Analysis of 5-fluorouracil in plasma and urine by high-performance liquid chromatography.

A relatively simple, sensitive and rapid high-performance liquid chromatographic method is described for measuring the anticancer drug 5-fluorouracil (5-FU) in human plasma and urine. The procedure includes liquid-liquid extraction using ethyl acetate-methanol (95:5) and preparative column chromatography to separate 5-FU from constituents normally occurring in these biological samples. The columns contained a specially modified form of diatomaceous earth, which requires no pre-conditioning washes. Reversed-phase high-performance liquid chromatography was performed on a C18 column (70 mm x 4.6 mm I.D.) with a mobile phase of water-methanol (95:5) and ultraviolet detection (268 nm). The overall recovery from plasma and urine was 91 and 94%, respectively, at the concentration of 50 ng/ml. The determination limit of the assay for 5-FU was 10 ng/ml of plasma and urine. Concentrations of 5-FU between 10 and 500 ng/ml were measured in plasma and urine with a relative standard deviation of 6.8%. In order to evaluate the procedure, plasma and urine samples from three patients treated with 5-FU by continuous intravenous perfusion, were investigated.     

Determination of pirlindole in plasma and urine by high-performance liquid chromatography

A high-performance liquid chromatographic method is described for the analysis of pirlindole [2,3,3a,4,5,6-hexahydro-8-methyl-1H-pyrazino(3,2,1-jk)carbazole hydrochloride], a new antidepressive drug. The drug was extracted from plasma into dichloromethane, and the analysis was carried out on a reversed-phase column, the effluent being monitored by fluorescence detection. The method is selective and sensitive (limit of detection 1-2 ng/ml plasma). Urine analysis was done by direct injection of the diluted sample. The method was applied to the analysis of plasma and urine samples of eight healthy male volunteers who received a 75-mg oral dose of a tablet formulation of pirlindole. The method was also applied to a study in three beagle dogs which received pirlindole (1 mg/kg) by infusion (0.1 mg/kg/min) and orally (10 mg/kg) to estimate the absolute bioavailability of the drug.     

Determination of amdinocillin in plasma and urine by high-performance liquid chromatography

A rapid, sensitive and specific high-performance liquid chromatographic (HPLC) assay was developed for the determination of amdinocillin (formerly mecillinam) in human plasma and urine. The assay is performed by direct injection of a plasma protein-free supernatant or a dilution of urine. A 10 micrometer muBondapak phenyl column with an eluting solvent of water--methanol--1 M phosphate buffer, pH 7 (70:30:0.5) was used, with UV detection of the effluent at 220 nm. Azidocillin potassium salt [potassium-6-(D-(-)-alpha-azidophenyacetamido)-penicillanate] was used as the internal standard and quantitation was based on peak height ratio of amdinocillin to that of the internal standard. The assay has a recovery of 74.4 +/- 6.3% (S.D.) in the concentration ranges of 0.1-20 microgram per 0.2 ml of plasma with a limit of detection equivalent to 0.5 microgram/ml plasma. The urine assay was validated over a concentration range of 0.025-5 mg/ml of urine, and has a limit of detection of 0.025 mg/ml (25 microgram/ml) using a 0.1-ml urine specimen per assay. The assay was applied to the determination of plasma and urine concentrations of amdinocillin following intravenous administration of a 10 mg/kg dose of amdinocillin to two human subjects. The HPLC and microbiological assays were shown to correlate well for these samples.     

Determination of picotamide in human plasma and urine by high-performance liquid chromatography.

A high-performance liquid chromatographic method for the determination of picotamide in human plasma and urine is described. After addition of an internal standard (bamifylline), the plasma and urine samples were subjected to liquid-liquid extraction and clean-up procedures. The final extracts were evaporated to dryness and the resulting residues were reconstituted in 100 microliters of methanol-water (50:50, v/v) and chromatographed on a LiChrosorb RP-SELECT B reversed-phase column coupled to an ultraviolet detector monitored at 230 nm. Chromatographic analysis takes about 10 min per sample. The assay was linear over a wide range and has a limit of detection of 0.005 and 0.1 micrograms/ml in plasma and urine, respectively. It was selective for picotamide, accurate and robust and thus suitable for routine assays after therapeutic doses of picotamide.     

Determination of cibenzoline in plasma and urine by high-performance liquid chromatography

A rapid, sensitive and selective high-performance liquid chromatographic (HPLC) assay was developed for the determination of cibenzoline (CipralanTM) in human plasma and urine. The assay involves the extraction of the compound into benzene from plasma or urine buffered to pH 11 and HPLC analysis of the residue dissolved in acetonitrile-phosphate buffer (0.015 mol/l, pH 6.0) (80:20). A 10-microns ion-exchange (sulfonate) column was used with acetonitrile-phosphate buffer (0.015 mol/l, pH 6.0) (80:20) as the mobile phase. UV detection at 214 nm was used for quantitation with the di-p-methyl analogue of cibenzoline as the internal standard. The recovery of cibenzoline in the assay ranged from 60 to 70% and was validated in human plasma and urine in the concentration range of 10-1000 ng/ml and 50-5000 ng/ml, respectively. A normal-phase HPLC assay was developed for the determination of the imidazole metabolite of cibenzoline. The assays were applied to the determination of plasma and urine concentrations of cibenzoline and trace amounts of its imidazole metabolite following oral administration of cibenzoline succinate to two human subjects.     

Quantitative determination of the loop diuretic bumetanide in urine and pharmaceuticals by high-performance liquid chromatography with amperometric detection.

