Gas chromatography — mass spectrometry of catechol estrogens

The gas chromatographic and mass spectrometric properties of 19 catechol estrogens and catechol estrogen methyl ethers are reported. The gas chromatographic behaviour of the TMS-derivatives on the stationary phases OV-1, OV-3, OV-7, and OV-17 is examined and correlated with their molecular weight, shape, and polarity. The characteristic mass spectrometric features of the compounds result from the aromatic ring A, which is able to stabilize positive charge within the molecular ions. Consequently the molecular ions form the base peaks of the spectra. Fragmentation patterns highly specific for the catechols as well as for their monomethyl and dimethyl ethers are discussed and substantiated by determination of metastable ions and high resolution mass measurements.     

Three series of high molecular weight alkanoates found in Amazonian plants

Electron impact mass spectra were measured by high temperature high resolution gas chromatography-mass spectrometry (HT-HRGC-MS) for three homologous series of high molecular weight compounds present in the Amazonian plants Marupá (Simaruba amara) and Brazil nut (Bertholettia excelsa). Based on their mass spectra, the compounds were identified as three wax ester series of alpha-tocopherol (vitamin E), beta-tocopherol and phytol (2,6,10,14-tetramethylhexadec-14-en-16-ol). The interpretations are supported by high resolution mass spectrometry and GC retention indices of authentic standards.     

Chemical ionization-mass spectra of aldobiouronic acids and dialdose dianhydrides

Chemical ionization (c.i.) mass spectra with isobutane as the reagent gas are reported for the peracetates of aldobiouronic acids and related compounds, and for peracetates and permethylated derivatives of dialdose dianhydrides. Ions (M⁺ + 43) having relatively high intensities were detected in the spectra of disaccharides lacking the dianhydride structure. Peracetylated dialdose dianhydrides showed very weak (M⁺ + 43) ions, and permethylated dianhydrides did not show them. The (M⁺ + 43) ion consisted of molecular ion and acetoxyl radical (but not of the reagent gas). In the c.i. mass spectra of the usual disaccharide peracetates, (M⁺ ? 31) and (M⁺ ? 60) ions had large intensities. In contrast, c.i. mass spectra extremely similar to the corresponding e.i. mass spectra were obtained for dialdose dianhydrides.     

Chemical ionization mass spectrometry of trimethylsilylated carbohydrates and organic acids retained in uremic serum

After appropriate sample pretreatment and derivatization, uremic serum was investigated by combined high resolution gas chromatography and mass spectrometry, using both electron impact and chemical ionization methods. Electron impact and chemical ionization spectra of a number of identified (trimethylsilylated) carbohydrates and organic acids are compared. The utilization of chemical ionization mass spectrometry, with isobutane as the reagent gas, is discussed in detail. The influence of the reagent gas pressure on the total ion current and on the spectral appearance was studied. The identification of compounds, based on electron impact mass spectral data, was confirmed and often aided appreciably by using this technique. The chemical ionization spectra of trimethylsilyated alditols and aldonic acids, as well as of other organic acids showed protonated molecular ions, whereas aldoses did not. Differences with electron impact spectra are found mainly in the high mass region. The loss of one or more trimethylsilanol groups becomes the predominating fragmentation route at higher reagent gas pressures.     

Determination of lorazepam in plasma and urine as trimethylsilyl derivative using gas chromatography–tandem mass spectrometry

A procedure based on gas chromatography–tandem mass spectrometry for identification and quantitation of lorazepam in plasma and urine is presented. The analyte was extracted from biological fluids under alkaline conditions using solid-phase extraction with an Extrelut-1 column in the presence of oxazepam-d₅ as the internal standard. Both compounds were then converted to their trimethylsilyl derivatives and the reaction products were identified and quantitated by gas chromatography–tandem mass spectrometry using the product ions of the two compounds (m/z 341, 306 and 267 for lorazepam derivative and m/z 346, 309 and 271 for oxazepam-d₅ derivative) formed from the parent ions by collision-induced dissociation in the ion trap spectrometer. Limit of quantitation was 0.1 ng/ml. This method was validated for urine and plasma samples of individuals in treatment with the drug.     

