Document 93bwxZ72m70L80gQ4vZdMze2e
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. AR&R6 -- 1370
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ANALYTICAL METHOD VALIDATION FOR THE DETERMINATIONF
PERFLUOROOCTANESUALRFTOINFAICTIEA,LPSOOTILASSIUM SALT (PFOS) IN
WILDLIFE INTERNATIONAL, LTD. PROTECT NUMBER: 454C-120 3M Environmental Lab Project No. 02723
AUTHORS: ReymondL Van Hove, PAD.
Wied Non Po. Jon A. MacGregor, B.S.
STUDY INITIATION DATE: February 5, 2001
STUDY COMPLETION DATE: May 2, 2003
Submited a EvirWoMnmCeonptorLisobonntor St.Pa5u3l5 MBiuhmoAsvaene55106
Wildlife International, Ltd.
E8a5o9n8,CoMmamyearncde D2r1i6v0e1
(410) 822-8600
Page 10146
002328
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Wildlife International, Ltd.
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Project Number 454C-120
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT
SPONSOR: 3M Corporation TITLE: ASnaatl(yFcOalS)MeitniAodvVialiadatSioolnfo ih Determinationofesuosoosanssulfonts, Possum
`WILDLIFE INTERNATIONAL, LTD. PROJECT NUMBER: 454C-120
3M ENVIRONMENTAL LAB PROJECT NUMBER: U2723 STUDY COMPLETION: May 27, 2003
1GooUdSThL.iasEbnosvrtiutdroyorymweaBnsrtacacoltncPderuoc(ttEeecNdtiVionAncMAoCgmIeplaCicHayEnacMe4(w05i8tC)hF1GR7o)PowadirttLha1b6ho0er,a1Dt7oolrAyluogPwruoascgttei1xc9ce8eS9pttaainnonddsaO:rdEsCsDpPurbilsicisphleedsboyf cord"aTnhecse uwibthiGoofodtheLatbeosrtastuobrsytParnaccetiocdeeSrasnodraag.e condiions ai te test ite was not determined in
STUDYDIRECTOR:
A Fayed Van Hoven, Fo.
SWeinienaeration, Ld
ATsEhols
SPONSORAPPROVAL:
Sh. r a.nd
Bca as [03
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Wildlife International, Ltd.
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Project Number454C-120
`QUALITY ASSURANCE STATEMENT
he U"SST,iEsnsvtiurdoynwmaesateaxaPrmoitneecdtifoonrAcgoempnlciyiannc4e0wCitFhRGPorotd L160i, 1b7 AougPurasctts19t8S9toaanndrdaOrydEsCsDs Ppurbilniisphlbesydf aGtoeosdhLaatboaryatoGroydiPnrgasctwiecere(rEeNpoVr/teMdCo/tChHeESMtud(y98D)iv17e)t.orThaeddaLtaebsoroafatlorlyiMnsapeactaiognsearnwmdeearuaditts anldlst:he
Aciry Matix Ferifnion DuadDar Reet Fst Repent
DATECONDUCTED: Feraey 7,200 Febuary 14m 15,2000 May 27,200
STUDY DIRDEACTTOERR:EPORMATNTEAODGEMENT:
Forayn 0 Fey 15,200 May 1,200
Fey, 00 Febery 153000 May 27,208
SuKs.Ca olmn an SeSnuisaonrQLu.aCloiltyemAasns,uBr.aAn.ce Representative
5-21-03
DATE
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Wildlife International, Ltd.
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Project Number454C:120
REPORT APPROVAL
SPONSOR: 384 Corpor ITE: Ao sp r Method Vn alduinfoheDeterminsonofPefurocsslns,Psion WILDLIFE INTERNATIONAL, LTD, PROTECTNUMBER: 45C-20 JM ENVIROLNARMPREOTENCTTNUMABELR: 2723
STUDYDIRECTOR:
RA n/a
=
MANAGEMENT:
a lilE t(pe
Ashales
I
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Wildlife International, Ltd. oe
TAOFBCOL NTENETS
EE Project Number 454-120
QualASTER cos RANAApi mmmm------------------------------
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Pri anm dSeacor nday ryStockSolutions...... wrmn---------- nemo TL
CLaMlAaitbraotBFilQoannUSktsaAanndNdarMdGasat0ixdCFUOLrVUrEBSeentrioens cummo inr s eseer 11| 22
R Methode VAWg iRnSBe HlIaPkSSd UESre er ee snecee en 122
REETENCES.....oocnsurrrsnssssen eov 484003394 EOE SSSR ESSA
14
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Project Number454C-120
"TOA F CB ONL TENE TS
=~ Continue-d
TABLES
Tabl1e MethodValidation Recoveriesfor PEOSiaNoo-TargetTestis Peat Sil....... 15
Table 2 Method Validation Recoveriesfo PFOS in Invercbrste Testing Sai...
--
APPENDICES
ADDER1 Pro88tAoTcEoBlADED...cvrcnreeiT
ON
Appendix AnalysesofPesticides, OrgaanndiMectasls in Wilde Inermtional, Lid
Appendi4x AsaMelthoydViARcONaDatl
33
Appendiz 4.1 Analytical Method Flowshart (0 he Apalysis ofPFOS in Actfcal Sol... 34
Appendix 42Typical HPLCIMS/MS Options PATE cr
Appendix 4.3 A Typical CalibCUrVeaOtrBiEDoS n ecru
36
Appendix 4.4 Example Calcfoural Repa rest enti atio ve ns Sample...........cmuwrmrmmmsssmsssssssnns 37
ADPEDGI4.x5 TonCHOofRLOWE ACEL PFOS SUBS.
