Differential assay of human pancreatic and salivary alpha-amylases in serum using a new fluorogenic substrate

The difference in the mode action of human pancreatic and salivary alpha-amylases on O-6-deoxy-6-[(2-pyridyl)amino]-alpha-D-glucopyranosyl-(1----4)-O-alpha-D - glucopyranosyl-(1----4)-O-alpha-D-glucopyranosyl-(1----4)-O-alpha-D- glucopyranosyl-(1----4)-O-alpha-D-glucopyranosyl-(1----4)-D-glucitol FG6R), a fluorogenic derivative of maltohexaitol, was found. The products of the enzymatic hydrolysis were analyzed by high-performance liquid chromatography (HPLC) in 8 min. FG6R was hydrolyzed by these enzymes to O-6-deoxy-6-[(2-pyridyl)amino]-alpha-D-glucopyranosyl- (1----4)-O-alpha-D-glucopyranosyl-(1----4)-D-glucose (FG3) and maltotriitol, or O-6-deoxy-6-[(2-pyridyl)amino]-alpha-D- glucopyranosyl-(1----4)-O-alpha-D-glucopyranosyl-(1----4)-O-alpha-D- glucopyranosyl-(1----4)-D-glucose (FG4) and maltitol. Pancreatic alpha-amylase produced more FG4 than salivary alpha-amylase. Taking advantage of the differences in action of the two amylases, a differential alpha-amylase assay in serum was performed. The method is simple and rapid and can be used for routine clinical assays of alpha-amylases.