沙棘清除DPPH自由基动力学特性研究及活性成分筛选  

Study on DPPH Kinetic Characteristics and Screening of Active Components of Frutus Hippophae

作　　者：李倩茹 李生茂[1,2] 黄婷 吴婷 LI Qianru;LI Shengmao;HUANG Ting;WU Ting(School of Pharmacy,North Sichuan Medical College,Nanchong 637000,China;Institute of Materia Medica,North Sichuan Medical College,Nanchong 637000,China)

机构地区：[1]川北医学院药学院,四川南充637000 [2]川北医学院药物研究所,四川南充637000

出　　处：《食品科技》2021年第1期225-231,共7页Food Science and Technology

基　　金：2017年四川省教育厅科研项目(17ZB0166);2018年四川省中医药管理局科研项目(2018JC017);2017年南充市市校合作科研项目(NSMC20170201);2018年四川省省级大学生创新创业训练计划项目(201810634012)。

摘　　要：为研究沙棘清除DPPH自由基的动力学规律,并初步明确其抗氧化活性成分,对样品不同初始浓度、反应温度和反应时间进行考察,研究了沙棘清除DPPH自由基的动力学,并在此基础上评价了其清除DPPH自由基的活性。采用DPPH-HPLC法,在单因素实验对DPPH自由基浓度、反应时间和反应温度等条件优化的基础上,通过对比分析沙棘与DPPH自由基反应前后相应色谱峰峰面积的变化(减小或消失),明确其清除DPPH自由基的活性特征色谱峰,再根据特征峰的保留时间,与对照品进行比对初步明确其抗氧化活性成分。沙棘清除DPPH自由基的动力学结果表明:样品初始浓度、反应温度和反应时间对沙棘清除DPPH自由基的速率均有影响,在实验条件范围内,沙棘与DPPH自由基反应的速率随初始浓度增大和反应温度增高而加快,随反应时间延长而减慢,反应的活化能Ea为52.12 kJ/moL,接近二级反应动力学方程,其清除DPPH自由基的IC50值为5.87×102 μg/mL(相当于生药量),具有较强的抗氧化活性;DPPH-HPLC研究结果表明:沙棘与DPPH自由基的最佳反应条件为:DPPH自由基浓度35.5 mmol/L、反应时间90 min以及反应温度40 ℃,此时沙棘样品中12个抗氧化活性成分的色谱峰完全消失或峰面积明显减少,其中初步确认9号和11号色谱峰分别是芦丁和槲皮素的色谱峰。该方法具有试剂用量少、操作简单方便等特点,能快速、灵敏、高通量筛选中药复杂体系中的抗氧化活性成分。该研究为初步明确沙棘清除DPPH自由基的动力学规律、抗氧化活性成分及与活性结合的质量控制提供了一定实验数据。In order to study the kinetics of DPPH· scavenging activity of Frutus Hippophae and preliminarily determine its antioxidant active components,the experiment investigated the effects of initial concentrations,reaction temperature and reaction time on scavenging DPPH· of Frutus Hippophae and established a DPPH-HPLC method to observe the changes of peak area before and after the reaction of Frutus Hippophae with DPPH· on the basis of optimizing the reaction conditions by single factor test.Finally,according to the retention time of the characteristic peak,the antioxidant active components was preliminarily determined by comparing with the reference substance.The results showed that the reaction rate between Frutus Hippophae and DPPH· was accelerated with the increase of initial concentration and reaction temperature,while slowed down with the extension of reaction time.The Ea values was 52.12 kJ/moL and the scavenging reaction was close to the second-order reaction kinetics equation.The IC50 of DPPH· scavenging activity was 0.587 mg/mL,which showed strong antioxidant activity.Moreover,the optimal experimental conditions for the reaction between Frutus Hippophae and DPPH· were as followed:35.5 mmol/L DPPH free radical,reaction time 90 min and reaction temperature 40 ℃.Under these conditions,HPLC peaks of 12 antioxidants in Frutus Hippophae extract decreased or disappeared,among which the No.9 and 11 were preliminarily confirmed as Rutin and Quercetin,respectively.This method can quickly,sensitively and high-throughput screening of antioxidant active components from complex matrices,and the results provided some experimental data for the preliminarily determination of the kinetics of DPPH· scavenging activity,antioxidant active components and the quality control of Frutus Hippophae.

关 键 词：DPPH-HPLC法 沙棘 抗氧化 动力学 IC50 

分 类 号：TS201.4[轻工技术与工程—食品科学]