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

食品科学 ›› 2005, Vol. 26 ›› Issue (8): 255-259.

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

刘剑虹, 庞广昌, 吴瑞巍

天津商学院食品科学与工程系

出版日期:2005-08-15 发布日期:2011-09-19

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

LIU Jian-Hong, PANG Guang-Chang, WU Rui-Wei

The Department of Food Science and Engineering , Tianjin University of Commerce

Online:2005-08-15 Published:2011-09-19

摘要/Abstract

摘要： 本文探讨了利用了(NH4)2SO4分级沉淀、透析脱盐及Q-SepharoseFF层析分离纯化酪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CGMPQ-SepharoseFF层析条件为:pH8.5、20mmol/LTris缓冲液平衡上样,用含0.3mol/LNaCl、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%(以酪蛋白计),总结合态唾液酸回收率为87.4%(以酶解液中的唾液酸计),糖基化度可提高到0.472以上。整个工艺路线简便、快捷、成本较低,适合工业化生产。

关键词: 酪蛋白糖巨肽(CGMP), 胃蛋白酶, 盐析, 离子交换层析, 唾液酸

Abstract: 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.

Key words: casein glycomacropeptide (CGMP), pepsin, salting-out, ion-exchange chromatography, sialic acid

刘剑虹, 庞广昌, 吴瑞巍. 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[J]. 食品科学, 2005, 26(8): 255-259.

LIU Jian-Hong, PANG Guang-Chang, WU Rui-Wei. Isolation and Purification of Casein Glymacropeptide from Pepsin Casein Hydrolysate by Salting-out and Q Sepharose FF[J]. FOOD SCIENCE, 2005, 26(8): 255-259.

[1]	范三红，王娇娇，白宝清，张锦华. 三相盐析萃取分离辣椒红色素和辣椒碱及不同辣椒品种的聚类分析[J]. 食品科学, 2020, 41(20): 270-277.
[2]	王婷，田迎樱，李雪静，王菲，王静凤. 体外模拟胃肠道对鲫鱼卵唾液酸糖蛋白的降解作用[J]. 食品科学, 2020, 41(17): 145-152.
[3]	吴伟，何莉媛，黄慧敏，吴晓娟，林亲录. 米糠酸败对米糠蛋白体外胃蛋白酶消化产物结构特征的影响[J]. 食品科学, 2019, 40(17): 14-21.
[4]	吴金勇，陈祥松，余超，陆姝欢，李翔宇，姚建铭. 大肠杆菌离子束诱变结合呼吸商在线实时调控高产聚唾液酸[J]. 食品科学, 2019, 40(14): 54-62.
[5]	曾颖，余垒，朱新儒，李雄，张海德. 盐析法联合离子液体双水相纯化木瓜蛋白酶[J]. 食品科学, 2018, 39(24): 261-267.
[6]	程芬芬，刘春，杨晓泉. 大豆胰蛋白酶抑制剂的制备及性质[J]. 食品科学, 2017, 38(3): 37-44.
[7]	王中江，张潇元，隋晓楠，齐宝坤，江连洲，李欣芮，姜楠，黄天赐，张鑫，李杨. 热处理大豆蛋白体外消化产物结构特征分析[J]. 食品科学, 2017, 38(1): 20-26.
[8]	何传波，魏好程，熊何健，吴国宏. 枇杷叶多糖酶法提取工艺优化及其离子交换层析纯化[J]. 食品科学, 2016, 37(8): 45-50.
[9]	王璐莎1，张首玉2，黄明1,*，周光宏1. 鸭肉蛋白源抗氧化肽的酶法制备工艺[J]. 食品科学, 2015, 36(7): 90-96.
[10]	田盼盼，程超，汪兴平. 逐级盐析法结合双水相萃取纯化葛仙米藻蓝蛋白[J]. 食品科学, 2015, 36(24): 16-22.
[11]	苏伟，杨旭卉，王瑜，母应春，邱树毅. 响应面试验优化基于胶原特性酶解猪皮工艺及其抗氧化特性[J]. 食品科学, 2015, 36(24): 70-76.
[12]	胡志和，夏磊，孙振刚，武文起，冯永强，薛璐，刘蓄瑾，贾莹. 酪蛋白水解物中ACE抑制肽分离及氨基酸序列分析[J]. 食品科学, 2015, 36(24): 156-163.
[13]	张发宇，赵冰冰，陈裕，袁梦媛，汪家权. 两步盐析联合双水相萃取提取纯化蓝藻中藻蓝蛋白[J]. 食品科学, 2015, 36(22): 6-10.
[14]	刘小红，李诚*，付刚，苏赵. 猪股骨头胶原蛋白降血压肽的分离纯化[J]. 食品科学, 2014, 35(6): 50-54.
[15]	董颖，张鹤，周光新，赵大庆*，赵雨，丛德毓. 正交试验优化哈蟆油蛋白双酶法水解工艺[J]. 食品科学, 2014, 35(6): 36-39.