Fatty acid amide hydrolase assay

Fatty acid amide hydrolase in brain Profenofos, Iribufos, ethyl octylphusphonofluoridaie Activity assay Quistad et al. (2001)... 

Anandamide (AEA) is the most studied member of a new class of lipid mediators, collectively called endocannabinoids. The biological activity of AEA at cannabinoid and noncannabinoid receptors depends on its Hfe span in the extracellular space, which is regulated by a rapid cellular uptake, followed by intracellular degradation by the enzyme AEA hydrolase (fatty acid amide hydrolase). Here, we present the methodological details of the procedures that we have developed to assay fatty acid amide hydrolase activity and to characterize AEA transport through cell membranes in a new ideal ex vivo system like brain synaptosomes. 

The discovery of anandamide (arachidonoyl ethanolamide, AEA) and of its manifold roles in the central nervous system and in the periphery (reviewed in refs. 1 and 2) prompted several researchers to develop analytical methods to assay and characterize the activity of the enzymes responsible for AEA metabolism in various cells and tissues. Fatty acid amide hydrolase (arachidonoyl ethanolamide amidohydrolase, EC 3.5.1.4 FAAH) has emerged as the key AEA hydrolase, showing a molecular mass of approx 64 kDa and an optimum pH of around 9.0 (3). Recently, FAAH has been crystallized,and its three dimensional structure has been determined at 2.8A resolution (4). This enzyme cleaves the amide bond and releases arachidonic acid and ethanolamine. High-performance liquid chromatography (HPLC) is the most widely used method to determine FAAH activity from different sources. We developed a new method (5) based on reversed-phase (RP)-HPLC and on-line scintillation counting, which combines the need for high resolution, reproducibility, and sensitivity... 

Maccarrone, M Bari, M., and Finazzi Agro A. (1999) A sensitivity and specific radiochromatographic assay of fatty acid amide hydrolase activity Anal. Biochem. 267, 314-318. 

Rakers C, Zoerner AA, Engeli S, Batkai S, Jordan J, Tsikas D. Stable isotope liquid chromatography—tandem mass spectrometry assay for fatty acid amide hydrolase activity. Anal Biochem 2012 421 699-705. 

Yapa U, Prusakiewicz JJ, Wrightstone AD, Christine LJ, Palandra J, Groeber E, et al. High-performance liquid chromatography—tandem mass spectrometry assay of fatty acid amide hydrolase (FAAH) in blood FAAH inhibition as clinical biomarker. Anal Biochem 2012 421 556-65. 

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