilaanttedglcryicstoage.en Npairnteitceleesni(n19) Z15716: Sinusoid with lining endothelial cell. Cellular fixation is poor. Adjacent OhhefeppRaatEtooRccyyattreeesspacrroeensetnnaoti.tnimnMapunhloyttiompgilrteoacpphheo.rnodxriisao,mResEaRrapnrdesgelnytc,ogbeunt-nloitkecopaurnttiecldesb.ecDaiulsaetewdhoblleebs Conclusions: f1i7xMateipoant.ocyte H(4epcaetlloscyctouensteadp)p.earCetlolscocnotnatianininacbruenadsaedntnugmlbyecrosg.eonf.peSruobxoipstoimmeals,cealvleurlaarge 01509 418-014:PAGE F-23 `TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.: 418-014;EM-9946 AnimalNo: _18952:2 Treaument Group: 16mak.highdose Sex: non applicable `Species/Strain: Cr:CDBRVAF/Plus Ra `Tissue: Lung BlockNots): __99.Z46i.518707PhotoNegative No(s):ZISTLL Significant Lesions (check one): __X Yes ______ No Interpreting Pathologist (signature anddate): npFurecoalbteauubrsle,ysal:nadmeal1ll3s1ao07blodiSiipensgslwechlPiecaNth vvlaeicrunsoglceaxsr,vawdeesdhnsoiCoeltmlehoecsaohllnvyeaoslnamrlsypiaocreg.coTrmhoeecreh.voaTnchudoelaeesrwoseureneldinoeed Sc3n7d0vo8it;inlCeollmlcaelplmibewrdiaanheva.enwhyserpseearpsaa.hecAmytohevhsloyneedr,iesdoPhnvNeicdnoogvelurebslmteeomltbearcabo:ndye:BeCneoatmh ies yan an rca bioofndtfiheeas,tupaepbdeorubiaggshhe,mtehWnehytiacmphepmcebaorrna1naen6mmantaeanryicalmatcpoocnhaSiaTnnidCnrgeax.meanTnsoyibocneoollfughaenofubdtaehlneac.teiAorddea1koCw2i5epr:event FZLNSTli0ni5nA-g tneclaievtoeldes,FXG onaiing mule lela bis (9) appr 0 ne oven by 3 ZcdooUrvSseTar1le0cd:obnAytlaivi'ensionlcgayrtaonspprlaaycsbmrinsotacoynpercxinrgdahcteesnldilonuteahdrelrbiaelacywhli.itnh SPseeNvveerrcasll]l3.clAsmdewjaliclneoncbtoodoiltielssiPeANecaopnpdtlraeyoisk eeZImSaTpLlLt:yyvdaceAuonlsieinsgs,lewehdi3acshkhtPacNve2elaippaesrqeunetelzyedebxeurwdeeedn tmwoelPaNb.odiicssc.yTtooptlhasmicgoonhsfistfsseevelrea BvsNhoLor.aogmgraeacprhloapmhealgleae,boordimeasybwietehxscomeelps: dAkesmmaalelrcaalp,iTlhyefeinbotdhieeslomwaeyr ibgehitn mtahergaidnfsocfenhte Conclusions: There appear to be increased numbersoftype I pneumocytes (PN2s), and `more lamellar bodies were seen in this sample. Otherwise, cellular morphology appeared. essentially normal. Fixation appeared satisfactory. 01580 418-014:PAGE F-24 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 418-014:EM-0946 Animal No.: __18956Treatment Group:_L.6 mg/kg,high-dose Sex: _non applicable Species/Strain:CEaUt.CDOBR VAFPlus Tissue: Liver Block Nofs).: __99.46-13 Photo/Negative No(s).Z:1_5ZL753723-6 Significant Lesions (check one): _X__ Yes No Interpreting Pathologist (sigaature and date): Fmeaattuurreeesr:yt2h1ro5c7y3t2e;s. HSeopamteiclasmienlulsaoriddecbornitsaiisnipnrgesneunctleinattehdeesriyntuhsrooiyd.ciTewaondenadcotohuepllieal cell 2n1uc5l7e3i3:are Hpreepsaetnotc.ytTehecroentaariensnoabcuonmdpalnettegrhaenpualtaorcypatretsicilnesthciosnpshiostteongtrawpiht.h glycogen, and annudmedriloautsedRcEriRstpaea.lisEaldeesv.enM(i1t1o)cpheornodxriisaoampepsecaorudnetnesd.e aAnbdilheavceansaolmiecuglursaniulparredseepnotsits 2b1e5t7w3e4e:n two hepatocytes. Hepatocyte. Similar to Z15733. Numerous dilated blebs of RER are present. `Twenty-four (24) peroxisomes. pZeIrSo7x3i5s:omHeespacotuonctyetde.with similar