Novel, fully automatic hydrophilic interaction/reversed-phase column-switching high-performance liquid...

In this study we developed and optimized a column-switching high-performance liquid chromatographic (HPLC) system for the complementary analysis of polar and apolar compounds in complex samples. The polar compounds are separated on a hydrophilic interaction chromatographic (HILIC) column; the part of the sample non-retained on HILIC is transferred into and separated on a reversed-phase (RP) column through an interface, thus preventing the loss of analytes eluting at dead time in common single-column mode. The signals are in turn recorded in the same chromatogram either by a UV or by a mass spectrometric detector. Applying a mixture of 25 standard compounds (12 hydrophilic and 13 hydrophobic) the system proves to be reliable, robust and easy-operating. The investigation of urine revealed an increase in the detected metabolite ion masses compared to the single-column HILIC-electrospray ionization (ESI)-MS analysis. Utilizing online the complementary selectivity of HILIC and RPLC in a fully automatic mode, this approach holds out the prospect to enlarge the number of detectable compounds in non-targeted "-omics" studies and sophisticated target-driven approaches by a single injection.