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A liquid chromatographic assay for the determination of CGP 61755 (I) in plasma and urine is described. A similar method for CGP 53437, another HIV-1 protease inhibitor, has been developed and reported previously. After a deproteinization step, a liquid-liquid extraction is performed. Compound I and the internal standard CGP 55749 (II) are hydrolyzed and the primary amine group derivatized using fluorescamine. Chromatography is achieved by isocratic elution with a mobile phase of 30 mM borax buffer (pH 9)-acetonitrile (58:42, v/v). The derivatives of the compounds I and II fluoresce at 480 nm, on excitation at 395 nm and the retention times under these conditions were approximately 6 and 8 min, respectively. The limit of quantitation (LOQ) which is the lowest concentration of the analyte that can be measured with a coefficient of variation and a deviation from theory of less than 20%, was 15 ng/ml plasma and 20 ng/ml urine. The analyte is stable for at least four months in human plasma and sixteen months in dog plasma samples. Different human plasma sources and three different species (rat, rabbit and dog) were tested and no interference between analyte and plasma constituents was observed.

A rapid and selective high-performance liquid chromatographic assay for determination of a new antimalarial drug (benflumetol, BFL) is described. After extraction with hexane-diethyl ether (70:30, v/v) from plasma, BFL was analysed using a C18 Partisil 10 ODS-3 reversed-phase stainless steel column and a mobile phase of acetonitrile-0.1 M ammonium acetate (90:10, v/v) adjusted to pH 4.9 with ultraviolet detection at 335 nm. The mean recovery of BFL over a concentration range of 50-400 ng/ml was 96.8 +/- 5.2%. The within-day and day-to-day coefficients of variation were 1.8-4.0 and 1.8-4.2%, respectively. The minimum detectable concentration in plasma for BFL was 5 ng/ml with a C.V. of less than 10%. This method was found to be suitable for clinical pharmacokinetic studies.

A specific and sensitive liquid chromatographic assay for the determination of 4-amidino-1-indanone-2'-amidinohydrazone (CGP 48 664, I) and a potential metabolite, 2-(4-carbamoyl-2,3-dihydro-1H-inden-1-yliden) hydrazine carboximidamide (CGP 53 391, II), in human and animal plasma was developed. CGP 51 467, a structural analog, was added to the plasma samples (up to 1 ml) as an internal standard. After mixing, the samples were processed automatically using an ASPEC solid-phase extraction system. The final extracts were chromatographed on a 5 microns Purospher RP-18 HPLC column. Chromatography was performed using a gradient system and UV detection. The described HPLC method is suitable for specific and quantitative measurement of concentrations of I, as well as its potential metabolite II down to 5-10 ng/ml in human and animal (dog, rat) plasma with acceptable reproducibility and accuracy.

A specific and sensitive liquid chromatographic assay for CGP 53,437 (I), a potent HIV protease inhibitor, is described. The method is based on a deproteinization step, followed by a liquid-liquid extraction with diisopropyl ether. Then a deprotection step of the primary amine and derivatization using fluorescamine is performed. Chromatography is achieved by isocratic elution with a mobile phase of 63 mM borax buffer (pH 9)-acetonitrile (58:42, v/v). The flow-rate of the mobile phase is 1 ml/min. The derivatives of compound I and its internal standard CGP 54,451, II, fluoresce at 480 nm on excitation at 395 nm. The limit of quantitation which is the lowest concentration of the analyte that can be measured with a coefficient of variation and a deviation from theory of less than 20%, was 5 nmol/l plasma. The analyte is stable for at least seven months in spiked human plasma samples. It is also stable after freezing and thawing cycles. Different human plasma sources and plasma samples from three different species (dog, marmoset, and rat) were tested and no interferences from plasma constituents was observed.

An enantioselective liquid chromatographic assay for the simultaneous determination of the S-(+) and R-(-) enantiomers of the monohydroxylated metabolite of oxcarbazepine in human plasma is described. The metabolite is the active principle. The method is based on the extraction of plasma with diethyl ether-dichloromethane (2:1, v/v), separation of the organic phase, evaporation of the solvent and dissolution of the residue in the mobile phase. The two enantiomers were resolved on a Chiralcel OD (250 mm x 4.6 mm I.D.) high-performance liquid chromatographic column. The separation was achieved by isocratic elution with n-hexane-2-propanol (77:23, v/v). The flow-rate of the mobile phase was 1.0 ml/min and the two enantiomers were detected by ultraviolet absorbance at 210 nm. The analytical method is suitable for the quantitative and simultaneous determination of the two enantiomers in plasma at concentrations down to 0.4 mumol/l after administration of oxcarbazepine.