Inhibitive activities detection of monoamine oxidases (MAO) A and B inhibitors in human liver MAO incubations by UPLC-ESI-MS/MS

A sensitive UPLC-ESI-MS/MS method was developed for determining and screening of inhibitors of monoamine oxidase (MAO) by using mix MAO enzymes prepared from human liver. 5-Hydroxytryptamine (5-HT) and 2-phenethylamine (2-PEA) were used as substrates for MAO-A and MAO-B in incubations, and 5-hydroxyindole-3-acetic acid (5-HIAA) and phenylacetic acid (PAA) resulting from 5-HT and 2-PEA were used as markers to evaluate inhibitive activities of test compounds on MAO-A and MAO-B. Proper separation was achieved for positive multiple reaction monitoring of 5-HIAA (m/z 192.1 -> 146.1) and negative multiple reaction monitoring PAA (m/z 135.0 -> 91.0) via isocratic elution (0.1% fromic acid:acetonitrile = 60:40) on a HSS T3 column following a simple precipitation of proteins for sample treatment. The relative standard deviations of intra- and inter-day precisions were ranged from 1.74% to 6.76% and 0.77% to 9.35%. The mean accuracies for the quality control samples were 101.37 +/- 6.60% and 101.39 +/- 2.85%, respectively. This method exhibited characteristics of small total reaction volume (100 mu l), short analysis time (3.5 min), highly sensitivity, low cost and without matrix effect (103.56 +/- 1 2.33% to 112.63 +/- 8.57% for 5-HIAA and 105.68 +/- 8.75% to 112.76 +/- 4.67% for PAA). The developed method was successfully applied for detection of the MAO-A and MAO-B inhibitive activities by model drugs, including pargyline, clorgyline, as well as p-carboline alkaloids from Peganum harrnala. (C) 2015 Published by Elsevier B.V.