透膜法生产L-天冬氨酸的工艺研究

Study on the enzymatic synthesis process of aspartic acid by increasing cell membrane permeability of Escherichia coli

为提高天冬氨酸的转化效率,对大肠杆菌发酵产酶条件和发酵转化液的酶促反应条件进行了优化,结果表明,在大肠杆菌在10 L发酵罐上溶氧维持在25%,发酵周期为8 h,发酵3 h时添加0.2%的吐温-60的条件下,天冬氨酸酶产率最高。大肠杆菌所产天冬氨酸酶为胞内酶,因此,采用表面活性剂、有机溶剂及超声波破碎法对大肠杆菌的细胞膜透性进行处理,结果表明:反应结束后发酵转化液用0.5%的甲苯处理1 h后进行底物转化,转化效果最好。在此条件下优化了酶促反应温度和pH,结果表明:温度为45℃,pH为8.5时,发酵转化液对底物的转化效果最佳。在上述条件下,10 mL发酵转化液转化90 mL质量浓度为200 g/L的富马酸底物时,转化周期在6 h以内,底物转化效率大于99%。

In order to increase the productivity of aspartic acid,the optimization of the fermentation conditions with Escherichia coli and the conditions of enzymatic reaction are investigated. The optimal fermentation condition is 25% of dissolved oxygen in 10 L fermenter for 8h,with the addition of 0. 2% Tween-60 after 3 h fermentation. As the aspartase produced by Escherichia coli is intracellular,the addition of surfactant,organic solvent and ultrasonic to increase the cell membrane preability are tested. The result suggests that the cell treated with 0. 5% toluene for 1h,keeping at 45℃ and pH 8. 5,is the most favorable for substrate transformation. Using 10 m L fermentation broth to transform 90 m L substrate（ 200 g/L fumaric acid） under the optimal condition,the conversion rate could reach 99. 6% within 6 h.