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目的优选出半夏总蛋白提取的最适方法并研究不同生长时期总蛋白量及成分的变化,为半夏药用成分的监控及合理采收提供科学依据。方法采用丙酮沉淀法、95%(NH4)2SO4盐析法、TCA-丙酮法和Tris-饱和酚法提取半夏总蛋白,筛选半夏总蛋白提取的最佳方法。分别于半夏出芽期、全苗期、珠芽期、佛焰苞期及倒苗期采集样品,分为新鲜组和干燥处理组,提取样品总蛋白,进行SDS-PAGE电泳检测及Bradford浓度测定。结果丙酮沉淀法、95%(NH4)2SO4盐析法、TCA-丙酮法和Tris-饱和酚法提取的半夏总蛋白量分别为0.369、0.678、1.082、0.493 mg/mL;新鲜半夏5个不同生长时期总蛋白量占块茎的质量分数分别为出芽期4.53 mg/g、全苗期2.28 mg/g、珠芽期2.79 mg/g、佛焰苞期7.61 mg/g、倒苗期10.21 mg/g;干燥半夏5个不同生长时期总蛋白量占块茎的质量分数分别为出芽期31.85 mg/g、全苗期18.52 mg/g、珠芽期42.08 mg/g、佛焰苞期28.56 mg/g、倒苗期56.84 mg/g;新鲜半夏块茎中蛋白含有15条蛋白带,干燥处理半夏块茎中蛋白含有5条蛋白带。结论 TCA-丙酮法是半夏总蛋白提取的最佳方法,不同生长时期的半夏块茎中蛋白成分各异,倒苗期总蛋白量最高。
Objective To optimize the extraction method of total protein of Pinellia ternata and to study the changes of total protein and composition in different growth stages so as to provide a scientific basis for the monitoring and reasonable harvest of the medicinal components of Pinellia ternata. Methods The best method for extracting total protein from Pinellia ternata was selected by acetone precipitation method, 95% (NH4) 2SO4 salting-out method, TCA-acetone method and Tris-saturated phenol method. The samples were collected at the sprouting stage, the whole seedling stage, the budding stage, the spatheraceae stage and the inverted seedling stage respectively. The samples were divided into fresh group and dry treatment group. The total protein of the samples was extracted for SDS-PAGE and Bradford determination. Results The total protein of Pinellia ternata extracted by acetone precipitation method, 95% (NH4) 2SO4 salting-out method, TCA-acetone method and Tris-saturated phenol method were 0.369,0.678,1.082,0.493 mg / The content of total protein in tubers at different growth stages were 4.53 mg / g, 2.28 mg / g, 2.79 mg / g, 2.71 mg / g respectively in budding stage and 10.21 mg / g.The total protein content in 5 different growth stages of Pinellia ternate were 31.85 mg / g, 18.52 mg / g, 42.08 mg / g and 28.56 mg / g respectively in budding stage , And 56.84 mg / g in the inverted seedling stage. The protein contained 15 protein bands in tuber of fresh Pinellia ternata, and the protein contained 5 protein bands in the tuber of pinellia tuber. Conclusion TCA-acetone method is the best method for extracting total protein from Pinellia ternata. The protein content of tuber of Pinellia ternata differs in different growth stages, and the total amount of protein is the highest under inverted seedling stage.