Microbial hydroxylation of pregnenolone derivatives

Pregnenolone (1) and pregnenolone acetate (2) were incubated with the fungi Cunninghamella elegans, Rhizopus stolonifer and Gibberella fujikuroi. Incubation of 1 with C elegans yielded metabolites, 3 beta,6 beta,11 alpha-trihydroxypreg-5-en-20-one (3), 3 beta,6 alpha,11 alpha,12 beta,15 beta-pentahydroxypreg-4-en-20-one (4) and 3 beta,6 beta,11 alpha-trihydroxypreg-4-en-20-one (5), while incubation with G. fujikuroi yielded two known metabolites, beta,7 beta-dihydroxypregn-5en-20-one (6) and 6 beta,15 beta-dihydroxypreg-4-ene-3,20-dione (7). Metabolites 4 and 5 were found to be new. Fermentation of 2 by C elegans yielded four known oxidative metabolites, 1, androsta-1,4-diene-3,17-dione (8), 6 beta,15 beta-dihydroxyandrost-4-ene-3,17-dione (9) and 11 alpha,15 beta-dihydroxypreg-4-ene-3,20-dione (10). Fermentation of 2 with R. stolonifer yielded two known metabolites, 11 alpha-hydroxypreg-4-ene-3,20-dione (11) and 7. Compounds 1-11 were screened for their cholinesterase inhibitory activity in a mechanism-based assay.