Metabolic Profiling of C. albicans

An overnight culture of C. albicans was diluted into YNB-CSM-2% glucose media at an OD⁶⁰⁰ of 0.25, grown for 3hr to mid-log phase, and then split into parallel 5 mL cultures for treatment with compounds (DMSO, 20µM rotenone, 1µM antimycin, 5mM potassium cyanide, 20µM oligomycin, 25 µg/mL 2,4-dinitrophenol, 32 µM FCCP, 2.5 µM ML316). After growth with compounds for the indicated time intervals, cultures were pelleted, washed once in PBS, and frozen in liquid nitrogen. Polar metabolites were extracted by re-suspending cells in 600 µL methanol, 300 µL water, and 400 µL chloroform, and adding 100 µL of glass beads before vortexing for 60 seconds. Samples were then centrifuged for 10 minutes at 13,000g, and the supernatant (polar) layer was moved to a new microcentrifuge tube, and evaporated by Speedvac. Samples were then subjected to LC-MS/MS profiling on a Thermo Orbitrap platform, as described previously.^{37 (link)}