Background: We previously described that 2ME have protective vascular effects in males and estrogen-deprived females (Bonacasa, 2008; Fenoy, 2010). Cathecol-O-methyltransferase (COMT) introduces a methyl group to 2-hydroxyestradiol to form 2-methoxyestradiol (2ME). Preeclampsia is a major obstetric problem and a significant source of maternal and fetal morbidity and mortality (Sibai, 2005), complicating an estimated 2% of all pregnancies(Duley, 2003). A key factor connecting the angiogenic imbalance with generalized endothelial dysfunction in preeclampsia has been suggested to be a reduction in the synthesis of 2ME (Kanasaki, 2008). Aim: The present study evaluated whether COMT inhibition in pregnant rats results in endothelial dysfunction as a consequence of a high levels of oxidative stress and a reduction of nitric oxide bioavailability. Methods: Pregnant Sprague Dawley rats were given entacapone (a COMT inhibitor) by gavage from the 10th to the 20th day of pregnancy at a dose of 150 mg/kg.day. Endothelium-dependent relaxation of pressurized mesenteric small arterial vessels was assessed in the absence and presence of 50 µM tempol (a superoxide anion scavenger). Nitric oxide release in aorta by electrochemical NO measurements in response to a calcium ionophore (A23187) was assessed in the absence and presence of tempol as well. Results: Vessels from entacapone-treated pregnant rats had diminished relaxations to acetylcholine compared with vehicle-treated pregnant rats. The presence of tempol improved endothelial function (Log EC50 : – 7.85 + 0.27 vs. – 6.96 + 0.13). Endothelial release of nitric oxide induced by A23187 was significantly greater in aortas from vehicle-treated pregnant rats (209.8 + 17 nM) than in pregnant rats given entacapone (159 + 11 nM), and was prevented by addition of tempol (279.17 + 27 vs 212.5 + 19 nM). Conclusion: The present study suggests an important role of superoxide anion in endothelial dysfunction observed in this COMT inhibition model of preeclampsia.