3
Appendix 46 TonChroof amHight-levoel FgOSSraadm ccc
39
Appendix 47 Too Chromatogram ofa NowTarget Teresa Plant Soil Matrix Blagk 40.
Appendix 438 Ton Chromsogram of 8Ivercbrate TsingSoilMatrixBAgk vv41
Appendix 49 Too Chromsiogram ofa Low-level Noa-Terget Testa Plant Soi Fortification
Appendix 4.10 TFooorCthartioomnatS ogrofuHighp -levell Noo-Tai getTe estarPlants Sil
8)
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Wildlife International, Ltd. "7
Projet Numbes 454C-120
`TOA F CB ONL TENE TS
Continued APPENDICES
Appendix 4.1 Too Chromatogram ofa Low-level Invertebrate Testing Sol Fortification Sampl....44 Appendis 4.2 Ton Chromatogram of aHigh-level Invertebrate Testing Sol Friication Sample... 45 APPEAIxS Personnel IvOilnthveSeGdccccreercrncrcnerrrd
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Wildlife International, Ltd. 5
Project Number454-120
SUMMARY
SPONSOR: 3M Corporation TITLE: Asalyical Method Validation forthe DeterminationofPerlorooctanesufonae, Potassium
Salt (PFOS) in Artificial Soil
WILDLIFE INTERNATIONAL, LTD, PROJECT NUMBER: 454C-120
3M ENVIRONMENTAL LAB PROJECT NUMBER: U2723
TEST DATES:
ExperiSStmuadeykI(nnOttiEaCtaDio)ln:: FFeebbruraaryr6y5,, 22000011 EExxppeerriimmeennttaallTSeramrin(aBtPiAo)n:: FFeebbrruuaarryy,, 22000011
TESTSYSTEM: NooTarge Temestial lant Soil and Iversbrte Testing Soi
FORTIFIED TEST Non-Targe Temestisl
vertebrate Testing
w(eClOwNeCgEhNtT(RdAyT-IveOiNgSht:buss `PlantSoi21l0(0m00g00a31.71i)9/Kg) 11000001151)80)
Soil(m21g00a00.i@0/31Kg79) 1100000011188)0)
RESULTS:
Nontarge teresa pant soil samples and inverse tesin sil samples wer cach Vf1oa0rc0ti0ufuimemdgfin3ter1ri/pelKdigccaotenenwtaiftwohegtPe-F,wOedSgthttordebfanlsest.cthnoeTmoinensasrlammecponlntecsaenlwtercraalteiiboernxasttorifao2cn.tae0dn0g,wei1tw0h.i0mt,et1ih0a0unaoinld,
oTsifoqlPuvieEdnGtcSh(r5ion0mo%at-moeagtrrhaagpnheoyelt:isp5lt0e %qaNupAlNanOatpuodmrlaessrsysiwuepaletdcepetrdr)olaamnentdoerveyarna(alHllyPmzLeeCdaInbMpySeIrhMciSeghn)t.perRreefccooovrvemeraryniceoesf o9i8l.1y%ie(lSdDed-a32o7ve;raClVl m3e.a3n3pe%r;cNent=re12c)o.verReycoofv9er6i.e5s% (ofSPDFO=S3.6i 1;iCnvVert=eb3r.a7t3e%e;tNn=g u1e2l)ori`fRieeadgeanotnltaarkgset,tceonnestisspolfasnolsvoeint4s5,dainndvemrattebrriaxtebleasatkinsgamspaille,sw,ecroensdiesvtoiiogd ooff tioatienretrhiorgoucgohmtphoenemnettsh.oTdohleogpyrewcissiotnypoifcaillpyltchaetse pfeorrtciefnitcarteiloantsivae,aoribetaeroil marx
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Projet Number454C-120
INTRODUCTION "This study was conducted by Wildlife International, Ld. for 3M Corporation at the Wildlife International, Ltd. analytical chemistry facility in Easton, Maryland. Validation samples were fortified `with Perfluorooctanesulfonate, Potassium Salt (hereafter referred to as PFOS) and analyzed with high
`mpeerofodrmfanrcetlhiqdueitdecrhmrionamtaitoongorfapPhRymOaSsisnspoenc-ttarrogmeettrtye(HrPeLsCt/MpSl/anMsSo)iltsoae nd vinva etrhil ebpretu refeosra tmiaont gceoioe sf.a Validation samples were prepared an analyzed between February 7, 2001 and February 5, 2001, Raw
datagenerated byWildlifeInternational, Ltd. and acoopfty he finalreportarefiled under Project Number
454C-120 in archives locatedo te Wildlife Intemational, Lid. st.
OBJECTIVE `The objective ofthisstudywas to verify the performance ofa HPLC/MS/MS methodology for the
analyses ofPEOS in non-target teresaplatsil and invertebrate estingsoils.Thevalidated method will be used in supportof plaat and invertebrate toxicology tests to be conducted withthe tet substance,
EXPERIMENTAL DESIGN Noo-target teretial plant sols and invertebrate testing soils wer each fortified at fou diffrent
concentrations in triplicate and analyzed based on HPLC/MS/MS methodology developed at Wildlife
InternationaLtld,. The fortification levelsbracketed theanticipatedconcentrationsofsamples from future
aplnaanlty
zaendd
cinovnecrutrerbernattelytooxiceivtaylteusattse.
pAotreetaigaelntabnallayntkiscaalmpmletehaonddmianttreirfxerbelnacnekss.a
mplesineachmatrixwere.
