Computerized Gas Chromatography-mass Spectrometry Research Articles

Identification and quantitative determination of steroids in bovine corpus luteum during oestrous cycle and pregnancy.

Neutral steroids in bovine corpus luteum were isolated by liquid-gel chromatography on hydrophobic Sephadex, and were analyzed by computerized gas chromatography-mass spectrometry. The presence of progesterone and 20beta-hydroxy-4-pregnen-3-one was confirmed. In addition, 3beta-hydroxy-5-pregnen-20-one, 5-pregnene-3beta,20beta-diol, 3beta-hydroxy-5alpha-pregnan-20-one and 5alpha-pregnane-3beta,20beta-diol were fully identified, and 3-hydroxy-4-pregnen-20-one, 4 pregnene-3,20-diol, 22-hydroxycholesterol and 20,22-dihydroxycholesterol were partially characterized. Steroid sulphates were not detected. Quantification of the six fully identified steroids was based on peak areas in specific fragment ion current chromatograms constructed by the computer. During the 4th-19th day of the oestrous cycle the steroid concentrations varied as follows: progesterone 6.0-36.7 mug/g wet luteal weight, 20beta-hydroxy-4-pregnen-3-one 0.8-5.5 mug/g, 3beta-hydroxy-5-pregnen-20-one 1.0-7.1mug/g, 5-pregnene-3beta,20beta-diol smaller than 0.2-0.9 mug/g, 3beta-hydroxy-5alpha-pregnan-20-one 1.7-8.6 mug/g, and 5alpha-pregnane-3beta,20beta-diol smaller than 0.2-1.2 mug/g. The concentrations of progesterone and 3beta-hydroxy-5-pregnen-20-one seemed to vary in parallel and were low during days 11-17. During this period the concentrations of 5-pregnene-3beta,20beta-diol and 5alpha-pregnane-3beta,20beta-diol were highest as was the relative contribution of all three 20beta-hydroxysteroids to the total amount of steroids. The relative amount of 3beta-hydroxy-5alpha-pregnan-20-one seemed to be highest during days 4-6. The total steroid concentration in corpora lutea taken in early pregnancy (75-105 days) was 18-47 mug/g. In the period 75-90 days, progesterone constituted only 35-42% of the total steroids, 3beta-hydroxy-5alpha-pregnan-20-one as much as 23-40% and the total 20beta-hydroxysteroids 18-30%. The total steroid concentration in corpora lutea taken in midterm and late pregnancy was 21-77 mug/g. In this period progesterone was by far the predominant steroid and constituted about 80-90% of the total steroids in corpora lutea taken between days 150 and 240. Possible correlations between luteal growth, steroid oxidoreductases and steroid concentrations are discussed.

OXIDATIVE DEGRADATION OF HUMIC AND FULVIC ACIDS EXTRACTED FROM TROPICAL VOLCANIC SOILS

To obtain a better understanding of the unusual accumulation of organic matter in tropical volcanic soils in the West Indies, humic and fulvic acids were extracted with 0.5 N NaOH under N2 from four surface and two subsurface horizons of four such soils from the island of Dominica and degraded by KMnO4 oxidation of unmethylated and methylated materials. The oxidation products were fractionated by solvent extraction and chromatographic methods and 52 compounds were identified on a gas chromatography–mass spectrometry–computer system. Major oxidation products (identified as esters and ethers) were benzenecarboxylic, phenolic and aliphatic mono- and dicarboxylic acids. Smaller amounts of n-alkanes, furan derivatives and dialkyl phthalates were also identified. The major chemical structures detected in the tropical volcanic humic and fulvic acids were aromatic rings substituted by: (a) three to six C atoms; (b) one OCH3 group and three, four and six C atoms; and (c) two OH groups and one, two, three and five C atoms. Judging from the qualitative and quantitative distribution of the major oxidation products, the chemical structure of tropical volcanic humic and fulvic acids did not appear to differ significantly from that of humic and fulvic acids extracted from soils from widely differing climatic zones. We were unable to detect any effect of depth of sampling or soil pH on the chemical structure of the humic materials extracted from the tropical volcanic soils.

Specific deuterium labelling and computerized gas chromatography -- mass spectrometry in studies on the metabolism in vivo of a steroid sulphate in the rat.

