Alpha-amylase immobilization onto chitosan coated magnetic nanoparticles

In this study, Fe3O4 nanoparticles were coated with chitosan (CS) by reversed-phase suspension method. ?-Amylase from Bacillus subtilis was immobilized onto chitosan coated Fe3O4 (Fe3O4-CS) nanoparticles by adsorption method and by covalent binding onto Fe3O4-CS nanoparticles which were activated by N-(3-dimethylaminopropyl)-N'- ethyl-carbodiimide hydrochloride (EDAC) and cyanuric chloride (SC). The effect of pH, temperature, substrate concentration, storage time on the activities of free and immobilized ?-amylases (Fe3O4-CS-A, Fe3O4- CS-EDAC-A, Fe3O4-CS-SC-A) were investigated. The optimum pH and optimum temperature were determined as 6,9; 8,0; 6,5; 6,0 and as 40°C, 30°C, 40°C ve 40°C for free ?-amylase, Fe3O4-CS-A, Fe3O4-CS-EDAC-A, Fe3O4-CS-SC-A respectively. Retained activities for immobilized systems were found to be 83%-97% after 10 runs, and 59%-80% after 30 runs in a day. The kinetic parameters were searched for free and Fe3O4- CS-A, Fe3O4-CS-EDAC-A, Fe3O4-CS-SC-A and calculated as 4,14; 2,41 2,3; 2,31 mg/mL for Km, 2,57; 6,06; 5,52; 4,43 U/mg for Vmax respectively.Key Words : Magnetic nanoparticles, chitosan, ?-amylase, enzyme immobilization, adsorption