Classification of bacteria by simultaneous methylation–solid phase microextraction and gas chromatography/mass spectrometry analysis of fatty acid methyl esters

Direct methylation and solid-phase microextraction (SPME) were used as a sample preparation technique for classification of bacteria based on fatty acid methyl ester (FAME) profiles. Methanolic tetramethylammonium hydroxide was applied as a dual-function reagent to saponify and derivatize whole-cell bacterial fatty acids into FAMEs in one step, and SPME was used to extract the bacterial FAMEs from the headspace. Compared with traditional alkaline saponification and sample preparation using liquid–liquid extraction, the method presented in this work avoids using comparatively large amounts of inorganic and organic solvents and greatly decreases the sample preparation time as well. Characteristic gas chromatography/mass spectrometry (GC/MS) of FAME profiles was achieved for six bacterial species. The difference between Gram-positive and Gram-negative bacteria was clearly visualized with the application of principal component analysis of the GC/MS data of bacterial FAMEs. A cross-validation study using ten bootstrap Latin partitions and the fuzzy rule building expert system demonstrated 87 ± 3% correct classification efficiency.      

Carbohydrate analysis of hemicelluloses by gas chromatography–mass spectrometry of acetylated methyl glycosides

A method based on gas chromatography-mass spectrometry analysis of acetylated methyl glycosides was developed in order to analyze monosaccharides obtained from various hemicelluloses. The derivatives of monosaccharide standards, arabinose, glucose, and xylose were studied in detail and (13)C-labeled analogues were used for identification and quantitative analysis. Excellent chromatographic separation of the monosaccharide derivatives was found and identification of the anomeric configuration was feasible through a prepared and identified pure methyl 2,3,4,6-tetra-O-acetyl-β-D-glucopyranoside. The electron ionization mass spectrum and fragmentation path was studied for each monosaccharide derivative. Fragment ion pairs of labeled and unlabeled monosaccharides were used for quantification; m/z 243/248 for glucose, 128/132 for xylose, and 217/218 for arabinose. Using the intensity ratios obtained from the extracted ion chromatograms, accurate quantification of monosaccharide constituents of selected hemicelluloses was demonstrated.     

Characterisation of historical organic dyestuffs by liquid chromatography–mass spectrometry

This review discusses the characterisation of natural organic dyestuffs of historical interest by liquid chromatography–mass spectrometry. The structures of the most important natural organic dyestuffs traditionally used are presented and discussed from the perspective of their analytical chemical determination. The practical aspects of the determination of this inhomogeneous range of compounds with different structures, such as anthraquinones, flavonoids, indigoids or tannins, are discussed with their implications for sample preparation, liquid chromatographic separation and mass spectrometric detection. The particular focus of this review is the discussion of the mass spectral fragmentation patterns of the different classes of natural organic dyestuffs, which in the ideal case allow the identification of the dyestuff actually used, and thereby provide a key to the better characterisation and understanding of historical objects dyed with natural organic dyestuffs. Figure LC-MS allows characterisation of natural dyestuff constituents: the MS spectrum of alizarin is superimposed over a photo of a textile coloured using this red dye     

Characterisation of permanent markers by pyrolysis gas chromatography–mass spectrometry

Pyrolysis gas chromatography–mass spectrometry (PyGC-MS) was used as a rapid method for the characterization of permanent marker ink. Twenty-four samples of various colours purchased from different manufacturers were characterised. Four main typologies of polymer-binding medium could be distinguished on the basis of the pyrolysis products, and differentiation between permanent markers of different manufacturers could be accomplished. For some permanent marker samples, PyGC-MS analysis allowed pigment identification as well.     

Comparison of liquid chromatography-microchip/mass spectrometry to conventional liquid chromatography–mass spectrometry for the analysis of steroids

The feasibility of a microfluidic-based liquid chromatography-electrospray ionization/mass spectrometric system (HPLC-Chip/ESI/MS) was studied and compared to a conventional narrow-bore liquid chromatography-electrospray ionization/mass spectrometric (LC-ESI/MS) system for the analysis of steroids. The limits of detection (LODs) for oxime derivatized steroids, expressed as concentrations, were slightly higher with the HPLC-Chip/MS system (50–300 pM) using an injection volume of 0.5 μL than with the conventional LC-ESI/MS (10–150 pM) using an injection volume of 40 μL. However, when the LODs are expressed as injected amounts, the sensitivity of the HPLC-Chip/MS system was about 50 times higher than with the conventional LC-ESI/MS system. The results indicate that the use of HPLC-Chip/MS system is clearly advantageous only in the analysis of low-volume samples. Both methods showed good linearity and good quantitative and chromatographic repeatability. In addition to the instrument comparisons with oxime derivatized steroids, the feasibility of the HPLC-Chip/MS system in the analysis of non-derivatized and oxime derivatized steroids was compared. The HPLC-Chip/MS method developed for non-derivatized steroids was also applied to the quantitative analysis of 15 mouse plasma samples.     

