Document gQXm6GDbm0z0953DXQp78KZa

BAKT MAIN 3M Environmental Laboratory BaciCopyof wtaBHaHv r SJlOO M ethod D e t e r m in a t io n o f P e r fl u o r o o c t a n e SULFONATE (PFO S), P e r f l u o r o o c t a n e SULFONYLAMIDE (PFO SA ), AND PERFLUOROOCTANOATE (POAA) IN WATER BY LlQUIDSo u d E x tr a c tio n and H ig h -P e r fo r m a n c e L iq u id C h ro m a to g ra ph y /T a n d em M a ss Sp e c t r o m e t r y (H PLC/M S/M S) M ethod N um ber: ETS-8-154.0 A doption Dote: / looq A uthor: K risten J. H ansen/H arold O . Johnson Revision D ate: Approved By: W illiam K. Reagen, Kent R. Lindstrom William K. Reagen, Laboratory M anagement 4 /p b A ) D ate Ik * Kristen J\ Hansen, PhD ., Group Leader Date Kent R. Lindstrom, Technical Reviewer M M co ____________________ . Date MS Word 97 ETS-8-154.0 Page 1 of 17 ^term ination o f PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LG/MS/MS BACK TO MAIN 1.0 S c o p e a nd Ap p l ic a t io n ____________________________________________________ 1.1 This method provides collection, extraction, and analytical procedures for the determination o f Perfluorooctane sulfonate (PFOS), Perfluorooctane Sulfonylamide (PFOSA), and Perflwnooctanoate (POAA) in groundwater, surface water, and drinking w ater samples. 1.2 This method was prepared according to die EPA document, "Guidelines and Form at for M ethods to be Proposed at 40 C FR Part 136 or Part 141" (see Reference 18.1), and is based in part on the report "M ethod o f Analysis for the Determination o f Perfluorooctane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluorooctanoate (POAA) in Water" (see Reference 18.2). 2.0 Su m m a r y o f M e th o d _____________________________________________________________ 2.1 W ater samples are collected from a site o f interest and shipped cold to an analytical facility. PFOS, PFOSA, and POAA are extracted from 40mL w ater samples using Cu solid phase extraction (SPE) cartridges. The compounds are eluted from die C ,gcartridge, using methanol. Separation, identification, and measurement are accomplished by high performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) analysis using m ultiple response m onitoring (MRM). The concentration o f each identified component is measured by comparing the MS response o f the quantitation ion produced by that compound to the MS response o f the quantitation ion produced by the same compound in an extracted calibration standard (external standard). 3.0 D e f in it io n s ________________________________ '_____________________ 3.1 A nalytical Sam ple--A portion o f an extracted Laboratory sample prepared for analysis. 3.2 C alibration S tandard--A solution prepared from the Working Standard (W S) and extracted according to this method. The calibration standard solutions are used to calibrate the instrument response w ith respect to analyte concentration. 3.3 D uplicate Sam ple (DS)--A separate aliquot o f a sample, taken in the analytical laboratory and analyzed separately w ith identical procedures. Analysis o fD S s compared to that o f the first aliquot give a measure o f the precision associated w ith laboratory procedures, but not w ith sample collection, preservation, o r storage procedures. 3.4 Field B lank C ontrol Sam ple (FB)--Type I w ater placed in a Sample container in the laboratory and treated as a sample in all respects, including exposure to sam pling site conditions, storage, preservation and all analytical procedures. The purpose o f the FB is to determine if test substances o r other interferences are present in the field environment. MS Word 97 ETS-8-154.0 Page 2 of 17 Determination of PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 3.5 Field D uplicate (FD)--A sample collected in duplicate at die same tim e as the sample and placed under identical circumstances and treated exactly die same throughout field and laboratory procedures. Analysis o f FD compared to that o f the first sample gives a measure o f the precision associated with sample collection, preservation and storage, as w ell as w ith laboratory procedures. 3.6 Field M atrix Spike (FM S)--A sample collected in duplicate to which known quantities o f the target analytes are added in the field at the tim e o f sample collection. The FMS should be spiked at approximately 50-150% o f the expected analyte concentration in die sample. The FMS is analyzed to ascertain if any matrix effects, interferences, o r stability issues m ay complicate the interpretation o f the sample analysis. 