Determination of retigabine and its acetyl metabolite in biological matrices by on-line solid-phase extraction (column switching) liquid chromatography with tandem mass spectrometry
Knebel, N.G.; Grieb, S.; Leisenheimer, S.; Locher, M.
Journal of Chromatography. B, Biomedical Sciences and Applications 748(1): 97-111
ISSN/ISBN: 1387-2273 PMID: 11092590 DOI: 10.1016/s0378-4347(00)00272-3
A HPLC assay with tandem mass spectrometric detection in the positive-ion atmospheric pressure chemical ionisation (APCI) mode for the sensitive determination of retigabine ((I), D-23129) and its acetyl metabolite ((II), ADW 21-360) in plasma was developed, utilising the structural analogue (D-10328), (III), as internal standard. Automated on-line solid-phase extraction of diluted plasma samples, based on 200-mul plasma aliquots, at pH 6.5, allowed a reliable quantification of retigabine and the acetyl metabolite down to 1 ng/ml. Injection of 500 mul of diluted plasma onto a C2 stationary phase-based column switching system in combination with a 75 mm X 4 mm reversed-phase analytical column at a flow-rate of 0.5 ml/min provided cycle times of 4 min per sample. The standard curves were linear from 1 to 1000 ng/ml using weighted linear regression analysis (1/chi2). The method is accurate (mean accuracy ltoreq+-10%), precise (RSD <+-15%) and sensitive, providing lower limits of quantification in plasma of 1 ng/ml for retigabine (I), and 2.5 ng/ml for the metabolite (II) with limits of detection of 0.5 ng/ml for both analytes. Up to 200 unknowns may be analysed each 24 h per analyst.