Metabolite identification by liquid chromatography-mass spectrometry

Metabolite identification (Met ID) is important during the early stages of drug discovery and development, as the metabolic products may be pharmacologically active or toxic in nature. Liquid chromatography-mass spectrometry (LC-MS) has a towering role in metabolism research.This review discusses current approaches and recent advances in using LC-MS for Met ID. We critically assess and compare various mass spectrometers, highlighting their strengths and limitations. Citing appropriate examples, we cover recent LC and ion sources, isotopic-pattern matching, hydrogen/deuterium-exchange MS, data dependent analyses, MS^E, mass defect filter, 2D and 3D approaches for the elucidation of molecular formula, polarity switching, and background-subtraction and noise-reduction algorithms. A flow chart outlines a comprehensive strategy for Met ID, including a focus on reactive metabolites.     

高温气相色谱和高温气相色谱-质谱法对费托合成蜡的分离和鉴定

费托合成蜡是费托合成反应中的重要产物之一。采用高温气相色谱与冷柱头进样相结合的方式,建立了一种分离分析费托合成蜡的气相色谱方法。该方法无需对费托合成蜡进行预处理,使用氦气为载气,选用更长的高温色谱柱,具有平稳的色谱基线,对费托合成蜡中正构烷烃和其他未知组分有很好的分离效果,能够洗脱费托合成蜡中碳数大于C₉₀的重组分。用高温气相色谱-质谱法对费托合成蜡馏分进行定性分析,其组分有烷烃、烯烃和含氧化合物。该方法对了解费托合成蜡组分的详细信息和费托合成工艺的开发有重要意义。     

Trace identification of plant substances by combining gas chromatography-mass spectrometry and direct deposition gas chromatography-Fourier transform infrared spectrometry

The advantages of combined gas chromatography-mass spectrometry and direct deposition gas chromatography-Fourier transform infrared spectrometry at the same level of sensitivity are demonstrated using an apolar capillary column. This technique was applied to allelochemicals present in minute concentrations. The identification of volatiles in green leaves and an alkaloid is described.     

Detection and identification of thyreostats in the thyroid gland by gas chromatography-mass spectrometry

Because thyreostatic compounds, also named thyreostats, are banned in Europe (directive 86/469/EEC), methods have to be developed to prevent the illegal use of these substances. The analytical procedure described herein involves the detection and identification at the low ng g⁻¹ level of the main thyreostats known to be used for growth promotion by gas chromatography coupled to mass spectrometry (GC-MS). The assay is based on a liquid/liquid extraction of the thyroid gland, derivatization with pentafluorobenzyl bromide (PFBBr), purification on a silica solid phase extraction column and finally a trimethylsilylation prior to GC-MS. Good thyreostat recoveries were obtained (from 40% to 70%) as well as at acceptable repeatability. The target analytes were detectable below the 1 ng g⁻¹ level on a quadrupole mass spectrometer with negative chemical ionization (NCI) using ammonia as reagent gas and the selected ion monitoring (SIM) acquisition mode. This limit of detection was also reached in the SIM high resolution mode. An improved specificity (more diagnostic ions) was obtained under electronic impact (EI) conditions and positive chemical ionization (PCI) with methane as reagent gas. Identification of thyreostats according to the EU (European Union) criteria (93/256/EEC decision) was made on the basis of two independent GC-MS techniques; the limit of identification was close to 5 ng g⁻¹ for most thyreostats, which represents a real improvement for their control.     

Detection of soil pollution by hydrocarbons using headspace-mass spectrometry and identification of compounds by headspace-fast gas chromatography-mass spectrometry

The direct coupling of a headspace sampler with a mass spectrometer is proposed as a screening tool for the rapid detection of soil pollution by hydrocarbons from petroleum and derivatives. The samples are subjected to the headspace generation process, with no prior treatment, and the volatiles generated are introduced directly into the mass spectrometer, thereby obtaining a fingerprint of the sample analysed. Suitable treatment of the signal by chemometric techniques allows unequivocal characterisation of the different types of sample. The use of fast gas chromatography with a mass spectrometer detector coupled to the headspace sampler allows identification of the major hydrocarbons present in the mineral and organic polluted samples, interpretation of the results obtained, and demonstrates the analytical potential of headspace-mass spectrometry coupling.     

GC-MS直接测定芥酸酰胺

通常衍生后的酰胺才能用气相色谱测定.在实验几种酰胺衍生化方法时发现衍生反应不彻底剩余的芥酸酰胺也能出峰.进一步实验表明:芥酸酰胺在295 ℃气化后可直接测定,该法以邻苯二甲酸二丁酯为内标,质量浓度在115.4～923.2 mg/L 范围内线性关系良好,样品回收率94.85%～98.04%,芥酸酰胺的检出限5.77 mg/L.     