morphologyasdescribed for above cells. Sixteen (16) g2r1a5n7u3l6a:r dHeepopsaittoscyatned. dDilialtaetdedcrbiisteabes.ofEiRgEhtReepnre(s1e8n)t.peMriotxoicshoomnedsr.ia are dense with `Conclusions: per hepatocyte I(nacvreeraasgeedonfufmobuerrcsouonftpeedrcoexlilss)o.meSsubpoeprthiempaaliofciyxiatei.on,17.25 peroxisomes 001518 418-014:PAGE F-25 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No.:_418:014; EM-99.46 Animal No: _18956:1 Treatment Group:_L.6mg/kg,high-dose Sex: _nonapplieable Species/Strsin;RCaALCDOBR VAFPLs Tissue: ung. Block No(s): __99.46:14 PhotofNegative Nos).:21521752773 Sigaificant Lesions (check one): __X__ Yes __ No Interpreting Pathologist (signawre and dey: aFreeatPuNr2esw:it1h5l7a27m;ellAalrveboodliuess.witShoamdejoafcetnhtecobloldaipesseadrseeppatrat.iaTllhyeetmwpotyb,oatniodminproenuemcoecllyies alaprpgeearPNtLo.beAetxtthreudliefntgimsaatcearpiiallleaxrtyrawcieltluhalanrulcylewaittehdoetrhyetrhrloicpyitdem.aterial. At the topis a 2la1m5e7l2l8ar: bCoodlylaappspeedaraslvteooblearwsietphtian. tSheevleuramlencoelflsaccaopnitlalianryl.amellar bodies. At left one e2n1d5o7t2h9e:liaSlocmelelwnhuactleTu-ssihsavpeendtrcaalpitlolatrhye ecroynttharionciyntge.anWeiltehcitnrotnh-edelnusmeeenryotfhrtohceyctaep.ilTlahrey aortehesrecveerlaslalpapmeealrlaor bboedisees.ctiAontsltefhtrooufgfhiepldeuismtoheeyttheinscayttvoaprliaosumsicanlgilneisn.g ofaPN1. The s2m1a5l7l30l:amelAllavreobloadrielsunmeexnt ao ndislinnuicnlgecuesl.s.ThCeendtarrakl-lsytaisinainsgmaclyltPopNl2ascmonttoatihneinrgigahtcoofutphlies pcerleluampopceyatress,10abnedatnheeyxtceonnstiaoinnfraocmouapnleallavmeeolllaarrmbaocdrioesp.hage. To the left are a couple ZIST31: bodies. * A rAilgvhetocleanrtrlaul,mepanlea-nsdtaliinniinngg, cpenlel.umTocoyptleefatlcseollciosntaaiPnNs2sewvietrhaltswmoalllamlealmlealrlar bodies. The septa are collapsed inthisarea. LCaomnecllluasriboondsi:esInnoctreedasiendcnapuimlblaerrieos.f paeumocyies containing lamellar bodies, Some co1512 418-014:PAGE F-26 TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION Study No: 418-014; EM.99.46 AnimalNo: _18973:1 Treatment Group:_L.6mg/kg,high-dose: Sex: __non applicable Species/Strain:RCa.tCDOBRVAFPIs Tissue: Live Block No(s). __99.46-11 Photo/Negative No(s).Z:1_5Z1754724-7 Significant Lesions (check one): __X Yes ____ No Interpreting Pathologist (signawre and date): _______ gFelaytcuorgeesn:i2n1t5he74c2y;topHleapsma.tocTyhteerecoanrteainnusmearbuonudsadnitlaftiende bglreabnsuloafrRmaEtReroinalccroonsssissetcetnitonw.ith pMeirtooxcihsoonmdersiacoaupnpteeadr.deSnosmeeanldamhealvlearsmoamteerigarlannuoltaerddeexptorsaictesl.luSleavrleyntaenednp(o1s7s)ibly within 2c1le5a7r43s:paceHsepiantothceytlee,ftopvoerrtailolnosifmtihlaerhetpoaZt1o5c7y4t2e,. although fixation appears better. Nine 2(91)57pe4r4o:xiHseopmaetsoccyoutnet.edA.tlLeafmtiesllaarhemmaatteorpioalieattitcocpelalppineasrisntuosobied eaxntdraactelulpuplearr.right Napipneeatresento(1b9e)apdeirloaxtiesdobmielesccaonuanltiecdu.lus. Abundant area of glycogen in cytoplasm, C2e1a5t7r4a5l:ly Sisinaucsloeiadr,ceyntdoopthlealsimailcacerlelawanidthinmcaomnpyledtielaptoerdtbiloenbsosfofacRoEuRp.leAhfepeawtocytes. Zp1e5ro4x6i;soHmeepsaatroecyprteesecnotn.taNinos