Abstract: Micro-size magnetic polyvinyl acetate microspheres were prepared by a modification suspension polymerization, the magnetic beads were further modified with hydrolysis and aminization reactions, and magnetic affinity microspheres suitable for nattokinase purification were obtained by covalent immobilization of p-aminobenzamidine to the amino-modified magnetic beads by the glutaraldehyde method. The properties of magnetic beads were examined by SEM, XRD, FT-IR and VSM. The results showed that these magnetic beads were typically superparamagnetic and the specific saturation magnetization of ligand-attached magnetic beads was 32.4 emu/g. It took only 40 min to reach adsorption equilibrium and less than 15 min to achieve desorption equilibrium. The purification factor and the recovery rate of enzyme activity were 10.3 and 73.4%, respectively. The purified nattokinase gave a single sharp band on SDS-PAGE.

Key words: nattokinase, affinity separation, purification, adsorption, kinetics

摘要： 采用改进悬浮聚合法制备了磁性聚醋酸乙烯酯微球，经水解、氨基化修饰，以戊二醛作间隔臂偶联配基对氨基苯甲脒，制得磁性PVA亲和微球，用于纳豆激酶粗酶液的分离纯化，对所制微球进行了表征，研究了纳豆激酶的吸附平衡、吸附和解吸动力学、酶的纯度和酶活回收. 结果表明，磁性微球具有较高的比饱和磁化强度(32.4 emu/g)，40 min即可达到纳豆激酶的吸附平衡，15 min内完成酶的解吸，酶活回收率接近75%，纯化因子为10.3，酶的分子量为28 kDa，电泳分析为一条带.

关键词: 纳豆激酶, 磁性PVA微球, 亲和分离, 吸附, 动力学