Organophosphates (OPs) have been in recent news with several emerging cases of nerve agent poisoning of significant personalities and national leaders. These small organic molecules act as potent acetylcholinesterase (AChE) inhibitors causing muscle contractions, blurry vision, seizures, respiratory failure and eventually death in severe exposure cases. To date, very little is known about the metabolism of these toxic compounds in humans and the aim of this study is explore the interaction of OPs with their major metabolizing enzymes, cytochrome P450 (CYPs/P450), inside the human body. To that extent we intend to determine the rate of metabolism of OPs and nerve agent simulants in human liver microsomes (HLMs) using 1D-NMR and LC-MS/MS metabolomics. NMR studies were completed with more than 20 OPs (acute, high dose exposure) and allowed for continuous monitoring across long time periods (up to 7 days) whereas approximately 50 OPs with a diverse range of structures were analyzed by LC-MS/MS (low dose exposure) for a period of at least 24 h. For example, the comparative analysis of the clearance rate of ³¹P NMR (at 5 mM) and MRM data (at 0.76 µM) of EMP, a nerve agent simulant of V_x, indicated that at higher concentration a two-phase decay of the simulant was observed compared to a monophasic decay observed at lower concentrations. This finding suggests two CYPs metabolizing EMP at higher concentration, one metabolizes EMP at a faster rate compared to the second CYP. These results drive the future studies including the identification of the different metabolites formed using 2D NMR, semi-targeted and untargeted metabolomics. Furthermore, these clearance rate values will be used for developing in silico metabolism models to enable predicting the likely metabolic pathways for newly emerging OP threats.

Clearance rate comparison of EMP (VX nerve agent simulant)

Garima Agarwal

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