paebruonxdiasnotmfeinceogurnatnumlaadre.material consistent with glycogen in athpepecayrtodpelnassem.anTdhhearveeasreomneumgerraonuulsardidleaptoesdibtlseabsnodfsoRmEeR diolnatcerdoscsrissetcat.ionE.leMvietnoc(h11o)ndria peroxisomes 215747: No counted. photograph. Negative wastoodark. With five photographs already evaluatedi.ti not necesary toretake ortry to print thisimage. cCeolnlsclcuosuinotneds.: SAupbpoepatri(m0ablefiixnactrioena.sed peroxisomes per hepatocyte. Average 14/cell (four 001513 418-014:PAGE F-27 `TRANSMISSION ELECTRON MICROGRAPH INTERPRETATION `Study No.: _418-014; EM-09.46 Avimal No.:_ 18973: `Treatment Group: 1.6mg/kg,high-dose Sex: _nonapplicable Species/Strain: C:CD@BRVAERlus Rat Tissue: __Lung 215737 Block No(s).: __99.d6-12 Photo/Negative No(s).;_Z15741 Significant Lesions (check one): __X__ Yes No Interpreting Pathologist (signature and date): eiFxegtahrttaucrleeallmsue:llal2ra1rs5pb7aoc3de7i.e;s,VCeoonlftlwaahlpilscyehdaraseelvvaeelrosalolarsaepsgeppmeteaanr.ttsCoeobnfetraracelellleylai'sssincagytPtohNep2ilrawcsiomtnhttheaanpttpsarpoipnxetiaomratsthee0ly caonndtaacianpsilolmaerylawmietlhlaerrybtohdryocvyatceuaolneds.endAotchoeluipallecPeNlIiss aprrespernetseinntlaowdejralceeft(nc0tentthrealPNar2e.n of pZhIoStTo3g8r:aphA.lveolus with lining cells. The large mucieus belongs 10 a PN1 whose cboydtioepsl,aaslmtlhionuegshtih'esaiirnsapascueb.jacBeennteacatphiltlhairsyc.eTloatnhdejmuisdtdtloetheeftriigshat craepislelvareyrawliltahmaelnlar 2e1yr5i7h3r0o:cytAelvtheaotlduoseasncdonctoalilnapSseodmeselpaam.elSairmidlearbtio i21t5h7e38l,umseonme lamellar bodies appear 2to15b7e4w0i:thiLnatrhgeelcuapmielnlaorfy awictahpislelavreyr.alPeNry1tshrloicnyettehseoalvteheolrairghstp.acLe.eft of this is a cellular elixnteesnstihoen,alpvreoolbaarblsypaaceP.N2,tht contains adistinct lamella body. A tia las ofPN1 ZalIo5n7g41t:heAllivienogl.arbutspnaocePcNo2ntaarienspraebseunntd.anLtamlealmellalrarbomdaiteesriaar.e pSreevseerntalinPNonIescaarpilplraersyent lumen and in some interstitial spaces in the right sideofthe photograph. sCaomnpclleu,siaolntsh:ougAhppmeaanry1webreeiinncrceapaisleldarlyamleulmlianaboadnidessoinmebeinUaslsvueeolcaormapnadriendtewrsittihticalomo] spaces. 001514 418-014:PAGE F-28 IIL Transmission Electron Micrographs 001514 418-014:PAGE F-29 NOTE: `The Dr. original glossy figure photographs (Figures 1- 12) MarvinT. Case withthedraftpathologyreport on were delivered to March 19, 1999. 001518 418-014:PAGE F-30 : PrimedicaAnAcrlgulsryStPuadtyhoNloo.gy41R8e.p0o1r4t Figure Legends EmiitgocuhrLoendrLiivaerr.eprHeespeanttoscoymtee sdihloawtiatnigonnoofrmcarilstcayet.opSloamsmeicpeorrogxanieslolmeess. aWrheiitdeentsipfoitesd.wiTthoin the righti an endothelial 1.0 mg/kg/day (control), c1e1l,l2a0n0dXemraygtnhirfoiccyatteisono.f adjacent sinusoid. Animal No. 18904- rFiegtuirceul2u,m (Lbivleerb.binHge)paitnodciyctaete.s Dsiulbaotpattiimoanloffiaxuatciloena.r eMnivteloocpheoannddrariroaeugdheensnedoapnldassmhiocw sHeommaetogproainueltaircdceeplolssiatrseparnedsdeinltaitnedscirniusstaoei.d aStoumpeperreprriegshteaotfatpihvoetopgerraopxhi.soAmnesimaarle lNaob.eled. 18912-1,0 mg/kg/day (control, 11,200X