Differentiation between isomeric oxidative metabolites of a piperidine-containing drug by liquid chromatography-tandem mass spectrometry with stable-isotope tracer technique.
This report describes the application of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to differentiation of hydroxylations and N-oxidations and of two different aliphatic hydroxylations in the investigation of the metabolism of pibutidine hydrochloride, a novel H2 antagonist, the structure of which includes a piperidine ring. Pibutidine metabolites in urine samples from adult male volunteers after oral administration of pibutidine hydrochloride were separated by reversed-phase LC and ionized using an electrospray ionization (ESI) interface. A hydroxylated form of pibutidine was distinguished from the N-oxide by comparison of their product ion spectra, although their mass-to-charge ratios of protonated molecules were identical. Further, two hydroxylated compounds were present in rat microsomal incubation mixtures with pibutidine. The distinction between their positions of hydroxylation (beta- and gamma-carbon hydroxylation) on the piperidine ring was studied using [piperidine-2H10] pibutidine as incubation substrate. The production of the beta-hydroxylated form was accompanied by the elimination of three 2H, resulting from a mechanism including the formation of iminium/enamine. The participation of the iminium ion intermediate in the beta-hydroxylation was confirmed by the observation that a cyanide adduct of pibutidine was formed instead of the beta-hydroxylated form when another incubation was performed in the presence of cyanide.
Kato
Koji
K
Drug Metabolism Laboratory, Pharmaceutical Research Laboratories, Taisho Pharmaceutical Co. Ltd, Saitama, Saitama 330-8530, Japan. s15379@ccm.taisho.co.jp
Jingu
Shigeji
S
Ogawa
Naoyoshi
N
Higuchi
Shohei
S
D001711:Biotransformation; D002853:Chromatography, Liquid / Q000379:methods*; D006635:Histamine H2 Antagonists / Q000737:chemistry / Q000378:metabolism* / Q000493:pharmacokinetics; D006801:Humans; D006900:Hydroxylation; D066298:In Vitro Techniques; D007536:Isomerism; D007554:Isotopes; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D010084:Oxidation-Reduction; D010880:Piperidines / Q000737:chemistry* / Q000378:metabolism* / Q000493:pharmacokinetics; D011725:Pyridines / Q000737:chemistry / Q000378:metabolism* / Q000493:pharmacokinetics
D006635:Histamine H2 Antagonists; D007554:Isotopes; D010880:Piperidines; D011725:Pyridines; C063924:IT 066