Comprehensive analysis of the lipidome is addressed here by analysing lipid subclasses not easily detected by current high-throughput methods. Abundant lipid subclasses in human plasma are chromatographically separated from low abundance lipids prior to detection, avoiding the need for derivatisation. Lipid subclasses from the de novo lipogenesis and sphingolipids pathways are presented in this work. Three chromatographic methods here were implemented using a tertiary pumping system to allow for the inclusion of a gradient for analyte separation using A and B pumps, while an isocratic wash elutes interfering compounds. The isocratic wash enabled elution of lipid subclasses not targeted within the method that would otherwise cause background signal in the subsequent sample injection and reduction in column lifetime. Four chromatographic methods coupled with mass spectrometry using targeted and untargeted approaches to separate high and low abundance lipid subclasses are described here. An optimised method for the extraction of lysolipids is also used in addition to Folch extraction in human plasma.
Comprehensive lipidome of human plasma using 1 minimal sample manipulation by liquid chromatography coupled with mass spectrometry