Miniaturized biocatalysis on polyacrylate-based capillary monoliths

Abstract

We developed a synthetic concept for the immobilization of enzymes onto monolithic supports. In addition, we elaborate on a generally applicable method for the rapid screening of the activity of such immobilized enzymes. For these purposes, we prepared monolithic acrylate-based systems by electron-beam (EB)-triggered free-radical polymerization within the confines of 200-μm capillary columns. Aiming for protein immobilization, we subjected the polyacrylate-based monoliths to EB-mediated grafting processes to introduce functional surface-located groups suited for the subsequent generation of functional units that themselves could bind different proteins. For the generation of the functional units, we used ring-opening metathesis polymerization and free-radical polymerization. The produced systems were tested for their ability to bind or repel proteins as exemplified by the use of the serine protease trypsin, which was used to catalyze the hydrolysis of N-α-benzoyl-L-arginine ethyl ester (BAEE). Finally, the monolith-immobilized trypsin systems were used for enzymatic peptide synthesis purposes, such as the acyl transfer of BAEE to amino acid amides. Complementarily, we used an immobilized trypsin variant, which we additionally subjected to on-column chemical modification with succinic anhydride to alter its synthetic properties. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011