A spectrophotometric assay for pyrethroid-cleaving enzymes in human serum

Abstract

A direct continuous spectrophotometric assay to measure pyrethroid-cleaving enzymes in human serum was developed using cis- and trans-α-naphthyl-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylate ( cis- and trans-naphthyl-Cl2CA). These substrates show a structure very similar to the pyrethroids most often used (e.g. permethrin, cyfluthrin). The method is based on an increase in absorbance at 321 nm which occurs with the hydrolysis of the α-naphthyl esters to α-naphthol. The assay was optimised regarding type of buffer, pH and substrate concentrations, it was linear for at least 10 min at 37°C. These esterases were completely inhibited by bis-(4-nitrophenylphosphate), a specific carboxyesterase inhibitor. They displayed a great individual variability in human serum, activities were between less than 40 and 1000 U/l for cis-naphthyl-Cl2CA, between less than 40 and 2000 U/l for trans-naphthyl-Cl2CA, respectively. However, a correlation of enzyme activity to sex or age could not be observed. Furthermore, the activity of pyrethroid-cleaving esterases did not correspond to the activities of acylesterase, arylesterase, acetylcholinesterase or butyrylcholinesterase.