Chemical Ionization Mass Fragmentography Research Articles

Isotope dilution ammonia chemical ionization mass fragmentographic analysis of urinary 3-O-methylated catecholamine metabolites: Rapid sample clean-up by derivatization and extraction of lyophilized samples

We developed a method for simultaneous quantification of the urinary 3-O-methylated catecholamine metabolites 3-methoxytyramine, normetanephrine and metanephrine by stable isotope-dilution ammonia chemical ionization mass fragmentography. Prepurification of lyophilized samples was done by simultaneous deconjugation and pentafluoropropionylation, followed by extraction and rederivatization. Compared with our previously described method, based on acid hydrolysis, alkaline extraction, derivatization and electron-impact mass fragmentography, the present method was found to be less laborious, more sensitive and presumably more accurate. New urinary excretion values were established for apparently healthy adults. The present prepurification method may prove applicable for profiling of a variety of naturally occurring mono-, di- and polyamines in biological samples.

Stable isotope dilution analysis of very long chain fatty acids in plasma, urine and amniotic fluid by electron capture negative ion mass fragmentography

A sensitive and selective stable isotope dilution electron capture negative ion chemical ionization mass fragmentography method applying pentafluorobenzyl derivatives was developed for the accurate quantitation of very long chain fatty acids. This technique allowed detection of 1–5 pg of each compound and was applied to plasma (100 μl), amniotic fluid (1 ml) and urine (1 ml). Normal concentrations were established and the concentrations in samples of selected patients with classified peroxisomal disorders were determined. In plasma samples of all patients the C26:0/C22:0 ratios were elevated (range 0.03–0.43), compared to the control ratios (range 0.003–0.021). The ratio C26:0/C22:0 was elevated in four of five amniotic fluid samples from fetuses with peroxisomal disorders (range 0.18–0.54) when compared with controls (range 0.05–0.25). An elevation of the ratio C26:1/C22:0 was observed in all five amniotic fluid samples (range 0.22–0.60 vs. 0–0.08 in controls). Urinary C26:0 concentrations were lower than in plasma and amniotic fluid and diagnostic ratios were not elevated in patients with peroxisomal disorders.

Determination of plasma bile acids by capillary gas-liquid chromatography-electron capture negative chemical ionization mass fragmentography.

Combined capillary gas-liquid chromatography-electron capture negative chemical ionization mass spectrometry of pentafluorobenzyl ester-TMSi ether derivatives of bile acids and isotope dilution using deuterated internal standards are introduced as a sensitive and selective analysis technique for plasma bile acids. As a result of the high ionization efficiency of pentafluorobenzyl derivatives under electron capturing conditions and minimal fragmentation, the detection limit of this technique is low: 1 pg for each bile acid. The high sensitivity enabled the detection and quantitation of atypical bile acids in 200-microliters aliquots of plasma from fasting healthy adults as exemplified by trihydroxycoprostanic acid (0.002 +/- 0.001 mumol/l) and dihydroxycoprostanic acid (0.013 +/- 0.002 mumol/l).

Detection of methamphetamine and amphetamine in a skeletonized body buried for 5 years

A 28-year-old male methamphetamine abuser, who had been buried for 5 years after being killed by strangulation, was found skeletonized. Methamphetamine and amphetamine in the significantly denatured fatty material of the bone marrow were analyzed by gas chromatography and gas chromatography-mass spectrometry. The confirmation of the chemicals was carried out by chemical ionization (CI) mass chromatography, CI mass spectrometry and CI mass fragmentography. The concentrations of methamphetamine and amphetamine determined by CI mass fragmentography were 1.0 μmol/100 g and 0.1 μmol/100 g, respectively. The method used would seem to be very useful for determination of methamphetamine and amphetamine in marked putrefied biological materials.

A specific and sensitive determination of gamma aminobutyric acid in CSF and brain tissue by gas chromatography-mass spectrometry

The determination of γ-aminobutyric acid (GABA) in cerebrospinal fluid and brain extracts is described. Its heptafluorobutyryl-isobutanol derivative was measured both by electron impact and chemical ionization mass fragmentography using GABA-d 6 as internal standard. The derivatization product is stable for several days. The method is sensitive (1 ng absolute in cerebrospinal fluid and 30 pg in standard GABA solutions) and specific, when chemical ionization mode is applied. Normal values of GABA are in rat brain extracts (1.40±0.32 μmol/g fresh weight) and human CSF (18.3±10.0 ng/ml).

