A gas chromatographic mass spectrometric chemical ionization assay for haloperidol with selected ion monitoring

A new gas chromatographic mass spectrometric chemical ionization assay for haloperidol with selectedion monitoring is presented which provides for better combined selectivity and sensitivity than previous assays. Levels of haloperidol in 2 ml of human plasma were reproducibly measured down to subnanogram levels. Both methane and methane--ammonia chemical ionization spectra are presented for haloperidol and the internal standard trifluperidol.     

Biological monitoring of exposure to n-heptane by gas chromatographic mass spectrometric determination of its metabolites

Urine samples from 10 workers that had been exposed to n-heptane were analysed by the GC/MS technique to verify the concentrations and the relative abundances of its metabolites. The procedure of sample preparation has undergone some modifications with respect to the Perbellini method and the mass spectrometric detection was carried out in selected ions monitoring conditions. The analyses of samples collected during three different workshifts showed that 2 heptanol was not the main metabolite and that the remains of 2 heptanone, valerolactone and 2,5 heptanedione were present at the beginning of the successive work week at 12, 34 and 39 of the average values found at the end of the previous week. Overall, a very slow excretion rate was detected for the last metabolite. The main and significant metabolite at the end of the two workshifts was 2 heptanone which was detected in urine at average values of 413 and 238 μg g^-1 creatinine. This urinary ketone correlated better than other metabolites with respect to the airborne n-heptane at the end of both the workshift and work week. These preliminary data suggest that further studies should be carried out to confirm whether 2 heptanone is really useful as an n-heptane marker in biological monitoring.     

Gas chromatographic mass spectrometric identification of N-dicarboxylmonoglycines

A number of N-dicarboxylmonoglycines of biological interest have been synthesized. They were characterized by means of mass spectrometry. Gas chromatography of the methyl esters of methylmalonyl-, succinyl-, glutaryl-, adipyl-, suberyl- and sebacylglycines showed a single sharp peak for each compound on Dexsil 300 and OV-17 columns. Methylene unit values and mass spectra of the six methyl esters are reported.     

Capillary gas chromatographic/mass spectrometric analysis of abnormal metabolites in hypoglycin-treated rat urine

Numerous abnormal metabolites were identified in large amounts in the urine of hypoglycin-treated rats using capillary gas chromatography/mass spectrometry-computer analysis. These metabolites are not detectable in significant amounts in normal rats' urine. Ten of them have not been previously associated with hypoglycin administration: these are several hydroxy compounds, including those from the valine and isoleucine pathways, 2-oxo-adipic acid, n-butyrylglycine and isovaleryl glucuronide. These results indicate that the pathways of isoleucine and valine metabolism are inhibited at their respective acyl-CoA dehydrogenation steps, as is the case for fatty acid, leucine and lysine metabolism, as previously shown. The mass spectra of the trimethylsilyl derivatives of cis, cis-4,7-decadiene-1,10-dioic, cis-4-decene-1,10-dioic, cis-4-octene-1,8-dioic acids, and (methylenecyclopropyl)acetylglycine, which were previously identified using nuclear magnetic resonance and oxidative cleavage or acid hydrolysis, are presented for the first time.     

Gas chromatographic and mass spectrometric analysis of conjugated steroids in urine

This study was carried out qualitatively and quantitatively to investigate the presence and the concentrations of anabolic steroids in urine collected from orally administered humans. Microanalysis of conjugated steroids by gas chromatography and mass spectrometry (GC/MS) has been carried out. Following oral administration three major metabolites of anabolic steroid drugs have been detected and partially characterized. The six steroids can be analysed at the same time in 17 min. The lower detection limit was 10 ng/ml in 5 ml of urine. The conjugated steroids from urine were centrifuged to 2,430g for 10 min, the supernatant solution passed through Amberlite XAD-2 column and the steroids eluted fraction esterified by using MSTFA and TMSI. The rate of metabolism and urinary excretion seem to be reasonably fast.     

