Immobilization of Candida antarctica lipase B onto modified silica nanoparticles and its application for the synthesis of l-ascorbyl oleate

Abstract

Lipases immobilized onto nonporous fumed silica nanoparticles (FNS) proved to be suitable catalysts for the various commercially important esters synthesis. This inert hydrophilic material is usually modified by different organosilanes, in order to assure formation of strong covalent bonds and/or hydrophobic interactions with enzyme surface amino acid residues. Hereby, we present methods for the modification of FNS surface by 3-aminopropyltrimethoxysilane (APTMS) and/or (Nphenylamino) propyltrimethoxysilane (PAPMS), and subsequent functionalization of amino-modified FNS nanoparticles with cyanuric-chloride (CC). Selected modified nanocarriers were applied for the immobilization of Candida antarctica lipase B (CALB). Influence of chemical modification of FNS surface on the CALB immobilization process, protein loading, and hydrolytic activity of obtained preparations were examined. Furthermore, chosen derivatives were tested in the production of valuable food additive, liposoluble antioxidant, L-ascorbyl oleate. Results showed that highest protein loadings were accomplished with nanocarriers modified with combination of both APTMS and PAPMS. However, probably due to hindered approach of substrate molecules to the enzyme active sites, their hydrolytic and esterification activities were not proportionally high. All preparations obtained with modified FNS nanoparticles were more active in the hydrolysis of p-nitrophenyl butyrate (p-NPB) comparing to CALB immobilized onto unmodified FNS. The highest hydrolytic activity of 1518.6 p-NPB/g was demonstrated by CALB covalently immobilized onto nanoparticles modified by APTMS and CC. Meanwhile, best results in the synthesis of L-ascorbyl oleate were achieved when CALB hydrophobically immobilized onto FNS nanoparticles modified by only PAPMS, with hydrolytic activity of 1200.2 p-NPB/g, was used as a catalyst and yield of 23.4 mM was accomplished. Therefore, it was proven that different modification strategies ensured formation of different interactions between lipase molecules and silica nanoparticles during immobilization, which led to diverse hydrolytic and esterifying characteristics of obtained preparations. CALB immobilized onto nanocarrier of highest hydrophobicity showed greatest potential for the production of L-ascorbyl oleate.