粉被虫草液体培养条件的优化

Studies on Optimization Conditions of Liquid Culture of Cordyceps pruinosa

通过摇瓶液体培养正交试验获得了粉被虫草的液体培养条件,即蔗糖2.5%,马铃薯淀粉2%,黄豆0.5%,牛肉膏0.5%,酵母膏0.1%,K2HPO4 0.1%,KCl 0.02%,MgSO4·7H2O 0.05%,pH5～7,装液量(100ml/250ml摇瓶),加15颗玻璃珠作分散剂,5%接种量,旋转式摇床150r/min,28℃培养7d,达到最大菌丝干重31.86g/L,于9d获得最大胞内产物3.13g/L。同时,对摇瓶液体培养和生物反应器液体深层分批培养的动力学做了较系统的研究,无论生物反应器还是摇瓶,其培养过程中相对应的pH、残糖、氨基氮、菌丝干重、胞内产物等参数变化趋势基本一致,但生物反应器培养时间大大缩短,48h即可达到最大菌丝干重29.97g/L,54h达到4.95g/L的最高胞内产物产率。生物反应器设定培养温度28℃、装液量65%、接种量10%、0.2%的消泡剂、搅拌转速350r/min、通气量(12±3)L/min、罐压1.1MPa、初始pH6.5,培养基配方用食用蔗糖3%、蔗糖糖蜜2%、花生0.5%分别替代蔗糖、马铃薯淀粉、牛肉膏以降低成本。实验表明,粉被虫草的液体培养条件基本能达到工业发酵水平的要求。

Liquid culture conditions of Cordyceps pruinosa Petch were investigated through orthogonal test in shake flaskculture. The contents and constituents of the optimal liquid culture medium were sucrose 2.5%, potato starch 2%, soybean0.5%, beef extract 0.5%, yeast extract 0.1%, K2HPO4 0.1%,KCl 0.02%,MgSO4·7H2O 0.05% and so on. The suitableconditions included initial pH 5.0～7.0, inoculation size 5%(V/V), medium capacity 100ml in 250ml flask, and 15 glass beadsas dispersants, cultured at 28℃ on rotary shaker at 150r/min for 7 days. The maximum mycelial production indicated 31.86g/Lin shake flask culture. And in 10th day, its intracellular product reached the maximum yield of 3.13g/L. During shake flask cultureand bioreactor submerged batch culture,the kinetics of growth period of the fungus were analysed with pH, residual sugarconcentration, amino-nitrogen concentration, mycelial dry weight, intracellular product and so on. Then, the variations ofbiochemical parameters were similar under different culture conditions. However, the culture time of bioreactor submerged batchculture was less than shake flask culture. The maxmium mycelial dry weight 29.97g/L for 48h and the maximum intracellularproduct yield 4.95g/L for 54h was obtained. Liquid submerged batch culture was carried out in a autocontrol bioreactor andculture was conducted under the following conditions: temperature, 28℃; medium capacity, 65%(V/V); aeration size,(12±3)L/min; agitation speed, 350r/min; jar-press, 1.1MPa; inoculation size, 10%; defoamer, 0.2%; initial pH, 6.5 with mediumcomponents (edible sucrose 3%, sucrose molasses 2%, peanut 0.5%, soybean 0.5%, beef extract 0.5%, yeast extract 0.1%,K2HPO4 0.1%, KCl 0.02%, MgSO4·7H2O 0.05%.). The results of this study suggested that the liquid culture conditions ofC. pruinosa have come up to the mid-test level of industrial fermentation of fungal mycelia.