菌株Arthrobacter sp.DNS10降解酶固定化条件的响应面优化

发布时间：2018-07-17 05:30

【摘要】：以前期分离得到阿特拉津降解菌株Arthrobacter sp.DNS10为供试菌株,选取海藻酸钠作为固定化材料,采用响应面法优化该菌株所包含的降解酶的固定化条件。借助Plackett-Burman设计筛选确定海藻酸钠浓度、固定化体系的p H值、加酶量和Ca Cl2溶液质量分数4个因素作为影响固定化酶比酶活的典型因素。借助Box-Behnken设计及响应面回归分析拟合,确定适宜上述降解酶的最佳包埋固定化条件及方法为:在每10 m L海藻酸钠浓度为1.93%,p H为8.5的固定化基质中加入983μL的降解酶液(蛋白浓度为88μg·L-1),然后利用注射器将上述混合溶液滴加到质量分数为2.7%的Ca Cl2溶液中即可制备出比酶活最高的固定化酶,实际测定固定化酶的最优比酶活为0.190 2 U·mg-1(预测值为0.187 4U·mg-1)。上述固定化酶平均粒径约为(0.44±0.01)cm,其在连续6次的使用过程中比酶活仍能保持在初始值的77.5%以上。上述固定化处理可有效的改善降解酶的环境贮存特性,固定化酶在常温下保存35 d后比酶活仍能保持在其初始状态的12.34%,而游离酶则无活性检出。
[Abstract]:Atrazine degrading strain Arthrobacter sp. DNS10 was selected as immobilized material, and the immobilization conditions of degradation enzyme were optimized by response surface method. The concentration of sodium alginate, pH value of immobilized system, the amount of enzyme added and the mass fraction of CaCl _ 2 solution were determined by Plackett-Burman design as the typical factors affecting the specific enzyme activity of immobilized enzyme. With Box-Behnken design and response surface regression analysis, The optimum immobilization conditions and methods were determined as follows: adding 983 渭 L degradation enzyme solution (protein concentration 88 渭 g L-1) to the immobilized substrate with concentration of 1.93 渭 L sodium alginate per 10 mL alginate, then using syringe to The immobilized enzyme with the highest enzyme activity can be prepared by dropping the above mixed solution into a 2.7wt% CaCl2 solution. The optimum specific activity of immobilized enzyme was 0.190 U mg-1 (prediction value 0.187 4 U mg-1). The average particle size of the immobilized enzyme was about (0.44 卤0.01) cm, and the specific enzyme activity remained above 77.5% of the initial value in the course of six successive uses. The immobilized enzyme could effectively improve the environmental storage characteristics of the enzyme. After 35 days of storage at room temperature, the immobilized enzyme could maintain its activity at 12.34% of its initial state, but the free enzyme could not be detected.
【作者单位】：东北农业大学资源与环境学院;
【基金】：黑龙江省教育厅“长江学者后备”计划项目(2012CJHB001) 黑龙江省高校科技创新团队建设计划项目(2013TD003) 东北农业大学“青年才俊”项目黑龙江省自然科学基金面上项目(C2016020)
【分类号】：X172;X592

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