黑豆黄嘌呤氧化酶抑制肽的制备及体外降尿酸活性  

Preparation of Xanthine Oxidase Inhibitory Peptide from Black Bean Protein by Enzymatic Hydrolysis and Its Uric Acid-Lowering Activity in Vitro

作　　者：孙菁茹 孙铭爽 吕文庆 曹荣安[1] 刁静静 王长远[1,3] SUN Jingru;SUN Mingshuang;LÜWenqing;CAO Rong’an;DIAO Jingjing;WANG Changyuan(College of Food Science,Heilongjiang Bayi Agricultural University,Daqing 163319,China;Heilongjiang Ounengda Dairy Co.Ltd.,Harbin 150025,China;National Coarse Cereals Engineering Research Center,Daqing 163319,China;Heilongjiang Coarse Grain Processing and Quality Safety Engineering Technology Research Center,Daqing 163319,China)

机构地区：[1]黑龙江八一农垦大学食品学院,黑龙江大庆163319 [2]黑龙江欧能达乳业有限公司,黑龙江哈尔滨150025 [3]国家杂粮工程技术研究中心,黑龙江大庆163319 [4]黑龙江省杂粮加工及质量安全工程技术研究中心,黑龙江大庆163319

出　　处：《食品科学》2024年第23期72-80,共9页Food Science

基　　金：黑龙江省重点研发计划项目(2022ZX02B18);黑龙江八一农垦大学研究生创新基金项目(YJSCX2023-ZX19)。

摘　　要：以黑豆蛋白为原料,以黄嘌呤氧化酶(xanthine oxidase,XOD)抑制率和水解度为指标筛选最佳蛋白酶和酶解工艺参数,在此基础上采用膜分离技术对黑豆蛋白酶解产物进行分离,获得不同分级的酶解产物,根据XOD抑制活性确定黑豆蛋白酶解产物的分子质量范围,并对其氨基酸组成、分子质量分布、肽序列以及活性稳定性进行分析。结果表明,黑豆XOD抑制肽的最佳酶解工艺为:碱性蛋白酶加酶量1.5%、酶解时间4 h、酶解温度50℃、pH 9.0、底物质量分数3%,此条件下黑豆蛋白酶解物的XOD抑制率和水解度分别为73.61%和21.29%。超滤产物中F3组分的XOD抑制活性最高(半抑制浓度为8.76 mg/mL),分子质量≤1500 Da,该组分的疏水性氨基酸和碱性氨基酸占氨基酸总量的56.66%和20.16%;该组分在高温热处理、胃、胰蛋白酶消化作用下均具有良好的稳定性。液相色谱-串联质谱法鉴定出F3组分中有18条肽段,平均分子质量主要集中在500~1400 Da。该系列肽段N-端和C-端疏水性氨基酸分别占44.86%和33.14%,碱性氨基酸分别占33.57%和39.29%。该研究结果可为黑豆蛋白的高值化利用提供一定的基础理论依据。In this study,black soybean protein was enzymatically hydrolyzed to prepare xanthine oxidase(XOD)inhibitory peptide.Based on XOD inhibitory activity and degree of hydrolysis(DH),various proteases were screened,and the hydrolysis parameters were optimized.Furthermore,the protein hydrolysate was fractionated by membrane separation into several fractions.The molecular mass ranges of the resulting fractions were determined based on XOD inhibitory activity,and their amino acid composition,molecular mass distribution,peptide sequences,activity and stability were analyzed.The results showed that the optimum enzymatic hydrolysis conditions were as follows:alkaline protease dosage of 1.5%,hydrolysis time of 4 h,temperature of 50℃,pH 9.0,and substrate concentration of 3%.Under the optimized conditions,the XOD inhibition rate and DH of the protein hydrolysate were 73.61%and 21.29%,respectively.The XOD inhibitory activity of F3,obtained from ultrafiltration of the hydrolysate,was the highest(with a half-maximal inhibitory concentration(IC50)of 8.76 mg/mL),with the molecular mass≤1500 Da.In F3,hydrophobic and basic amino acids accounted for 56.66%and 20.16%of the total amino acids,respectively.This fraction had good stability under high-temperature treatment,simulated gastric and trypsin digestion.Liquid chromatography-tandem mass spectrometry(LC-MS/MS)identified 18 peptides in F3,with an average molecular mass of approximately 500–1400 Da.In these peptides,hydrophobic amino acids at the N-terminus and C-terminus accounted for 44.86%and 33.14%,respectively,and basic amino acids accounted for 33.57%and 39.29%of the total amino acid residues,respectively.The results of this study provide a theoretical basis for the high value utilization of black bean protein.

关 键 词：黑豆蛋白 酶解工艺 黄嘌呤氧化酶抑制肽 氨基酸组成 肽序列 

分 类 号：TS214.9[轻工技术与工程—粮食、油脂及植物蛋白工程]