A calibration curve was
generated from analysesofstandard solutions of PFOS with each seriesofsamples analyzed.
MATERIALS AND METHODS
`Thisstudywasconductedaccordingtothe procedures outlined inthe protocol, "Analytical Method.
Validation for the DeterminationofPerfluorooctanesulfonate, Potassium Salt (PFOS)inArtificial Soil"
(Appendix 1).
Test Substance The tet substance was eceived from 3M Corporation on Octaber 29, 1998, asigaed Wilde
Interastonsl, Lu. identification number 4675,and stored under ambient conditions. The test substance 002336
Wildlife International, Lid. "10
Project Number 454-120
was described ss awhite powder.Ttwas identified as FC-95 from lot umber 217, witnh expiration date ofAugast 31, 2001. Information fromthe most recent Certificate of Analysis provided by the Sponsor indicateda purty of 86.9% (Appendix 2). Reagents and Solvents
Al solveats usd i this study were HPLC grade or equivalent. Al eagens were ACS reagent grade or equivalent. NANOpurewater (equivalent to ASTM Type Il Desigaation DI193.91)wassed Io Test Systems feat
"The artificalsolusedfor validating the methodology fo invertebratetestingwas composed na atoofapproximately 10 pats sphagnum pest, 20 pats kaolin clay, and 70 parts industial quartz Sand. The dy constituents ofthesil were mixed (Soil Mixer #2) and the pHofthe dymixture was adjusted 0 55 with calcium carbonate powder prior to hydration. Afe hydration, the maistre contentofthe soil(30.5%)was determinedbyweighingsoil samplesbeforeandafter drying at 105C.
NToneTrargret esirPliansaSolil "The artifical soil useforvalidatingthemethofdo olanlt esotinggwyas composed naraoof
approximatel4ypartskaolin lay, 0pars industrialquartzsand and paris peat, Crushed limestone was added to buff the pH ofthe sol, anda slow-release frtlizer was added o provide utils essential for plant growth. A sil sample represcataive ofthat used in this studywas s1o0Agvtise Laboratories, In. in Northwood, North Dakot, for analysis of th paricle size distbuton and organi conten ofthesil. Reports forthe atestsoil charateizatisorn stored in thesechivesathe Wildlife nterationl, Li. st in Easton, Maryland. The resultsofthe most recent GLP-complint analyses performed to measure the conceonfsetlecrtedacotntiamionanntsinsolused fr non-target {emestial lat testing at Wilde Itermatona, Lid. i rested in Appendix 3. No asalyes were measured at levels that were expected to bave an impact on the study.
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Analytical Validation Procedures
Wildlife International, Ltd. conducted avalidation trial using procedures developed at Wildlife
Inteasionl, Lid. (Appeodix 4). Matrix blanks and mais: fortificationsampleswer prepared i sn target terrestrial plat sil an invertebrate testing soils. A rsgeat blank contaiingeverything except PFOS and soil was carried troughthe entre procedure foreachsoil matrix. Thesampleswer extracted
with methanol. Thesampleswerethea vacuum filtered, dilutedwithmethanadcneatorifluge,d. Dilutions
into the calibraion eageofthe HPLCMS/MS methodology were performed with solution of 50%
`methanol (HPLC grade, 99.9+%) and 50% NANOpure water. Samples were then analbyyzdieredct
injection. Concentrations of PFOS were determined by reverse-phase high performance liquid chromatography using a Hewlett-Packard Model 1100 High PerformanceLiquidChromatogrph (HPLC)
with PerkinElmer API 3000LC Mass Spesromete quipped with Perkin-ElmerTurbalonSpray on source. Chromatographic separwaerteaichoienvesd usingaKeystone BetCayscoiurl (50mm 2.0
`mm, 3-um particle size) fitted with a Keystone Javelin Cis guard column (20 mm x 2.0 mm). A method
flowchart is provided in Appendix 4.1 aad the instrument parameters are summarized in Appendix 4.2.
Primary and Secondary Stock Solutions
All primary and secondary stock preparations were adjustedforthe purity ofthe test substance (86.9%). A 10.0 mg a.i/mL primary stock solutionof PFOS in methanol was prepared by weighing
11508 g ofthe testsubstance and bringing 0 inal volumeof100 milwlithemetrhansol. Secondary V`sotolckusomleuttiodnislu(ti1o0n00r,o1m00t,he10p.r0i,m1a.r0y0,saonckd.0.m1g20 .i.0 /L)ofPFOS in methanol were preparbeyd serial
Calibration Standards and Curves
Calibrationstandardswere prepared in 50:50 methanol: NANOpure waterbyappropriatedilutions
of the 1.00 mg a.i/L stock solutionof PFOS in methanol. Calibration standardsof PFOS, ranging in concentration from 1.00 to 10.0 ug #.i/L, were analyzed with each sample set. The same and most
promineenat eposse for PFOS ws tized tomositor PFOS inal calibration adtdysaps. No amp was made 1 quantify PFOS on the bass of individual isomeric componeats. The calibration
`standard series was injected at the beginning and end ofeach run, and one standard was injected,at a
nimimum, afte very five samples.Linerregressioncqutions were geared usapears espoases Versus he respective concentrationsofthe calibration staadds. A ypical calibration curve is presated
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Prsiec:Number 454C:120
inAppendix 4.3. The concentrationof PFOS ithesamplwaes determinedbysubstitheuptkinrega:
responses into the applicable linear regression equation (Appendix 4.4). Representative ion chromatograms of low aad high calibration standards are presented in Appendices 4.5 and 4.6,
--
Matrix Blanks and Matrix Fortifications Selected 10-5 aliquots ofcach artifial oil type were forifed with the appropriate primary or
secstoocknsoludtioan orf PFyOS prepared in methazousing gus-ght syringe, The friedsoilswere allowed 0ai dry thea manually mixed prior to processing. The marx blanksfo cach oi ype were unfortifid ois.
