Quantitation of levorphanol in human plasma at subnanogram per milliliter levels using capillary gas chromatography with electron-capture detection

A gas chromatographic method for the determination of levorphanol in human plasma is described. The method utilizes extractive alkylation with tetrabutylammonium cation as the phase-transfer catalyst and pentafluorobenzyl bromide as the alkylating agent, and employs a structural analog, d-3-hydroxy-N-ethylmorphinan, as the internal standard. The pentafluorobenzyl ethers formed are separated by capillary gas chromatography and detected by electron capture. The method has good precision and accuracy for concentrations ranging from 0.25 ng/ml to 100 ng/ml and has been used to measure plasma concentrations as part of a study to evaluate the management of chronic neuropathic pain with levorphanol.     

Determination of acetaldehyde in biological samples by gas chromatography with electron-capture detection

A simple specific assay was developed for the determination of acetaldehyde in biological samples. Acetaldehyde was derivatized to 2,4-dinitrophenylhydrazone, which was determined by gas chromatography with electron-capture detection. The use of this detection method is an important device to which no one drew notice. This procedure was very simple and so sensitive that as little as 500 fmol of acetaldehyde could be measured in aqueous solution. The calibration curve of acetaldehyde was linear at least up to 40 μM. Its recoveries from human plasma and rat liver homogenate were 96.5 and 95.7%, respectively.     

Assay of plasma catecholamines by liquid chromatography with electrochemical detection

A method was developed for the simultaneous assay of noradrenaline and adrenaline in 2 ml of human plasma. The method involves adsorption of the catechols onto alumina, desorption, lyophilizing, reconstitution, and injection into a reverse-phase ion-pairing liquid chromatography system. Sensitivity and selectivity are introduced using direct electrochemical detection of the column eluant.     

Determination of a new blood glucose-lowering agent in biological fluids by capillary gas chromatography using electron-capture detection

A gas chromatographic method for the determination of 5-(4-chlorophenyl)-2-[(4-methylphenyl)sulphonyl]-4-pentynoic acid (I) in plasma (serum) and urine has been developed. After an extraction process, the cleaned-up organic extract was derivatized with diazomethane at ambient temperature. Results are evaluated from peak-height ratios with respect to the appropriate internal standard. The detection limit following extraction of a 1-ml plasma sample is about 20 ng/ml.     

Simultaneous determination of griseofulvin and 6-desmethylgriseofulvin in plasma by electron-capture gas chromatography

Two methods have been developed for the simultaneous determination of griseofulvin and its major metabolite 6-desmethylgriseofulvin in plasma using electron-capture gas chromatography. The first method was based on the quantitative reversion of the 6-desmethyl metabolite to griseofulvin by diazomethane. Plasma extract was chromatographed both before and after treatment with diazomethane, the former being the measure of griseofulvin and the latter representing the sum of the two compounds. In the second method, plasma extract was treated with diazobutane and griseofulvin and the butylated metabolite were separated by gas chromatography. The sensitivity for griseofulvin was 20 ng/ml by both methods and that for the metabolite was 20 ng/ml and 40 ng/ml by the first and the second method, respectively. The concentrations of the metabolite as well as griseofulvin were determined in dog and human plasma after oral administration of griseofulvin.     

Simplified method for determination of the tetracyclic antidepressant mianserin in human plasma using gas chromatography with nitrogen detection

A simplified gas chromatographic method for determination of the antidepressant drug mianserin in human plasma is described. Application of a nitrogen-sensitive detector reduces the assay procedure to extraction, concentration and gas chromatographic determination. The method is suitable to determine mianserin in human plasma at the 1 ng/mol level on a routine basis. At the 20 ng/ml level the deviation of the mean from the true value and the relative standard deviation amount to 1.0% and 6.8%, respectively.     

Determination of pilocarpic acid in human plasma by capillary gas chromatography with mass-selective detection

A novel, highly sensitive method for the determination of pilocarpic acid (PA) in human plasma is described. In addition, the method provides for the conversion of the lactone, pilocarpine (P), to PA so that a total drug presence can be determined. Using novel high-performance liquid chromatographic conditions capable of separating P, isopilocarpine (I-P), PA and isopilocarpic acid (I-PA) from each other and from endogenous plasma impurities, it was confirmed that P exclusively and quantitatively converts to PA in heparinized human plasma during storage. For the determination of PA, the selective extraction of PA from protein-free plasma was accomplished using two different solid-phase extraction (SPE) cartridges in two consecutive SPE steps. After extraction, PA was lactonized with trifluoroacetic acid back to P, and both P and an internal standard were acylated using heptafluorobutyric anhydride (HFBA). The trifluoroacetylated derivatives were monitored using gas chromatography (GC) with mass spectrometric (MS) detection. This procedure allowed the sensitive and reliable determination of PA with a limit of quantification (LOQ) of 1 ng/ml, which could not be achieved using previously described methods. The assay was validated in the concentration range of 1 to 10 ng/ml with an intra-day precision (expressed as the coefficient of variation, C.V.) ranging from 9.9 to 0.5%. Inter-day precision for the quality control standard at 2.5 ng/ml showed a C.V. of 10.2%. Accuracy ranged from 94 to 102%. The assay was used to monitor the maximum systemic exposure to P, administered by the ocular route, in terms of total plasma PA (P and PA).     

