Analysis of Titan tholin pyrolysis products by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

The reddish brown haze that surrounds Titan, Saturn's largest moon, is thought to consist of tholin-like organic aerosols. Tholins are complex materials of largely unknown structure. The very high peak capacity and structured chromatograms obtained from comprehensive two-dimensional GC (GC x GC) are attractive attributes for the characterization of tholin pyrolysis products. In this report, GC x GC with time-of-flight MS detection and a flash pyrolysis inlet is used to characterize tholin pyrolysis products. Identified pyrolysis products include low-molecular-weight nitriles, alkyl substituted pyrroles, linear and branched hydrocarbons, alkyl-substituted benzenes and PAH compounds. The pyrolysis of standards found in tholin pyrolysate showed that little alteration occurred and thus these structures are likely present in the tholin material.     

Statistical methods for comparing comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry results: metabolomic analysis of mouse tissue extracts

The potential for the utilization of GC x GC-time-of-flight (TOF) MS for high-resolution metabolomics studies is discussed, with the implementation of some statistical comparisons for biomarker detection. Metabolite profiles from NZO obese mice versus BL/6 control mice are compared and contrasted using a number of chromatogram comparison routines, including direct chromatogram comparisons, chromatogram subtraction and averaging routines, as well as a method for generating relative weighted peak surface difference chromatograms, and a more conventional Student's t-test statistical approach.     

Solid phase microextraction-comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry for the analysis of honey volatiles

Head-space solid phase microextration (SPME), followed by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GCxGC-TOFMS), has been implemented for the analysis of honey volatiles, with emphasis on the optimal selection of SPME fibre and the first- and second-dimension GC capillaries. From seven SPME fibres investigated, a divinylbenzene/Carboxen/polydimethylsiloxane (DVB/CAR/PDMS) 50/30 microm fibre provided the best sorption capacity and the broadest range of volatiles extracted from the headspace of a mixed honey sample. A combination of DB-5ms x SUPELCOWAX 10 columns enabled the best resolution of sample components compared to the other two tested column configurations. Employing this powerful analytical strategy led to the identification of 164 volatile compounds present in a honey mixture during a 19-min GC run. Combination of this simple and inexpensive SPME-based sampling/concentration technique with the advanced separation/identification approach represented by GCxGC-TOFMS allows a rapid and comprehensive examination of the honey volatiles profile. In this way, the laboratory sample throughput can be increased significantly and, at the same time, the risk of erroneous identification, which cannot be avoided in one-dimensional GC separation, is minimised.     

Characterization of complex hydrocarbons in cigarette smoke condensate by gas chromatography-mass spectrometry and comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

Gas chromatography-mass spectrometry with electron ionization and positive-ion chemical ionization and comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC x GC-TOF-MS) were applied for the characterization of the chemical composition of complex hydrocarbons in the non-polar neutral fraction of cigarette smoke condensates. Automated data processing by TOF-MS software combined with structured chromatograms and manual review of library hits were used to assign the components from GC x GC-TOF-MS analysis. The distributions of aliphatic hydrocarbons and aromatics were also investigated. Over 100 isoprenoid hydrocarbons were detected, including carotene degradation products, phytadiene isomers and carbocyclic diterpenoids. A total of 1800 hydrocarbons were tentatively identified, including aliphatic hydrocarbons, aromatics, and isoprenoid hydrocarbons. The identified hydrocarbons by GC x GC-TOF-MS were far more than those by GC-MS.     

Application of comprehensive two-dimensional gas chromatography mass spectrometry and different types of data analysis for the investigation of cigarette particulate matter

In tobacco research, the comparison of different tobacco blends as well as the puff-dependent behaviour of cigarettes is a matter of particular interest. For the investigation of smoke characteristics, GC x GC offers different ways for data analysis, namely, compound target analysis, automated peak-based compound classification and comprehensive pixel-based data analysis. This study will show the application as well as the pros and cons of these types of data analysis for very complex matrices like cigarette particulate matter. In addition, new aspects about the recently discovered puff-dependent behaviour of compounds in cigarette smoke will be presented. Automated peak-based compound classification including mass spectrometric pattern recognition is used for the classification of tobacco particulate matter samples and the puff-dependent investigation of different compound classes. This compound group specific analysis is further reinforced by applying an even more comprehensive pixel-based analysis. This kind of analysis is used to generate fingerprints of different types of cigarettes. The combination of fast feature reduction methods like analysis of variance (ANOVA) and t-test with multivariate feature transformation methods like partial least squares discriminate analysis (PLSDA) for feature selection provides a powerful tool for a detailed inspection of different types of cigarettes.     

Application of comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry in the analysis of volatile oil of traditional Chinese medicines

This paper reports comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC x GC-TOF MS) analysis of Pogostemon cablin Benth (Cablin Parchouli) volatile oil. The suitable column system and operation conditions were chosen on the basis of the properties of composition of the volatile oil. One-dimensional gas chromatography (ID-GC) and GC x GC, GC-MS and GC x GC-TOF MS were compared under appropriate conditions, and the enhanced sensitivity and superior resolution of GC x GC were demonstrated. 394 components were tentatively identified by GC x GC-TOF MS.     

