Medical Aspects of Chemical Warfare (2008) - The Black Vault

Medical Aspects of Chemical WarfareEXHIBIT 22-4SAMPLE PREPARATION METHODS FOR Gas Chromatographic/MassSpectrometric ANALYSIS OF chlorovinylarsonous acidSPE procedure of Logan et al for CVAA in urine:• Precondition a C18 SPE cartridge with methanol followed by water.• Adjust pH of 1 mL of urine to pH 6 using HCl.• Add PAO as an internal standard.• Transfer sample to C18 SPE cartridge for sample cleanup/extraction.• Elute analytes using methanol.• Evaporate to dryness under nitrogen.• Reconstitute with ethanol containing EDT.• Analyze using GC-MS with EI ionization. 1SPME headspace procedure of Wooten et al for CVAA in urine:• Place 1 mL of urine into a 10-mL vial.• Add ammonium acetate buffer-water solution.• Add PAO as an internal standard.• Add PDT in order to derivatize CVAA and PAO.• Crimp-seal vial and heat at 70°C for 20 min.• Insert a 100-μM poly(dimethylsiloxane) SPME fiber into the sample vial headspace for 10 min.• Remove the SPME fiber from the vial and insert into the injection port of a GC-MS with EI ionization. 2SPE procedure of Fidder et al for free and bound CVAA in blood:• Add 2,3-dimercapto-1-propanol (BAL) to a 2-mL blood sample. The BAL causes displacement of bound CVAAand derivatizes both the free and released CVAA.• Add phenylarsine-BAL as an internal standard.• Shake the sample at room temperature overnight.• Dilute the sample with water.• Transfer sample to C18 SPE cartridge for sample cleanup/extraction.• Elute the analytes from the cartridge using dichloromethane/acetonitrile mixture.• Concentrate the extracted sample to dryness.• Redissolve in toluene.• Add HFBI to derivatize the BAL hydroxyl group.• Incubate the sample for 1 hour at 50°C and then cool to room temperature.• Wash with water and then dry over MgSO 4.• Analyze using GC-MS with EI ionization. 3BAL: British anti-LewisiteCVAA: chlorovinylarsonous acidEDT: ethanedithiolEI: electron impactGC: gas chromatographyHCl: hydrochloric acidHFBI: heptafluorobutyryl imidazoleMS: mass spectrometryPAO: phenylarsine oxidePDT: 1,3-propanedithiolSPE: solid phase extractionSPME: solid phase microextractionData sources: (1) Logan TP, Smith JR, Jakubowski EM, Nielson RE. Verification of lewisite exposure by the analysis of 2-chlorovinylarsonous acid in urine. Toxicol Meth. 1999;9:275–284. (2) Wooten JV, Ashley DL, Calafat AM. Quantitation of 2-chlorovinylarsonousacid in human urine by automated solid-phase microextraction-gas chromatography-mass spectrometry. J Chromatogr B Analyt TechnolBiomed Life Sci. 2002;772:147–153. (3) Fidder A, Noort D, Hulst AG, de Jong LP, Benschop HP. Biomonitoring of exposure to lewisitebased on adducts to haemoglobin. Arch Toxicol. 2000;74:207–214.730