ABSTRACT Purpose: Since sexual dysfunction related to vas deferens smooth muscle contractility is a possible side effect of St. John’s wort (SJW) (Hypericum perforatum) we evaluated the effect of this herbal antidepressant on rat and human vas deferens contractility. Materials and Methods: The effect of SJW was evaluated on contractions induced by electrical field stimulation or exogenous agonists (α,β-methylene adenosine triphosphate and phenylephrine) in isolated rat and human vas deferens. Results: SJW (1 to 300 μM) decreased in a concentration dependent manner the amplitude of electrical field stimulation and agonist induced contractions with the same potency, suggesting direct inhibition of rat vas deferens smooth muscle. Of the chemical constituents of SJW tested hyperforin but not hypericin or the flavonoids quercitrin, rutin and kaempferol inhibited phenylephrine induced contractions. SJW and hyperforin also inhibited phenylephrine induced contractions in human vas deferens Conclusions: The results of our study demonstrate that SJW directly inhibits rat and human vas deferens contractility. If confirmed in vivo, these results suggest that SJW might affect sexual function in humans. These results might explain delayed ejaculation described in patients receiving SJW.