High-performance liquid chromatographic analysis of Peptide T in rabbit plasma with on-line column enrichment

The present paper describes the development of a simple and sensitive analytical method for quantification of Peptide T (PT) in rabbit plasma, using standard analytical equipment and on-line column enrichment, without prior extraction, clean-up or derivatization. The analytical procedure was found to be accurate, precise and linear. The accuracy was 100% (range 97–103%) and the mean precision was 8% (range 3–14%) for all (n=6) concentrations (0, 15, 50, 100 and 200 ng/ml). The total recovery was found to be approximately 80%, and it was found to be dependent upon the injection rate onto the extraction column. The correlation between added and found concentrations was 0.9982, and the limit of detection was estimated to be around 5 ng/ml. The method is therefore found to be suitable for bioavailability studies, involving Peptide T, in rabbits.     

High-performance liquid chromatographic determination of diclofenac in human plasma using automated column switching

A validated high-performance liquid chromatographic procedure employing ultraviolet detection for the analysis of diclofenac in human plasma is reported. The method is rapid and, coupled with column switching, leads to a sensitive, accurate and reproducible assay. The retention times of diclofenac and the internal standard (4′-methoxydiclofenac, CGP-4287) are 6.4 and 7.6 min, respectively. The peak height versus plasma concentration is linear over the range 5.0–2000 ng/ml with a detection limit below 2.5 ng/ml. The mean absolute recovery of diclofenac using the described assay is 96.5% (n = 24). The inter- and intra-day accuracy and precision are within 8.3% of the actual values for all concentrations investigated. Furthermore, this procedure is applied to assess the pharmacokinetics of a single 75-mg oral dose of diclofenac sodium.     

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 determination of finasteride in human plasma using direct injection with column switching

A fully automated column-switching high-performance liquid chromatographic (HPLC) method was developed for the quantification of finasteride [N-(1,1-dimethylethyl)-3-oxo-4-aza-5α-androst-1-ene-17β-carboxamide] in human plasma. Plasma samples were diluted with an equal volume of ethylene glycol-water (40:60, v/v), then the diluted sample (150 μl) was injected into the HPLC system without clean-up. The analyte was retained on a pretreatment column, whereas plasma proteins and other endogenous components were washed out to waste. The analyte was transferred to the analytical column in the heart-cut mode and then detected at 210 nm. A quantification limit of 1 ng/ml was attained. There was a linear relationship between peak height and drug concentration in plasma in the range 1–50 ng/ml. This method was validated and applied to the assay of plasma samples to characterize pharmacokinetic parameters in clinical studies.     

High-performance liquid column and thin-layer chromatographic determination of human serum glibenclamide at therapeutic levels

For glibenclamide bioavailability studies in serum, high-performance liquid column and thin-layer chromatographic methods were introduced. Both methods are specific, accurate and sensitive with detection limits of at least 5 ng of glibenclamide per ml of serum. Detection is performed in the ultraviolet at wavelengths of 200 nm for liquid chromatography or 300 nm for thin-layer chromatography.Serum levels determined by either method correlated well with those determined by an already existing radioimmunoassay. Some pharmacokinetic data were computed using a one-compartment open model.     

High-performance liquid chromatographic analysis of glycoamines in serum

This report describes the development of an HPLC-UV method for studies of glycoamines and glycoamine-like compounds in normal human serum and osteosarcoma patient serum as potential biological markers of cancer. The glycoamines, a newly recognized class of endogenous, low-molecular-mass biopolymers, are conjugates of amino acids and sugar units, containing 5 to 29 amino acid and 1 to 17 sugar units. After ultrafiltration of serum samples, reversed-phase HPLC separation with diode-array detection was used to obtain standard profiles of serum ultrafiltrates below M_r 10 000 in healthy subjects. These highly reproducible profiles utilized two-dimensional peak identification and were used to develop a statistical profile of the major glycoamine peaks in normal serum. This newly developed analytical method was subsequently used to address a key question: whether or not there is a single tumor-specific glycoamine or a family of tumor-specific glycoamines in cancer patient serum. Preliminary results suggest that this method can separate and detect glycoamines and glycoamine-like compounds in various types of cancer patient serum with a high degree of reproducibility on the basis of comparative two-dimensional identification of natural compounds and a panel of synthetic glycoamine analogs. Moreover, the method is useful for following the relative changes in the amount of a given glycoamine over an extended clinical time course. Initial results suggest that a glycoamine or glycoamine-like compound, GA-4.63, may have clinical utility in human osteosarcoma studies.     

High-performance liquid chromatographic determination of ambroxol in human plasma

Ambroxol has been determined in biological fluids using a rapid and sensitive high-performance liquid chromatographic method. The samples prepared from plasma by liquid—liquid extraction were analysed on reversed-phase silica gel by competing-ion chromatography with ultraviolet detection. The method was applied to the determination of ambroxol levels in twelve healthy volunteers after oral administration of 90 mg of ambroxol in tablets of Mucosolvan and Ambrosan.     