magnification. FIiagbeulreed3,,anLdivaeprp.eaCrelmlourlaernmuomreprhooulsogtyhainsisinmiclonatrrtools.FiAgurbeil2e.caSnoalmiecupleursoixsipsroemseesntarien: uApnpiemralriNghot.of18p9h5o2t-o2g,r1a.p6hmagn/dkgh/edmaatyo(phoiigeht-idcoscee)l,ls1a1r,e2p0r0eXsemnatgniinfsiicnautsiooni.d at upper lft. pFeirgouxries4o, meLsi,veri.ncrDeialsaetdatiinonnuomfbreoru,gahreenldaobeplleads.miAcniremtailcuNlou.m1i8s9e5v6id1e,nt1..6Smogm/ekg/day (high-dose), 11,200X magnification. dFiisgpuerresi5,onLoifveorr.ganHeelpleast.ocyCtleeadremsopnascteraattitonpg osfocmeellw,hwahtibcehttceornftiaxiantsiosnowmiethmemmobrreaenvoeuns `dmeber/iksg,/daapype(ahrisght-odobsee)e,xt1r1a,c2e0l0lXulimaanrgntihfeicpeartiisoinn.usoidal space. Animal No. 18973-1, 1.6 Epingeuurmeoc6y. teL.ungL.amTeyllpaerIblopdnieeusmwoictyhitnetihepcreysteonptlaaslmonigdetnhieifaylvtehoilsacrelwlasll2,saadjtaycpeentIIto type I preumocyte. Animal No. 18904-1, 0 mg/kg/day (control, 15.680X magnification. pEingeuurmea7.cyLeunigs.eviTdherneteacluobnogidtahle vpenneturamlocpyotrteisonlionfe tthhiesaaiirrwwaayyssppaaccee.. TAwfolasnmtaelnledlatmyeplle[ar obfoidtiseslaimneollnaercbuobdoiiedsalancdellissduegsgteisntedthtiso biseactoympeeaIltyppaecuI mporeeyutmeocwyhtiec.h AhnasimsaelcrNeot.ed1m8o9s0t42,0 mg/kg/day (control), 11,200X magnification. Fboidgiuerse,8a, ppLeuanrgs.sqAuemeozreed dbeentsweeley-nsttawionitnygpetyIppenIelupmooecuytmeosc.yteS,ocmoentlaaimneilnlgartwmoatlearimaellliasr tahlesotpyrpeesIeTntpnwcituhmionctyhee.caApnililmaarly lNuom.en1o8f9t15h-e1,se0pmtgal/kwga/lldabyot(hcoanbtorvoe),an1d1,t2o0t0hXe right of magnification 0015: 14 418-014:PAGE F-31 PrimedicaAnAcriglulasryStPuadtyhoNloo.gy41R8e.p0o1r4t IFipgnueruemo9.cytLeusn.g. FDoeunrsdei-stsiunicntilnagmecellllairsbaodtiyepseaIrleppareeusmenotcyitnethpeosciyttioopnleadsbmeotfwetheenttywpoe ItIype tphneeurimgohctyotfet,hiasltchelolugahretthewoldaamerlklalrammaetlelraira-ltyipsepbaortdiiaelslysuwrarsohuenddeodutbyinstohemsee lviapicdu.oleTsh.isTo `Aanpipmeaarls Ntoo.be1w8i9t5h2i-n2,a 1c.e6llmign/kacgo/ldlaayps(ehidgahr-edaoosef),se1p1ta..2a0n0dXmmaagynibfeicaamtaiocnr.ophage. Flaimgeulrlear10b,odLieusn.g. ACtoylpleapIspendeaurmeoaocfytseepitsatoshthoewilenfgt otfwtohetsyepceelIIlsp.neCuampoilclyatreiesscaonntdaaidnjiancgent a1l,v1e.o6lamrgc/eklgs/daarye (phriegshe-ndtodsoer)s,al1l1y,a2n0d0tomatghaeirfiigchattioofn.the photograph. Animal No. 8952Faippgeuarres11t.obeLuangt.ypeCIepnutreaulmlyoicsyatcee.ll Gcontlainiyng scevemraoatlerlgiamaellaelpapre-aalirkset-ostfrliulcltmuiurecshkwohfietchhe bcryotnocphliaoslma.rAcdejllasc.eCnutbcoelildsalteoptihteherliigahltceclolnstaairnecteonttrhieolleefstaannddrveepnrtersaelnttocitlhieattyedpe IT pneumocyte. Animal No. 18952-1, 1.6 mg/kg/day (high-dose), 11,200X magification. Fcoingsuirsete1n2.t wLiutnhgs.urfAalcvteaonltasrecsrpeatceedcfornotamitnyspaebIulnpdnaenutmolcamyetlelsa.r-lAiketympaeteIripanle,uwmhoicchyeislines mthaetesrpiaacleapopnetahretloefbteanpdneriugmhotc.ytTews,opcruobboaibdlayl tcyepllesIcIos nwthaiicnihnhgagvleycroegleena-slektehegirranlualmealrlar bodies. Animal No. 18973-1, 1.6 mg/kg/day (high-dose), 11.200X magnification. co1 51% Abbreviation A BC BE a CoaNp cp D DD pC DN DR EC ER G HC L LB M N Pp