Covalent immobilization of Aspergillus tamarii β-fructofuranosidase on Fe3O4-chitosan magnetic nanoparticles enhances thermostability and catalytic performance for inverted sugar production.

Abstract

β-fructofuranosidases (FFases) catalyze the hydrolysis of sucrose, producing commercially valuable products such as invert sugar. However, the use of enzymes in industry faces limitations due to operational instability. Immobilization emerges as an alternative to reduce these instabilities, improving stability and enabling reuse. The objective of this study was to immobilize the FFase from A. tamarii URM4634 onto Fe3O4 magnetic nanoparticles coated with chitosan, to biochemically characterize it, study its kinetic and thermodynamic parameters, and apply it in sucrose hydrolysis. Immobilization was carried out according to a full 22 factorial design with variations in glutaraldehyde concentration and contact time. A high activity yield was obtained (94.45%), which exhibited optimal activity at 60 °C and pH 7.0, maintaining full catalytic activity over 8 consecutive reuse cycles. Immobilization significantly increased thermostability at 50 °C, evidenced by the increase in thermal denaturation activation energy (E*d = 322.16 kJ mol-1) and a stabilization factor of 1.92 relative to the free enzyme. The application of immobilized FFase in invert sugar synthesis achieved a maximum sucrose conversion of 49.39%. These results, combined with the ease of magnetic separation from the reaction medium, highlight the biotechnological potential of the immobilized enzyme for the industrial production of invert sugar.