摘要
γ-聚谷氨酸(γ-PGA)作为一种对人体和环境无毒害的新型高分子材料，被广泛应用于农业、化妆品、医药制造、生物医学、食品加工和水处理等许多领域，具有极大开发利用价值和广阔应用前景。
本文探讨了γ-聚谷氨酸的强酸水解条件，确定了γ-PGA的最佳水解工艺为：加入4倍体积的6mol/L HCl、90℃的恒温加热下水解24h，并使用生物传感分析仪快速准确测定水解产物谷氨酸的含量。通过实验研究绘制出纳豆芽孢杆菌生长曲线，从而获得菌种的生长对数期末期在培养15h左右，并以此作为接种材料。并通过单因素实验和正交实验优化了培养基组成和培养条件，在优化条件下固态发酵生产γ-聚谷氨酸的粗产量为105.37g/（kg固体基质），比实验优化前的平均水平85.52 g/（kg固体基质）增加了18.8%，经生物传感分析仪测定产品水解前后的含量，结果显示粗产品中γ-PGA含量为45.6%。
关键词：γ-聚谷氨酸；生物传感器；谷氨酸；纳豆芽孢杆菌；固态发酵
Abstract
Poly-γ-glutamic acid(γ-PGA)，as a new polymer material， is widely applied in many areas such as agriculture，cosmetics and pharmaceutical manufacturing, biological medicine, food processing and water treatment， due to its harmless property to human body and environment. Then, it shows great exploitation value and broad application prospect.
In this paper, the condition of acid hydrolysis of γ-PGA was researched, and the optimal condition of acid hydrolysis was obtained as follows: the ratio of a 6 mol/L hydrochloric acid to sample solution 4∶1 (V/V), hydrolyzing 24 h at 90℃. Subsequently, glutamic acid of hydrolysate was rapidly and accurately measured with the SBA-biosensor. The growth curve of bacillus subtilis natto was drew by experimental research, and final logarithmic growth phase, in which the strain could be well material for inoculation, was discovered at about 15th hour from inoculation. The optimal culture medium and conditions were investigated by one-factor experiment and orthogonal experiment. The higher output of γ-PGA by solid fermentation in the optimum conditions was 105.37 g/(kg solid substrate), increased 18.8 percent comparing to average revel 85.52 g/(kg solid substrate) before optimizing. Then the γ-PGA content was measured by analyzing glutamic acid in hydrolysate of product before and after hydrolysis in SBA-biosensor，and the results showed that the content of product was 45.6% .
Key words: Poly-γ-glutamic acid；SBA-biosensor；glutamic acid；bacillus subtilis natto；solid fermentation
目录
- 引言 5
1.1γ- PGA的结构与性质 5
1.2γ-PGA的发酵生产 5
1.2.1产生菌 6
1.2.2生物合成途径和机制 6
1.2.3生产工艺流程 7
1.2.4γ-PGA的提取 7
1.2.5产品含量的检测 7
1.3、γ-PGA的应用 8
1.3.1 γ-聚谷氨酸在农业中的应用 8
1.3.2 γ-聚谷氨酸在日用品中的应用 8
1.3.3γ-聚谷氨酸在医药中的应用 9
1.3.4γ-聚谷氨酸作为生物医学材料的应用 9
1.3.5γ-聚谷氨酸在食品工业中的应用 9
1.3.6γ-聚谷氨酸在水处理行业的应用 10
1.3.7γ-聚谷氨酸在其它领域的应用 10
1.4展 望 11
1.5本课题的研究意义 11
2.材料与方法 11
2.1菌种 11
2.2材料与试剂 11
2.3仪器设备 12
2.4培养基 12
2.5培养方法 12
2.6生长曲线的绘制方法 13
2.7γ-PGA的提取和粗产量测定 13
2.8γ-PGA含量的测定方法 13
3.结果与讨论 13
3.1γ- PGA水解条件的探讨[17] 13
3.1.1γ-PGA水解温度的确定 14
3.1.2γ-PGA水解时间的确定 14
3.1.3γ-PGA水解酸用量的确定 15
3.2纳豆芽孢杆菌生长曲线的绘制 15
3.3培养条件的优化 16
3.3.1 250 mL摇瓶装量优化 16
3.3.2 初始水份的优化 16
3.3.3初始pH值的优化 17
3.3.4接种量的优化 18
3.3.5培养温度对γ-PGA产量的影响 18
3.3.6培养时间对γ-PGA产量的影响 19
3.4培养基组成的优化 19
3.4.1培养基固体基质的优化 20
3.4.2外加碳源的优化 21
3.4.3外加氮源的优化 22
3.4.4谷氨酸钠添加量对γ-PGA产量的影响 24
3.4.5柠檬酸钠添加量对γ-PGA产量的影响 24
3.4.6 NaCl的加入量对γ-PGA产量的影响 25
3.4.7葡萄糖、酵母粉、谷氨酸钠、柠檬酸钠含量正交实验优化 26
3.5最优化条件验证实验 27
3.6双缩脲法比较初步确定产品成分 27
4.结论 28
5.展望 28
参考文献： 29
致谢 31
