The strong anti-hyaluronidase effect of ellagic acid markedly decreases polyspermy during in vitro fertilization, resulting in sustainment of the developmental potency in porcine oocytes.

The present study investigated the effects of ellagic acid, a type of polyphenol that does not have a glycan and is composed of four hydroxyl groups and two lactone functional groups, on porcine in vitro fertilization (IVF) by focusing on its anti-hyaluronidase activity. A comparative analysis of ellagic acid and apigenin, which is commonly used as a hyaluronidase inhibitor, was performed. It compared the effects of ellagic acid and apigenin on hyaluronidase activity at different concentrations. The results showed that 10, 20, and 40 μM ellagic acid strongly reduced hyaluronidase activity (P < 0.05). The addition of 20 μM ellagic acid, but not apigenin, to porcine IVF medium effectively reduced polyspermy without decreasing sperm penetration or the formation rates of male pronuclei in cumulus-free oocytes. However, neither ellagic acid nor apigenin affected the number of sperm that bound to zona pellucida (ZP) or the induction of zona hardening and protease resistance. The percentage of acrosome-reacting sperm that bound to the ZP was markedly lower in the presence of 20 μM ellagic acid than in the untreated and apigenin-treated groups, even though the antioxidant capacity of ellagic acid was weaker than that of apigenin. Furthermore, a markedly higher percentage of embryos developed to the blastocyst stage in the ellagic acid-treated group, and the apoptotic indexes of expanded blastocysts produced by the ellagic acid treatment during IVF were significantly low. Therefore, the anti-hyaluronidase effect of ellagic acid markedly suppressed the induction of the acrosome reaction in sperm that bound to the ZP, resulting in a marked decrease in polyspermy under conditions that maintained high sperm penetrability during IVF and sustainment of the developmental potency in porcine oocytes.