A fatal methamphetamine poisoning associated with hyperpyrexia

After self-administration of 0.05g of methamphetamine hydrochloride intravenously on three occasions at intervals of 3h, a 25-year-old female methamphetamine abuser ingested approximately 1.5 g of methamphetamine hydrochloride, and was found dead 3–4 h later. Complete rigor mortis was observed 1–2 h after death and the rectal temperature was 38.4°C 3–4 h after death. Distribution of methamphetamine and amphetamine in the body was analyzed by chemical ionization mass fragmentography. Amphetamine/methamphetamine concentrations (μ mol/100 g) were 0.26 28.8 in blood, 0.64 68.2 in brain, 0.96 117.1 in liver, 0.53 50.6 in kidney, and 1.49 1045 in stomach contents. Total amount of methamphetamine hydrochloride in stomach contents was 11.6mg. Amphetamine in tissues was a metabolite of methamphetamine, and amphetamine in stomach contents resulted from excretion into saliva and gastric mucous excretion. With rectal temperature at death estimated at more than 41°C, it would seem that hyperpyrexia played an important role in causing death from methamphetamine poisoning.

Detection and measurement of opium alkaloids and metabolites in urine of opium eaters by methane chemical ionization mass fragmentography

A gas chromatographic—mass spectrometric assay for eight opium alkaloids in human urine following opium ingestion is described. The compounds were extracted from urine with methylene chloride—isopropanol (7:3, v/v) at pH 9.5, evaporated, derivatized with Tri-Sil Z and analyzed by methane chemical ionization mass fragmentography. The method is sensitive to ca. 0.01 μg/ml for morphine and codeine and ca. 0.05 μg/ml for the other compounds. Adsorption problems on the gas chromatography column prevented obtaining reproducible results for the measurement of noscapine. Extraction efficiencies over the pH range of 8–11 for the eight compounds are reported. Retention times of the opium alkaloids were determined using five different liquid phases (3%) on Gas-Chrom Q (100–120 mesh) and two column lengths (36 cm and 183 cm). The 36-cm column packed with OV-210 was selected for use in the assay. Ions were selected for monitoring for each component from their methane chemical ionization spectrum to provide the needed sensitivity and specificity for analysis of a multi-component mixture. The assay was used for the analysis of an “opium eater's” urine. Morphine, codeine, nomorphine, norcodeine and noscapine were detected; however, no evidence was obtained for thebaine, papaverine or oripavine. Unconjugated morphine (0.64 μg/ml) was present at nearly twice the concentration of codeine (0.37 μg/ml) and normorphine and norcodeine were present in equal amounts (ca. 0.15 μg/ml).

Simultaneous measurement of acetylcholine and choline in brain by pyrolysis-gas chromatography-mass spectrometry

Pyrolysis-gas chromatography and chemical ionization mass fragmentography were combined to develop a specific, simple and rapid method for simultaneously measuring endogenous and stable isotopic variants of acetylcholine and choline with a detection limit of approxiamtely 10 −12 mol. The recovery and reproductivity of the method are excellent, and the method is suitable for measuring acetylcholine and choline in discrete regions of rat brain and to measure incorporation of choline into acetylcholine, both of which uses are demonstrated. This method affords easy analysis of 40 samples in a working day. The new technique used to extract compounds from tissues and the modified gas flow arrangement may be useful to measure other compounds as well.

