Kronkvist K, Lövgren U, Edholm L E, Johansson G
Department of Analytical Chemistry, University of Lund, Sweden.
J Pharm Biomed Anal. 1993 Jun;11(6):459-67. doi: 10.1016/0731-7085(93)80158-w.
A competitive ELISA with electrochemical detection in a flow injection system (FIA) has been developed for determinations of the steroid drug budesonide in biological samples. Plasma samples were cleaned from interfering and cross-reacting compounds by two pretreatment steps consisting of a solid-phase extraction and a liquid chromatography fractionation. The enzyme label was alkaline phosphatase, which was used with p-aminophenyl phosphate (PAPP) as a substrate. The product, p-aminophenol, was detected electrochemically at a glassy carbon electrode at 250 mV (vs Ag/AgCl). The limited stability of both the substrate and the product influenced the performance of the method and had to be taken into account in the procedure by a normalization with time. Budesonide could be quantified in plasma samples down to 10 pM. The major sensitivity-limiting factor was the amperometric background response, probably due to spontaneous hydrolysis of PAPP to p-aminophenol.
已开发出一种在流动注射系统（FIA）中进行电化学检测的竞争性酶联免疫吸附测定法（ELISA），用于测定生物样品中的甾体药物布地奈德。血浆样品通过固相萃取和液相色谱分级分离这两个预处理步骤，去除干扰和交叉反应的化合物。酶标记物是碱性磷酸酶，它与对氨基苯磷酸（PAPP）作为底物一起使用。产物对氨基酚在玻碳电极上于250 mV（相对于Ag/AgCl）进行电化学检测。底物和产物的有限稳定性影响了该方法的性能，因此在程序中必须通过时间归一化来加以考虑。布地奈德在血浆样品中的定量下限可达10 pM。主要的灵敏度限制因素是安培背景响应，这可能是由于PAPP自发水解成对氨基酚所致。
