conditions. In this study, carboxyl-functionalized chitosan-coated magnetic nanoparticle was synthesized and used as a support for the immobilization of laccase. The synthesized nanoparticle was characterized by transmission electron microscopy and FT-IR spectroscopy. In order to investigate the stability of the enzyme to pH changes, the activities of free and immobilized enzymes were measured at different pH values ranging from 2.0 to 8.0. The thermal stability and storage stability of the enzymes were also studied. Kinetic parameters and reusability were evaluated for both free and immobilized enzymes. The results showed that the magnetic nanoparticles and carboxyl-functionalized chitosan-coated magnetic nanoparticles are spherical with dispersed size distribution and zeta-potential of − 33.4 and + 1.95 mV, respectively. Based on our observations, the maximum activity of the enzyme was shifted toward higher temperatures after the immobilization. Furthermore, the immobilized laccase retained 80% of its initial activity after 120 days, while free enzyme maintained only 50% of its initial activity during the same period. The values of Michealis-Menten constant and maximum velocity for immobilized laccase were 0.113 mM and 1.27 μM min−1, respectively, whereas the values for free enzyme were 0.093 mM and 1.79 μM min−1,respectively. Both free and immobilized laccase exhibited optimum activities at pH 3.0 and decreasing trends were found at pH values more than 5 but immobilized laccase preserved its high activity at a wide range of pH compared with that of freelaccase. In particular, it can be used at higher temperatures and broader pH with improved storage stability and reusability, demonstrating the superior potential of bioremediation utilizing this immobilization method.
