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目的从蝎毒中寻找一种新的镇痛多肽,考察其分离纯化的最佳条件,并对其生物活性进行研究。方法通过应用Superdex 75凝胶层析预装柱,对样品的不同处理方法、洗脱流速、不同洗脱液以及样品浓度对分离条件进行考察,采用SDS-PAGE电泳鉴定各组分纯度以及分子量。热板法和醋酸扭体法检测生物活性。结果蝎毒溶解后直接低温离心,过滤,上层析柱;样品浓度为10 mg/ml,以0.05 M PBS p H=7.2缓冲液洗脱,洗脱流速为1.0 ml/min,为其最佳分离、纯化条件。分离纯化所得的5号组分,具有很好的镇痛作用,SDS-PAGE电泳显示其呈单一条带。分子量大致为10KDa-15KDa之间。结论选用Superdex 75凝胶层析预装柱可获得一种新的具有镇痛作用的蝎毒多肽,具有良好的开发应用前景。
Objective To search for a new analgesic peptide from scorpion venom, investigate its optimal conditions for isolation and purification, and study its biological activity. Methods By using Superdex 75 gel prepacked column, the separation conditions of different samples, elution flow rate, different eluents and sample concentrations were investigated. The purity and molecular weight of each component were determined by SDS-PAGE electrophoresis. Hot plate method and acetic acid writhing method to detect biological activity. Results Scorpion venom was directly lysed by centrifugation, filtered and chromatographed on the column. The sample concentration was 10 mg / ml and eluted with 0.05 M PBS p H = 7.2 buffer at a flow rate of 1.0 ml / min, which was the best Isolation, purification conditions. Isolation and purification of the resulting No. 5 components, has a good analgesic effect, SDS-PAGE electrophoresis showed a single band. The molecular weight is roughly between 10KDa and 15KDa. Conclusion A new kind of scorpion venom peptide with analgesic effect was obtained by using Superdex 75 gel prepacked column, which has a good prospect of development and application.