Immobilization of horseradish peroxidase in Ca-alginate beads: Evaluation of the enzyme leakage on the overall removal of an azo-dye and mathematical modeling

Abstract

Horseradish peroxidase (HRP) was immobilized in calcium alginate beads by extruding an alginate/HRP solution through a syringe onto calcium chloride. The aim of this study was to evaluate their use to remove the azo-dye Orange II (OII), under different experimental conditions, such as the number of beads, pH, reuse of the biocatalyst, hydrogen peroxide feeding strategy. A mathematical model that takes into account the diffusion of reactants, products, and HRP throughout the beads and the effect of the enzyme leakage on the observable oxidation rate was also developed. Results indicated that the immobilization efficiency decreased as a function of the incubation time in CaCl2. Although enzyme-containing beads had a low OII adsorption capacity, under the presence of hydrogen peroxide, the dye removal was highly increased due to the catalytic activity of HRP. Besides, the initial OII oxidation rate (RS0) using 3 beads at pH 9 was higher than at pH 7. Also, RS0 increased as a function of the number of tested beads. The developed model was fitted to experimental data obtained at different conditions. Then, the model was validated against a new data set that was not used for the calibration of the model, obtaining a satisfactory agreement between simulation results and these new data. Simulations demonstrate that 70–90% of the total OII removed was due to its oxidation by hydrogen peroxide catalyzed by HRP in the liquid phase.