Limit of Quantitation
"Themethod validation levels in both atifiial ol ypes were well bovethemethodcapabilities for quaaiationofth es substance, Therefore he method litofquantitation (LOG)i eachmarx wis 1.00 mg/Kgom awet-weigh bass called beprodofutcelowestcalibrationstandard0.00100 mg 2.1) and the overall dilution factorofthe max: blaak samples (1000 LIK). Fortheno-trget
terrestrial plant artificial soil matrix with reported moisture content of 15.5%, the LOQ on a dry-weight
ais ws calculated as 1.18 mg 81/Ke. Fo the invertebrate esting aia soil matrix with reported `moisture conicatof30.5%, he LOQ 01 dy-weight bass was cleulated 8 1.4 mg Ke.
Reagent and Matrix Blank Samples RESULTS A reagent blak ad threemati blank samples in cachmtxwere analyzedto determineposse
iatecerence. No interferences wee observed ai or above the respective LOQs (Tables 1 ad 2). Representative on chromatogramsofponargt erstral lat sil and invertebrate tstin sil mat blank samples are presented ia Appendices 4.7 aod 4.8, respectively.
Method Validation Samples Now-arget eretal plat sols were fried i trplicte 812.3, 118, 118 ad 130 mg/Kg
(@y-weight basis) resulin in mean recoveries of98.3, 101, 97.5, ad 96.6%, respectively (Table 1).
Represcotaive on chromatogramsof lw aad high-level fortification i non-target eresral EYER3 3.9
Wildlife International, Ltd. u
Prsect Number454C:120
are presented in Appendices 4.9 and 4.10, respectively. Invertebrate testing soils were fortified in
eipcne 12.88, 144,144 0d 1440 mg Kg (dry-weght bass) resulting fo mean esoveiesof 9.7, 100,93.1 and 95.1%, respecivey (Table 2), Representative ion chromatograms ofowadhighevel
fortifications in invertebrate testing soils are presented in Appendices 4.11 and 4.12, respectively.
"Theprecisionofthmethodology,as demandb the alysis ofplicat samples win esc forifation evel, was re erent lativeobeter, for al fouleovf reifilctisoni he inverchrae etn sil mati. With he exception of oo evel (1180 mg Kg 00 a dry-welght busi = 40%), precision i he non-arge csetvia lat mati was lotr percent lative, orbeter
concLusions Noo-argettrea plant nd vertebrate estingarf soilswere fortifiedwihPROS tole sominal concentrations of2.0, 10.0, 100d 1000 mg 8.7K 00 we-weigh basi, The spl were extracted with methanol. The samples were th vacuum flere,dite withethaandncootilfg,ed Dion tohcalibration ageofthe HPLCMSMS methowderoelesoorgmeyd with solution
50% methanol and50% NANOpure water. Sampleswere then analyzed by high performance liquid
chromatography mass spectrometry (HPLC/MS/MS). RecoveriesofPFOS innon-targetterrestrial plant
soilyiealnodveeradll meanpercent recovery on awet-wbeasiisgofht 98.1% (SD = 3.27%; CV =3.33%;
N= 12). Recoveriesof PFOSin iverehae tessoitlyiieldnedag overallmesspertrecover8y
`wet-weight
and mari
basis
black
of 96.9% (SD
sample, consi
=
st
3.61%;
ing ofu
nCfVor=i3e.d7a3%r;fNal=
s12l)s.,
Rweeargeendt belanvokfs,aotccfoienrdisoicosofsmtoploivnennetngss,
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REFERENCES 1. A'mDeIr1i3.c9a1n,SAocSieTtMyfoSreTcetsitoIinngWaatnedsMaantderEinavlisr.on1m9e9]n.taSltaTnedcahrndolSopegcyi,fiVcoalt.io1nf1o.0r1R:e4a5-g4e7n.t Water.
002341
Wildlife International, Ltd. 1s.