3.7 Field Spike C ontrol Sam ple (FSCS)--A n aliquot o f type I w ater to which known quantities o f the target analytes are added in the field at the tim e o f sample collection (at an appropriate concentration to be determined by the project lead). The FSCS is extracted and analyzed exactly Hire a sample to determine whether a loss o f analyte could be attributed to sample storage and/or shipm ent 3.8 L aboratory C ontrol Sam ple (LCS)-- An aliquot o f type I water to which known . quantities o f the target analytes are added in the laboratory. Two levels are included, one at die LOQ (approx. 25Pg/mL), the other at a concentration o f approx. 100-250Pg/mL or another concentration to be determined by the project lead. The LCS is extracted and analyzed exactly like a laboratory sample to determine whether the methodology is in control, and whether the laboratory is capable o f making accurate measurements at the required method detection lim it and higher. 3.9 L aboratory Sam ple---A portion o f a sample received from die field for testing. 3.10 L im it o f D etection (LOD)--The lowest concentration o f an analyte that can be measured and reported w ith 99% confidence that the analyte concentration is greater than zero. The LOD can be determined in several ways, including signal-to-noise ratio and statistical calculations. 3.11 L im it o f Q uantitation (LO Q )--The lowest concentration (LLOQ) or highest concentration (ULOQ) that can be reliably achieved within the specified lim its o f precision and accuracy during routine operating conditions. N ote: The LLOQ is generally 5-10 tim es die LOD. For many analytes, die LLOQ analyte concentration is selected as the low est non-zero standard in die calibration curve. How ever, it m ay be nom inally chosen w ithin these stated guidelines to sim plify data reporting. Sam ple LLOQs are m atrix-dependent 3.12 M atrix Spike (MS)-- A n aliquot o f a sample, to which known quantities o f target analytes are added in the laboratory. The MS is extracted and analyzed exactly like a laboratory sample to determine whether the sample matrix contributes bias to the analytical results. The background concentrations o f die analytes in the sample m atrix m ust be determined in a separate aliquot and the measured values in the MS corrected for background concentrations. MS Word 97 ETS-8-154.0 Page 3 o f 17 Detenmnatian ofPFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LCYMS/MS BACK TO MAIN 3.13 M ethod B lank--An aliquot o f type I w ater that is treated exactly like a laboratory sample including exposure to all glassware, equipment, solvents, reagents, internal standards, and surrogates that are used w ith other laboratory samples. The m ethod blank is used to determine if test substances or other interferences are present in the laboratory environment, die reagents, o r die apparatus. 3.14 M ethod Detection L im it (M DL) D eterm ination--One o f several processes that m ay be used to establish a LO D value. The statistically calculated minimum am ount o f an analyte that can be measured w ith 99% confidence that the reported value is greater than zero. This term is usually associated w ith the EPA definition in 40 GFR Part 136 Appendix B . 3.15 Sam ple--A sample is a small portion collected from a larger quantity o f m aterial intended to represent die original source material. 3.16 Spiking Stock S tan d ard (SSS)--A solution prepared from stock standards used to prepare die working standard. . 3.17 Stock S tandard OSS)--A concentrated solution o f a single analyte prepared in the laboratory w ith an assayed reference compound. . 3.18 W orking S tandard (W S)--A solution o f several analytes prepared in the laboratory from SSs and diluted as needed to prepare calibration standards and other required analyte solutions. 4.0 W a rn in g s and c a u t io n s_____________________________________________________ 4.1 H ealth and Safety W arnings 4.1.1 The acute and chronic toxicity o f die standards t o this method have not been precisely determined; however, each should be treated as a potential health hazard. 4.1.2 Unknown samples m ay contain high concentrations o f volatile toxic compounds. Sample containers should be opened in a hood and handled with gloves to prevent exposure. 4.1.3 T he laboratory is resp o n sib le fo r m aintaining a safe w ork environm ent an d a curren t awareness o f local regulations regarding the handling o f the chem icals used in this method. A reference file o f material safety data sheets (M SDS) should be available to all personnel involved in these analyses. 