Enantioselective in-vitro elimination kinetics of nerve agents in blood monitored by derivatization and LC-MS/MS...

We present a simple method for chiral separation and analysis of organophosphorus nerve agents and apply it to monitor the enantioselective blood elimination kinetics of sarin in-vitro. The method is implemented in standard reverse phase LC-MS operating conditions, relieving the user of the dedicated operating conditions frequently demanded in chiral LC-MS analysis. The method consists of formation of diastereomers by a rapid derivatization with (R)-2-(1 aminoethyl) phenol, followed by LC-MS/MS analysis. Derivatization enantioselectivity was studied by comparing the reaction of optically pure sarin and racemic sarin, proving no substantial enantiomeric preference in the reaction and demonstrating the enantiomeric discrimination abilities of the technique. Enantioselective sarin elimination pathways were probed in-vitro by following the fast elimination kinetics of the two sarin enantiomers as well as its hydrolysis metabolite (isopropyl methyl-phosphonic acid, IMPA) in whole blood and plasma compared to water. Sarin enantiomers showed the known marked differences in elimination kinetics with rapid elimination of the (+) enantiomer and slower elimination of the (-) enantiomer in whole blood and plasma as well as dose-dependent kinetics (faster elimination at lower concentrations). We found that small amounts of acetonitrile in plasma prevent the rapid elimination of the (+) enantiomer, resulting in similar, slower elimination kinetics for both enantiomers.