Combined assay for phenacetin and paracetamol in plasma using capillary column gas chromatography—negative-ion mass spectrometry

A gas chromatographic—mass spectrometric assay has been developed for the measurement of phenacetin and its major metabolite paracetamol in plasma. Phenacetin and unconjugated paracetamol are analysed in a single chromatographic run while total paracetamol is measured separately after enzymatic hydrolysis. The two compounds, and the deuterated analogues used as internal standards, are analysed as their trifluoroacetyl derivatives and the mass spectrometer is operated in the electron-capture negative-ion chemical ionisation mode. The negative-ion mass spectra of the derivatives contain fragment ions, formed by loss of an acetyl group from the respective molecular ions, which are the base peaks in the spectra. When these ions are specifically monitored, amounts of derivative equivalent to 1 pg of parent compound can be detected. This allowed the development of an assay for phenacetin, unconjugated paracetamol and total paracetamol in plasma having a precision of 2.6, 1.4 and 2.4%, respectively, and preliminary results for a subject given a 100-mg oral dose of phenacetin are reported.     

Negative ion ammonia chemical ionization and electron impact ionization mass spectrometric analysis of steryl fatty acyl esters

Synthesis of steryl palmitates, varied in the nature of the steryl moiety, provided model compounds for investigation of the mass spectrometric behavior of steryl long-chain fatty acyl esters. The structure of the steryl moiety was varied according to: (i) position and degree of unsaturation in the steroid nucleus and C-17 side-chain, (ii) position and degree of methylation, (iii) presence or absence of a 9 beta, 19-cyclopropane ring. Compounds were chosen so as to be representative of biochemically important steryl esters. Electron impact (EI) behavior of steryl palmitate esters closely resembles that of their short-chain (e.g. acetate) counterparts. M+.ions were generally weak or absent and the major high mass ions arose from characteristic fragmentations of the steroid nucleus following loss of the acyl moiety ([M-RCO2H]+.). Fragment ions characteristic of the acyl moiety were lacking. Negative ion chemical ionization (NICI) using ammonia as reagent gas, on the other hand, afforded spectra containing characteristic fragment ions [RCO2]-, [RCO2-18]-, and [RCO2-19]- from which the nature of the fatty acyl moiety can be readily deduced. Hence, NICI and EI provide complementary means of ionization for the mass spectrometric determination of structures of steryl esters.     

Bis(trifluoromethyl)aryl derivatives for drug analysis by gas chromatography electron capture negative ion chemical ionization mass spectrometry. Application to the measurement of low levels of clonidine in plasma

A gas chromatographic mass spectrometric assay for clonidine in plasma with a detection limit of a few picograms per ml was required. The p-trifluoromethylbenzyl, pentafluorobenzyl and pentafluorobenzoyl derivatives of clonidine were synthesized and the electron capture negative ion chemical ionization mass spectra of these compounds show extensive fragmentation with prominent ions at m/z 35 and 37 due to the two chlorine atoms in the clonidine molecule. Selected ion monitoring of specific high mass ions in these mass spectra indicated that the required sensitivity could not be obtained with these derivatives. Several bis(trifluoromethyl)pyrimidines were synthesized and these compounds were found to give an intense negative ion current under conditions of resonance electron capture. Consequently, a derivative of clonidine containing a bis(trifluoromethyl)aryl group was synthesized by reacting the drug with 3,5-bis(trifluoromethyl)benzoyl chloride. The negative ion mass spectrum of the reaction product has a base peak at m/z 673 and, when this ion is specifically monitored, an amount of derivative equivalent to 1 picogram of clonidine can be detected. This allowed the development of an assay for clonidine in plasma with a precision of 8% (SD) at 50 pg ml-1, 22% (SD) at 20 pg ml-1 and a lower limit for quantitative determination of 10 pg ml-1. Plasma concentrations of clonidine in 10 subjects given a single 50 micrograms oral dose are reported.     

