Gas Chromatography-negative Ion Chemical Ionization Research Articles

Quantitative analysis of S3341 in human plasma and urine by combined gas chromatography-negative ion chemical ionization mass spectrometry: 15 month inter-day precision and accuracy validation.

A new quantitative assay for the determination of S3341, an alpha-2 agonist antihypertensive drug, has been developed using combined gas chromatography-negative ion chemical ionization mass spectrometry. The [M]-. ions from TFA derivatives of S3341 (m/z 276) and the internal standard (2H4)S3341 (m/z 280) are monitored simultaneously by selected ion monitoring. For S3341 concentrations ranging from the limit of detection (0.2 ng ml-1 using 1 ml of plasma) to 5 ng ml-1, the average assay precision (CV) is approximately 7% while the average assay accuracy (percentage of error) is 4%. Validation of the day-to-day precision and accuracy was realized after analysing control plasma samples (n = 295) concurrently with the biological samples collected during the pharmacokinetic studies conducted over 15 months. The average day-to-day precision (CV) and accuracy (percentage of error) are 10% and 6% respectively, thus indicating that this assay procedure routinely provides reliable analytical data.

Species-specific variations in the molecular heterogeneity of the platelet-activating factor.

The molecular heterogeneity of platelet-activating factor (PAF) synthesized by unstimulated and Ca2+ ionophore (A23187)-stimulated PMN from rat, mouse, and guinea pig and by rat basophilic leukemia (RBL) cells was investigated by gas chromatography-negative ion chemical ionization mass spectrometry. Several molecular species of PAF ranging from C14:0 to C19:0 were detected in all of the cells studied. PAF produced by each cell type exhibited a unique pattern of molecular species distribution. Although C16:0 was the major PAF molecular species of rat PMN and RBL cells representing 96% and 85% of the total PAF, respectively, PAF from mice PMN contained 81% of C16:0, 10% of C18:1, and 6% of C18:0. Alternatively, A23187-stimulated guinea pig PMN yielded PAF molecular species 35% in C16:0, 35% in C17:0, 8% in C18:1, and 3% in C18:0. Small but significant differences in the PAF molecular species distribution of resting and ionophore stimulated cells were also observed. In contrast to the PAF molecular species composition, the precursor 1-O-alkyl-2-acyl-glycero-3-phosphocholine of all the cell types was predominantly hexadecyl (C16:0) alkyl chain in the sn-1 position, representing 60 to 80% of the total 1-O-alkyl-2-acyl-glycero-3-phosphocholine. Thus, these results not only indicate a high degree of selectivity for utilization of precursor substrates for PAF biosynthesis, but also demonstrate that the selectivity is species specific.

Confirmation of clorsulon residues in cattle kidney by capillary gas chromatography—negative-ion chemical-ionization mass spectrometry

A confirmatory assay for residues of the anthelmintic agent clorsulon [4-amino-6-(trichloroethenyl)-1,3-benzenedisulfonamide] in cattle kidney tissue has been developed. The assay involves isolation of a drug-containing fraction by solvent extraction, methylation of the analyte, and fused-silica capillary column gas chromatography—negative-ion chemical-ionization mass spectrometry of the pentamethyl derivative of clorsulon. The intensities of four negative ions [ m/z 406 and 408 (trichloro species) and m/z 413 and 415 (dichloro species)] are monitored. Confirmation of the presence of drug in an analyte requires that all four ions appear at the appropriate retention time with their intensity ratios within 10–15% of those arising from analysis of the reference standard, methylated clorsulon; the lower limit of detection is 3 ppb. Quantification of the drug is based on the intensity of the m/z 406 ion. Identification and quantification of residues by the gas chromatographic—mass spectrometric assay gave results in good agreement with those obtained with an electron-capture gas chromatographic assay.

