低温炮制工艺对水蛭水溶性蛋白组成及纤溶活性的影响

目的:研究低温炮制工艺对水蛭水溶性蛋白组成及其纤溶活性的影响。方法:水蛭的低温炮制采用液氮快速冷冻和冷冻干燥技术;传统炮制参照2005药典;不同炮制工艺水蛭的水溶性蛋白组成的差异检测采用SDS-PAGE,定量分析采用Scion Image软件;不同炮制工艺水蛭的水溶性蛋白组分的纤溶活性鉴定采用纤维蛋白平板法,溶栓活性分析采用体外溶栓试验。结果:低温炮制工艺的水蛭水溶性蛋白组成比传统炮制工艺丰富,不仅SDS-PAGE蛋白条带的数量与浓度显著高于传统工艺,而且纤溶活性也显著高于传统炮制工艺。结论:不同炮制工艺对水蛭水溶性蛋白组成与溶栓活性有显著影响;在防止水蛭中蛋白质类活性成分的降解、变性与失活方面,低温炮制工艺优于传统炮制工艺。该工艺适用于以蛋白为主要活性成分的动物药研究。

Effect of Cryogenic Processing Technology of Leech on the Components of Water-solubililty Proteins and the Fibrinolytic Activity

Objective: To research the effect of cryogenic processing technology of leech on the components of water-solubility proteins and the fibrinolytic activity.Methods: Leech was respectively processed by traditional processing technology(TPT) and cryogenic processing technology(CPT).The former was done according to the method described in 2005 pharmacopeia of our country,and the latter adopted the quick-liquid-nitrogen-frozen technology and the freeze-drying technology.The difference of the components of water-solubility proteins in leech between the two processing technology was qualitatively and quantitatively determined by SDS-PAGE and Scion Image software.Fibrin plate method and thrombolysis test in vitro were used to qualitatively and quantitatively determine the difference between the two processing technologies in the fibrinolytic and thrombolysis activity.Results: Water-solubility proteins in leech processed by CPT were more abundant than those of TPT.Not only the number and concentration of protein bands in SDS-PAGE gel,but also the fibrinolytic and thrombolysis activity of CPT,were markedly higher than those of TPT.Conclusion: CPT is advantaged to TPT in assuring activity of peptides and proteins of leech.CPT can be theoretically applied into the processing of other animal Chinese traditional medicine in which the main active components are peptides or proteins.