New fluorogenic triacylglycerol analogs as substrates for the determination and chiral discrimination of lipase activities

A new type of fluorogenic and isomerically pure 1(3)-O-alkyl-2,3 (3,2)-diacyl glycerols was synthesized that can be used as substrate for the determination of lipase activities. These compounds contain a fluorescent pyrene acyl chain and, as a potent quencher of pyrene fluorescence, a trinitrophenylamino acyl residue. In their intact form, the fluorogens show only low fluorescence intensity. Upon lipase-induced or chemical hydrolysis of the substrates, however, the fluorophore and quencher separate from each other. This leads to a gradual increase in pyrene fluorescence, reflecting the time-dependent progress of lipolysis and, under substrate saturation conditions, lipase activity. This lipase assay is continuous and does not require separation of substrate and reaction products. Short- and long-chain homologues as well as optical isomers of the fluorogenic alkyldiacyl glycerols were hydrolyzed by pancreatic lipase, hepatic lipase, and lipoprotein lipase at highly different rates depending on the substrate or enzyme preparation and source (e.g., postheparin plasma or cultured cells). It is proposed that a useful set of enantiomeric and/or homologous substrates in combination with appropriate reaction media might be applied to the selective determination of a lipase in a mixture of lipases, e.g., hepatic and lipoprotein lipase in PHP, for medical diagnostics.-Duque, M., M. Graupner, H. Stutz, I. Wicher, R. Zechner, F. Paltauf, and A. Hermetter. New fluorogenic triacylglycerol analogs as substrates for the determination and chiral discrimination of lipase activities.