不同聚合度葡萄籽原花青素对三种消化酶抑制作用机制
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摘要
目的利用多种光谱及色谱技术研究不同聚合度葡萄籽原花青素与α-淀粉酶、脂肪酶、胃蛋白酶的作用机制,探讨聚合度与抑制作用的关系。方法采用紫外分光光度法和荧光法测定经不同聚合度原花青素对三种消化酶的体外半数抑制率;采用动态光散射法测定原花青素与消化酶聚集体的粒度;应用圆二色谱法测定原花青素作用后三种消化酶蛋白二级结构变化;利用AutoDock Vina软件进行原花青素与消化酶作用的分子模拟对接。结果体外酶活实验证明原花青素抑制作用随着聚合度的增加而增强;随着原花青素聚合度的增加,溶液中平均粒径增大,多分散指数(polymer dispersity index,PDI)降低;原花青素改变了三种消化酶的二级结构中α-螺旋、β-折叠等结构单元的含量,使其发生相互转化,且聚合度越大,转化量越大,结构破坏越严重;分子对接评分数值均为负数,表明原花青素与酶产生亲和力,且聚合度越大与消化酶中的关键氨基酸形成的氢键越多、疏水作用越强。结论首次研究并描述了不同聚合度原花青素与三种消化酶的作用机制,证明原花青素的聚合度越大对消化酶的抑制作用越强,对原花青素保健品的开发应用具有重要意义。
Objective To evaluate the interaction of grape seed procyanidins with different degrees of polymerization on α-amylase,lipase and pepsin,a variety of spectra and chromatographic methods were used.The relationship between degree of polymerization and inhibition was explored.Methods The particle size was measured by dynamic light scattering method;The changes of secondary structure of three kinds of digestive enzymes were determined by circular dichroism;Molecular docking was performed using AutoDockVina software.Results With the increase of the degree of polymerization of procyanidins,the diameter of digestive enzymes increased and the polydispersity index(PDI)decreased.The data of circular dichroism spectroscopy showed that procyanidins changed the proportion of α-helices and β-sheets in the secondary structure of three kinds of digestive enzymes.The higher the degree of polymerization was,the more contents of α-helices and β-sheets were changed and the more serious the secondary structures of the enzymes were destroyed.The docking scores were all negative,indicating that procyanidins had an affinity with the enzymes.The higher the degree of polymerization was,the more hydrogen bonds were formed with the amino acids in the digestive enzymes and the stronger the hydrophobic effect was.Conclusion In this paper,the interaction between procyanidins with different degrees of polymerization and three kinds of digestive enzymes is firstly studied.It is speculated that the greater the degree of polymerization of procyanidins is,the stronger the inhibitory effect on digestive enzymes is,which is of great significance in promoting the development of health product industry.
Objective To evaluate the interaction of grape seed procyanidins with different degrees of polymerization on α-amylase,lipase and pepsin,a variety of spectra and chromatographic methods were used.The relationship between degree of polymerization and inhibition was explored.Methods The particle size was measured by dynamic light scattering method;The changes of secondary structure of three kinds of digestive enzymes were determined by circular dichroism;Molecular docking was performed using AutoDockVina software.Results With the increase of the degree of polymerization of procyanidins,the diameter of digestive enzymes increased and the polydispersity index(PDI)decreased.The data of circular dichroism spectroscopy showed that procyanidins changed the proportion of α-helices and β-sheets in the secondary structure of three kinds of digestive enzymes.The higher the degree of polymerization was,the more contents of α-helices and β-sheets were changed and the more serious the secondary structures of the enzymes were destroyed.The docking scores were all negative,indicating that procyanidins had an affinity with the enzymes.The higher the degree of polymerization was,the more hydrogen bonds were formed with the amino acids in the digestive enzymes and the stronger the hydrophobic effect was.Conclusion In this paper,the interaction between procyanidins with different degrees of polymerization and three kinds of digestive enzymes is firstly studied.It is speculated that the greater the degree of polymerization of procyanidins is,the stronger the inhibitory effect on digestive enzymes is,which is of great significance in promoting the development of health product industry.
引文
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[23] LIU Y,CHEN D,YAN Y,et al.Biodiesel synthesis and conformation of lipase from Burkholderia cepacia in room temperature ionic liquids and organic solvents[J].Bioresour Technol,2011,102(22):10414-10418.
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[25] JAMIL A,ERIKA F,GABOR L,et al.From carbohydrates to drug-like fragments:Rational development of novel alpha-amylase inhibitors[J].Bioorg Med Chem,2015,23(20):6725-6732.
[2] BASHIR N,MANOHARAN V,MILTONPRABU S.Grape seed proanthocyanidins protects against cadmium induced oxidative pancreatitis in rats by attenuating oxidative stress,inflammation and apoptosis via Nrf-2/HO-1 signaling[J].J Nutr Biochem,2016,32:128-141.
