The beneficial cardiovascular effect of polyphenol-rich food and beverages has been attributed, at least in part, to the improvement of the vascular function through the induction of two major endothelial vasoprotective mechanisms. The aim of the present study is to determine whether a polyphenol-rich blackcurrant juice (PRBJ, 2.7 g/l), improves the vasoprotective endothelial function, and, if so, to characterize the underlying mechanism. The reactivity of porcine coronary artery rings was assessed in organ chambers, and the expression and phosphorylation levels of proteins in cultured porcine coronary endothelial cells by Western blot analysis. PRBJ caused potent endotheliumdependent relaxations that were significantly reduced by an eNOS inhibitor, not affected by inhibition of endothelium-dependent hyperpolarization, and abolished by both treatments. PRBJ-induced NO-mediated relaxations were significantly reduced by chelators of either copper or iron, membrane permeant analogues of superoxide dismutase and catalase, inhibitors of either Src or PI3-kinase, and by calmidazolium, a calmodulin inhibitor. The NO-mediated relaxation was not affected by inhibitors of either PKC, EGFR, IGFR, or of several endogenous enzymes involved in the formation of ROS (NADPH oxidase, xanthine oxidase, mitochondrial respiration chain, cytochrome P450). In cultured endothelial cells, PRBJ increased the formation of NO as assessed by electron paramagnetic resonance spectroscopy. Moreover, PRBJ induced the phosphorylation of Akt and eNOS on activator sites, which were inhibited membrane permeant analogues of superoxide dismutase and catalase and inhibitors of either Src or PI3-kinase. PRBJ is a potent inducer of endothelium- dependent NO-mediated relaxations in porcine coronary artery rings. The NO-mediated relaxation involves an intracellular copper- and iron-dependent redox-sensitive activation the Src/PI3-kinase/Akt pathway leading to activation of eNOS and subsequent formation of NO.