5.0 In t e r fe r e n c e s________________________________ ^_____________________________ _____ 5.1 During extraction and analysis, m ajor potential contaminant sources are reagents and liquid-solid extraction devices. 5.2 A ll materials used in the analyses shall be demonstrated to be free from interferences under conditions o f analysis by running method blanks. 5.3 Teflon* containing m aterials (e.g. caps, w ash bottles) contain fluorocompounds w hich m ay cause interferences and should not be used during collection, storage, extraction, o r analysis o f the samples. MS Word 97 ETS-8-154.0 Page 4 o f 17 Determination ofPFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 6 .0 E q u ipm e n t , S u p p l ie s , and M a t e r ia l s ____________________________________________ N ote: Brand names, suppliers, and part num bers are for illustrative purposes only. Equivalent perform ance m ay be achieved using apparatus and m aterials other than those specified here, but dem onstration o f equivalent performance th at m eets the requirem ents o f this m ethod is the responsibility o f the laboratory. 6.1 Sam pling E quipm ent 6.1.1 Sam ple collection b o ttles-- L D PE (e.g ., N algeneTM ) narrow -m outh bottles w ith screw cap. N ote: Do not use Teflon bottles o r Teflon lined caps. 6.1.2 Coolers for sample shipm ent 6.13 Ice for sample shipm ent . 6.1.4 Bottles must be lot-certified to be free o f artifacts by running M ethod blanks according to this method. , 62 L aboratory E quipm ent (E xtraction and A nalytical) 6.2.1 Balance, analytical (display at least O.OOOlg), M ettler. 6.2.2 Vacuum pump, Bfichi. 6.2.3 Visiprep vacuum manifold, Supelco. 6.2.4 Sep Pak Vac 6cc (lg ) tC ,, cartridges (part # W AT 036795),W aters. 6.2.5 50mL disposable polypropylene centrifuge tubes, VWR. . 6.2.6 15mL disposable polypropylene centrifuge tubes, VWR. 6.2.7 Disposable micropipettes (50-1 OOpL, 100-200pLX Drummond. 6.2.8 Class A pipettes and volumetric flasks, various. 6.2.9 Hypercarb drop-in guard column (4mm) (part # 844017-400), Keystone. 6.2.10 Stand-alone drop-in guard cartridge holder, Keystone. 6.2.11 125mL LDPE narrow-mouth bottles, Nalgene. 6.2.12 HPLC pump (LC10AD), Shimadzu. 6.2.13 2mL clear HPLC vial kit (cat # 5181-3400), Hewlett Packard. 6.2.14 Standard lab equipment (graduated cylinders, disposable tubes, etc.), various. 6.2.15 LC/MS/MS and HPLC systems, as described in section 10.1. 6 3 E quipm ent Notes 63.1 In order to avoid contamination, the use o f disposable labware is highly recommended (tubes, pipettes, etc.). 6 3 .2 Teflon or Teflon-lined containers or equipment, including Teflon-lined HPLC vials o r caps for the HPLC auto sampler m ust not be used. 6 3 3 Type I w ater used during the sample and standard extraction should be filtered through a Hypercarb guard column using a HPLC pump. This w ater is referred to as "filtered type I water", hereafter in this repeat 6 3 .4 It is necessary to check the solvents (methanol) for the presence o f contam inants (especially POAA) by LC/MS/MS prior to use. Certain lot numbers have been found to be unsuitable for use. 6 3 .5 Use disposable micropipettes or pipettes to aliquot standard solutions to m ake calibration standards and matrix spikes. MS Word 97 ETS-8-154.0 PageS o f 17 Detenninatian o f PFOS, FFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 7.0 R e a g e n t s a nd S ta n d a r d s ___________________ ___________________________________ N ote: Suppliers and catalog num bers are for illustrative purposes only. Equivalent perform ance m ay be achieved using chem icals obtained from other suppliers. Do not use a lesser grade o f chem ical than those listed. 7.1 Chem icals 7.1.1 Methanol (MeOH), HPLC grade, JT Baker, Catalog No. JT9093-2. 1.12 Ammonium Acetate, Reagent grade, Sigma-Aldrich, Catalog No. A -7330. 7.1.3 Water, type I, prepared in-house. 7.1.4 Sodium Thiosulfate, Reagent grade, JT Baker. 12 Standards 7.2.1 Potassium perfluorooctane sulfonate (see Attachment A, Figure 1). 7.2.2 Perfluorooctane sulfonylamide (see Attachment A, Figure 2). 7.2.3 Ammonium perfluorooctanoate (see Attachment A, Figure 3). 13 Reagent Preparation 7.3.1 250mg/mL sodium thiosulfate solution (E xtraction)--D issolve 25g o f sodium . thiosulfate in lOOmL reagent water. 7.3.2 40% m ethanol (E xtraction)-- M easure 400mL methanol and adjust the volume to 1.0L w ith reagent water. 