大规模茶叶细胞悬浮培养茶氨酸合成工艺优化研究

以婺源绿茶为材料进行茶叶愈伤组织悬浮培养,采用正交实验设计进行了大规模茶叶细胞悬浮培养合成茶氨酸工艺条件优化研究。结果显示,NH4+/NO-30.0/60.0mmol/L、K+100.0mmol/L、Mg++3.0mmol/L、H2PO-43.0mmol/L、蔗糖30.0g/L、水解酪蛋白2.0g/L条件下,茶叶细胞生长量(速率)和茶氨酸积累量均达到最高值;提高培养基中蔗糖和水解酪蛋白浓度可延长细胞对数生长期和稳定生长期,从而有利于茶氨酸积累;H2PO-4浓度主要影响细胞生长速率和茶氨酸积累速率的同步性,低H2PO4-浓度环境中茶氨酸积累速率峰值滞后于细胞增长速率峰值,高H2PO4-浓度环境中早于细胞生长速率峰值出现时间;K+和Mg++对细胞生长的影响不明显,但影响茶氨酸合成酶活性,维持适量的K+和Mg++有利于茶氨酸积累。先加入一定量盐酸乙胺再每天进行少量补充,茶氨酸合成量比一次性加入的效果要好。从生产效率考虑,培养周期以19～22d为宜。

Study on theanine biosynthesis in tea cell suspension culture

The suspension culture on the callus obtained from Wuyuan tea were conducted in a fermenter and then the effects of different factors on the growth of tea cells and the amount of synthesized theanine were analyzed.The result showed:the theanine accumulation peaked between nineteenth day and twenty-second day in a culture period.The growth of tea cells and the amount of theanine presented an optimal increment,when there are NH 4/NO-30.0/60.0 mmol/L,K 100.0 mmol/L,Mg 3.0 mmol/L,H2PO-4 3.0 mmol/L,sugar 30.0 g/L,protein 2.0 g/L in the medium.The log phase and the stationary phase were enhanced when the amount of sugar and protein raised in the medium.The simultaneity of tea cells growth and theanine accumulation was changed by the different amount of H2PO-4 in the medium.The activity of theanine-synthatase(L-glutamate:ethylamine lingase)was effected by K and Mg .Accumulating-content of theanine could greatly raised when ethyl-amine(ZtNH2-HCl)added,with multi-addition ZtNH2-HCl performing better than the one-time addition of ZtNH2-HCl at a specific amount at first and then a smaller amounts.