EFFECT OF SUBSTRATE CONCENTRATION ON SUCROSE TRANSFRUCTOSYLATION IN A PACKED-BED REACTOR CONTAINING CATALYTIC BIOMASS OF Aspergillus oryzae IPT-301 IMMOBILIZED IN Luffa cylindrica

The implementation of packed-bed reactors (PBR) to increase industrial production volume, compared to the batch system, has been investigated for the production of fructooligosaccharide (FOS). Therefore, this study aimed to use biomass of Aspergillus oryzae IPT-301 containing fructosyltransferase (FTase, EC 2.4.1.9) for the transfructosylation of sucrose, by immobilizing the biomass in vegetable sponge (Luffa cylindrica) and applying it as a biocatalytic bed in continuous reactors. In addition, the effect  of varying sucrose concentration in the reactor feed solution and its impact  on sucrose transfructosylation was evaluated. The biocatalyst was obtained in a single step through  simultaneous cultivation and immobilization of the catalytic biomass. Loofah sponge cubes (1.0 cm edge length) were used, and the culture lasted 28 h at 30 °C and 200 rpm. A PBR reactor was used, filled with nine vegetable sponges containing immobilized biocatalytic cells, totaling a biocatalytic bed volume of 3.0 cm. The operation was carried out at a flow rate of 6 mL min^-1, and samples were taken at 0, 5, 10, 15, 25, 40 and 60 minutes. The substrate used was sucrose solutions (pH 5.5, concentrations of 200 g/L and 600g/L), kept at 50 °C. The experiments were conducted in triplicate, under the same operating conditions for both trials. After collection, the samples were subjected to enzymatic deactivation and kept for 15 minutes in a boiling bath, followed by immersion  in an ice bath for 5 minutes. The concentrations of glucose and reducing sugars were quantified using the glucose oxidase enzyme kit and 3.5 dinitrosalicylic acid colorimetric method to calculate the transfructosylation activity, defined as the amount of enzyme that transfers 1 μmol of fructose per minute under the experimental conditions. According to the ANOVA statistical test, at a significance level of 0.05, both the 200 g L^-1 and 600 g L^-1 concentration tests showed similar transfructosylation activity values over time, with the highest value obtained for the 200 g L^-1 concentration (281.17 ± 30.46 U g^-1). Thus, it was observed that using lower concentrations of sucrose solution favors enzymatic activity, contributes to minimizing the formation of by-products along the catalytic bed, and reduces raw material consumption in the process, making it a promising for FOS production.