1 3 3 lOOmM am m onium acetate solution (Analysis)--W eigh 7.71g o f ammonium acetate and dissolve in 1.0L o f reagent water. Dilute the lOOmM solution by a factor o f 50 to make the 2mM ammonium acetate solution used for m obile phase A N ote: A lternative volum es m ay be prepared as long as the ratios o f die solvent to solute ratios are maintained. 7.4 Spiking Stock S tan d ard (SSS) P reparation 7.4.1 lOOpg/mL each PFO S, P F O S A and POAA SSSs--W eigh out lOmg o f analytical standard (corrected for percent salt and purity--Le., 10 mg Q F ^ O jK purity 90% * 8.35mg C ^ tSOj- ) and dilute to 1OQmL, w ith methanol in a 1OOmL volumetric flask. Transfer to a 123mL UDPE bottle. Prepare a separate solution for each analyte. Store solutions in a refrigerator at 42C for a maximum period o f 6 months from the date o f preparation. 7.42 lpg/m L m ixed SSS--Add l.OmL each o f the lOOpg/mL SSSs (from 7.4.1) to a lOOmL volum etric flgek and bring up to volume w ith methanol. 1.43 0.1fig/mL m ixed SSS--Add lO.OmL o f the l.Opg/mL-mixed solution (from 7.4.2) to a lOOmL volumetric flask and bring up to volume w ith methanol. 7.4.4 O.Olpg/mL m ixed SSS--Add lO.OmL o f the 0.1fig/mL-mixed solution (from 7.4.3) to a 100mL volumetric flask and bring up to volume w ith methanol. 7.4.5 Storage C onditions--Store all SSSs in a refrigerator in 125mL LDPE bottles at 42C for a maximum period o f 3 months from foe date o f preparation. MS Word 97 ETS-8-154.0 Page 6 of 17 Determination of PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 7.5 C alibration S tandards 7.5.1 100pg/mL each PFO S, PFO SA , and POAA stock stan d ard solutions--W eigh out lOmg o f analytical standard (corrected for percent salt and purity) and dilute to lOOmL w ith methanol in a lOOmL volumetric flask. Transfer to a 125mL LDPE bottle. Prepare a sep arate solution for each analyte. Store solutions in a refrigerator at 42C for a maximum period o f 6 months from foe date o f preparation. ' 7.5.2 lfig/m L W orking S tan d ard --Add l.QmL each o f the 10pg/mL SS solutions (from 7.5.1) to a lOOmL volum etric flask and bring up to volume w ith m ethanol. 7.5.3 O.lpg/m L W orking S tan d ard -- Add 10.OmL o f the 1.Opg/mL m ixed solution (from 7.52) to a lOOmL volum etric flask and bring up to volume w ith methanol. 7.5.4 0.01fig/mL W orking S tan d ard --Add lO.OrhL o f the O.lpg/mL m ixed solution (from 7.5.3) to a lOOmL volumetric flask and bring up to volume w ith methanoL 7.5.5 Storage C ondition*--Store all W Ss in a refrigerator (in 125mL LDPE bottles) at 42C for a maximum period o f 3 months from the date o f preparation. 7.5.6 C alibration S tan d ard --Prepare a minimum o f five calibration solutions in filtered type I w ater according to the following table: Concentration Volume of Final Calibration Standard Final Concentration of of WS, fig/mL W S,|tL Volume, mL Calibration Standard, Pg/mL 0.0 0 40 0 0.010 100 40 25 0.010 0.010 200 400 40 40 50 100 0.10 100 40 250 0.10 200 40 500 0.10 300 40 7501 0.10 400 40 10002 1 May be preptred to extend the range beyond SOOPg/mL. 2 Mybe prepared to extend tbe rang* beyond 750Pf/inL. Note: The absolute volumes of the standards may be varied by the analyst as long as foe correct proportions o f solute to solvent are maintained. 7.5.7 The standards are processed through the extraction procedure (Section 9.0), identical to foe laboratory samples. Tim extracted concentration o f foe calibration standard is equal to 8X foe initial concentration, due to foe concentration o f foe standard during foe extraction process. 725.8 Storage C onditions--Store all extracted calibration standards in ISmL polypropylene tubes at 420C, for a maximum period o f tw o weeks from the date o f preparation. MS Word 97 EIS-8-154.0 p age 7 o f 17 Determination ofPFOS, PFOSA, POAA in Water by Liquid-Solid Extraction *nd LC/MS/MS BACK TO MAIN 8.0 Sa m p l e C o l l e c t io n , P r e s e r v a t io n a n d S t o r a g e _________________________ Note: Sampling equipment, including automatic samplers, must be free ofTeflon tubing, gaskets, and other p ots that may leach interfering analytes into toe water sample. Automatic samplers that composite samples over time should use refrigerated polypropylene sample containers if possible. Sample bottles should not be rinsed before sample collection. 8.1 T ap W ater--Open the tap and allow the system to flush until toe w ater tem perature (1510Q has stabilized (usually about two minutes). Adjust toe flow to about 500mL/min and collect samples from toe flowing stream. 