斑点免疫印迹法快速测定地龙炮制品中蚓激酶及其水解产物

Rapid determination of lumbrukinase and its hydrolysates in processed earthworm by dot blotting

目的测定参环毛蚓、秉氏环毛蚓、日本杜拉蚓和天锡杜拉蚓炮制品中蚓激酶及其水解产物,建立斑点免疫印迹法评价相应生药材药效成分明确的中药炮制品质量新方法。方法以兔抗蚓激酶单抗为一抗,应用斑点免疫印迹法对地龙炮制品中蚓激酶及其水解产物作定性检测,杂交信号应用Quantity One软件作定量分析。结果现行中药材参环毛蚓与山东产地方品种:秉氏环毛蚓、日本杜拉蚓、天锡杜拉蚓炮制品中蚓激酶及其水解产物的质量分数分别为125.23、118.34、81.05、72.13μg/g。结论秉氏环毛蚓炮制品中蚓激酶及其水解产物与现行地龙炮制品参环毛蚓无显著差异,理论上可作为广地龙的替代品。斑点免疫印迹法简便快捷,结果准确,无需大型设备,可用于地龙与类似中药材炮制品的品质评价。

Objective To determine the lumbrukinase contents and its hydrolysates in processed earthworm, set up a new evaluating method of dot blotting to evaluate the qualities of those processed Chinese medicines as earthworm. Methods Contents of lumbrukinase and its hydrolysates in processed earthworm were determined by dot blotting with rabbit antibody of antidumbrukinase as the first antibody. Blotting signal was analyzed by Quantity One software. Results Contents of lumbrukinase and its hydrolysates in processed earthworm of Pheretima aspergilum, P. pingi, Drawida japonica, and D. gisti were 125.23, 118.34, 81.05, and 72.13 μg/g, respectively. Conclusion There was no significant difference between the processed earthworms of P. aspergilum and P. pingi in the content of lumbrukinase and its hydrolysates, both of which were higher than those of the other two species in Shandong. Theoretically, the processed products of P. pingi could be used as the succedaneum of P. aspergilum in the clinical of traditional Chinese medicine. Dot blotting, as a convenient and accurate method, could be broadly used for the evaluation of the processed products of Chinese crude drugs similar to earthworm.