Document 65RO12RJ4RgGZ4g8D1OmZg5KE

3M ENVIRONMFNTAL LABoRAToRy Bw copoyfod inal Tew oLffIz@el lnftw DaW- AUTHOD DFTERAGNATION OF PERFLUOROOCTANE STILFONATF, (PFOS), PERFLUOROOCTANF, SuLFoNYLAmIDE (PFOSA), AND PERmuoRoocTANoATE (POAA) IN WATER BY LIQUID- SOLiiDExTRAcTioN AND,HiGH-PERFoRmANCE LiQuiD CHRomAToGRAPHyfrANDEm MASs SPECTROMETRY (HPLC/MSIMS) Method Number: ETS-8-154.0 Adoption Date: Author: KristenJ.Hansen/Harold 0. Johnson Revision Date: Approved By: William K. Reagen, Kent R. Lindstrom WilliamK. Reagen, LaboratoryManagement Date 'K?istenJ.Hansen, Ph.D., Group Leader Kent R..Lindstrom, Technical Reviewer Date oqlzglc)o Date MS Word 97 ETS-9-154.0 Page I of 17 DetemiinatioonfPFOS, PFOSA, POAA inWaterby Liquid-SodliExtractioannd LC/MS/MS 1.0 SCOPE AND APPLicATioN 1.1 This method providescollectione,xtractiona,nd analyticaplroceduresforthe determinatioonf Perfluorooctanseulfonate(PFOS), PerfluorooctanSeuffonylamide (PFOSA), and PerfluorooctanoatOe?OAA) ingroundwater,surfacewater,and dfinldng water samples. 1.2 This method was preparedaccordingtotheEPA document,"Guidelinesand Format for Methods to be Proposed at40 CFR Part 136 or Part141" (seeReference18.1),and is based inparton thereport"Method of AnalysisfortheDeterminationofPerfluorooctane sulfonateO?FOS),Perfluorooctanseulfonylaraid(ePFOSA), and Perfluorooctanoate (POAA) inWatee'(seeReference18.2). 2.0 SUMMARYOFMETHOD 2.1 Water samples arecollectedfrom a siteof interesatnd shippedcoldto an analytical facilitPyF.OS,,PFOSA, and POAA are extractefdrom 40mL water samples usingC,, solidphase extractio(nSPE) cartridgesT.he compounds areelutedfrom the C,,cartridge, usingmethanol.Separationi,dentificatioann,d measurement areaccomplishedby highperformanceliquidchromatography/tandemmass spectrometry(B:PLC/MS/MS) analysis usingmultipleresponsemonitoring(N4RM). The concentratioonfeach identifiecdomponent ismeasured by comparing theMS responseof thequantitatioinonproduced by thatcompound totheMS responseof the quantitatioinonproduced by the same compound in an extractedcalibratiosntandard (externasltandard). 3.0 DV-MITIONS 3.1 AnalyticalSample-A portionof an extracteLdaboratorysample preparedforanalysis. 3.2 CalibrationStandard-A solutionpreparedfrom theWorking Standard(WS) and extractedaccordingto thismethod. The calibratiosntandardsolutionsareused to calibratteheinstrumentresponsewith respectto analyteconcentration. 3.3 Duplicate Sample (DS@--A separatealiquotof a sample,takenin theanalytical laboratoryand analyzedseparatelwyith identicaplroceduresA.nalysisofDSs compared tothatofthefirsatliquotgive a measure oftheprecisionassociatewdith laboratory procedures,but not with sgmple collectionp,reservationo,r storageprocedures. 3.4 FieldBlank Control Sample (FB@-Type Iwaterplacedin a sample containerin the laboratoryand treatedas a sample in allrespectsi,ncludingexposureto sampling site conditionss,toragep,reservationand allanalyticaplroceduresT.he purpose oftheFB isto determineiftestsubstancesor otherinterferenceasrepresentin thefieldenvironment. MS Word 97 ETS-8-154.0 Page 2 of 17 Determinationof PFOS, PFOSA, POAA in Water by Liquid-SoliEdxtractionand LC/MS/MS 3.5 3.6 3.7 3.8 3.9 3.10 3.11 3.12 FieldDuplicate(FD)-A sampldcollecteidnduplicataetthesame time asthesample and placedunder identicaclircumstancesand treatedexactlythesame throughoutfield and laboratorpyrocedures.AnalysisofFD compared tothatofthefirsstwnple givesa measure of theprecisionassociatedwith sxnple collectionp,reservatioannd storage,as well aswith laboratorpyrocedures. FieldMatrix Spike (FMS)-A sample collecteidnduplicatteowhich known quantities ofthe targetanalytesareadded inthefieldatthetime of sample collectionT.he FMS shouldbe spikedat'approximatel5y0-150% oftheexpectedanalyteconcentratioinnthe sample.The FMS isanalyzedtoascertaiinfany matrixeffectsi,nterferenceosr, stability issuesmay complicatetheinterpretatiofnthe sample analysis. FieldSpike ControlSample (FSCS)-An aliquootftypeIwatertowhich known quantitieosfthetargetanalytesareadded inthefieldatthetime