The major 17 alpha-ethinyl estradiol 2-hydroxylase is humans is the hepatic enzyme cytochrome P-450 IIIA4 (P-450NF), which is known to be inducible by rifampicin or barbiturates. The literature indicates that 17 beta-estradiol, progesterone, and norgestrel are competitive inhibitors and that primaquine and tolbutamide are rather weak noncompetitive inhibitors. Recent experiments in this laboratory indicate that gestodene is a relatively potent mechanism-based inactivator of cytochrome P-450 IIIA4 in vitro. Inhibition requires incubation with the reduced form of nicotinamide adenine dinucleotide phosphate, is time and concentration dependent, and can be partially blocked by the presence of noninhibitory cytochrome P-450 IIIA4 substrates. The in vitro activation by gestodene provides a possible explanation for the increase in plasma estrogen levels reported in women administered gestodene along with 17 alpha-ethinyl estradiol.