MCS固定化木瓜蛋白酶和菠萝蛋白酶的初步研究.doc

PAGE  PAGE   PAGE 19 Page  PAGE 0  DATE \@ M/d/yyyy 4/22/2017 M—CS固定化木瓜蛋白酶和菠萝蛋白酶的初步研究 王 志 彬 邵阳学院生物与化学工程系湖南邵阳422000 指导老师：赵良忠 摘 要 以自制壳聚糖为载体，戊二醛为交联剂,并以Span-80为乳化剂，通过化学共转化法将Fe3O4磁性纳米粒子吸附在壳聚糖微球表面；经活化，得到一种生物亲和性好、分散性好、粒度均匀、表面富含功能环氧基的磁性壳聚糖微球；对木瓜蛋白酶和菠萝蛋白酶固定化，并初步研究了木瓜蛋白酶的用量和固定时间对木瓜蛋白酶固定效果及活性的影响，研究了固定时戊二醛的浓度和固定温度对菠萝蛋白酶固定效果及活性的影响。结果表明：制备微球时，壳聚糖的最佳浓度为3%，戊二醛质量百分数为25%。制备磁性壳聚糖时，适宜操作条件为：在壳聚糖微球中加入摩尔比为1：1的Fe2+ 和Fe3+ 的混合溶液，并60℃下浸泡40min，然后加入 2mo1／L的NaOH溶液。60℃水浴中反应40min，所得的磁性壳聚糖微球外观为球形、颗粒大小均匀，具有强磁响应性，加酶量为200mg/g载体。固定化木瓜蛋白酶，最适固定温度为5℃,pH为8；固定化菠萝蛋白酶蛋白酶最适温度为23℃，pH为6.0-9.0。酶活回收率为23.7%，固定化酶在热稳定性、pH稳定性和操作稳定性上较游离酶均得到了较大幅度的提高。 关键词：磁性壳聚糖；木瓜蛋白酶；菠萝蛋白酶；酶活力 The study of Magnetic Chitosan Immobilized Trypsin and Bromelain Wang zhi-bin (Department of Biology and Chemistry Engineering,Shaoyang University,Shaoyang 422000,China) the guided teacher:Zhao liang-zhong Abstract Summarize:The study used Magnetic Chitosan asa carrier, Glutaraldehyde as a crosslingking agent and Span-80 as a emulsifier to embed the chitosan microballoons，and the chitosan microballoons of Fe3O4 that passed by the objects surface by suspengding crosslingking method ,which synthesized a type of Magnetic Chitosan Microspheres of a high-hydrophilic ,uniform size ,dispersion .At the same time ,the surface of composition was full of Funcional Epoxy .Whats more,It also took Papain and Bromelain as Model Encyme to immobilize Protease ,and the study of the effect when it immobilize Papain and Bromelain. The results showed that the best concentration of Chitosan was 3% ,the volume of Glutaraldehyde was 25% ,to make Magnetic Chitosan, the best concentration of Fe 2+Fe3+Towas 1:1, immobilize Trypsi ,the most suitable temperature was 5 centigrade ,pH 8 ;To immobilize Bromelain ,the most suitable temperature was 55 centigrade ,pH for 6.0-9.0 ,and the recovery of enzyme activity was 23.7% .It also showed a more substantional increase than Free Enzyme in thermal stability ,pH stabili