ofsample collectio(nat an appropriatceoncentratiotnobe determinedby theprojectlead).The FSCS isextracted and analyzedexactlylikea sample todeterminewhethera lossof analytecould be attributetdo sample storageand/orshipment. Laboratory Control Sample (LCS)-An aliquotof typeIwaterto which known quantitieosf thetargetanalytesareadded inthelaboratoryT.wo levelsareincluded,one attheLOQ (approx.25Pg/mL), theotherata concentratioonfapprox.100-250PS/mL or anotherconcentratiotno be determinedby theprojectlead.The LCS isextractedand analyzedexactlylikea laboratorsyample todeterminewhetherthemethodology isin control,and whether thelaboratoryiscapableofmaking accuratemeasurements atthe requiredmethod detectionlimitand higher. Laboratory Sample-A portionof a sample receivedfrom thefieldfortesting. Limit ofDetection(LOD)-The lowestconcentratioonf an analytethatcan be measured and reportedwith 99% confidencethattheanalyteconcentratioinsgreaterthanzero.The LOD can be determinedinseveralways, includingsignal-to-noirsaetioand statistical calculations. Limit of Quantitation(LOQ)-The lowestconcentratio(nLLOQ) or highest concentratio(nULOQ) thatcan be reliablaychievedwithinthespecifieldimitsof precisionand accuracyduringroutineoperatingconditions. Note:The LLOQ isgenerally5-10 timestheLOD. Formany analytest,heLLOQ analyte concentratioinsselectedasthelowestnon-zerostandardinthecalibratiocnurve.However, it may be nominallychosen widiinthesestatedguidelinetsosimplifydatareportingS.ample LLOQs arematrix-dependent. Matrix Spike (MS)-An aliquootf a sample,towhichknown quantities'toafrget analytesareadded inthe laboratoryT.he MS isextractedand analyzedexactlylikea laboratorysmnple to determinewhether thesample matrixcontributebsiasto the analyticarlesultsT.he background concentrationosfthe analytesinthesample matrix must be determinedin a separatealiquotand themeasured valuesin theMS correctedfor background concentrations. MS Word 97 ETS-8-154.0 Page 3 of 17 DetemiinatioonfPFOS, PFOSA, POAA inWaterby Liquid-SohdExtractioanndLC/MS/MS 3.13 Method Blank-An aliquotof type I water thatistreatedexactlylikea laboratory sample includingexposureto.allglassware,equipment,solventsr,eagents,internal standardsa,nd surrogatetshatareused with otherlaboratorysamples.The method blank isused todetermineiftestsubstancesor otherinterferenceasrepresentin thelaboratory environment,thereagents,or theapparatus. 3.14 Method DetectionLimit (MDL) Determination--One of severalprocessesthatmay be used toestablisahLOD value.The statisticaclallyculatemdinimum amount of an analyte thatcan be measured with 99% confidencethatthereportedvalueisgreaterthan zero. Thisterm isusuall'yassociatewdiththeEPA definitioinn40 CFR Part136 Appendix B. 3.15 Sample-A sample isa smallportioncollectefdrom a largerquantityofmaterial intendedtorepresenttheoriginalsource material. 3.16 Spiking Stock Standard (SSS)-A solutionpreparedfrom stockstandardsused to preparetheworking standard. 3.17 Stock Standard (SS)-A concentratedsolutionof a singleanalytepreparedin the laboratorwyith an assayedreferencecompound. 3.18 Working Standard (WS)-A solutionof severalanalytepsreparedinthelaboratory from SSs and dilutedas needed to preparecalibratiosntandardsand otherrequired analytesolutions. 4.0 WARNINGS AND CAUNONS 4.1 Health and SafetyWarnings 4.1.1 The acuteand chronictoxicityof thestandardsforthismethod have not been preciselydetermined;however, each shouldbe treatedas apotentialhealth hazard. 4.1.2 Unknown samplesmay containhigh concentrationosfvolatilteoxiccompounds. Sample containersshouldbe opened in a hood and handledwithglovesto preventexposure. 4.1.3 The laboratoriysresponsibleformaintaininga safework environmentand a currentawarenessof localregulationrsegardingthehandlingof the chemicals used inthismethod.A referencefileofmaterialsafetydatasheets(MSDS) shouldbe availableto allpersonnelinvolvedintheseanalyses. 5.0 INTERFERENCES 5.1 During extractioannd analysism,ajor potentialcontaminantsourcesarereagentsand liquid-soleixdtractiodnevices. 