Chromatographic study on the specificity of bis- p-nitrophenylphosphate in vivo Identification of labelled proteins of rat liver after intravenous injection of [formula omitted] as carboxylesterases and amidases

Abstract

The procedure established to isolate the carboxylesterase E 1, E 2 and EA from rat liver (Arndt, R. and Krisch K. (1972) Hoppe-Seyler's Z. Physiol. Chem. 353, 589–598) was applied to characterize in vivo bis-p- nitro[ 14C]phenyl -P- labelled proteins. The peaks of radioactivity and of residual enzyme activities (hydrolysing methylbutyrate, p-nitrophenylacetate and acetanilide) were found in the same peaks after column chromatography and could be related to the well-defined esterases E 1, E 2 and EA. There is no indication of a nonspecific binding of bis- p-nitrophenyl- P or of one of its metabolites. The relative quantitative amounts of E 1, E 2 and EA were calculated to represent 40, 14 and 46%, respectively, of the total carboxylesterase content of rat liver. The relative amount of bound (not dialysable) radioactivity in rat liver depended on the survival time. During purification, the yield of enzyme activities corresponded to that of bound radioactivity, confirming the specificity of bis- p-nitrophenyl- P in vivo. Hence the radioactive metabolites of the inhibitor obviously do not possess binding affinities of quantitative importance to the rat liver proteins.