Development of Polymer Monoliths for the Analysis of Peptides and Proteins

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Brigham Young University - Provo

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Gu, Binghe
Brigham Young University - Provo
monolith; proteins; peptides; biocompatible; liquid chromatography; size exclusion chromatography; strong cation-exchange chromatography; polyethylene glycol diacrylate; 2-acrylamido-2-methyl-1-propanesulfonic acid; sulfoethyl methacrylate; vinyl sulfonic acid; capillary columns; peak capacity; Biochemistry; Chemistry
thesis / dissertation description
Several novel polymer monoliths for the analysis of peptides and proteins were synthesized using polyethylene glycol diacrylate (PEGDA) as crosslinker. Photo-initiated copolymerization of polyethylene glycol methyl ether acrylate and PEGDA yielded an inert monolith that could be used for size exclusion liquid chromatography of peptides and proteins. This macroscopically uniform monolith did not shrink or swell in either water or tetrahydrofuran. More importantly, it was found to resist adsorption of both acidic and basic proteins in aqueous buffer without any organic solvent additives. A strong cation-exchange polymer monolith was synthesized by copolymerization of 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPS) and PEGDA. A ternary porogen (water, methanol and ethyl ether) was found suitable to prepare a flow-through monolith with moderate pressure drop in aqueous buffer. The resulting monolith showed excellent ion exchange capillary liquid chromatography of peptides using a simple salt gradient. Extremely narrow peaks were obtained for the analysis of synthetic peptides, natural peptides and a protein digest. A peak capacity of 179 was achieved. Although the poly(AMPS) monolith demonstrated extraordinary performance, one main drawback of this monolith was its relatively strong hydrophobicity. A decrease in hydrophobicity was achieved by using more hydrophilic monomers (e.g., sulfoethyl methacrylate or vinyl sulfonic acid). The most hydrophilic poly(vinyl sulfonic acid) monolith provided high resolution cation-exchange liquid chromatography of protein standards and lipoproteins. Use of the new PEGDA biocompatible crosslinker over the conventional ethylene glycol dimethacrylate crosslinker for the preparation of polymer monoliths was found to be advantageous for the analysis of biological compounds in several chromatography modes.