Purification and quantitative analysis of urinary prostanoids in human and in rat by packed and capillary gas chromatography—negative-ion chemical-ionization mass spectrometry

We describe a method for the quantitative analysis of prostaglandin (PG) E 2 and the major urinary metabolites PGI 2 and thromboxane (Tx) A 2 in human and in rat by combined gas chromatography and negative-ion chemical-ionization mass spectrometry. The procedure is based on the sequential use of small columns with distinct properties combined with a thin-layer chromatography step, for the extraction and the purification of urinary prostaglandins. The compounds are then analysed as their pentafluorobenzyl ester-O-methyloxime-trimethylsilyl ether derivatives, using either packed or capillary columns. Deuterated analogues are used as internal standards. The method was established by using tritiated prostaglandins covering the extremes of polarity in order to optimize the recovery of prostanoids as well as the quality of the chromatograms and spectra. The overall recovery was 24%. Standard curves were obtained by the same procedure and found to be reproducible, with a maximal day-to-day variation of ±5%. The relatively simple approach required for the sequential extraction and purification of prostaglandins on small columns of distinct properties, combined with the highly specific and highly sensitive method of detection, places this procedure among the most reliable method for measuring urinary prostanoids in both humans and animals. In addition, the procedure is faster than classical approaches and necessitates smaller amounts of samples and solvents.

Profiling of prostaglandin biosynthesis in biopsy fragments of human lung carcinomas and normal human lung by capillary gas chromatography-negative ion chemical ionization mass spectrometry

Methods for the profiling of prostaglandin F 2α (PGF 2α), prostaglandin D 2 (PGD 2), prostaglandin E 2 (PGE 2), thromboxane 3 2 (TxB 2) and 6-keto-prostaglandin F 1α (6KPGF 1α) biosynthesis in tissue samples of clinical origin by capillary gas chromatography- negative ion chemical ionization mass spectrometry (CGC-NICIMS) are detailed. Alquouts (25μ1) of incubates (1 ml volume) of human lung carcinoma and normal lung tissue fragments (total protein content = 0.2 to 2.0 mg) were derivatized for vapor phase analysis in the presence of 0.75 to 1.60 ng of tetradeuterated analogs of PGE 2, PGF 2α and 6KPGF 1α without prior extraction and/or chromatography. The derivatized analytes and internal standards were detected by simultaneous monitoring of ions at six different masses characteristic for each of the derivatized prostanoids. The inter-sample and intra-sample coefficients of variantion for the assay method were typically less than 12%. The analysis of biological samples were completed with less than 2.5% of each derivatized sample per injection. The samples were of adequate purity for the identification and quantitation of each of the eicosanoids. The methods described in this report are highly sensitive with detection limits of 0.1 to 0.2 picograms per injection. The analytical procedures provide the basis for comparisons of the qualitative and quantitative profiles of prostaglandin biosynthesis and should be adaptable for use in a variety of biological and clinical studies.

Platelet-activating factor and leukotriene biosynthesis is inhibited in polymorphonuclear leukocytes depleted of arachidonic acid.

Rat peripheral or elicited polymorphonuclear leukocytes 90% deficient in arachidonic acid incorporate, after stimulation with the calcium ionophore A23187, 86% less acetate into platelet-activating factor than control. The total amount of platelet-activating factor in the ionophore stimulated elicited polymorphonuclear leukocytes deficient in arachidonate, measured by gas chromatography-negative ion chemical ionization mass spectrometry, was 84% less than that of control. The mass spectrometry also revealed the presence of various molecular species of platelet-activating factor ranging from 1-O-tetradecyl to 1-O-nonadecyl forms in both the deficient and control cells. However, the 1-O-hexadecyl was the predominate molecular species representing 79 and 96% of the total platelet-activating factor in the respective deficient and control cells. Other molecular species were less than 1.5 and 8.5% of the total for control and deficient polymorphonuclear leukocytes, respectively. Leukotriene B4 formation was also inhibited by 90% in the deficient cells. Both platelet-activating factor and leukotriene B4 biosynthesis could be partially restored in arachidonic acid-deficient cells by prelabeling the cells with arachidonate. This represents the first dietary link with platelet-activating factor biosynthesis.

