Lipid Profiling by Gas Chromatography

Lipids were isolated from cell pellets by standard Folch extraction. An aliquot of the pellet was used for cell protein determination by the Bradford assay. Triglycerides were directly analyzed by GC as described [84 (link)]. In brief, lipids were separated using a GC-2010 gas chromatograph (Shimadzu) equipped with a programmed temperature vaporizer injector and a ZB-5HT capillary column (15 m x 0.32 mm x 0.1 μm; Phenomenex). Trinonadecanoin (Sigma) was used as standard. For fatty acid analysis, FOLCH-extracts were trans-esterified using boron trifluoride-methanol solution (Sigma) at 80°C for 2 hrs followed by extraction with hexane. Lipids were separated on a ZB-FFAP capillary column (15 m x 0.32mm x 0.25 μm; Phenomenex) using pentadecanoin (Sigma) as standard. Chromatograms were analyzed using GC Solutions 2.3 (Shimadzu) and values were normalized to cell protein.