High-performance liquid chromatographic determination of tramadol in human plasma

Tramadol has been determined in human plasma samples using a sensitive high-performance liquid chromatographic method. The plasma samples were extracted with tert.-butylmethyl ether in one-step liquid-liquid extraction (recovery 86%) and analyses of the extracts were performed on reversed-phase silica gel using ion-pair chromatography (verapamil as an internal standard) and fluorescence detection. The method was applied to the determination of tramadol levels in twelve healthy volunteers after oral administration of 100 mg of tramadol in capsules of Protradon and Tramal.     

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 determination of beta-phenylethylamine in human plasma with fluorescence detection

A simple and highly sensitive method for the determination of beta-phenylethylamine in human plasma is investigated. The method employs high-performance liquid chromatography with fluorescence detection. beta-Phenylethylamine and p-methylbenzylamine (internal standard) in human plasma are isolated by cation-exchange chromatography on a Toyopak SP cartridge and then converted into the corresponding fluorescent derivatives with 3,4-dihydro-6,7-dimethoxy-4-methyl-3-oxoquinoxaline-2-carbonyl chloride, a fluorescence derivatization reagent for amines. The derivatives are separated within 30 min on a reversed-phase column, TSK gel ODS-120T, with isocratic elution, and detected fluorometrically. The detection limit of beta-phenylethylamine is 0.3 pmol/ml in plasma (S/N = 3).     

High-performance liquid chromatographic determination of phenylephrine in human serum with coulometric detection

A high-performance liquid chromatographic method for the determination of phenylephrine (PE) in human serum using coulometric detection is described. PE and internal standard, orciprenaline, were extracted from serum by solid-phase extraction and separation achieved on a coupled column system consisting of two C₁₈ cartridge columns (250×4.6 mm I.D. coupled to a shorter 50×4.6 mm I.D. column) using a mobile phase of methanol-50 mM phosphate buffer (pH 3.2; 10:90) at 36°C. Dual electrode coulometric detection was used in the “oxidative screen” mode. Calibration curves were linear over the range 0.3–4 ng/ml with a limit of quantification (LOQ) of 0.35 ng/ml. The method has a greater degree of sensitivity, precision and accuracy compared to previously published methods for PE and is suitable for use in pharmacokinetic and bioequivalence studies in humans.     

High-performance liquid chromatographic determination of levodropropizine in human plasma with fluorometric detection

The present describes a new high-performance liquid chromatographic method with fluorescence detection for the analysis of levodropropizine [S-(−)-3-(4-phenylpiperazin-1-yl)-propane-1,2-diol] (Levotuss), an anti-tussive drug, in human serum and plasma. A reversed-phase separation of levodropropizine was coupled with detection of the native fluorescence of the molecule, using excitation and emission wavelengths of 240 nm and 350 nm respectively. The analytical column was packed with spherical 5 μm poly(styrene-divinylbenzene) particles and the mobile phase was 0.1 M NaH₂PO₄ pH 3-methanol (70:30, v/v), containing 0.5% (v/v) tetrahydrofuran. For quantitation, p-methoxylevodropropizine was used as the internal standard. Samples of 200 μl of either serum or plasma were mixed with 200 μl of 0.1 M Na₂HPO₄ pH 8.9 and extracted with 5 ml of chloroform-2-propanol (9:1, v/v). The dried residue from the organic extract was redissolved with distilled water and directly injected into the chromatograph. The limit of detection for levodropropizine, in biological matrix, was about 1–2 ng/ml, at a signal-to-noise ratio of 3. The linearity was satisfactory over a range of concentrations from 3 to 1000 ng/ml (r² = 0.99910); within-day precision tested in the range 5–100 ng/ml as well as day-to-day reproducibility proved acceptable, with relative standard deviations better than 1% in most cases. Interferences from as many as 91 therapeutic or illicit drugs were excluded.     

High-performance liquid chromatographic determination of pipotiazine in human plasma and urine

A high-performance liquid chromatographic method has been developed for the determination of pipotiazine in human plasma and urine. After selective extraction, pipotiazine and the internal standard (7-methoxypipotiazine) are chromatographed on a column packed with Spherosil XOA 600 (5 μm) using a 7:3 (v/v) mixture of diisopropyl ether—isooctane (1:1, v/v) + 0.2% triethylamine and diisopropyl ether—methanol (1:1, v/v) + 0.2% triethylamine + 2.6% water. The eluted compounds are measured by fluorescence detection. The sensitivity of the method was established at 0.25 ng/ml pipotiazine in plasma and 2 ng/ml pipotiazine in urine (C.V. < 5%). The method has been successfully applied to a pharmacokinetic study following a single oral administration of 10 mg of pipotiazine.