title: Column-switching high-performance liquid chromatographic analysis of carbamazepine and its principal metabolite in human plasma with direct sample injection using an alkyl-diol silica (ADS) precolumn.
abstract: In this paper, the on-line coupling of solid-phase extraction, based on a restricted-access support with high-performance reverse phase chromatography for the analysis of carbamazepine (CBZ) and carbamazepine-10,11-epoxide (CBZ-E) in human plasma samples is described. A precolumn packed with 25 mum C(18) alkyl-diol support is used for direct plasma injection. Using column-switching techniques, the analytes were enriched on the precolumn by a 5 mM phosphate buffer (pH 7) with 2% of methanol solution at a flow-rate of 0.8 ml min(-1), while proteins and endogenous hydrophilic substances in plasma were washed off to waste. The enriched analytes were then back-flushed onto the analytical C(18) column, separated by a mixture of 10 mM phosphate buffer (pH 7) acetonitrile (70:30 v/v) solution at a flow-rate of 1.0 ml min(-1) and detected by the ultraviolet absorbance set at 212 and 285 nm and without transfer loss. Linear calibration graphs were obtained for sample injection volumes of 50 (0.2-4.0 of mug of CBZ ml(-1) and 0.1-5.0 mug of CBZ-E ml(-1), respectively), and 20 mul (5.0-20.0 mug of CBZ ml(-1)); in either case the r-value was >0.9963. Recoveries from spiked plasma samples were quantitative for both analytes and the coefficients of variation were below 3.83%. The lowest samples concentrations that can be quantified with acceptable accuracy and precision was 0.2 mug CBZ ml(-1) and 0.1 mug CBZ-E ml(-1) when a sample volume of 50 mul was injected. Concentrations of 0.08 and 0.05 mug ml(-1) of CBZ and CBZ-E were considered the limit of detection for a signal-to-noise ratio of 3. Furthermore, the developed column-switching method was successfully applied to the determination of CBZ and CBZ-E in plasma samples of patients submitted to CBZ therapy.
authors.affiliation: IVAIQUIM (Venezuelan Andean Institute for Chemical Research), Faculty of Science, University of Los Andes, Ipostel La Hechicera, PO Box 3, Mérida 5101-A, Venezuela.
authors.forename: M R
authors.initials: MR
authors.lastname: Brunetto
authors.forename: M A
authors.initials: MA
authors.lastname: Obando
authors.forename: A
authors.initials: A
authors.lastname: Fernández
authors.forename: M
authors.initials: M
authors.lastname: Gallignani
authors.forename: J L
authors.initials: JL
authors.lastname: Burguera
authors.forename: M
authors.initials: M
authors.lastname: Burguera