Imobilização da enzima frutosiltransferase extracelular de Aspergillus oryzae IPT-301 em sílica gel pura e funcionalizada para produção de frutooligossacarídeos

Abstract

Fructooligosaccharides (FOS) are low-calorie, non-cariogenic prebiotic sugars that can be consumed by diabetics and, as they are not hydrolyzed by gastrointestinal enzymes, they promote the selectivity of probiotic bacteria in the intestinal microbiota, helping to eliminate pathogenic microorganisms and prevent cancer of colon. They are commercially produced by microbial enzymes such as fructosyltransferase (FTase, E.C.2.4.1.9) using sucrose as substrate. The immobilization of these enzymes on porous supports, allows to increase the enzymatic stability, to reuse the biocatalyst and to protect the enzyme from adverse conditions of the reaction medium. Therefore, this work aimed to immobilize, by physical adsorption and covalent bonding, the extracellular FTase of Aspergillus oryzae IPT-301 on silica gel, pure (untreated) and functionalized with glutaraldehyde, in order to obtain an active and stable heterogeneous biocatalyst for FOS production. The thermal stability, against the incubation, operational and storage pH of the immobilized enzyme, as well as the kinetic profiles of the biocatalyst, were evaluated. The best conditions obtained for the immobilization of FTase on pure silica gel (35 ºC, pH 5.5 and 175 rpm) and on silica gel functionalized with glutaraldehyde were used to determine the immobilization yield (RI) and the recovered activity (AR). For the FTase immobilized on pure silica gel, RI and AR values equal to 13% and 10%, respectively, were reached, while for the enzyme immobilized on the functionalized support, an RI of 38% and an AR of 7, were obtained. 5%. The enzyme immobilized on both supports showed kinetic behavior described by the Hill corporate model, whose highest activity values were obtained for a substrate concentration range between 400 g L-1 and 600 g L-1. FTase immobilized on functionalized silica gel showed greater reuse capacity over 8 consecutive reaction cycles, in addition to exhibiting greater thermal, storage and pH stability compared to the biocatalyst adsorbed on the pure support. The results obtained suggest a high potential for the application of functionalized silica gel as an FTase immobilization support for the production of FOS.