Analysis of pH and buffer effects on flucytosine activity in broth dilution susceptibility testing of Candida albicans in two synthetic media

Abstract

We examined the influences of different pH levels and three different buffers on flucytosine activity against 12 isolates of Candida albicans in two synthetic media, yeast nitrogen base (YNB) and synthetic amino acid medium-fungal (SAAMF), using broth dilution techniques and measuring the endpoints of visual MICs and turbidimetric 50% inhibitory concentrations. The two media were originally prepared as follows: YNB, unbuffered, pH 5.6; SAAMF, buffered with morpholinepropanesulfonic acid-Tris, pH 7.4; the resultant geometric mean MIC and 50% inhibitory concentration of 5-FC were 78- and 32-fold higher, respectively, in SAAMF. Raising the pH of YNB or lowering the pH of SAAMF had virtually no effect on these differences in MIC and 50% inhibitory concentration in the two media. In contrast, virtually all of the discrepancy appeared to be due to morpholinepropanesulfonic acid-Tris, which exerted concentration-dependent inhibition of flucytosine activity not evident when N-2-hydroxyethylpiperazine-N'-ethanesulfonic acid or phosphate buffer systems were substituted. In other turbidimetric studies, growth was slowed more than 50% in YNB as the pH was raised to 7.4, regardless of which buffer was used. Based on our studies, we recommend modifying the composition of SAAMF by substituting a nonantagonistic buffer if any buffer is to be used with SAAMF in the testing of flucytosine. With this modification, SAAMF warrants further study as a generally applicable medium for fungal-susceptibility testing.