白斑狗鱼鱼肉中抗冻蛋白的分离纯化

Isolation and purification of antifreeze protein from muscle tissues of Esox lucius

利用单因素试验及响应面分析法将从白斑狗鱼鱼肉组织中获取的抗冻蛋白进行分离纯化。试验以鱼肉抗冻蛋白的热滞活性(THA)为响应值,对提取温度、pH值、提取时间、料液比4个因素进行优化。结果表明:白斑狗鱼鱼肉中抗冻蛋白提取的最佳工艺条件为温度5.14℃、pH值8.40、提取时间为1.88 h,料液比1∶2(g∶mL),此时热滞活性达到0.061 2℃。验证试验结果显示热滞活性(THA)为(0.059 4±0.001 3)℃,与理论预测值基本吻合。经DEAE-Sepharose Fast Flow阴离子交换柱和Sephadex G-50分子筛层析柱纯化后获得了电泳纯的抗冻蛋白,其相对分子质量约为7.6 ku,热滞活性大约为(0.052±0.0015)℃。

Antifreeze protein （AFP） from muscle tissues of Esox lucius was isolated and purified by single factor experiment and response surface methodology. Furthermore, the extraction time, extraction temperature, extraction buffer pH and water to material ratio was comprehensively detected with thermal hysteresis activity （THA） as the evaluation index. Results showed that the THA of AFP from muscle tissues of E. lucius was up to 0.061 2℃ under the optimized condition of extraction temperature 5.14℃, sample solutions pH 8.40, extraction time 1.88 h and solid to liquid ratio 1：2（g：ml）. In addition, the verification experiment showed that the THA was （0.059 4±0.001 3）℃, similar to the experiment value. After DEAE- Sepharose fast flow anion exchange column and Sephadex G-50 molecular chromatography column purification, AFP of electrophoresis level with 7.6 ku molecular weight was obtained by further chromatographic purification, and the THA was about （0.052±0.001 5）℃.