A 4-methoxy-2-naphthylamide substrate for the histochemical localization of esteroproteases in the submandibular gland of the rat.

The 4-methoxynaphthylamide (MNA) derivative of D-Val-Leu-Arg-4-MNA has been used as a substrate for the histochemical localization of esteroproteases in the submandibular gland of rats, and compared with the substrate alpha N-O-met. The hydrolysis of Val-Leu-Arg-4-MNA by esteroproteases was investigated using spectrophotometry and isoelectric focusing. Both methods demonstrated that the substrate is cleaved by different enzymes and is not a monospecific kallikrein marker, although Val-Leu-Arg-4-MNA had a much smaller spectrum of enzyme activities than alpha N-O-met. The activity against Val-Leu-Arg-4-MNA was totally inhibited by aprotinin but only partly by soybean trypsin inhibitor. A comparison of the histochemical localization of esteroproteases given by the substrate with the immunofluorescence localization of kallikrein showed that all the enzymes that hydrolysed Val-Leu-Arg-4-MNA and kallikrein were present in the same tissue structures.