【作者】 芦国营；

【导师】 张朝晖；

【作者基本信息】 浙江工业大学 ， 生物化工， 2006， 硕士

【摘要】 本文首先利用固定化黄孢原毛平革菌限碳培养合成木素过氧化物酶，经过酶的初步分离后，对其进行固定化；研究了单酶、双酶固定化的条件、固定化酶性质；最后利用固定化酶对染料进行了降解脱色。 通过单因素和正交实验，确定聚氨脂泡沫固定黄孢原毛平革菌生产木素过氧化物酶的最优条件是：泡沫尺寸0.5×0.5×0.5cm；摇动前先静止6小时；最佳的接种量是3×10⁵个／mL；最佳温度梯度是37-33℃；最佳转速梯度是140-120rpm。 产酶培养基优化后结果是：最适碳源、氮源分别是葡萄糖和酒石酸铵，最适葡萄糖浓度2.0g／L；酒石酸铵浓度2.0g／L；维生素B₁ 0.0030g／L；苯甲醇5.0mmol／L；KH₂PO₄ 1.0g／L；MgSO₄·7H₂O 0.025g／L；CaCl₂ 0.0075g／L；微量元素液20mL／L。 采用冷冻除多糖、硫酸铵盐析、透析、超滤等方法进行发酵液进行了初步分离纯化。确定硫酸铵盐析的最佳浓度是70％，粗酶液经过三步纯化后，比酶活提高1.60倍，酶活回收率达57.9％。 以壳聚糖作载体，戊二醛作活化剂，对木素过氧化物酶进行了固定化：戊二醛浓度0.4％时效果最好，酶交联时间为150min，反应pH6.5，更多还原

【Abstract】 Lignin peroxidase （Lip） can degrade artificial synthesized substance. In this thesis, Lip production in C-limited cultures by immobilized white-rot fungus Phanerochaete chrysosporium was first studied. Secondly, Lip in the broth was simply purified, then immobilized on chitosan. Finally, the immobilized enzyme was used to decolorize dyes.The fermentation conditions were optimized by single-factor experiment and orthogonal experiment. The optimum conditions were as follows: size of polyurethane foam was 0.5×0.5×0.5cm; flasks should stay 6 hours still before rotation; amounts of inoculation was 3×10⁵/mL; temperature shifting was 37-33℃; rotate speed shifting was 140-120rpm.The optimum carbon and nitrogen source was glucose and ammonium tartrate respectively. The optimum fermentation medium were composed of as follows: glucose 2.0g/L; ammonium tartrate 2.0g/L; VB₁ 0.0030g/L; benzyl alcohol 5.0mmol/L; KH₂O₄ 1.0g/L; MgSO₄·7H₂O 0.025g/L; CaCl₂更多还原