Polymer Monoliths with Low Hydrophobicity for Strong Cation-Exchange Capillary Liquid Chromatography of Peptides and Proteins

Abstract

Two polymer monoliths were designed and synthesized from commercially available monomers with an attempt to decrease hydrophobicity for strong cation-exchange chromatography. One was prepared from the copolymerization of sulfoethyl methacrylate and poly(ethylene glycol) diacrylate, and the other was synthesized from vinylsulfonic acid and poly(ethylene glycol) diacrylate. Both of the monoliths were synthesized inside 75-microm i.d., UV-transparent fused-silica capillaries by photopolymerization. The hydrophobicities of the two monoliths were systematically evaluated using standard synthetic undecapeptides under ion-exchange conditions and propyl paraben under reversed-phase conditions. The poly(sulfoethyl methacrylate) monolith demonstrated similar hydrophobicity as a monolith prepared from copolymerization of 2-acrylamido-2-methyl-1-propanesulfonic acid and poly(ethylene glycol) diacrylate, and 40% acetonitrile was required to suppress any hydrophobic interactions with peptides under ion-exchange conditions. However, with the use of vinylsulfonic acid as the functional monomer, a monolith with very low hydrophobicity was obtained, making it suitable for strong cation-exchange liquid chromatography of both peptides and proteins. It was found that monolith hydrophobicity could be adjusted by selection of monomers that differ in hydrocarbon content and type of vinyl group. Finally, excellent separations of model protein standards and high-density lipoproteins were achieved using the poly(vinylsulfonic acid) monolith. Five subclasses of high-density lipoproteins were resolved using a simple linear NaCl gradient.