GNPS - A comparative analysis of mycobacterial lipids using liquid chromatography mass spectrometry-based lipidomics

The lipid content of twenty different species across the genus Mycobacterium was analysed by LC-MS. Single colonies were inoculated into Middlebrook 7H9 medium and grown to an OD 600nm of one. One mL of culture was transferred onto a filter and filters were placed on 7H10 plates and incubated for 6 doubling times. Lipids were extracted with chloroform: methanol in 2:1, 1:1 and 1:2 ratios. Extracts were pooled and dried and resuspended in LC mobile phase A. Analysis was done using a BETASIL Diol-100 column on an Ultimate 3000 HPLC system attached to a Thermo Q Exactive Orbitrap Mass Spectrometer. Mobile phase A was isopropanol:hexane at 70:30 and mobile phase B was isopropanol: methanol at 70:30, both phases contained 0.02% [m/v] formic acid and 0.01% [m/v] ammonium hydroxide.