含羧基中药成分荧光标记方法的建立及优化

用荧光标记技术对含羧基类中药成分进行标记,探索中药成分荧光标记方法,提高其检测灵敏度、为中药药代动力学研究奠定基础。以荧光试剂8-氨基芘-1,3,6-三磺酸三钠盐(APTS)和绿原酸组成研究体系,对标记条件进行优化。反应体系中加入1-乙基-3-(3-二甲氨丙基)碳二亚胺盐酸盐(EDC.HCl)与N-羟基丁二酰亚胺(NHS)可使衍生反应在室温下进行。APTS标记绿原酸的较优反应条件为:绿原酸与APTS的浓度比为1∶5,EDC的浓度为5 mmol/L,NHS的浓度为0.33 mmol/L。以0.1mol/L、pH5.50的NaH2PO4-Na2HPO4为缓冲溶液,绿原酸首先与EDC、NHS避光反应4 h,再与APTS反应4h,即可达到较好的标记效果。此标记方法操作简便,灵敏度高,可以用于含羧基类中药成分的荧光标记研究。

Establish and optimization of fluorescent labeling of carboxylfractions in Chinese medicine

Labeling of carboxyl fractions is important to the research of traditional Chinese medicine.It can improve the detection sensitivity and perfect the pharmacokinetics studying of Chinese medicine.The labeling method and its evaluation system are presentedin this paper.In the work,trisodium 8-aminopyrene-1,3,6-trisulfonate(APTS)and chlorogenic acid(CGA)were chosen to forma labeling system.If EDC and NHS exist,the labeling reaction can take placeat room temperature.Experimental resultsshowthat the prefer buffer system was NaH2PO4-Na2HPO4(0.1mol/L,pH5.50).CGA reacted with EDC and NHS in above buffer for 4 h at room temperature.The concentration of EDC and NHS was 5 mmol/L and 0.33 mmol/L,respectively.Then,APTS(concentration ratio of CGA and APTS was 1∶5)was added to above reaction systems and continue reacted for 4 h.Conclusions The labling method is not only simple but also sensitive,and can be possibly used in study ofcarboxylfractionof Chinese medicinelabeling.