Sensitive determination of deuterated and non-deuterated phenylalanine and tyrosine in human plasma by combined capillary gas chromatography—negative ion chemical ionization mass spectrometry

A combined capillary gas chromatography—negative ion chemical ionization mass spectrometric method for the determination of deuterated and non-deuterated phenylalanine and tyrosine in plasma was developed. Phenylalanine and tyrosine were converted to pentafluorobenzyl—trifluoroacetyl (PFB—TFA) and PFB—TFA—trimethylsilyl (TMS) derivatives, respectively, after prepurification with a Bio-Rad AG 50W-X2 cation-exchange column. These derivatives showed good gas chromatographic separation properties and provided intense (M  PFB) − ions. These ions were ideal for the specific and sensitive determination of deuterated and non-deuterated phenylalanine and tyrosine by selected ion monitoring assay. [2,2′,3,3,3′,4′,5′,6′- 2H 8] l-Phenylalanine and [2,2′,3,3,3′,5′,6′- 2H 7] l-tyrosine were used as internal standards. This method was used to determine the plasma levels of deuterated and non-deuterated phenylalanine and tyrosine, after oral administration of [2′,3′,4′,5′,6′- 2H 5] l-phenylalanine to a healthy person.

Determination of peripheral plasma prostanoid concentration: an unreliable index of 'in vivo' prostanoid activity.

Plasma concentrations of PGE2, PGF2 alpha, 6-keto-PGF1 alpha and TxB2 were determined in 4 healthy volunteers by gas chromatography-negative ion chemical ionization mass spectrometry. TxB2 concentrations in all volunteers increased with time during blood collection, increases occurred more sporadically in the case of PGE2, PGF2 alpha and 6-keto-PGF1 alpha. Rapid changes in plasma prostanoid concentrations within a sampling series were unpredictable and were inexplicable. The measured plasma prostanoid concentrations apparently depended on the sampling conditions, which could not be adequately standardized and controlled. However, very short term changes in plasma prostanoid levels cannot be excluded.

Determination of ketocyclazocine in human plasma by gas chromatography—negative-ion chemical-ionization mass spectrometry

Determination of ketocyclazocine in human plasma by gas chromatography—negative-ion chemical-ionization mass spectrometry

Sensitive determination of tyrosine metabolites, p-hydroxyphenylacetic acid, 4-hydroxy-3-methoxyphenyl-acetic acid and 4-hydroxy-3-methoxymandelic acid, by gas chromatography—negative-ion chemical-ionization mass spectrometry : Application to a stable isotope-labelled tracer experiment to investigate their metabolism in man

A method has been established for studying the dynamic metabolism of tyrosine to its metabolites in humans using a deuterium-labelled amino acid. Phenylalanine-d 5 was administered orally to human subjects (5 mg/kg) and the levels of p-hydroxyphenylacetic acid-d 4, 4-hydroxy-3-methoxyphenylacetic acid-d 3, and 4-hydroxy-3-methoxymandelic acid-d 3 excreted into urine every hour were determined by gas chromatography—negative-ion chemical-ionization mass spectrometry. This method was also applied to some patients with depression and it was possible to detect a slight alteration in the excretion of some compounds compared with the control.

