Document jgGwOpn1Y3Ko8YE9VJrqKOJR9

AfUAG- 3lf7 / 0 a*K 000152 ftW 6 - RESEARCH i Prets Research. Proven Resuns. IH3 by LCZMS/MS- Revision 2 AUTHOR Emily Decker DAm T O i t January 8,2004 SPONSOR DuPont Nyaric, DP.EO.1B9o7x145-00050USA in Serum Sciences g LABORATORY St3a0tEe5xC8yogRleleefscgeReaVrecsPheAaDr1cr6hiv8e01 ^1M >JW M BER ExM-008-276Revision 2 TOTAL NUMBER OFPAGES 20 CompanySanitized. Does not contain TSCACB1 000153 MANAGEMENT APPROVAL EVxicyegPenreRsiedseenatrch DSS^p.uMoPno.asAnrokt*r*RK*e*ep~nrn.e*s.eednytati'vfey _______ f-JfDtaaJte-ZoQij Exygen Research Page2of20 000154 TABLE OF CONTENTS TMEATNUAE.G...E...M....E..N...T....A...P...P..R...O....V...A...L..................................................................................................................................... TABLE OF CONTENTS............................................................................ LIST OF TABUES............................................................ ,.......................... LIST OF FIGURES-- .................................- ........................................... 1. SUMMARY............................................................................. -............. 2. EXPERIMENTAL COMPOUNDS..................................................... 3. CHEMICALS AND SUPPLIES.......................................................... 3 .1. Chemicals -- ..........................................................-- 3.2. 3.3. EStqaunipdmaerndtsa..n...d...S..u...p..p..l..i.e.s......................................................................................................................... 33..45.. PSorelpuatrioantsio...n..o...f...S..t..a...n.--dar.d..s...a..n...d...F..o...r..t..t.h,..c..a..t..t..o..n...S...o..l..u...t..i.o...n..s. 33..55..12.. FStoorctkifsicoaltuitoinoSno...l..u..t...i.q..n..s..--....................................................................-............ 3.5.3. Caubrahon Standards -- ................................. . 4. M44..12E..TFSHalOomwDpl.D.e..i.Pa..r.g..or...ca..em..s...s....i..n....g...................................................................................................................................................................................... 4.3. SamplePreparation................................................... . 4.4. ExiRAcnoN..................... ................................................... 4 .5 . 4Q.5o.1a.noLtCa/MtiqS/nM...S...S..y...st..e.m....a..n...d...O...p..e..r..a..t.in--g ........................ . Conditions...... 4.5.2. TuneFile Parameters...--------- --------- -----------4.5.3. CalibrationProcedures................. .................... 4.6. 4A.5c.c4e.pStaamncpleeCArnitaelryiasi..s.......................................................................................................... 5. 6. 4C4S..AA87.F.LEDPCTeUmrYLfeoARrTemIqOaunNircSee_dC_f_ro_irt_eA_r_ni_aa_.l._.y.._.s..i_.s._..._.--.._.._.._.-._.-.-._-..-_.-.-._-..-_.-._-..-. i+**t ...1 ......23 ....4 .......54 ...6 ....7 ....7 ....7 ..............,,......11..1111.......18891990000 .......,............11111111111553424512 Bxygen Rjcaeareh Page 3 of20 000155 LIST OF TABLES Table!. Recovery ofPFOA from Fortifications In Monkey Serum........... ................. 16 LIST OFFIGURES Figure 1, Calibrationcurve forPFOAin Control MonkeySerum. 17 Figure2. RPFepOrAesewntiatthiv-e5CnhgrtoomLaotofg^rCam-PoFfOaAC..o.n..t.r..o..l.M....o.n..k..e.y...S..e..r.u..m....S..a..m...p..l.e...f.o..r.. 18 Figure 3. RMeopnrkeseeynStaetriuvme Cwhirtohm~a5tonggratomLooffau1C0-pPpFbOSAta.n..d.--ard forPF.O...A..,i.n...C. .o..n..t.r.o..l.....19 Figure4. RweitphrePsFeOntAatiavnedCwhirtohm~a5tonggra/mmLoofCf ,o3nCt-rPolFMOAon..k.--ey S.e.r..u..m...F..o..r..t.if.i.e..d...a..t..1.0...p..p..b20 Rxygea Research Page 4 of20 000156 1. SUMMARY Tsehruismr.eport details a method of analysis for perfluorooctanoic add (PFOA) in PQsTpFhueOeacntcArthoiefmimciseaittcriayeolxn(ftLoorarCfmcPftMeuFdlOaS/oAMffroiPSsmF)aOacnAcsaeolrimysusmpgilsiivsuehbsneyidningbpsyrseoeclitlteqeioiucnntied2dpcorhreefrcoatihcmpitisiatoatmnotigoemrtnahoponhdiiny.to/traainncdgeetom(SnRimtrMialse)s.. Tsehreumlo.wo: limit of quantitation (LLOQ) for this method is 10 ppb for PFOA in Qinutearnntaiflisctaatniodnarids. performed using extracted calibration standard^ containing an Tst6han.i9sd%mared(tThdoaedvbilwaetaIio)s.ndfeavrelPoFpOedAuisninmgomnokenykesyersuemrumat.1T0h.1e0o0v.earnadll1p0e0rc0epnptbrewcoavse9r4y% FA1.igreuRprreeespsr2eentstoeant4itv.aetivcaelicbhrraotimonatcougrrvaemfsorfoPrFOPFAOinAminonmkeoynskeeryumseirsumshoawren isnhoFwignurien i Exygen Research Page S o f 20 000157 2. EXPERIMENTAL COMPOUNDS Tgihveenstrbueclotuwre. ofperfluorooctanoic acid (PFOA)and die internal standard are MFCFhoeOlmeAcicualalrNwadmgeht -- 4p1er4fluorooctanoic add FFFF 13C -P F O A CMhoelmeciuclaalrNwaemigeht == p41er6fluorooctanoic add, Exygen Research Page 6 of20 000158 3. CHEMICALS AND SUPPLIES 3.1. Chemicals