Chiral high-performance liquid chromatographic analysis of antifungal SCH 56592 and evaluation of its chiral inversion in animals and humans

SCH 56592 is a novel triazole antifungal agent that is active both orally and intravenously in animal models of infection. This compound is in Phase II-III clinical trials for the treatment of systemic fungal infections. SCH 56592 is a single enantiomer with four stereogenic centers; therefore, it was necessary to evaluate the possible chiral inversion of this drug candidate in animals and humans. Thus, chiral high-performance liquid chromatographic (HPLC) methods have been developed to separate SCH 56592 from its diastereomers and to evaluate its chiral inversion in rats, dogs, cynomolgus monkeys, and humans. Chiral HPLC analysis involved the use of a Chiralcel OD column set at 39 degrees C with a mobile phase of hexane-ethanol-diethylamine and a fluorescence detector set at an excitation wavelength of 270 nm and an emission wavelength of 390 nm. Plasma or serum samples were subjected to solid phase extraction on a C(2) cartridge followed by HPLC analysis. The method was sensitive with a limit of quantitation of 0.1 microg/ml in dog serum. The linearity was satisfactory, as shown by correlations of >0.997 and by visual examination of the calibration curves. The precision and accuracy were satisfactory, as indicated by coefficients of variation (CV) ranging from 1.1 to 12.1% and bias values ranging from -11.0 to 9.0%. Chiral HPLC analysis indicated that SCH 56592 was not subjected to chiral inversion in rats, dogs, cynomolgus monkeys, and humans.     

High-performance liquid chromatographic analysis of the D4 receptor antagonist SCH 66712 in rat plasma

SCH 66712 is a potent and selective dopamine D₄ receptor antagonist. An HPLC method was developed for the analysis of SCH 66712 in the plasma of rats, a species used for safety evaluation of this compound. The method involved solid-phase extraction on an ethyl cartridge and HPLC separation on a reversed-phase C₈ column with quantitation using a fluorescence detector. The calibration curve was linear over a concentration range of 5–100 ng/ml. The limit of quantitation was 5 ng/ml, where the coefficient of variation (C.V.) was 2.9% and the bias was 6%. The precision of the method was satisfactory as indicated by an intra-day C.V. of ≤4% and an inter-day C.V. of ≤6%. The accuracy was also satisfactory as shown by an intra-day bias of ≤8% and an inter-day bias of ≤9%. The assay was shown to be sensitive, specific, accurate, precise, and reliable for use in pharmacokinetic or toxicokinetic studies.     

High-performance liquid chromatographic determination of the calcium channel blocker nimodipine in monkey plasma

A new high-performance liquid chromatographic (HPLC) assay was developed for the determination of nimodipine in monkey plasma. An ethyl acetate extraction procedure was employed with a reversed-phase HPLC separation for the analysis. Absolute recovery of nimodipine from plasma was over 95% with a lower limit of quantitation of 10 ng/ml. This method was applied to a preliminary pharmacokinetic study in which 0.25 mg/kg nimodipine was administered intravenously to three monkeys. Protein binding and stability of nimodipine in monkey plasma were also examined. The pharmacokinetic parameters of nimodipine in monkeys were similar to those obtained in humans and indicate that monkeys are an appropriate animal model for further pharmacokinetic investigations.     

High-performance liquid chromatographic analysis of nitroxoline in plasma and urine

A high-performance liquid chromatographic method has been developed for the determination of nitroxoline in 50-μl plasma and urine samples.A structural analogue of nitroxoline, 8-hydroxyquinoline, was added to the eluent in order to suppress peak asymmetry. Several parameters of the eluent were studied for the optimisation of the chromatographic system.Plasma concentration—time curves were constructed for three volunteers after they had received an oral dose of 100 mg of nitroxoline. Plasma half-life was about 1 h. Within 12 h, about 1% of the dose was excreted in the urine as free nitroxoline and about 30% as conjugated metabolite of the parent compound.     

High-performance liquid chromatographic analysis of isoxicam in human plasma and urine

A sensitive, selective, and rapid high-performance liquid chromatographic procedure was developed for the determination of isoxicam in human plasma and urine. Acidified plasma or urine were extracted with toluene. Portions of the organic extract were evaporated to dryness, the residue dissolved in tetrahydrofuran (plasma) or acetonitrile (urine) and chromatographed on a μBondapak C₁₈ column preceded by a 4–5 cm × 2 mm I.D. column packed with Corasil C₁₈. Quantitation was obtained by UV spectrometry at 320 nm. Linearity in plasma ranged from 0.2 to 10 μg/ml. Recoveries from plasma samples seeded with 1.8, 4 and 8 μg/ml isoxicam were 1.86 ± 0.077, 4.10 ± 0.107 and 8.43 ± 0.154 μg/ml with relative standard deviations of 3.3%, 2.5% and 5.4%, respectively. The linearity in urine ranged from 0.125 to 2 μg/ml. The precision of the method was 3.3–9.0% relative standard deviation over the linear range.     