Development of high throughput dispersive LC-ion mobility-TOFMS techniques for analysing the human plasma proteome.

A technique that combines ion mobility spectrometry (IMS) with reversed-phase liquid chromatography (LC), collision-induced dissociation (CID) and mass spectrometry (MS) has been developed. The approach is described as a high throughput means of analysing complex mixtures of peptides that arise from enzymatic digestion of protein mixtures. In this approach, peptides are separated by LC and, as they elute from the column, they are introduced into the gas phase and ionised by electrospray ionisation. The beam of ions is accumulated in an ion trap and then the concentrated ion packet is injected into a drift tube where the ions are separated again in the gas phase by IMS, a technique that differentiates ions based on their mobilities through a buffer gas. As ions exit the drift tube, they can be subjected to collisional activation to produce fragments prior to being introduced into a mass spectrometer for detection. The IMS separation can be carried out in only a few milliseconds and offers a number of advantages compared with LC-MS alone. An example of a single 21-minute LC-IMS-(CID)-MS analysis of the human plasma proteome reveals approximately 20,000 parent ions and approximately 600,000 fragment ions and evidence for 227 unique protein assignments.     

An Interpretation of the Mass Spectra of Salinomycin and Its Derivatives

Salinomycin is a new polyether antibiotic elaborated by Streptomyces albus. An interpretation of the mass spectra of salinomycin and its derivatives has been achieved with the aid of high resolution mass spectrometry along with measurement of the metastable ions. Interesting aspects of the mass spectral data of these compounds are that the characteristic cleavage occurred at the ring C of the tricyclic spiroketal system, and that the sodium salts of salinomycin itself and some derivatives thereof are volatile enough to give distinct spectra which showed fragmentations comparable with those of the free acid and the methyl ester.

An interpretation of the principal fragment ions and their formation mechanism for salinomycin and its derivatives is discussed.     

Trennung und charakterisierung saurer harnbestandteile

Separation and characterization of acidic urine constituentsThe acidic compounds of urine were separated by thin-layer chromatography in eight fractions. Each fraction was investigated separately by the combination glass capillary gas chromatography—mass spectrometry. About 500 compounds were detected, 2/5 of these could be characterized by their mass spectra. Retention data and key fragments of the mass spectra were tabulated. Many of the detected compounds are still unknown.     

The mass spectra of diethylstilbestrol and related compounds

The low resolution mass spectra of E-3,4-bis-(p-hydroxyphenyl)-hex-3-ene (diethylstilbestrol), E-[1,1,1-3H3]3,4-bis-(p-hydroxyphenyl)-hex-3-ene, E-2,3-bis-(p-hydroxyphenyl)-but-2-ene (dimethylstilbestrol), E,E-3,4-bis-(p-hydroxyphenyl)hexa-2,4-diene (dienestrol) and 3,4-bis-(p-hydroxyphenyl)-hexane (hexestrol) were examined as the parent compounds, their diacetates, dimethyl ethers, and bis-trimethylsilyl ethers. In addition, the mass spectra of the diethyl ether and the hexadeuteriodimethyl ether of E-3,4-bis-(p-hydroxyphenyl)-hex-3-ene were studied. Each compound gives rise to several sets of characteristic fragment ions associated with loss of alkyl groups, loss of aryl groups and rearrangements. An ion of m/e 165 (C13H9) was found in the spectra of all the compounds studied. With the aid of high resolution mass spectrometry empirical formulae were assigned to major ions of the free diphenols.     

Gas chromatography mass spectrometry studies on biologically important 8-aminoquinoline derivatives

Several substituted 8-aminoquinolines related to known antimalarial drugs have been studied by gas chromatography mass spectrometry. 5,6-Dihydroxy-8-aminoquinoline, a possible metabolite of Primaquine, can be detected by single ion monitoring after conversion to a trimethylsilyl ether derivative. The mass spectra obtained in this study indicate that there are certain ions which are characteristic of the trimethylsilyl ethers of hydroxylated 8-aminoquinolines and 5,6-dimethoxy-8-aminoquinolines. These compounds should thus be amenable to analysis if they were produced during in vivo metabolism studies. Using selected ion monitoring the derivatized compounds can be detected at submicrogram levels.     