Ultra-high-pressure liquid chromatography–tandem mass spectrometry for the analysis of six resorcylic acid lactones in bovine milk

This work reports a rapid, reliable and sensitive multi-residue method for the simultaneous determination of six resorcylic acid lactones in bovine milk by ultra-high-pressure liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS). The resorcylic acid lactones were extracted, purified, and concentrated from milk samples in one step using a solid-phase extraction (SPE) cartridge that contained a polymeric mixed-mode anion-exchange sorbent. The analysis was performed on a Waters Acquity BEH C₁₈ column utilizing a gradient elution profile. Each LC run was completed in 3.5 min. The analytes were detected by multiple reaction monitoring (MRM) using electrospray ionization (ESI) negative mode. Mean recoveries from fortified samples ranged from 92.6% to 112.5%, with relative standard deviations lower than 11.4%. Using 5 mL bovine milk, the limits of detection and quantification for resorcylic acid lactones were in the ranges of 0.01–0.05 and 0.05–0.2 μg/L, respectively. The application of this newly developed method was demonstrated by analyzing bovine milk samples from markets.     

Oligosaccharide analysis by graphitized carbon liquid chromatography–mass spectrometry

Structural analysis of complex mixtures of oligosaccharides using tandem mass spectrometry is regularly complicated by the presence of a multitude of structural isomers. Detailed structural analysis is, therefore, often achieved by combining oligosaccharide separation by HPLC with online electrospray ionization and mass spectrometric detection. A very popular and promising method for analysis of oligosaccharides, which is covered by this review, is graphitized carbon HPLC–ESI-MS. The oligosaccharides may be applied in native or reduced form, after labeling with a fluorescent tag, or in the permethylated form. Elution can be accomplished by aqueous organic solvent mixtures containing low concentrations of acids or volatile buffers; this enables online ESI-MS analysis in positive-ion or negative-ion mode. Importantly, graphitized carbon HPLC is often able to resolve many glycan isomers, which may then be analyzed individually by tandem mass spectrometry for structure elucidation. While graphitized carbon HPLC–MS for glycan analysis is still only applied by a limited number of groups, more users are expected to apply this method when databases which support structural assignment become available.

Comparison of orthogonal liquid and gas chromatography–mass spectrometry platforms for the determination of amino acid concentrations in human plasma

Concentrations of amino acids in a human plasma pool were determined using four independent quantification methods. Orthogonal separation schemes (LC, GC, or GC×GC) and detection systems (triple quadrupole or time-of-flight mass spectrometry) are shown to demonstrate excellent consistency among platforms for quantifying 18 amino acids in NIST Standard Reference Material (SRM) 1950 Metabolites in Human Plasma using a well-characterized isotope dilution (ID) quantification method. Measured levels were consistent with reference values in plasma from the literature. Individual amino acid concentrations in plasma varied by over an order of magnitude ranging from 1.83 μg/g to 28.0 μg/g (7.78 μmol/L to 321 μmol/L). Average variability (coefficient of variation) between experimental amino acid concentrations (excluding cysteine) among all methods was 6.3%. Certified mass fraction values for amino acids in NIST SRM 1950 will be established from statistically weighted means of all experimental results.     