MAAOCmhcmeeetmnhtmoiaoSnninciootairlulvli(lmWeM(AeAaOtceCeHrtNa)t)e RGHHHePPPra3agLLLdeCCCent SEEEigMMMmSoaSSS-ucccAriiiceeeldnnenrcccieeech CWAJaTAXXta9-00l70o10394g033540N---211o. 3.2. Standards Standard pplCeeiPrrgFfRlOuunAoor)reooooiccttaannooiicc add acid, Lot Number 21002 NA P(u9%r7i)ty 96.35 1 Source OPDarokodwPuoocontsdt Exygen Research Page 7 of20 0001SS 3.3. E quipm ent and Su pplies DBH2CC(2LSSd1pi2ttCaieylmmsaasaal5npp/nnpasrMLL-toendstdorm#iiesc-asAccfSaapLeaurl8a/bled,bpMgpor4Llaableelein4eDarnpSned0tbHmaeduP1radltbeooPysE7inrcqeproLt-dafrinus4-pcnoCHciai0e-andpperi0tPlvnrmincgv)pEo(Li.tudaog)ewrewCiaillutnfsus-kruatpmidsm(irag(lyt5dgacomees(0yorcuttcta-erlauteaa1uidmhrcbtnt0mut#brefls0taeilnsoduta5aetsgL(1tsdkl4et8,esc011shmy.-0o0l3m0il0n4d-)02d0e10e0rr0)gsu),L) DvruamrEiKSomMpuuVepsoypeWenssptntutdollRdpie(neorpVerrlfWierRs ) variKoNueasyHlgssPuteopnnpeeliers Assedctet|isocnri4be.5d.in Nifothtee:y cEaqnubiveaslehnotwmnatotepriraolsdumcaeysabteissfaucbtsotrityurteedsuflotsr.those specified in this method N1.otes: 2. 3. 4. 5. TrccoUIPcIHfenotoaofThcPsPulnneieioFoLnbtFtsmaarEddCrOdmmainhmtsetvAoeoiypiirnnoeicproab.tananeeoslenndsPsxasstteuabcTtsfsaadovanlrF(nebiyorse(bEdytumiltudehaittbLvroielaccdcieaCbnaHrosstcolree/n,eaPMhtpddprtneLfaiiidbopSdcmpCcgaekr/eosMisteutaannsessdsmuaSt,loeaiitteeonr.ipibtosnsepclheneaes).if)oo,pmrfmo.souldervoroltiedosteelvienfruteriuotscmsbteseaoaeqe.tul(iuiwomosqiCtnpfciuetnesmhhodt.orhaititentsaansnpbnitg1natoeoe,nmsldul)daoisnibatnefclrowduendlur.tuhesldmaeodfibnnlribuwoegetmiartopsrhPenrdehTesiiaseFstevoEesnehyl-cmuilsbgeittnaeiehokeemolnndyef 3.4. Solutions (1) 20.15mgMofamammmoonniuiummaceatacteetaatneddsiossluotlivoinngiins 1LproefpOarmedniSboylv Wwaetiegrh. ing Nmoayteb:eTphreepaafroerdemaselnotinognaeds tehxearmatpiolesiosfpdroievsidoelvdefnotrtgousiodlauntecea,raeltmeraninattaivineevdo.lumes Exygen Research Page 8 of20 000160 3.5. Preparation of Standards and Fortification Solutions Anal213y...ticLCsactiMesaonenaofaalrfmanacbsdlueiltotcoebmcyrpaenitrtrsnlxenaeatiodarrsttstSmoofr.inaoaaprltrdihtyndtminikoesakeCSoenrnsatesanosaoatnek-mnnwcuaetudosTprlnyyesarolhrederstlceiededocmsSstsrpnaofeupea~lcomiaftcekrnrorTeani.etdrxchltshtiiroirnbeaof-evgirtsneciaeeoTattrapoentyshnuisto.etatar.hhnsplnLyeeeosMdtasifaarLceboearraLsdoesrt:lrtpsOripirafxoeraiQtnceocrsaepsorpayetaviipnkorecoredenroefdysspnI.tbQaahtaryrrextoeeelsdLdupuseLsiipsnkteOeuidikccnaQetoltgolsonyrieatnenrpurvtoedorsaleeplatpmduhrraeaeeoirptneneufadiksrtetheehelaydddeet