Simultaneous determination of ketobemidone and its N-demethylated metabolite in patient plasma samples by gas chromatography mass spectrometry with selected ion monitoring

An analytical procedure has been developed for the simultaneous determination of ketobemidone and its N-demethylated metabolite, norketobemidone. After isolation from plasma and re-extraction to acidic aqueous phase, the two aminophenols were extracted as ions pairs with tetrabutylammonium to dichloromethane, where derivatization with ethyl chloroformate took place. Determination was performed by gas chromatography mass spectrometry with selected ion monitoring. Ketobemidone and norketobemidone could be detected in plasma in a concentration of 1 ng ml-1 and 3 ng ml-1, respectively. Determinations were performed down to 5 ng ml-1. The relative standard deviation of the method in the analysis of 10 ng ml-1 of ketobemidone and norketobemidone, respectively, was 8% and 9% (n=10). The absolute recovery of unconjugated ketobemidone and norketobemidone through the method at the 100 ng ml-1 level was 91% and 85%, respectively. The method was applied to the determination of ketobemidone and norketobemidone in plasma from patients given ketobemidone. The concentrations of unconjugated norketobemidone was too small to be detected.     

Biological monitoring of dinitrotoluene by gas chromatographic–mass spectrometric analysis of 2,4-dinitrobenzoic acid in human urine

The method of analysis described permits the determination of 2,4-dinitrobenzoic acid down to the lower μg l⁻¹ range in the urine of persons exposed to dinitrotoluene. 2,4-Dinitrobenzoic acid is the main metabolite of 2,4-dinitrotoluene and technical dinitrotoluene. After acidic hydrolysis, which served to release the conjugated part of the 2,4-dinitrobenzoic acid, the analyte was selectively separated from the urine matrix via various extraction steps and then derivatised to the methyl ester. Quantitative analysis was carried out using capillary gas chromatography and mass selective detection. 3,5-Dinitrobenzoic acid was used as an internal standard. The detection limit was 1 μg l⁻¹ urine. The relative standard deviations of within-series imprecision were between 5 and 6%. The relative recoveries were between 91 and 110% depending on the concentration. The analytical method developed as part of this study was used to investigate a collective consisting of 82 urine samples from persons working in the area of explosives disposal. The concentrations of 2,4-dinitrobenzoic acid determined ranged from the detection limit to 95 μg l⁻¹ urine. The method allowed the quantification of low-level internal exposure to dinitrotoluene.     

Rapid determination of para-phenylenediamine by gas chromatography-mass spectrometry with selected ion monitoring in henna-containing cosmetic products

A rapid method for the determination of para-phenylenediamine (PPD) in cosmetic products, such as henna tattoos has been developed and evaluated. This analytical procedure involved extracting a 10mg test portion of cosmetic product in 10 mL of ethyl acetate, followed by determination by gas chromatography-mass spectrometry in the selected ion monitoring mode (GC/MS-SIM). 1,4-Phenylenediamine-2,3,5,6-d(4) was selected as an internal standard that was added at the beginning of the extraction procedure and used to correct for recovery and matrix effects. The linearity ranged from 1.0 to 1275 μg/mL with a coefficient of determination (r(2)) greater than 0.999. LOQ and LOD were 1.0 and 0.10 μg/mL, respectively. The recovery in a tattoo product containing PPD was 94% and that for a tattoo product containing no PPD reached 105%. Extraction efficiency of 98% was obtained. This method has been successfully applied to henna temporary tattoo and other henna-related cosmetic products for the determination and quantitation of PPD.     

Simultaneous determination of urinary androgen glucuronides by high temperature gas chromatography-mass spectrometry with selected ion monitoring

An efficient procedure is described for the simultaneous determination of 9 androgen glucuronides including androsterone, etiocholanolone, 11-ketoandrosterone, 11-ketoetiocholanolone, 11beta-hydroxyandrosterone, 11beta-hydroxyetiocholanolone, and dehydroepiandrosterone (DHEA) in 3-glucuronide form and dihydrotestosterone (DHT) and testosterone in 17-glucuronide form from urine specimens. The method involves solid-phase extraction of the urinary steroids using Serdolit PAD-1 resin, with subsequent conversion to methyl ester-trimethylsilyl (Me-TMS) ether derivatives for the direct analysis by gas chromatography-mass spectrometry (GC-MS) using high temperature MXT-1 (Silcosteel-treated stainless steel) capillary column. Upon split injection of Me-TMS steroids at 330 degrees C into the MXT-1 capillary column initially maintained at 300 degrees C then programmed to 322 degrees C at 2 degrees C/min, each androgen glucuronide was well separated in excellent peak shape. The characteristic ions at m/z 217 constituting the base peaks in the electron-impact (20 eV) mass spectra for most steroids permitted their sensitive detection by GC-MS with selected-ion monitoring (SIM), whereas base peak ion at m/z 271 was used for the SIM of dehydroepiandrosterone-3-glucuronide. The detection limits for SIM of most of the steroids were 15 pg except for the 3-glucuronides of 11-ketoandrosterone and 11-ketoetiocholanolone, which could be detected down to 20 pg. The SIM responses were linear with correlation coefficients varying from 0.981 to 0.993 in the concentration range of 20 to 3000 ng/ml for the androgens studied. When applied to urine samples, the present method allowed rapid screening for the 7 androgens in their glucuro-conjugated forms simultaneously with good overall precision and accuracy within the normal concentration ranges of 15.1 to 3124.6 ng/ml.     