8.2 G round W ater--Purge toe well o f standing w ater using a pump or a bailer. C ollect toe sample directly from the pump or from toe bailer. 8 3 Surface W ater--W hen sampling from an open body o f water, fill toe sample container w ith water from a representative area. 8.4 Sam ple D echlorination--All samples should be iced or refrigerated at 42C and kept in toe dark from toe tim e o f collection until extraction. Residual chlorine should be reduced by adding 200jiL o f a 25Qmg/mL sodium thiosulfate solution to each w ater . sample, FB, and FSCS (which may be placed in each bottle before leaving for toe sampling site.). 8 3 H olding Tim e (H T)-- Results o f the time/storage study o f all target analytes showed that toe three compounds are stable for 14 days in water samples when toe sam ples are dechlorinated and stored as described in section 8.4 (see also reference 18.3). Therefore, laboratory samples m ust be extracted w ithin 14 days and toe extracts analyzed w ithin 30 days o f sample collection. I f the HT exceeds 14 days, great care is used when evaluating field spikes to avoid misrepresentation o f the sample concentration. 8.6 Field B lanks . 8.6.1 Process a Field Blank Control Sample (FB) along w ith each sample set (samples collected from toe same general sample site at approximately toe same tim e). A t the laboratory, prior to sample collection, fill a sample container w ith filtered type I w ater, seal, and ship the FB to the sampling site along w ith the em pty sample containers. Return the FB to the laboratory with the filled sam ple bottles. 8.6.2 When sodium thiosulfate is added to samples, use toe same procedure to preserve theFB . 8.7 Field D uplicates 8.7.1 Collect a Field Duplicate (FD) for every ten (10) samples collected o r per each sampling set, if less than 10 samples are collected. 8.7.2 Separate FDs m ust be collected for each type o f water sample (ground, tap, etc.) collected. 8 .7 3 Collect toe FD immediately after toe sample. 8.7.4 Preserve, store and ship FD using the same procedures as used for the samples. MS Word 97 ETS-8-154.0 Page 8 of 17 Determination of PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS7M^ BACK TO MAIN 8.8 Field Spike C ontrol Sem ple (FSCS) 8.8.1 A Field Spike Control Sample (FSCS) m ust be prepared for each sample shipm ent I f m ultiple coolers are used to ship a set o f samples, each cooler m ust contain a FSCS. 8.8.2 A t the laboratory, fill a sample container 'with lOOmL o f type I water. Seal and ship to file sampling site along w ith the empty sample containers and FBs. 8.8.3 W hen sodium thiosulfate is added to samples, use the same procedure to add the same amounts to file FSCS. 8.8.4 Seal and gently invert the FSCS to mix. Store and ship the FSCS using file same procedures as used for the samples. MS Word 97 ETS-8- 154.0 Page 9 o f 17 Determination ofPFOS, PFOSA, POA in Water by Liquid-Solid Extraction and LC/MS/MS 9.0 Extraction Procedure 9.1 E xtraction Scheme 9.1.1 Allow samples to equilibrate to room temperature. Thoroughly m ix samples by gently inverting the sample bottle. 9.1.2 M easure 40mL o f sample into 50mL polypropylene centrifuge tubes (Spike the QC and M atrix spikes as required*, replace lid and mix well). N ote: * Samples m ay need to be prescreened to determ ine an appropriate m atrix spike level (typically 50-150% o f sam ple concentration). 9.1.3 Condition the C,t SPE cartridges (Ig , 6mL) by passing 1OmL methanol followed by 5mL filtered type I w ater (~2drop/sec). Do not let column run dry. N ote: F or the following steps, mmniain a ~ldrop/sec flow rate. Do not allow the colum n to run dry at any tim e. 9.1.4 Load the analytical sample onto the Cu SPE cartridge. Discard ehiate. 9.1.5 W ash w ith ~5mL 40% methanol in water. Discard ehiate. 9.1.6 Elute w ith ~5mL 100% methanol. Collect 5xnL o f eluate into graduated 15mL polypropylene centrifuge tubes. This is the target elution fraction (final volum e = 5mL). 9.1.7 Analyze a portion o f the target elution fraction eluent using negative electrospray HPLC/MS/MS (Section 10.2). N ote: Samples are concentrated by a factor o f eight during the extraction; Initial V ol -4 0 m L -> Final Vol. * 5mL. 9.1.8 Samples are stable at room temperature for at least 24 hours. Analytical sam ples may be stored in a refrigerator at 42C until analysis. 9.1.9 S tandardization o f Clg SPE colum ns--I f poor recoveries are observed, it may be necessary to standardize the C ,, SPE columns in the following m anner before analyzing samples. 