Tcmreohcareoyreradcbbetesdporpluoirntpeepotvarhtroieeoldunrmsaawoessflodsoofanlttguah.teeassttNoatnhosdeotealvprdaereslnspmtoaarartayeotiobmotenavainadantdraidieitnidocebnodayn. lctAehcneolttneraacrntneaioanlytntersat(atcairooesnnlcsaoecnnmcgturaaaryastitotehbnlyees 3.5.1. Stocksolution PpPErferuFraoferOcrmipihtgAayter)hserteabaointydncodakdrwtdiaevsietliooiudg2lftuuhepatCiirntloeogptnsota1dor0(6caoi0ntktiCmo~sn1Lao.2nl15uwd0-timiiotmshnLgsmst1aoeoXbtffh3le1a~a%nnfoo1alrlb0yi0oantitctmpalaegal1s/x0)rsia0tmail-s.nmudomLtaofrvpdPboeFelr(OuicomosAdrteoroetrarcefinctded1fdliy1an3efsCokaa-rr. 3.5.2. Fortification Solutions a. b. c. d. e. P0PPP0d033dtdaP1n...cre..eirrr0r05005leLeeeseesu.0,00.ppc0ppctp22wto555arraaiaaJ.iinnmbrrbrribrbpjpeeiegeeirehetttgLggoordadaaaf1i//aatlmmi1oi.1nceff0nfnfo0oe0fLoofLLir0ton0r33mrrotttttorieo.h.immiianf5t5frLffiefiffiiiiJ.fcvLcLtcc21i?!,roaPoac333aa..iwaCwtaCtlfaCltFtieiuiiotoaoa-o-Oiti-imtihFtPnPnonnnnhhnAeFdFnFdessaaaOstsOOt0t01rcctsata3sivaa.Ae.oAeAnGcn5t5nontatlddoof*ubdldbmnmblauanPntaayaydyrimriLFisLrrodadttdddkdrdOrnreoioii.odioilltlloAolefruefudffuifftttoicetith0ish1nini0nsfno1fn.e0ec0l.gagl.al1gjar0u10311isv3v10tb1.CkCijpo0pp0o.oei..1g-l0-ggg.ldnuP0uPfprffmmimFmiFomodirgnniOiOeLL/eLLeLLLmsttAA3rrcoofffifiLo.rccooo5osfsfifobor.rfrrftt1ltflehllhPahPuPauPodeesseFttFFtFrokiikPPoOiOoOOP..nlnFFn3FA1AAAOCO3Od0d.e0-beAA5ccAcPssyoo.ocmF1cnssnnsrrdOootoiitLttabibaallolAluuiiueiuennnwttdtdt1iiiiiiiononobnin0iitngnnngngyh0g BxygenResearch Page 9 of20 000161 3d.e5s.c2r.ibbetodi1n003.m5.L2.awaitnhda0c.e5tomnLitroilfetihnea13vCo-luPmF0etAricsofllaustkio.n described in pSbthroteeotptlsleaetorsaac)ltlkiaofstonor2,ltuiafCtfiictoeatnroti.wo6nhCiscthafnotduranaredmistaoixlsuimtnioeucnmessipsnaeraryiortedofromigfae1kraeytoenraer(wifnrso1tam2n5dt-hamredLdsLauDtseiPnoEgf 3.5.3. Calibration Standards Tiedxhetenratcicactlaioilbnrtaiostiodtnoonesetasanacdmcaoprrdldesisna.gretoTptrhooeeceffososllerotdiwftihcinarogtiuotagnbhleot:ofetehxetrasctatinodnarpdrsocbedefuorree, FoCSrMootiilftiuioxct.eiaoOdtinofn VFoorlutimficea(tpioLn) SVamColopunlmetr(eomloLf) CESCaxtloaitbnrnaredca,attoeriodfdn (tt00g000.f.0...0i111n1iL) 21500000 2to0o0 000...111 00..11 <21P25100P00000b) 4. METHOD 4TdehLsecrFifpllotoiwowndDoifaieagagrcarhmasmteopf. the method is given below, followed by a detailed Measure0.1P(mfrooLrtetoiifnMfypssearatmehmcopipdplleieFtsa(luoilt4wi*swoihnneDginnidadaigescrpsaeiogmtosnanabittlereidlm)e icropipette) LC/MS/MS analysis 4 2 . Sample Processing Nmousstabmepalellopwroecdestsoincgomispnleeteedlyedthfaowr ,seurnu-maidseadm, paltesr.ooHmowtemevpeer,raftruorzee.n Ssaammppleles Exygen Research Page 10of20 000162 storedrefrigerated should also be allowed to equilibrate to room temperature. All 4 3 . Sample Preparation a. bc.. EAAoarAceonnadcttndceoldcheltvireewateeaibnsoarostatygnrtotmaeacfnotnlohiaenomtrtoenrstciahafxotmsersnaaciabxpmtnomraldnoseptppctlplrihle(ockeea.alsercesrsbexerfmtiaoxtoerttradcnartcahiyifttciresbtetihhdelederodopasubueh(tlltrgdokayfhounnpborlkiedmcw)dea,eixnlboeldtyencraaoe3psctne0rttmceohpedoceaanertrisratdnpirltxuaeeodtrslniuecysospo)nolfnmstirocotrtrsauoottirslefevvti(qeeeimundrriiecffesyylatutht.pdorreadeoccabkoetlnvdaloneuewrakryas)n.l,t 4.4. ErntAcnoN i 1. 2. 43.. MAA(aCwanendciedtncadhatovlsryauouieznfnrrnueatetoeglis0ynuxea.tggm1ffehoofapmroslar-erLnc-1sfee1ou0oet0rosdftsminiensefidiagcitcnr,moaieculntlepltiedoeolcsescsnt.eoriaonnvtlistot-dopa1l,iurn2a4amiyn,nm(edLWg)LvCOtioo/eniMrppttmeepmSrxean/.Mnk~adelS1oes.0rxtfastcnreaecdcnoattnriroddifnsua)g.vteo0lmu.0mi0see5s (fortify 5p0g0/mpLL 4 5 . Quantitation 4.5.L LC/MS/MSSystem and Operating Conditions Instrument: KSCBISECXITEuSrAboPlIo4t0S00prBayiolmiqouleidcuIlnatrrModauscstiAonnaInlytezrefrace Computer Deli OptiPlex G X110 Software: PE Sciex Analyst 1.2 HPLC Equipment- HACeQVwouaulcltauetutrtmpiutnumPnjmeaCcDcptokoeGmargr1Gapd3sa1(1srH3e1tm1rAP3Ge)An1St3eG2ri21eA3s1161A00 ai8nSn4jsd4em/c0mt1o7Hr-TP4pw0oL0orCt) 4tsaoyrxesttreaa1mpt0ta.amcnhymedrehisynipd-leuimneeacroabnfttdearrmotpihn-eiannptgsuurtghaeradtvmcaalavryterihdaegnedisnb(teKhfeoeyrmestoothbneiel,espapmahrapt il#fe ExygeaResearch Pagali of20 000163 HCMIMnoPjooelLubbcCmiitllieeonCnPPTohhVleuaamossmleeupn(Cme:ArBae))t:::ure:25G3M0mjejenLtMhCesaiAnsomCl ms (oJnoinuems CAhcreotmateationgOramphnyi)S,o2l.v1Wmamtexr 50 mm,4jr lo033791739.....s255507e Iptemrfoarymbaencnee.cessary to adjust the %444444000000000A HPLC %111666666000000000B000 gradient in order to Row (00000000m........33355355L/minl optim1iz0e3instrument tons m onitored: 13ACPn-FFaOFlyAOteA NNeeMggaaottdiivveee Tran44si11ti53o--n44M3367o90nitored ReAt~-e2p2np..t88rioommxniiiTmnni..amtee Tmainshtoeelbrosrinlepegteeprnashtseaidostnhetheetritodmcur.iegfsDht omcruioaftntytthiinvneaurareenytsae, lntoyhtftirtioocanuagldtihramaynt-eh.tseoi-esdnaatycircbeeapastinasab,llydesewipseitanhndidningathnoeansnttaahlneydtbiacoradtlcshruaornef, Neqouteiv:aAlenntalternative LC/MS/MS system may be used