Estimation of inhibitory organophosphates with purified pig liver carboxylesterase

Abstract

Organophosphates that inhibit acetylcholineesterase normally also inhibit pig liver carboxylesterase irreversibly. Since this liver esterase is well characterized and easily accessible in large amounts, we propose the use of this enzyme for the quantitation of low concentrations of such organophosphates. The principle of two estimation methods is described. Both methods involve the addition of an unknown amount of organophosphate to an assay mixture of purified esterase, buffer and a low affinity esterase substrate. In the first of these methods, the inhibitor concentration is calculated from the esterase activities before and after the addition of the inhibitor. In the second method, the amounts of inhibitor or of enzyme are changed in several assays, until equimolar conditions can be detected from the observed reaction kinetics. The theoretical background of these methods is discussed and practical examples for the estimation of paraoxon (order of 0.1 nmoles) are given.