一种用于昆虫性信息素成分单不饱和双键定位的简便方法

昆虫利用特定的性信息素成分实现种内信息交流和种间生殖隔离,性信息素成分异构体的细微差异都能导致其生物活性的很大改变。本研究利用二甲基二硫醚（DMDS）甲硫基化反应和气相色谱-电子轰击质谱（GC-EI-MS）的方法对脂肪族单不饱和醇、醛和乙酸酯类昆虫性信息素成分双键在碳链中的位置异构进行系统研究。质谱分析结果表明,含有醇和乙酸酯官能团的单不饱和烯烃甲硫基化衍生物的质谱特征碎片分子离子峰（M+）及2个主要的诊断离子m/z 61+14m ［H-（CH₂）_m-CH=S⁺ CH₃］和77+14n［CH₃S⁺ =CH（CH₂）_nOH］（或119+14n,［CH₃S⁺ =CH（CH₂）_nOOCCH₃］）丰度都很强,可以据此直接推断双键在碳链中的位置。尽管单不饱和醛类化合物DMDS衍生物的特征碎片峰也很明显,但是由于羰基的质量数和2个亚甲基的质量数相同,不能由低分辨质谱的特征碎片峰直接推断双键在碳链中的位置。DMDS甲硫基化反应结合GC-EI-MS分析方法鉴定昆虫性信息素成分单不饱和双键在碳链中的位置异构,该方法简单快捷,为昆虫性信息素的结构鉴定工作奠定了坚实的基础。     

TagFinder for the quantitative analysis of gas chromatography--mass spectrometry (GC-MS)-based metabolite profiling experiments

MOTIVATION: Typical GC-MS-based metabolite profiling experiments may comprise hundreds of chromatogram files, which each contain up to 1000 mass spectral tags (MSTs). MSTs are the characteristic patterns of approximately 25-250 fragment ions and respective isotopomers, which are generated after gas chromatography (GC) by electron impact ionization (EI) of the separated chemical molecules. These fragment ions are subsequently detected by time-of-flight (TOF) mass spectrometry (MS). MSTs of profiling experiments are typically reported as a list of ions, which are characterized by mass, chromatographic retention index (RI) or retention time (RT), and arbitrary abundance. The first two parameters allow the identification, the later the quantification of the represented chemical compounds. Many software tools have been reported for the pre-processing, the so-called curve resolution and deconvolution, of GC-(EI-TOF)-MS files. Pre-processing tools generate numerical data matrices, which contain all aligned MSTs and samples of an experiment. This process, however, is error prone mainly due to (i) the imprecise RI or RT alignment of MSTs and (ii) the high complexity of biological samples. This complexity causes co-elution of compounds and as a consequence non-selective, in other words impure MSTs. The selection and validation of optimal fragment ions for the specific and selective quantification of simultaneously eluting compounds is, therefore, mandatory. Currently validation is performed in most laboratories under human supervision. So far no software tool supports the non-targeted and user-independent quality assessment of the data matrices prior to statistical analysis. TagFinder may fill this gap. Strategy: TagFinder facilitates the analysis of all fragment ions, which are observed in GC-(EI-TOF)-MS profiling experiments. The non-targeted approach allows the discovery of novel and unexpected compounds. In addition, mass isotopomer resolution is maintained by TagFinder processing. This feature is essential for metabolic flux analyses and highly useful, but not required for metabolite profiling. Whenever possible, TagFinder gives precedence to chemical means of standardization, for example, the use of internal reference compounds for retention time calibration or quantitative standardization. In addition, external standardization is supported for both compound identification and calibration. The workflow of TagFinder comprises, (i) the import of fragment ion data, namely mass, time and arbitrary abundance (intensity), from a chromatography file interchange format or from peak lists provided by other chromatogram pre-processing software, (ii) the annotation of sample information and grouping of samples into classes, (iii) the RI calculation, (iv) the binning of observed fragment ions of equal mass from different chromatograms into RI windows, (v) the combination of these bins, so-called mass tags, into time groups of co-eluting fragment ions, (vi) the test of time groups for intensity correlated mass tags, (vii) the data matrix generation and (viii) the extraction of selective mass tags supported by compound identification. Thus, TagFinder supports both non-targeted fingerprinting analyses and metabolite targeted profiling. AVAILABILITY: Exemplary TagFinder workspaces and test data sets are made available upon request to the contact authors. TagFinder is made freely available for academic use from http://www-en.mpimp-golm.mpg.de/03-research/researchGroups/01-dept1/Root_Metabolism/smp/TagFinder/index.html.     