Incorporation of2H from2H2O into ABA in Tomato Shoots: Evidence for a Large Pool of Precursors

Incorporation of²H from²H₂O into ABA in Tomato Shoots: Evidence for a Large Pool of Precursors

Residue Methodology for AMDRO Fire Ant Insecticide (AC 217,300) in Pasture Grass and Crops

Residue methodology is described for the determination of AC 217,300 residues in pasture grass and crop samples. After extraction and subsequent cleanup on an XAD-2 column, residues of AC 217,300 are determined by liquid chromatography (LC), using a reverse phase paired-ion chromatographic system and detection at 300 nm. The method has a validated limit of sensitivity of 0.05 ppm with corresponding control values for the commodities analyzed of less than 0.01 ppm. Apparent residues over 0.05 ppm can be confirmed by either gas chromatography with an electron capture detector (GC-EC) or gas chromatography-negative ion chemical ionization mass spectrometry (GC-NICI). The direct GC-NICI method circumvents the need for sample cleanup on the XAD-2 column, and offers a greatly simplified procedure that is useful for screening samples. Recoveries of AC 217,300 from the commodities analyzed have been satisfactory with all methods of analysis.

Determination of acetyltrimoprostil and its metabolite trimoprostil in human or dog plasma by gas chromatography—negative-ion chemical ionization mass spectrometry

A sensitive and specific procedure is described for the determination of the antisecretory prostaglandin acetyltrimoprostil and its metabolite trimoprostil in human or dog plasma using gas chromatography—negative-ion chemical ionization mass spectrometry (GC—NICI- MS). Trideuterated analogues of both compounds are added to plasma as the internal standards. The plasma is extracted at pH 7.3 with benzene—dichloromethane (9:1), and the residue of the organic extract is reacted at room temperature with pentafluorobenzyl bromide in the presence of 18-crown-6-ether and potassium acetate. The derivatives are reconstituted in heptane, and appropriate aliquots are analyzed by GC—NICI-MS with selected-ion monitoring of the intense (M - C 6F 5CH 2) − fragment ions of acetyltrimoprostil ( m/ z 419), trimoprostil ( m/ z 377), and their respective trideuterated analogues ( m/ z 422 and m/ z 380, respectively). Quantitation of an experimental plasma sample is based on a comparison of the m/ z 419 versus m/ z 422 and m/ z 377 versus m/ z 380 ion ratios in each sample to that obtained from the analysis of drug-free plasma fortified with various amounts of both protio compounds, and a fixed amount of each trideuterated internal standard. The limit of quantitation of the assay for human plasma is 0.2 ng ml −1 with mean relative standard deviations at this concentration of 15.5% and 9.7% for acetyltrimoprostil and trimoprostil, respectively.

Determination of low plasma concentrations of 3-methoxy-4-hydroxyphenylethylene glycol using gas chromatography—negative-ion chemical ionization mass spectrometry

Gas chromatography—negative-ion chemical ionization mass spectrometry (GC—NICI-MS) allowed the detection of extremely low plasma concentrations of 3-methoxy-4-hydroxy-phenylethylene glycol (MHPG). Glucuronide and sulphate conjugates of MHPG were determined after enzymatic hydrolysis of plasma with β-glucuronidase—arylsulphatase. A 1-ml plasma sample was extracted at the pH of the hydrolysis (pH 4.8) with ethyl acetate, and the dry extract was derivatized with pentafluoropropionic anhydride in ethyl acetate. After evaporation of the solvent, the residue was dissolved in benzene and an aliquot was analysed by GC—NICI-MS. A trideuterated analogue of MHPG was used as an internal standard. Negative-ion chemical ionization of the pentafluoropropionyl derivatives was carried out using ammonia. The ion—molecule adducts at m/e 766 and 785 (MHPG) and m/e 769 and 788 (internal standard) were formed from the pentafluoropropionyl derivatives with the ions of m/e 163 (CF 3,CF 2,COO −) and m/e 144 (loss of fluorine from m/e 163). The concentrations of the ions of m/e 163 and 144 play a major role in the sensitivity and precision of this technique, which allows the detection of free MHPG plasma concentrations as low as 100 pg/ml in routine analysis.

