Allicin selectively enhances the fungicidal activity of amphotericin B (AmB). It also accelerates AmB-induced vacuole disruption but does not affect AmB-induced potassium ion efflux in Saccharomyces cerevisiae and Candida albicans. The fungicidal activity of AmB alone or combined with allicin was further evaluated based on the relationship among cell viability, vacuole disruption and potassium ion efflux in S. cerevisiae. Lethality and vacuole disruption caused by AmB alone were completely restricted when K(+) and Mg(2+) were added to the growth medium. On the other hand, in identical conditions, the combination of AmB and allicin induced both lethality and vacuole disruption. S. cerevisiae Δerg6 cells, which lack ergosterol in plasma membrane, were mostly resistant to AmB as well as the combination of AmB and allicin against both lethality and vacuole disruption. The incorporation of AmB into the cytoplasm of Δerg6 cells was significantly reduced in comparison with that in parent cells, regardless of the presence of allicin. Our results suggest that the fungicidal activity of AmB combined with allicin is involved in vacuole disruption but not in potassium ion efflux, and that the expression of allicin-mediated activity of AmB requires the presence of ergosterol in the plasma membrane.