A high-performance liquid chromatographic method with amperometric detection has been developed for the determination of the diuretic bumetanide using a microBondapak C18 column. The mobile phase consists of a 50:50 acetonitrile-water mixture containing 5mM KH2PO4-K2HPO4 (pH 4.0). The compound is monitored at +1350 mV with an amperometric detector equipped with a glassy carbon working electrode. A liquid-liquid or solid-liquid extraction is done prior to chromatographic analysis in order to avoid the interferences found in the urine matrix. The percentages of recovery obtained are 71%+/-1% for liquid-liquid extraction and 84.2%+/-0.7% for solid-liquid extraction. The method developed has a linear concentration range from 50 to 499 ng/mL with a reproducibility in terms of relative standard deviation of 1.73% and 3.85% for a concentration level of 70 ng/mL and 237 ng/mL, respectively, and a detection limit of 0.25 ng/mL (3:1 signal-to-noise ratio). The method is applied to the determination of bumetanide in pharmaceutical formulations and urine obtained from hypertensive patients and healthy volunteers after the ingestion of a therapeutic dose of Fordiuran (1 mg bumetanide).     

Determination of trichlormethiazide in human plasma and urine by high-performance liquid chromatography

Summary This paper describes a high-performance liquid chromatographic (HPLC) assay method for the determination of trichlormethiazide (TCM) in human plasma and urine. After extraction and separation on an ODS column TCM from plasma was detected by oxidation in an electrochemical detector (ECD) by a porous graphite electrode. The sensitivity was better than HPLC with UV detection, enabling the determination of 2 ng ml^–1 TCM in human plasma. This method also allows determination of TCM at higher concentrations by exchanging the UV for the electrochemical detector. To study the pharmacokinetics, TCM in plasma and urine was assayed with coefficients of variation in the range 2–3%. The method has the advantages of high sensitivity for plasma assay and high precision with a simple procedure for both plasma and urine samples. Small samples of 0.5 ml plasma per assay also reduced the total volume of plasma needed.     

Determination of piquindone in canine plasma and urine by high-performance liquid chromatography

This report describes a rapid, sensitive and selective method for the determination of piquindone in canine plasma and piquindone and the N-demethyl metabolite of piquindone in canine urine, utilizing normal-phase high-performance liquid chromatography (HPLC) with isocratic elution at ambient temperature and monitoring the ultraviolet absorbance of the eluent at 254 nm. The trimethyl analogue of piquindone is used as the internal standard in the HPLC assay of plasma. The assay was applied to the measurement of concentrations of piquindone in the plasma and urine of a dog following single intravenous and oral administration of 5 mg/kg doses of piquindone hydrochloride dihydrate.     

Determination of 2-mercaptopropionylglycine in plasma and urine by high-performance liquid chromatography

Methods for quantitative analysis of total and non-protein-bound 2-mercaptopropionylglycine (2-MPG) in plasma, and total 2-MPG in urine, have been developed. By reduction of urine, plasma or deproteinized plasma samples with tributylphosphine, 2-MPG is liberated from its disulphides, and after clean-up of the sample, 2-MPG is derivatized with N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM). The 2-MPG-DACM derivative is then quantified by high-performance liquid chromatography (HPLC) with fluorimetric detection. Both ion-suppression and ion-pair HPLC gave satisfactory chromatograms. The precision of the methods was satisfactory (coefficient of variation 3.1-5.8%), analytical recovery was quantitative (85-99%) and the two HPLC techniques were well correlated (r = 0.99). Five healthy subjects receiving 500 mg of 2-MPG showed maximal total plasma concentration of 13.8-26.9 mumol/l at 3-5 h after intake, and their non-protein-bound 2-MPG was, at the same time, 62-77% of the total 2-MPG. The urinary excretion was 27.8 +/- 3.8% (mean +/- S.D.) of the given dose, most of it excreted within 12 h after intake.     

Determination of cefotaxime and desacetylcefotaxime in plasma and urine by high-performance liquid chromatography

A high-performance liquid chromatographic method is described for the analysis of the anti-bacterial agent cefotaxime and desacetylcefotaxime in physiological fluids. Plasma or serum samples were mixed with chloroform--acetone to remove proteins and most lipid material. The aqueous phase was then freeze-dried, reconstituted in mobile phase and chromatographed on a reversed-phase column using UV detection at 262 nm. Urine was analysed directly after centrifugation to remove particulate matter. The detection limit was 0.5--1.0 micrograms/ml for serum and 5 micrograms/ml for urine. The method has been applied to the analyses of cefotaxime and desacetylcefotaxime in plasma, serum, urine, cerebrospinal fluid, saliva, and pus from infected wound secretions. Two additional metabolites, which are lactones in which the beta-lactam ring has been opened, could be separated by this method.     

Determination of cabergoline in plasma and urine by high-performance liquid chromatography with electrochemical detection.

A sensitive and selective high-performance liquid chromatographic method for the determination of cabergoline in plasma and urine has been developed. After buffering plasma and urine samples, cabergoline was extracted with a methylene chloride-isooctane mixture, back-extracted into 0.1 M phosphoric acid, then analysed by reversed-phase high-performance liquid chromatography. Quantitation was achieved by electrochemical detection of the eluate. The linearity, precision and accuracy of the method were evaluated. No interference from the biological matrices (human plasma and urine) was observed. The assay was still inadequate in terms of sensitivity for the quantitation of cabergoline plasma concentrations after a single oral dose of 1 mg of the drug to humans, but was successfully used in the determination of the urinary excretion of the drug.