The interactions of the azole antifungal agents fluconazole, itraconazole, ketoconazole, or miconazole with amphotericin B (AmB) in their effect on Candida albicans were investigated. These four azoles antagonized the fungistatic activity of AmB at sub-MICs if both substances acted simultaneously. This coincubation test was primarily developed to observe the azole-mediated demethylase inhibition quantitatively by bioassay. Interestingly, the occurrence of azole-AmB antagonism depended on azole lipophilia if specially selected test conditions were applied. By a consecutive incubation regimen, preincubation at high azole concentrations (1 to 50 g/ml) and then subsequent incubation with AmB (1 g/ml), only preincubation with the three lipophilic azoles decreased the fungicidal activity of AmB but not that of FCZ. It was shown that yeasts absorb only lipophilic azoles to a remarkable extent. This fact might be responsible for the absence of antagonism of FCZ to AmB when yeasts were incubated consecutively. It can be concluded with caution that consecutive treatment of candidiasis with FCZ and AmB does not necessarily result in a clinically relevant antagonism.Alterations of the effects of amphotericin B (AmB) on pathogenic fungi by azoles were observed repeatedly in vivo as well as in vitro. The majority of reports mention antagonisms (2-4, 6, 7, 11, 12, 16, 17), but indifference (1, 10, 18) and synergism (10, 14, 18) have also been described. The results were influenced by a variety of factors, especially experimental conditions as well as the particular fungal strains or species. The fact that the results depend on the properties of antifungal agents has been described sporadically (12). At first, a postexposure antagonism of azoles to AmB was assumed. In experiments dealing with this question, it was found that lipophilic azoles are able to influence the fungicidal activity of subsequently applied AmB on yeasts but not that of fluconazole (FCZ). To detect this behavior of lipophilic azoles, special test conditions (unusually high drug concentrations as well as a high density of organisms) had to be selected. Decreasing both inoculum size and antifungal concentration resulted in a failure to detect an influence of azoles on the fungicidal activity of AmB. For this aim of the investigation, the test conditions had to be very different from those that work with drug concentrations similar to those found in vivo, that is, sub-MICs, to observe postantifungal effects (9) with possible clinical significance.Additional tests that were performed in order to explain the dependence of the long-lasting effects of azoles on their lipophilicities also required large quantities of test organisms.The results presented in this report focus on a further aspect of the azole-AmB antagonism. The antagonism between azoles and AmB has been explained by the inhibition of ergosterol synthesis and then a disappearance of ergosterol from the cell membrane (12,13,17,19). The affinity of AmB for ergosterol precursors is not as high as that fo...