A rapid and sensitive technique for assessing exposure to VX via GC-MS-MS analysis.

A rapid and sensitive method for the determination of the chemical warfare agent VX in plasma taken from Göttingen minipigs has been developed using isotope-dilution gas chromatography-tandem mass spectrometry (GC-MS-MS). Chromatographic separation was achieved on a 5% diphenyl/95% dimethyl polysiloxane capillary column with a total run time of about 11 min. The analyte was detected using ammonia chemical ionization in the multiple reaction monitoring mode, following a simple extraction with 10% 2-propanol in hexane. A good linear relationship was obtained in the quantitative concentration range of 10 ng/mL to 1000 ng/mL (r(2) = 0.9998) with an average slope of 1.275 +/- 0.037 (n = 7), and an absolute detection limit of 0.4 pg on column. The average recovery for VX was 95% in saline in the concentration range of 50-100 ng/mL. The method was successfully applied to the analysis of VX in minipig plasma in a preliminary toxicokinetic study.

Validation and application of a GC-MS method for determining soman concentration in rat plasma following low-level vapor exposure.

A method for determining the chemical warfare agent soman (GD) in rat plasma has been validated and applied to low-level inhalation exposure studies currently being conducted. This method utilizes a fluoride ion-based regeneration assay with isotope dilution followed by large volume injection gas chromatography with ammonia chemical ionization mass spectrometric detection. Following sample preparation by solid phase extraction, chromatographic separation was achieved using a 14% cyanopropylphenyl/86% dimethyl polysiloxane capillary column with a total run time of 18.16 min. Soman and the deuterated isotope ((2)H(4)-soman) internal standard were detected using the selected ion monitoring mode and quantitated using the ammonia adduction ratio of m/z ions 200/204. A reproducible linear relationship was obtained for the quantitative concentration range of 10 pg on-column to 1000 pg on-column (r(2) = 0.9995) for standards in ethyl acetate with a detection limit of 5.65 pg on-column, and an average recovery of 93% in plasma. This sensitive method was successfully applied to the analysis of soman in rat plasma immediately post-exposure, resulting in the construction of dose-response plots.

Determination of VX-G analogue in red blood cells via gas chromatography-tandem mass spectrometry following an accidental exposure to VX.

A sensitive method for determining exposure to the chemical warfare agent VX is described in which the biomarker ethyl methylphosphonofluoridate (VX-G) is measured in red blood cells (RBCs) following treatment with fluoride ion using isotope-dilution gas chromatography-tandem mass spectrometry. The analyte was isolated via solid-phase extraction and detected using ammonia chemical ionization in the multiple reaction monitoring mode. A good linear relationship was obtained in the quantitative concentration range of 4 ng/mL to 1000 ng/mL with an absolute detection limit of < 1 pg on column. The method has been applied to the analysis of RBCs from a laboratory worker accidentally exposed to VX vapor. Detection and quantitation of VX-G were possible in samples taken as late as 27 days following exposure.

Quantification of nerve agent VX-butyrylcholinesterase adduct biomarker from an accidental exposure.

The lack of data in the open literature on human exposure to the nerve agent O-ethyl-S-(2-diisopropylaminoethyl) methylphosphonothioate (VX) gives a special relevance to the data presented in this study in which we report the quantification of VX-butyrylcholinesterase adduct from a relatively low-level accidental human exposure. The samples were analyzed by gas chromatography-high resolution mass spectrometry using the fluoride ion regeneration method for the quantification of multiple nerve agents including VX. Six human plasma samples from the same individual were collected after the patient had been treated once with oxime immediately after exhibiting signs of exposure. Detection limits of approximately 5.5 pg/mL plasma were achieved for the G-analogue of VX (G-VX). Levels of the G-VX ranged from 81.4 pg/mL on the first day after the exposure to 6.9 pg/mL in the sample taken 27 days after the exposure. Based on the reported concentration of human butyrylcholinesterase in plasma of approximately 80 nM, it can be calculated that inhibition levels of >or= 0.05% of BuChE can be accurately quantified. These data further indicate that the fluoride ion regeneration method is a potentially powerful tool that can be used to assess low-level exposure to VX.

Kinetics of sarin (GB) following a single sublethal inhalation exposure in the guinea pig.

To improve toxicity estimates from sublethal exposures to chemical warfare nerve agents (CWNA), it is necessary to generate mathematical models of the absorption, distribution, and elimination of nerve agents. However, current models are based on representative data sets generated with different routes of exposure and in different species and are designed to interpolate between limited laboratory data sets to predict a wide range of possible human exposure scenarios. This study was performed to integrate CWNA sublethal toxicity data in male Duncan Hartley guinea pigs. Specific goal was to compare uptake and clearance kinetics of different sublethal doses of sarin (either 0.1 x or 0.4 x LC50) in blood and tissues of guinea pigs exposed to agent by acute whole-body inhalation exposure after the 60-min LC50 was determined. Arterial catheterization allowed repeated blood sampling from the same animal at various time periods. Blood and tissue levels of acetylcholinesterase, butyrylcholinesterase, and regenerated sarin (rGB) were determined at various time points during and following sarin exposure. The following pharmacokinetic parameters were calculated from the graph of plasma or RBC rGB concentration versus time: time to reach the maximal concentration; maximal concentration; mean residence time; clearance; volume of distribution at steady state; terminal elimination-phase rate constant; and area under plasma concentration time curve extrapolated to infinity using the WinNonlin analysis program 5.0. Plasma and RBC t(1/2) for rGB was also calculated. Data will be used to develop mathematical model of absorption and distribution of sublethal sarin doses into susceptible tissues.