Pusest Numbes 454-120
Tabie1
Method Validation RecoveriesforPOS in Non-Targt Terestcal Plant Sil
TTR
Tome (454C-120)
Type
VRERT Reger Blank
Wgerinm s
Fortified
0
Measured
eur
Peco
Recovered'
Pp
MMeow smaMenm - _(mgai/Ke) -Recovery
VVIAMARR? MbaeiBllak VMABS MaiBak
oo <Suis z- - o pi
WVAASST: MaMamFFooimfoisiioonn 23273 223m0 aw1 asnome om WAS Man Forionion 237 2% wa Clow
VVAASSA MMuaxmFoFroinfiifcasiieonn 111183 112200 WAS Mun Fomifonion 114 its
wo oum sw ws om
VVMMAASSST MMaaFxoFroinfibecaatioonn 111188 uwms 5s2o amss ois VASS MamxPomibeuion 18 1s se Lim
VVVMMMAAASSSTII20 MMuoMawxxFoFForomimibibboeemmsiiiooonnn 11U188000 1l1200i25 99w 175 edaomon ses SuntSDer viS mt= A A1327
C--Foo_vr--eenoeeywvtirdveing Temw itaprngtWNolNue vS niotT& vie Rohe arvon 7ra0edtitOg0an0msag 2n1(L1OGwsve1r3mdgns e20row fhe s kctd(100pLEr)idoefd ovespcltosnls
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PrsictNumber 454-120
Tabiez
Method Validation Recoveries or PFOS in Invertebrate Testing Sol
-- em
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(@s4c120)
Type
eATe ) Pec MMeeand TMeeww
Forifid __ Messured __ Recovered! _(mgai/Ke) Recovery
'VREB-2 Reagent Blank
0
<144
-
-
-
VVMMAABB4S VAD
vMaariebBalkk Vox Blk
;00 SSpoie z-z -
-
VVWAASIHS4 MMaaFsorFiofriifacsiioonn 228888 2wn 5wi5 soaows oer VALS Mark Forifcsion 288 2% we
VVVWAAAISIIS?6 MMvaaaswiiFFFooorrriiifffcccnusiiiooonnn 111444444 j10r03 WWoo osadoeie
MVMAASSITS MMaakFForoibuesinon 114644 11336 w25s aom0e oe VASE Ma Fonifcsion 144 i oe ise
VVWVAAASSSTTZH MMuMaskFFFooonrriiitffeeesasiiioonnn 11l4d60o40130D0o1l 5wss1ra aimomow es SuniGwipoonmsi&oins 93d61w
N _o--meRo esveeSedo oe Dn iVuN,vwon Se er he Camere bin wees potSi te Tae da (e 305mg Thelimitofquantitation ror (LOQ)was1.44 in eof ar Sank le (1000LXEdied mg 2.1/Kgon adry-weightbasiscalculatedastheproductofthe oepen ss lowestcalibration
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Wildlife International, Ltd. re
Appendi1x
Protocol and Amendments.
Project Number454-120
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Project Number 454C-120
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002345
Wildlife International, Ltd.
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Project Number 454C-120
Wildlife International, Led.
a T ANALYTE ICALMETR LHEOODNVAALIFR DOATTAISONSOOSRO ATLHTEDFErToR ERMADNAATIS COONF S01.
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002346
Wildlife International, Ltd. 2
Project Number 454C:120
Wildlife International, Led.
a WA en',4 r[oiartai it pres os id FeoammgSukOmma icreryed (1LOWSP)FeOh)oi clemlidtfrofxetcroa, miyta hxT,y hMatyk.obTtpriimdadW vdsWAeeret,sL,d.arlied eramby Wiioieimla,4 natrctpoofrlseoFObSe. e byricfu rsthvtk= cpiveavsiuimytnoWikies,1. t,o 1stebin o eid ecb ofts dy 1veriloynmpcnevre roLENSmeebo orses of PROS iacl i ed Wiki mil, Ltcontep. vt asd vet.
Wht, ans!Li.EixirmsEeNlsTkArLDtEoSIeGsNwd carb ndmv. rosisnmtete vinl boeftidedr.fuWficl,samcilis1deMedo wi 1LiEMS nrimongstwial ekawdi bevsendt srliny. ifnrioineofPpEOrtbi t te. aMist.t To eleoefbriiot aswtiircheees. sReegdsdemsmttirasofwainlbesfeemexdteormead riesofmsptitciinstrisapo.hno.tTlhehacear,vriik,ranri 1of uta inoch
ite mit.
PROTOCOLNO. 45412101 5UBISH Em Lab i PecNon r E73 002347
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ldlife International, Ltd.
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Project Number 454C-120
Wildlife International, Ltd.
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PRONO:TSAOMCYOSULBS Em Larryi PoeNen erU3 002348
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Wildlife International, Ltd 2
ProjectNumber454C-120
Wildlife International, Ltd.
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PROTOCONOL: S412TNANYS5UBH Enema iborioryPe Nom ETD 002349
Wildlife International, Ltd. 5.
Project Numbes 454C-120
Wildlife International, Ltd.
FoThre SteoiclkSsboeloatonisdwith rinein msconto ith chided fronye mbes.
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002350
ildlife International, Ltd. ae
PrsietNumber 454-120
Wildlife International, Ltd.
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PROTOCONOL: 43010700NY-S3UBASH Savina LabiPr NeseUET3 002351
Wildlife International, Ltd. 5.
Project Number454-120
Wildlife International, Ltd.
a rt ootec mids ndrhsd i hls oesey i
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ect,tsccacmp,cmoengss1, evlatspetn,ssaofrwgeaagt 4 Giln,t Aidiitin dcmom.s ttyefc lyrity lhin. 7. Themeott SoyDion, sansofce isprofits,todem dnepnii eebavid,eesos., po etn ei 5. inTteeistkyl edeme iof nt rit et pti rd 1101. setbmtwperecyomeQuyApcrcesnUoe orge.i yeti wee
ke te otBs ei oeSyDich.
Pron re optCuwitbnfonroiomrnoofcogrLi. sms Sy wDieei adSonemftosc rRee.a iAo.Aervegos illlbetfeed pwrtenofdbeevrsend pyaSetnyDit1kihhere a. AL gnfpr wi ct FROTOCOLNO. SUNYS5UMSK Em Latei Pr n er 73
002352
life International, Lt
-
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Project Number 454C-120
Wildlife International, Led.
-
ty it co0e0dDLcABoOReATOiRYGPRoAdCaTIbCrES PriceSonus EPA E0uCsRtPytc1o60d1bOyRWiPkicsmoof Gnou,dLa4rr+ytPiync(FaVnMdCbTyEN 0W9I) 7A: Oorieg,Pot1iQusboe s mciUspdroree.lsastwloefcdLlerswisPaGcoekmL,aSttde Smciciil berealne spironn i ofsty lttwbhyGSodlLratoar,icTo Lre sppyeote ooteerior.RepweorehkepreesdtWdilnretWA,
tyarie, Lfo t,permetaiWoieiaioe,id fogS.cAt790 mtr.
