Reversed-phase liquid chromatography of proteins and peptides using multimodal copolymer-encapsulated silica

Multimodal copolymer-encapsulated particles for liquid chromatography were prepared by bonding 1-octadecene and unsaturated carboxylic acids on silica particles (5 μm diameter, 300 Å pores) for liquid chromatography of proteins. These multimodal copolymer-encapsulated particles can provide both hydrophobic and hydrogen bonding interactions with polar compounds. The chromatographic performance of these multimodal copolymer-encapsulated particles for peptide and protein separations was evaluated under reversed-phase conditions. Compared with typical C8-bonded silica, polymer-encapsulated particles were more stable in acidic mobile phases and provided better recoveries, especially for large proteins (Mr>0.5·106). Totally hydrophobic polymer-encapsulated particles were found to produce broad peaks for proteins, and significant improvements were observed by introducing hydrophilic groups (–COOH) onto the polymer-encapsulated surface to form a multimodal phase. For the reversed-phase liquid chromatography of peptides and proteins, improved selectivity and increased solute retention were found using the multimodal polymer-encapsulated particles. More peaks were resolved for the separation of complex peptide mixtures such as protein digests using the multimodal polymer-encapsulated particles as compared to totally hydrophobic polymer-encapsulated particles.