采用盐析及Q Sepharose FF分离纯化酪蛋白胃蛋白酶水解物中的酪蛋白糖巨肽

本文探讨了利用了（NH4）2SO4分级沉淀、透析脱盐及Q—Sepharose FF层析分离纯化酩蛋白的胃蛋白酶水解物中的CGMP的工艺。结果表明：采用60％饱和度的（NH4）2SO4盐析，可从上清液中直接回收高纯度CGMP，得率为0．71％。低饱和度的（NH4）2SO4可有效脱除杂蛋白。脱盐粗提物CGMP Q—Sepharose FF层析条件为：pH8．5、20mmol／L Tris缓冲液平衡上样，用含0．3mol／L NaCl、20mmol、pH7．1的Tris缓冲液洗脱：浓缩物最大上样量为34．2mg蛋白／ml树脂。利用优化的工艺对水解液层析纯化，纯化产物得率约为2．19％（以酪蛋白汁），总结合态唾液酸回收率为87．4％（以酶解液中的唾液酸汁），糖基化度可提高到0．472以上。整个工艺路线简使、快捷、成本较低，适合工业化生产。

Isolation and Purification of Casein Glymacropeptide from Pepsin Casein Hydrolysate by Salting-out and Q Sepharose FF

In this paper, the isolation and purification conditions for CGMP from pepsin casein hydrolysate were studied by ammonium sulfate precipitation, dialysis desalting and Q Sepharose fast flow chromatography. The results show that 0.71% recovery rate CGMP with high purity could be obtained immediately from salting-out supernate by 60% saturation degree ammonium sulfate precipitation;and non-CGMP proteins could be separated with 20% saturation degree ammonium sulfate effectively;Desalting fraction of crude separating mixture could be chromatographed by Q Sepharose FF equilibrated and added samples with pH 8.5, 20mM Tris buffer and eluted with 20mM Tris buffer, pH 7.1containing 0.3M NaCl; and the maximal bearing capacity was 34.2mg protein for 1ml resin. With that optimizing purification conditions, the estimated yield of CGMP fraction was approximately 2.19% of casein; its sialic acid recovery rate to entire content in hydrolysate supernatant was 87.4% and glycosylation ratio of purified CGMP could be raised to over 0.472. The entire processing condition was simple, fast ,low cost and suitable for industry.