氨基磁性微球固定化木瓜蛋白酶及其应用

Immobilization of Papain on Amino Magnetic Microsphere and Its Application Study

目的:采用实验室自制氨基磁性微球固定化木瓜蛋白酶,并将其应用于啤酒中蛋白质的水解。方法:考察不同给酶量、戊二醛浓度、固定时间等因素对固定化木瓜蛋白酶的影响。将固定化酶用于啤酒的处理,测定其游离氨基酸变化。结果:实验表明给酶量1.2mg/ml,戊二醛浓度5%,固定化时间4h为固定化最佳条件。酶活力测定表明,在最优化条件下制备的固定化酶较溶液酶具有更低的Km值(0.489%∶3.412%),更好的酸碱(6.8∶7.2)和温度耐受性(77℃∶67℃),操作和储藏稳定性也得到了很大提高。固定化酶水解啤酒中蛋白质后,啤酒混浊度降低了0.013个单位,酪氨酸和苯丙氨酸等游离氨基酸有不同程度的增加,啤酒色泽和口味得到了改善。结论:磁性固定化木瓜蛋白酶较溶液酶有更佳的稳定性和更好的可操作性,具有广泛的应用前景。

Objective：To immobilize papain with the amino microspheres prepared self-made in lab,and then it was applied to hydrolysis of protein in beer.Method：The effect of those factors such as different proportion of enzyme,concentration of glutaraldehyde and immobilization time to papain immubilization was investigated.The immobilized protein was used to manipulation of beer,and the diversity of free aminoacid in beer was determined too.Result：The optimal immobilization condition is that：1.2mg/ml for proportion of enzyme,5% for concentration of glutaraldehyde and 4h for immobilization time.The measurement of enzyme activity indicated that,compared to free enzyme,immobilized enzyme which prepared on the optimal condition had lower Km （0.489%：3.412%）,better tolerance of acid base （6.8：7.2） and temperature （77℃：67℃）,the stability of manipulation and preservation enhanced greatly too.After hydrolysis of protein in beer by means of immobilized enzyme,the turbidity of beer decreased by 0.013 units,the free amino acid such as tyrosine and phenylalanine increased in different extent,both the color and taste of beer were improved.Conclusion：Magnetic immobilized papain possessed better constancy and can be manipulate much more easily compared to sluble enzyme.Magnetic immobilized enzyme had extensive application prospect.