不同等电点沉淀法和超速离心法提取牛奶乳清蛋白的双向电泳分析

doi:10.7506/spkx1002-6630-201420036

食品科学

不同等电点沉淀法和超速离心法提取牛奶乳清蛋白的双向电泳分析

陈静廷1,2，卜登攀1,3,4,*，马露1，杨永新1，李发弟2

1.中国农业科学院北京畜牧兽医研究所，动物营养学国家重点实验室，北京 100193；
2.甘肃农业大学动物科学技术学院，甘肃兰州 730070；
3.中国农业科学院-世界农用林业中心，农用林业与可持续畜牧业联合实验室，北京 100193；
4.东北农业大学食品安全与营养协同创新中心，黑龙江哈尔滨 150030

出版日期:2014-10-25 发布日期:2014-11-07
基金资助:
中国农业科学院科技创新工程项目（cxgc-ias-07）；“十二五”国家科技支撑计划项目（2012BAD12B02-5）；
动物营养学国家重点实验室自主研究课题（2004DA125184G1103）

2-DE Analysis of Whey Protein Extracted by Different Isoelectric Precipitations and Ultracentrifugation Methods from Cow Milk

CHEN Jing-ting1,2, BU Deng-pan1,3,4,*, MA Lu1, YANG Yong-xin1, LI Fa-di2

1. State Key Laboratory of Animal Nutrition, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing
100193, China; 2. College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China;
3. CAAS-ICRAF Joint Laboratory on Agroforestry and Sustainable Animal Husbandry, Beijing 100193, China;
4. Synergetic Innovation Center of Food Safety and Nutrition, Northeast Agricultural University, Harbin 150030, China

Online:2014-10-25 Published:2014-11-07

摘要/Abstract

摘要：

为探索牛奶乳清蛋白提取的最适方法，以生鲜荷斯坦牛奶为对象，采用不同等电点（pH 4.6和pH 4.8）沉淀法和超速离心法提取牛奶乳清蛋白样品，并对提取效果进行双向凝胶电泳（two-dimensional electrophoresis，2-DE）图谱分析。依据凝胶图谱上蛋白斑点比较图谱质量，采用PDQuest 8.0凝胶图像分析软件进行凝胶图谱分析。结果显示：2 种方法均能有效提取牛奶乳清蛋白，且获得的2-DE凝胶图谱背景清晰、蛋白点个体独立、无明显的拖尾现象，且重复性强，但都存在一定的酪蛋白残留。与超速离心法相比，pH 4.6沉淀法和pH 4.8沉淀法提取乳清蛋白的2-DE凝胶图谱可检测到较多的蛋白质斑点。pH 4.6沉淀法提取乳清蛋白制备的2-DE凝胶图谱中蛋白点的表达丰度略高于pH 4.8沉淀法。研究表明：pH 4.6沉淀法提取乳清蛋白制备2-DE凝胶图谱略优于pH 4.8沉淀法和超速离心法。但2 种等电点沉淀法和超速离心法都可有效去除牛奶中的高丰度酪蛋白，提高低丰度蛋白的检出敏感性。

关键词: 牛奶, 乳清蛋白, 等电沉淀, 超速离心, 提取方法, 双向凝胶电泳图谱

Abstract:

The objective of this study was to investigate the effect of extraction methods on two-dimensional electrophoresis (2-
DE) maps of whey proteome in cow milk and to explore an optimal extraction method. Samples were extracted from cow milk
by different isoelectric precipitations and ultracentrifugation methods. 2-DE maps were analyzed by PDQuest 8.0 software. The
results of 2-DE maps showed that milk whey protein could be effectively extracted by the above methods with less background,
no significant strips and good repeatability. However, there were still some residual caseins appearing in each map. 2-DE maps
of whey protein refined by isoelectric precipitations were relatively informative when compared with the gel maps obtained by
ultracentrifugation. Moreover, the richness of different whey proteins in various maps extracted by adjusting pH to 4.6 as isoelectric
point was slightly higher than adjusting pH to 4.8. The results indicated that adjusting pH to 4.6 as isoelectric point to extract whey
protein had some advantages than adjusting pH to 4.8 and ultracentrifugation. However, all the methods used in this study could
effectively remove high abundant casein to improve the detection sensitivity of low abundance proteins.

Key words: cow milk, whey protein, isoelectric precipitation, ultracentrifugation, extraction methods, two-dimensional electrophoresis map

中图分类号:

S823

陈静廷1,2，卜登攀1,3,4,*，马露1，杨永新1，李发弟2. 不同等电点沉淀法和超速离心法提取牛奶乳清蛋白的双向电泳分析[J]. 食品科学, doi: 10.7506/spkx1002-6630-201420036.

CHEN Jing-ting1,2, BU Deng-pan1,3,4,*, MA Lu1, YANG Yong-xin1, LI Fa-di2. 2-DE Analysis of Whey Protein Extracted by Different Isoelectric Precipitations and Ultracentrifugation Methods from Cow Milk[J]. FOOD SCIENCE, doi: 10.7506/spkx1002-6630-201420036.

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不同等电点沉淀法和超速离心法提取牛奶乳清蛋白的双向电泳分析

2-DE Analysis of Whey Protein Extracted by Different Isoelectric Precipitations and Ultracentrifugation Methods from Cow Milk

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