摘要
目的:探究高核苷酸酵母水解物的体外抗氧化活性。方法:采用化学法测定高核苷酸酵母水解物对二苯代苦味酰基自由基(DPPH)和羟自由基的清除能力及总还原力。建立游离脂肪酸(FFA)诱导HepG2细胞氧化应激模型,采用噻唑蓝溴化四唑(MTT)法检测高核苷酸酵母水解物对HepG2细胞的活力影响,荧光探针DCFH-DA法检测HepG2细胞内活性氧的变化,商业试剂盒检测细胞内氧化损伤标志物丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-PX)的活力。实时定量PCR检测keap1,Nrf2,HO-1和NQO-1的mRNA表达水平。免疫印迹检测Nrf2活化与转位情况。结果:5~50 mg/mL高核苷酸酵母水解物以质量浓度依赖性方式清除体外活性氧自由基,具有显著抗氧化能力。500~1000μg/mL高核苷酸酵母水解物可显著降低细胞内丙二醛(MDA)的含量,降低活性氧自由基(ROS)水平,提高细胞中超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-PX)的活性。抗氧化酶基因NQO1,Nrf2和HO-1的mRNA表达水平显著增加,keap1 mRNA表达水平显著降低。高核苷酸酵母水解物能够显著增加HepG2细胞核内Nrf2的蛋白质含量,降低其胞浆中的蛋白质含量,进一步阐明高核苷酸酵母水解物通过激活Nrf2通路减轻FFA诱导的HepG2氧化应激损伤作用。
Purpose:To explore the in vitro antioxidant activity of high-nucleotide yeast hydrolysate.Methodology:The scavenging activity of diphenyl bitter acyl radical(DPPH),hydroxyl free radical as well as the total reducing power were measured by chemical method.The oxidative stress model of the HepG2 cell induced by free fatty acid(FFA)was established.The effects of high nucleotide yeast hydrolysate on HepG2 cells were detected by thiazolyl blue tetrazolium bromide(MTT).And the changes of reactive oxygen in the HepG2 cells were measured by fluorescent probe method.Commercial kits were used to detected the activities of intracellular oxidative damage markers such as malondialdehyde(MDA),superoxide dismutase(SOD)and glutathione peroxidase(GSH-PX).Real-time quantitative PCR was used to detect mRNA expression levels of keap1,Nrf2,HO-1 and NQO-1.The excitation and transposition of Nrf2 was dected by western-blot.Conclusion:5~50 mg/mL high nucleotide yeast hydrolysate could scavenge oxygen free radicals in vitro in amass concentration-independt manner and had significant antioxidant capacity.500-1000μg/mL high nucleotide yeast hydrolysate could significantly decrease the content of malonaldehyde(MDA)in the cells as well as the level of reactive oxygen species(ROS);additionally and increase the activities of superoxide dismutase(SOD)and glutathione peroxidase(GSH-PX).The expression mRNA levels of the antioxidant enzyme genes NQO1,Nrf2 and HO-1 were also significantly increased,and the expression mRNA level of keap1 was significantly decreased.High nucleotide yeast hydrolysate could increase the protein content in the endonuclear of HepG2 and decrease the protein content of Nrf2 in the endochylema,and this results could futher clarified that high nucleotide yeast hydrolysate could reduce FFA-induced HepG2 oxidation by activating the Nrf2 pathway injury effect.
作者
潘聪
张大力
段盛林
苑鹏
夏凯
李占东
于伟厚
刘美宏
于寒松
赵可心
Pan Cong;Zhang Dali;Duan Shenglin;Yuan Peng;Xia Kai;Li Zhandong;Yu Weihou;Liu Meihong;Yu Hansong;Zhao Kexin(School of Food Science and Engineering,Jilin Agricultural University,Changchun 130118;China National Research Institute of Food and Fermentation Industries,Beijing 100015;School of Food Engineering,Jilin Engineering Normal University,Changchun 130052;Dalian Shuang Di Technology,Dalian 116635,Liaoning)
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2020年第9期10-18,共9页
Journal of Chinese Institute Of Food Science and Technology
