PT  - JOURNAL ARTICLE
AU  - Sophie C. Turfus
AU  - Mark C. Parkin
AU  - David A. Cowan
AU  - John M. Halket
AU  - Norman W. Smith
AU  - Robin A. Braithwaite
AU  - Simon P. Elliot
AU  - Glyn B. Steventon
AU  - Andrew T. Kicman
TI  - Use of Human Microsomes and Deuterated Substrates: An Alternative Approach for the Identification of Novel Metabolites of Ketamine by Mass Spectrometry
AID  - 10.1124/dmd.108.026328
DP  - 2009 Aug 01
TA  - Drug Metabolism and Disposition
PG  - 1769--1778
VI  - 37
IP  - 8
4099  - http://dmd.aspetjournals.org/content/37/8/1769.short
4100  - http://dmd.aspetjournals.org/content/37/8/1769.full
SO  - Drug Metab Dispos2009 Aug 01; 37
AB  - In vitro biosynthesis using pooled human liver microsomes was applied to help identify in vivo metabolites of ketamine by liquid chromatography (LC)-tandem mass spectrometry. Microsomal synthesis produced dehydronorketamine, seven structural isomers of hydroxynorketamine, and at least five structural isomers of hydroxyketamine. To aid identification, stable isotopes of the metabolites were also produced from tetra-deuterated isotopes of ketamine or norketamine as substrates. Five metabolites (three hydroxynorketamine and two hydroxyketamine isomers) gave chromatographically resolved components with product ion spectra indicating the presence of a phenolic group, with phenolic metabolites being further substantiated by selective liquid-liquid extraction after adjustments to the pH. Two glucuronide conjugates of hydroxynorketamine were also identified. Analysis by LC-coupled ion cyclotron resonance mass spectrometry gave unique masses in accordance with the predicted elemental composition. The metabolites, including the phenols, were subsequently confirmed to be present in urine of subjects after oral ketamine administration, as facilitated by the addition of deuterated metabolites generated from the in vitro biosynthesis. To our knowledge, phenolic metabolites of ketamine, including an intact glucuronide conjugate, are here reported for the first time. The use of biologically synthesized deuterated material as an internal chromatographic and mass spectrometric marker is a viable approach to aid in the identification of metabolites. Metabolites that have particular diagnostic value can be selected as candidates for chemical synthesis of standards.