Altis triple quadrupole mass spectrometer

Lipids were extracted using a 1:1 mixture of methanol and butanol containing a panel of internal lipid standards (Avanti^® Polar Lipids). Sphingolipids were quantified using targeted lipidomics on a TSQ Altis triple quadrupole mass spectrometer, following lipid separation using an Agilent Eclipse Plus C8 column on a Vanquish^™ ultra high-performance liquid chromatography system [35 (link)]. Peaks were integrated using Xcalibur [35 (link)]. Absolute concentrations were determined by normalization against corresponding internal standards and tissue weight. Hierarchical clustering was employed to group similar patterns of lipid changes using Euclidean distance and Ward’s algorithm, where Z-scores were calculated to measure the relative deviation of individual lipid concentrations from the mean values (Metaboanalyst 5.0).

Lipids were extracted from mouse liver tissues or Huh7 cells in 1 mL methanol containing a cohort of internal lipid standards (Avanti^® Polar Lipids, Alabaster, USA). The liver tissues isolated from the first 8 animals of each genotype were analysed. Hepatic levels of TG, CE, and phospholipids were quantified using untargeted lipidomic profiling on a Q Exactive HF-X mass spectrometer, following lipid separation on a Waters Acquity C18 UPLC column [53 ]. LipidSearch software was used for lipid annotation, chromatogram alignment and peak integration [75 (link)]. In contrast, FFAs, DG, FC, and sphingolipids were determined by targeted lipidomics on a TSQ Altis triple quadrupole mass spectrometer, following lipid separation on an Agilent Eclipse Plus C8 column [28 (link), 53 ]. Peaks were integrated using Xcalibur (Thermo Fisher, Waltham, USA) [28 (link)].