Antifungal Susceptibility of Oceanalin B

The fungal isolate used in this study was a Candida glabrata clinical isolate (University of California Davis Medical Center, UCDMC). The fungi were grown and maintained in Sabouraud dextrose agar and incubated at 30 °C for 24 h. The in vitro susceptibility of oceanalin B was determined by the broth microdilution method [20 (link)]. Briefly, 2-fold serial dilutions of oceanalin B were prepared in 96-well microtiter plates from stock solutions in an RPMI-1640 broth medium (Sigma, St. Louis, MO, USA) buffered to a final pH of 7.0 with 0.165 M morpholinepropanesulfonic acid (MOPS; Sigma, St. Louis, MO, USA) to a final volume of 100 μL. A stock solution of oceanalin B was prepared in dimethyl sulfoxide and amphotericin B (AMB) (Sigma, St. Louis, MO, USA) was prepared as a positive control. C. glabrata cells (5 × 10⁶/mL) in 100 μL suspension was added to the wells. The final concentrations tested were from 0.062 to 64 μg/mL and from 0.0078 to 8 μg/mL for amphotericin B. The experiment was performed in triplicates in each run of the experiments. Cell growth was determined by the OD at 600 nm using a Spectramax Plus 384 microplate reader (Molecular Devices, San Jose, CA, USA). The MIC end point was defined as the lowest concentration with complete (>90%) growth inhibition.