Use of chemical ionization in gas chromatographic/mass spectrometric screening of human urine for disaccharides containing inositol

Gas chromatography/mass spectrometry (GC/MS) in the positive chemical ionization (CI) mode was used to screen normal urine for inositol-containing disaccharides in the form of permethylated derivatives, after borodeuteride reduction of the reducing saccharides. Ammonia was the reactant gas. The results revealed the existence of deoxyhexosyl-inositol and hexosyl-inositol disaccharides, and of a new compound, N-acetylhexosaminyl-inositol disaccharide. Up to four isomers of deoxyhexosyl-inositol could be present in the same sample even though only one of them has so far been fully characterized in man. As regards the hexosyl-inositols, one to three isomers were present in the same sample and probably corresponded to the three isomers of galactosyl-inositol recently described in man. N-Acetylhexosaminyl-inositol (identified elsewhere by us as N-acetylgalactosaminyl-alpha (1-1)-myo-inositol) was seen in only a few samples. No relationship can be found between the excretion of all these inositol-containing disaccharides on the one hand, ABO(H) blood group and 'Secretor' status (Se or sese) of the donors on the other. The gas chromatographic CI mass spectrometric technique used here with various ammonia pressures can be applied to the screening of other biological fluids or tissues for inositol glycosides.     

Analysis of the thromboxane/prostacyclin balance in human urine by gas chromatography/selected ion monitoring: abnormalities in diabetics

We microanalyzed 2,3-dinor-6-keto-prostaglandin F_1α (2,3-dinor-6-keto-PGF_1α 1) and 11-dehydrothromboxane B₂ (11-dehydro-TXB₂, 2) in human urine. Samples containing a [²H₄]-analogue as an internal standard were extracted by chromatography using Sep Pak tC18 and silica gel. The compounds were then analysed by means of the lactone ring opening reaction and dimethylisopropylsilylation. The conversion of 1 to 1-methyl ester (ME)-propylamide (PA)-9,12,15-dimethylisopropylsilyl (DMIPS) ether derivative and of 2 to 1-ME-6-methoxime (MO)-9,12,15-tris-DMIPS ether derivative was followed by gas chromatography/selected ion monitoring (GC/SIM). Interfering substances from the urine matrix were eliminated during GC/SIM analysis using a DB-5 column. We were able to detect 1 (222–1031 pg/mg creatinine) and 2 (18–155 pg/mg creatinine) in human urine. Furthermore, the thromboxane/prostacyclin (IX/PGI) ratio in the urine of diabetics was higher than that of healthy volunteers. This method can be used to determine the TX/PGI balance in human urine.     

Keto acids in tissues and biological fluids: O-t-butyldimethylsilyl quinoxalinols as derivatives for sensitive gas chromatographic/mass spectrometric determination

Quinoxalinol t-butyldimethylsilyl ethers were prepared from three branched-chain and from two aliphatic unbranched 2-keto acids. The electron impact (EI) mass spectra display pronounced [M-57]+ ions. With 39-51% of total ion current contained within them, sensitivity is greater than on chemical ionization (CI) mass spectrometry of O-trimethylsilyl derivatives. Mass spectra and chromatographic behaviour of these novel keto acid derivatives are discussed and preliminary quantitative data from rat muscle are given.     

