板蓝根蛋白的表征及响应曲面法优化其超声辅助提取工艺研究

目的?表征板蓝根蛋白的分子量组成，优化板蓝根蛋白超声提取工艺。方法?运用响应曲面法，以蛋白得率为评价指标，对提取时间、液料比及pH值3个影响因素进行考察，优化板蓝根蛋白的提取工艺，另以Design Expert软件对数据进行综合统计分析；采用聚丙烯酰胺凝胶电泳的方法表征板蓝根蛋白的分子量分布；应用扫描电镜技术观察超声提取前后板蓝根药材粉末的微观结构。结果?最优工艺条件是以pH值为7.8的50?mmol/L Tris-HCl为溶剂，液料比为80∶1，超声提取2次，每次提取65?min，按该工艺进行试验所得板蓝根蛋白得率为0.705%；板蓝根蛋白5个分子量段的蛋白分别为19.2、21.5、24.8、34.0~43.0?kDa 和>170?kDa。结论?运用响应曲面法优选的板蓝根蛋白提取工艺稳定可行，可用于板蓝根蛋白的提取。 

Characterization of Radix Isatidis Protein and its Ultrasound-assisted Extraction Process Optimization with Response Surface Methodology

OBJECTIVE?To characterize the molecular weight of Radix Isatidis protein (RIP) and optimize the extraction process of protein from Radix Isatidis. METHODS?Response surface methodology (RSM) was applied to optimize the extraction of protein from Radix Isatidis. Extraction time， liquid-to-solid ratio and pH value were set as the investigated factors with respect to the protein yield. In addition， Design Expert software was used for data analysis. The RIP was characterized for composition using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Scanning electron microscopy (SEM) analysis was performed to observe microstructure of the Radix Isatidis powder before and after ultrasound-assisted extraction (UAE). RESULTS?Based on the RSM analysis， optimum conditions were determined as follows: twice ultrasonic extraction in 50?mmol/L Tris-HCl buffer solution， pH at 7.8， liquid-to-solid ratio at 80∶1 and extraction for 65?min each time. Under the optimized conditions， the experimental values were 0.705%， which is in close agreement with values predicted by the model. The characterization of the RIP demonstrated that it contained five major groups of protein bands， namely bands of 19.2?kDa， 21.5?kDa， 24.8?kDa， 34~43?kDa and >170?kDa respectively. CONCLUSION?RSM can be applied for the optimization of extraction process of RIP， which is effective， stable and feasible.