title: The use of 18O-exchange and base-catalyzed N-dealkylation with liquid chromatography/tandem mass spectrometry to identify carbinolamide metabolites.
abstract: Oxidation of N-alkyl-substituted amides is a common transformation observed in metabolism studies of drugs and other chemicals. Metabolism at the alpha carbon atom can produce stable carbinolamide compounds, which may be abundant enough to require complete confidence in structural assignments. In a drug discovery setting, rapid structural elucidation of test compounds is critical to inform the compound selection process. Traditional approaches to the analysis of carbinolamides have relied upon the time-consuming synthesis of authentic standards or purification of large enough quantities for characterization by nuclear magnetic resonance (NMR). We describe a simple technique used in conjunction with liquid chromatography/tandem mass spectrometry (LC/MS/MS) which demonstrates the chemical identity of a carbinolamide by its distinctive ability to reversibly exchange [(18)O]water through an imine intermediate. A key advantage of the technique is that the chromatographic retention times of metabolites are preserved, allowing direct comparisons of mass chromatograms from non-treated and [(18)O]water-treated samples. Metabolites susceptible to the treatment are clearly indicated by the addition of 2 mass units to their original mass. An additional test which can be used in conjunction with (18)O-exchange is base-catalyzed N-dealkylation of N-(alpha-hydroxy)alkyl compounds. The use of the technique is described for carbinolamide metabolites of dirlotapide, loperamide, and a proprietary compound.
authors.affiliation: Department of Pharmacokinetics, Dynamics, and Metabolism, Pfizer Global R&D, Groton, CT 06340, USA. andrew.j.bessire@pfizer.com
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Bessire
authors.forename: Alfin D N
authors.initials: AD
authors.lastname: Vaz
authors.forename: Gregory S
authors.initials: GS
authors.lastname: Walker
authors.forename: Wei Wei
authors.initials: WW
authors.lastname: Wang
authors.forename: Raman
authors.initials: R
authors.lastname: Sharma
mesh_terms: D000818:Animals; D002219:Carbamates / Q000008:administration & dosage / Q000737:chemistry* / Q000378:metabolism*; D002384:Catalysis; D002853:Chromatography, Liquid / Q000379:methods*; D003640:Dealkylation; D005260:Female; D007211:Indoles / Q000008:administration & dosage / Q000737:chemistry* / Q000378:metabolism*; D008297:Male; D015394:Molecular Structure; D010103:Oxygen Isotopes / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D002219:Carbamates; D007211:Indoles; D010103:Oxygen Isotopes; C521290:dirlotapide