Enhancement of the Yu and Ledeen gas-liquid chromatographic method for sialic acid estimation: use of methane chemical ionization mass fragmentography

The sialic acid present in erythrocyte ghosts was estimated by methane chemical ionization mass fragmentography of the trimethylsilyl methyl glycosides. The internal standard was 3,4,6-tris-trimethylsilyl-alpha-phenyl-2-deoxy-2-acetamide-D-glucosaminide, as proposed by Yu and Ledeen (J. Lipid Res. 1970. 11: 506-516). The [MH-16]+ions (m/e 610 for the TMS-methylglycoside of N-acetylneuraminic acid and m/e 498 for the internal standard) were used for quantifying nanogram levels of sialic acid in the presence of other contaminating substances. This effectively raises the signal-to-noise ratio for the Yu and Ledeen method by at least two orders of magnitude. The sensitivity and linearity of the method, without use of isotopic carriers, were tested using known quantities of N-acetylneuraminic acid. The limit of detection was below 0.4 nanograms (approximately one picomole). The useful range of detection was 10 ng-1 microgram, showing a large dynamic range.

Determination of N-Nitrosodimethylamine in Human Urine by Chemical Ionization-Mass Fragmentography

N-Nitrosodimethylamine (NDMA) in human urine was determined by chemical ionization-mass fragmentography (CI-MF), and the results were compared with those of thermal energy analyzer (TEA). Urine samples were extracted with dichloromethane and the concentrate was cleaned up on a silica gel column. CI-MF was very sensitive (detection limit was 1.2 pg at S/N=2) and the selectivity was comparable with that of TEA. NDMA levels in urine were ND-0.14 ppb. The values determined by CI-MF were similar to those by TEA, and this method was applicable to trace analysis of urinary NDMA with high sensitivity (above 0.002ppb).

Simultaneous determination of endogenous norepinephrine and dopamine-β-hydroxylase activity in biological materials by chemical ionization mass fragmentography

A new clean-up procedure is described for the quantitation of biogenic amines and their metabolites by gas chromatography-chemical ionization mass fragmentography. The procedure is carried out in an acidic, non-aqueous medium. The metabolites as well as the amines are extracted with greater than 92% recovery. Using a CPG-10 column, the procedure is most suitable for the purification of these amines and their metabolites from the extract. It has been applied to the simultaneous determination of endogenous norepinephrine and dopamine-β-hydroxylase (DBH) activity in rat adrenal by use of [ 2H 2]dopamine as a substrate and [ 2H 6]-norepinephrine as an internal standard. The quantitation limit of the DBH activity is 0.1 pmole/mg of rat adrenal per hour.

Assessment of serum cholesterol by two methods: Gas—liquid chromatography on a capillary column and chemical ionization-mass fragmentography with isotopic dilution of [3,4- 13C]cholesterol as internal standard

A gas—liquid chromatographic (GLC) method and an isotopic dilution—mass fragmentographic (ID—MF) procedure using the same capillary chromatographic separation are described for serum cholesterol assay. GLC included silylation and separation on a highly efficient glass capillary column which allowed the separation of cholesterol from cholestanol and the use of epicoprostanol as internal standard. The concentrations were calculated from the areas of the signals and digitalized by a reporting integrator. The reproducibility was 0.5% and the correlation with the ID—MF technique was 0.997. The ID—MF technique was characterized by the use of [3,4- 13C]cholesterol as the labelled standard and a chemical ionization mode. The reproducibility was 0.8%.

Quantitative analysis of berberine in urine samples by chemical ionization mass fragmentography

A highly specific and sensitive method has been developed for the quantitative determination of berberine in human urine. In order to carry out the microdetermination of berberine by chemical ionization mass fragmentography, berberine was reduced with sodium borohydride in methanol to tetrahydroberberine and subjected to gas chromatography—mass spectrometry. Berberine concentrations as low as 1 ng/ml urine can be measured by this method, with [ 2H 3]berberine chloride as an internal standard.

Determination of Terbutaline in Postmortem Human Tissues by Gas Chromatography-Mass Spectrometry

A method is described for the quantitative analysis of terbutaline in human tissues. Gas chromatography and chemical ionization mass fragmentography are used for selective and sensitive detection. A detection limit of 1.5 ng of the free drug per gram tissue is observed when 0.5 g of the dry tissue is used. No interference from the major metabolite, the sulphate conjugate of terbutaline, is found. Finally the method is applied to the postmortem tissues of two persons who used terbutaline frequently.

