The metabolomics of (±)-arecoline 1-oxide in the mouse and its formation by human flavin-containing monooxygenases

The alkaloid arecoline is a main constituent of areca nuts that are chewed by approximately 600 million persons worldwide. A principal metabolite of arecoline is arecoline 1-oxide whose metabolism has been poorly studied. To redress this, synthetic (±)-arecoline 1-oxide was administered to mice (20 mg/kg p.o.) and a metabolomic study performed on 0–12 h urine using ultra-performance liquid chromatography-coupled time-of-flight mass spectrometry (UPLC-TOFMS) with multivariate data analysis. A total of 16 mass/retention time pairs yielded 13 metabolites of (±)-arecoline 1-oxide, most of them novel. Identity of metabolites was confirmed by tandem mass spectrometry. The principal pathways of metabolism of (±)-arecoline 1-oxide were mercapturic acid formation, with catabolism to mercaptan and methylmercaptan metabolites, apparent CC double-bond reduction, carboxylic acid reduction to the aldehyde (a novel pathway in mammals), N-oxide reduction, and de-esterification. Relative percentages of metabolites were determined directly from the metabolomic data. Approximately, 50% of the urinary metabolites corresponded to unchanged (±)-arecoline 1-oxide, 25% to other N-oxide metabolites, while approximately, 30% corresponded to mercapturic acids or their metabolites. Many metabolites, principally mercapturic acids and their derivatives, were excreted as diastereomers that could be resolved by UPLC-TOFMS. Arecoline was converted to arecoline 1-oxide in vitro by human flavin-containing monooxygenases FMO1 (KM: 13.6 ± 4.9 μM; VMAX: 0.114 ± 0.01 nmol min−1 μg−1 protein) and FMO3 (KM: 44.5 ± 8.0 μM; VMAX: 0.014 ± 0.001 nmol min−1 μg−1 protein), but not by FMO5 or any of 11 human cytochromes P450. This report underscores the power of metabolomics in drug metabolite mining.