Recent advances in application of hydrophilic interaction chromatography for quantitative bioanalysis

Hydrophilic interaction chromatography (HILIC) provides a complementary separation mode to RPLC and thus has been gaining increased utilization in LC-MS/MS based quantitative bioanalysis. It has proven to be a powerful tool for separation of polar compounds and has afforded increased selectivity, higher sensitivity, and improved efficiency for quantitation of drug and their metabolites in complex biological matrices. Practical knowledge has been gained for some of the challenges with HILIC applications and effective remedies have been adopted to overcome these challenges. Due to its orthogonality to RPLC, HILIC has been coupled with RP sample preparation or separation techniques, either off-line or on-line to achieve better assay selectivity. The low back-pressure with HILIC enables fast separations under higher flow rates. Small particle columns can be operated under HILIC conditions with regular system back-pressure, eliminating the requirement for ultra-high-pressure liquid chromatographic systems. Matrix effects under HILIC have also been systematically investigated. Effective approaches to reduce or eliminate matrix effects have included optimizing sample preparation, modifying chromatographic conditions, and/or using valve-switching techniques. Finally, the utilization of HILIC is not limited to quantitation of polar drugs and their metabolites, but extended to quantitation of relatively non-polar compounds, peptides and biomarkers.