In the present study, the effects of the bioflavonoid chrysin (5,7-dihydroxyflavone) were analysed on nitric oxide (NO) production from vascular endothelium. In aortic rings, incubation with chrysin or acetylcholine (both at 10 microM) increased L-NAME-sensitive endothelial NO release as measured using the fluorescent probe 4,5-diaminofluorescein diacetate (DAF-2 DA). Moreover, chrysin increased cGMP accumulation only in aortic rings with endothelium. However, at this concentration, chrysin had no effect either on basal or on NADPH-stimulated vascular superoxide production. Moreover, at this low concentration, chrysin, similar to acetylcholine, induced aortic relaxation, which was abolished by both endothelial deprivation and NO synthase inhibition. Endothelium-dependent relaxation induced by chrysin was unaltered by removal of extracellular calcium and incubation with the intracellular calcium chelator BAPTA, while the phosphatidylinositol (PI)-3 kinase inhibitor wortmannin suppressed the endothelial dependence. In conclusion, chrysin stimulated NO release from endothelial cells leading to vascular cGMP accumulation and subsequent endothelium dependent aortic relaxation. Chrysin-stimulated NO release is calcium independent and possibly mediated via PI3-kinase.