郑鹏 王波 王前

摘 要：沙棘（Hippophae rhamnoides）是一種具有药用价值的植物，沙棘果油具有抗氧化、抗炎症及抗肿瘤等多种药理作用。为了探讨沙棘果油对H2O2造成氧化性损伤的细胞生长的影响及其抗氧化性，该研究选择H2O2对RAW264.7细胞氧化损伤模型，通过DPPH（1，1-二苯基-2-三硝基苯肼）自由基清除实验检测沙棘果油体外抗氧化能力，用[3-（4，5-二甲基噻唑-2）-2，5-二苯基四氮唑溴盐]MTT法和流式细胞仪检测超氧化物阴离子荧光探针（DHE）信号，分别检测不同浓度沙棘果油对H2O2损伤细胞的存活率和超氧化物阴离子水平。结果表明：（1）在DPPH自由基清除实验中，当沙棘果油浓度小于4.9%时，沙棘果油的抗氧化能力大于维生素C。（2）通过MTT法发现，浓度为3.125%的沙棘果油对H2O2损伤细胞的存活率显著升高（P<0.01）。（3）从DHE检测发现，在同一检测时间点，随着沙棘果油浓度增加，DHE阳性细胞比例显著下降（P<0.01）;在不同检测时间点，随着沙棘果油浓度增加，DHE阳性细胞比例显著升高（P<0.01）。沙棘果油对过氧化氢诱导的RAW264.7细胞氧化损伤模型有一定修复作用，可能与细胞内超氧化物阴离子水平受到抑制有关，它具有抗氧化性损伤的潜能。

关键词：沙棘果油，抗氧化性，MTT法，DHE，过氧化氢损伤模型

中图分类号：Q943.2

文献标识码：A

文章编号：1000-3142（2020）09-1349-08

Abstract：The sea buckthorn fruit （Hippophae rhamnoides） oil is medically claimed as having phytoantioxidant，antiinflammatory， and anticancerous properties in humans. To explore the hydrogen peroxide of the sea buckthorn fruit oil on mouse monocyte macrophages （RAW264.7 cells） oxidative damage model. By spectrophotometry，scavenging effects of sea buckthorn fruit oil on DPPH free radicals; In vitro antioxidant activity was assessed on RAW264.7 cells against hydrogen peroxide （H2O2） induced cytotoxicity. The antioxidant effect was determined by measuring the cell dye MTT[3-（4，5-dimethylthiazol-2-yl）-2，5-diphenyltetrazolium bromide] reducing assay. Then the survival rate of RAW264.7 cells was determined by MTT assay，and the expression of superoxide anion was determined by flow cytometry labeling with dihydroethidium（DHE）. The results were as follows：（1）Compared with the antioxidant capacity of vitamin C，when the concentration of sea buckthorn fruit oil was less than 4.9%，the antioxidant capacity of sea buckthorn fruit oil was higher than vitamin C. （2） Comparing to the H2O2 group，3.125% sea buckthorn fruit oil group had higher survival rate（P<0.01）. （3） Comparing to the H2O2 group，the proportion of DHE-positive cell was significantly decreased in the same time （P<0.01） and it was negative effect with concentration of sea buckthorn fruit oil，the proportion of DHE-positive cell was significantly increased at the same concentration（P<0.01），and which was related positive effect with concentration of sea buckthorn fruit oil （P<0.01）. These results support the protective effects of sea buckthorn fruit oil against hydrogen peroxide-induced toxicity in vitro with the underlying mechanisms of inhibiting oxidative stress. Taken together，these results clearly indicate that sea buckthorn fruit oil has significant potential as a natural antioxidant agent，which could be a potential source for the discovery of natrual antioxidant health product.

Key words：

sea buckthorn fruit oil，oxidation resistance，MTT，DHE，hydrogen peroxide damage model

人体在代谢过程中会产生活性氧自由基（ROS），包括超氧阴离子（O-2） 、羟自由基（·OH）、过氧化氢

（H2O2） 和脂质过氧化物的中间产物（Chauhan et al.，1997）。在正常的生理情况下，线粒体需要消耗约2% 的氧产生的ROS，由呼吸链底物端释放出电子还原氧分子单电子，产生ROS的前体

—

O-2和H2O2。在很多疾病以及衰老时，体内氧自由基会产生异常积累。过量的ROS会导致细胞损伤或细胞凋亡。ROS的活泼性和氧化反应能力都很强，能通过氧化作用攻击细胞的生物大分子物质（ 核酸、蛋白质、糖类、脂质） 。当人体处于亚健康状态时，自由基异常累积会引发疾病。近年来，天然抗氧化剂因其安全、高效、能满足亚健康人群日常保健需求的特点而受到广泛关注。……