Studies with the Hydroxysteroid Dehydrogenases: I. A Simplified Method for the Enzymatic Estimation of 3- and 17-Hydroxysteroids

Abstract

A simplified method for the estimation of [alpha] and [beta] substrates of the hydroxysteroid dehydrogenases using the enzymes recovered from the acetone powder of Pseudomonas testosteroni is described. In terms of sensitivity, accuracy, reproducibility, simplicity and speed, the method offers significant advantage over most techniques for the assay of steroids separated chromatographically, and is particularly valuable in the determination of certain steroids for which no satisfactory colorimetric method is available. p-Hydroxymercuribenzoate is a potent inhibitor of the a-hydroxysteroid dehydrogenase activity when added to the assay system over a wide range of concentrations. This selective inhibition of the [alpha]-activity allows the determination of [beta]-hydroxyl functions in a mixture of steroids and provides structural information previously requiring complete separation of the enzymes.