Comparison of supercritical fluid chromatography–tandem mass spectrometry and liquid chromatography–tandem mass spectrometry for the stereoselective analysis of chlorfenvinphos and dimethylvinphos in tobacco

This study used reversed-phase liquid chromatography–tandem mass spectrometry and supercritical fluid chromatography–tandem mass spectrometry for determination of the stereoisomers of chlorfenvinphos and dimethylvinphos in tobacco. Tobacco samples were extracted and purified with a modified quick, easy, cheap, effective, rugged, and safe technique using spherical carbon. The performance of both methodologies was comprehensively compared in terms of methods validation parameters (separation efficiency, linearity, selectivity, recovery, repeatability, sensitivity, matrix effect, etc.). Under optimized conditions, the calibration curves of the stereoisomers of chlorfenvinphos and dimethylvinphos in the range of 10–500 ng/mL showed excellent linearity with R² ≥ 0.997 in both methods. The adequate recoveries of analytes from three different spiked tobaccos were obtained using reversed-phase liquid chromatography–tandem mass spectrometry (86.1–95.7%) as well as supercritical fluid chromatography–tandem mass spectrometry (86.5–94.0%). The relative standard deviations for spiked samples were all below 7.0%. Compared with supercritical fluid chromatography–tandem mass spectrometry, lower matrix effects and LODs can be obtained in reversed-phase liquid chromatography–tandem mass spectrometry.     

Fast,sensitive and highly discriminant gas chromatography–mass spectrometry method for profiling analysis of fatty acids in serum

A method for the determination of fatty acids in serum based on GC–MS (micro-SIS detection mode) has been developed and the separation and cis/trans isomers have been identified. A prior two-step extraction/derivatization procedure accelerated by ultrasound allows individual determination of esterified (EFAs) and non-esterified fatty acids (NEFAs), and shortening of the derivatization steps to 5 min for EFAs and 15 min for NEFAs. The total analysis time for 39 fatty acids was 61 min. The minimum LOD and LOQ values were 0.002 and 0.006 μg/ml, respectively. The proposed method was validated for EFAs and NEFAs using two different methods and the results show no statistical differences between the proposed method and those used as reference. The proposed derivatization–extraction methodology is suitable for fatty-acid analysis of human serum, and can be applied to nutritional and epidemiological studies.     

Determination of trace volatile fatty acids in ambient air by capillary gas chromatography–mass spectrometry in SIM mode

A simple, rapid and sensitive method was developed and validated for the analysis of C₂–C₅ volatile fatty acids (VFAs) in ambient air. This method involves preconcentration of VFAs with a sodium carbonate-impregnated silica gel tube, ultrasonic extraction with pure water, partition of VFAs to diethyl ether and determination using gas chromatography with a mass selective detector in the selected ion monitoring mode. A water-resistant free fatty acid phase capillary column was used to directly separate C₂–C₅ VFAs without the time-consuming derivatisation process. The limits of detection ranged from 0.001 to 0.003 µg m^?3 and the limits of quantification ranged from 0.003 to 0.010 µg m^?3. The validated method was successfully applied to the analysis trace-level VFAs in ambient air and in air samples from a landfill with perceived odour pollution.     

Development of novel fiber-packed needle interface for off-line reversed-phase liquid chromatography–capillary gas chromatography

For two-dimensional reversed-phase liquid chromatography–gas chromatography (2D RPLC–GC), a specially-designed needle packed with a polymer-coated fibrous stationary phase was introduced as a novel interface. The bundle of synthetic fibers coated with polydimethylsiloxane (PDMS) was packed into the head section of the needle, and served as the extraction medium. Using the post-column dilution of the LC eluent by water and subsequent extraction with the needle interface, the analyte was successfully concentrated to the PDMS phase on the fibrous support in the needle. The concentrated analytes were directly injected to GC system by inserting the needle to a heated GC injector. 2D separations of aliphatic and aromatic hydrocarbons, and also kerosene-extract were performed with the off-line RPLC–GC system interfaced by the needle extractor. The results suggested that the fiber-packed needle interface could be one of the simple and effective approaches to develop an on-line coupled LC–GC system.     

Elucidation of n-butyl benzyl phthalate biodegradation using high-performance liquid chromatography and gas chromatography–mass spectrometry

n-Butyl benzyl phthalate (BBP) is an endocrine-disrupting chemical. A bacterium species capable of using BBP as the sole source of carbon and energy was isolated from mangrove sediment. Effects of BBP concentration, pH, temperature, and salinity on BBP biodegradation were studied. The optimum pH, temperature, and salinity for the BBP biodegradation were 7.0, 37°C, and 15 g L⁻¹, respectively. BBP was completely degraded within 6 days under optimum conditions, and the biodegradation of BBP could be fitted to a first-order kinetic model. The major metabolites of BBP biodegradation were identified as mono-butyl phthalate, mono-benzyl phthalate, phthalic acid, and benzoic acid by using high-performance liquid chromatography and gas chromatography–mass spectrometry. A preliminary metabolic pathway was proposed for the biodegradation of BBP.      

