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7-hydroxycoumarinyl-arachidonate (7-HCA), a fluorogenic substrate for monoacylglycerol lipase (MAGL), can be used with MAGL as a novel fluorescence-based assay to screen small molecule inhibitors of MAGL.

MAGL protein catalyzed the hydrolysis of 7-HCA to generate arachidonic acid and the highly fluorescent 7-hydroxyl coumarin (7-HC). 7-hydroxycoumarin (7-HC) can be monitored spectrophotometrically (excitation at 335 nm, emission at 450 nm). Release of 7-HC was measured using a fluorometer. MAGL protein catalyzed the hydrolysis of 7-HCA with an apparent Km of 9.8 μM and Vmax of 1.7 mmol/min/mg of protein. The assay is specific for MAGL as assay buffer alone or heat-denatured MAGL protein showed no significant activity against 7-HCA [1]. 7-hydroxycoumarinyl Arachidonate is the arachidonic acid ester of 7-hydroxycoumarin (umbelliferone) and can be used as a substrate for cPLA2. Hydrolysis of 7-hydroxycoumarinyl arachidonate by phospholipase resulted in the release of the fluorescent compound [2,3].

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