Determination of ritodrine in biological fluids of the pregnant sheep by fused-silica capillary gas chromatography using electron-capture detection

A sensitive and selective gas chromatographic assay method employing splitless injection, fused-silica capillary columns and electron-capture detection is reported for the quantitation of the tocolytic drug, ritodrine, in a variety of biological fluids obtained from the pregnant ewe and fetus. This method has improved sensitivity and selectivity over previously published assay procedures. A 25 m × 0.31 mm I.D., cross-linked 5% phenylmethylsilicone, fused-silica capillary column was employed for all analyses. Linearity of response was observed over the range 2.5–75 ng of ritodrine base per 0.05–0.5 ml of biological fluid, representing ≈ 1–75 pg at the detector. The coefficient of variation was less than 10% over the range 2.5–75 ng of added ritodrine. The minimum quantifiable amounts is ≈ 2.5 ng from a 0.5-ml biological fluid sample. Applicability of this method to biological fluids, obtained from ovine subjects, is demonstrated by the analysis of samples obtained during the course of ritodrine placental transfer studies.     

Determination of estradiol 2- and 4-hydroxylase activities by gas chromatography with electron-capture detection

A highly sensitive assay has been developed for measuring the rate of formation of 2-hydroxyestradiol and 4-hydroxyestradiol from estradiol by microsomal preparations. Catechol estrogens were converted to heptafluorobutyryl esters, which were separated by capillary column gas chromatography and quantified using electron-capture detection. 2-Hydroxyestradiol 17-acetate was used as an internal standard. The identity of catechol estrogen derivatives was verified by gas chromatography—mass spectrometry using negative-ion chemical ionization. Estrogens were identified by negative molecular ions and/or by characteristic fragments. This procedure permits quantification of catechol estrogens at the subpicogram level. The assay was validated by comparing estrogen 2- and 4-hydroxylase activities in microsomes from hamster and rat liver with values reported previously.     

Analysis of selenium in bovine liver by gas chromatography with mass-selective, electron-capture and nitrogen-phosphorus detection

The concentration of selenium (Se) in liver was determined by gas chromatography (GC) with mass-selective (GC-MS), electron capture (GC-ECD) and nitrogen-phosphorus (GC-NPD) detection. Liver samples were digested in a mixture containing HNO₃ and Mg(NO₃). Se^VI was converted to Se^IV. Se^IV was derivatized with 4-nitrophenylenediamine and then extracted in toluene. A 1-μl volume of the toluene extract was analyzed by the GC-MS, GC-ECD or GC-NPD methods. The detection limits of the GC-ECD, the GC-NPD and the GC-MS methods were 25, 50 and 800 pg, respectively. The GC-NPD method was more selective for the derivatized Se than the GC-ECD method. The GC-MS method had the advantage of using the ⁷⁶Se isotope as the internal standard. Se concentrations in liver samples determined by the three methods were comparable.     

Determination of urinary methylhistamine excretion in male and female rats by gas chromatography with electron-capture detection

A gas-liquid chromatographic method for the determination of methylhistamine in urine is described. Methylhistamine is reacted with 2,6-dinitro-4-trifluoromethylbenzene sulfonic acid and the derivative thus formed is quantitated with electron-capture detection. The twenty-four hour urinary excretion of methylhistamine in the male rat is about five-fold greater than that in the female rat. Amino-guanadine, a diamine oxidase inhibitor, causes a four-to five-fold increase in methylhistamine excretion in both male and female rats. Treatment with pargyline, a monoamine oxidase inhibitor, causes only a small increase in methylhistamine excretion in male and female rats thereby suggesting that oxidative deamination of methylhistamine is a relatively minor pathway.     

Analysis of malondialdehyde in biological matrices by capillary gas chromatography with electron-capture detection and mass spectrometry

A gas chromatographic method is described for the quantification of free and total malondialdehyde (MDA) in biological materials. The procedure involves derivatization of the analyte with 2,4,6-trichlorophenylhydrazine, extraction with n-hexane, and separation of the cyclic derivatization product on a OV-5 gas chromatographic column. Concentration of the derivatization reagent, pH, reaction time, and temperature were investigated to determine the optimal derivatization conditions. Under these conditions, the method allows for the selective detection of free and total MDA at femtomole levels in several biological materials without any interferences. The procedure yields relative standard deviation values for the intra- and interassays in the range 3.3 and 3.9%, respectively, for the electron-capture and mass-selective (SIM mode) detection systems. Recoveries of MDA from spiked matrices reached 96%. The present method offers the advantage of the alternative use of either electron-capture or mass-selective detection. Furthermore it avoids overestimation of MDA since it employs mild conditions for sample processing and there is no need for preventing protein separation for the assessment of free MDA.     

Determination of tianeptine in human plasma using high-performance liquid chromatography with fluorescence detection

A new, selective and sensitive high-performance liquid chromatography (HPLC) method with fluorimetric detection was developed for the determination of tianeptine (TIA) in human plasma using solid phase extraction (SPE) procedures. The method is based on the derivatization of TIA with 4-chloro-7-nitrobenzofurazan (NBD-Cl) in borate buffer of pH 8.5 to yield a yellow, fluorescent product. The HPLC separation was achieved on a Phenomenex C(18) column (250 mm x 4.6 mm) using a mobile phase of acetonitrile-10mM orthophosphoric acid (pH 2.5) (77:23, v/v) solvent system at 1 mL/min flow rate. Gabapentin (GA) was used as the internal standard. The fluorometric detector was operated at 458 nm (excitation) and 520 nm (emission). The assay was linear over the concentration range of 5-300 ng/mL. The detection limit (LOD) was found to be 2 ng/mL. The mean recovery was determined to be 88.6%. The proposed method was applied for pharmacokinetic study of 12.5mg TIA in a healthy volunteer.