Theoretical kinetics of sequential metabolism in vitro. Study of the formation of 16 alpha-hydroxyandrostenedione from testosterone by purified rat P450 2C11
Sugiyama, K.; Nagata, K.; Gillette, J.R.; Darbyshire, J.F.
Drug Metabolism and Disposition the Biological Fate of Chemicals 22(4): 584-591
1994
ISSN/ISBN: 0090-9556 PMID: 7956734 Document Number: 422711
P450 2C11 from rat liver is known to metabolize testosterone to 2 alpha-, 16 alpha-, and 6 beta-hydroxytestosterone, and to androstenedione and 16 alpha-hydroxyandrostenedione. Because Waxman (J. Biol. Chem. 259, 15481-15490) has reported that the enzyme converts androstenedione to 16 alpha-hydroxyandrostenedione, it seemed likely that the metabolite was formed from testosterone by way of androstenedione. Indeed, we have found that P450 2C11 does not convert 16 alpha-hydroxytestosterone to 16 alpha-hydroxyandrostenedione to any significant extent and, therefore, that the metabolite is formed from testosterone almost solely by way of androstenedione. To determine whether some of the 16 alpha-hydroxyandrostenedione might be formed directly from the androstenedione-enzyme complex, we developed an approach by which it is possible to calculate the amount of the androstenedione, released into the medium, relative to the amount of the androstenedione-enzyme complex that is converted directly to 16 alpha-hydroxyandrostenedione under initial conditions when the concentration of released androstenedione will be negligible. The approach uses two factors: factor A is the androstenedione/(androstenedione + 16 alpha-hydroxyandrostenedione) present at the end of the incubation, and factor B corrects for the amount of released androstenedione that recombines with the enzyme and is converted to 16 alpha-hydroxyandrostenedione. Although the values of both factors A and B will vary with the concentrations of testosterone and preformed androstenedione present in the incubation mixtures and with the duration of incubation, the value of A*B will be independent of these parameters.