A Comparison of Three Methods for Arsenic Speciation in Biological - PowerPoint PPT Presentation

A Comparison of Three Methods for Arsenic Speciation in Biological Tissues May Nguyen Brooks Rand Labs Seattle, WA

Select Arsenic Species Inorganic Species • arsenite [As(III)] Methylated Species • arsenate [As(V)] • monomethylarsine [MMA] • dimethylarsine [DMA] Organic Species • trimethylarsine [TMA] • arsenobetaine [AsB] • trimethylarsine oxide [TMAO] • arsenocholine [AsC]

Relative Toxicity Species Charge Toxicity AsB cation non ‐ toxic AsC cation non ‐ toxic MMA anion moderately toxic DMA anion moderately toxic TMA cation moderately toxic TMAO cation moderately toxic As(V) anion toxic As(III) anion very toxic

EPA Method 1632 • hydride generation reaction with volatile species • cryogenic trap • heating element – different boiling points for different species • atomic absorbance spectrophotometer

Sample A ‐ H ‐ R1 As(III) & As(V) Analysis 300000 • As(III) and As(V) have the same boiling point 250000 • As(In) = As(III) + As(V) As(In) • For biota, As(III) and As(In) As(III) prepared by the same digestion 200000 Signal [digital counts] method. • As(III) directly quantifiable – 150000 analysis within very specific pH range 100000 • requires separate prep • As(In) – As(III) = As(V) 50000 • two complete digestions and analyses 0 0 10 20 30 40 50 60 70 Time [s]

Sample S ‐ H ‐ R1 MMA & DMA Analysis 45000 As(In) • For biota, separate digestion 40000 method – NaOH. 35000 • MMA and DMA have very different boiling points 30000 Signal [ digital counts] • able to analyze for both in the same run 25000 • not easy to achieve baseline 20000 separation 15000 DMA MMA 10000 5000 0 0 10 20 30 40 50 60 Time [s]

EPA Method 1632 – Summary Pros Cons very narrow calibration range: 0.5 very low detection limits: 0.5 • • to 30 ng ng or 0.025 µg/L in reaction – in other words, 0.025 to 1.5 µg/L in vessel reaction vessel demonstrated method for As • – necessitates dilutions speciation – first drafted 1998 multiple digestions for multiple • As(III) – the most toxic species analytes • – is directly quantifiable As(V) is not directly quantified • MMA and DMA analysis is As(III) analysis requires titration • • pretty good no analysis for arsenic cation • species

Initial Demonstration of Proficiency for the Multilaboratory Validation of Arsenic Speciation Methods 3110 and 6870 EPA INTERCOMPARISON STUDY

Extraction by EPA 3110 heated digestion Certified Reference Certified Average • Material Value (mg/kg) Recovery (%) centrifugation of sample • DOLT ‐ 3 Dogfish Liver 10.2 84 material DOLT ‐ 4 Dogfish Liver 9.66 88 neutralization and heating of • digestion extract DORM ‐ 2 Dogfish Muscle 18 85 – Per EPA Sec. 11.2.3, it is noted DORM ‐ 3 Fish Protein 6.88 93 that some arsenicals are lost in the neutralization process. GBW 08571 Mussel 6.1 99 centrifugation and further • IAEA ‐ 407 Fish Tissue 12.6 97 heating of neutralized extract TORT ‐ 2 Lobster 21.6 82

Total Arsenic Recoveries Total Arsenic in Sample Total Arsenic in Extraction Sample Sample (ng/g) Extract (ng/g) Efficiency (%) A ‐ L ‐ R1 Rep 1 34900 31700 91 A ‐ H ‐ R1 Rep 1 164000 151000 92 A ‐ H ‐ R2 Rep 2 161400 155000 96 S ‐ L ‐ R1 Rep 1 8490 6980 82 S ‐ H ‐ R1 Rep 1 63600 60400 95 S ‐ H ‐ MS Matrix Spike 82130 75330 92 LCS BCR ‐ 627 LCS 4940 4020 81

Extraction by EPA 3110 HGAAS As(III) HGAAS As(V) HPLC As(III) HPLC As(V) Per EPA Sec. 1.2, digestion • 3.63 extract (TMAOH) favors 2.61 2.50 As(V) stability at higher pH. 1.56 1.17 0.13 0.12 0.08 TMAOH can act as an • oxidizing agent and push A ‐ L S ‐ L conversion of As(III) to 38.33 As(V). 21.60 18.53 15.20 10.10 0.60 0.84 A ‐ H S ‐ H

Extraction by EPA 3110 Pros Cons • single digestion for cation • unknown stability of species and anion analysis over time • Bigger mention for cation • conversion of As(III) to As(V) analysis!