once demonstrated to be 4.5.2. TuneFUeParameters Tos0tauor.fh3enlemuesmtdmaoiovlfbVaneoisdlmrsoetafishpnspPehaefFaplcOotsrtueorwAnodcemuro(afacneitttlteteeia1.o.ri0nniTTsi.pnhhtlgOu/eismn6nate0cunide%nnalf,edyouimftereisletiienhintshgeissatiraanntunnhoimtaelminlaaeylinnultseydtssiebits4oudy0tnndu%Ieupnndurefiu2dmfnmso,giptrnMhr)gtoevhuaaeiotamip-pnatmaiem1r"oaeTpinnnzg"aiteu/liidmynmostpnLoisaaa.rascanteamsdttnaredttehetaaeemrrandst 4.53. Calibration Procedures a. LInCjZecMtSt/hMeS.same aliquot (S pL) of each calibration standard into the Exygen Research Page 12 of20 000164 b. UnccUga(smotoanueasalrnnajmteiyOiblceesyberrQtbqltaanieieceunttrtmeiarieoedvalopaunxtaoefrifsclooasuetlsntskrunbtattleadteanio)eanneadrfrcdddesrahabcoatarefnayrdfrarmdt/dasonwiuslnmeyatschatdltieiyurenan.met/trtghjcevecaeostbecohlynstycueescatsdllefeadlitoinnampbabrtneprrn.eadaarqdorttaeiuiprrooxAaaedrnncngtinlatruyopcltieeetdufeasaceroastivasidnikuootletitmne.nbloai,rreununraHresesatiitaoltlnoeincwsngnsaotvteealt1ei(vsnab/eertxedarsrx.rgaauncw,tr.ssdeiddtHeoaeaiinrduentgudhsdhg2tesiaote0nftiraotn%gdOarnugalAdnutoocniadtonuflfuriradettmtvhlhoroyeeeabTsfsterbtteoarest1attthhtoiar.)o2aeeel,f c. 5Tap0hp.e9r9oc2po5rriare(trel*atsi>ote0np.9sc8o5me).uffsitcIifebnectatla(ibrk)reanftoiotrnocraaedlsijbuurlsatsttioifnnasltlrcuoumruvtesenisdtegoetphneeersraaetitoeldnim, miatsnu,dstthtbhenee 4.5.4. SampleAnalysis I a. IinntjoecthtethICe s/MamSe/aMliqSusoyts(t5empL. ) ofeach standard, sample,recovery, control, etc. b. iSntcalnuddaerddsincoanrreasnpaolyntdicinagl stcoLat least five or more concentration levels must be c. Aacssinaecanjmclteiobcefpurotnlianelotltitoinrosewfeinonste.resdattaaIosbnnasfydmeceacapcirotldalhnielbsedisrbrmaerstcateaiatoytisnooebns,fetcassianttnaaldicnnbalddurraadadrrtsedidosdssnh).aionstuttaAeltdhnrssedbpaaebernerdisgnsaeijlnedmtcenteuriennvsdagtetrabiayvtenet,dh,5the-aa1enbt0efteighsrnaeisntmtindraepiannldesgdseootlaf(fotsoaaft d. nTsrwetehasictnpehedoiscnansorastndehrcyseec,snsuhdttraroivanluuetdilt,odaenrbbdtarohcasefuceekdrsvaeacetomhrrneapssnlapteghmoseen.pisnlpeees/f5aoo0krf:t5tiahf0riecemaaretiesootiindfu/utceneoaonfclotrhrwuonaladtniesairnldyteeotteae.cgrhmivsTieanhmeaedprflseertosaspmneodtnathsrIdeef e. FthoertLifJiLca0tQio)narreeccoovnesirdieesrefdalalicncgepwtaibthlein. 