Differentiation of Xylene Isomers by Chemical Ionization Mass Fragmentography

Three kind of isomers of xylene and ethylbenzene were analyzed using a gas chromatogrph-chemical ionization mass spectrometer with mass fragmentograhy technique to find any difference between these isomers in the various conditions, such as different reagent gases(CH4, i-C4H10 and NH3)and ion source temperature(200°C∼280°C). With iso-buthane as a reagent gas, the peak intensity of m/e 107(QM+)and the peak intensity ratio of m/e 106 to m/e 107 show remarkable differences between m-xylene and the other isomers at 200°C of the ion source temperature, and the differences decrease with increasing temperature. With ammonia as a reagent gas, the peak intensity of m/e 106 and the peak intensity ratio of m/e 108 to m/e 106 show much differences between p-xylene and the other isomers, and the differences are unchanged by the ion source temperature 200°∼280°C. Peak intensity ratio of m/e 105 to m/e 107 of ethylbenzene is much higher than the one of xylene isomers with iso-buthane as a reagent gas at 200°C of ion source temperature, and the differences decrease with increasing temperature.

Determination of 1-β- d-ribofuranosyl-1,2,4-triazole-3-carboxamide (virazole) in blood and urine by chemical ionization-mass fragmentography

A combined gas chromatographic-mass spectrometric technique is described for the quantification of virazole in serum and urine. Proteins are removed by molecular filtration, lipids by extraction with dichloromethane and interfering endogenous constituents by acidic and basic ion-exchange resins. Virazole is quantified by monitoring the protonated molecular ions of the fully silylated derivatives of virazole ( m/e 533) and the arabinose analog (internal standard) obtained by methane chemical ionization. The detection limit is 150 pg (0.6·10 −12 mole) of virazole injected. In serum 10 ng/ml (4·10 −8 mole) can be detected, 25 ng/ml quantified. In urine 0.5 μg/ml can be quantified without preconcentration. Virazole was detected in serum for at least 96 h at the 70-ng/ml level.

Simultaneous measurement of secondary and tertiary tricyclic antidepressants by GC/MS chemical ionization mass fragmentography.

We present a method for measuring seven commonly used tricyclic antidepressants in plasma. This method is suitable for monitoring therapeutic concentrations and for screening drug overdoses in cases where the identity of the abused tricyclic antidepressant may not be known. Drugs from alkalinized plasma are extracted into hexane in one step; two injections into the gas chromatograph/mass spectrometer follow. The tertiary amines (amitriptyline, doxepin, and imipramine) are analyzed by direct injection; the secondary amines (nortriptyline, desmethyldoxepin, desipramine, and protriptyline) are analyzed after derivitization with trifluoroacetic anhydride. Clomipramine and desmethyltrimipramine are suitable internal standards. Chemical ionization mass fragmentography, with methane as the reactant gas, is used. While maintaining specificity for these drugs, concentrations in human plasma ranging from 5 to 500 microng/liter can be measured. The coefficients of variation are about 4 to 11%.

Quantitative determination of sulpyrid in biological samples from rats by gas-liquid chromatography and chemical ionization-mass fragmentography

Quantitative determination of sulpyrid in biological samples from rats by gas-liquid chromatography and chemical ionization-mass fragmentography

Quantitation of glycine in plasma and urine by chemical ionization mass ofragmentography

A new technique has been developed for rapid and precise quantitation of glycine in micro samples of biological fluids. Glycine is measured as the n-butyl N-trifluoroacetyl (TAB) derivative by chemical ionization mass fragmentography, using both methane and isobutane as carrier/reagent gases, with the analogous derivative of 2,2-dideuterioglycine added as the internal standard. This technique is also readily applicable, with sensitivity to the femtomole level, to determinations of other amino acids, as, for example, in studies of congenital errors of metabolism.

Mass fragmentographic determination of d- and l-chloropheniramine with aid of the stable isotope technique.

To investigate the pharmacokinetics of d-chlorpheniramine (d-CPA) and its l-isomer, d-CPA-d6 and l-CPA-d0 were administered orally and simultaneously to healthy adult volunteers. Both isomers in the serum were measured by chemical ionization-mass fragmentography using dl-CPA-d13 as an internal standard. The concentration of d-isomer was higher than that of l-isomer and the ratio (d/l) of both isomers was 1.9-3.1, and the biological half-lives of d- and l-isomer were 24 hr and 15 hr, respectively.