摘要
利用等电点沉淀法提取鸭肝中的蛋白质,经碱性蛋白酶和风味蛋白酶同步水解,得到鸭肝蛋白多肽组分。水解物经凝胶渗透色谱测定分子质量分布后,采用凝胶过滤色谱与反相高效液相色谱结合的方法对潜在抗炎活性的肽逐步分离纯化。并结合液相色谱-串联质谱联用技术共鉴定筛选出10种分子质量低于1 300 Da的新肽。合成多肽在脂多糖诱导的RAW264.7细胞中验证其抗炎活性。其中,LVYPFPGPI和VIESPPEI剂量依赖性抑制炎症细胞中NO的释放,质量浓度为100μg/mL时,抑制率分别为48.24%、56.32%。而VIESPPEI在质量浓度为100μg/mL时,对TNF-α释放的抑制率达42.48%。抑制IL-6释放能力最强的是IDVSPDSPDHY,在质量浓度为25μg/mL时,抑制率达到27.04%。研究结果为鸭副产物高值化综合利用和炎症的安全治疗提供新思路。
In order to obtain peptide fractions,duck liver proteins were recovered by isoelectric precipitation,and then hydrolyzed simultaneously by alcalase and flavorzyme.The molecular mass distribution of the hydrolysate was determined by using a gel permeation chromatography(GPC)system followed by fractionation using a combination of gel filtration chromatography(GFC)and reversed phase-high performance liquid chromatography(RP-HPLC).Ten novel peptides with molecular mass less than 1300 Da were identified from the subfraction with high inhibitory effect on cytokine release from macrophages using liquid chromatography-tandem mass spectrometry(LC-MS/MS).The anti-inflammatory activity of the synthetic peptides was tested in lipopolysaccharide(LPS)-stimulated RAW264.7 macrophage cells.Peptides LVYPFPGPI and VIESPPEI inhibited the release of NO from inflammatory cells in a dose-dependent manner,with inhibition percentages of 48.24%and 56.32%at the concentration of 100μg/mL,respectively.VIESPPEI at the concentration of 100μg/mL inhibited TNF-αrelease by 42.48%.Notably,IDVSPDSPDHY showed the strongest ability to inhibit the release of IL-6,with an inhibition percentage of 27.04%at the concentration of 25μg/mL.This study may provide new ideas for the high-valued comprehensive utilization of duck by-products and their application in safe treatment of inflammation.
作者
张来弟
孙杨赢
周昌瑜
潘道东
何俊
ZHANG Laidi;SUN Yangying;ZHOU Changyu;PAN Daodong;HE Jun(Key Laboratory of Animal Products Food Deep Processing Technology of Zhejiang Province,College of Food and Pharmaceutical Sciences,Ningbo University,Ningbo 315800,China)
出处
《食品科学》
EI
CAS
CSCD
北大核心
2023年第12期150-156,共7页
Food Science
基金
浙江省重点研发计划项目(2019C02085)
国家水禽产业技术体系肉类加工岗位科学家基金项目(CARS-42-25)。
关键词
鸭肝蛋白
酶解
抗炎肽
RAW264.7巨噬细胞
duck liver proteins
enzymatic hydrolysis
anti-inflammatory peptides
RAW264.7 macrophages
