The mechanism of the microbial hydroxylation of steroids. Part 4. The C-6 β hydroxylation of androst-4-ene-3,17-dione and related compounds by Rhizopusarrhizus ATCC 11145

Abstract

The steroid analogue 4,4a,5,6,7,8-hexahydro-2(3H)-naphthalenone was hydroxylated at C-8α, C-8β, and C-4a by Rhizopusarrhizus. Similar products were obtained by peracid oxidation of the corresponding enol ethers: hydroxylation of estr-4-ene-3,17-dione by the same fungus occurred at the analogous C-6 and C-10 positions. These results are consistent with a mechanism of microbial hydroxylation involving the enol form of the Δ4-3-ketone. Data from the incubations with R. arrhizus of androst-4-ene-3,17-dione specifically labelled with deuterium at C-4, C-6α, or C-6β and from those of other deuterium labelled substrates have been interpreted in terms of a mechanism of C-β hydroxylation involving a rate-determining step before enolization of the ketone, followed by rapid enolization and oxidation of the enol to give the 6β-hydroxy-Δ4-3-ketone. The kinetic isotope effect, kH/kD, for the hydroxylation of androst-4-ene-3,17-dione at C-6β has been found to be 1.2 ± 0.1.