响应面法优化鹿筋蛋白提取工艺及体外抗类风湿性关节炎活性  

Optimization of Protein Extraction Process from Deer Sinew by Response Surface Methodology and Its Activity of Anti-rheumatoid Arthritis in Vitro

作　　者：郅慧 兰梦 李晶峰 张晔 杨小倩 张辉 李春楠 ZHI Hui;LAN Meng;LI Jing-feng;ZHANG Ye;YANG Xiao-qian;ZHANG Hui;LI Chun-nan(Jinlin Ginseng Academy,Changchun University of Chinese Medicine,Changchun 130177,China;Jilin Xinshui Science and Technology Development Co.,Ltd.,Changchun 130000,China)

机构地区：[1]长春中医药大学,吉林省人参科学研究院,吉林长春130117 [2]吉林鑫水科技开发有限公司,吉林长春130000

出　　处：《食品工业科技》2020年第22期178-184,共7页Science and Technology of Food Industry

基　　金：国家中医药行业专项(201507002)。

摘　　要：目的:优化鹿筋蛋白提取工艺,探究其对类风湿性关节炎成纤维样滑膜细胞(MH7A)增殖及炎症因子分泌的影响。方法:以料液比、提取温度、提取时间为自变量,鹿筋蛋白含量为因变量,进行单因素实验,结合Box-Behnken试验设计,获得鹿筋蛋白最佳提取工艺。采用60 ng/mL肿瘤坏死因子-α(TNF-α)体外诱导MH7A细胞,MTT法测定不同浓度的(25、50、100、200μg/mL)鹿筋蛋白对MH7A细胞增殖抑制活性的影响;酶联免疫法测定细胞上清液中NO(一氧化氮)、白细胞介素-6(IL-6)、TNF-α含量。结果:鹿筋蛋白最佳提取条件为料液比1∶21 g/mL、提取温度88℃、提取时间8.6 h,此条件下测得鹿筋蛋白含量为12.53%。体外活性实验结果表明,与模型组比较,不同浓度(25、50、100、200μg/mL)的鹿筋蛋白均可显著(P<0.05)抑制MH7A细胞的增殖和NO、IL-6、TNF-α的释放(P<0.05),模型组的NO、IL-6、TNF-α释放量分别为:4.07、21.95、16.31 pg/mL,在100μg/mL质量浓度下,鹿筋蛋白对上述三种炎症因子释放的抑制作用最显著(P<0.05),其释放量分别为:1.60、13.60、7.29 pg/mL。结论:鹿筋蛋白通过抑制TNF-α诱导的MH7A细胞增殖,同时减少炎症因子NO、IL-6、TNF-α的释放,发挥其抗类风湿性关节炎的作用。本研究为研究鹿筋蛋白的制备工艺提供参考,并证明鹿筋蛋白具有较好的体外抗类风湿性关节炎活性。Objective:To optimize the extraction processing of deer sinew,and study its effects on the proliferation of MH7A cells and the secretion of inflammatory factors.Methods:Single factor extraction was carried out with solid-liquid ratio,extraction temperature,time as independent variables and deer sinew contents as dependent variables.And Box-Behnken experiment design was used to obtain the optimum extraction processing of deer sinew.MH7A cells were induced by 60 ng/mL tumor necrosis factor-α(TNF-α).MTT assay was used to determine the effects of different concentrations of deer sinew protein(25,50,100,200 ug/mL)on the proliferation of MH7A cells,and ELLSA was used to determine the contents of NO,IL-6 and TNF-αin cell supernatant.Results:The optimum extraction conditions of deer sinew protein were 88℃,8.6 h,1∶21 g/mL,and the protein content of deer sinew was 12.53%.In vitro anti-inflammatory experiments showed that different concentrations(25,50,100,200 ug/mL)of deer sinew could inhibit the proliferation of MH7A cells and the release of NO,IL-6 and TNF-α(P<0.05).The release of NO,IL-6 and TNF-αin the model group were 4.07,21.95,16.31 pg/mL,and deer sinew could inhibit the proliferation of MH7A cells at 100μg/mL,the release of inflammatory factors in the deer sinew group were 1.60,13.60,7.29 pg/mL,respectively.Conclusion:Deer sinew protein plays an anti-rheumatoid arthritis role by inhibiting the proliferation of MH7A cells induced by TNF-αand reducing the release of inflammatory factors NO,IL-6 and TNF-α.This study would lay a foundation for the extraction of deer sinew protein,provide a reference for further study of the preparation process of deer sinew protein,and prove that deer sinew protein have good anti rheumatoid joint activity.

关 键 词：鹿筋蛋白 响应面优化 抗类风湿性关节炎 MH7A 细胞 

分 类 号：TS201.2[轻工技术与工程—食品科学]