Simple and rapid chemiluminescence assay for lipase activity in pharmaceutical preparations using proenhancer substrate

Abstract

A novel chemiluminescence (CL) assay method, using a HDI-laurate [lauric acid ester of 2-(4-hydroxyphenyl)-4,5-diphenylimidazole (HDI)] as a proenhancer substrate, was applied to the determination of lipase (triglycerol lipase, EC 3.1.1.3) activity in pharmaceutical preparations. The method is based on an enhanced CL reaction of luminol-horseradish peroxidase (HRP)-hydrogen peroxide with HDI, which is liberated from the proenhancer substrate by enzymatic hydrolysis. The proposed method involves a homogeneous reaction system in which enzymatic hydrolysis of HDI-laurate and enhanced CL reaction with HDI occur in the same reaction mixture. The lipase activities of three commercially available preparations were measured by the proposed CL method. Linear relationships were obtained (r > 0.977) between the concentrations and CL intensities in all of the tested preparations. The results obtained by the proposed method were compared with those by the titration method in Japanese Pharmacopoeia, and good correlations were obtained between both methods (r > 0.919). The CL method is simple and rapid, permitting the completion of single assay within 5 min. The sensitivity and repeatability of the CL method were also superior to those of the titration method. © 2006 The Japan Society for Analytical Chemistry.