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目的研究一种基于超分子“印迹模板”黄芩苷与酶具有高度特异性亲和作用的分离方法,能实现黄芩苷超分子对其代谢酶勾钓分离。方法通过共反应剂法对DEAE-52纤维素进行化学改性键合黄芩苷,合成DEAE-52纤维素黄芩苷酯,用红外光谱分析检验键合效果;比较DEAE-52纤维素黄芩苷酯与DEAE-52纤维素分别作为固定相制备层析色谱柱对黄芩中代谢酶的分离富集效果,检验DEAE-52纤维素黄芩苷酯与DEAE-52纤维素对代谢酶的特异性吸附性的性能;通过分析获得的酶组分的酶解率检验酶组分的酶解活性,并用电泳检验其纯度。结果红外结果显示,与DEAE-52纤维素相比,形成的DEAE-52纤维素黄芩苷酯在1740 cm-1左右出现了酯基的特征吸收峰,1248 cm-1、1060 cm-1出现C-O-C的伸缩振动峰;对酶的分离效果显示,DEAE-52纤维素黄芩苷酯对黄芩的某种酶组分具有特异性吸附。结论运用超分子“印迹模板”的自主识别规律,黄芩苷与代谢酶可按其超分子“印迹模板”进行勾钓分离。本研究为中药代谢酶的寻找、分离纯化的研究提供了新的方法,为中药成分的生化代谢途径的研究奠定基础。
OBJECTIVE To study a separation method based on supramolecular “blotting template” and baicalin, which has a highly specific affinity to the enzyme, and to realize the separation of its metabolic enzymes by baicalin supermolecules. Methods Baicalin was chemically modified with co-reactant to synthesize DEAE-52 cellulose baicalin. The binding effect of DEAE-52 cellulose baicalin ester was evaluated by infrared spectroscopy. DEAE-52 cellulose were used as stationary phase preparative chromatography column for the separation and enrichment of the metabolic enzymes in Scutellaria baicalensis Georgi, DEAE-52 cellulose and DEAE-52 cellulose were tested for their specific adsorbability The enzymatic activity of the enzyme components was checked by analyzing the enzymatic hydrolysis rate of the obtained enzyme components and the purity of the enzyme components was examined by electrophoresis. Results The infrared results showed that the characteristic absorption peak of ester group appeared at about 1740 cm-1 in DEAE-52 cellulose baicalein compared with DEAE-52 cellulose, and the COC appeared at 1248 cm-1 and 1060 cm-1 The results showed that DEAE-52 cellulose baicalin has specific adsorption to some enzyme components of Scutellaria baicalensis Georgi. Conclusion According to the autonomic recognition rules of supramolecules “blotting template”, baicalin and metabolic enzymes can be separated by hook-and-loop method based on their supramolecular “blot template”. This study provides a new method for the search of traditional Chinese medicine metabolic enzymes and the study of isolation and purification, which lays the foundation for the study of the biochemical metabolic pathways of traditional Chinese medicine.