EPA Method 6870 • HPLC ‐ ICP ‐ MS • encompasses 3 analyses: total arsenic in extract (via ICP), cations, and anions • separate ion ‐ exchange columns for anionic and cationic analysis • isocratic separation of the mobile phase

Cations – Calibration 5 µg/L 80000 AsB – reference peak 70000 60000 50000 Signal [cps] 40000 30000 AsB 20000 TMAO AsC TMA 10000 0 0 100 200 300 400 500 600 700 800 Time [s]

Cations – Sample S ‐ H ‐ R1 140000 120000 unknown species 100000 AsB – reference peak Signal [cps] 80000 60000 40000 TMAO AsC TMA AsB 20000 0 0 100 200 300 400 500 600 700 800 Time [s]

Cations – QC Results Sample Description AsB TMAO AsC TMA A ‐ L ‐ MS Matrix Spike 89% 94% 92% 94% A ‐ L ‐ MSD MS Duplicate 88% 88% 80% 89% A ‐ H ‐ MS Matrix Spike 67% 67% 63% 66% A ‐ H ‐ MSD MS Duplicate 77% 69% 66% 59% S ‐ L ‐ MS Matrix Spike 105% 69% 95% 95% S ‐ L ‐ MSD MS Duplicate 100% 70% 92% 93% S ‐ H ‐ MS Matrix Spike 77% 64% 84% 86% S ‐ H ‐ MSD MS Duplicate 74% 57% 64% 69% LCS BCR ‐ 627 ‐ MS LCS Spike 223% 119% 136% 137% BLANK SPIKE ‐ R1 Rep 1 116% 99% 97% 98% BLANK SPIKE ‐ R2 Rep 2 115% 99% 98% 99% BLANK SPIKE ‐ R3 Rep 3 113% 99% 98% 98%

Anions – Calibration 10 µg/L 80000 As(V) – reference peak 70000 60000 50000 Signal [cps] 40000 30000 As(III) DMA 20000 MMA As(V) 10000 0 0 200 400 600 800 1000 1200 1400 Time [s]

Anions – Sample S ‐ H ‐ R1 120000 unknown species 100000 80000 As(V) – reference peak Signal [cps] 60000 DMA 40000 As(V) MMA 20000 As(III) 0 0 200 400 600 800 1000 1200 1400 Time [s]

Anions – QC Results Sample Description As(III) DMA MMA As(V) A ‐ L ‐ MS Matrix Spike 4% 101% 108% 204% A ‐ L ‐ MSD MS Duplicate 3% 105% 108% 212% A ‐ H ‐ MS Matrix Spike 1% 73% 85% 151% A ‐ H ‐ MSD MS Duplicate 0% 90% 124% 207% S ‐ L ‐ MS Matrix Spike 8% 66% 69% 197% S ‐ L ‐ MSD MS Duplicate 8% 68% 68% 189% S ‐ H ‐ MS Matrix Spike 15% 81% 85% 215% S ‐ H ‐ MSD MS Duplicate 9% 64% 71% 175% LCS BCR ‐ 627 ‐ MS LCS Spike 12% 201% 199% 439% BLANK SPIKE ‐ R1 Rep 1 7% 150% 153% 308% BLANK SPIKE ‐ R2 Rep 2 9% 155% 154% 309% BLANK SPIKE ‐ R3 Rep 3 7% 150% 151% 303%

EPA Method 6870 – Summary Pros Cons anion and cation analyses reference peak does not • • potentially covers 8 species monitor for within run matrix HPLC ‐ ICP ‐ MS has a wider effects • calibration range: 0.25 ‐ 10 µg/L close peaks for As(III) and • direct quantification of all • DMA – no baseline separation species ICP ‐ MS standard mode is • ease and simplicity of use: • susceptible to polyatomic – standard mode for ICP ‐ MS interferences leading to biased – isocratic separation for HPLC results

A Comparison: EPA 1632 vs EPA 6870 EPA 1632 EPA 6870 species 4 8 digestions 3 1 analyses 3 2

Our Recommendations EPA 3110 EPA 6870 • Different digestion solution? • continuous internal standard introduction to – HNO 3 monitor matrix effects – HCl – NAOH • gradient ‐ step separation to • test for preservation get baseline separation of properties as well As(III) and DMA • DRC mode to alleviate polyatomic interferences

Interference Reduction Technology Average Recovery in Average Recovery Certified Reference Material Certified Value (mg/kg) Standard Mode (%) in DRC Mode (%) DOLT ‐ 4 Dogfish Liver 9.66 88 91 DORM ‐ 2 Dogfish Muscle 18 85 92 DORM ‐ 3 Fish Protein 6.88 93 97 IAEA ‐ 407 Fish Tissue 12.6 97 107 TORT ‐ 2 Lobster 21.6 82 97

Gradient ‐ Step Separation 10ppb As mix run1 10ppb As mix run2 Blank 1000 20 DMA MMA 18 900 As(III) 16 800 14 700 Standards Signal [Kcps] Blank Signal [cps] 12 600 As(V) 10 500 8 400 6 300 4 200 2 100 0 0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Time [min]

Questions?