Characterization of archaeological frankincense by gas chromatography-mass spectrometry

A simple gas chromatography-mass spectrometry (GC-MS) method has been developed for the characterization of frankincense in archaeological samples. After trimethylsilylation of the methanolic extract, 15 triterpenoids have been found among the chemical constituents of commercial olibanum (alpha-boswellic acid, 3-O-acetyl-alpha-boswellic acid, beta-boswellic acid, 3-O-acetyl-beta-boswellic acid, alpha-amyrin, beta-amyrin, lupeol, 3-epi-beta-amyrin, 3-epi-beta-amyrin, 3-epi-lupeol, alpha-amyrenone, beta-amyrenone, lupenone, 3alpha-hydroxy-lup-20(29)-en-24-oic acid and 3-O-acetyl-hydroxy-lup-20(29)-en-24-oic acid). These compounds have been unequivocally identified by retention time and mass spectral comparison with pure standards previously isolated, for the most part, in our laboratory. Within these triterpenes, acid ones, the corresponding O-acetates, and their products of degradation were found to be characteristic of frankincense (Boswellia resin). The presence of these unusual triterpenic compounds in an archaeological resinous sample, recovered during excavations from Dahshour site (Egypt, XIIth Dynasty), enabled us to identify unambiguously frankincense resin among several other materials. Additional chromatographic peaks of this sample were assigned to broad chemical classes using retention time and mass spectra features.     

Determination of selenoamino acids by gas chromatography-mass spectrometry

The application of the recently introduced ethylchloroformate derivatization method for the separation and determination of selenomethionine and selenocystein in selenium-enriched yeast and yeast-free tablets by means of a gas chromatography-mass spectrometry (GC-MS) system has been studied. The efficiency of three methods for the extraction of selenomethionine from the tablets were compared. Total selenium content of the same tablets were measured using inductively coupled plasma (ICP)-MS and it was found that in the selenized yeast tablets about 80% of the total selenium is present as selenomethionine. The results were in agreement with the values in the labels and with the literature. The accuracy of the total selenium analysis was controlled by the analysis of a reference material.     

Determination of ink photoinitiators in packaged beverages by gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry

A new analytical method, using gas chromatography-mass spectrometry (GC/MS) and liquid chromatography-mass spectrometry (LC/MS) techniques, was developed for the determination in packaged food beverages of five ink photoinitiator residues: 2-isopropylthioxanthone (ITX), benzophenone, 2-ethylhexyl-4-dimethylaminobenzoate (EHDAB), 1-hydroxycyclohexyl-1-phenyl ketone (IRGACURE 184) and ethyl-4-dimethylaminobenzoate (EDAB). Samples were extracted from selected beverages (milk, fruit juices and wine) and relative packagings, using n-hexane and dichloromethane, respectively, purified on solid-phase extraction (SPE) silica gel cartridges, and then analyzed in GC/MS and LC/MS. The recovery percentages, obtained spiking the beverage samples at concentrations of 4 and 10 microgl(-1) with a standard mixture of photoinitiators, were in the range 42-108% (milk), 50-84% (wine), and 48-109% (fruit juices). The repeatability of the method was assessed in all cases by the % of correlation value, that was lower than 19%. The lowest limits of detection (LODs) and limits of quantification (LOQs), obtained using GC/MS, were in the range 0.2-1 and 1-5 microgl(-1), respectively. The method was applied to the analysis of forty packaged food beverages (milk, fruit juices and wine samples). The most significant contamination was that of benzophenone, found in all samples in a concentration range of 5-217mugl(-1). Its presence was confirmed by an LC/Atmospheric-Pressure PhotoIonization (APPI)/MS/MS analysis. The photoinitiator (EHDAB) was found in eleven out of forty beverages in a concentration range of 0.13-0.8 microgl(-1). Less important was the ITX contamination, found in three out of forty samples in a range 0.2-0.24 microgl(-1). The work proposes a new method to analyze ink photoinitiator residues in polycoupled carton packaging and in contained food beverages.     

Solid-phase microcolumn extraction and gas chromatography-mass spectrometry identification of volatile organic compounds emitted by paper

A rapid non-destructive sampling technique for the analysis of volatile organic compounds (VOCs) emitted by paper sheets is described. A capillary, which is connected to a microcolumn packed with Tenax TA, is inserted between two sheets at the centre of a paper stack encapsulated inside a PET/Al/PE composite foil. The other end of the microcolumn is connected to a gas-tight syringe and an appropriate volume of gaseous phase is aspirated. The microcolumn is then thermally desorbed in a modified GC inlet (modification is presented) and analysed by gas chromatography-mass spectrometry (GC-MS). In the chromatogram from the analysis of artificially aged paper sample 21 compounds were identified. Advantages of the method including the short sampling time (1 min), simplicity and economic aspect are discussed.     

