[Solid-phase extraction and RP-HPLC screening procedure for diuretics, probenecid, caffeine and pemoline in urine].

A solid-phase extraction and reversed phase high performance liquid chromatographic method (RP-HPLC) was developed for the rapid determination of 13 diuretics (belonging to five different pharmacological groups), probenecid, caffeine and pemoline in urine. Two ml urine sample was first adsorbed on a XAD-2 column, then eluted with ether-ethyl acetate (1:1). The eluate was evaporated to dryness and reconstituted in methanol. The methanolic solution was injected into a HP LiChrosorb RP-18 column, using phosphate buffer (pH 3) and acetonitrile as the mobile phase and monitored at 216 nm, 230 nm, and 275 nm on a diode array ultraviolet detector. The extraction recoveries of 16 drugs were above 75%. The limits of detection ranged from 0.3-3.0 micrograms/ml of urine. All drugs were separately administered to healthy volunteers, positive urine samples were collected, and urinary excretion-time curves of some drugs were reported.

Determination of pemoline in plasma, urine and tissues by reversed-phase high-performance liquid chromatography.

A simple and selective high-performance liquid chromatographic method with ultraviolet detection at 215 nm for the determination for pemoline in rat plasma, urine and tissues is described. Pemoline in the samples was extracted with methylene chloride at pH 10 and the organic phase was evaporated after adding 5-methyl-5-phenylhydantoin used as an internal standard. Pemoline and the internal standard were separated on a Kaseisorb LC C8-60-5 reversed-phase column. The limits of determination of pemoline in 0.1-0.2 ml of plasma, urine and tissue homogenates were 2, 100 and 20 ng, respectively. The method should be useful for studies of the pharmacokinetics and distribution of pemoline in small animals.

Screening of amphetamines by gradient microbore liquid chromatography and pre-column technology.

Amphetamine-type drugs with a wide polarity range have been screened in both human and horse urine using on-line pre-concentration on pre-columns packed with hydrophobic and cation-exchange sorbents in series and gradient microbore high-performance liquid chromatography. The underivatized amphetamines were identified by UV detection at 210 nm. The method has potential for the automated liquid chromatographic screening of amphetamines in urine, e.g., for doping control.

Determination of pemoline in plasma, plasma water, mixed saliva, and urine by high-performance liquid chromatography.

A new determination method of pemoline in plasma, plasma water, mixed saliva, and urine using high-performance liquid chromatography was developed. The detection limit of pemoline using 0.2 ml of the sample was 0.02 microgram/ml in plasma, plasma water, saliva, and urine. The recoveries of pemoline added to plasma, saliva, and urine (each 6 micrograms/ml) were more than 98%. The coefficients of variation of within-run and between-run precisions for 5 concentrations of pemoline in plasma were less than 5 and 6%, respectively. The elimination half-lives of pemoline obtained from the plasma concentration-time curves after a single oral administration of pemoline in two subjects were 6.7 and 10.3 h. Mean values of the ratio between saliva and plasma total concentration in two subjects were 0.55 and 0.64, and mean values of plasma protein binding percent were 35.7 and 25.2%, respectively. The saliva concentrations were in proportion to the plasma unbound pemoline concentrations at simultaneous samplings, and the ratios between saliva and plasma unbound concentration were about 0.9 in two subjects. Usefulness of saliva in the estimation of plasma protein-unbound pemoline concentration was noted.

Determination of the psychostimulants pemoline, fenozolone and thozalinone in human urine by gas chromatography/mass spectrometry and thin layer chromatography.

Detection of the psychostimulants pemoline (Tradon), fenozolone (Ordinator) and thozalinone (Stimsen) in biological samples (urine) presents certain experimental difficulties. These substances were therefore converted by acid hydrolysis to their common hydrolysis product 5-phenyl-2,4-oxazolidinedione, which can be detected with high sensitivity and selectivity by thin layer chromatography. After silylation with N-methyl-N-trimethylsilyltrifluoroacetamide/trimethylchlorosilane quantitative results can be obtained from the same extract by the proposed GC/MS-method. After a single dose of 30 mg pemoline, levels ranging from one to four mg/l urine were observed.

Gas/liquid chromatographic analysis of pemoline in biological fluids using electron capture detection.

An analytical gas/liquid chromatographic (GLC) protocol is described for the quantitation of pemoline in biological fluids of the horse. Plasma samples containing known quantities of pemoline and its analog as an internal standard (IS) were deproteinized with 5-sulfosalicylic acid, heated at 80 degree C, and centrifuged. 5-Phenyl-2,4-oxazolidinedione, the hydrolytic product of pemoline in acid medium, was extracted with dichloromethane (DCM). The organic layer was in turn re-extracted with 1% NaHCO3. The aqueous layer was acidified with HCI, and re-extracted with DCM, which was evaporated to dryness. The resulting residue was methylated with ethereal diazomethane. The methylated dione was dissolved in benzene, and analyzed with a gas/liquid chromatograph equipped with a tritiated scandium foil (Sc3H) electron capture detector (ECD). Urine samples to which pemoline was added were hydrolyzed with hydrochloric acid and carried through the analytical procedure in a manner similar to the plasma, except that the urine was further washed with lead acetate solution prior to the sodium bicarbonate extraction. The lower limits of detection were found to be 0.05 microgram/ml for plasma and 0.1 microgram/ml for urine when 1.0 ml of sample was used. This procedure can be used for pemoline detection control in international sporting events.

Pharmacokinetics of pemoline in plasma, saliva and urine following oral administration.

