The application of a new synthetic substrate to the determination of enteropeptidase in rat small intestine and human intestinal biopsies. 
The application of a new synthetic substrate to the direct determination of enteropeptidase is described.  The substrate Gly-(L-Asp)4-L-Lys-2-naphthylamide contains the amino acid sequence of the activation peptides of trypsinogen linked via an amide bond to the fluorophore 2-naphthylamine.  The sequence of amino acids is responsible for the specificity and substrate recognition of the enteropeptidase-catalyzed activation of trypsinogen.  Interference in the assay by trypsin is prevented by the addition of soybean trypsin inhibitor to the substrate solution.  The fluorimetric determination of the liberated 2-naphthylamine allows the direct observation of the reaction kinetics.  For the hyrolysis of the synthetic substrate by purified enteropeptidase the pH optimum was 8.2 and the Km 0.17 mmol/l. The new substrate was used to determine the distribution of enteropeptidase along the rat small intestine and also to measure enteropeptidase activity in human intestinal biopsies.