5.2 All materialsused inthe analysesshallbe demonstratedtobe freefrom interferences under conditionosf analysisby runningmethod blanks. 5.3 Tefloe containingmaterials(e.g.caps,wash bottlesc)ontainfluoro'compoundswhich may causeinterferenceasnd shouldnot be used duringcollections,torage,extractiono,r analysisof thesamples. MS Word 97 ETS-8-154.0 Page 4 of 17 DeterminationofPFOS, PFOSA, POAA inWater by Liqiiid-SolEixdtractionand LC/MS/MS 6.0 EQuipmENT, SUPPLIES, AND MATERIALS Note: Brand names, suppliers,and partnumbers are forillustratipvuerposes only.Equivalent performance may be achievedusing apparatusand materialsotherthanthose specifiedhere,but demonstrationof equivalentperformance thatmeets the requirementsof thismethod isthe responsibilitoyf the laboratory. 6.1 Sampling Equ,.*pment . 6.1.1 Sample collectionbottles-LDPE (e.g.,NalgenenA) narrow-mouth bottleswith screw cap. Note: Do not use Teflonbottlesor Teflon linedcaps. 6.1.2 Coolers forsample shipment. 6.1.3 Icefor sample shipment. 6.1.4 Bottlesmust be lot-certifiteod be freeof artifactbsy running Method according to thismethod. blanks 6.2 Laboratory Equipment (Extraction and Analytical) 6.2.1 Balance, analytical(displayat least0.0001g), Mettler. 6.2.2 Vacuum pump, Bdchi. 6.2.3 Visiprep vacuum manifold, Supelco. 6.2.4 Sep Pak Vac 6cc (1g)tC,,cartridges(part# WAT 036795),Waters. 6.2.5 50mL disposablepolypropylene centrifugetubes,VWR. 6.2.6 15mL disposablepolypropylene centrifugetubes,VWR. 6.2.7 Disposable micropipettes(50-IOOgL, 100-200jiL),Drummond. 6.2.8 Class A pipettesand volumetric flasks,various. 6.2.9 Hypercarb drop-inguard column (4mm) (part# 844017-400), Keystone. 6.2.10 Stand-alone drop-in guard cartridgeholder,Keystone. 6.2.11 125mL LDPE narrow-mouth bottles,Nalgene. 6.2.12 BPLC pump (LC IOAD), Shimadzu. 6.2.13 2mL clearBPLC vialIdt(cat# 5181-3400), Hewlett Packard. 6.2.14 Standard lab equipment (graduatedcylinders,disposabletubes,etc.)v,arious. 6.2-15 LC/MS/MS and BPLC systems,as described in section10.1. 6.3 Equipment Notes 6.3.1 In order to avoid contamination,the use of disposablelabware ishighly 6.3.2 recommended (tubes,pipettes,e,tc.). Teflon or Teflon-linedcontainersor equipment, includingTeflon-linedBPLC vialsor caps forthe BPLC auto sampler must not be used. 6.3.3 Type I water used during the sample and standard extractionshould be filtered through a Hypercarb guard column using A,BPLC pump. This water is referredto as "filteredtype I water",hereafterin thisreport. 6.3.4 Itisnecessaryto check the solvents(methanol) forthe presence of contaminants (especiallyPOAA) by LC/MS/MS priortouse.Certainlotnumbers have been 6.3.5 found to be unsuitableforuse. Use disposablemicropipettesor pipettesto aliquotstandard solutionsto,make calibrationstandards and matrix spikes. MS Word 97 ETS-8-154.0 Page 5 of 17 Determinationof PFOS, PFOSA, POAA in Water by Liquid-SolidExtractionand LC/MS/MS 7.0 7.1 7.2 ,7.3 7.4 REAGENTSANDSTANDARDS Note: Suppliersand catalognumbers are forillustratipvuerposes only.Equivalentperformance may be achieved using chemicals obtainedfrom othersuppliersD.o not use a lessergrade of chemical than those listed. Chemicals 7.1.1 Methanol (MeOH), BPLC grade, JT Baker, Catalog No. JT9093-2. 7.1.2 7.1.3 7.1.4 Ammonium Acetate,Reagent grade,Sigma-Aldrich,Catalog No. A-7330. Water, type I,prepared in-house. Sodium Thiosulfate,Reagent grade,JT Baker. Standards 7.2.1 Potassium perfluorooctanesulfonate(seeAttachment A, Figure 1). 7.2.2 Perfluorooetanesulfonylamide (seeAttachment A, Figure 2). 7.2.3 Ammonium perfluorooctanoate(seeAttachment A, Figure 3). Reagent Preparation 7.3.1 250mg/mL sodium thiosulfatesolution(Extraction)-Dissolve 25g of sodium thiosulfatien lOOmL reagentwater. 7.3.2 40% methanol (Extraction)-Measure 400mL methanol and adjustthe volume to I.OL with reagentwater. 