Separation of nucleobases, nucleosides, and nucleotides using two zwitterionic silica-based monolithic capillary columns coupled with tandem mass spectrometry

• Two kinds of zwitterionic monolithic capillary columns were prepared.
• Columns were evaluated in HILIC mode.
• The sample contained a mixture of nucleobases, nucleosides, and nucleotides.
• LC–ESI–MS/MS allowed detection and identification of 21 highly polar compounds.

The capability of employing synthesized zwitterionic silica-based monolithic capillary columns (140 mm × 0.1 mm) for separation of highly polar and hydrophilic nucleobases, nucleosides, and nucleotides in hydrophilic interaction chromatography is reported. The suitability of the columns for on-line conjunction with electrospray tandem mass spectrometry was explored. Our results show that the grafted layer of zwitterionic monomer ([2-(methacryloyloxy)ethyl]-dimethyl-(3-sulfopropyl)-ammonium hydroxide or 2-methacryloyloxyethyl phosphorylcholine) on the silica monolithic surface significantly improved the separation selectivity and reproducibility, as compared to the bare silica monolith. The stepwise elution from 90% to 70% of acetonitrile enabled separation of a complex sample mixture containing 21 compounds with a total analysis time less than 40 min.