Damsma G, Westerink B H, Horn A S
J Neurochem. 1985 Nov;45(5):1649-52. doi: 10.1111/j.1471-4159.1985.tb07238.x.
A simple, efficient, economic, and sensitive method is presented for the detection of choline and acetylcholine in neuronal tissue using HPLC, a postcolumn enzyme reactor with immobilized enzyme, and electrochemical detection. The method is based on a separation of choline and acetylcholine by cation exchange HPLC followed by passage of the effluent through a postcolumn reactor containing a mixture of acetylcholinesterase and choline oxidase; the latter enzyme converts choline to betaine and hydrogen peroxide, the former enzyme hydrolyzes acetylcholine to acetate and choline. The hydrogen peroxide produced is electrochemically detected. A simple and efficient preparation of neuronal tissue is described using an optional prepurification step on Sephadex G-10 columns, offering the possibility to detect choline and acetylcholine as well as catecholamines and their related metabolites in the same tissue sample. The sensitivity of the assay system is 250 fmol for choline and 500 fmol for acetylcholine.
本文介绍了一种简单、高效、经济且灵敏的方法，用于使用高效液相色谱（HPLC）、固定化酶的柱后酶反应器和电化学检测法来检测神经元组织中的胆碱和乙酰胆碱。该方法基于通过阳离子交换HPLC分离胆碱和乙酰胆碱，然后使流出物通过含有乙酰胆碱酯酶和胆碱氧化酶混合物的柱后反应器；后一种酶将胆碱转化为甜菜碱和过氧化氢，前一种酶将乙酰胆碱水解为乙酸盐和胆碱。产生的过氧化氢通过电化学检测。描述了一种简单高效的神经元组织制备方法，该方法在Sephadex G-10柱上使用可选的预纯化步骤，使得在同一组织样品中检测胆碱、乙酰胆碱以及儿茶酚胺及其相关代谢物成为可能。该检测系统对胆碱的灵敏度为250飞摩尔，对乙酰胆碱的灵敏度为500飞摩尔。