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002353
Wildlife International, Ltd. -
ProjectNumber 454-120
Wildlife International, Ltd.
[--
Page of
AMENTODSTUMDYPEROTNOCTOL
STFEURDYTOIRTLOEC,TANAAELYSTOILCSANLAMTEETTHOOTDAVAALLIDATSAILOTN(FFOoRTRHEDAERTIECRAMASTO0I.OONF
PROTOCOL NO: Iz OMY.S5UBH SPONSOR: 3MCopeion sronsonsruov vo; vs
Jr---- FROmCTNUMBER. 34.20 EITECTIVEDATE Fama,2801
AMEPNg.eDSMNeoETagsNTeT:Pls CHANGE: TJheowri bsicomspo olecy, ls dodpo mcd TO "E Tovm ecompetofa cy, hs ar ddpe in
REASON:Ta ose m aol ont be alchinsy vi uessdpci
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002354
Wildlife International, Ltd. xe
Project Number454C-120
Wildlife International, Ltd.
ProjectNumber 454C-120
Pee tot1
AMENTODSTUMDYEPRONTOCTOL
STUDYTITLEp: ricMtootVilloS nFTDr imi OfPec
F-------- SPONSOR:MCops E----
AMENOMENTNO.2 re ---- EFFEDACTET:MaIy1V,2E03
J-- Core RaeRaids Te Sabet
Reason: ReseofrRpts posh tofeS. AHEOreM,ECh NPT;t
ag "a hondhgsSpTrvilbSeythfrem ofwis sede Te J"oPimtochesetippe iwili.Tofftr eof ibrsots Reason. --
Liah oF
2AGEMENT
DATE
[2
He 5-19-03
002355
-----
ildlife
International, Ltd. 29-
Project Number 454C-120
Appendi2x CenifoifAcnaalytsies
INTERIMCERTIFOIFACNAALYTSIES
ContrAnstcIlMLRRPeuervfoiedsbruiecon:nocPe1:CR(r9OO/S7AS/Dt0R0L0)e1aof8t2e1si7ces#: T8018
i 363i%e Twa)
[PT ir ESii taeae mman w%i|
on [Appenrance
F T EET IT 71
WhiteCrystallinePowder ____| Conforms |
"MetaD2l1..s(aMCIaaCmlgPoeMmnsS.i)o.m f[derie=eW .ima SyiEm nRiA a sRtASEfihSABeERY]L 321. 00L00o80wwriAamKn
4 Poussum
AmalSRE SEG] cum
lls aE HE 5
Fonfoesfagiieemsil Sool
EE J EEEear e LETTS es|
[ResidualSolvents(TGA)bogs %"Giogggo |
C PAe ) r |S aETR SiE [PustybyDSC_r_i_p_e_soln oii ag JC]
| oiswow NooDetected |
NotApplicable" |
2.Floods
3. BNroimriode
([EEElSliS SiAdE ]
press EI TPR
32 o 00005owmmm ml
5i5. SoNomwee EEEpSmiRmsicaeSSisERnelietoteoininnnl] 0 oeuSon hewn
5 2epTm WO (a ( EES n EapeI iEELrERR ARE| 2| l armws
3 WBA
Epa SIE EEReES 3 olownk
ANA EE TNR RISER 4 028whe
i 33 FNioonen 5 Pe
FP PoR Ror emme ea i33 sa heera Temmvimie | 3 ium
commana
ret 002356
Wildlife International, Ltd. "30-
Project Number 454C-120
CenItrNeTAEuRsItMinCLEaRTbIoFIrCaCAOtTAERoeOfreFrieAnecNeAs#:LY62S3I01S84 Dut ofLastAssis: 04310
ExpirationDate: 08/31/01
StorageContons: Frozen 10 [S------
Fo"Pourkity,= 01050%7-M(suRm ofpmeatailinmpur1i5ti3e4s,s1g.a45i%c+iLCdIiMmSpimuprueri,i0s,JE84P1%O+AIRno,rganic:
033%)
ToFulaiimpur1it0y 0Boma13l.l0e9sts -= 1836.50%9%
Potassiumisexpectedinthissaltform andisthereforenotconsideredanimpurity.
ch or isele 3pusityby DSCisgenerallynotapplicabletomaterialsof lowpurity.Noendothermwas ce{"eSunlfiaurnainottiohenosranmmthpgecleagpapweoeairnisiltosocbnle.cyoTn,voelraiteondntgrocSolOndasenrdnebenwcieldewtteiechrtepdrualstiinogrnt.heBis on eel,the SOc otconnedonEmp
STFINFAPBR.AAA "eNPToepraiiseftlolbuiorororboorpuaeocaoeapuytcomirocdplieiacdscniiedd
"Theoreticalvaluecalculationsbasedootheempiricalformuls, CyFiSOyK"(MW=538) "ThisworkwasconductedunderEPAGoodLaboratoryPracticeStandards (40CFR 160).
commana
fo 002357
c-- e ------------------------------------------------
Wildlife International, Ltd.
