Université Libre de Bruxelles, Faculty of Pharmacy, Brussels 1050, Belgium.
Talanta. 2013 May 15;109:116-20. doi: 10.1016/j.talanta.2013.01.062. Epub 2013 Feb 8.
This paper describes the use of a commercially available thin-layer flow through a detector with the sensing block customized in an original design for acetylcholinesterase (AChE) immobilization and suitable for inhibition studies by flow injection analysis (FI). AChE was chemically linked onto a gold disk substrate adjacent to a silver disk electrode. The downstream positioned silver electrode, poised at 0.08 V vs. Ag/AgCl, KCl 3M, permitted the sensitive amperometric detection of liberated thiocholine (TCh) using acetylthiocholine (ATCh) as enzyme substrate. A typical Michaelis-Menten curve was obtained for ATCh within the concentration range 1×10(-5)-1×10(-2)M with a Kmapp of 5.93×10(-4)M. ATCh quantification was achieved with a limit of detection (LOD) of 5.3×10(-6)M. The utility of the developed FI setup was demonstrated for AChE inhibition studies using neostigmine as model compound. The IC50 for neostigmine was obtained to be 1.45×10(-7)M.
本文介绍了使用市售的薄层流动通过检测器，以及在原始设计中定制的传感块，用于乙酰胆碱酯酶（AChE）固定化，并适合通过流动注射分析（FI）进行抑制研究。AChE 通过化学方法连接到金盘基底上，与银盘电极相邻。位于下游的银电极，相对于 Ag/AgCl、3MKCl 电极为 0.08V，允许使用乙酰硫代胆碱（ATCh）作为酶底物，对释放的硫代胆碱（TCh）进行灵敏的电流检测。在 1×10(-5)-1×10(-2)M 的浓度范围内获得了 ATCh 的典型米氏曲线，Kmapp 为 5.93×10(-4)M。ATCh 的定量检测的检测限（LOD）为 5.3×10(-6)M。所开发的 FI 装置的实用性通过使用新斯的明作为模型化合物进行 AChE 抑制研究得到了证明。获得的新斯的明 IC50 为 1.45×10(-7)M。
