This work compares the separation selectivity of two complementary densely bonded zwitterionic columns for hydrophilic interaction liquid chromatography (HILIC). One column type was a newly developed densely bonded zwitterionic stationary phase with phosphorylcholine functional groups (ZIC-cHILIC), comprising inner negatively charged phosphate groups and distal positively charged quaternary ammonium groups in 1:1 ratio. The second column type was based on similar technology, but the zwitterionic functional groups were sulfobetaine (ZIC-HILIC), i.e., inner positively charged quaternary ammonium groups and distal negatively charged sulfonate groups.

Chromatographic studies under typical HILIC eluent conditions showed that the spatial arrangement of the charges influenced the separation selectivity, especially for charged polar hydrophilic molecules. The distal charge dominated slightly over the inner charge on both columns, although the close proximity of the two charges resulted in overall weak ionic interactions. The two columns were thus providing different, and complementary, selectivity for charged polar hydrophilic molecules, with more retention of anionic compounds on the phosphorylcholine-based ZIC-cHILIC and more retention of cations on the sulfobetaine-based ZIC-HILIC.

These documented selectivity features were used for development of HPLC separations of compounds such as aminoglycosides and organic acids, and can also be applied to analyze polar drugs and impurities, biomarkers, food constituents and for applications in proteomics and metabolomics.