85 to 115% (80 to 120% fear levels at f. Eanxatrlyascitse.d samples m ist be stored refrigerated between 2C to 6C until g. cSSoaarmmreppsllpeeossniidnninwwghhaiiccnhhalnypoteeapkreesatkeansretaiordenedtteiemctteeecdstewadti(ltilh.ebe.ecsroiegrprneoasrplt:eodnnodaiissnegNrDaatnioa(nly<otte3d:re1et)teecanttteitdoh)ne. Exygen Rew ard) Page 13 of20 00016 t(i1m0epspbbuwthaererelLesLsOthQan-1th0eplpobw)ewstilclobnecreenptorartteiodnaso<f 1th0epcpabl.ibration standards 4.6. A cceptance C r iter ia TFfhOeAfo:llowing criteria must be met to assure rite presence of PFOA and 13C- 1. TPuhFCeO-cFhAFr:oOmaAad:taouaggrdahamtuegrshimotenursaitotsn3h6ao9tw3at7mh0euaffmorolulmofwraoimnpgaarpepenaatkroesfn:4t1o3f4a1m5uamu 2. AfthoneldyLlaoLnwOaelQyr.ttehAapnrnetysheeannLtaLilnyOtmeQpe. trheosednbtliannkthsemreuasgt ebnetabtlalneka:stm5u-sftolbdelaotwleearstht a5n3. Rmlod1ime1ueet5tcetrih%otrisdmovewde(irn8aitfe0ehrosr-ier1afton2irfafte0isc-c%lercaaexebt-ipftoeroctaxnaorctbnrtldlaieteorcavontbeirloialsssnncpwogaoinketaftertrtrshsohahelenaontesLeudnpdlLdit.mkiOreeabQ)tear)infxaeao,lvlflsyiapnttliihguckeaaeitlosresudmketsntuibodsywAet bnnttheyhveembaelaaautwnctercasiexe.leypnsstApat8bink5tloyee4. Ataecopxantlpcaiyebrloercnapdaturili2moiab0ntber%aseottraiuoonotnfdlfiateshcrrtdaeatlneaitbdstorttat,ahartmedlioLnanfuLyomuOsbtnaeQbdnedm'txaoocurfldsbutssdebtaetehnadradeftsarttaorcadimotnsiusetiltdindhc.jeaeblccetuoereudvxtelcai.elnurddHbeaoydtwlmueesvauiesnsrttg,ontahnoneet 5. T2astp0hap.en9rd9oca2por5rdrirsae(tfole*arts2ittoh0ene.p9rsc8eo5lmee)v.fufasinIctftibesceneattltoia(brkfr)esanaftiomotronpclreaaesdlsijsbuuhrlsaottstuiioflndanslbtlrceuoumrurevtaesennisdatelgoyetpzhneeeedrsar.eatitoelindm,miatsnu,dstthtbhenee 4.7. Pekeqkmance Cbitehia 1tbh7e1ee0ncfouomslleomdwefnioncrgetmhtwiesnomtceortifhteaorndiaa.lymsiusswt bheenpeursfionrgmaendiansstarusmysetnetmatisounitsaebti-luitpyttheastt,hbaesfonroet fpRathinaurirdsenamtOaCcberttisraetteiirantrsnei.oadfnaseridgcnsaaonll-nutootit-onnobieosenmrLaetCito,/MooSfp/aRtitm4lSeiazcseto9rar:en1sd,pcooncmhdaipnnaggreetdoittnohseatrureemsatigemnentatteObdlpaLenrLka.tOinQIgf Exygen Research ) Page 14 of20 000166 SRrcsinetreuagictdrnreoteiirennsiaodsgainsnoCaeaintrtslioytowmersfribieistsoeht.,tlnaosGa:nawdmceanotphreedlerfesafsLitcoemLifeaOanfycQitvaboleueifbpaordntaroeatitmloefyirnrozmerecehidunicrwgavothiientoehcfsnoetsnct(rarote2nrn)aadctcoaiehorfndnatasrnaLtiantelilevtoyeaentrelssslt,pea0evns.rped9sale8ontd5obn..tibaneigOninnaacclelriuandnteiegnadesr 4.8. Time Required fo r analysis pAlarbosoceretadtouofrrey35icnosnaatpmropprlolesxspiim(k1eastre,elaaygne8dn3ht 0ocuosrnastmrbopyll,eo1s)nmecaapnterirbxseocntoa.nkterHonil,eth1LroClau/bgMohrSat/htMoerSyexsatprnaiakclytei,soi2ns 