7.3.3 lOOmM ammonium acetate solution (Analysis)-Weigh 7.71g of ammonium acetateand dissolve'm I.OL of reagent water.Dilute the 1OOMM solutionby a factorof 50 to make the 2mM A. ammonium acetatesolutionused formobile phase Note: Alternativevolumes may be prepared as long as theratiosof the solventto soluteratios are maintained. Spiking Stock Standard (SSS) Preparation 7.4.1 lOOgg/mL each PFOS, PFOSA, and POAA SSSs-Weigh out 10mg of analyticalstandard (corrected forpercent saltand purity-i.e., 10 mg C,F,7SOK purity 90% = 8.35mg C,F,7SOI-) and diluteto IOOML with methanol in a 1OOML volumetric flask.Transferto a 125mL LDPE bottle.Prepare a separate solution foreach analyte.Store solutionsina refrigeratoart4*2C fora maximum period of 6 months from the date of preparation. 7.4.2 lpg/mL mixed SSS-Add 1.0niL each of the lOOgg/inL SSSs (from 7.4.1)to a IOOML volumetric flaskand bring up to volume with methanol. 7.4.3 O.Iftg/mL mixed SSS-Add 1O.OmL of the l.Ogg/mL-mixed solution(from 7.4.2)to a IOOML volumetric flaskand bring up to volume with methanol. 7.4.4 0.01;tg/mL mixed SSS-Add 10.0niL of the O.lgghnL-mixed solution(from 7.4.3)to a IOOML volumetric flaskand bring up to volume with methanol. 7.4.5 Storage Conditions-Store allSSSs in a refiigeratoirn 125mL LDPE bottlesat 4"-2'C fora maximum period of 3 months from the date of prepara@on. MS Word 97 ETS-8-154.0 Page 6 of 17 Determinationof PFOS, PFOSA, POAA inWater by Liquid-SohdExtractionand LC/MS/MS 7.5 CalibrationStandards 7.5.1 lOOgg/mL each PFOS, PFOSA, and POAA stockstandardsolutions-Weigh out10mg ofanalyticasltandard(correctefdorpercentsaltand purity)and dilute to 1OOML with methanol in a 1OOML volumetricflaskT.ransferto a 125mL LDPE bottleP.reparea separatesolutionforeach analyte.Storesolutionsina refrigerataotr4*2"C fora maximum periodof6 months from thedateof preparation. 7.5.2 Ittg/mL Working Standard-Add l.OmL each of the 106PLghnL SS solutions (from7.5.1)toa IOOML volumetricflaskand bringup tovolume with methanol. 7.5.3 0.1pglml Working Standard -Add IO.OniLofthe l.Ogg/mLmixed solution (from7.5.2)toa IOOML volumetricflaskand bringup tovolume with methanol. 7.5.4 O.Ollig/mLWorking Standard -Add 1O.OmL ofthe0.1gghnl mixed solution (from7.5.3)toa 1OOML volumetricflaskand bringup tovolume withmethanol. 7.5.5 StorageConditions-StoreallWSs ina refrigerat(oirn125ML LDPE bottles) at4*:L2'Cfora maximum periodof 3 months from thedateofpreparation. 7.5.6 CalibrationStandard-Prepare a minimum offivecalibratisoonlutionsin filteretdypeI wateraccordingto thefollowingtable: Concentration Volume of FinalCalibratioSntandard FinalConcentrationof ofWS, iLglni]L WS, tL Volume, nlJL CaUbration Standard,Pg/n2L 0.0 0 40 0 0.010 100 40 25 0.010 200 40 so 0.010 400 40 100 0.10 100 40 250 0.10 200 40 500 0.10 300 40 0.10 400 40 1 May be prepared to extend the range beyond 50OPg/mL. 2 May be preparedtoextendtherangebeyond 75OPghnL. 7501 10002 Note: The absolutevolumes ofthe standardsmay be variedby theanalystas longasthecorrect proportionosfsolutetosolventaremaintained. 7.5.7 The standardsareprocessedthroughtheextractiopnrocedure(Section9.0), identicatlothelaboratorysamples,The ext. ractecdoncentratioonf thecalibration standardisequalto8X theinitiacloncentrtaion,due tothe concentrationofthe standardduringtheextractionprocess. 7.5.8 StorageConditions-Store allextractecdalibratiosntandardsin ISmL polypropylenetubesat41:i:2"Cf,ora maximum periodoftwo weeks from the dateof preparation. MS Word 97 ETS-8-154.0 Page 7 of 17 DeterminationofPFOS, PFOSA, POAA inWater by Liquid-SoliEdxtractionand LC/MS/MS 8.0 SAMPLE COLLECTION@ PRESERVATION AND STORAGF, Note: Sampling equipmen@ includingautomatic samplers,must be freeof Teflon tubing, gaskets,and otherpartsthatmay leach interferinagnalytesintothe water sample. Automatic samplers thatcomposite samples over time should use refrigeratepdolypropylene sample containersifpossible.Sample bottlesshould not be rinsedbefore sample collection. 8.1 Tap Water--Open the tap and allow the system to flushuntilthe water temperature (15* IO'C) has stabilized(usuallyabout two minutes).Adj ustthe flow to about 500mL/min and collectsamples from the flowing stream. 8.2 Ground Water-Purge the well of standing water using a pump or a bailer.Collect the sample directlyfrom the pump or from the bailer. 