DPPH自由基清除活性的光度微量滴定模型及应用  

Photometric Micro-Titration Model of DPPH Radicals Scavenging Activity and Its Application

作　　者：高云涛[1,2] 魏薇[1,2] 叶丽清[1,2] 李晓芬[1,2] 刘萍[1,2] 张宏教[1,2] 杨露[1,2] 虞姣姣 茶家伟 

机构地区：[1]云南民族大学化学与生物技术学院,云南昆明650500 [2]云南民族大学民族药资源化学国家民委一教育部重点实验室,云南昆明650500

出　　处：《光谱学与光谱分析》2015年第2期492-496,共5页Spectroscopy and Spectral Analysis

基　　金：国家自然科学基金项目(21367025);云南省创新团队项目(2011HC008);云南省教育厅创新团队项目(2010UY08);国家民委教育部民族药资源化学重点实验室开放基金项目(MZY1201)资助

摘　　要：传统的DPPH自由基清除活性评价方法以半数清除浓度EC50为评价指标,但EC50随DPPH初始加入量增加而增加,随分析体积增加而减小,因此,不同条件EC50值不具有可比性。提出以DPPH与抗氧化剂相互反应的化学计量数比（R）作为评价DPPH清除活性的指标,该指标只与DPPH与抗氧化剂相互反应的化学计量关系有关,与DPPH初始加入量和分析体积等因素无关,解决了EC50可比性差的问题。提出了测定化学计量数比（R）的光度微量滴定法,建立了利用滴定过程吸光度差（ΔA）与抗氧化剂加入量之间的滴定方程计算R值、以R计算EC50的光度微量滴定模型,并利用芦丁对模型进行验证。结果：芦丁与DPPH反应R值在1.817~1.846之间,当DPPH加入量为1.12×10-7,2.24×10-7,4.48×10-7和6.72×10-7mol时,分别计算得EC50值分别为1.196×10-3,2.392×10-3,4.819×10-3和7.292×10-3 mg·mL-1。在此基础上,基于文献报道的芦丁清除DPPH条件,利用得到的芦丁R值计算出相应EC50,结果与文献报道EC50值相当。方法可比性好,样品消耗量明显降低,简单、成本低,结果可靠,为自由基清除活性评价提出了一种新的思路。In the present paper, the stoichiometric ratio （R） for the interreaction of DPPH radicals with the antoxidant was em- ployed as a evaluation index for DPPH radicals scavenging activity of antioxidants. This evaluation index was related only with the stoichiometric relationship between DPPH radicals and the antioxidant, not the relationship with the initial DPPH amount and the volume of sample, which could offer a solution for the problem of poor comparability of EC50 under different conditions. A novel photometric micro-titration method was proposed for the determination of the stoichiometric ratio （R） for the interreac- tion of DPPH radicals with the antoxidant. The titration equation was established based on the absorbance difference （AA） of DPPH radicals in the titration process and the added amount of antoxidant. The stoichiometric ratio （R） for the reaction of DP- PH radicals with the addition amount of antoxidant was determined by the titration equation obtained, while, the DPPH median elimination concentration （EC50） of antoxidant can be calculated by the stoichiometric ratio （R）. The above photometric micro-ti- tration model was verified using rutin as DPPH radicals scavenger. As experiment results, the stoichiometric ratio （R） of DPPH radicals to rutin was determined to be in the range of 1. 817 ~ 1. 846. The calculated value of EC50 was 1. 196 × 10-3 , 2. 392 × 10-3 , 4. 819×10-3 and 7. 292×10-3 mg ·mL-1 for 1.12×10 7, 2.24×10-7 , 4. 48×10-7 and 6.72×10-7 mol of the addition amount of DPPH radicals, respectively. The proposed method has better precision and reliability with smaller amount of sample than conventional method. While, the obtained stoichiometric ratio value （R） of rutin was employed to calculate the rutin median elimination concentration for DPPH （EC50） according to the conditions as reported in the literatures, and the calculated results were consistent with that reported in the literatures.

关 键 词：DPPH自由基 光度微量滴定 滴定曲线 化学计量数比 芦丁 

分 类 号：O657.3[理学—分析化学]