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In vitro formation of phase I and II metabolites of propranolol and determination of their structures using chemical derivatization and liquid chromatography tandem mass spectrometry
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
2009 (English)In: Journal of Mass Spectrometry, ISSN 1076-5174, E-ISSN 1096-9888, Vol. 44, no 5, 742-754 p.Article in journal (Refereed) Published
Abstract [en]

Derivatization with 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC)  has been successfully used as a tool to differentiate between aromatic and aliphatic O-glucuronides of hydroxypropranolol. The analyses were   performed with liquid chromatography-electrospray ionization-tandem  mass spectrometry (LC-ESI-MS/MS) with both a triple quadrupole and an   ion trap instrument. Hydroxylated forms of propranolol can be glucuronidated in aliphatic as well as aromatic positions. These isoforms are not distinguishable by tandem MS alone, as they both   initially lose 176 Da, i.e. monodehydrated glucuronic acid, giving back   the aglycone. Two in vitro systems were set up for the production of  propranolol metabolites. The obtained isomers of 4'-hydroxypropranolol   glucuronide were determined to correspond to one aliphatic and one aromatic form, using chemical derivatization with DMISC and LC-MSn. DMISC was shown to react with the secondary amine in the case where the   naphtol was occupied by the glucuronyl moiety, resulting in a different fragmentation pattern compared with that of the aliphatic glucuronide, where the naphtol group was accessible to derivatization.

Place, publisher, year, edition, pages
2009. Vol. 44, no 5, 742-754 p.
Keyword [en]
derivatization, 1, 2-dimethylimidazole-4-sulfonyl chloride (DMISC), LC-MS/MS, structural investigation, propranolol, glucuronides
National Category
Pharmaceutical Sciences
Identifiers
URN: urn:nbn:se:uu:diva-97128DOI: 10.1002/jms.1551ISI: 000266681600016OAI: oai:DiVA.org:uu-97128DiVA: diva2:171929
Available from: 2008-04-25 Created: 2008-04-25 Last updated: 2010-07-27Bibliographically approved
In thesis
1. Chemical Derivatization in Combination with Liquid Chromatography Tandem Mass Spectrometry for Detection and Structural Investigation of Glucuronides
Open this publication in new window or tab >>Chemical Derivatization in Combination with Liquid Chromatography Tandem Mass Spectrometry for Detection and Structural Investigation of Glucuronides
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis presents novel approaches for structural investigation of glucuronides using chemical derivatization in combination with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MSn).

Today, LC-ESI-MSn is the dominant technique for quantitative as well as qualitative analyses of metabolites, due to its high sensitivity and selectivity. However, for compounds without an easily ionizable group, e.g., steroids, the sensitivity is limited. In the work presented in this thesis, a derivatization procedure forming a basic oxime significantly increased the detection sensitivity for the altrenogest glucuronide.

Furthermore, in structural evaluations of glucuronides, the limitation of LC-MSn becomes evident due to the initial neutral loss of 176 u, i.e. monodehydrated glucuronic acid, which often makes it impossible to elucidate the structures of the conjugates. To solve this problem, the main part of the work described in this thesis was devoted to chemical derivatization as a means of facilitating the determination of the site of conjugation.

For the first time, the isomeric estriol glucuronides were evaluated using a combination of three reagents 2-chloro-1-methylpyridinium iodide (CMPI), 1-ethyl-3-(3-dimethyl- aminopropyl)-carbodiimide (EDC), and 2-picolylamine (PA). Interestingly, the derivatization gave a selective fragmentation pattern leading to differentiation of the isomers.

Another derivatization reagent, 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC), was also tested for the first time in structural investigations. The isomeric glucuronides of morphine, formoterol, and hydroxypropranolol were evaluated. They can all be conjugated in aliphatic as well as aromatic positions. DMISC was proven to be useful in two ways. Firstly, the morphine and formoterol glucuronides that contained a free phenol could be differentiated from those that were conjugated in the aromatic position based on different reactivity. Secondly, for the aromatic O-glucuronide of 4’-hydroxypropranolol, DMISC was proven to react with the amine. This product gave a different fragmentation pattern compared to the corresponding derivative of the aliphatic glucuronide.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2008. 75 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 72
Keyword
Pharmaceutical chemistry, Derivatization, mass spectrometry, glucuronide, drug metabolism, increased sensitivity, structural investigation, 2-chloro-1-methylpyridinium iodide (CMPI), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC), 2-picolylamine (PA), 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC), Farmaceutisk kemi
Identifiers
urn:nbn:se:uu:diva-8670 (URN)978-91-554-7178-1 (ISBN)
Public defence
2008-05-16, B42, BMC, Husargatan 3, Uppsala, 09:15
Opponent
Supervisors
Available from: 2008-04-25 Created: 2008-04-25Bibliographically approved

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