An improved method for the determination of cytoplasmic 3 alpha- and 17 beta- and microsomal 3 alpha-hydroxysteroid dehydrogenase activities in rat liver.

This paper described a modified method for the radiometric determination of hydroxysteroid dehydrogenase activities in rat liver. The principle advantages of this method are the improved precision and a radical reduction in the time involved in performing the assay. The procedure comprises the following steps: incubation of 14C-labelled substrate with coenzyme and cell fraction under optimized conditions; termination of the reaction by addition of organic solvent containing a defined amount of 3H-labelled reaction product; removal of precipitated protein and coenzyme by centrifugation; paper chromatographic isolation of the product; direct quantitation of 14C activity in the product zone of the paper chromatogram. The assay systems have been applied to elucidate and quantitate sex and strain differences in the activities of the above enzymes in Chbb/THOM and Sprague-Dawley rats.