鹿血抗运动疲劳肽分离纯化及其肽段分析  

Isolation and purification of anti-exercise fatigue peptides from Cervus nippon blood and its peptide fragment analysis

作　　者：杜小琴 夏炎 张小琴 吴中宝[1] DU Xiao-qin;XIA Yan;ZHANG Xiao-qin;WU Zhong-bao(Bio-resource Research and Utilization Joint Key Laboratory of Sichuan and Chongqing,Chongqing Institute of Medical Planting Material,Nanchuan 408435,China)

机构地区：[1]重庆市药物种植研究所特色生物资源研究与利用川渝共建重点实验室,南川408435

出　　处：《天然产物研究与开发》2025年第3期481-490,共10页Natural Product Research and Development

基　　金：国家重点研发计划(2021YFD1601005);重庆市科技局绩效激励项目(2022JX010);重庆市科技局基本科研业务费项目(2024jbky-015)。

摘　　要：从鹿血中筛选出具有抗运动疲劳活性的肽段。以冻干鹿血粉为原料,选用碱性蛋白酶制备抗运动疲劳多肽,采用透析和葡聚糖凝胶Sephadex G-25色谱分离纯化,以对小鼠负重力竭游泳时间,肝糖原、血乳酸及血尿氮含量的影响为指标,获得抗运动疲劳活性较强的肽段。采用氨基酸自动分析仪检测游离氨基酸与水解氨基酸含量;高效液相色谱法检测γ-氨基丁酸(γ-aminobutyric acid,GABA)与牛磺酸含量;凝胶渗透色谱法检测肽段分子量及其分布;液相色谱-串联质谱法鉴定多肽序列。透析分离得到分子量为500~3500 Da(M-1)、3500~8000 Da(M-2)和大于8000 Da(M-3)的三个组分,葡聚糖凝胶Sephadex G-25色谱进一步分离M-1得到P-1、P-2两个组分。与生理盐水给药相比,M-1给药在增加小鼠力竭游泳时间、肝糖原含量和降低血乳酸、血尿氮含量上差异均达到显著(P<0.01);P-2给药在增加小鼠力竭游泳时间和降低血乳酸、血尿氮含量上差异达到显著(P<0.05)。M-1中亮氨酸含量最高(12.14%),P-2中天冬氨酸含量最高(13.54%)。M-1的GABA、牛磺酸含量分别比P-2多178.39%、51.04%。M-1由小于20个氨基酸,分子量低于2000 Da的肽段组成。筛选出高活性的抗运动疲劳肽段M-1,可为鹿血天然抗运动疲劳产品开发提供理论依据。This study aims to screen anti-exercise fatigue peptides from enzymatic hydrolysates of Cervus nippon blood.The anti-exercise fatigue peptides were analyzed by swimming time(ST)under load,blood urea nitrogen(BUN)content,blood lactic acid(BLA)and hepatic glycogen(HG)content of the mice.Dialysis and Sephadex G-25 dextran gel chromatography were applied for isolation and purification.The contents of free amino acid and hydrolyzed amino acid were detected by automatic amino acid analyzer.The contents ofγ-aminobutyric acid(GABA)and taurine were determined by high performance liquid chromatography.Gel permeation chromatography was utilized for the molecular weight distribution and liquid chromatography-tandem mass spectrometry was utilized for the identification of peptides.Dialysis produced three fractions:molecular weights of 500-3500 Da(M-1),3500-8000 Da(M-2)and>8000 Da(M-3)and Sephadex G-25 gel chromatography produced two fractions from M-1:P-1 and P-2.Compared with the normal saline group,M-1 significantly(P<0.01)increased the ST under load,the HG content and decreased the BUN content,the BLA content in mice,P-2 could significantly(P<0.05)increase the ST under load and decrease the BUN content,the BLA content in mice.The content of leucine from M-1 was the highest(12.14%).The content of aspartate from P-2 was the highest(13.54%).Compared with P-2,the content of GABA and taurine from M-1 were increased by 178.39%and 51.04%respectively.M-1 composed of peptides with amino acids less than 20 and molecular weight less than 2000 Da.M-1 were screened from the enzymatic hydrolysates of Cervus nippon blood.The results of this study could provide a theoretical basis for the development of anti-exercise fatigue products from Cervus nippon blood.

关 键 词：鹿血 抗运动疲劳肽 分离纯化 氨基酸 鉴定 

分 类 号：S879.9[农业科学—畜牧兽医]