9.1 A 1 U se a standard w ith an analyte concentration between 1000 and 4000 Pg/mL. Follow tile extraction scheme as outlined from steps 9.1.1 to 9.1.6t except, collect the eluate fraction separately (approx. 5mL), as w ell as the target elution fraction. 9.1.9.2 A fter step 9.1.6, collect a post-elution fraction by, eluting w ith an additional 5mL o f 100% methanol. 9.1.93 Analyze all three fractions by HPLC/MS/MS. I f the target fraction contains a minimum o f 85% o f the respective analytes, it m ay be considered acceptable. 9.13.4 I f the wash contains significant standard (>15%), either the w ash volume o r percentage ofM eO H should be decreased. 9.1.93 I f the post-elution fraction contains significant standard (>15%), the target elution volume should be increased. MS Word 97 ETS-8-154.0 Page 10 o f 17 Determination of PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and L O M S /^j BACK TO MAIN 10.0 C a l ib r a t io n and St a n d a r d iz a t io n (a n a l y t ic a l S e tu p) _________________________ N ote: Other instruments m ay be used and the equipm ent and conditions m ay be very different as long as the method criteria are m e t The operator m ust optim ize and document the equipm ent and settings used. . 10.1 Establish the LC/MS/MS system and operating conditions equivalent to the following: M ass Spec: M icromass Quattro Ultima (Micromass) ' Interface: Electrospray (Micromass) M ode: Electrospray Negative, M ultiple Response M onitoring (MRM) Harvard infusion pump (Harvard Instruments), for tuning C om puter COMPAQ Professional W orkstation AP200 Software: Windows NT, M assLynx 3.3 HPLC: Hewlett Packard (HP) Series 1100 HP Quat Pump HP Vacuum Degasser HP Autosampler HP Column Oven ' N ote: A 4 x 10mm Hypercarb drop-in guard cartridge (Keystone, part # 844017-400) is attached on-line after the purge valve and before the sample injector port to trap any residue contam inants that m ay be in the m obile phase and/or HPLC system. HPLC Column: Genesis C , (Jones Chromatography), 2.1mm x 50mm, 4jim Column Temperature: 35C Injection Volume: 15pL M obile Phase (A): 2mM Ammonium Acetate in filtered type I w ater (See 7.3.1) M obile Phase (B): M ethanol HPLC Gra d ia it Program : Tim e, P ercent M obile P ercent M obile m in Phase A Phase B 0.0 60 0.4 60 40 40 1.0 10 90 7.0 10 90 7J 0 100 9.0 0 100 9J5 60 40 13.5 60 40 14.0 60 40 Flow R ate, m i 7mfn 0.3 0.3 0.3 0.3 0.3 0.4 0.4 0.4 0.3 N ote: O ther HPLC gradients m ay be used as long as the method criteria are m e t MS Word 97 ETS-8-154.0 Page 11 of 17 Determination ofPFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN It m ay be necessary to adjust die HPLC gradient in order to optimize instrument perform ance. Columns w ith different dimensions (e.g. 2.1mm x 30mm) and columns from different m anufacturers (Keystone Betasil C ,, etc.) m ay be used. Ions M onitored: A nalyte P rim ary Ion P ro d u ct Ion POAA PFOS PFOSA 413.0 499.0 498.0 169.0 99.0 78.0 A pproxim ate R etention Tim e 5.0 5.2 5.8 Other product ions may be chosen at the discretion o f the analyst, although m /z 99 is suggested for PFOS. U se o f the suggested prim ary ion is recommended. Retention tim es m ay vary slightly, on a day-to-day basis, depending on the batch o f m obile phase etc. D rift in retention tim es is acceptable w ithin an analytical run, as long as die drift continues through the entire analysis and the standards are interspersed throughout the analytical run. ` . 10.2 T one File Param eters 10.2.1 The following values are provided as an example. Actual values m ay vary from instrument to instrum ent Also, these values may be changed from tim e to tim e in order to optimize for greatest sensitivity. A nalyte POAA PFOS PFOSA Dwell, sec 0.2-0.4 0 .2 -0 .4 0.2-0.4 C o llisio n E nergy, eV 10-25 30-60 20-50 Cone, V 20-30 50-80 30-60 Source C ap illary Hexapole 1 Aperture 1 Hexapole 2 Source Block Temp. Desolvation Temp. Set 2.56-3.5kV 0.5V 0.2V 0.8V 100-150C 250-400C MS Word 97 ETS-8-154.0 Page 12 of 17 Determination o f PFOS, PFOSA, PQAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN A nalyzer LM Res 1 HM Res 1 DEnergy 1 Entrance E x it LM Res 2 HMRes 2 IEnergy 2 M ultiplier Set 12.0-15.0V 12.0-15.0V 0.7V -2V IV . 