A reverse-phase HPLC assay for measuring the interaction of polyene macrolide antifungal agents with sterols.
A quick and simple affinity chromatography method for gauging the interaction of polyene antifungal agents with sterols has been developed. The required affinity columns are prepared from a standard C-18 reverse-phase HPLC column by injecting a measured quantity of sterol under conditions where it is completely retained. After the assay, the sterol is eluted with a less polar solvent and the column reused. By comparing the elution volume of a polyene injected onto the sterol-free column (Ve) with that of the polyene injected onto the sterol-doped column (V), an association constant (Ka) for the polyene-sterol complex was determined. Association constants of different amphotericin B-sterol and pimaricin-sterol complexes were determined and correlated with the polyene's ability to induce membrane permeability and its antifungal properties. This procedure provides a new tool for screening polyene macrolides for antifungal therapy.
Backes
B J
BJ
Department of Chemistry, University of Minnesota, Minneapolis 55455.
Rychnovsky
S D
SD
D000666:Amphotericin B / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004347:Drug Interactions; D010866:Natamycin / Q000378:metabolism*; D013261:Sterols / Q000378:metabolism*
D013261:Sterols; D000666:Amphotericin B; D010866:Natamycin