新型层状交联酶聚集体的制备条件优化与性质研究  

Optimized preparation and characterization of novel layered cross-linked enzyme aggregates of Candida sp. lipase

作　　者：尹春华 马烨炜 赵志敏 张海洋 闫海 YIN Chunhua;MA Yewei;ZHAO Zhimin;ZHANG Haiyang;YAN Hai(Department of Biological Science and Engineering, School of Chemical and Biological Engineering, University of Science and Technology Beijing, Beijing 100083, China)

机构地区：[1]北京科技大学化学与生物工程学院生物科学与工程系,北京100083

出　　处：《化工学报》2018年第12期5192-5198,共7页CIESC Journal

基　　金：国家自然科学基金项目(21606016)~~

摘　　要：以假丝酵母脂肪酶(Candida sp. lipase)为研究对象,开发了一种新型层状交联酶聚集体。用蛋白或氨基酸对纳米氧化锌粒子进行修饰,继以交联剂交联后作为核芯,酶分子再交联在纳米核表面形成层状结构。实验结果表明牛血清白蛋白(BSA)是纳米氧化锌适宜的修饰剂。并且对纳米芯层状交联酶聚集体(BSA-N-LCLEAs)其他制备条件进行了优化,优化后BSA-N-LCLEAs制备条件为:沉淀剂硫酸铵饱和度为58%,交联剂戊二醛浓度为3.5%,交联温度和时间分别为0℃和2h。BSA-N-LCLEAs酶活收率较传统CLEAs提高了196.5%。扫描电镜表征表明BSA-N-LCLEAs较传统CLEAs孔道大幅增加。纳米芯层状CLEAs的pH稳定性和热稳定性也都比传统CLEAs有所提高,并将该固定化酶用于催化维生素E琥珀酸酯的合成,反应五批次后反应产率还能达90%左右,说明该新型交联酶聚集体具有良好的催化活性和操作稳定性。A novel layered cross-linked enzyme aggregates(CLEAs)of Candida sp.lipase was developed.In the preparation of the novel enzyme aggregates,nano-ZnO particles were modified and cross-linked,which were used as the core of the immobilized lipase,then enzyme molecules were cross-linked on the surface of the nanoparticles,which formed a layered structure.The obtained results showed the bovine serum albumin(BSA)was the suitable agent to modify nano-ZnO particles among the test agents.The other factors affecting the activity of nano layered CLEAs(BSA-N-LCLEAs),such as ammonium sulfate saturation,concentration of glutaraldehyde,temperature and time for cross-linking reaction,were investigated.The optimum conditions were as follows:ammonium sulfate saturation58%,glutaraldehyde concentration3.5%,cross-linking temperature0℃and time2h.Compared with the traditional CLEAs,the recovered activity of BSA-N-LCLEAs increased about196.5%.Scanning electron microscopy(SEM)characterization indicated that BSA-N-LCLEAs increased significantly compared to conventional CLEAs.Both pH stability and thermo stability of BSA-N-LCLEAs were improved.Furthermore,the layered CLEAs was used to synthesize astaxanthin succinate.The yields of astaxanthin were about90%.The immobilized lipase could be reused for five batches,which indicated it had good operational stability.

关 键 词：固定化 酶 生物催化 纳米粒子 交联酶聚集体 

分 类 号：TQ[化学工程]