-
ae
Project Number 454C-120
CentreAnalytical Laboratories, Inc.
fo
Pro 0) 31852
FoxGre) 31-11022051800
INTERIM CERTIFICATE OF ANALYSIS
ContreAntLabCOrAReetirtee 6230184
[v--
er wm
mw
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war]
Sm gma ee Nt: Th OClewe clcnine4 rc
fromtheC6 andC8 standard carves.
my
SEpi Lorin
a iieeu cee ic Ltrse2ly
fre
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002358
r --r ------------------------------------
Wildlife International, Ltd. ae
Project Number 454-120
Appendix
AnalysesofPesticides, Organics and Metals in Wildlife Intemational, Ltd. Greenhouse Soil"
Component
TTeroore pGrreosot DeEirtbioar EBueiesodsotnuintinpisc EBrainden aftiicn sLHiienpcadcoioirct) Heiamshora-- nes rMarorryir
Tews
`Sodium Caleivm
IFKroownnsoin CBuMeraenitgoonens Co
[r-- MeadConcmion Comprar
Goollmugihss SColoelne SQoollomgehtse SCoolinnes Gooloignhges GCoolimngghsy GCSoooillnmas SGooimnges GSoolmmgaee
ek
60 mg/kg. 4,700 mg/kg
sa6,Dm10oa0oanmgiin/keeeg SCoougnhsa songs
CCollrotmevroiprter DCilenporsei DiPsenonnie MVeidaosin GnTo PFuuttiinormet SFairoemen rrroaaonmm rrraaiiinn rai eas Nea
Copper Zine
pMCoruliayaebiedveenum SeMmacian
Meant Comenion a pon p0r0y1aegd Poorl aigni NP ootoDtnerahisnd 000011ntetes o0o1t mmagtae SSoolol ormaaettes Soooooltrametsie Sot mrs
Goma
<omgg 6.1 mphkg
<SSSlos2aommagekasgg otmosee
hos perormed TNONein Foodavian pecll on Oe 4 15,199.
002359
Wildlife International, Ltd. sie
Appendi4s Aasyical Method Validation Data
Pret Number 454C:120
002360
Wildlife International, Ltd e
Projet Number 454C-120
Appendix 4.1
Analytical Method Flowchartforthe Analysis of PFOS in Artificial Soil
1W0e1i-g0h2 Fapeparcohx-imsaqeuleyb1o0.t0sg.raFmosromfastiilxpefrrsiacmtpileontosaampalre,dfwerigyhcbaocsh,saRmepcloerdwiwtehigthhtes,apTprranospfreri volume ofPROS stock soltion i methanorl wilgh gda-1ghmt sryringe. Allow tai dry. The max For cach sil sample, measure 100 miltersofmethanol wit graduate cylinder ad rans no the French-square botatle.forAtdhde s1i0l0 msialmlpillietse.rsTohfemeatdhdaiyntoonlalinstoamapnleemipthyeFrreenacghe-nstqbulaarnek.bottle and process ap botes and place ona shake ale.appArlolsoiwmatthelsya2m3p0lepsmt.o shake fo minimum of30 ious at Vacuum filter with qualitative filter paper and ine4retained soil thre ines with methanol nto filrate. Transfer the lt oto a 200-mL voluvmeetrri!c Treapseka4tanvderrsiinognso. volume with methanol. Mix well wit Tracnsifenr atpparovsiiamaatnldyctpw.entCyenmtirliftuegressoufmspclehifsoarmapplperonxiomateslpyafrvseemgliasessceati2f0g0e0 tpurb.e or Dilute samples nt te calbraion rangofhe PROS LC/MS meodolgy: Paral illClassA vvoolluummeetorficsafmlapslkes wwiithh diglaut-iiognhstolsvyernitng(e5a0n%dmberthaagnool:v5ol0u%'meNAwiNtOhpluurteionwastoelrv)e.nt.AdPdroapcpersoparitatxe
`blank samples using the same dilution and aliquot volumes as for the lowest fortification level,
Ampulate samples and submi1t or HPLCIMS/MS analysis
002361
Wildlife International, Ltd. 235+
Projet Number 454C-120
Appendix 4.2
INSTRUMENT:
Typical HPLC/MS/MS Operational Parameters.
Holt PackardMode 1100HighPeformance LiquidChromalograph `Prweeiartckhtiiano-nPEmelormnkeiirtno-TrEuilrnmbgeor(loMAnPRSIMpr)3a0ym0oi0doenMassosurcSep,ectOrpoemrestieerdeqinuimpupeldilweithfoan
ANALYTICAL COLUMN: Keystone Betasi Cs column (0mmx 2 mnLD. um pice size)
GUARD COLUMN:
Keystone Javelin Cis column (20 mm x 2 mm LD.)
OVEN TEMPERATURE:
STOP TIME: FLOW RATE: MOBILE PHASE:
INIECTION VOLUME:
PFOS RETENTION TIME:
PFOS MONITORED MASS:
40C
500 minutes 250pmLioute 70.0% Methanol:
30.0% NANOpure Water containing 0.1% Formic Acid
too
Approximately 4.0 minutes
499.0amu 99.1 amu
002362
TO
Wildlife International, Ltd.
:
-36-
Project Number454C-120
Appendix 43 ATypicalCalibCurrvae ftoxiPoFOnS
a5ne4s55.10200.1cNoos r 7l0S0r
.
Sf e atA kanoo
1000 000m sano sano an 200
ko 1.00 | 2.00 3.00 4.00 Cone br 5.00 6.00 7.00 9.00 9.00 10.00
002363
Wildlife International, Ltd. a.
Projet Number 454C-120
Appendix 44 Example Calculations or a Represeotative Sample
Sample number 454C-120-VMAS-1, nominal concentration of 200 mg &./Kg on 8wet-weight bass plant soil YPnetakerAereepat==2764311.701 Slope = 1194438 Primary nDiilautlioWne:ight Wy = 100g SecondarOryiDgiilnuatlioFni:nal Volume (V;)= 200 mL.