1 Pemoline concentrations were measured in plasma and saliva following a single oral dose (37.5 or 50.0 mg) to healthy volunteers. In addition urinary excretion rates and cumulative urinary excretion of the parent compound and its oxazolidinedione metabolite were determined. 2 The plasma curves exhibited a mean elimination half-live of 11.0 +/- 1.2 h (n=4). Peak levels were reached at 2.7 +/- 0.6 h (n=4). The saliva concentrations were about 50% lower than the corresponding plasma concentrations during the elimination phase. During the absorption phase irregularities in the saliva to plasma concentration ratios were observed. 3 In urine 47.0 +/- 8.4% of the dose (n=6) administered was excreted as unchanged drug and only 3.7 +/- 0.8% (n=3) as the oxazolidinedione metabolite. Urinary half-lives were slightly shorter than the corresponding plasma half-lives.

Sensitive GLC assay for pemoline in biological fluids using nitrogen-specific detection.

Extractive alkylation was used to determine intact pemoline in serum and urine. Pemoline was extracted into methylene chloride as an ion-pair with tetrapentylammonium hydroxide under alkaline conditions. Evaporation of the solvent at 70 degrees in the presence of methyl iodide yielded the N,N-dimethylpemoline derivative. GLC analysis was performed on a 5% FFAP column with nitrogen-specific detection. Sensitivity was 0.05 microgram/ml with 1 ml of urine of serum. Calibration curves were linear to at least 4 microgram/ml with serum and 15 microgram/ml with urine. Precision was excellent with a pooled relative standard deviation of +/- 7.5% for serum samples in a 0.1-4 microgram/ml range.

Assay of pemoline in human plasma, saliva and urine by capillary gas chromatography with nitrogen-selective detection.

A simple gas chromatographic assay of the psycho-stimulant pemoline in human urine, plasma and saliva has been developed. Instead of direct extraction of the drug from urine, plasma and saliva, it is hydrolyzed to 5-phenyl-2,4-oxazolidine-dione with 1 N hydrochloric acid. After extraction this compound is methylated with diazomethane and determined by gas-liquid chromatography using a capillary SCOT column with a mixed stationary phase, a solid injection system and a nitrogen-selective detector. 5-Phenyl-2,4-oxazolidinedione, which was also found to be a metabolite of pemoline, could be determined quantitatively in human urine.

GLC determination of pemoline in biological fluids.

A specific GLC method for the determination of microgram quantities of the central stimulant pemoline in biological fluids is described. Extraction problems due to the low solubility of pemoline are avoided by acid hydrolysis of the drug to 5-phenyl-2,4-oxazolidinedione, which then can be easily isolated by two-phase extraction with dichloromethane, ether, or chloroform. Amide-imide tautomerism enables a cleanup of the extract. Quantitative determination at the microgram level is done on a methylated fraction of the dichloromethane extract by GLC using a suitable internal standard. For supporting evidence of the GLC method's specificity, the compound is also identified by examining an aliquot of the final dichloromethane extract by TLC.

Gas chromatographic determination of pemoline as 5-phenyl-2,4-oxazolidinedione in human urine.

A simple gas chromatographic assay of the psychostimulant pemoline in human urine has been developed. Instead of extraction of the drug from urine, it is hydrolysed to 5-phenyl-2,4-oxazolidinedione with 1 N hydrochloric acid. After the extraction, this compound is methylated with diazomethane and determined by gas-liquid chromatography using a nitrogen-selective detector and a solid injection system. The method has been applied in preliminary human pharmacokinetic studies, by measuring the urinary excretion rate of pemoline following oral administration. At present, the screening procedures for doping control do not involve the detection of pemoline, but the method described can easily be incorporated in such procedures.

Gas chromatographic determination of pemoline in biological fluids using electron capture detection.

A simple and sensitive gas chromatographic (GC) method for the determination of pemoline in biological fluids, utilizing electron capture detection is described. Plasma samples with a pemoline analog added as internal standard were deproteinized with sulfosalicylic acid, and the supernatants were heated at 80 degrees. The pemoline-dione formed was extracted with benzene, and the extract was analyzed on a gas chromatograph equipped with a tritium foil electron capture detector. Poly A-103 (3%) on Gas-Chrom Q (100-120 mesh) packed in a 3-ft. silanized glass column was used as the stationary phase, with nitrogen serving as carrier gass. Under the same GC conditions, benzene extracts of pemoline-dione from acid-hydrolyzed urine samples were analyzed. Us-ng 1 ml of plasma or urine, the lower limit for the assay was about 0.1 microgram/ml. The method is accurate and reproducible, with a relative standard deviation with +/-4%. Mandelic acid (a metabolite of pemoline) does not interfere with the assay.

Combined gas-liquid chromatographic-mass spectrometric analysis of pemoline in biological samples.

A description is given of a gas-liquid chromatographic (GLC) method for the detection and determination of pemoline in biological samples. On treatment with hydrochloric acid, pemoline is converted into 5-phenyl-2,4-dioxooxazolidine, an acidic compound, which can be easily extracted with dichloromethane and determined by GLC. A combined GLC-mass spectrometric method is described.

[Quantitative determination of pemoline in serum and urine after ingestion of therapeutic doses (author's transl)]. / Quantitative Bestimmung von Pemolin in Serum und Harn nach Einnahme therapeutischer Dosen.

Serum and urine 5-phenyl-2-imino-4-oxazolidone (pemoline, Tradon) levels have been determined in normal subjects after therapeutic doses (40 mg). Peak levels in serum were reached within 4-6 h (1.1-1.5 mg/1). The half-life was 16-18 h. After the oral administration of 40 mg of pemoline 35-50% of the dose was excreted in urine within 32 h.