纳米磁球固定化木瓜蛋白酶的制备及酶学性质研究  

Preparation and enzymatic properties of papain immobilized by nano-magnetic spheres

作　　者：梁杰 林国荣 周凤超 杨磊 邹汉勋 刘涛 章丽 郭天章 蔡力锋 LIANG Jie;LIN Guorong;ZHOU Fengchao;YANG Lei;ZOU Hanxun;LIU Tao;ZHANG Li;GUO Tianzhang;CAI Lifeng(College of Environmental and Biological Engineering,Putian University,Putian 351100,China;Key Laboratory of Ecology-toxicological Effects&Rubber Research Institute,Putian 351100,China;Key Laboratory of Ecological Environment and Information Atlas,Fujian Provincial University,Putian 351100,China;Fujian Province Sailor Foods Co.Ltd.,Putian 351100,China)

机构地区：[1]莆田学院环境与生物工程学院,福建莆田351100 [2]福建省新型污染物生态毒理效应与控制重点实验室,福建莆田351100 [3]生态环境及其信息图谱福建省高校重点实验室,福建莆田351100 [4]福建省水手食品有限公司,福建莆田351100

出　　处：《食品与发酵工业》2024年第8期161-168,共8页Food and Fermentation Industries

基　　金：福建省自然科学基金(2021J011107,2022J011160)。

摘　　要：为实现木瓜蛋白酶(papain,PAP)的固定化,通过悬浮聚合法在磁性微球Fe_(3)O_(4)表面均匀包覆聚甲基丙烯酸甲酯(polymethylmethacrylate,PMMA),制备核-壳结构的磁性纳米粒子Fe_(3)O_(4)@PMMA,进一步使PMMA微球表面酯解形成羧基,接枝聚乙烯亚胺(polyethyleneimine,PEI),将PAP物理吸附在PEI—1800长链上,通过交联剂戊二醛(glutaraldehyde,GA)对PEI进行交联封装,构筑具有磁响应且表面富含氨基的亲水性三维立体结构的载体Fe_(3)O_(4)@PMMA-PEI 1800,实现PAP在纳米磁球表面的封装固定。对载体的形貌特征和理化性质表征分析,评价固定化酶Fe_(3)O_(4)@PMMA-PEI 1800-PAP的酶学性质。结果表明,Fe_(3)O_(4)@PMMA-PEI 1800比表面积为151.30 m^(2)/g,孔容为0.27 cm^(3)/g。Fe_(3)O_(4)@PMMA-PEI 1800-PAP最适温度为60℃,最适pH为6.0,在40~80℃维持80%以上的相对酶活力,在90℃时依然能保持66.84%以上的相对酶活力。Fe_(3)O_(4)@PMMA-PEI 1800-PAP热稳定性良好,在60℃下温育2 h后能保留80%以上的酶活力,在90℃温育2 h后仍能保留36.63%酶活力。Fe_(3)O_(4)@PMMA-PEI 1800-PAP在pH 5.0~pH 9.0均能维持80%以上相对酶活力。经过7次重复使用与洗脱之后,Fe_(3)O_(4)@PMMA-PEI 1800-PAP酶活力依然保留在70%以上。To realize the immobilization of papain(PAP),core-shell magnetic nanoparticles Fe_(3)O_(4)@PMMA were prepared by uniform coating of polymethylmethacrylate(PMMA)on the surface by suspension polymerization.Furthermore,the surface of PMMA microsphere was deoxidized to form carboxyl group,following by grafted on Polyethyleneimine(PEI).To construct a hydrophilic three-dimensional spatial structure Fe_(3)O_(4)@PMMA-PEI 1800 with magnetic response and amino rich surface,the PAP was physically absorbed on PEI-1800 chain and was crosslinked by glutaraldehyde(GA).The PAP was then encapsulated and fixed on the surface of the nano-magnetic ball.The morphology and physicochemical properties of the carrier were analyzed,and the enzymatic properties of the immobilized enzyme Fe_(3)O_(4)@PMMA-PEI 1800-PAP were evaluated.Results indicated that the specific surface area of Fe_(3)O_(4)@PMMA-PEI 1800 was 151.30 m^(2)/g and the pore volume was 0.27 cm^(3)/g.The optimum temperature of Fe_(3)O_(4)@PMMA-PEI 1800-PAP was 60℃,with an optimum pH at 6.0,the relative enzyme activity above 80%was maintained in the range of 40~80℃,and it can be maintained above 66.84%at 90℃.Fe_(3)O_(4)@PMMA-PEI 1800-PAP showed good thermal stability and 80%and 36.63%enzyme activity was retained after incubation at 60℃and 90℃for 2 h,respectively.The relative enzyme activity of Fe_(3)O_(4)@PMMA-PEI 1800-PAP was maintain over 80%during the pH range from 5.0 to 9.0.After 7 times of reuse and elution,the activity of Fe_(3)O_(4)@PMMA-PEI 1800-PAP enzyme remained above 70%.

关 键 词：纳米磁球 固定化 木瓜蛋白酶 接枝 聚乙烯亚胺 交联封装 酶学性质 

分 类 号：TS201.25[轻工技术与工程—食品科学]