Comparative Lipid Profiling of E. coli

Total lipid extract of E. coli was analyzed by multiple precursor ion scanning [16 (link)] and by data-dependent acquisition [10 (link)] on a QSTAR Pulsar i mass spectrometer. The same extract was analyzed by data-dependent HCD at the LTQ Orbitrap Velos mass spectrometer. Each analysis was performed in four replicates. Datasets of shotgun MS and MS/MS spectra were imported into MasterScan files built separately for each mass spectrometer and lipid species identified by MFQL queries (see Additional file 14 for the import settings and Additional file 11 for the queries). Lipid species were quantified in MS mode by using the intensities of their molecular ions. For MS/MS quantification, MFQL queries recognized and reported the sum of abundances of acyl anion fragments for each individual precursor. Relative quantities of individual lipids were calculated by normalizing to the total abundance of all species of the same lipid class. Parameters of linear correlation of lipid species profiles obtained by different methods (correlation coefficient R² and slope) were computed by Microsoft Excel (see Additional file 14).