8.3 Surface Water-When sampling from an open body of water,filtlhe sample container with water from a representativearea. 8.4 Sample Dechlorination-All samples should be iced or refrigerate-dat4'-+20C and kept inthe dark from thetime of collectionuntilextractiorRlesidualchlorineshould be reduced by adding 200gL of a 250mg/mL sodium thiosulfatseolutionto each water sample, FB, and FSCS (which may be placed in each bottlebefore leavingfor the sampling site.). 8.5 Holding Time (HT)- Resultsof the time/storagestudy of alltargetanalytesshowed thatthe threecompounds are stablefor 14 days in water samples when the samples are dechlorinatedand stored as described in section 8.4 (seealsoreference 18.3).Therefore, laboratorysamples must be extractedwithin 14 days and the extractsanalyzed within 30 days of sample collectionI.fthe HT exceeds 14 days, greatcare isused when evaluating fieldspikesto avoid misrepresentationof the sample concentration. 8.6 Field Blanks 8.6.1 -Process a Field Blank Control Sample (FB) along with each sample set(samples collectedfrom the same general sample siteat approximately the same time).At the laboratory,priorto sample collectionf,illa sample containerwith filtered type I water, seal,and ship the FB to the sampling sitealong with the empty smnple containers.Return the FB to the laboratorywith the filledsample bottles. 8.6.2 When sodium thiosulfaties added to samples, use the same procedure to preserve the FB. 8.7 Field Duplicates 8.7.1 CoRect a Field Duplicate(FD) forevery ten (10)samples collectedor per each sampling set,iflessthan 10 samples arecollected. 8.7.2 Separate FDs must be collectedforeach type of water sample (ground, tap,etc.) collected. 8.7.3 Collectthe FD immediately afterthe sample. 8.7.4 Preserve,storeand ship FD using the same procedures as used for the samples. MS Word 97 ETS-8-154.0 Page 8 of 17 DetenninationofPFOS, PFOSA, POAA inWater by Liquid-SoliEdxtractionand LC/MS/M 8.8 Field Spike Control Sample (FSCS) 8.8.1 A FieldSpike ControlSample (FSCS) must be preparedforeach sample shipment.Ifmultiplecoolersareused to shipa setof samples,each coolermust containa FSCS. 8.8.2 At thelaboratoryf,ilal sample containerwith 1OOML of typeIwater.Seal and shiptothesamplingsitealongwiththeempty s=ple containerasnd FBs. 8.8.3 When sodium thiosulfatiesadded to samples,use thesame procedureto add the same amounts to theFSCS. 8.8.4 Sealand gentlyinverttheFSCS tomix. Storeand shiptheFSCS usingthe same proceduresas used forthesamples. MS Word 97 ETS-8-154.0 Page 9 of 17 DeterminatioonfPFOS, PFOSA, POAA inWaterby Liquid-SoliEdxtractioanndLCtMS/MS 9.0 EXTRACTION PROCEDURE 9.1 ExtractionScheme 9.1.1 Allow samples to equilibratteo room temperature.Thoroughly mix samples by gentlyinvertingthesample bottle. 9.1.2 Measure 4On3.Lof sample intoSOML polypropylenecentrifugetubes (Spike the QC and Matrix spikesas required*,replacelidand mix well). Note: * Samples may need to be prescreened to determine an appropriatematrix spike level (typically50-150% of sample concentration). 9.1.3 ConditiontheC,a SPE cartridge(s1g, 6mL) by passing IOmL methanol followed by SmL filteretdypeI water (-2drop/sec)D.o not letcolumn run dry. Note: For the followingsteps,maintain a -1 drop/secflow rate.Do not allow the column to run dry at any time. 9.1.4 Load the analyticalsample onto the Cle SPE cartridge.Discard eluate. 9.1.5 9.1.6 Wash with -5n-iL40% methanol in water. Discard eluate. Elute with -5mL 100% methanol. Collect 5mL of.eluateinto graduated 15mL polypropylene centrifagetubes.This isthe targetelutionfraction(finalvolume 5mL). 9.1.7 Analyze a portion of the targetelution fractioncluentusing negative electrospray B:PLC/MS/MS (Section 10.2). Note: Samples are concentratedby a factorof eightduring the extraction;InitiaVlol = 40mL Final Vol. = 5mL. 9.1.8 Samples are stableat room tmnperature for at least24 hours. Analytical samples may be storedin a refrigeratorat 412'C untilanalysis. 