Microbial metabolism of steviol and steviol-16α,17-epoxide

Abstract

Steviol ( 2) possesses a blood glucose-lowering property. In order to produce potentially more- or less-active, toxic, or inactive metabolites compared to steviol ( 2), its microbial metabolism was investigated. Incubation of 2 with the microorganisms Bacillus megaterium ATCC 14581, Mucor recurvatus MR 36, and Aspergillus niger BCRC 32720 yielded one new metabolite, ent-7α,11β,13-trihydroxykaur-16-en-19-oic acid ( 7), together with four known related biotransformation products, ent-7α,13-dihydroxykaur-16-en-19-oic acid ( 3), ent-13-hydroxykaur-16-en-19-α- d-glucopyranosyl ester ( 4), ent-13,16β,17-trihydroxykauran-19-oic acid ( 5), and ent-13-hydroxy-7-ketokaur-16-en-19-oic acid ( 6). The preliminary testing of antihyperglycemic effects showed that 5 was more potent than the parent compound ( 2). Thus, the microbial metabolism of steviol-16α,17-epoxide ( 8) with M. recurvatus MR 36 was continued to produce higher amounts of 5 for future study of its action mechanism. Preparative-scale fermentation of 8 yielded 5, ent-11α,13,16α,17-tetrahydroxykauran-19-oic acid ( 10), ent-1β,17-dihydroxy-16-ketobeyeran-19-oic acid ( 11), and ent-7α,17-dihydroxy-16-ketobeyeran-19-oic acid ( 13), together with three new metabolites: ent-13,16β-dihydroxykauran-17-acetoxy-19-oic acid ( 9), ent-11β,13-dihydroxy-16β,17-epoxykauran-19-oic acid ( 12), and ent-11β,13,16β,17-tetrahydroxykauran-19-oic acid ( 14). The structures of the compounds were fully elucidated using 1D and 2D NMR spectroscopic techniques, as well as HRFABMS. In addition, a GRE (glucocorticoid responsive element)-mediated luciferase reporter assay was used to initially screen the compounds 3– 5, and 7 as glucocorticoid agonists. Compounds 4, 5 and 7 showed significant effects.