食品科学 ›› 2018, Vol. 39 ›› Issue (20): 92-98.doi: 10.7506/spkx1002-6630-201820014

• 食品化学 • 上一篇    下一篇

麦胚降血糖肽的分离纯化及鉴定

颜辉1,张琦1,江明珠1,朱胜虎2,聂旭东2,余永建2,张佳鑫1,贾俊强1,熊孟1   

  1. (1.江苏科技大学生物技术学院,江苏?镇江 212018;2.江苏恒顺集团有限公司,江苏?镇江 212028)
  • 出版日期:2018-10-25 发布日期:2018-10-24
  • 基金资助:
    江苏省镇江市重点研发计划项目(GY2015006)

Isolation and Structural Identification of Hypoglycemic Peptides from Wheat Germ Protein

YAN Hui1, ZHANG Qi1, JIANG Mingzhu1, ZHU Shenghu2, NIE Xudong2, YU Yongjian2, ZHANG Jiaxin1, JIA Junqiang1, XIONG Meng1   

  1. (1. School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang 212018, China;2. Jiangsu Hengshun Group Co. Ltd., Zhenjiang 212028, China)
  • Online:2018-10-25 Published:2018-10-24

摘要: 建立麦胚降血糖肽的分离纯化方法及鉴定活性多肽的结构组成。采用胰蛋白酶酶解麦胚蛋白,得到降血糖多肽,并进行降血糖动物实验。超滤获得高活性组分,离子交换吸附实验选择最佳的树脂,并对麦胚降血糖肽进行离子交换吸附分离,SephadexG-25和SephadexG-15分离,再经过反相高效液相色谱(reverse phase-high performance liquid chromatography,RP-HPLC)分离高活性成分。最后采用高效液相色谱-电喷雾-质谱(HPLC-electrospray ionization-mass spectrometry,HPLC-ESI-MS)鉴定活性多肽的结构组成。结果显示酶解麦胚蛋白对α-葡萄糖苷酶的抑制性IC50为10.98?mg/mL,能够缓解糖尿病小鼠的症状。超滤获得分子质量小于5?kDa的高活性组分,IC50为1.60?mg/mL。732型阳离子交换树脂的分离效果最好,2.5%氨水的洗脱率为98.13%,通过动态离子交换吸附分离后,活性显著提高,IC50为0.30?mg/mL。通过SephadexG-25和SephadexG-15分离得到高活性组分,经RP-HPLC分离得到8?个组分(峰),其中1号峰的活性和含量都最高,IC50为0.098?mg/mL。HPLC-ESI-MS结果显示m/z?274.45的丰度比较高,经离子碎片拼接,共有7?种多肽,其中二肽有1?种,三肽有6?种。本研究对于开发具有降血糖功效的新型功能性食品有一定的作用。

关键词: 麦胚, 降血糖多肽, α-葡萄糖苷酶, 分离纯化, 结构鉴定, 动物实验

Abstract: The objective of the present study was to purify and structurally identify hypoglycemic peptides from wheat germ protein. The protein was hydrolyzed with trypsin to obtain the hypoglycemic peptides, and the hypoglycemic effect was evaluated by animal test. The active peptides were obtained by ultrafiltration. The optimum resin was selected for purification of the peptides by ion exchange adsorption, and further purification was carried out by sequential column chromatography on SephadexG-25 and SephadexG-15 before separation by reverse phase-high performance liquid chromatography (RP-HPLC). Finally, the structures of the purified peptides were identified by HPLC-electrospray ionization-mass spectrometry (HPLC-ESI-MS). The results showed that the enzymatic hydrolysate had α-glucosidase inhibitory activity with an IC50 of 10.98 mg/mL, and could relieve symptoms in diabetic mice. The highly active peptides with molecular mass less than 5 kDa were obtained by ultrafiltration, and their IC50 was 1.60 mg/mL. 732 cation exchange resin showed the best separation efficiency, and the elution percentage with 2.5% ammonia was 98.13%. After ion exchange adsorption, the hypoglycemic activity was significantly improved, yielding an IC50 of 0.30 mg/mL. Eight peaks were obtained after purification by RP-HPLC and among these the amount and activity of peak 1, with an IC50 of 0.098 mg/mL, were both highest. HPLC-ESI-MS analysis showed that the abundance at m/z 274.45 was high and that 7 peptides including 1 dipeptide and 6 tripeptides were obtained by ion fragment re-assembling. The findings obtained in this work are of significant importance for the development of novel functional foods with hypoglycemic efficacy.

Key words: wheat germ, hypoglycemic peptides, α-glucosidase, isolation and purification, structural identification, animal test

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