High-performance liquid chromatographic analysis of mibefradil in dog plasma and urine

The objective of the study was to develop a sensitive and specific assay for studying the pharmacokinetics of a novel calcium antagonist, a benzimidazolyl-substituted tetraline derivative, mibefradil (I) in the dog. The assay involves liquid-liquid extraction of a biological sample, reversed-phase HPLC separation and fluorescence detection (λ_ex = 270 nm and λ_em = 300 nm) of a sample components. Each sample was eluted with a mobile phase pumping at a flow-rate of 2 ml/min. The mobile phase composition was a mixture of acetonitrile and aqueous solution (38:62, v/v). The aqueous solution contains 0.0393 M KH₂PO₄ and 0.0082 M Na-pentanesulphonic acid. The retention times were 10.7 min for I, and 12.2 min for internal standard Ro 40–6792. Calibration curves with concentrations of I ranging from 10 to 500 ng/ml were linear (r² > 0.99). The detection limit for I was 0.5 ng/ml when 0.5 ml of plasma or urine was used. Intra- and inter-day accuracy and precision were within 10%. The assay was successfully applied to the pharmacokinetic studies of I in dogs.     

High-performance liquid chromatographic analysis of dipyridamole in plasma and whole blood

A rapid, sensitive, and specific high-performance liquid chromatographic method is described for the quantitative analysis of dipyridamole in plasma and whole blood. The method involves a single extraction of an alkalinized sample with diethyl ether followed by evaporation of the organic solvent and ion-pair chromatography using fluorescence detection. The lower limit of sensitivity for dipyridamole is 1 ng/ml. Concentrations of dipyridamole between 1 and 500 ng per sample are measured with an average coefficient of variation of 4.5% in plasma and 7.4% in whole blood.     

High-performance liquid chromatographic determination of fluconazole in plasma

A high-performance liquid chromatographic method with ultraviolet absorbance detection at 260 nm was developed for the analysis of fluconazole in plasma. The method involves sample clean-up by liquid-liquid extraction. The proposed technique is reproducible, selective, reliable and sensitive. Calibration standards were prepared in the range 1.25-20 mg/l. The limit of quantitation was 0.4 mg/l. The coefficients of variation were 5% between measurements of a single extract injected in duplicate, and 7% between two extractions of spiked samples at the same concentrations. The separation between fluconazole and endogenous substances was satisfactory. This method was designed in order to minimise the risk of interference from substances that could be co-administered to critically ill patients undergoing hemodiafiltration. With a run time below 5 min, the present method is rapid and easy to use for later clinical studies, as well as for routine monitoring.     

High-performance liquid chromatographic determination of ganciclovir in plasma

A rapid, selective and sensitive isocratic reversed-phase high-performance liquid chromatographic method for the determination of ganciclovir in plasma samples was developed. This method, which was applied to the analysis of plasma ganciclovir from heart transplant patients under ganciclovir therapy for cytomegalovirus infections, represents a suitable analytical tool for drug monitoring and pharmacokinetic investigations.     

High-performance liquid chromatographic determination of pentamidine in plasma

This report describes the analysis of pentamidine by isocratic reversed-phase high-performance liquid chromatography (HPLC) using a commercially available compound (melphalan) as the external standard. Previously described assays use ion-pairing HPLC, an internal standard (hexamidine) that is not readily available, and require a relatively large sample size. In the present assay, pentamidine was extracted from plasma using solid-phase extraction and was analyzed using a C₁₈ column and a mobile phase containing 18% acetonitrile, 2% methanol, 0.2 M ammonium acetate and 0.5% triethylamine. The identity of the eluting peaks was verified using a diode array detector. The extraction yield of pentamidine was 82%. The limit of detection was 8.6 ng/ml with a sample size of 100 μl. The inter-day and intra-day coefficients of variation ranged between 0.3% and 10% with an average of 5%. This method was applied to study the pharmacokinetics of pentamidine in rodents.     

High-performance liquid chromatographic method for the estimation of the novel investigational anti-cancer agent SR271425 and its metabolites in mouse plasma

A simple and reliable HPLC method was developed for the estimation of a new anti-cancer agent that belongs to the thioxanthone class, SR271425 in mouse plasma. SR271425, it’s metabolites and internal standard (SR233377) were separated from plasma by liquid–liquid extraction using dichloromethane after quenching the plasma proteins with acetonitrile. Chromatography was performed on a reversed-phase C₁₈ column using methanol–10 mM phosphate buffer, pH 3.5 (45:55) as mobile phase at a flow-rate of 0.8 ml/min for first 10 min and 1.4 ml/min for the next 15 min with UV–Vis detection at 264 nm and SR233377 as internal standard. The retention times of SR271425 and internal standard were 18.6 and 14.8 min, respectively. The limit of detection was 40 ng/ml and the limit of quantification was 78 ng/ml. This method was also able to detect the three metabolites of SR271425. The intra- and inter-day relative standard deviations were less than 13% at all concentrations. This analytical method was precise and reproducible for pharmacokinetics and metabolism studies of the drug in mice. SR271425 is proceeding to phase I clinical trials in 2001.