Imobilização de células de Aspergillus oryzae IPT-301 em esponja de poliuretano para a produção enzimática de fruto-oligossacarídeos

Abstract

Fructooligosaccharides (FOS) are fructose oligomers beneficial to human health and nutrition by exhibiting nutraceutical properties. Their production occurs by transfructosylation reaction on sucrose molecules catalyzed by fructose-transferase enzymes (FTase, E.C.2.4.1.9) adhered to microbial cells. Immobilization of cells on support materials allows to obtain heterogeneous biocatalysts that are robust and resistant to the adverse conditions of the reaction medium. Polyurethane (EPU) sponges are suitable supports for immobilization because they present thermal and mechanical stabilities, besides pore size and distribution that allow gas and substrate diffusion, essential for microbial growth. Therefore, the objective of this work was to evaluate the production of Aspergillus oryzae IPT-301 cells, by submerged cell culture, and its concomitant immobilization in EPU sponges. To this end, the growth capacity and uptake of the cells as a function of culture time and transfructosylation (TA) activity were investigated, as well as characterization studies of the biochemical properties and stability of the immobilized and immobilized/reticulated cells, aiming to obtain robust, stable and active biocatalysts for FOS synthesis. The production of the microbial cells, confined in EPU, occurred in synthetic culture medium, pH 5.5, 200 rpm at 30 ºC. The microbial growth curve showed that the best cell culture time occurred for 32 h of process. The temperature of 50 ºC, pH 5.5 and concentration of 480.2 g L-1 were the parameters of the reaction medium that provided the maximum values of AT. Evaluation of the enzyme kinetics showed that the Hill model fitted satisfactorily to the kinetic profiles presented for both biocatalysts. Thermodynamic parameters indicated that the immobilized cells provided an increase in the thermostability of the biocatalyst. The pH stability tests showed that both cells (immobilized and immobilized and reticulated) were stable over a pH range between 4.5 and 6.0 and retained up to 40% initial AT when stored for 28 days under refrigeration. Finally, the results of operational stability indicated that the immobilized and immobilized and reticulated cells retained 50.23% and 60.90% of their initial enzymatic activities, respectively, after 12 consecutive reaction cycles, reaching an average production of 100 g L-1 of FOS. This set of results shows the potentiality of the application of catalytic cells, immobilized in EPU, in the production of FOS and represents the beginning of a sequence of studies on the immobilization of these microbial cells, by adsorption, in different support materials to obtain biocatalysts in the synthesis of bioproducts of industrial interest and in the implementation of various bioprocesses.