野生蓝莓酚酸成分鉴定及其清除细胞内自由基活性研究

Identification of Phenolic Acids in Wild Blueberries and Their Scavenging Intracellular ROS Activity

酚酸具有良好的自由基清除活性,能有效降低非酒精性脂肪肝、心血管疾病、糖尿病等疾病发生风险。酚酸是大兴安岭野生蓝莓的主要酚类物质之一,其成分尚不清楚。本文采用柱层析法纯化蓝莓中的酚酸组分,用高效液相色谱法进行成分鉴定与含量检测。以AAPH诱导Hep G-2细胞氧化损伤模型评价酚酸清除细胞内自由基活性。从物质结构特性出发,通过循环伏安法分析酚酸的氧化-还原能力,探讨酚酸清除细胞内自由基活性机理。结果表明:蓝莓酚酸主要由咖啡酸、绿原酸、肉桂酸、阿魏酸、没食子酸、对香豆酸和原儿茶酸7种酚酸组成,含量较多的为绿原酸(21.2%)、阿魏酸(8.3%)、对香豆酸(7.0%)和咖啡酸(4.6%)。4种主要酚酸对细胞内自由基清除能力为咖啡酸>绿原酸>阿魏酸>对香豆酸,其失电子能力依次为绿原酸>咖啡酸>阿魏酸>对香豆酸。酚酸是蓝莓的主要抗氧化成分之一,其失电子能力是影响酚酸清除细胞内自由基活性的主要原因。

More and more studies suggest that phenolic acids exhibit high bioactivity in scavenging reactive oxygen species （ROS） and can effectively reduce risks in some diseases, such as nonalcoholic fatty liver disease, cardiovascular disease and diabetes. Phenolic acids were the main polyphenolic compounds in wild blueberries from the Greater Hinggan Mountains. However, the specific components were still unknown. In this work, column chromatography was employed to purify phenolic acid compounds from fresh blueberries; high performance liquid chromatography （HPLC） was used to i- dentify and quantitative analysis the phenolic acids; based on the in vitro model of 2, 2＇-azobis （2-amidinopropane） dihydrochloride （AAPH）-induced oxidative damage of HepG-2 cells, the scavenging intracellular ROS activity of phenolic acids from blueberries were subsequently studied; in further, the mechanisms of phenolic acids scavenging intracellular ROS were also discussed according to their reducing capacity revealed by cyclic voltammograms assay. The results indi- cated that 7 phenolic acids were identified in wild blueberries： caffeic acid, chlorogenic acid, cinnamic acid, ferulic acid, gallic acid, p-coumaric acid, and protocatechuic acid. And the main phenolic acids were chlorogenic acid （21.2%）, ferulic acid （8.3%）, p-coumaric acid （7.0%）, and caffeic acid （4.6%）. On the aspect of scavenging intracellular ROS activity, the maximal clearance rate of ROS for the above 4 phenolic acids were 58.4%, 43.6%, 38.2%, 28.9%, respectively, and their scavenging intracellular ROS capacity was caffeic acid〉 chlorogenic acid 〉 ferulic acid 〉 p-coumaric acid. The chemical assay results show that the reducing capacity of the above 4 phenolic acids was chlorogenic acid 〉 caffeic acid 〉 ferulic acid 〉 p-coumaric acid. According to the above described results, phenolic acids should be one kind of the main anti-oxidative compounds in wild blueberries and their scavenging intracellular ROS activity was closely related with their reducing capacity.