Complementary fragmentation pattern analysis by gas chromatography–mass spectrometry and liquid chromatography tandem mass spectrometry confirmed the precious lignan content of Cirsium weeds

In this paper, as novelties to the field, it is confirmed at first, that the fruits of Cirsium species, regarded as injurious weeds, do contain lignans, two, different butyrolactone-type glycoside/aglycone pairs: the well known arctiin/arctigenin and the particularly rare tracheloside/trachelogenin species. These experiences were supported by gas chromatography–mass spectrometry (GC–MS), by liquid chromatography tandem mass spectrometry (LC–MS/(MS)) and by nuclear magnetic resonance (NMR) spectroscopy. The study reflects the powerful impact of the complementary chromatographic mass fragmentation evidences resulting in the identification and quantification, the extremely rare, with on line technique not yet identified and described, tracheloside/trachelogenin pair lignans, without authentic standard compounds. Fragmentation pattern analysis of the trimethylsilyl (TMS) derivative of trachelogenin, based on GC–MS, via two different fragmentation pathways confirmed the detailed structure of the trachelogenin molecule. The complementary chromatographic evidences have been unambiguously confirmed, by ¹H and ¹³C NMR analysis of trachelogenin, isolated by preparative chromatography. Identification and quantification of the fruit extracts of four Cirsium (C.) species (C. arvense, C. canum, C. oleraceum, and C. palustre), revealed that (i) all four species do accumulate the tracheloside/trachelogenin or the arctiin/arctigenin butyrolactone-type glycoside/aglycone pairs, (ii) the overwhelming part of lignans are present as glycosides (tracheloside 9.1–14.5 mg/g, arctiin 28.6–39.3 mg/g, expressed on dry fruit basis), (iii) their acidic and enzymatic hydrolyses to the corresponding aglycones, to trachelogenin and arctigenin are fast and quantitative and (iv) the many-sided beneficial trachelogenin and arctigenin can be prepared separately, without impurities, excellent for medicinal purposes.     

Fast analysis of multiple pesticide residues in apple juice using dispersive liquid–liquid microextraction and multidimensional gas chromatography–mass spectrometry

A method for the rapid trace analysis of 24 residual pesticides in apple juice by multidimensional gas chromatography–mass spectrometry (MD-GC/MS) using dispersive liquid–liquid microextraction (DLLME) was developed and optimized. Several parameters of the extraction procedure such as type and volume of extraction solvent, type and volume of dispersive solvent and salt addition were evaluated to achieve the highest yield and to attain the lowest detection limits. The DLLME procedure optimized consists in the formation of a cloudy solution promoted by the fast addition to the sample (5 ml) of a mixture of carbon tetrachloride (extraction solvent, 100 μl) and acetone (dispersive solvent, 400 μl). The tiny droplets formed and dispersed among the aqueous sample solution are further joined and sedimented (85 μl) in the bottom of the conical test tube by centrifugation. Once extracted, all the 24 pesticides were directly injected and separated by a dual GC column system, comprising a short wide-bore DB-5 capillary column with low film thickness connected by a Deans switch system to a second chromatographic narrower column, with identical stationary phase. The instrumental setting used, in combination with carefully optimized operational fast GC and MS parameters, markedly decreased the retention times of the targeted analytes. The total chromatographic run was 8 min. Mean recoveries for apple juice spiked at three concentrations ranged from 60% to 105% and the intra-repeatability ranged from 1% to 21%. The limits of detection of the 24 pesticides ranged from 0.06 to 2.20 μg/L. In 2 of a total of 28 analysed samples were found residues of captan, although at levels below the maximum limit legal established.     

Identification of monoacylglycerol regio-isomers by gas chromatography–mass spectrometry

Monoacylglycerols (MAGs) are lipids found in trace amounts in plants and animal tissues. While they are widely used in various industrial applications, accurate determination of the regio-specific distribution is hindered by the lack of stable, commercially available standards. Indeed, unsaturated β-MAG (or Sn-2 MAG) readily undergoes isomerization into α-MAG (acyl chain is attached to the Sn-1 or the Sn-3 position). In the present study, we describe structural elucidation of α- and β-regio-isomers of monopalmitoyl-glycerol (MAG C16:0) as model compounds in their silylated forms using gas chromatography–mass spectrometry (GC–MS) with electronic impact (EI) ionization. MS fragmentation of α-MAG C16:0 is characterized by the loss of methylene(trimethylsilyl)oxonium (103 amu) and the consecutive loss of acyl chain yielding a fragment ion at m/z 205. The fragmentation pattern of β-MAG C16:0 shows a series of diagnostic fragments at m/z 218, 203, 191 and 103 that are not formed from the α-isomer and hereby enable reliable distinction of these regio-isomers. Possible fragmentation scenarios are postulated to explain the formation of these marker ions, which were also applied to characterize the regio-isomer composition of a complex mixture of MAG sample containing n-3 long-chain polyunsaturated fatty acids.     

