基于中药大分子探讨芍药甘草汤镇痛的物质基础及作用机制

Based on the macromolecules in traditional Chinese medicine exploring the material basis and mechanism of the analgesic effect of Shaoyao Gancao Decoction

为了研究芍药甘草汤的镇痛作用,本文从中药大分子角度探讨其物质基础及作用机制。本实验从中药复方煎煮后汤液浑浊的现象出发,以芍药甘草汤为研究对象,研究其煎煮时沉淀物的形成过程,结果表明汤剂在煎煮过程中会形成具有一定形貌的聚集体,通过静置离心得到沉淀物。定性、定量分析发现沉淀物主要由芍药苷、芍药内酯苷、甘草酸、甘草苷、异甘草苷和没食子酸6种小分子和蛋白质、多糖类大分子组成,虽然其成分与芍药甘草汤组成一致,但去除沉淀物的芍药甘草汤镇痛药效显著降低。在此基础上,本研究从芍药甘草汤中分离得到小分子化合物、多糖及蛋白质部位,各部位的含量分别为60.4、700.7和207.2 mg·g^(-1),大分子蛋白质、多糖含量较小分子含量高,按照多糖∶小分子=11.6∶1、蛋白质∶小分子=3.4∶1的比例在模拟煎煮的状态下制备沉淀物。粒径表征结果表明共同加热后两种沉淀物的粒径会发生明显变化,含量测定结果表明形成沉淀物后游离的小分子化合物芍药苷、芍药内酯苷、甘草酸、甘草苷、异甘草苷和没食子酸含量均明显降低,因此推测小分子成分分别与蛋白质、多糖发生了相互作用。醋酸扭体实验证明模拟制备的沉淀物具有很好的镇痛作用,并能有效降低炎症因子前列腺素E2和一氧化氮的水平,提高抗炎因子白介素-10水平。以上结果证明芍药甘草汤中的沉淀物是其发挥镇痛作用的重要物质基础,而大分子如蛋白质、多糖是构成沉淀物的主要成分,沉淀物发挥镇痛作用的化学机制是大分子与小分子间发生了相互作用。对于芍药甘草汤沉淀物中大分子的研究,不仅为阐明芍药甘草汤的药效物质基础提供新的思路和方法,也为解析传统汤剂起效的科学性提供了参考。

In order to study the analgesic effect of Shaoyao Gancao Decoction,this paper discussed material basis and mechanism from the perspective of macromolecules in traditional Chinese medicine.Inspired by the phenomenon of turbidity after boiling Chinese medicine,this experiment took Shaoyao Gancao Decoction as the research object to study the formation process of precipitation during boiling.The results showed that aggregates with a certain shape were formed in the solvent during the boiling process,and the precipitate was obtained by standing and centrifuging.Analysis found that the precipitation was mainly composed of small molecules such as paeoniflorin,albiflorin,liquiritin,glycyrrhizic acid,isoliquiritin and gallic acid,and macromolecules such as protein and polysaccharide.The composition of precipitate was consistent with that of Shaoyao Gancao Decoction,but the analgesic effect of Shaoyao Gancao Decoction after removing the precipitate was significantly reduced.Based on these results,we isolated small molecular compounds,polysaccharides and protein from Shaoyao Gancao Decoction and their contents are 60.4,700.7 and 207.2 mg·g-1 respectively.We get the ratio,polysaccharide:small molecule=11.6∶1,protein:small molecule=3.4∶1,the precipitate is prepared in the state of boiling.The characterization results showed that the particle size of the precipitate will change significantly after co-heating,and the content determination results showed that the content of the six small molecular compounds which was free in solvent was significantly reduced after the formation of the precipitate.The acetic acid writhing experiment proved that the precipitate has a good analgesic effect,and effectively reduced the levels of inflammatory factors prostaglandin E2 and nitric oxide,and increased the level of anti-inflammatory factor interleukin-10.These results proved that the precipitate in Shaoyao Gancao Decoction is an important material basis for analgesic effect,and macromolecules such as protein and polysaccharide are the main components of the precipitate.The study of macromolecules in the precipitate of Shaoyao Gancao Decoction not only provides new ideas and methods for elucidating the pharmacodynamic material basis of Shaoyao Gancao Decoction,but also provides a reference for analyzing the scientificity of traditional decoction.