Comparison of electron and chemical ionization mass spectrometry of sialic acids

Sialic acids were analyzed as per-O-trimethylsilylated compounds by gas-liquid chromatography/mass spectrometry either on electron or chemical ionization by isobutane. Electron ionization mass spectra of these derivatives are very similar to those of the corresponding methyl esters described earlier whereas chemical ionization mass spectra are characterized in the high mass range by loss of the C-2 and the C-4 substituents from the M + 1 ion. Together with other fragment ions the seven different sialic acids analyzed could be clearly identified.     

The Mass Spectra of the Lankacidin Antibiotics

Bundlins A and B, antibiotics elaborated by Streptomyces sp. 6642 GC₁, have the unique structures which possess a seventeen-membered carbon skeleton fused with an unusual β-keto-δ-Mactonic system and a pyruvamide side chain. In the course of the structural studies of the antibiotics, we found that these compounds showed the interesting fragmentations in their mass spectra and in consequence, the investigation about the interpretation of the principal peaks together with their formation mechanism was undertaken by the aid of high resolution mass spectrometry and the measurement of meta stable ions. In addition to the two antibiotics, the mass spectra of related compounds designated T–2636 D and T-2636 F were also investigated.     

Characterization of gangliosides by direct inlet chemical ionization mass spectrometry

Permethylated derivatives of N-acetylneuraminic acid-containing GM3, N-glycolylneuraminic acid-containing GM3, GD1a, and GD1b, were analyzed by direct inlet ammonia chemical ionization (CI) mass spectrometry. These compounds were found to yield simple fragmentations, prominent fragment ions in the high mass region, and characteristic fragment ions corresponding to the cleavage of glycosidic linkages. This method also provides detailed information on the carbohydrate sequence and lipophilic composition in gangliosides.     

Application of gas chromatography-surface ionization organic mass spectrometry to forensic toxicology

Surface ionization (SI), which consists in the formation of positive and negative ions along the course of thermal desorption of particles from a solid surface, was first applied as a detector for gas chromatography (GC), GC-surface ionization detection (SID); we developed many new sensitive methods for the determination of abused and other drugs by GC-SID. Recently, Fujii has devised a combination of SI and a quadrupole mass spectrometer and named this system a surface ionization organic mass spectrometer (SIOMS), which is highly selective and sensitive for organic compounds containing tertiary amino groups. We have tried to apply this mass spectrometer to forensic toxicological study; so far we have succeeded in determining important drugs-of-abuse and toxic compounds, such as phencyclidine (PCP), pethidine, pentazocine, MPTP and its derivatives from human body fluids with high sensitivity and selectivity. In this review, we describe our recent studies on the application of GC-SIOMS to forensic toxicology.     

Estrogens in seminal plasma of human and animal species: Identification and quantitative estimation by gas chromatography—Mass spectrometry associated with stable isotope dilution

Estrone, 2-methoxyestrone and estradiol-17β have been definitively identified in seminal plasma of man, bull, boar and stallion by high resolution gas chromatography associated with selective monitoring of characteristic ions of suitable derivatives. Quantitative estimations were performed by isotope dilution with deuterated analogues and by monitoring molecular ions of trimethylsilyl ethers of labelled and unlabelled compounds. Concentrations of unconjugated and total estrogens are reported together with the statistical evaluation of accuracy and precision.