Identification of some N-hydroxylated metabolites of (±)-3,4-methylenedioxymethamphetamine in horse urine by gas chromatography-mass spectrometry

Abstract

1. The in vivo biotransformation of (±)-3,4-methylenedioxymethamphetamine [(±)-MDMA] in the thoroughbred horse was determined after oral administration.

2. Unconjugated compounds and aglycones were isolated from enzyme-hydrolysed urine by solid-phase extraction using mixed-mode cartridges. The basic isolates were derivatized (trimethylsilylether, TMS) and analysed by positive-ion electron ionization/gas chromatography-mass spectrometry (EI+/GC-MS). MDMA and 10 Phase I metabolites containing the arylisopropylamine substructure were detected.

3. N-Hydroxy amphetamine and N-hydroxymethamphetamine were synthesized. The EI + mass spectra of their O-TMS derivatives showed characteristic α-cleavage ions at m/z 132 and 146, respectively, as base peaks. Based upon these data, five putative N-hydroxylated metabolites of MDMA were detected.

4. In the horse, (±)-MDMA is metabolized by oxidative N-demethylation to form the primary amine methylenedioxyamphetamine (MDA). Both MDMA and MDA are further metabolized by oxidative demethylenation (cleavage and O-demethylation of the benzodioxole moiety) to form the corresponding catechols, 3-O-methylation to form the guaiacols and N-oxidation of the secondary and primary amine metabolites to form the hydroxylamines.

5. Both phenolic and N-hydroxy metabolites of (±)-MDMA undergo Phase II conjugation before excretion in urine.