Record Details

A colorimetric assay method for 3beta-hydroxy-delta5-steroid dehydrogenase.

IR@CSIR-CFTRI

View Archive Info
 
 
Field Value
 
Relation http://ir.cftri.com/2211/
 
Title A colorimetric assay method for 3beta-hydroxy-delta5-steroid dehydrogenase.
 
Creator Shivanandappa, T.
Venkatesh, S.
 
Subject 16 Enzyme Chemistry
05 Calorimetric Analysis
 
Description 3beta-Hydroxy-Delta5-steroid dehydrogenase is an important enzyme of steroid hormone biosynthesis present in steroidogenic tissues like adrenal, testis, and ovary of vertebrates. The enzyme is assayed mainly by radiochemical substrates. Spectrophotometric assay is not adequately sensitive to detect the enzyme activity since it is often present in low levels. We have developed a simple colorimetric assay based on formazan formation due to the reduction of the tetrazolium salt. The reaction mixture containing the substrate, pregnenolone or dehydroepiandrosterone, NAD and iodonitrotetrazolium in 0.1 M Tris-HCl buffer (pH 7.8), and the enzyme extract is incubated for 1 h at 37 degrees C. Absorbance at 490 nm is read in a spectrophotometer. The enzyme activity was linear with time and protein concentration. The assay works well with adrenal tissue extract, whereas in the case of testis, Leydig cell preparation may be required. We have assayed the enzyme activity in the adrenal of rat, mouse, and gerbil. The method is two- to threefold more sensitive than the spectrophotometric assay.
 
Date 1997
 
Type Article
PeerReviewed
 
Format application/pdf
 
Language en
 
Rights —
 
Identifier http://ir.cftri.com/2211/1/Analytical%20Biochemistry%2C%20Volume%20254%2C%20Issue%201%2C%201%20December%201997%2C%20Pages%2057-61.pdf
Shivanandappa, T. and Venkatesh, S. (1997) A colorimetric assay method for 3beta-hydroxy-delta5-steroid dehydrogenase. Analytical Biochemistry, 254 (1). pp. 57-61. ISSN 0003-2697