Unknown residual solvents identification in drug products by headspace solid phase microextraction gas chromatography-mass spectrometry

Summary A sensitive headspace SPME method for the extraction of residual solvents from pharmaceutical products has been developed and optimized. It was found that minimizing sample and headspace volume has a beneficial effect on extraction efficiency. At the same time the method reproducibility was seriously affected by reducing sample and headspace volume. The added air volume was not found to have any significant influence on method sensitivity. The method showed reproducibilities of less than 10% and detection limits as low as 1 ppb for benzene and dichloromethane. The headspace SPME method is around 1000 times more sensitive than static headspace. The optimized parameters were headspace volume 1.5 mL, sample volume 10 μL, and extraction time 30 min. The method was successfully applied to the identification of unknown residual solvents in three different proprietary active drug substances and was successfully applied to the confirmation of the presence of benzene in a proprietary drug substance. Presented at Balaton Symposium '01 on High-Performance Separation Methods, Siófok, Hungary, September 2–4, 2001     

Novel gas chromatography-mass spectrometry database for automatic identification and quantification of micropollutants

A novel gas chromatography-mass spectroscopy (GC-MS) database for identification and quantification of micropollutants in environmental and food samples is reported. GC retention times, calibration curves, and mass spectra of nearly 700 chemicals were registered in the database, and the GC retention times of registered chemicals in actual samples were predicted from the retention times of n-alkanes measured before sample analysis. Differences between predicted and actual retention times were less than 3 s, an accuracy that is nearly identical to that obtained by analysis of standard substances. After the retention times were predicted, a calibration file for the GC-MS instrument was created from the predicted retention times, calibration curves, and mass spectra of the registered chemicals. With the resulting calibration file, automated identification of all the chemicals in actual samples was possible without the use of standards, and the identification method was as reliable as conventional methods. When the GC inlet, column, and tuning conditions were adjusted using GC-MS performance check standards, relative standard deviations of 20% or less for determination values could be obtained. More than 90% of the chemicals in the database could be detected at a sensitivity sufficient for all practical purposes (100 pg or less). Because each chemical in the database, to which new substances can easily be added, can be determined in 1 h, micropollutants in samples can be analyzed efficiently and inexpensively.     

裂解气相色谱-质谱法分析裂解气压缩机垢样的组成

采用裂解气相色谱-质谱联用技术对大庆石化公司裂解气压缩机EC301不同压缩段的垢样进行分析。实验使用了RJ-1型管炉式裂解器,在500℃下对垢样进行裂解,色谱柱为60 m DB-1型毛细管色谱柱,质谱采用电子轰击电离源,电离能70 eV。研究表明,垢样的形成与环戊二烯密切相关。裂解气相色谱-质谱法可以作为分析该垢样成因的有效手段。     

Characterisation of beeswax in works of art by gas chromatography-mass spectrometry and pyrolysis-gas chromatography-mass spectrometry procedures

Pyrolysis (Py) with in situ derivatisation with hexamethyldisilazane-gas chroma-break tography-mass spectrometry (GC-MS) and a gas chromatography-mass spectrometry procedure based on microwave-assisted saponification were used to identify the organic components in small sized beeswax samples. With the latter procedure quantitative recoveries can be made and hydrocarbons, alcohols and omega-1-diols in the neutral fraction, and fatty acids and omega-1-hydroxy acids in the acidic fraction can be efficiently separated and detected. Both procedures were used to characterise a wax anatomic sculpture "The Plague" (1691-1694) by Gaetano Zumbo, resulting in the identification of beeswax and a Pinaceae resin. The GC-MS analysis brought to light some essential differences in beeswax composition between the raw material and the old modelled wax thus giving some clear indications about the recipe used by the sculptor.     

气相色谱-质谱法测定化妆品中的三氯叔丁醇

建立了测定化妆品中三氯叔丁醇的气相色谱-质谱分析方法.膏霜、水剂、散粉、香波、唇膏等不同类型的化妆品样品加入50％甲醇-无水乙醇或无水乙醇超声提取后,样品提取液高速离心处理,取上清液经无水硫酸钠脱水,进行气相色谱-质谱定性及定量分析.选用DB-1701(30 m×0.25 mm×0.25μm)石英毛细管柱,程序升温,流...     

气相色谱-质谱法测定水中痕量的四乙基铅

建立了气相色谱-质谱(GC-MS)测定水中痕量四乙基铅的分析方法。用正己烷萃取水样中的四乙基铅,萃取液浓缩后加入同位素内标萘-d8,采用GC-MS选择离子方式(SIM)进行检测,在200 mL水样中四乙基铅的检出限可达0.04 μg/L;添加回收率为92.2%～103%,准确度好;平行5次测定的相对标准差为4.4%～13.3%。结果表明: 方法简便、快速、准确、实用,可用于水中痕量四乙基铅的测定。