The determination of amino acid pool sizes and turnover rates by gas chromatographic mass spectrometric analysis of stable isotope enrichment

Gas chromatography mass spectrometry has been used to determine the 15N enrichment of plasma glycine of rabbits at various times following the intravenous administration of 15N-glycine. These data were used to prepare isotope enrichment time decay curves for eleven individual animals. The slopes and intercepts of least squares lines that describe the decay curves were considerably more accurately than those reported in similar studies employing radioactive tracers. Individual glycine pool sizes (13.8-37.4 micronmoles per 100 g body wt), turnovers rates (2.66-3.36 pools h-1) and flux (50.4-99.7 micronmoles h-1 per 100 g body wt) were estimated from these parameters in a group of animals and compared with the literature values. These results demonstrate that low risk non-radioactive stable isotopes can be substituted for radioactive tracers in studies of human amino acid metabolism, with considerable saving in time and without loss in accuracy, when gas chromatography mass spectrometry is used to determine plasma amino acid and stable isotope enrichment.     

Liquid chromatography–electrospray ionization mass spectrometric analysis of corticosterone in rat plasma using selected ion monitoring

A simple and fast yet highly sensitive and specific method based on HPLC coupled to electrospray ionization mass spectrometry has been developed for the quantitation of corticosterone in rat plasma. After extraction of rat plasma (100 μl) with diethyl ether using 5-pregnen-3β-ol-20-one-16α-carbonitrile (Sigma) as internal standard, HPLC was performed on a short C₈ column (Zorbax-Eclipse, 50×4.6 mm I.D.) using a steep methanol–water gradient (methanol 54% to 90% in 6 min). Detection was performed on a single quadruple mass spectrometer in selected ion monitoring mode (m/z 369 for corticosterone and 364 for the internal standard). The detection limit of the assay was 9 fmol (3 pg) of corticosterone on column. In vitro data were subjected to curve fitting (cubic, r²=0.9999). Recovery of corticosterone after extraction ranged from 81 to 93%. The relative standard deviations for intra- and inter-assay precision ranged from 0.8 to 3.6% and 5.2 to 12.9%, respectively. Corticosterone did not undergo any appreciable degradation when stored in plasma at −20°C for 2 months. The assay is routinely used in our laboratory to examine corticosterone levels as a marker of stress in rats and may also be used for the determination of 18-hydroxy-11-deoxycorticosterone.     

Development of a gas chromatographic selected ion monitoring assay for thyroxine (T4) in human serum

The components of a gas chromatographic mass spectrometric-selected ion monitoring (SIM) assay for thyroxine (T4) in human serum are described. The internal standard for the assay was synthesized from deuterium-labelled 3,5-diiodotyrosine and 3,5-diiodo-4-hydroxyphenylpyruvic acid. A novel method was developed for isolating the products of the coupling reaction. The results obtained by gas chromatography mass spectrometry SIM were compared with those of radioimmunoassay. The gas chromatographic mass spectrometric SIM assay would form the basis of a reference assay for T4.     

Measurement of propericiazine in plasma by capillary gas chromatography and selected ion monitoring detection

A new method, based upon a selective extraction and gas chromatographic/mass spectrometric analysis, was developed to monitor the effect of combined haemoperfusion-haemodialysis treatment in a case of propericiazine poisoning. The method relies on the selected ion monitoring of the acetate derivatives of propericiazine and its internal standard fluphenazine, after their extraction from 1 ml of alkalinized plasma with n-hexane:isopropanol (8:2, v/v), back-extraction into an acidified water phase, realkalinization and extraction with n-hexane: isopropanol, derivatization with acetic anhydride and gas chromatography on a short (12 m) OV-101 fused silica capillary column. The described procedure is specific and provides between-assay variability of 4.8% CV at 5 micrograms 1(-1) plasma concentration. The method enables quantification down to 1 microgram 1(-1) and hence demonstrates sufficient sensitivity to permit pharmacokinetic or drug monitoring studies.     

Quantitative gas chromatographic mass spectrometric determination of mandelic acid in blood plasma. Comparison of deuterated and homologous internal standards

Quantitative gas chromatography/mass spectrometry/selected ion monitoring of mandelic acid in plasma in the lower micromolar range has been investigated using both the deuterated compound and a homologue, 2-phenyllactic acid, as internal standards. On chromatography of the TMSi-ester ethers, the latter is less stable than the former. The chromatographic isotope effect observed for deuterated mandelic acid does not suggest a carrier function for this compound. Normal plasma levels of mandelic acid are about 0.5 microM. Only a minor portion of phenyramidol is metabolized to mandelic acid. Preliminary in vivo data indicate the presence of a stereoselective transport system for D(-)-mandelic acid in gastrointestinal tract and possibly kidney.