PNI PN2 mM RER 418-014:PAGE F-32 PrimedicAanAcrilgluasrySuPadtyhoNloo.gy18R.ep0o1r4t Index of Labels for Ultrastructural Structures Struerure Alveolar space Bile canaliculus Bronchiolar epithelium Capillary lumen CCaepnitliloalreys Cuboidal pueumocyte Desmosome Dense deposits Dilated cristae: Dilated nuclear envelope: Dilated RER Endothelial cell Erythrocyte Glycogen Hematopoietic cells Lipid droplet LMaimteolclhaorndbroiday Nucleus Peroxisome Type I preumocyte Type Il pneumocyte Plasma membrane Rough endoplasmic reticulum 001519 BEST COPY AVAILABLE 418014:PAGE F-33 mec WETF yg GCE EE RB AG oE o A AR pt AT TIR ) pW vy eZ Zi L Le Bh tT 3 Jo gS RC WL Jes Sr Cid erragAsuEo GSTEExRgE. TRHee Za 1LaGigI[Ra MEA bg CER CEE ER R NgRdEWhologtYei: VAR Ee NRT A ESoNN IWa s dT eKP AT h RET . 3Rg AHnL ) ET ai ua H PLdLe , I. Ed Lo hol. 5 ON LoL TR TE ly ob a GJ ETE EY Cg Lg ws. ' : co1520 BEST COPY AVAILABLE 418.014PAGE F-34 FR NA ig > Te TR, - .9 gh eT 4 x Ch 2 AW E ad lSACRREEWS f5ast pr 4 BhE T ameE nRTuk he 0 LaF ENE 208 B75 or Bion Rng ah SAMBA 0s on Ape IR allo oc A aad py 5. TR Load Rw Ny co PSLN S AoR gE SEiRlIyE. eaa %: ` P E oa REe CT B enR SA CA N RNE NEElS sC ra R td E ,oD Vy F A mJL NRE a BoeA JELIFe N2%, a E Es5 IRE AOR SNS BP ATTRan R e aeAv AryaP eel =] SN RR Ea STE Bl ET TE agEee ed Eds. TE Fe. SREY oe : sempre Joes Seng , a Ya ed SGD Reity ABT Seve : o"v,reaifleD Sh CENERSiM yE UEC Rage ERR " | 001528 BEST COPY AVAILABLE 418014PAGE F-35 ! Ca, ETw A eoe . a.e <, SE Le TIRTYER Nea, RN a SB Tica 3 Gi d ; SCL x. 3 SH{ p- Bld PENS gr ERIS igh BRITE 5 ha AG fa one SSF5 (4. Lo | nnada yyRNE T AN ESRfeAe SigRG A) FRE ENE: eR. k: y i PE RD yP A gh i SRSA. B BC SSpigk I<a L3 R PRL.ny gl SEE] EW Tamas Fr g5 arPreess OANOrNeE CR YE a oR d Ban A aL iL Xn k=, 2 oeig pSrA Ton . uTeh LE BsE +d RA A SRR R . ARR Re A EL N Sa Ro: ORO Rh IS A . )5. S IR RS R ErLa IEg LEEEC San pvEJTa 8 Ta hg ad Ba = 00152), JEST COPY AVAILABLE 418-014PAGE F-36 AESE A VSreTaRER. r. A gRaaaN "or7S of ; EBisds R Rese T i EINEY Bards + Lilie =SETA maRrRAy BR ay Se x Ft C| 5BNEWSaTESTRdERERE | ST CTR rE i tls ae Rade RC eT a EST SEEN oT ARE k AR 7 ATERYN.R, Pn Foasa. a Seg8a M BRRt d Sa Se hE a SRTLo ' A ThI FE W EIREatnae2 a - 3 ad LEDM S J esa isiolaenaPee TRhIal NEoa. S a ii TOSopHE eRSEa"Tou bed FE a PRRRe "- " 3 BEA E EaRa eRe ae 001523 BESTCOPY AVAILABLE 418.014PAGE F-37 P Ne S ERN R SES he en, TPL SE Ee Rare: 9 ay SH a = BEL Pra i ae } aa ht Bra eli BRE4 i. por av-a, lle. LR AET GeheR a, DEG Wah C 77 R Pea aeoEs Sx ane E li CREAS ol L SER ER, sel A SEALER Th Re SRO aEe Se an RUE Ne ak Bebeia1.lo SoawRea Rns l Sne akRy FRET a ab E Ec S 2NS NTC. SE ia Sl Ae Lhdd i CadaSrRanG sSeE ga ae od Ses S BEEy TEokNe E iBY a ATNaeS SR S CS AER R a pe EeSou aEE g= t Wns0 Ba ISONC SE e eS a i dToAra fi ea TySeeing a NE Beeler wR i ee SM SFasOSaCE a a cva oSSsIe MCGd Re Bes CI SEPTaka ren 2 Sy eA] wT Re a se 001524 BEST COPY AVAILABLE rrooreonrcaes "F EFl FTRERR | iBi.RAgr Ce Lo Se L S R R ASC aEgsY rR || ATSOCAs SRT hae FN oS+ "gad c= alee g hA es AYE EC y e DES ` sate= AfPEo2 dShT .i Ne ee mR Coen aEl a ER a BSc if i Rs : x ro HET4 CIR > Be an Sg SE 6 rs] ia - ar Ri L eR 0 SLE = RN Ry ( ts rT BEARS 4 Pa Sn CAL Be 3 eg IP onea L STR LER LoTR SR 01524 BEST COPY AVAILAIE #18.016PAGE 30 BE IPT 4 RE x Re} 7 2 AF Re A Pa f re) ee, vt 6,OB En yay JER 0 EET sa hy Ra YP AO A EE Be i Vor Rs #C3e itSin A RT ER Sl He S ee Stg NPE ERANW ER) aRa ameeeops SPR SN fo Gan SEPR RL SJ A a N PEERS if a 2 LA0T0A2 Rada MD Yen SON NY AGE ARS L R R NPe E SR o EEsRRE CERL e SeC r pRE eAeR eaEe RS P5PERTORPRPRRoSdE non, Beotk IE aie ob i joe NL 2) 001528 BESTCOPY AVAILABLE "BORAGE F40 Rog DW EN Chart SATE a oY PER TO ERSg BE Ct3 d k Ph t Wx a Ss UN . Leiter 1 BJ BT ZA De iy?F5"niioyg S PR VogR T ; C R 2 s ENe n aRdA pr || a AEE RLgSe ET STE TR Sae lsSspistas g | LEEl RY i Ue. ull BREAS fee ror | | E WEEE N Tf E h SORy IE CYRAL0E ERE | le or 0152% _ BEST COPY AVAILABLE EO PAGE Fit Wik bow CEA of Ny SE ANE La od GT i 2d alFr SRARN SCEEI rl iae [EAA ORRANNE HS Bart a Mee 0 Ri TH A [I SAN ei 2a ; ion RO BR E RE NN as" y| Nei TEE HER SENG, FREYCR ShaRe R Sa oR Fa i IEE EN. : 5N 2 ER ien RR eSa 4 b p a 5 NN IE ek SE Hla N copra eet Wf 2 EE B.C we FA (Re Nr A RE lYee ITgy aYP vB3on EE) BEX C0 IR af eniy c Ey EhpARMERENITE ARES PiRbEe S so e Es wl Sa Cl e ATn~z 5EESd ge ACpEoRl (rEsde gaeBA aniaCRTN IE Ng E INERT ANRERAS G Te co152 BEST COPY AVAILABLE #1014PAGE Fiz y SA Sue a feelin RE ok Au Ti i ea aaa hdEl ge BN NL A N oh E ONG aid Eg EN SSRI hssFORSLA A oF] (EEF TREE en NN {RE Ee hos SRA A > "iS JFA LefGhETIrITER pk YE TPES NC SENEE RA RFA paRe gosh Aas ae LSCoady ERCIeY So aagsof yan 4 oun Cai TO or TEA Se Uh Ly aN a Ray Aker B ReonNTi PM e oi re An anae BEINNKRS ado oYi Ng) f a MON dR a Set EE BR 5 x + SAR RataTe a) fone ATE A = Mo Nm BREA, he Ts Si FoR ech a El Bl as LET ade ES Peters ZR To i od gr ESF 01524 BESTCOPYAVAILABLE -014:PAGE F-43 i |/ !| o i Ou CF SRN [. 3% Pi bo Er ne Se RRR CS W RN e 3 Lg oo] 2, = Ny RrE iE Sa || SEE aEeaehieRaflsFeeA. WsN, RSEeEg Thay 3 Serr Se so iy) BREEEe Cp at SH i SA { CSE EE E E TehHTS PYasS"SSeRNy WES oN CNS | Co FER TER LL A STE Shanty WN neni RE a aSeas anpli ve RS eEE R R y SRA SOpRNe yST pr uE LkNSEG a30)Rgi EiR] 9 Eine ates ! En Sp 4% ge mt Ba 2 ENT WIT 001530 BEST COPY AVAILABLE 418.014PAGE Fis Tis tats TL WY PC fpoeotteeda . Tye 24 A Garay ME] LPR SA Rally Ga i Jh yRE RN R Rk Nr aAkE e, TE Tan ge . cEBelpasPdoBRN no St E S A een b. S NOW S T iE e J es Rh : 5 I]Af ei La2i OPEERB cc A nany pier' - L iES efi SpIeCH) . hy Ta JL; a `Tee J Fo PSR Ra R nnre hst co Eiaa: iYi 5 Eg rB iw S A es gr NP CiIA EEA EER ae Q ANS, 3 Beg y NE H SOER SeJCTAa i Sp TY ARRRNA Eor = 5. k BN i KE. SE PAVE7 oe 7 cA NY - ET SOE praes) C fi 255 oR AA RW ei CR RTTL 2 ihcFe A eon cc1538 418-014:PAGE F-45 IV. QualityAssuranceStatement 001532, 77 418-014:PAGE F-46 Pathology Associates International AComaanyofScienceAppicationsimematienal Corporation == Ae omen GoresComa. nancoes 2 he US: UFfnoieotcdatsaonnrdemqDuiircrurQegoUdsbAAcdLyomIpiinyTnieYssttGruAaodtSyoiSodhnUa.LsRabAbPoaNArIsaCntEhoiranySsapPTerAucanTtcecEidctMeaoEsniNdnPTgaeugdiatesdubsy tpheruialnlyg oSfailnstpyecAtsisonusr/aanucdeitUsniatnwdhhiceh arpeostrisngdirreecptolryt0: management. Theafnaollnepnromrecord Dateof Inspection' 218,199 anes 7s 4/5/99 PhaseInspected