A new procedure for the zymography of monophenolase and o-diphenolase activities of polyphenol oxidase (PPO), and peroxidase (POX) is proposed using a highly sensitive, chromogenic nucleophile 3-methyl-2-benzothiazolinone hydrazone (MBTH), which traps quinones. The procedure allowed the distinction between PPO isoenzymes from sorghum and mushroom in the same gel, as well as between monophenolase and o-diphenolase activities of PPO isoenzymes from crude extracts of mushroom. Three isoforms were detected with monophenolase activity, and at least seven isoforms were detected with o diphenolase activity. The procedure also allows the identification of PPO isoforms exhibiting monophenolase activity from a crude extract. The sensitivity, speed and ability to discriminate between mono and o-diphenolase activities could make the newly developed procedure a universal and powerful method for the routine zymography of PPOs and POXs in biological materials. The assay also discriminates the activities of PPOs, POXs and laccases.

polyphenol oxidase peroxidase zymography activity staining isoenzymes mushroom