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Identification of ketobemidone metabolites by capillary LC-MS/MS: Part I: Method development for analysis of in vivo micro-dialysates
Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
Manuscript (Other academic)
URN: urn:nbn:se:uu:diva-95953OAI: oai:DiVA.org:uu-95953DiVA: diva2:170343
Available from: 2007-05-10 Created: 2007-05-10 Last updated: 2010-01-13Bibliographically approved
In thesis
1. Liquid Chromatography-Mass Spectrometry as a Tool for Drug Metabolite Identification in Biological Fluids: With Application to Ketobemidone
Open this publication in new window or tab >>Liquid Chromatography-Mass Spectrometry as a Tool for Drug Metabolite Identification in Biological Fluids: With Application to Ketobemidone
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Electrospray ionization (ESI) mass spectrometry (MS) in combination with liquid chromatography (LC) is an excellent tool for the identification of drug metabolites. Utilizing this hyphenated technique in combination with proper sample pretreatment, the metabolic pathways of the analgesic drug ketobemidone were investigated in human urine and rat microdialysate from blood and brain. Two novel phase I metabolites (ketobemidone N-oxide and meta-hydroxymethoxyketobemidone) and three novel phase II metabolites (glucuronic acid conjugates of ketobemidone, norketobemidone and hydroxymethoxyketobemidone) were identified in human urine. Further, norketobemidone and ketobemidone N-oxide were identified in rat microdialysate from brain after regional distribution of ketobemidone in striatum. This indicates that the brain itself has the possibility to metabolize ketobemidone.

Synthetic ketobemidone metabolites were used for comparison of retention times and tandem MS spectra with the possible metabolites recovered from the biological samples. The conjugated metabolites were identified by accurate mass measurements and tandem MS spectra of the aglycones. The accuracy of the estimated masses was better than 2.1 ppm for two out of three conjugates in presence of internal standard.

On-line micro-SPE was successfully used for trapping and desalting of the microdialysates. The small SPE pre-column made it possible to inject approximately 100 times more sample on the analytical column compared to injection without pre-column. Selective trapping was demonstrated for the polar catechol amine metabolite, dihydroxyketobemidone, which forms covalent complexes with phenylboronic acid (PBA). A fluorinated silica type stationary phase was the only column out of several tested that was able to separate ketobemidone and all relevant phase I metabolites.

Liquid chromatography and mass spectrometry are independently valuable tools in the field of analytical pharmaceutical chemistry. The present study showed that the combination of LC-MS, with its excellent selectivity and sensitivity, offers an outstanding tool in the qualitative analysis of drugs and metabolites in biological fluids.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2007. 46 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 57
Analytical chemistry, LC-MS/MS, accurate mass measurement, drug metabolites, microdialysate, brain, fluorinated stationary phase, phenylboronic acid, retention mechanism, Analytisk kemi
urn:nbn:se:uu:diva-7909 (URN)978-91-554-6908-5 (ISBN)
Public defence
2007-06-01, B42, BMC, Husargatan 3, Uppsala, 13:15
Available from: 2007-05-10 Created: 2007-05-10Bibliographically approved

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