9.1.9 Standardization of Cl,,SPE columns-If poor recoveries are observed, itmay be necessary to standardize the C,, SPE columns in the following manner before analyzing samples. 9.1.9.1 Use a standard with an analyte concentration between 1000 and 4000 Pg/inL. Follow the extraction scheme as outlined from steps 9.1.1 to 9.1.6, except, collectthe eluatefractionseparately (approx..5mL), as well as the 9.1.9.2 target elution fraction. After step 9.1.6,collecta post-elutionfractionby, elutingwith an additional5mL of 100% methanol. 9.1-9.3 Analyze allthreefractionsby BPLC/MS/MS. Ifthe targetfraction contains a minimum of 85% of the respective analytes,itmay be considered acceptable. 9.1'.9.4 If the wash contains significantstandard (>15%), eitherthe wash volume or percentage of MEOH should be decreased. 9.1-9.5 Ifthe post-elutionfractioncontains significantstandard (>15%), the target elution volume should be increased. MS Word 97 ETS-8-154.0 Page 10 of 17 Determinationof PFOS, PFOSA, POAA in Water by Liquid-SolidExtractionand LCIMSNI 10.0 CALIBRATION AND STANDARDIZATION (ANALYTICAL SETUP) Note:Otherinstrumentmsay be used and theequipmentandconditionmsay be very differenats longasthemethod criteriaaremet.The operatormust optimizeanddocument theequipment and settingussed. 10.1 EstablishtheLC/MS/MS system and operatingconditionesquivalentothefollowing: Mass Spec: NficromassQuattroLntima (Micromass) InterfaceE:lectMray (Mcromass) Mode: ElectrospraNyegative,MultipleResponse Monitoring(NflZK Harvard infusiopnump (HarvardInstruments)f,ortuning Computer: COMPAQ ProfessionaWlorkstationAP200 Software:Windows NT, MassLynx 3.3 I-IPLC:HewlettPackard (B?) SeriesI100 B:P Quat Pwnp I-IPVacuum Degasser B? Autosampler E:P Column Oven Note:A 4 x.10mm Hypercarbdrop-inguardcartridg(eKeystonep,art# 844017-400)isattached on-lineafterthepurgevalveandbeforethesample injectoprorttotrapanyresiduecontaminants thatmay be inthemobilephase and/orBPLC system. li?LC Column: GenesisC, (JonesChromatography),2.lmm x 50mm, 4@Lm Column Temperature:350C InjectioVnolume: 15gL Mobile Phase (A): 2mM Ammonium AcetateinfilteretdypeIwater(See 7.3.1) Mobile Phase(B):Methanol HPLC GradientProgram: Time, PercentMobile min Phase A 0.0 60 0.4 60 1.0 10 7.0 10 7.5 0 9.0 0 9.5 60 13.5 60 14.0 60 PercentMobile Phase B 40 40 90 90. 100 100 40 40 40 Flow Rate, mL/min 0.3 0.3 0.3 0.3 0.3 0.40.4 0.4 0.3 Note: OtherI-IPLCgradientsmay be used as longasthemethod criteriaaremet. MS Word 97 ETS-8-154.0 Page IIof 17 DetemiinatioonfPFOS, PFOSA, POAA inWaterby Liquid-SohdExtractioanndLC/MS/MS Itmay be necessaryto adjusttheB:PLC gradientin orderto optimizeinstrumentperformance. Colun-inswith differendtimensions(e.g.2.lnun x 30mm) and columns from different manufacturers(KeystoneBetasilCs etc.)may be used. Ions Monitored: Analyte Primary Ion Product Ion POAA' PFOS PFOSA 413.0 499.0 498.0 169.0 99.0 78.0 Approximate RetentionTime 5.0 5.2 5.8 Otherproductionsmay be chosenatthediscretioonf the analys@althoughm/z 99 issuggested forPFOS. Use ofthesuggestedprimary ion isrecommended. Retentiontimesmay vary slightly, on a day-to-&y basis,dependingon thebatchofmobile phase etc.Driftinretentiotnimes is acceptablewithinan analyticarlun,as long as the driftcontinuesthroughthe entireanalysisand the standardsareinterspersetdhroughoutthe analyticanlm. 10.2 Tune FileParameters 10.2.1 The followingvaluesareprovidedas an example.Actualvaluesmay vary from instrumenttoinstrumentA.lso,thesevaluesmay be changed from time totime in orderto optimizeforgreatestsensitivity. Analyte POAA PFOS PFOSA Dwell, sec 0.2-0.4 0.2-0.4 0.2-0.4 Collision Energy, eV 10-25 30-60 20-50 Cone, V 20-30 50-80 30-60 Source Capillary Hexapole 1 Aperture 1 Hexapole 2 SourceBlock Temp. DesolvationTemp. Set 2.56-3.5kV 0.5V 0.2V 0.8V 100-1500C 250-4000C MS Word 97 ETS-8-154.0 Page 12 of 17 DeterminatioonfPFOS, PFOSA, POAA inWaterby Liquid-SoliEdxtractioanndLC/MS/MS Analyzer LM Res I HM Res I EEnergy 1 Entrance Exit LM Res 2 HM Res 2 lenergy2 Multiplier Set 12.0-15.OV 12.0-15.OV 0.7V -2V IV 1i.ov 1i.ov i.ov 650V Gas Flows Cone Gas Desolvation Set 150L/br 70OL/hr Pressures Gas Cell Set 3.0e-3mbar 11.0 ANALYTicAL QUALrff CONTROL 11.1 AnalyticalresultsoftheFB, FMS, FD, and FSCS shouldbe evaluatedattheconclusion of thestudyto helpinterpretthedataqualityof samples data.Analyticarlesultsforthese control/duplicastaemplesmust be reportedwith thesample data. MS Word 97 ETS-8-154.0 Page 13 of 17 Deterniinatioonf PFOS, PFOSA, POAA in Water by Liquid-SoliEdxtractionand LCIMS/N4S 12.0 ANALYTICAL PROCEDURE 12.1 Sample Analysis 12.1.1 Setup analysissample queue. 