AbstractStudies were carried out to evaluate the effects of cadmium in vitro on microsomal steroid metabolism in the inner (zona reticularis) and outer (zona fasciculata and zona glomerulosa) zones of the guinea pig adrenal cortex. Microsomes from the inner zone have greater 21‐hydroxylase than 17α‐hydroxylase activity, resulting in the conversion of progesterone primarily to 11‐deoxycorticosterone and of 17α‐hydroxy progesterone principally to its 21‐hydroxylated metabolite, 11‐deoxycortisol. Microsomes from the outer zones, by contrast, have far greater 17α‐hydroxylase and C17,20‐lyase activities than 21‐hydroxylase activity. As a result, progesterone is converted primarily to its 17‐hydroxylated metabolite, 17α‐hydroxyprogesterone; and 17α‐hydroxyprogesterone is converted principally to δ4‐androstenedione, with only small amounts of 21‐hydroxylated metabolites being produced. Addition of cadmium to incubations with inner zone microsomes causes concentration‐dependent decreases in 21‐hydroxylation and increases in 17α‐hydroxylase and C17,20‐lyase activities, resulting in a pattern of steroid metabolism similar to that in normal outer zone microsomes. Cadmium similarly decreases 21‐hydroxylation by outer zone microsomes but has no effect on the formation of 17‐hydroxylated metabolites or on androgen (Δ4‐androstenedione) production. In neither inner nor outer zone microsomes did cadmium affect cytochrome P‐450 concentrations, steroid interactions with cytochrome(s) P‐450, or NADPH–cytochrome P‐450 reductase activities. The results indicate that cadmium produces both quantitative and qualitative changes in adrenal microsomal steroid metabolism and that the nature of the changes differs in the inner and outer adrenocortical zones. In inner zone microsomes, there appears to be a reciprocal relationship between 21‐hydroxylase and 17α‐hydroxylase/C17,20‐lyase activitie