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Appl Environ Microbiol. 1975 October; 30(4): 530-535
Copyright © 1975 American Society for Microbiology. All Rights Reserved.

Convenient Non-Chromatographic Assays for the Microbial Deconjugation and 7-OH Bioconversion of Taurocholate

Departments of Medicine and Microbiology, Dalhousie University, Halifax, Nova Scotia, Canada

ABSTRACT

We described two convenient assay methods to estimate bile acid deconjugation and bile acid bioconversion at the 7-OH position by individual microorganisms grown in media containing taurocholic acid. The methods are based on (i) a selective chemical assay for taurine conjugates previously described and (ii) the use of a cell-free preparation of 7-hydroxysteroid dehydrogenase from Escherichia coli to directly quantify 7-OH groups. These non-chromatographic approaches have been applied to the study of three model strains of intestinal organisms, E. coli, Bacteriodes fragilis, and Clostridium perfringens, grown in standard media in the presence of purified tritiated taurocholate. Assay results were confirmed by thin-layer chromatography solvent systems designed to separate conjugated from unconjugated bile acid and unmodified cholic acid nucleus from 7-OH bioconversion product(s) (primarily 3,12 dihydroxy, 7-keto-cholanoic acid). In addition, 7-hydroxysteroid dehydrogenase activity was demonstrated in cell-free extracts of all three organisms. Of the three organisms, only C. perfringens was demonstrated to (i) deconjugate taurocholic acid, (ii) contain 3-hydroxysteroid dehydrogenase activity, (iii) convert cholic acid into at least five labeled metabolites visible on thin-layer chromatography, and (iv) catalyze significant tritium exchange with water in the medium.