Synergistic Fungicidal Activities of Amphotericin B and N-Methyl-N″-dodecylguanidine: A Constituent of Polyol Macrolide Antibiotic Niphimycin

Abstract

The synergy between the alkylguanidinium chain of niphimycin (NM), a polyol macrolide antibiotic, and polyene macrolide amphotericin B (AmB) without such an alkyl side chain was examined using N-methyl-NЉ-alkylguanidines as its synthetic analogs. Among the analogs, N-methyl-NЉ-dodecylguanidine (MC12) most strongly inhibited the growth of Saccharomyces cerevisiae cells and those of other fungal strains in synergy with AmB. MC12 itself was not lethal but the analog could be a cause of a rapid cell death progression of yeast cells in the presence of AmB at a nonlethal concentration. Their combined actions resulted in the generation of NM-like fungicidal activity that depended on plasma membrane disability and cellular reactive oxygen species production. We also found an aberrant vacuolar morphogenesis and an associated vacuolar membrane disability in cells treated simultaneously with MC12 and AmB, as in the case of NM-treated cells. These findings support the idea that the alkylguanidinium chain plays a major role in the fungicidal activity of NM in cooperation with the polyol lactone ring as its enhancer.

Visualization analysis of the vacuole-targeting fungicidal activity of amphotericin B against the parent strain and an ergosterol-less mutant of Saccharomyces cerevisiae

“…A similar pattern of vacuole disruption is more markedly observed with the polyol macrolide antibiotic niphimycin (NM; Fig. 1b) in S. cerevisiae cells (Nakayama et al, 2002;Ogita et al, 2007). The molecular structure of NM is characterized by the presence of an alkylguanidium chain attached to its polyol lactone ring, which is absent in the polyene macrolide structure of AmB (see Fig.…”

Enhancement effect of N-methyl-N″-dodecylguanidine on the vacuole-targeting fungicidal activity of amphotericin B against the pathogenic fungus Candida albicans

“…The lethal action of MC12 in C. albicans cells was distinguishable from its static growth inhibition pattern observed in S. cerevisiae cells. 17 This difference may depend on different functional mechanisms for protection against MC12-induced oxidative stress in C. albicans and S. cerevisiae, as discussed below. C. albicans cells were extremely sensitive to the lethal action of AmB in the presence of MC12, suggesting a synergistic relationship in the mechanism of cell death progression between this polyene macrolide antibiotic and an analog of alkylguanidium chain in NM (Figure 2c).…”

“…Effects of AmB and MC12 alone and in combination on vacuole morphology Electron microscopic observation clearly revealed fragmentation of a membrane-enclosed rounded architecture of the vacuole in NMtreated cells of S. cerevisiae. 17 This drastic morphological change of the organelle could be also visualized under a microscope with the aid of a fluorescent probe. The vacuole was therefore evaluated as a target of NM lethality against S. cerevisiae, although it remains unknown whether or not vacuole disruption occurs as a result of NM-induced ROS production.…”

“…16 A comparison of AmB lethality in the absence and presence of MC12 additionally suggested the role of alkylguanidium chain in the vacuole disrupting activity of NM. 17 MC12 may be more useful and valuable as an enhancer of AmB lethality than allicin because MC12 lacks odorous and volatile properties. In this study, we attempted to find the role of MC12 as an enhancer of AmB lethality against C. albicans cells.…”

Dependence of vacuole disruption and independence of potassium ion efflux in fungicidal activity induced by combination of amphotericin B and allicin against Saccharomyces cerevisiae

“…15 Cells from the overnight culture in YPD medium were suspended in a freshly prepared medium to obtain a density of 1Â10 7 cells per ml. After incubation with 5 mM FM4-64 at 30 1C for 30 min, the cells were collected by centrifugation, washed twice and then suspended in distilled water at a final density of 1Â10 7 cells per ml.…”