改性磁性壳聚糖微球固定化乳糖酶

Immobilization of lactase on modified magnetic chitosan microspheres

通过反相悬浮聚合法,以甲基丙烯酸2-羟乙酯(HE-MA)与甲基丙烯酸缩水甘油酯(GMA)为单体,过硫酸铵为引发剂制备得到改性磁性壳聚糖微球。进一步以改性磁性壳聚糖微球为载体,通过吸附、共价结合以及戊二醛交联反应三方协同作用固定乳糖酶。对影响固定化的各种因素进行优化,确定固定化乳糖酶最适条件为:载体在0.1 mol/L、pH 7.0的磷酸缓冲液中充分溶胀后,按2.0 U/mg载体的添加量加入乳糖酶,4℃吸附3 h,再添加0.1%戊二醛交联4 h;最终所得的固定化乳糖酶活为685 U/g载体,酶活回收率为34.3%。固定化后的乳糖酶的pH稳定性和热稳定性都较游离酶有明显提高;连续操作10次后,固定化酶活仍保持在70%以上,具有良好的操作稳定性。

Modified magnetic chitosan microspheres（MMCM） were prepared with reversed-phase suspension polymerization method using HEMA and GMA as monomers and ammonium persulfate as initiator.Lactase was immobilized on MMCM via adsorption,covalent bonding and cross-linking with glutaraldehyde.The optimum conditions of lactase immobilization were： MMCM were previously swelled in 0.1 M pH 7.0 phosphate buffer sufficiently,lactase was added to 2.0 U per mg of carrier and incubated at 4 ℃ for 3 h,reacted with 0.1% glutaraldehyde for 4 h.The activity of immobilized lactase was 685.14 U/g and the activity recovery was 34.3%.In comparison with the free enzyme,the pH and thermal stability of immobilized lactase were increased.After continuous operation for 10 cycles,the immobilized lactase activity was remained over 70%.