In the present investigation, we studied the inhibitory effects of three tea catechins [catechin, epicatechin, and (-)-epigallocatechin-3-O-gallate] and two bioflavonoids (quercetin and fisetin) on the O-methylation of 2- and 4-hydroxyestradiol (2-OH-E(2) and 4-OH-E(2), respectively) by human liver cytosolic catechol-O-methyltransferase (COMT). We found that catechin and epicatechin each inhibited the O-methylation of 2-OH-E(2) and 4-OH-E(2) in a concentration-dependent manner. The IC(50) values for inhibition of 2-OH-E(2) methylation by catechin and epicatechin were 14 to 17 microM and 44 to 65 microM, respectively, and their IC(50) values for inhibition of 4-OH-E(2) methylation were 5 to 7 microM and 10 to 18 microM, respectively. Our data showed that these two catechins had 2- to 6-fold higher inhibition potency for the O-methylation of 4-OH-E(2) than for the O-methylation of 2-OH-E(2). (-)-Epigallocatechin-3-O-gallate was found to have a distinctly high inhibition potency for the O-methylation of 2- and 4-OH-E(2) (IC(50) values of 0.04-0.07 microM and 0.2-0.5 microM, respectively). The crude extracts from green tea and black tea also showed very strong activity in inhibiting human liver COMT-mediated O-methylation of catechol estrogens. We also determined, for comparison, two common bioflavonoids (quercetin and fisetin) for their inhibitory effects on human liver COMT-mediated O-methylation of catechol estrogens. The IC(50) values for quercetin and fisetin were 0.9 to 1.5 microM and 3.3 to 4.5 microM, respectively, for inhibiting the O-methylation of 2-OH-E(2), and 0.5 to 1.2 microM and 2.6 to 4.2 microM, respectively, for inhibiting the O-methylation of 4-OH-E(2). Enzyme kinetic analyses showed that both tea catechins and bioflavonoids inhibited human liver COMT-mediated O-methylation of 4-OH-E(2) (a representative substrate) with a mixed mechanism of inhibition (competitive plus noncompetitive). In summary, the catechol-containing tea catechins and bioflavonoids are strong inhibitors of human liver COMT-mediated O-methylation of catechol estrogens. More studies are warranted to determine the extent of such inhibition in human subjects and the potential biological consequences.