Assay methods for 6-keto-prostaglandin F 1α in human urine : Comparison of chromatographic techniques with radioimmunoassay and gas chromatography—negative-ion chemical-ionization mass spectrometry

6-Keto-prostaglandin F 1α (6-keto-PGF 1α) in human urine is considered to be a reflection of renal prostacyclin production. Because of the large amounts of unidentified eicosanoid metabolites in urine that may potentially bind to 6-keto-PGF 1α antisera, most radioimmunoassays include chromatographic purification of urine. A comparison of chromatographic techniques and of antisera to 6-keto-PGF 1α for the assay of human urine is described. Gas chromatography—negative-ion chemical-ionization mass spectrometry (GC—NICI-MS) was used as the reference method. Radioimmunoassays were performed with each of four antisera combined with each of three chromatographic purification systems (silicic acid, Sephadex LH-20, reversed-phase high-performance liquid chromatography). There was marked variability in the results; however, there was at least one chromatographic method for each antiserum that yielded results comparable to GC—NICI-MS. Direct radioimmunoassay of urine without chromatography yielded markedly elevated and variable results for the four antisera. In contrast, the four antisera gave very similar results with direct assay of media from isolated perfused organs. Thus, for the radioimmunoassay of 6-keto-PGF 1α in human urine, each antiserum is sensitive to different contaminants in urine and must be individually matched to a chromatographic purification system.

Determination of nilvadipine in human plasma by capillary column gas chromatography—negative-ion chemical-ionization mass spectrometry

A highly sensitive and specific method for the determination of nilvadipine, a new dihydropyridine calcium antagonist, in human plasma is described. A deuterated analogue of nilvadipine is added to the plasma as an internal standard. The agent and its internal standard are extracted at pH 9 from the plasma into a benzene— n-hexane (1:1) mixture. The extract is analysed by fused-silica capillary column gas chromatography—negative-ion chemical-ionization mass spectrometry with methane as the reagent gas. The mass spectrometer is set to monitor the negative molecular ions of the agent and internal standard. Quantitation is possible down to 0.01 ng/ml using 1 ml of plasma. The coefficients of variation of the method are 6.4 and 2.1% at the 0.01 and 0.1 ng/ml levels, respectively. Plasma levels obtained with this method are given for four healthy volunteers who had received a 6-mg oral dose of nilvadipine.

Quantitation of cibenzoline in human plasma by gas chromatography—negative-ion chemical-ionization mass spectrometry

Quantitation of cibenzoline in human plasma by gas chromatography—negative-ion chemical-ionization mass spectrometry

Pentafluorobenzyl derivative of histamine for determination by gas chromatography—negative ion chemical ionization mass spectrometry

Pentafluorobenzyl derivative of histamine for determination by gas chromatography—negative ion chemical ionization mass spectrometry

Measurement of Plasma Prostaglandin E2 Using Capillary Gas Chromatography Negative Ion Chemical Ionization Mass Spectrometry

A stable isotope dilution assay for the measurement of plasma prostaglandin E2 (PGE2) employing capillary column gas chromatography-negative ion chemical ionization mass spectrometry (GC-NICIMS) is described. PGE2 was extracted from plasma using C18 and silica SEP-PAKS. Further purification and separation was accomplished by thin layer chromatography. The prostaglandin was analyzed as its pentafluorobenzyl ester-methoxime-trimethyl-silyl ether, using fragment ions at m/e 524 (protium) and m/e 528 (deuterium) for quantitation. The mean plasma concentration of PGE2 determined in 8 healthy volunteers was 2.8 +/- 2.0 pg/ml.

Determination of Organobromine Compounds in Water by Gas Chromatography-Negative Ion Chemical Ionization Mass Spectrometry

Gas chromatography-negative ion chemical ionization mass spectrometry (GC-NCIMS) was evaluated as a method for qualitative and quantitative determination of organobromine compounds in water. The negative ion mass spectra of bromine containing trihalomethanes (CHBrCl2, CHBr2Cl and CHBr3) consisted of two peaks of m/z 79 and 81, corresponding to mass numbers of bromine atom. The detection limit for these compounds was 1.0 ppb. The results in this study showed that organobromine compounds in water could be assayed by GC-NCIMS.