Initial Volume (V)) = 0.200 mL Final Volume (V9 = 10.0 mL.
`Overall DilutionFactor(Vy/W, x VV:) = 1000 ml/g = 1000 LKKg
1 _ (peaks - yinteroep PFOS at instrument (mg a.i/L) = ated itercepy
eauT.i7gaa0 0001978 mgaiL PROS in sample (mg 1.Ke) = PFOS at instrumet (mg .1/L) x overal dilution futor (UKE) PinFsamplS e(mg arykg)- OSULIE BEmga, 11000000LL 197 mgaine
PercentofNomin. al Conccoraio=nFESBS (Bng eaiL1innsa)msamleple
11897 0 100-587
Instrument Software: MacQuan, version 1.6.
002364
Wildlife International, Ltd. 8e
Appendis 45
Ton Chromatogram ofaLow-level PFOS Standard
Projeet Number454C-120
10 intensity: 10000 cps
a
7
5
s
4
a
2:
201
ozT O4h8 Efh iB1h2h6 8Wh177 aW28h08 AeWdt 2P51hSiamn
Nominal concentration: 0.00100mg aL
002365
--re ----------------------------
Wildlife International, Ltd.
9.
Appendix46 on Chromatogram ofa High-level PEOS Standard
ProjectNumbes 454.120
100
I2n4t0osiy: 10000 cps
90
i
8700
ii
so
i
50
th
3400
ikiil)
po
0 Bf
biole si eo syst _tea/ a\N. AGEN, one
ToLh TGe)h TBLH 1P81h 2F51b 2UW41 2V8)hiSeman
Nominalconcentration: 001mg0ai/0L
002366
Wildlife International, Ltd. ~0-
ProjectNumber454C-120
Appendix 47
Ion Chromatogramof Non-Target Terrestrial Plant Soil Matrix Blank
: | 10
o s 7 o 5 5 a " ole
Tohey8f2he
intensity: 10000 cps 1TU1u9kh 1s0A_WTh 1872X821b11 2"U4214L 22W88)0hiSamn
Sample umber 454C-120-VMAB-1 Thearrow indicatestespproximte retain imeofPEOS.
002367
r -- r--------------------
Wildlife International, Ltd.
.
ae
Pusiect Number454C-120
Appendix 48 on Chromatogram ofanInvertebrate Testing Soil Mati Blaak
intensity: 10000cps
: | 10
o o 7 o 5 a a ' o 34C0sT64T3106TZTon 151481ho74 2B82a11 22G4de2s 2A1hiSTiamen
Sample umber 454C-120-VMAB-, The arow indicates the approximate retention time ofPOS.
002368
E-- r --------------------------------------
Wildlife International, Ltd. a
Project Number454-120
Appendix 49 Ton Chromatogramof Low-level Non-Targe Teresa lat Soil Fortification Sample
intensiy: 10000 cps
10
o
o
7
o
5
.
a 2
240
1o
TEL
10391183181167
210
Te)
.
241
228013 San
0.69 136 203 270 337 404 471 Time
GSampile nfalucmtobre=ur41504o0C0-1K2n0g-.VMAS-1, 2.00mg Kgwe-weight nominalconceotraton, overall
002369
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Wildlife International, Ltd.
ae
Project Number 454C-120
Appendix 4.10 Ton Chromatogramof aHighlevel Non-Target Terestial lant Soil Fofcaion Sample
100 90 80 70
60
50
40
30
20
10
ols ss
4G160
200
7808 138 183/\
8113s 12201a 126710 230317
intensity: 10000 cps
242
i
i
fl
fil
A
AN
244014
248711 STciamne
`Samplenumber454C-120-VMAS-10,1000mg i/Kgwet-weight nominalconcentration,overal dilution
fa=c 200t 000oL/r Kg.
002370
--ee-------- ee ----------------
Wildlife International, Ltd. aa
ProjectNumber 454C-120
Appendix 4.11
Ton Chromatogramof Low-level aversbrute Testing Soi Fortification Sample
10 intensity: 10000 cps
o
7
o
5
a 2
22
+
o 20 TGTeo86 aSm 1HBoa139 1f81r17o1 2B021y4 2UW41 2W81heSaen
`Sfaamcptlo=ernu1m0b0e0r 4K5g4.C-120-VMAS-13,2.00mg a.i/Kgwet-weight nominalconcentration,overalldilution
voL371
Wildlife International, Ltd. ase
Proiee Number 454C-120
Appendix 412 oo ChromaotfoaHigghr-laevmel averebrte Testing Soil Fortification Suple
inensty: 10000 cps 100 90 a 70
s6o
)242
"
30
20
A
Tlo in se rr ie ies A2MEg,\
T th LTh EFLh ETLh B3T 2Udh aWhSiemn
Sample number454C-120-VMAS-22, 1000mg Kiwet-weghtnomial concentration, overaldition
fa=c 200t 000oL/r Kg.
VUE rg
J
-----------mGi-e
Wildlife International, Ltd.
:
-46-
Project Number 454C:120
Appendix 5 Personnel Involved in the Study
manaTgheemefoolflohwiisngsukde:y Wildife Internationa, Ltd. personcel were invalved in the conduct or
1. Willd B. Nixon, Ph.D, Director, Analytical Chemistry 23.. RJoanyMmacoGnrLde.goVra,nBHSo.veScni,enPht.iDs.,t Scientist
4. FrJ.a Lezn ottek ,B.S.,Chemist
002373
