Development of a single mobile phase for LC-IM-MS-based discovery lipidomics and metabolic phenotyping: Application to methapyrilene hepatotoxicity in the rat

•A generic mobile phase has been developed for metabolomic and lipidomic separations•Isopropyl alcohol was replaced by acetonitrile as the organic modifier for lipidomics•Lipidomics on a phenyl-hexyl column increased peak capacity (58 %) and detection (61 %)•RPLC on a C8 stationary phase was used for moderately polar-nonpolar analytes•Polar metabolites were separated by HILIC on an amide stationary phase The untargeted global profiling of endogenous metabolites and lipids has the potential to increase knowledge and understanding in many areas of biology. LC-MS/MS is a key technology for such analyses however, several different LC methodologies, using different mobile phase compositions, are required to cover the diversity in polarity and analyte structure encountered in biological samples. Most notably many lipid screening methods make use of isopropanol (IPA) as a major component of mobile phases employed for comprehensive lipidomic profiling. In order to increase laboratory efficiency, and minimize opportunities for errors, a suite of methods, based on a single acetonitrile (ACN)-aqueous buffer mobile phase combination, has been developed. This mobile phase can be used for hydrophobic interaction liquid chromatography on an amide stationary phase (for polar analytes), reversed-phase (RP) LC analysis on a C8 stationary phase (for moderately polar-non-polar compounds) and RPLC using a CSH phenyl-hexyl bonded column (for lipids). All of these sub 10 minute separations had good throughput and reproducibility with CV's of analyte response