Validated capillary electrophoresis assay for the simultaneous enantioselective determination of propafenone and its major metabolites in biological samples

A robust, inexpensive, and fully validated CE method for the simultaneous determination of the enantiomers of propafenone (PPF), 5‐hydroxy‐propafenone (5OH‐PPF) and N‐despropyl‐propafenone (NOR‐PPF) in serum and in in vitro media is described. It is based upon liquid–liquid extraction at alkaline pH...

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Bibliographic Details
Published in:Electrophoresis 2006-04, Vol.27 (8), p.1517-1525
Main Authors: Afshar, Minoo, Thormann, Wolfgang
Format: Article
Language:English
Subjects:
5-Hydroxy-propafenone
Anti-Arrhythmia Agents - analysis
Cytochrome P-450 CYP2D6 - metabolism
Cytochrome P-450 CYP3A
Cytochrome P-450 Enzyme System - metabolism
Electrophoresis, Capillary - methods
Humans
In vitro metabolism
Microsomes, Liver - metabolism
N-Despropyl-propafenone
Propafenone - analogs & derivatives
Propafenone - analysis
Propafenone - blood
Propafenone - metabolism
Reproducibility of Results
Serum
Stereoisomerism
Stereoselective metabolism
Sulfated β-cyclodextrin
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Summary:A robust, inexpensive, and fully validated CE method for the simultaneous determination of the enantiomers of propafenone (PPF), 5‐hydroxy‐propafenone (5OH‐PPF) and N‐despropyl‐propafenone (NOR‐PPF) in serum and in in vitro media is described. It is based upon liquid–liquid extraction at alkaline pH followed by analysis of the reconstituted extract by CE in presence of a pH 2.0 running buffer composed of 100 mM sodium phosphate, 19% methanol, and 0.6% highly sulfated β‐CD. For each compound, the S‐enantiomers are shown to migrate ahead of their antipodes, and the overall run time is about 30 min. Enantiomer levels between 25 and 1000 ng/mL provide linear calibration graphs, and the LOD for all enantiomers is between 10 and 12 ng/mL. The assay is shown to be suitable for the determination of the enantiomers of PPF and its metabolites in in vitro incubations comprising human liver microsomes or single CYP450 enzymes (SUPERSOMES). Incubations with CYP2D6 SUPERSOMES revealed, for the first time, the simultaneous formation of the enantiomers of 5OH‐PPF and NOR‐PPF with that enzyme. CE data can be used for the evaluation of the enzymatic N‐dealkylation and hydroxylation rates.
ISSN:0173-0835
1522-2683
DOI:10.1002/elps.200500663