Determination of plasma amino acids by gas chromatography

A complete methodology, incorporating a novel clean-up technique, for quantitative determination of amino acids in plasma by gas chromatography is described. Glucose, a component causing major analytical interference, is removed by an enzymic reaction included in the pre-chromatographic clean-up. The procedure for derivatisation of amino acid standards is shown to be reproducible down to a level of 2.5 micrograms for each amino acid, relative standard deviations for all amino acids except arginine and histidine being 3% or lower. For the entire procedure applied to plasma, relative standard deviations for most amino acids are below 5% with recoveries ranging from 90 to 120%. Normal values, obtained using eighteen plasma samples, are in reasonable agreement with published data. Plasma amino acid values were determined simultaneously by gas chromatographic and ion-exchange chromatographic techniques. Statistical evaluation shows there to be no significant difference between corresponding values for eleven amino acids. Values for tyrosine, histidine and particularly phenylalanine show significant differences (p less than 0.001).     

Assay of underivatized intrazepam and clonazepam in plasma by capillary gas chromatography applied to pharmacokinetic and bioavailability studies in humans

The assay procedure of underivatized, intact nitrazepam and clonazepam in human plasma is described, using gas chromatography with a support-coated open tubular column (OV-17), a solid injection system and electron-capture detection. Clonazepam is used as a internal standard in the assay of nitrazepam and vice versa. Linear calibration curves after a single extraction step were obtained in the concentration range 10--100 ng/ml plasma, with standard deviations less than 4.9%. The sensitivity limit of the method is about 1 ng/ml plasma for both drugs. The method was applied to pharmacokinetic and bioavailability studies of nitrazepam in humans. Seven healthy volunteers received two nitrazepam-containing tablet preparations (5 mg) and plasma concentrations were determined regularly from 15 min to 80 h following drug administration. The mean elimination half-life of nitrazepam was 27 h (range 13-34 h). Considerable intra-individual differences in peak level times between the two preparations were observed, whereas the extent of bioavailability was rather similar.     

Quantitative analysis of ethylenediamine in plasma by capillary column gas chromatography

A specific and sensitive method for the determination of ethylenediamine in human plasma is described. The aqueous sample is treated with m-toluoyl chloride, yielding the N,N′-bis(m-toluoyl) derivative of the diamine, which is extracted into dichloromethane and quantitated after “on column” methylation by capillary gas liquid chromatography with alkali flame ionization detection. The corresponding derivative of putrescine serves as internal standard. The assay is reproducible and calibration curves are linear over the concentration range 0.05 to 10 μg · ml^?1. The lower detection limit is about 10 ng · ml^?1. The structures of the compounds of interest eluting from the capillary column are examined by gas liquid chromatography/mass spectrometry. The assay has been applied to the analysis of ethylenediamine in plasma following the administration of aminophylline and ethylenediamine in a cross-over study to patients with bronchopulmonary diseases. The method also proves suitable for measuring other primary and secondary amines and diamines in aqueous solutions by gas liquid chromatography.     

Sensitive determination of piritramide in human plasma by gas chromatography.

A selective and sensitive method for the determination of piritramide in human plasma is described. A 1-ml aliquot of plasma was extracted with 10 ml of hexane-isoamyl alcohol (99.5:0.5, v/v) (extraction efficiency 86%) after addition of 50 microliters of 2 M ammonia and 20 microliters of aqueous strychnine solution (100 ng per 10 microliters) as internal standard. Gas chromatography was performed with J&W DB-1, 30 m x 0.53 mm I.D. separation column, film thickness 1.5 microns, using an nitrogen-phosphorus-sensitive detector. The assay was linear in the concentration range 3.75-2250 ng/ml (r = 0.999), with a lower limit of detection of 1-2 ng/ml. The precision was determined using spiked plasma samples (10 and 50 ng/ml), with coefficients of variation of 3.5 and 3.1% (intra-day; n = 5) and 4.6 and 4.1% (inter-day; n = 4). In the range 3.75-150 ng/ml, the accuracy of the assay was 3.36%. The method was used for the determination of piritramide plasma concentrations in patients receiving intra- or post-operative analgesia.     

Determination of oxprenolol in human plasma by high-performance liquid chromatography, in comparison with gas chromatography and gas chromatography-mass spectrometry

A high-performance liquid chromatographic method for the quantitative assay of oxprenolol in human plasma is described. After addition of alprenolol as internal standard, the compounds are extracted from plasma at alkaline pH into an organic phase and back-extracted into an acidic aqueous phase. Separation of the plasma components and metabolites was achieved on a reversed-phase column. Concentrations down to 66 nmol/l (20 ng/ml) can be determined with UV detection at 222 nm. This technique compares favourably with gas chromatographic and gas chromatographic-mass spectrometric methods.     

Determination of busulfan in human plasma by gas chromatography with electron-capture detection

A simple and highly sensitive gas chromatographic method has been developed for the determination of busulfan in human plasma. After extraction of plasma specimens (clinical or spiked) with ethyl acetate, busulfan and the internal standard [1,8-bis(methanesulfonyloxy)octane] were derivatized with 2,3,5,6-tetrafluorothiophenol to yield compounds monitored by a 63Ni electron-capture detector. Sample recoveries from extraction and derivatization were greater than 78 and 91%, respectively. The limit of quantitation was 0.01 microgram/ml (0.04 microM) in 1 ml of plasma with a linear relationship over the 0.01-1.0 micrograms/ml (0.04-4 microM) concentration range. The method has been applied to analyze the plasma versus time profile of busulfan in human subjects following administration of an oral dose of 4 mg/kg per day as a marrow ablative chemotherapy for bone marrow transplantation.