Novel derivatisation technique for the determination of chlorophenoxy acid type herbicides by gas chromatography–mass spectrometry

The analytical detection of chlorophenoxycarboxylic-acid-type herbicides (2,4-D, dichloprop, MCPA, etc.) in environmental samples is often a problem in instrumental analysis, as these compounds containing free carboxylic groups require chemical derivatisation prior to gas chromatographic (GC) methods. Nine chlorophenoxy-acid-type herbicide active ingredients have been derivatised successfully with trimethylsilyl N,N-dimethyl carbamate and t-butyldimethylsilyl N,N-dimethyl carbamate by forming their trimethylsilyl (TMS) and t-butyldimethylsilyl (TBDMS) esters, respectively. The detection and determination of the derivatives were performed by capillary gas chromatography–mass spectrometry. The study included determination of retention indices, mass spectral properties and comparison of derivatives produced. The mass spectra of TBDMS derivatives are usually dominated by very characteristic ions [M-57]⁺ resulting from the cleavage of t-butyl moiety during electron impact (EI) ionisation in the mass spectrometer. Limits of detection were 5 to 100 pg applying GC with EI-MS detection in full scan mode. The method, using SPE sample preparation, was applied for the analysis of 115 ground water and surface water samples collected in Békés County, Hungary in 2009.     

Oxidative degradation products analysis of polymer materials by pyrolysis gas chromatography–mass spectrometry

Pyrolysis gas chromatography–mass spectroscopy (PGC–MS) has been proved to be a powerful method to analyze both the volatile additives and the macromolecular structure of polymer materials. In this paper, flash evaporation technique was used to analyze the volatile degradation products of polymer materials during natural and artificial aging. In high density polyethylene (HDPE) composites, mainly n-alkanes with carbon number from 14 to 29 were detected after natural aging, while no oxidative product was found. Different composites have different n-alkane distributions. In contrast, various oxidative products including ketones, alcohols, esters and unsaturated species could be found in aged polypropylene (PP) nanocomposites. Nanoparticles accelerated the chain scission of PP and increased the formation of oxidative products significantly. During thermal oxidation of nitrile rubber (NBR) seal rubbers, heat/oxidation-induced extra crosslinking predominated and no volatile degradation products was detected. The main change happened in the volatiles is the decrease of additives, especially paraffins, antioxidant RD and hindered phenol. This resulted in the hardening of the rubber and the weakening of the protection from oxidation. Furthermore, the additive distribution along the depth was investigated, showing different migration speeds of different additives. From the additive levels remained in the NBR rubber, it is possible to predict the degradation status. In summary, PGC–MS can supply abundant information of polymer degradation and is helpful for mechanism research.     

Nanoscale reversed-phase liquid chromatography–mass spectrometry of permethylated N-glycans

Reversed-phase liquid chromatography on the nanoscale coupled to electrospray tandem mass spectrometry was used to analyse a mixture of four commercial glycan standards, and the method was further adapted to N-glycans enzymatically released from alpha-1-acid glycoprotein and immunoglobulin gamma. Glycans were permethylated to enable their separation by reversed-phase chromatography and to facilitate interpretation of fragmentation data. Prior to derivatization of glycans by permethylation, they were reduced to cancel anomerism because, although feasible, it was not desired to separate α- and β-anomers. The effect of supplementing chromatographic solvent with sodium hydroxide to guide adduct formation was investigated. Raising the temperature in which the separation was performed improved chromatographic resolution and affected retention times as expected. It was shown by using the tetrasaccharides sialyl Lewis X and sialyl Lewis A that reversed-phase chromatography could achieve the separation of methylated isobaric glycan analytes. Isobaric glycans were detected among the N-glycans of immunoglobulin gamma and further analysed by tandem mass spectrometry.