Immobilization of horseradish peroxidase by entrapment in natural polysaccharide



Horseradish peroxidase (HRP), which catalyzes oxidation reduction reactions of large number of substrates, was entrapped in K-carrageenan beads using polyethyleneimine as hardening agent. The heat and storage stability was found to be better for entrapped horseradish peroxidase than free enzyme. The entrapped enzyme showed 50% retention of its activity after 4 cycles. Effective diffusion coefficient for diffusion of hydroquinone into K-carrageenan beads was found to be 0.27 × 10−10 m2/s during enzyme-catalyzed oxidation of hydroquinone. Kinetic parameters calculated from Lineweaver–Burke plots were observed to be Km = 8 × 10−5 and Vmax = 1.53 for free enzyme, and Km = 8.3 × 10−5 and Vmax = 2.18 for entrapped enzyme when enzyme concentration was kept constant and Km = 4 × 10−11 and Vmax = 0.45 for free enzyme and Km = 4.5 × 10−11 and Vmax = 0.58 for entrapped enzyme when substrate concentration was kept constant. This indicates that there is no conformational change during entrapment. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 91: 2063–2071, 2004