11.0V 11.0V 1.0V 650V Gan Flows Cone Gas D esolvation Set 150L/hr 700L/hr Pressures Gas Cell Set 3.0e-3m bar 11.0 A n a l y t ic a l Q u a l it y C o n t r o l __________________________________________________ 11.1 Analytical results o f the FB, FM S, FD, and FSCS should be evaluated at die conclusion o f the study to help interpret the data quality o f samples data. Analytical results for these control/dupHcate samples m ust be reported w ith the sample data. MS Word 97 ETS-8-154.0 . pige 13 of 17 Detennmttkm ofPFOS. PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 12.0 A n a l y t ic a l P r o c e d u r e _______________________________ 12.1 SampleAnalysis 12.1.1 Set up analysis sample queue. 12.1.2 Inject the same aliquot (between 5-25 jxL) o f each standard, analytical sample, recovery, control etc. into the LC/MS/MS system. 12.1.3 A ll samples showing a response for one or more analytes above the response o f die highest, active calibration curve level m ust be diluted and reanalyzed. 122 C alibration C urve 12.2.1 Starting with the standard o f lowest concentration, inject the same size aliquot (between 10-25pL) o f each extracted calibration standard according to Section 12.1 and tabulate the response (peak height or area) versus the concentration in die standard. U se linear standard curves for quantitation generated for each . analyte by linear regression w ith 1/x weighting o f peak area versus calibration standard concentration. The correlation coefficient (r) for the calibration curves . m ust be 0.990 (r*0.980). I f calibration results fall outside these lim its, then appropriate steps m ust be taken to adjust instrument operation and the standards reanalyzed. 12.2.2 C urve--The measured value for each curve point must be w ithin 30% o f theoretical values when curve is evaluated over a range appropriate to die data. High or low points m ay be deactivated to achieve these criteria, but an acceptable curve m ust contain at least five active curve points. ` 12.2.3 C ontinuing C urve V erification (CCV)--M id- and low-level calibration checks should be analyzed every 5-10 injections. The analyte level measured in the CCVs should be w ithin 30% o f theoretical values. I f CCVs fall outside o f this range, data collected subsequent to the last passing CCV should not be used. Only data collected between acceptable CCVs or the initial curve can be used. 13.0 D a ta A n a ly sis and C a lc u la t io n s 13.1 C alculate th e analytical sam ple (ex tract) concentration from th e standard cu rv e u sin g tire following equation: (slope) 13J2 Calculate the percent recovery o f the FSCS using the following equation: rFoSwC S %/oricwc.. = ((CFSoCncS cone.,_PPggffealLL)) 13J Calculate the percent recovery o f die M Ss using die following equation: M S/rec ( ^ <^ c > P g ^ - S^ leC^ p gftpL) jc10q ' (C one, added, Pg/m L ) MS Word 97 ETS-8-154.0 Page 14 o f 17 Determination ofPFOS, PFOSA, PQAA in Water by Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN 14.0 M e t h o d P e r fo r m a n c e p a r a m e t e r s ______________________________________________ N ote: Any method performance param eters that are not achieved m ust be considered in the evaluation o f the data. Nonconform ance to any specified param eters m ust be described and discussed in any reporting o f the data. 14.1 L inearity--Linear standard curves for quantitation generated for each analyte by linear regression with 1/x weighting o f peak area versus calibration standard concentration. The correlation coefficient (r) for fee calibration curves m ust be 0.990 (i*0.980). 14.2 C alibration C urve Standards--The measured value for each curve point m ust be w ithin 30% o ftheoretical values when curve is evaluated over a range appropriate to the data. High or low points may be deactivated to achieve these criteria, but an acceptable curve m ust contain at least five active curve points. 143 C CV Perform ance--M id and low level calibration checks to be analyzed every 5-10 injections. The analyte level measured in fee CCVs should be within 30% o f theoretical values. I f CCVs fall outside o f this range, data collected subsequent to the last passing CCV should not be used. Only data collected between acceptable CCVs can be used. 14.4 L im it of D etection (LOD)-- The low est calibration standard w ife a peak area at least 2X fee peak area o f fee extraction blank that can be measured at a concentration greater than zero. . 14.5 L im its o f Q uantitation (LO Q)--The lower LOQ (LLOQ) is the lowest non-zero active standard in fee calibration curve; fee peak area o f fee LLOQ m ust be at least 2X feat o f fee extraction blank. B y definition, fee measured value o f fee LLOQ m ust be w ithin 30% o f fee theoretical value. 14.6 M atrix Spikes--M atrix spike percent recoveries m ust be within 30% o f fee spiked concentration. 