Box-Behnken响应面法优化葛根异黄酮提取纯化工艺及损伤心肌细胞治疗作用研究

Optimization of Extraction Process of Isoflavones from Radix Puerariae Via Box-Behnken Response Surface Methodology and Study on Therapeutic Effect of Injured Myocardial Cells

目的:采用Box-Behnken响应面优化葛根异黄酮最佳提取工艺,并研究葛根异黄酮对损伤心肌细胞的治疗作用。方法:在单因素实验的基础上,以料液比、提取时间、溶剂浓度为考察因素,以3'-羟基葛根素、葛根素、3'-甲氧基葛根素、大豆苷的含量为评价指标,采用Box-Behnken响应面法设计实验,以葛根异黄酮总提取率为考察指标,通过分析回归方程模型,优选最佳提取工艺并验证工艺;采用MTT法检测葛根异黄酮对损伤心肌细胞的治疗作用。结果:Box-Behnken响应面法优化得到葛根异黄酮最佳提取工艺为:料液比为30 mg·mL^(-1),提取时间为100 min,乙醇体积分数为73%,葛根异黄酮的总提取率平均值为41.6 mg·g^(-1),与预测值42.08 mg·g^(-1),相对误差0.81%;葛根异黄酮对H 2O_(2)损伤的缺氧复氧模型和Na 2S_(2)O 4损伤的缺血再灌注模型OD值分别为0.7436±0.0292和0.7457±0.0094,细胞存活率分别为81.32%和73.41%。结论:优选葛根抗氧化活性异黄酮最佳工艺稳定,预测模型效果良好,且葛根异黄酮对心肌缺血有良好的治疗作用。

Objective:Box-Behnken response surface methodology was accepted to optimize the extraction process of Pueraria Isoflavon,and the study on the therapeutic effect of pueraria isoflavones on injured myocardial cells.Methods:On the basis of single factor experiment,taking the ratio of material to liquid,extraction time and solvent concentration as the investigation factors,taking the contents of 3'-hydroxypuerarin,puerarin,3'-methoxy puerarin and daidzein as evaluation indexes,by analyzing the regression equation model,the best extraction process was optimized and verified;MTT assay was used to detect the therapepredicted value was 42.08 mg·g^(-1),which was 0.81%.The OD values of the hypoxia reoxygutic effect of pueraria isoflavones on injured myocardial cells.Results:The optimal extracti on parameters were determined as follows:the material liquid ratio was 30 mg·mL^(-1),the extraction time was 100 min,and the ethanol concentration was 73%,the relative error of average total extraction rate of isoflavones from Pueraria lobata was 41.6 mg·g^(-1),and the enation model of H 2O_(2) injury and the ischemia reperfusion model of Na 2S_(2)O 4 injury induced by pueraria isoflavones were 0.7436±0.0292 and 0.7457±0.0094,respectively,thel survival rates were 81.32%and 73.41%,respectively.Conclusion:The optimal process of isoflavones with antioxidant activity of Pueraria lobata is stable,the prediction model is effective,and Pueraria lobata isoflavones have a good therapeutic effect on myocardial ischemia.