Horseradish peroxidase and chitosan: Activation, immobilization and comparative results

Abstract

Recently, horseradish peroxidase (HRP) was immobilized on activated wool and we envisioned that the use of chitosan would be interesting instead of wool owing to its simple chemical structure, abundant nature and biodegradability. In this work, HRP was immobilized on chitosan crosslinked with cyanuric chloride. FT-IR spectroscopy and scanning electron microscopy were used to characterize immobilized HRP. The number of ten reuses of immobilized HRP has been detected. The pH was shifted from 5.5 for soluble HRP to 5.0 for immobilized enzyme. The soluble HRP had an optimum temperature of 30°C, which was shifted to 35°C for immobilized enzyme. The soluble HRP and immobilized HRP were thermal stable up to 35 and 45°C, respectively. The apparent kinetic constant values (Km) of soluble HRP and chitosan–HRP were 35mM and 40mM for guaiacol and 2.73mM and 5.7mM for H2O2, respectively. Immobilization of HRP partially protected them from metal ions compared to soluble enzyme. The chitosan–HRP was remarkably more stable against urea, Triton X-100 and organic solvents. Chitosan–HRP exhibited large number of reuses and more resistance to harmful compounds compared with wool–HRP. On the basis of results obtained in the present study, chitosan–HRP could be employed in bioremediation application.