一种用于脂肪酸酰胺水解酶的简单终点测定法，避免使用氯仿萃取阶段。
A simple stopped assay for fatty acid amide hydrolase avoiding the use of a chloroform extraction phase.
作者信息
Boldrup Linda, Wilson Sandy J, Barbier Ann J, Fowler Christopher J
机构信息
Department of Pharmacology and Clinical Neuroscience, Umeå University, SE901 87 Umeå, Sweden.
出版信息
J Biochem Biophys Methods. 2004 Aug 31;60(2):171-7. doi: 10.1016/j.jbbm.2004.04.020.
A stopped assay for fatty acid amide hydrolase (FAAH) has been developed, whereby the enzyme reaction product ([(3)H]ethanolamine) was separated from substrate (anandamide [ethanolamine-1-(3)H]), by differential adsorption to charcoal. The assay gave a better extraction efficiency when acidic rather than alkaline charcoal solutions were used to stop the reaction, and a very good ratio of sample/blank was also seen. The acidic charcoal assay gave the expected sensitivities to compounds known to inhibit FAAH (palmitoyltrifluoromethyl ketone, arvanil, AM404 and indomethacin). It is concluded that the acidic charcoal extraction method provides a robust and simple stopped assay for FAAH without the need to use potentially hazardous solvents like chloroform.
已开发出一种用于脂肪酸酰胺水解酶（FAAH）的终止测定法，通过将酶反应产物（[³H]乙醇胺）与底物（花生四烯酸乙醇胺[乙醇胺 - 1 - ³H]）进行差示吸附到活性炭上而实现分离。当使用酸性而非碱性活性炭溶液终止反应时，该测定法具有更好的提取效率，并且样品/空白比值也非常理想。酸性活性炭测定法对已知抑制FAAH的化合物（棕榈酰三氟甲基酮、arvanil、AM404和吲哚美辛）具有预期的灵敏度。结论是，酸性活性炭提取方法为FAAH提供了一种稳健且简单的终止测定法，无需使用如氯仿等潜在危险溶剂。
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