发酵桑叶糖蛋白提取工艺优化及活性研究

Study of optimization and activity of extraction process of fermented mulberry leaf glycoprotein

目的筛选发酵桑叶糖蛋白(FMLGP)的最佳提取工艺并评价其活性。方法以蛋白浓度为指标,采用单因素结合响应面法优化FMLGP提取工艺;通过琼脂糖纤维蛋白平板法检测纤溶活性;利用ox-LDL诱导巨噬细胞RAW264.7泡沫化,实验分为空白组、模型组和FMLGP高、中、低剂量组(终浓度分别为15、10、5 mg/ml),细胞培养24 h,采用油红染色法考察各组细胞内脂质蓄积程度。结果FMLGP最佳提取条件为PBS缓冲液、料液比1∶6 g/ml、于25℃提取4 h,蛋白浓度为(57.92±1.14)mg/ml,纤溶活性为(1780.66±76.49)IU/ml,15 mg/ml FMLGP对巨噬泡沫细胞有明显促进脂质排出作用。结论本研究确定了FMLGP的最佳提取工艺,通过纤溶活性及巨噬细胞泡沫化实验提示FMLGP具有促进脂质排出作用。

Objective To determine the optimal extraction process of fermented mulberry leaf glycoprotein(FMLGP),and to evaluate the activity of glycoprotein.Methods Taking protein concentration as index,the extraction process of FMLGP was optimized by single factor combined with response surface method.Fibrinolytic activity was detected by agarose fibrin plate method.Macrophage RAW264.7 foam was induced by ox-LDL.The experiment was divided into blank group,model group,and FMLGP high-dose,medium-dose,and low-dose groups(the final concentration was 15,10,and 5 mg/ml,respectively).The cells were cultured for 24 h and the degree of lipid accumulation in each group was investigated by Oil red staining.Results The optimal extraction conditions of FMLGP were PBS buffer solution,solid-liquid ratio of 1∶6 g/ml,extraction at 25℃for 4 h,protein concentration of(57.92±1.14)mg/ml,fibrinolytic activity of(1780.66±76.49)IU/ml,and 15 mg/ml FMLGP could significantly promote lipid excretion in macrophage foam cells.Conclusion In this study,the optimal extraction process of FMLGP is determined,and the fibrinolytic activity and macrophage foaming experiments shows that FMLGP had potential anti-atherosclerosis effects.