Photochemical preparation of magnetic chitosan beads for immobilization of pullulanase

Magnetic chitosan beads were prepared via photochemical polymerization in Fe3O4 magnetite aqueous suspension, and characterized by Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), photocorrelation spectroscopy (PCS) and vibrating sample magnetometer (VSM). The FT-IR result indicated that the magnetic chitosan beads were synthesized successfully under UV irradiation in aqueous system. The magnetic chitosan beads were of regularly spherical shape, had a mean diameter of 86 nm, exhibited superparamagnetic property and the saturation magnetization was 52.6 emu/g. Pullulanase was covalently immobilized on magnetic chitosan beads by cross-linking with glutaraldehyde. The maximum quantity of pullulanase immobilized onto 1.0 mg magnetic chitosan beads was 180 μg. The optimum temperature of immobilized pullulanase was 50° C, which was identical to that of free enzyme, and the immobilized pullulanase exhibited a higher relative activity than that of free pullulanase over 50° C. The optimal pH for immobilized pullulanase was 5.0, which was lower than that of the free pullulanase (pH 5.5), and the immobilization resulted in stabilization of enzyme over a broader pH range. The kinetic constant value (Km) of immobilized pullulanase was three times higher than that of free pullulanase. However, the thermal and operational stabilities of immobilized pullulanase were improved greatly.