Document 0JZO5akZwYDkmbyEmjzvEBrmO

T ons R Comparative Molecular Biology Title: of Perfluorooctancsulfonamide (FOSA, T- 7132) in Rats following four consecutive daysof dosing. Final Report February 18, 2004 PSrtoutdoycoNlumAbmeern:dment NDuTImSber: 2. S3tMudMyedDiirceacltoDre:pAanrdtrmeenwtSSetaucdaty PNhu.mDb.er: T-7132.1 Analytical laboratories: I. Kendall B.Wallace, Ph.D. DAB. PS1rc0ohfUoenosilsvoeorrf,siMDteeydpitDcoriifnveeBiochemistry and Molecular Biology Duluth, MN 55812-2496 2 DrXinlu Departmentof Pharmacology and Physiology 6R0oc1heEsltmerw,oNodewAvYeo,rbkox14761412 OPrroilgiifnearlatSitonudiyn PRraotstoacnodlGTuiilen:eaCoPmipgasrative Molecular Biology of Peroxisome Original Study Initiated: In-Life Start Date: November 16, 1998 In-Life End Date: December 21, 1998 5 = 23 so "55 2- 30 5 TorRsIER Purpose: `(Tvheehipculrepcoosnetorfolt)hiosr 4s0tumdgy/wkags/dtaoytrpeeartftlhuroereoomcatlaenersautslftorneaamteiddew:itChFeIit7heSrOn2oNcHom2po(uFnOdSA, cPeFrOtaSiAn,toFxiOcSoAlmogiidceal,eTf7fe1c3t2s,).suTchheassthuedpaytwiacspedreosxiigsneodmetoprevoalliufaetreatthieoonmf,etthaibsocloismmpoaunndd and to compare these effects to other compounds dosed af the same concentration. Methods Test Materials: P1e2r0f0l6u7o-r1o0o8c,taG.nesMuolofroena1m0i/d2e8/(0F0O,STA-,71P3F2O)SwAa,sFiOnvSesAtmiigadtee,d.C8TFh1e7vSeOh2icNlHe2coNntBro#l was propylene glycol. Dose Groups: Three male rats received volume of mlfkg body wperiogphytl.ene glycol as the vehicle control by oral gavage at a `oTnheretehrmoaulgehrfatosurreocfetihveedstauddyo,seaonfd w4e0rmegeuFtOhaSnAiz/eKdgobnoddayywfeiivge.htA, vsiuasopreanlsgiaovnoagfe$onmgd/amyls o"TfhiFsOdSosAinign prreogpiymleentneagclhyiceovledwaascupmrueplaarteidv,eadnodseaovfol1u60memog/fk5gmalf/tkegr fwoausr asdumcicneissstievreedd.ays of of dosing. This this protocol dose was the same and is comparable as to the doseof PFOS administered effective dose levels for hepatic under amendment peroxisome | bpruotltifheercautimounlaintirvaesdoofsePFofOSFOfSoAundadimitnhiestleitreerdatuurned.eTrhtehisLpDroStocfoorl FisObSeAloiws ntohte kLnDoSw0n,for PFOS 251 mg/kg for PFOS in com oil, as pointof reference. Method ofSpecimen Collection: Tinhteof~ivIegr wpaisecresemaonvdefdlaassh-rfarpoizdelny adsirpeocstlsyibilnelaifqtuiedenuitthraongaseina.inTthareedlipvoelrsypwreorpeyldeinveided (Nalgene) containers. The containers were movedtodry ice and weighed afer all liquid nitrogen had evaporated. The tissue was stored at 709C and shipped on dry ice. Sera cUlpotttoing1,0tmhleobflboloodowdaswacsenctorlilfeucgteeddfforrom10emaicnhutaensimaatl1i1n0t0oxglgassats4eruC.m tthubeess.eraFowlaloswing. `tTrawnosfaelrirqeudottsoonfetwhetusbeersuamndsabmeplceen(t~ri0f.u7g5edmla)gawienrfeorsa1v0edmifnourtpeosssaitbl1e10m0etxagbolatit4e C. analysis TTI2S1ER Specimen Handling: sAhi1p-p2egdrianmdralyiiqcueottoofthteheanlailvyetriscaalmpllaebosratthaotriweesrleisftleadsahcfcroorzdeinngintolitqhueidlinvietrrsosgeanmpwleere identification chart below to Kendall B Wallace, Ph.D. D.A BLT. SPcrohfoeoslsoorf,MDeedpitcoifneBiUoncihveemrisisttyroyfanMdinMnoelseoctualar Biology 10 University Drive Duluth, MN 55812-2496 `LCioveArosxaimdpalseesacwteirvietyan(aPloyzoesdchbyanpdreYvaimoauszlaykipub1l9i8s6h)eadnmdeptrhootdesinfcoronPt4e5nt0 (cBornatednfto,rdLa1u9r7o6y)! in the laboratoryof University of MN. Ken Wallace Dept of Biochemistry and Molecular Biology at the Another 1-2 gram aliquotofthe liver sample was sent to: Dr. Michael Wempe Laboratory ofM. W. Anders 6D0e1paErltmmewnotoodfAPvhea,rmbaocxo7l1o1gy and Physiology Rochester, New York 14642 aDnr.alLysiinsXouf lpiveerrfsoarmmpeldetshferaomnraatlsogyifvstehniesaserasnagmeploefsfalnuodrporcoavribdoend. abTrhieefpasruenmtmaarnydofthe p`mreetvaibooulsiltyespoufbltihsehfeldumoreotchaordbson(sHawnesreendeeftealr.mi20n0e1d).in liver samples by LC-MS/MS using Liver Sample Identification Chartfor T-7132.1 [Sample#[Animal # [Species[Sex[DoseGroup | (1 TT 11 [1[iRoo7az[Rat |WJeontal | [2 [iRo07as [Rat |W Jeontol | [3 JiRo07a [Rat [TW fool | [--&|[ " RatR |W0 [40 mg0 kg/da7 y FOSA5| [5 [R074 [Rat|W 40 mgkgidayFOSA | [& [R077|" Rat|W|s0mghgidayFOSA | TorRsIER Results and Discussion Average body weights ofthe FOSA treatment group were significantly lower than the control group on days four and five (Table 1). Liver weights were not significantly different between the treated and control groups, but the liver weight as a percentage of body weight was significantly increased in the FOSA treatment group `The resouflPt45s0 content (Figure 1) and Acyl CoA oxidase activity (Figure 2) for the tlihveerrefsoarmepalehsepiantdiiccapteerdotxhiastoFmeOSprAoliifnedruacteodr tinheraetsx.pressionofthese proteins and FOSA is The concentrations of the parent compound and metabolites in livers were measured (Table 2). The data show that PFOS was the major metabolite found in the livers of rats. gAipvpernoxFiOmaStAelaynd0.3FpOeSrcAenNt-ogfltuhcuercounmiudleawtaisveiddeonsteifwiaeds apsreasmenitnoinr tmheetalbiovleirtea.s either the parent compound, FOSA, 3). The low percentage of or as the FOSA in metabolite PFOS, one day the liver and the apparent after the last dose low conversionof (Table FOSA to PFOS has also been noted in in-vitro microsomal and liver slice metabolism studies (Xu et al. 2003) which observed that FOSA was converted to PFOS in-vitro by liver slices at a low rate, but not by microsomes or cytosol . Itis noteworthy that the control animals (1R00742, 1R00743, and 1R00744) contained significant concentrations of FOSA. Furthermore, no PFOS or FOSA N-glucuronide were found in the control samples with a high backgroundof FOSA. A parallel analysis oflivers from Fischer 344 rats maintained in the University of RochesterVivariumdid not show detectable concentrations of FOSA. These results suggest that the control liver samples for this study were contaminated at some point time ex-vivo by trace quantities. soafmFpOlSesA, as no metabolism to PFOS or FOSA N-glucuronide had occurred in the control fCoin Sinaturs Report prepared by, n fr ds LNW]a. ri Andrew M. Seacat, PhD, DABT 2/sfor Date -- Di/ e 7 J Domaanmt oe alt > 2 <, : To1r3i21 FR References: pBrroatdefionrdu,ilMz.inM.g t(h1e97p6).rAiranpoifcdpraiontdpcsilenndsyeietivbeinmdeitngh.odAnfaoltBhieoqcuhaentmi7t2a,ti2o4n5o-f5m4i.crogram quantities of cHhaanrsaecnt,erKi.zaJt.iColneomfoerng,anLi.cAf,uoErloesshoenm,icMa.lsE.inabinodloJgoihcnaslonma,trHi.ce0s.. (E200n1.vCoimporunSodci:ensnpcemeciafenicd,nTqeucatnhtnioatlatoligve 35,766.70 uPsoionsgcha,Ma.urSo,y-aCnodA-Ybaamsaezdakfi,ioRo.mKe.t(i1c98a)ss.aDyetBeiromcihniamtiBoinoopfhpyesrAocxitso$m84a,l5f9a5y.a5cyl-CoA oxidase sciity ThXoyuxd,ircLoo.klyoScgetiachacylpaetSrc,TieunAo.cMr,eoBso,uctSicaunnpchpso,utl,Vfo.lo.nLa.7m,2i,adnpe.d(3A1n-4dEerAs,FM.ObSWy.E(a)2t00l3i)v.erBmiioctraonssofmoersm,actyitoonsoolf aNn-dEilheyel.