Influenza virus entry and replication inhibited by 8‐prenylnaringenin from Citrullus lanatus var. citroides (wild watermelon)

We previously demonstrated the anti‐influenza activity of Citrullus lanatus var. citroides (wild watermelon, WWM); however, the active ingredient was unknown. Here, we performed metabolomic analysis to evaluate the ingredients of WWM associated with antiviral activity. Many low‐molecular weight compounds were identified, with flavonoids accounting for 35% of all the compounds in WWM juice. Prenylated flavonoids accounted for 30% of the flavonoids. Among the measurable components of phytoestrogens in WWM juice, 8‐prenylnaringenin showed the highest antiviral activity. We synthesized 8‐prenylnaringenin and used liquid chromatography–mass spectrometry to quantitate the active ingredient in WWM. The antiviral activities of 8‐prenylnaringenin were observed against H1N1 and H3N2 influenza A subtypes and influenza B viruses. Moreover, 8‐prenylnaringenin was found to inhibit virus adsorption and late‐stage virus replication, suggesting that the mechanisms of action of 8‐prenylnaringenin may differ from those of amantadine and oseltamivir. We confirmed that 8‐prenylnaringenin strongly inhibited the viral entry of all the influenza virus strains that were examined, including those resistant to the anti‐influenza drugs oseltamivir and amantadine. This result indicates that 8‐prenylnaringenin may activate the host cell's defense mechanisms, rather than directly acting on the influenza virus. Since 8‐prenylnaringenin did not inhibit late‐stage virus replication of oseltamivir‐resistant strains, 8‐prenylnaringenin may interact directly with viral neuraminidase. These results are the first report on the anti‐influenza virus activity of 8‐prenylnaringenin. Our results highlight the potential of WWM and phytoestrogens to develop effective prophylactic and therapeutic approaches to the influenza virus.