[19-14C]androstenedione: a new substrate for assaying aromatase and studying its reaction mechanism

[19-14C]Androstenedione has been prepared and utilized as a substrate for assaying microsomal human placental aromatase. Enzyme activity is determined by measuring the rate at which [14C]formate is produced by aromatization of this 14C-labeled steroid. Isotope ratio experiments using [19-14C]androstenedione and [1 beta-3H]androstenedione demonstrate that an apparent kinetic hydrogen isotope effect exists for the aromatization of the tritiated steroid with kH/kT approximately 1.09. Metabolic switching occurs to a minor extent (approximately 3%) during aromatization of [1 beta-3H]androstenedione, but not during the aromatization of [19-14C]androstenedione.