12-1.2 Injecthesame aliquot(between5-25liL)of eachstandarda,nalyticaslample, recovery,controletc.intothe LC/MS/i\4S system. 12.1.3 All samplesshowing a responseforone or more analytesabove theresponseof thehighest,activecalibratiocnurve levelmust be dilutedand reanalyzed. 12.2 CalibrationCurve 12.2.1 Startinwgith thestandardoflowestconcentrationi,njecthesame sizealiquot (between10-25tiL)of each extractedcalibratiosntandardaccordingto Section 12.1and tabulatetheresponse(peakheightor area)versusthe concentrationin the standardU.se linearstandardcurvesforquantitatiogneneratedforeach analyteby linearegressiownith I/xweioifingofpeak areaversuscalibration standardconcentrationT.he correlatiocnoefficien(tr)forthecalibratiocnurves must be ;->0.99(0r'@:0.980I)f.calibratiornesultsfalloutsidetheselimitst,hen appropriatsetepsmust be takento adjustinstrumentoperationand thestandards reanalyzed. 12-2.2 Curve-The measured valueforeachcurvepointmust be within+-30% of theoreticavlalueswhen curveisevaluatedover a rangeappropriatteo the data. High or low pointsmay be deactivatedto achievethesecriteriab,ut an acceptable curvemust containatleastfiveactivecurvepoints. 12-2.3 Continuing Curve Verifleation(CCV)-Nfid- and low-levelcalibratiocnhecks shouldbe analyzedevery5-10 injectionsT.he analytelevelmeasured in the CCVs shouldbe within 30% oftheoreticavlalues.IfCCVs falloutsideofthis range,datacollectedsubsequentto thelastpassingCCV shouldnot be used. Only datacollectedbetween acceptableCCVs or theinitiaclurvecan be used. 13.0 DATA ANALYSIS AND CALCULATIONS 13.1 Calculatetheanalyticaslample (extractc)oncentratiofnrom thestandardcurveusing the followingequation: (Peakarea-intercept) ExtractConcentrationp,g/mL = (slope) 13.2 CalculatethepercentrecoveryoftheFSCS usingthefollowingequation: FSCS % rec.= (FSCS conc.,Pg/mL ) x 100 (Conc.added,Pg/mL) 13.3 CalculatethepercentrecoveryoftheMSs usingthefoiolwing equation: MS % rec.= (MS cone.,Pg/mL-SampleConc.,Pglr@L) X100 (Conc.added,Pg/mL) MS Word 97 ETS-8-154.0 Page 14 of 17 DetenninatioonfPFOS, PFOSA, POAA inWaterby Liquid-SoliEdxtractioanndLC/MS/MS 14.0 METHOD PERFORMANCE PARAMPTFRS Note:Any methodperformanceparametertshatarenotachievedmustbe considereidnthe evaluationof the data.Nonconformance to any specifiedparameters must be describedand discussedinany reportingof the data. 14.1 Linearity-Linear standardcurves forquantitationgeneratedforeach analyteby linear regressionwith 1/xweighting of peak area versus calibratiors.tandard concentration.The correlationcoefficient(r)for the calibratiocnurves must be >-0.990(r22:0.980). 14.2 Calibration Curve'Standards-The measured value foreach curve point must be within 30% of theoreticalvalues when curve isevaluatedover a range appropriateto the data. High or low pointsmay be deactivatedto achieve these criteriab,ut an acceptable curve must containat leastfiveactivecurve points. 14.3 CCV Performance-Md and low levelcalibrationchecks to be analyzed e*very5-10 injectionsT.he analytelevelmeasured in the CCVs should be within 30% of theoretical values.IfCCVs falloutsideof thisrange, data collectedsubsequent to the lastpassing CCV should not be used. Only data collectedbetween acceptableCCVs can be used. 14.4 Limit of Detection (LOD)-The lowest calibrationstandardwith a peak area at least2X the peak area of the extractionblank thatcan be measured at a concentrationgreaterthan zero. 14.5 LimitsofQuantitati(oLnOQ)-The lowerLOQ (LLOQ)isthelowesnton-zeraoctive standardinthecalibratiocnurve;thepeak areaoftheLLOQ must be atleast2X thatof theextractiobnlank.By definitiont,hemeasured valueofthe LLOQ must be within30% of the theoreticavlalue. 