Proteases Entrapped in Hydrogels Based on Poly(N-Vinyl Caprolactam) as Promising Biocatalysts in Water/Organic Systems

Chemico-enzymatic methods are a promising approach to obtain various biologically active compounds, including enantiomerically pure substances. Entrapment in gels is one of the most convenient methods to stabilize enzymes for their application in water/organic media. In this study, proteases (trypsin and chymotrypsin) were entrapped into a composite poly (N-vinyl caprolactam)-calcium alginate (PVCL-CaAlg) hydrogel, and properties of the biocatalysts obtained were studied. Both entrapped trypsin and α-chymotrypsin retained high activity (up to 85% of the initial value for the immobilized trypsin) and displayed high storage stability. The entrapped proteases were active over broad temperature ranges, and their heat tolerance was approximately 20-25°C higher than that of the soluble enzymes. The dependence of the activities of the soluble and entrapped enzymes on the organic solvent concentration was studied. It appeared that gel entrapment provided retention of enzyme activity in the media with high concentrations of organic solvents (up to 0.5% water content) and ensured high operational stability of α-chymotrypsin in a cyclic process (270 h in total). The entrapped α-chymotrypsin was used as a biocatalyst for obtaining optically pure L-phenylalanine (e.e. 87.8%) by enantioselective hydrolysis of Schiff's base of amino acid ethyl ester in an acetonitrile/water system.

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