黄刺多糖工艺优化及其对过氧化氢损伤的胰岛β细胞的保护作用  

Optimization of extracting Berberi dasystachya polysaccharides and its protection against H_(2)O_(2)-induced pancreaticβcell injury

作　　者：邓永蓉 韩丽娟 岳庆明 漆莹 马娜娜 赵玉欣 DENG Yongrong;HAN Lijuan;YUE Qingming;QI Ying;MA Nana;ZHAO Yuxin(College of Agriculture and Animal Husbandry,Qinghai University,Xining 810016,China;State Key Laboratory of Plateau Ecology and Agriculture,Qinghai University,Xining 810016,China)

机构地区：[1]青海大学农牧学院,青海西宁810016 [2]省部共建三江源生态与高原农业国家重点实验室(青海大学),青海西宁810016

出　　处：《食品与发酵工业》2023年第3期146-156,共11页Food and Fermentation Industries

基　　金：青海省科学技术厅自然科学基金青年项目(2020-ZJ-951Q);国家自然科学基金项目(31960088)。

摘　　要：为研究黄刺粗多糖(Berberi dasystachya polysaccharides,BDPs)对H_(2)O_(2)诱导的RIN-m5F细胞氧化损伤的保护作用,以黄刺浆果为原料,通过单因素试验和响应面Box-Behnken设计实验优化超声波辅助酶解提取黄刺果实多糖的工艺。采用高效体积排阻色谱、离子色谱、扫描电镜及刚果红实验对BDPs理化性质进行分析,以H_(2)O_(2)诱导RIN-m5F细胞建立氧化应激损伤模型,通过CCK-8法测定细胞存活率,DCFH-DA探针法检测细胞内活性氧(reactive oxygen species,ROS)水平、比色法测定细胞内超氧化物歧化酶(superoxide dismutase,SOD)、过氧化氢酶(catalase,CAT)活性和丙二醛(malondialdehyde,MDA)含量探讨BDPs对氧化损伤细胞的保护作用。结果表明,当加酶量为1.25%(质量分数),酶解时间57 min,酶解温度45℃,超声波功率164 W时,黄刺多糖提取得率为(3.478±0.075)%,与预测值3.451%相近。此外,BDPs的重均分子质量为10.2 kDa,主要由岩藻糖、鼠李糖、半乳糖、葡萄糖、木糖、甘露糖、半乳糖醛酸以及葡萄糖醛酸组成(摩尔比为1∶42.2∶32.6∶11.8∶4.4∶56.1∶5.1∶1.8)。刚果红实验结果表明,BDPs具有三螺旋结构。细胞实验结果表明,H_(2)O_(2)能够导致RIN-m5F细胞的氧化损伤,与模型组相比,经0.0625~0.5 mg/mL的BDPs干预后的细胞存活率增加(P<0.05),ROS、MDA水平呈下降趋势。BDPs能够提高细胞SOD、CAT活性,并且呈一定量效关系。综上所述,BDPs在一定剂量范围内可以提高细胞存活率,对H_(2)O_(2)诱导的氧化损伤具有保护作用,研究结果可为黄刺多糖功能研究提供参考。To investigate the protective effect of Berberi dasystachya polysaccharides(BDPs)against H_(2)O_(2)-induced oxidative stress in RIN-m5F cells,through a single-factor test and response Box-Behnken design test,the ultrasonic auxiliary enzyme extraction of BDPs was optimized using Berberi dasystachya Maxim berries as the raw material.The physicochemical properties were analyzed by high performance size exclusion chromatography(HPSEC),ion chromatography,scanning electron microscope,and Congo red.The oxidative stress injury model was established with H_(2)O_(2)-induced RIN-m5F cells,and the effect of different concentrations of BDPs on cell survival was determined by the CCK-8 method.The intracellular reactive oxygen species(ROS)level was detected by the DCFH-DA probe method and the activity of superoxide dismutase(SOD),catalase(CAT),and malondialdehyde(MDA)content in the cell culture supernatant was determined by colorimetric method.Results showed that the extraction rate of BDPs was(3.478±0.075)%,which was close to the predicted value of 3.451%when the addition amount of enzymes was 1.25%(mass fraction),the enzymatic time was 57 min,the enzymatic lysis temperature was 45℃,and the ultrasonic power was 164 W.RIN-m5F cells were susceptible to the oxidative damage caused by H_(2)O_(2),as the intervention of BDPs of 0.0625-0.5 mg/mL(P<0.05),the cell survival rate rose in comparison to the model group while the levels of ROS and MDA exhibited a declining trend.BDPs had been found to enhance the SOD and CAT activity of RIN-m5F cells.In conclusion,BDPs had been found to increase cell survival within a range of doses and had a protective effect against H_(2)O_(2)-induced oxidative damage,providing a reference for the development of BDPs function.

关 键 词：黄刺多糖 抗氧化 RIN-m5F细胞 氧化应激 

分 类 号：R285[医药卫生—中药学]