Assay of MAO Inhibition by Chromatographic Techniques (HPLC/HPLC-MS).

ABSTRACT

Monoamine oxidase (MAO) enzymes (MAO A and B) catalyze the oxidative deamination of biogenic amines, neurotransmitters, and xenobiotic amines and contribute to the regulation of the content of these active substances in mammalian organisms. The oxidation of biogenic amines by MAO produces hydrogen peroxide (H2O2) and aldehydes that represent risk factors for oxidative injury. The inhibitors of MAO are useful as antidepressants and neuroprotective agents. Usually, the assays of MAO determine amine deamination products or measure the H2O2 released by using direct spectrophotometric or fluorimetric methods. Direct methods are more prone to interferences and can afford inaccurate results. Those limitations can be avoided by using chromatographic techniques. This work describes a chromatographic method to assay MAO A and MAO B activity by using kynuramine as a nonselective substrate and the subsequent analysis of 4-hydroxyquinoline by RP-HPLC-DAD-fluorescence and mass spectrometry (MS). Alternatively, the assay uses the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin as a substrate of MAO that is oxidized (bioactivated) to neurotoxic pyridinium cations which are analyzed by HPLC. These methods are applied to assess the inhibition of MAO by bioactive ß-carboline alkaloids occurring in foods, plants, and biological systems.