酵母源葡萄糖耐量因子的提取纯化研究

发布时间：2018-05-23 07:25

  本文选题：富铬酵母 + 葡萄糖耐量因子　； 参考：《天然产物研究与开发》2016年11期

【摘要】：为探究葡萄糖耐量因子(GTF)分离纯化工艺,简化纯化步骤,以期得到GTF纯品,本实验在前人氨水提取基础上,采用有机溶剂沉淀除去杂蛋白,在有机溶剂的种类、溶剂的浓度、提取组分以及沉淀时间的确定等方面进行了试验,以有机铬含量(μg)与总蛋白含量(mg)的比值为纯化评价标准,高相液相色谱对纯化结果进行验证,同时将纯化前后的GTF作用于胰岛素抵抗型Hep-G2肝癌细胞,测定细胞葡萄糖消耗量,检测GTF纯化品对细胞葡萄糖代谢的调节活性。最终确定用30%低温乙醇沉淀20 min,10000×g离心取上清液分离纯化GTF,此方法相较于氨水直接提取,有机铬含量与总蛋白含量的比值提高了6倍多,GTF提取率和纯度都得到了显著提高,且纯化后的GTF对胰岛素抵抗型Hep-G2细胞葡萄糖代谢具有显著调节作用(P0.01)。
[Abstract]:In order to explore the separation and purification process of glucose tolerance factor (GTF) and simplify the purification procedure, we used organic solvent to precipitate and remove the impurity protein on the basis of ammonia water extraction, and to simplify the purification process, and to obtain the pure GTF product. The extraction components and the determination of precipitation time were tested. The ratio of organic chromium content (渭 g) to total protein content (渭 g) was used as the standard of purification evaluation. The purification results were verified by high phase liquid chromatography. At the same time, the purified GTF was applied to insulin-resistant Hep-G2 hepatoma cells, the glucose consumption of the cells was measured, and the regulating activity of the purified GTF on glucose metabolism of the cells was measured. It was determined that GTFs were separated and purified by centrifugation with 30% low-temperature ethanol for 20 min ~ 10000 脳 g centrifugation. Compared with direct extraction with ammonia, the ratio of organic chromium content to total protein content was increased by more than 6 times and the extraction rate and purity of GTF were significantly improved. The purified GTF could significantly regulate glucose metabolism in insulin resistant Hep-G2 cells.
【作者单位】： 天津科技大学食品工程与生物技术学院;中国农业科学院农产品加工研究所;
【基金】：国家自然科学基金(31371763)
【分类号】：R914
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本文编号：1923807