[目的]探索固定化漆酶酶促合成茶黄素的工艺.[方法]采用D152树脂固定化漆酶体外酶促氧化茶多酚合成茶黄素.通过单因素试验研究反应温度、pH、酶浓度、反应时间和通氧量对合成茶黄素的影响,在此基础上进行茶黄素的酶促氧化合成反应,之后测定茶黄素产量.[结果]采用树脂载体D152固定化得到的固定化漆酶,在重复使用5次后,固定化漆酶催化活性约为原始催化活性的70％;在重复使用10次后,酶催化活性仍然可以保留48％左右.固定化漆酶促氧化的最佳条件为:反应温度60 ℃,最佳pH 5,底物浓度2.0g/L,通氧量20 L/min和反应时间1.5h.[结论]采用树脂载体D152可有效固定漆酶.与游离漆酶氧化法相比,固定化法能有效提高漆酶的稳定性和利用效率,反应产物也更易于纯化.%[Objective ] The study aimed to explore the technology of synthesizing the theaflavins through the enzyme-catalysing oxidation with the immobilized laccase. [Method] The theaflavins was synthesized from tea polyphenol through in vitro enzyme-catalysing oxidation with D152 resin immobilized laccase. The effects of the reaction temperature, pH value, enzyme concn. , reaction time and speed of adding oxygen on synthesizing the theaflavins was studied though the single factor experiment, on base of which, the synthesizing reaction through the enzyme-catalysing oxidation was conducted on the theaflavins and then the yield of the theaflavins was determined. [Result] The immobilized laccase was got by the immobilization with the resin carrier D152,the enzyme-catalysing activity by the immobilized laccase was about 70% of the original catalytic activity after the repetition use of the resin for 5 times and that still retained 48% or so after the repetition use for 10 times. The best conditions for the enzyme-catalysing oxidation with the immobilized laccase were as followsjthe reaction temperature of 60 ℃ ,best pH value of 5.0,substrate concn. Of 2.0 g/L,adding oxygen speed of 20 L/min and reaction time of 1.5 h. [Conclusion] Using the resin carrier D152 could effectively fix the laccase. Compared with the method of the enzyme-catalysing oxidation with the free laccase, the immobilized method could effectively increase the stability and utilization efficiency of the laccase and its reaction products also could be easy for the purification.
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