条斑紫菜凝集素的提取及其性质研究

Researches on Extraction,Purification Technique and Properties of Lectin from Porphyra yezoensis

[目的]研究条斑紫菜(Porphyra yezoensis)凝集素的提取及分离纯化条件,并测定凝集素的性质。[方法]以比活力为指标,研究了缓冲体系、固液比、温度及时间等条件对条斑紫菜凝集素提取效果的影响。采用DEAE-Sepharose FF离子交换层析技术进行分离,确定了分离的最佳条件。以血凝集活性为指标,测定了凝集素的糖专一性、2价阳离子依赖性、温度及酸碱稳定性等性质。[结果]固液比为1∶5的Tris-HCl(25 mmol/L,pH值7.5)缓冲液在40℃条件下提取20 h得到的提取液具有最高比活力。条斑紫菜凝集素在50℃条件下加热30 min后活性保持不变,表现出一定的热稳定性,最适作用pH值为7～9。该凝集素能够与麦芽糖、半乳糖、阿拉伯糖及木糖结合,其中与麦芽糖的结合最强,凝集活性依赖Ca2+、Mg2+等2价阳离子。[结论]为阐明条斑紫菜的免疫机制及其高效综合利用提供了理论依据。

[Objective] The aim was to explore the extraction technique and purification method of lectin from Porphyra yezoensis,as well as to investigate the properties of lectin.[Method] The effects of four factors such as buffer system,solid-liquid ratio,temperature,and extracting time on the lectin extraction result from P.yezoensis were investigated by the use of specific activity as the indicator.Then the lectin was purified by DEAE-Sepharose FF chromatography.The hemagglutination activity was used as the indicator,and the properties of the lectin such as carbohydrate specificity,divalent cations dependence,temperature,acidic and alkaline stability and others were detected.[Result] The optimal extraction conditions were：extraction ageat Tris-HCl buffer（25 mmol/L,pH=7.5）,solid-liquid ratio=1∶5,extracting temperature 40 ℃,and extracting time 16 h.The activity of the lectin of P.yezoensis didn＇t changed after 30 min of heating at 50 ℃,which showed a certain extent of heat stability,and the suitable PH value was 7-9.The lectin of P.yezoensis could combine with maltose,D-galactose,D-xylose and L-Arabinose,in which the binding force with the maltose was the strongest.The lectin activity was depended on the addition of Ca2＋ and Mg2＋.[Conclusion] The research had provided theoretical basis for the clarification of immune mechanism of P.yezoensis,as well as its high effective utilization.