-NIen2- Tonsis Tables Table 1 Biological Parameters. [OAf(ToCIumormSpceaornnasteidcvuemtMioveledncultaarofBydzoissi_ong--of PT rfiroscanssuonaT mids (BFOS-A. -7152) ) n Rais Giving | I5TTSBoy Vlg (SW).Cor oh C0) Thyd Wag 1) + |-- = Ler Wel & -- [fSamopAmni#DmosaeGrloup- [BWTZBIWL T_O[[2B72W1/_01 2B72W2/01 - 2[7B2W3001 or L2W72(q3)L70W1B]WT| W l h | aday1 day2 do ays |de aya [days oo)- "12[11RR0007744Z3]ccoonnttrrooll re 22560 972507] 6"72680 5 228604 2562601 267] 200] 229752 10087233739%%[11006827 276 10.70] 3.9% 0.5 sAo "[0as aue ari Tzeis o2m uorssaseewe0.r14 4ROUAOTghAgGSay| 250 254 2500 280 227 077 47%1078 |TROJ 07ASr 40 mgo kGidas y|| a249224453]a23 12025 81o48%l 1088 s! anaFFSogsSA A _== wmsoTooassma e T0a Ta61s 00 7m10 ae11 ms15 f0o.r0e9 0d.3e4w%io00r9s Pama Ter 021 013 Gos 002 001 0.44 001032 foTmecsotno(rn,pn, on ied,equal variance) A Palaof 0.05 was consideredsignificantly ifr TorRs IR Table 2 Liver Fluorocarbon Concentration Hepatic Concentrations of Rats Given FOSA (40 of Fluorocarbons mg/kg/day) and Fluorocarbon Metabolites in Livers Sampl#e| Anim#a| _ Sex iversample mo FOpmS) A|| FOGpSAA| gluc-uronide [2Teor wi loon [0026| 0d 132 00 oq [5Tomoorss[wooo |10200| od oma 00 ood [so Tomooras [wt rosa ["oas2| tesa 1957 am] oud [5[owoorss[| rosa |0190s| 21a 134 om ond [Not6 e: nmL=wnoot wmearsu|redwv oss |osos| aes 1639 mm 03d Torsim Table 3 Percent Fluorocarbon Dose in Liver Hepatic Percent of Dosed Fluorocarbons and Fluorocarbon Rats Given FOSA (40 mg/kg/day) for Four Days Metabolites in Livers of onal#[D(Gorsoeup [[DToosre [[imPFOS [PLRieOrS [CPFoOS/[as7.--dose -- [CFOaSArve[FrOS-- A--FTeOaRoAaIas (mg) |doseTM|(ppm) |(mg) pirvoesr (Pm) (mg) [liver (%) I r [ilRoT oTazleonr or |ooT 0l 0 C 1 S | eas || W |SHRe O0Tadleo] nrol 00r 0] 0m |_a |amn a] fE [oaliroT uras|Fos fearedes|teT sz iT sl o1 z mees7 zi osm) B[ [esli[ 6 mour/ as OO0SSAA0T lsvs7 0.arell7 s.]19li22e01s24a83sT ]]2a23.u12s6]] 00o93s55 TTm6re3T a9n5i1i1s97i 0o0s2ms lsI o T est ozs 00T s tezsvs ang ToTisR Figures Figure 1 Cytochrome P450 content in Liver ome) Total CYTP450 Content in RaFtOLSiAver Following TreatmentWith AEvreriagper:et pr En i 7 oms0 3 | EE ` oom Control gl Fos Figure 2: Lauroyl CoA oxidase activity in liver rl3] LCoA-Oxidase Activity in Rat Liver Homogenate Following `Treatment With FOSA Swagione. * i: Few Feo, Foo 7 E=M | Eh Control in Eh vahios | : Ya FOSA oo Eegg|[g111a11e711y] Ali | | Fegesns[1][]]]] L sEeeEeem|]a]]]]n]]] [le fo Zl ls Ea S$ |g 3/3/9125 9 HEY a Hapa Beo FL]TBE ea| 111] HH ree ho fee i5grlec| hd EEN. H SEEN L | PGelAe ERpiE. Sel i XL] `| El EAN Md blo ol | rela hax il [ IT TaT TmeseeeT ]TTTTTT TILeTbtTeeTtTT [[hEraeeaseeeressseae1m1ee1s]1 WEBEERsEEEHoEgHe1E7]1q1 [| enpgetmagagz1]1e]s errr iL [HeE HII TTT ree nn TTT ped : EEE er 1 5 3 RR "Re i ; TTT Ee HIN CTT ioat |B rkppps | Ela Chel | Co|mHfpSer !|| HE ! | REE ii Ai RE URr IEe| H MaemE (5 W(INHTL igl | i 155i 7g4 BdINA WE : Hl | [1 TNL BL em imITT IH IITESR ET] nmin |fees resins | i || B-felelle]Tf