Development of a high throughput 96-well plate sample preparation method for the determination of trileptal (oxcarbazepine) and its metabolites in human plasma.
A high throughput preparation method for the determination of trileptal (oxcarbazepine, OXC) and its mono (MHD) and dihydroxy (DHD) metabolites in human plasma, using 96-well plate technology, has been developed and validated according to international regulatory requirements. Preparation of plasma samples (50 microl) containing the compounds to be analysed involved solid-phase extraction (SPE) on Empore C18 96-well SPE plates. Eluates from the plate were injected onto a reversed-phase column (Hypersil C18,3 microm) with UV detection at 210 nm. Detector response was linear over the ranges 0.2-10, 0.1-200 and 0.1-20 micromol/l, for OXC, MHD and DHD, respectively, with relative standard deviations from 1 to 10% and mean accuracies within 4% of the nominal values (number of standard curves=3 in duplicate). The limits of quantitation were 0.2, 0.1 and 0.1 micromol/l, respectively. The overall mean accuracies ranged from 96 to 106% and precision was in the range 4 to 11%. Cross validation indicated no significant difference between plasma concentrations obtained using the 96-well method and the previous method using a traditional SPE method with a 50 mg C18 cartridge. About a threefold increase in sample throughput and a twofold decrease of plasma volume required for the assays, were the main advantages obtained from the previous method. The method was applied for the determination of 3000 plasma samples from clinical studies.
Souppart
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Drug Metabolism and Pharmacokinetics, Novartis Pharma SA, Rueil-Malmaison, France.
Decherf
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Humbert
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Maurer
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D000927:Anticonvulsants / Q000097:blood*; D002220:Carbamazepine / Q000031:analogs & derivatives* / Q000097:blood*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D000078330:Oxcarbazepine; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet
D000927:Anticonvulsants; D002220:Carbamazepine; D000078330:Oxcarbazepine