The metabolism of 3beta-hydroxy-5alpha-pregnan-20-one sulphate was studied in bile fistula rats and in isolated perfused livers. Computerized gas chromatography--mass spectrometry, in combination with specific deuterium-labelling, was employed to follow the metabolic transformations. Male animals excreted metabolites into bile more rapidly than females, a finding which could be correlated with the preferential formation of glucuronide conjugates in the male liver. The major metabolic pathway in male rats involved the steps: hydrolysis, 2alpha-hydroxylation, oxidoreduction at C-3 and glucuronide conjugation, yielding 2alpha, 3alpha-dihydroxy-5alpha-pregnan-20-one glucuronide as the major metabolite. Only traces of the injected steroid sulphate were detected in bile from male animals. In contrast, the administered compound was the major steroid excreted in bile of female rats, where the main metabolite was identified as 3beta,15beta-dihydroxy-5alpha-pregnan-20-one sulphate. A minor metabolite, 3beta,16alpha-dihydroxy-5alpha-pregnan-20-one, was found as a monosulphate in female rats and as both a disulphate and a glucuronide conjugate in male rats. The deuterium content of the sulphated 15beta-and 16alpha-hydroxylated metabolites was consistent with metabolic pathways involving direct hydroxylation of the injected steroid sulphate. The results obtained from the liver perfusions were essentially the same as those from the experiments with bile fistula animals. This indicates that all the observed metabolic reactions took place in the liver.

Use of High-Resolution Open Tubular Glass Capillary Columns to Separate Acidic Metabolites in Urine

We used high-resolution glass capillary columns to study the trimethylsilyl derivatives of some acidic metabolites found in pooled urine specimens form control and postpartum subjects. About 30 compounds were identified by gas chromatography-mass spectrometry-computer techniques. In general, open tubular capillary columns effect better resolution of trimethylsilyl derivatives of organic acids than do conventional packed columns. GESE-30 proved to be a good general-purpose stationary phase, whereas OV-17 offered certain advantages in resolving aromatic acid components.

The recognition of organic pollutants in aquatic sediments.

Aquatic sediments contain small amounts of organic matter which have been contributed from the land by water and wind erosion, from the water column, and from the sediment itself by biological activity. These fluxes include variable loads of pollutants and their degradation products. Base-line studies of the organic components of contemporary surface sediments and of layers from cores are urgently needed so that the pollutants can be distinguished from the natural compounds. Such analyses require precise analytical procedures capable of identifying pollutants and natural compounds, both expected and unexpected. Computerized gas chromatography-mass spectrometry provides this capability and the option of further processing the stored data long after acquisition. The ways in which computers can be used to process mass spectral data are discussed and some applications are described. The use of one advanced computerized gas chromatography-mass spectroscopy system for the analysis of an estuarine sediment is described in detail. The presence of phthalate esters, polynuclear aromatic hydrocarbons and partially degraded crude oil, as well as certain natural lipids, is demonstrated.

Analysis of steroids by liquid-gel chromatography and computerized gas chromatography-mass spectrometry

Work carried out in this laboratory aiming at the development of generally applicable methods for analysis of steroids in biological materials is reviewed. Isolation and separation of groups of steroids is achieved by chromatography in miscible solvent systems on neutral or ion exchanging lipophilic derivatives of Sephadex. Reversed or straight-phase systems can be designed to obtain columns which act as filters to remove unwanted material. The hydrolyzed steroids are converted to O-methyloxime-trimethylsilyl ether derivatives which are purified by liquid-gel chromatography and analyzed by computerized gas chromatography-mass spectrometry. Repetitive magnetic scanning of the entire mass range or repetitive accelerating voltage scanning of a limited mass range is used to monitor individual fragment ion currents. Spectra are recorded on magnetic tape, and peaks in fragment ion current chromatograms are located by the computer which performs a preliminary search for potential molecular ions in spectra taken at the located peaks. Quantitative determinations are based on areas of peaks with correct retention time in chromatograms based on the current of specific fragment ions. Similar methods are used to determine the number and abundance of heavy atoms in steroids formed from precursors labelled with stable isotopes and in metabolites formed from steroids labelled with stable isotopes in specific positions.

Plasma amino acid analysis by isotope ratio gas chromatography mass spectrometry computer techniques.

A routine analysis for micro samples of plasma amino acids by a gas chromatography mass spectrometry computer system has been developed. Isotope ratio determination was used as the quantitating technique via multiple internal standards. The speed of the analysis is increased by omitting the ion exchange purification step and its flexibility is maximized by using repetitive scanning rather than selective ion monitoring. This procedure yields excellent precision and accuracy, as demonstrated by the analysis of a known amino acid mixture and of neonatal plasma.

Utilization of the carbon and hydrogen atoms of ethanol in the biosynthesis of steroids and bile acids.

