Diets rich in citrus and citrus-based products have been negatively correlated with the risk of cardiovascular disease, but so far no studies have been conducted to determine whether naringenin and hesperetin, two major flavanones in citrus plants, influence endothelium nitric oxide (NO) production. The aim of this study is to clarify estrogenic activities of naringenin and hesperetin and to examine whether they affect endothelial NO production via estrogen receptor (ER) activation. Both naringenin and hesperetin were observed to promote growth of MCF-7 cells under greatly reduced estrogen conditions and to suppress estrogen-induced response. Naringenin activated both ERalpha and ERbeta, whereas hesperetin exhibited stronger potential to activate ERalpha rather than ERbeta. Hesperetin, but not naringenin, increased NO releases from human umbilical vein endothelial cells in a dose-dependent manner. Hesperetin-induce responses were suppressed by ICI 182 780 and actinomycin D. Real-time reverse transcription polymerase chain reaction (RT-PCR) and western-blotting analysis revealed that hesperetin up-regulated endothelium nitric oxide synthase (eNOS) expression. These results suggested that hesperetin exerts an antiatherogenic effect, in part, via ER-mediated eNOS expression and subsequent increase of endothelial NO production. Distinct effects of naringenin and hesperetin on NO production also imply that ERalpha might play the major role in estrogen-induced eNOS expression. However, the inefficacy of naringenin on NO production remains to be elaborately studied. Our findings add more proof to the molecular explanations for the health benefits of citrus used to prevent cardiovascular disease, especially for postmenopausal women.