Abstract
Glucuronides of piperazine hydroxylamines are rarely reported in the literature, and even more rarely are their structures unambiguously identified. One major metabolite was detected by liquid chromatography/mass spectrometry-radioactivity in urine from monkeys treated with the aryl piperazine oral hypoglycemic agent 9-[(1S,2R)-2-fluoro-1-methylpropyl]-2-methoxy-6-(1-piperazinyl) purine hydrochloride (1). The mass spectrum of this metabolite indicated that it was both monooxygenated and glucuronidated on the piperazine ring. Possible structures included the N- or O-glucuronic acid conjugates of a carbinolamine, hydroxylamine, or N-oxide. Treatment with β-glucuronidase gave a monooxygenated derivative of the parent compound. 1H NMR analysis of either the glucuronic acid conjugate or the monooxygenated product provided insufficient evidence to unambiguously determine their structures. Incubation of 1 with pig liver microsomes resulted in formation of the same monooxygenated derivative derived from β-glucuronidase treatment of the glucuronide metabolite. This in vitro system was used to generate sufficient material for analysis by 13C NMR, and the metabolite was identified as a hydroxylamine derivative 2. Incubation of the hydroxylamine with monkey liver microsomes and uridine diphospho-5′-glucuronic acid gave the same glucuronic acid conjugate as that observed in monkey urine. 13C NMR analysis of this biosynthetic product led to its unequivocal structure assignment as the O-glucuronic acid conjugate of the hydroxylamine 3.
Footnotes
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↵1 Abbreviations used are: 1, 9-[(1S,2R)-2-fluoro-1-methylpropyl]-2-methoxy-6-(1-piperazinyl) purine hydrochloride; LC/MS, liquid chromatography/mass spectrometry; MS/MS, tandem mass spectrometry; [14C]1, 9-[(1S,2R)-2-fluoro-1-methylpropyl]-2-methoxy-6-(1-piperazinyl)-2-[14C]purine hydrochloride; HPLC, high-performance liquid chromatography; NH4OAc, ammonium acetate; TFA, trifluoroacetic acid; MeOH, methanol; UDPGA, uridine diphospho-5′-glucuronic acid.
- Received April 18, 2003.
- Accepted September 17, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
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