发酵透明质酸蛋白脱除方法的比较研究

Comparative Study on Deproteinization Technologies for Crude Fermentation Hyaluronic Acid

以发酵的透明质酸(HA)粗品为纯化对象,以改良的咔唑法和考马斯亮蓝法为检测手段,比较了Sevage法、固定化蛋白酶-Sevage法、Sevage-CPC法、冻融-Sevage法脱除HA粗品中蛋白的效果。研究了有机相比例、振摇时间、HA粗品浓度对Sevage法脱除蛋白的影响,结果表明,Sevage法脱除蛋白的最优条件为:HA粗品浓度10 g.L-1、正丁醇和三氯甲烷的体积比1∶4、振摇时间30 min。通过比较,发现Sevage法HA损失率低但蛋白脱除效果差;固定化蛋白酶-Sevage法蛋白脱除效果好,操作简便,适于工业化;冻融-Sevage法操作简单,蛋白脱除效果好,但成本高,适于小批量生产;Sevage-CPC法可得到最纯的HA,但成本高、工艺繁琐,CPC不能回收,有待于进一步的研究。

Taking the improved carbazole method and Coomassie brilliant blue method as test methods, the influences of sample concentration, shaking time and the volume ratio of n-butanol to trichloromethane on the crude fermentation hyaluronic acid purifying by Sevage method were studied. Taking the loss of HA and the removal of protein in sample as symbol,the different deproteinization technologies were compared. The optimum conditions of Sevage method were as follows： the volume ratio of n-butanol to trichloromethane of 1 ： 4, crude HA concentration of 10 g· L^-1 and shaking time of 30 min. The deproteinization efficiency of Sevage method is worse than other combined methods. Immobilized enzyme-Sevage method is simple and industrialized. Frozenthawed-Sevage method is simple and excellent, but it costs too much. Sevage-CPC method can be used to get almost pure HA, but it is complicated and costs too much. There is also a problem in CPC recovery which needs further research.