Continuous fluorometric assay of phospholipase A2 with pyrene-labeled lecithin as a substrate

1,2-Bis[4-(1-pyreno)butanoyl]-sn-glycero-3-phosphorylcholine was synthesized as a fluorogenic substrate for phospholipase A₂. It has a critical micellar concentration of 7.3 μm and gives only excimer fluorescent emission at 480 nm in aqueous micellar dispersion. When hydrolyzed by phospholipase A₂, the products give only monomer emission which is monitored best at 382 and 400 nm. Conditions were developed for an assay for phospholipase A₂ using this substrate. The assay was sensitive to as little as 8 ng of pure porcine pancreatic phospholipase A₂.