Influence of serum protein binding on the in vitro activity of anti-fungal agents.

Historically it has been assumed that the pharmacological effect is related to the free drug concentration. In exposing Candida albicans to itraconazole and ketoconazole serum concentration-time profiles, however, antifungal activity was not diminished despite intense albumin binding. The relevance of serum protein binding was further investigated, by in vitro susceptibility testing of C. albicans (40 clinical isolates) and Trichophyton rubrum (ten strains) against antifungal agents using microdilution tests allowing the determination of IC30- and MIC-values. The range of serum protein binding ranges from 11% with fluconazole to > 99% with itraconazole and terbinafine. The ratios of IC30- and MIC-values with and without serum protein (albumin, alpha- and gamma-globulin, human plasma) were related to the loss of susceptibility expected according to the free-drug hypothesis. A difference in the albumin effect with the test strains was not observed. With most antifungals including terbinafine, the activity declined as expected. IC30- and MIC-ratios for miconazole were 7 and 13 (observed) vs. 12-20 (expected), for fluconazole 1.5 and 3.5 vs. 1.1, for amphotericin B 10 vs. 11-20, for griseofulvin 3.6 vs. 4, and for terbinafine 61 vs. 100. Itraconazole activity, however, was not diminished by albumin (expected ratio 286), and ketoconazole effects decreased less than expected (ratio 5-15, expected about 100). alpha-globulin, but not gamma-globulin induced a major loss in anti-Candida activity of itraconazole and ketoconazole, which is paralleled by a decline in ketoconazole (but not itraconazole) activity due to plasma. With the other antifungals (except for ciclopiroxolamine) IC30-values for C. albicans increased, too. Due to the complete inhibition of T. rubrum growth by gamma-globulin, this species proved unsuitable for studying the gamma-globulin effects. The present study demonstrates that the effects of intense protein binding on drug activity are only partly predictable from binding studies in vitro.