[3] 高璐,王滢,饶胜其,等.葡萄籽原花青素提取物对衰老模型小鼠抗氧化作用[J].食品科学,2014,35(23):253-256.
[4] 张先杰,郭荣年,张妍,等.葡萄籽原花青素对肾血管性高血压大鼠血压和TNF-α水平的影响[J].中药药理与临床,2013,29(4):70-73.
[5] KATIATIYAR S K.Dietary proanthocyanidins inhibit UV radiation-induced skin tumor development through functional activation of the immune system[J].Mol Nutr Food Res,2016,60(6):1374-1382.
[6] NUNES M A,PIMENTEL F,COSTA A S,et al.Cardioprotective properties of grape seed proanthocyanidins:An update[J].Trends in Food Science & Technology,2016,57:31-39.
[7] WEBER H A,HODGES A E,GUTHRIE J R,et al.Comparison of proanthocyanidins in commercial antioxidants:Grape seed and pine bark extracts[J].Journal of Agricultural and Food Chemistry,2007,55(1):148-156.
[8] LUO L,CUI Y,ZHANG S,et al.Preparative separation of grape skin polyphenols by high-speed counter-current chromatography[J].Food Chem,2016,212:712-721.
[9] GU Y,HURST W J,STUART D A.Stuart Inhibition of key digestive enzymes by cocoa extracts and procyanidins[J].J Agric Food Chem,2011,59(10):5305-5311.
[10] HE Q,LV Y,YAO K.Effects of tea polyphenols on the activities of α-amylase,pepsin,trypsin and lipase[J].Food Chemistry,2007,101(3):1178-1182.
[11] LI Q,CHEN J,LI T,et al.Separation and characterization of polyphenolics from underutilized byproducts of fruit production(Choerospondias axillaris peels):inhibitory activity of proanthocyanidins against glycolysis enzymes[J].Food Funct,2015,6(12):3693-3701.
[12] SUI X,ZHANG Y,ZHOU W.In vitro and in silico studies of the inhibition activity of anthocyanins against porcine pancreatic α-amylase[J].Journal of Functional Foods,2016,21:50-57.
[13] SALVADO M J,CASANOVA E,FERNANDEZ A,et al.Roles of proanthocyanidin rich extracts in obesity[J].Food Funct,2015,6(4):1053-1071.
[14] STRUGALA V,KENNINGTON E J,CAMPBELL R J,et al.Inhibition of pepsin activity by alginates in vitro and the effect of epimerization[J].Int J Pharm,2005,304(1/2):40-50.
[15] ZHANG S,CUI Y,LI L,et al.Preparative HSCCC isolation of phloroglucinolysis products from grape seed polymeric proanthocyanidins as new powerful antioxidants[J].Food Chem,2015,188:422-429.
[16] 崔明超,陈少军,崔文,等.基于反向分子对接技术的异鼠李素靶标的预测[J].中国药房,2016,27(28):3921-3924.
[17] TROTT O,OLSON A J.AutoDock Vina:improving the speed and accuracy of docking with a new scoring function,efficient optimization,and multithreading[J].Journal of Computational Chemistry,2010,31(2):455-461.
[18] DILIP A,LE?NIK S,?TULAR T,et al.Ligand-based virtual screening interface between PyMOL and LiSiCA[J].Cheminform,2016,8(1):46.
[19] KUSANO R,OGAWA S,MATSUO Y,et al.α-Amylase and lipase inhibitory activity and structural characterization of acacia bark proanthocyanidins[J].Journal of Natural Products,2010,74(2):119-128.
[20] SUGIYAMA H,AKAZOME Y,SHOJI T,et al.Oligomeric procyanidins in apple polyphenol are main active components for inhibition of pancreatic lipase and triglyceride absorption[J].Journal of Agricultural and Food Chemistry,2007,55(11):4604-4609.
[21] HE,Q,LV Y,YAO K.Effects of tea polyphenols on the activities of α-amylase,pepsin,trypsin and lipase[J].Food Chemistry,2007,101(3):1178-1182.
[22] 朱书法,汤俊明,马晓明,等.碳酸钙结晶对胃蛋白酶二级结构的影响[J].光谱学与光谱分析,2003,23(3):477-480.
[23] LIU Y,CHEN D,YAN Y,et al.Biodiesel synthesis and conformation of lipase from Burkholderia cepacia in room temperature ionic liquids and organic solvents[J].Bioresour Technol,2011,102(22):10414-10418.
[24] SINGH S,BAJPAI U,LYNN A M.Structure based virtual screening to identify inhibitors against MurE Enzyme of Mycobacterium tuberculosis using AutoDock Vina[J].Bioinformation,2014,10(11):697-792.
[25] JAMIL A,ERIKA F,GABOR L,et al.From carbohydrates to drug-like fragments:Rational development of novel alpha-amylase inhibitors[J].Bioorg Med Chem,2015,23(20):6725-6732.