14.6 Matrix Spikes-Matrix spikepercentrecoveriemsust be within 30% of the spiked concentration. 14.7 SolventBlanks, Method Blanks, and Matrix Blanks-Values must be below the lowestnon-zeroactivestandardinthe calibratiocnurve.Matrixblanksareconsidered compliantifno testsubstanceisdetectedabove theLOD forthatanalyte. 14.8 ReproducibWty-Reproducibilityofthemethod isdefinedby theresultosfthematrix spikesand matrixspikeduplicatesT.he MS/MSD shouldbe reproducibletowithin20%. 14.9 Use ofConfirmatory Methods-None 14.10 D emonstrationofSpecificity-Specificiitsydemonstratedby chromatographic retentiotnime (within3% of standard)and themass spectrarlesponseofunique product ionsgeneratedfrom a characteristpircimaryion. 14.11 Documentation .14.11.1Ifcriterilaistedinthismethod performancesectionarenot me@ maintenance may be performed on thesystem and samplesreanalyzed,or otheractionstaken as determinedby the analyst.Document allactionsin theappropriatelogbook. 14.11.2Ifdataaretobe reportedwhen performancecriterihave not been met, the data must be footnotedon tablesand discussedin thetextof thereport. MS Word 97 ETS-8-154.0 Page 15 of 17 DeterminatioonfPFOS, PFOSA, POAA inWaterby Liquid-SoliEdxtractioannd LC/MS/MS 15.0 POLLUTION PREVENTION AND WASTF, MANAGEMENT 15.1 Sample extractwaste and flammable solventisdiscardedinhighBTU containersa,nd glasspipettewaste isdiscardedinbroken glasscontainerlsocatedin thelaboratory. 16.0 RECORDS 16.1 Each page generatedfora studymust have thefollowinginformationincluded,eitherin theheaderorhand-writtenon thepage: studyorpmject number, acquisitiomnethod, integratiomnethod,@samplename, extractiodnate,dilutiofnactor(ifapplicable)a,nd analyst. 16.2 Printthetunepage,sample lista,nd acquisitiomnethod from MassLynx toincludeinthe appropriatestudyfolder.Copy thesepages and tapeintotheinstrumentran log. 16.3 Plotthecalibratiocnurvesas describedin.thismethod,thenprintthesegraphsand store inthe studyfolder. 16.4 Printdataintegratiosniimmary,integratiomnethod, and chromatograms,from MassLynx, and storeinthestudyfolder. 16.5 Si,Tnmarie datausingsuitablseoftware(MS Excel 97) and storein'thestudyfolder. 16.6 Back up electronidcatatoappropriatmeedium. Record instudynotebook the.filnewne and locationofbackup electronidcata. 17.0 ArrAcEmENTs 17.1. AttachmentA: Figures-FluomchemicaClompounds 18.0 REFERENCES 18.1 "Guidelinesand Format forMethods tobe Proposed at40 CFR Part136 orPart 141", U.S.EnvironmentalProtectionAgency, OfficeofScienceand Technology Officeof Water,Washington,D.C. Draft1996. 18.2 'Method ofAnalysisfortheDeterminationofPerfluorooetanseulfonate(PFOS), Perfluorooctanseulfonylamide(PFOSA), and Perfluorooctanoa(tPeOAA) inWater",E. Wickrmnesinhe and J.Flaherty,Study Number 023-002,CentreAnalyticalLaboratories, Inc.,StateCollege,Pennsylvania,January 2000. 18.3 Validationreportforthe"N4ethodof AnalysisfortheDetenninationofPerfluorooctane sulfonat(ePFOS),Perfluorooctanseulfonylamide(PFOSA), and Perfluorooctanoate (POAA) inWatee',E. Wickremesinhe and J.FlahertyS,tudyNumber 023-002,Centre AnalyticalLaboratoriesI,nc.,StateCollege,Pennsylvania(,Approvalpending) 19.0 REVISIONS Revision Number. Reason For Revision Revision Date MS Word 97 M-8-154.0 Page 16 of 17 DetenninatioonfPFOS, PFOSA, POAA inWaterby Liquid-SoliEdxtractioannd LC/MS/MS Figure 1: PFOS Chemical Name Molecularion 0 Perfluorooctanseulfonate 499 (CsFl7SO3-) C8r 17S-0- 0 PFOS Note:Standardsaremade from thesaltp,otassiumperfluorooetasnuelfonat[eCgF17SO3K], m/w 538. Figure 2: PFOSA ChemicalName Molecularion Perfluorooctanseulfonylamide 498 (CgFl7SO2NH2) CBF17S- NH2 0 PFOSA Figure 3: POAA ChemicalName Molecularion Perfluorooctanoate 413 (c7FISCOO-) C7Fl5CC)7 POAA Note:Standardsaremade from thesalta,mmonium perfluorooetanoaItCe7FI5COONH4], m/w 431 MS Word 97 AttachmentA: ETS-8-154.0 pajze17 of17 DeterminationfPFOS, PFOSA, POAA inWaterby Liquid-SolEixdtractioand LC/MS/MS