5. CALCULATIONS a. UpbeysaedkEieaqrAueanat)aioluynssit1nsgotoftthcweaalcsrteualpnardtoeagrtrdhaemcua.mrvoeu(nlitnoefaarnreaglyrteessfioounndp(rinamtlge/tmeL),gbeanseerdatoedn EAqnuaalytitoenfo1u:nd (ppb) * (AnalytePeak areaflSSlpoepaekareal - Intercept x IS cone. (ng/mL) b. FEoqruasatimonpl2estofocartlicfuieldatewtihthe pkenrocwenntaremcoouvnertsy.of analyte prior to extraction, use ERqeucoatvieorny2(:%) = i AnalvtefoundfoobAlm- oauvngt.AAnnaallyvtteeafodudneddfinppcbo)ntrol (bob! x 100% 6. SAFETY TrhplaearhbaenegoccaUreaaunintnttoisagorlynysbstsceatwofnosadhhrtaeseor,ndusphsladeafarenfnrtdoeydlraimgndrelgiaantsshggeseeernustmth,msai,as.nitnedmrciglauellotdhvsionaedfgse..twyUoUdrskeasitenbaglosuoihnndeiv-efbetusormsmfaeolerhppoaaroletlhdcosastguaaetnninoddnhawsradneswdarlahiinnneggnd Exygen Research Page 15 of20 000167 Table I. Recovery ofPFOA from Fortifications In Monkey Serum Summary fPFOARecoveriesfor10ppbFortificationinMoulceySeram HxIyDgan 000333000000888000888 SSSpppkki ACB LLLoooSttt444p000noKK>n8s777o444r000000 E000Rx555EDt///r333Laa000cAt//et/00i0To33}InVESSTT000AAA555nDNN///333aaDD0l00yt/aA0A00si33RRsDDDDAEEFVVVoI(IrEAPAti.11RPOLTT00AbIIe)OOGveNNEl::: %Recovery 219999.854621 SummaryofPFOARecoveriesfor 100ppbFortificationin MonkeySeram 000333000000E888x0I00yD888gSeSSopppkkkDEF ' LLLoooSttt444p000oHXXn8>s777o444r000000 E000Rx555EDt//r/33L3aa00cA0te/t/00i0To833InVESSTT000AAA555nD/NN//333ao0DD00lyt0//eAA0s03i333RRs DDDDEEAFVVVoIIfrE1AAt1at.Ro00*TTLo0A0>IIeOOGveNNEl::: % Recovery 1 U19999.S72203 Summary of PFOA Recoveries for 1000 ppb Fortification In Monkey Seram 00033300000E0888x00I0yD888gSSeSapppkkkGfli LLLoooSttt444pQ00IoDKKKns777o444r000000 E000Rx555EDt///r333Laa000cAte///t000iTo333InVESSTT000AAA555DnNN///333auDD000ly///eAA000si333RRsDDDDEEAFVVVoIIfE111tAAtti000RLpTT000bAe00I0I)OOvGNdNE::: 9>Recovery r 111998330145-.780 OVERALLROEVLEARTAIVLELSSTTAAONNVDDEAARRRADDLDDLEEAVVVIIEAARTTAIIOOGNNE;:: 7693S4 Bxygen Research Page 16of20 000168 Figure 1. Calibration curve for PFOA in Control Monkey Seram OKJOOaAjdb(PFOAJ:1 W Ftogralgnn/ <UM2Mt14x*0.0130393(r<(UKMSS4a) I Exygee Research Page 17 of20 000169 Figure 2. RSaepmrpesleenfotartPivFeOCAhrwomitahto-5grnagm/moLf aoCf *oCn-tProFlOMAonkey Serum Exygm Research Page IS of20 000170 ZJ(roftd Figure 3. 1 Figure 4. RFeoprtriefsieedntaatti1v0epCphbrowmitahtoPgFrOamA oanf Cd ownitthro~l M5 nognAkneyLSoefrUuCmPFOA Exygen Research I Page20 of20 000172