The effect of pteridines and sex hormones on hydroxyindole-O-methyltransferase (HIOMT) catalysis was examined. Equivalent quantities of partially purified bovine HIOMT was incubated in the presence and absence of the various test substances and the results fitted to the Michaelis-Menten equation. The pteridines (triampterene and xanthopterin) had no noticeable effect on HIOMT catalysis, while all the sex hormones examined were found to be competitive inhibitors of the enzyme with respect to the hydroxy-substrate. Testosterone concentrations causing noticeable inhibition were above physiological while estrone, estradiol, and progesterone all caused inhibition within the physiological range; estrone was the most potent and progesterone the least potent inhibitor. The possible significance of these findings are discussed.