Processing and Embedding Data Audit Oratt Report Audit Final Report Audit 1D0atMeanFiangdeimngesnRteported andStudyDirector 2109 17s 1710s 4/5/99 &ne AdsesWuarradn,ceB.SS.p,eRcQaAltP-GLP GH Sponsor: 3M CorporateToxicology Test Article: PFOS. Study Number: 418.014, PAI Stuy EM-99.45 001533 49150 Prospectus Dre + Oumar, North Carcina 27713 + (579) 544-5257 979) 543278 Fo APPENDIX G STATEMENT OF THE STUDY DIRECTOR 001534 r PRIMEDICA - 418-014:PAGE G-1 WE ooI Beier Argus Research Laboratories. Inc. ee PROTOCOL 418-014: `ORAL (GAVAGE) CROSS-FOSTERING STUDY OF PFOS IN RATS SPONSOR'S STUDY NUMBER: T-6295.13 STATEMENT OF THE STUDY DIRECTOR This final report accurately reflects the raw data obtained during the performance of the study. No deviations from the U.S. Food and Drug Administration (FDA) Good Laboratory Practice Regulations; Final Rule, the Japanese Ministry of Health and Welfare (MHW) Good Laboratory Practice Standard forSafety Studies on Drugs" and the European Economic Community (EEC) Council decision on 28 July 1989 on the acceptance by the European Economic `Community of an OECD decision/recommendation on compliance with principles of good laboratory practice occurred that affected the quality or integrity of the study. sg etesaa-99 R: nd G. York, ., DABT Date Associate Director of Research and Study Director a U.S. Food and Drug Administration. Good Laboratory Practice Regulations; Final Rule. 21 CFR Part 58. b. Japanese Ministry of Health and Welfare (1988). Good Laboratory Practice Standard for Safety Studies on Drugs, MHW Ordinance Number 21, March 26, 1997. c. European Economic Community (1989). Council decision on 28 July 1989 on the acceptance by the European Economic Community of an OECD decision/recommendation on compliance with principles of good laboratory practice. Official Journal of the European Communities: Legislation. 32(No. L 315; 28 October): 1-17. 001535 APPENDIX H QUALITY ASSURANCE UNIT FINAL REPORT STATEMENT 0153% r,PRIMEDICA TeionT Baars Argus Research Laboratories. Inc. QUALITY ASSURANCE UNIT FINAL REPORT STATEMENT Study Director: Raymond G. York, Ph.D., DABT Executive Director of Research: Mildred S. Christian, Ph.D., Fellow, ATS Protocol 418-014: Oral (Gavage) Cross-Fostering Study of PFOS in Rats `Sponsor's Study Number: T-6295.13 The draft protocol for this study was audited for adherence to U.S. Food and Drug Administration (FDA) Good Laboratory Practice Regulations, Japanese Ministry of Health and Welfare (MHW); Good Laboratory Practice Standard for Safety Studies on Drugs, and European Economic Community (1989) council decision on 28 July 1989 on the acceptance by the European Economic Community of an OECD decision/recommendation on compliance with principles of good laboratory practice on 19 OCT 98. Critical phases of this study were inspected nine times; study information and raw data were audited twice (see tables 1 and 2 for dates and phases/data). The draft final report and the raw data for this study were compared and audited for accuracy, for adherence to protocol requirements, and for adherence to U.S. Food and Drug Administration (FDA) Good Laboratory Practice Regulations, Japanese Ministry of Health and Welfare (MHW); Good Laboratory Practice Standard for Safety Studies on Drugs, and European Economic `Community (1989) council decision on 28 July 1989 on the acceptance by the European Economic Community of an OECD decision/recommendation on compliance with principles of good laboratory practice between 21 MAY 99 and 10 JUN 99, and for revisions requested by the Sponsor on 20 JUL 99, 22 JUL 99, and for finalization on 23 JUL 99. 