青天葵甲醇提取物的体外抗氧化活性

In Vitro Antioxidant Activities of Nervilia Fordii Methanol Extracts

探讨了青天葵甲醇提取物的体外抗氧化活性。对二苯代苦味酰基自由基(1,1-diphenyl-2-picrylhydrazyl,DPPH)和羟自由基(hydroxylradicals,·OH)清除实验评价青天葵提取物体外抗氧化能力。过氧化氢(H_2O_2,150μmol/L)诱发Caco-2细胞建立损伤模型评估青天葵提取物的细胞保护效果,MTT法检测细p胞生存率。比色法测定超氧化物歧化酶(superoxide dismutase,SOD),过氧化氢酶(catalase,CAT)和谷胱甘肽过氧化酶(glutathione peroxidase,GSH-Px)活力。硫代巴比妥酸法(thiobarbituric acid reactive substance,TBARS)和2’,7’-二氢二氯荧光黄双乙酸钠(dichloro-dihydro-fluorescein diacetate,DCFH-DA)探针分别测定丙二醛(malondiadehycle,MDA)和活性氧(reactive oxygen species,ROS)水平。酶联法(EPnzyme linked immunosorbent assay,ELISA)测定细胞白介素(Interlukin,IL)-8和IL-10水平。青天葵提取物中含总黄酮(4.72±0.13 mg Rutin/g)和总多酚(4.25±0.46 mg GAE/g),有较好的体外抗氧化能力。青天葵提取物能明显提升细胞生存率及内源性抗氧化物酶活性。降低受损细胞内总ROS,MDA及IL-8水平,提高抗炎细胞因子IL-10的分泌量。结果提示,青天葵提取物能显著改善H_2O_2诱发的Caco-2细胞所氧化应激损伤。

The in vitro antioxidant activities of Nervilia fordii mthanol extracts（NFME） were investigated. Total polyphenols and flavonoids in NFME were determined by ultraviolet spectrophotometry. The antioxidant capability of NFME was evaluated according to the 1,1-diphenyl-2-picrylhydrazyl（DPPH） and hydroxyl radicals（·OH） scavenging assay, respectively. Protective effect of NFME was investigated using a model of hydrogen peroxide（H_2 O_2, 150 μmol/L） induced oxidative stress in intestinal epithelial Caco-2 cells. Cell viability was determined by MTT assay. The cellular levels of superoxide dismutase（SOD）, catalase（CAT） and glutathione peroxidase（GSH-Px） were determined by commercial assay kits according the manuscripts. The cellular levels of malondiadehycle（MDA） and reactive oxygen species（ROS） were determined by thiobarbituric acid reactive substance（TBARS） and dichloro-dihydro-fluorescein diacetate（DCFH-DA） assay, respectively. The levels of interlukin-8（IL-8） and IL-10 were determined used an enzyme linked immunosorbent assay（ELISA）. The NFME were enriched in total flavonoids（4.72 ± 0.13 mg Rutin/g） and poly phenols（4.25 ± 0.46 mg GAE/g）. NFME also exhibited a great DPPH and ·OH radical scavenging activity. NFME was able to increase the cell viability, and significantly increased the activity of SOD, CAT and GSH-Px in oxidative damaged Caco-2 cells. In addition, NFME treatment was also effectively decreased the generation of ROS, MDA and IL-8 secretion in H_2 O_2 treated Caco-2 cells. In contrast, NFME increased the IL-10 secretion in damaged Caco-2 cells. These results suggested that the NFME exhibits a great in vitro antioxidant activity and can effectively attenuate the H_2 O_2-induced oxidative stress in Caco-2 cells.