Impact of sulfonium versus ammonium groups in zwitterions on hydrophilic interaction chromatographic separation performance.

The separation performance of zwitterionic hydrophilic interaction chromatography (HILIC) is highly dependent on the molecular structure of zwitterion. Notably, sulfonium cations possess distinct physicochemical properties compared to ammonium cations in conventional stationary phases. In this study, the impact of sulfonium‑ vs. ammonium‑based zwitterions on hydrophilic interaction chromatographic performance was systematically investigated using monolithic column technology. Both sulfonium and ammonium-based zwitterionic monoliths were prepared by copolymerizing 3-(methyl{2-[(2-methylacryloyl)oxy]ethyl}sulfaniumyl)propane-1-sulfonate (MMSPS) and N,N-dimethyl-N-methacryloxyethyl-N-(3-sulfopropyl) ammonium betaine (SPE), respectively, with crosslinker N,N'-methylenebisacrylamide (MBA). The novel poly(MMSPS-co-MBA) monolith exhibited excellent column efficiency (∼100,000 plates/m), permeability, mechanical stability, and separation performance. Compared to the ammonium-based poly(SPE-co-MBA) monolith, the sulfonium-based monolith showed higher surface charge density, which resulted in prolonged retention of amine compounds and accelerated the elution of benzoic acid derivatives and phenolic analytes due to weaker hydrogen-bonding interactions, respectively. Moreover, the sulfonium-based monolith demonstrated superior performance in the enrichment of N-glycopeptide from a human immunoglobulin G tryptic digest, benefiting from its high hydrophilicity and enhanced electrostatic interaction. Overall, this study not only clarifies the significant influence of the cationic group on the chromatographic behavior of zwitterionic HILIC stationary phases, but also highlights the distinct hydrophilicity, electrostatic properties, and hydrogen-bonding characteristics of sulfonium-based zwitterion in comparison to ammonium-based counterparts.