14.7 Solvent B lanks, M ethod B lanks, an d M atrix Blanks--Values m ust be below fee lowest non-zero active standard in fee calibration curve. M atrix blanks are considered compliant if no test substance is detected above fee LOD for feat analyte. 14.8 R eproducibility--Reproducibility o f fee method is defined by fee results o f the matrix spikes and m atrix spike duplicates. The MS/MSD should be reproducible to w ithin 20%. 14.9 Use o f C onfirm atory M ethods--None 14.10 D em onstration o f Specificity-- Specificity is demonstrated by chromatographic retention time (w ithin 3% o f standard) and fee mass spectral response o f unique product ions generated from a characteristic prim ary ion. . 14.11 D ocum entation 14.11.1 I f criteria listed in this method performance section are not met, maintenance may be perform ed on fee system and samples reanalyzed, or other actions taken as determined by the analyst Document all actions in fee appropriate logbook. 14.11.2 I f data are to be reported when performance criteria have not been m et, fee data must be footnoted on tables and discussed in fee text o f fee report MS Word 97 ETS-8-154.0 Page 15 of 17 Determination of PFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LQMS/MS BACK TO MAIN 15.0 P o l l u t io n P r e v e n t io n a n p W a s t e M a n a g em en t _______________________________ 15.1 Sample extract waste and flammable solvent is discarded in high BTU containers, and glass pipette waste is discarded in broken glass containers located in the laboratory. 16.0 R ec o r d s_____________________________________ ____________________________________ 16.1 Each page generated for a study m ust have die following information included, either in the header or hand-written on the page: study or project number, acquisition method, integration m ethod,sam ple nam e, extraction date, dilution factor (if applicable), and analyst 16.2 Print die tune page, sample list, and acquisition method from MassLynx to include in the appropriate study folder. Copy these pages and tape into the instrument run log. 1 6 3 Plot the calibration curves as described in this method, then print these graphs and store in the study folder. 16.4 Print data integration summary, integration method, and chromatograms, from M assLynx, and store in the study folder. 1 6 3 Summarize data using suitable software (M S Excel 97) and store in the study folder. 16.6 Back up electronic data to appropriate medium. Record in study notebook the .file name and location ofbackup electronic data. 17.0 A t ta c h m en t s___________________________________________________________________ 17.1 A ttachm ent A: Figures--Fluorochemical Compounds 18.0 R e fe r e n c e s______________________________________________________________________ 18.1 "Guidelines and Format for M ethods to be Proposed at 40 CFR Part 136 o r P art 141", U.S. Environmental Protection Agency, Office o f Science and Technology O ffice o f W ater, W ashington, D.C. D raft 1996. 18.2 "M ethod o f Analysis for the Determination o f Perfluorooctane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluorooctanoate (POAA) in W ater", E. W ickremesinhe and J. Flaherty, Study Number 023-002, Centre Analytical Laboratories, Inc., State College, Pennsylvania, January 2000. 1 8 3 Validation report for the "M ethod o f Analysis for the Determination o f Perfluorooctane sulfonate (PFOS), Perfluorooctane sulfonylamide (PFOSA), and Perfluorooctanoate (POAA) in Water", E. W ickremesinhe mid J. Flaherty, Study Number 023-002, Centre Analytical Laboratories, Inc., State College, Pennsylvania, (Approval pending) 19.0 R e v isio n s R ev isio n N um ber. Reason For Revision R ev isio n D ate MS Word 97 ETS-8-154.0 Page 16 c 17 Determination o f PFOS, PFOSA, POAA in Waterby Liquid-Solid Extraction and LC/MS/MS BACK TO MAIN F ig u re 1: P F O S Chemical Name M olecular ion Perfluorooctane sulfonate 499 (C ^ ,7S 0 31 0 II II C 8F 17S -O - O PFOS N ote: Standards are made from die salt potassium perfluorooctane sulfonate [CgFjySC^K ], m/w 538. Figure 2: PFOSA Chemical Name M olecular ion * Perfluorooctane sulfonylamide 498 (C*F,7SOiNH2) 0T C8F17S-- - nh2 0 . PFOSA Figure 3: POAA Chemical Name = M olecular ion Perfluorooctanoale 413 (C7F15COO") I CtF15CO- POAA N ote: Standards are made from the salt, ammonium perfhiorooctanoate [C7F j 5COONH4], m /w 431 '' MS Word 97 Attachment A: ETS-8-154.0 page 17 of 17 Determinatimi o fPFOS, PFOSA, POAA in Water by Liquid-Solid Extraction and LC/MS/MS