[1‐2H2]Ethanol, [2‐2H3]ethanol and [1‐13C, 1‐2H2]ethanol were administered to bile fistula rats under conditions ensuring a maximal rate of oxidation for about 24 h. Cholesterol, cholic acid, chenodeoxycholic acid and 3β,11β,21‐trihydroxy‐5α‐pregnan‐20‐one were isolated from bile collected at intervals during the experiment. The fraction of molecules containing different numbers of heavy isotope atoms was measured by gas chromatography–mass spectrometry–computer techniques. The results were used to determine the origin of the cholesterol pools used as precursors for the different compounds isolated from bile.The bile acids were found to be formed from a cholesterol pool, the turnover of which was about 5 times more rapid than that of the cholesterol pool used for biliary excretion. The cholesterol pools used for biliary excretion and as corticosterone precursor seemed to be in equilibrium.The acetyl groups derived from ethanol were introduced into the steroids without exchange of hydrogens at C‐2, and the acetyl‐CoA pool used as precursor did not exchange with acetyl groups in e.g. citrate or fatty acids. A difference in incorporation of [2‐2H3]acetyl groups and [1‐13C]acetyl groups was interpreted to be due to an isotope effect. Ethanol contributed about 45% of the acetyl‐CoA pool used as precursor of cholesterol and bile acids.Deuterium from [1‐2H2]ethanol was incorporated in many positions of all the steroids. If it is assumed that the incorporation is mediated by NADPH, the deuterium excess of the hydrogens introduced was about 10 atoms%. The contribution via different pathways of an important part of the hydrogen and carbon atoms in cholesterol, bile acids and corticosteroids provides possible mechanisms by which ethanol may affect the biosynthesis and metabolism of steroids.

Separation and computerized gas chromatography-mass spectrometry of unconjugated neutral steroids in plasma

A method is described for analysis of unconjugated neutral steroids in plasma. It is based on extraction of the sample with Amberlite XAD-2 at elevated temperature, purification of the extract on lipophilic ion exchanging and neutral Sephadex derivatives, and gas chromatography-mass spectrometry-computer analysis of O-methyloxime-trimethylsilyl ether derivatives purified on hydrophobic Sephadex. Examples of the application of the method to analysis of steroids in plasma from pregnant women are given.

Analysis of tissue steroids by liquid-gel chromatography and computerized gas chromatography-mass spectrometry.

A method for analysis of unconjugated neutral steroids of low and medium polarity in tissues is described. Nonpolar lipids in the tissue extract are removed by reversed phase chromatography on hydroxyalkoxyalkyl Sephadex. Further purification is obtained by straight phase chromatography using the same gel. Acidic contaminants are removed by filtration of a reaction mixture with methoxyamine hydrochloride through the anion exchanger Amberlyst A-26. The steroids are then analyzed as MO-TMS derivatives using repetitive scanning gas chromatography-mass spectrometry. Spectra are recorded on magnetic tape and quantification of steroids is based on peak areas in specific fragment ion current chromatograms.

Analysis of molecular species of 2H-labelled phosphatidylcholines by liquid-gel chromatography and gas chromatography-mass spectrometry

A procedure has been developed for the study of deuterium incorporation into fatty acid and glycerol moieties of individual molecular species of phosphatidylcholines. The phosphatidylcholines are isolated by chromatography on a lipophilic Sephadex anion exchanger and are hydrolyzed with phospholipase C. The 1,2-diglycerides are separated as trimethylsilyl ethers by reversed-phase chromatography on hydroxyalkoxypropyi Sephadex. The deuterium excess of the glycerol and fatty acid moieties of the separated trimethylsilyl ethers is determined by computerized gas chromatography-mass spectrometry using repetitive magnetic scanning of whole spectra or repetitive accelerating voltage scanning of a limited mass range. The trimethylsilyl ethers are then hydrolyzed and 3-perdeuteriotrimethylsilyl ethers are prepared which are separated into individual molecular species by preparative gas chromatography. The 2-acylsn-glycerol 3-perdeuteriotrimethylsilyl ethers are then obtained by brief lipase digestion and isolation by rapid straight-phase chromatography on hydroxyalkoxypropyl Sephadex. The deuterium excess at different carbons of the glycerol moiety is determined by gas Chromatographic—mass spectrometric analysis of the 2-acyl- sn-glycerol 1-trimethylsily-3-perdeuteriotrimethylsily ethers. The analyses can be carried out with 5–400 μg of individual phosphatidyl-cholines in the original extract. Examples of the analysis of phosphatidylcholines in bile of bile fistula rats given [1,1- 2H 2]ethanol are given.

Identification of chlorinated styrenes in cormorant tissues by a computerized gas chromatography-mass spectrometry system

The results of a study concerning the identification of three unknown compounds which were detected in extracts of cormorant tissue are reported. From these results it has been concluded that one of the compounds is octachlorostyrene. It is highly probable that the other two compounds are heptachlorostyrenes. Octachlorostyrene was also detected in fish, seals and various other aquatic bird species.

Analysis of steroids by off-line computerized gas chromatography-mass spectrometry.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTAnalysis of steroids by off-line computerized gas chromatography-mass spectrometryRobert. Reimendal and Jan. SjovallCite this: Anal. Chem. 1972, 44, 1, 21–29Publication Date (Print):January 1, 1972Publication History Published online1 May 2002Published inissue 1 January 1972https://doi.org/10.1021/ac60309a014RIGHTS & PERMISSIONSArticle Views77Altmetric-Citations107LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (825 KB) Get e-Alerts Get e-Alerts