液氮快速冷冻对地龙中水溶性蛋白组成和活性的影响

Effect of liquid nitrogen rapid freezing technology on components of water-soluble proteins in ground dragon and their activities

目的探讨低温炮制工艺对地龙水溶性蛋白质组成以及溶栓与抗肿瘤活性的影响。方法地龙的炮制采用传统与低温两种工艺,低温炮制采用液氮快速冷冻与冷冻干燥技术;采用SDS-PAGE与ImageTool软件,对两种炮制工艺的地龙蛋白质组成的差异进行定性与定量分析;应用纤维蛋白平板法与体外溶栓试验,定性与定量分析两种炮制工艺的纤溶活性的差异;应用显微观察与MTT法,对两种炮制工艺体外抑制人肝癌细胞增殖活性进行定性与定量比较分析。结果低温炮制工艺地龙的水溶性蛋白组成比传统炮制工艺丰富,SDS-PAGE的蛋白条带数与浓度明显高于传统工艺;两种炮制工艺的地龙蛋白液均具有显著的抗人肝癌细胞与纤溶活性,但低温炮制工艺的两种活性显著高于传统工艺。结论不同炮制工艺对地龙水溶性蛋白组成及其抗肿瘤与溶栓活性有显著影响;在保全地龙的蛋白质类药效成分的生物活性方面,低温炮制工艺优于传统炮制工艺。

Objective To study the effect of cold-processing technology on the composition of water-soluble proteins in ground dragon and their activities of thrombolysis and anti-tumor activity. Methods Ground dragon was respectively processed by traditional processing technology and cold- processing technology. The latter was done by the liquid nitrogen rapid freezing technology and lyophiliz- ing technology. By means of qualitative and quantitative tests, SDS-PAGE and the software of ImageTool were applied to analyzing the difference of protein components in ground dragon from the two processing techniques. Fibrin plate method and thrombolysis test in vitro were carried out to determine the difference of the two processing techniques for the fibrinolytic activity. Micro-observation and MTT methods were used to assess the differences of antihepatic cancer activity between the traditional and the cold-processing groups. Results Compared with the traditional group, the components of water-soluble proteins in the ground dragon processed by cold-processing technique were more abundant than traditional one, and the number of protein bands and concentration in SDS-PAGE gel were more and much higher. Although the two kinds of protein solution from the different processing techniques had the visible fibrinolytic activity and antihepatic cancer activity, both activities of cold-processing technique were markedly higher than those in the traditional group. Conclusion Effects of different processing techniques of ground dragon on fibrinolytic activity and anti-tumor activity are significant. On protecting the bioactivity of proteins in ground dragon, the cold-processing techniques is advantage to the traditional processing technology.