01. 53% 418-014:PAGE H-2 This study was conducted according to U.S. Food and Drug Administration (FDA) Good Laboratory Practice Regulations, Japanese Ministry of Health and Welfare (MHW); Good Laboratory Practice Standard for Safety Studies on Drugs, and European Economic Community (1989) council decision on 28 July 1989 on the acceptance by the European Economic Community of an `OECD decision/recommendation on compliance with principles of good laboratory practice. Zoneybopluk irs Nancy J. Gongliewski Date Quality Assurance Manager tn fod 2:90 Heather L. Rabuttino, M.S. Date Quality Assurance Supervisor and Principal Auditor 001533 TABLE 1 CRITICAL PHASES INSPECTED 418-014:PAGE H-3 Test Article Administration - Gavage Date of inspection: 03 NOV 98 Date results reported to the Study Director and Management: 06 NOV 98 Test Article Preparation Date of inspection: 03 NOV 98 Date resus reported to the Study Director and Management: 06 NOV 98 Cohabitation Date of inspection: 17 DEC 98 Date results reported to the Study Director and Management: 18 DEC 98 Natural DeliverylLitter Observations Date of inspection: 06 JAN 9 Date results 19 JAN 99 reported to the Study Director and Management: Blood Collection - Dams and Pups Dates of inspection: 28 JAN 99, 28 JAN 99 Dates results reported to the Study Director and Management: 02 FEB 99, 02 FEB 99 001539 Eecal and Urine Collection 418-014:PAGE H-4 Date of inspection: 28 JAN 99 Date results reported to the Study Director and Management: 02 FEB 99 ScheduledSacrifice-Dams andPups Dates of inspection: 28 JAN 99, 28 JAN 99 Dates results reported to the Study Director and Management: 02 FEB 99, 02 FEB 99 co1s4o 418-014:PAGE H-5 TABLE 2 RAW DATA AUDIT(S) The following study information and raw data wereaudited from 26 FEB 99 to 10 MAR 99: Animal receipt,randomization,physical examination andacclimation. In-life transactionrecord. Feed consumption. Cohabitation. Natural delivery observations. LPiuttperboobdsyerwveaitgihotnss.and status. Table of random units. Milk collection data and packinglists. UBrlionoed aanndd fleicvaerl ccoolllleeccttiioonn ddaattaa aanndd ppaacckkiinngg lliissttss.. Necropsy. Tissue packing lists. Male breeder colonyrecords. `GSetnuedryamlacionmtemeenananntdcsre.elraetciovredsh.umidityreports. Temperatur Feed, water and bedding analyses. Edit requests. Dosage volumes. Data review pages. The results of this audit were reported to the Study Director and Management on 12 MAR 99. The following study information and raw data wereaudited from 26 FEB 99 to 10 MAR 99: TVeehsitcalretircelceeirpetc,epirpte,paprraetpairoantiaonndaunsde.use. Test article packing lists. The results of this. audit were reported to the Study Director and Management on 12 MAR 99. co1548