Comparison of comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry and gas chromatography-mass spectrometry for the qualitative characterisation of roasted barley by solid-phase microextraction

Volatile compounds of roasted barley used in the production of barley coffee, the most common coffee substitute, were analysed by using solid-phase microextraction (SPME) followed by GC-MS and comprehensive GC x GC-TOF-MS, respectively. The optimised SPME extraction conditions in terms of selection of the fibre coating, extraction time and extraction temperature allowed to obtain the highest GC response, thus enhancing the identification capabilities of the developed method. As for the SPME-GC x GC-TOF-MS analysis, 64 compounds with similarity, reverse and probability values above 800, 900 and 6000, respectively, were identified, by using a polar x apolar column set combination; in contrast, GC-MS was able to identify a lower number of compounds, i.e. 40 volatiles.     

Comparison of two algorithmic data processing strategies for metabolic fingerprinting by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

The alignment algorithm Statistical Compare (SC) developed by LECO Corporation for the processing of comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) data was validated and compared to the in-house developed retention time correction and data alignment tool INCA (Integrative Normalization and Comparative Analysis) by a spike-in experiment and the comparative metabolic fingerprinting of a wild type versus a double mutant strain of Escherichia coli (E. coli). Starting with the same peak lists generated by LECO's ChromaTOF software, the accuracy of peak alignment and detection of 1.1- to 4-fold changes in metabolite concentration was assessed by spiking 20 standard compounds into an aqueous methanol extract of E. coli. To provide the same quality input signals for both alignment routines, the universal m/z 73 trace of the trimethylsilyl (TMS) group was used as a quantitative measure for all features. The performance of data processing and alignment was evaluated and illustrated by ROC curves. Statistical Compare performed marginally better at the lower fold changes, while INCA did so at the higher fold changes. Using SC, quantitative precision could be improved substantially by exploiting the signal intensities of metabolite-specific unique (U) m/z ion traces rather than the universal m/z 73 trace. A list of 56 features that distinguished the two E. coli strains was obtained by the SC alignment using m/z U with an estimated false discovery rate (FDR) of <0.05. Ultimately, 23 metabolites could be identified, one additional and five less than with INCA due to the failure of SC to extract unitized m/z U's across all fingerprints with suitable spectral intensities for the latter metabolites.     

Optimization of separation and detection conditions for the multiresidue analysis of pesticides in grapes by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

A comprehensive GCxGC-TOFMS method was optimized for multiresidue analysis of pesticides using a combination of a non-polar (RTX-5MS, 10 m x 0.18 mm x 0.2 microm) and a polar capillary column (TR-50MS, 1 m x 0.1 mm x 0.1 microm), connected in series through a dual stage thermal modulator. The method resolved the co-elution problems as observed in full scan one-dimensional GC-MS analysis and allowed chromatographic separation of 51 pesticides within 24 min run time with library-searchable mass spectrometric confirmation. Four pesticides, viz. chlorpyrifos-methyl, vinclozoline, parathion-methyl and heptachlor could be baseline separated on GCxGC, which were otherwise closely eluting and interfering each other's detection in 1D GC-MS run. Similarly, it could be possible to separate myclobutanil, buprofezin, flusilazole and oxyfluorfen on GCxGC. Although in 1D GC-MS, these closely eluting compounds could be identified through deconvolution algorithm and 'peak-find' option of the Chromatof software but the spectral purity significantly improved on GCxGC analysis. Thorough optimization was accomplished for the oven temperature programming, ion source temperature and GCxGC parameters like modulation period, duration of hot pulses, modulation-offset temperature, acquisition rate, etc. to achieve best possible separation of the test compounds. The limit of detection significantly improved by 2-12 times on GCxGC-TOFMS against GC-TOFMS because of sharper and narrower peak shapes. The method was tested for grape matrix after preparing the samples using previously described method and recoveries of the entire test pesticides were within 70-110% at 10 ng/g level of fortification. GCxGC-TOFMS was found to be an excellent technique for library-based screening of pesticides with high accuracy and sensitivity.     

Multivariate selectivity as a metric for evaluating comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry subjected to chemometric peak deconvolution

Two-dimensional gas chromatography (GC x GC) coupled to time-of-flight mass spectrometry (TOFMS) [GC x GC-TOFMS)] is a highly selective technique well suited to analyzing complex mixtures. The data generated is information-rich, making it applicable to multivariate quantitative analysis and pattern recognition. One separation on a GC x GC-TOFMS provides retention times on two chromatographic columns and a complete mass spectrum for each component within the mixture. In this report, we demonstrate how GC x GC-TOFMS combined with trilinear chemometric techniques, specifically parallel factor analysis (PARAFAC) initiated by trilinear decomposition (TLD), results in a powerful analytical methodology for multivariate deconvolution. Using PARAFAC, partially resolved components in complex mixtures can be deconvoluted and identified without requiring a standard data set, signal shape assumptions or any fully selective mass signals. A set of four isomers (iso-butyl, sec-butyl, tert-butyl, and n-butyl benzenes) is used to investigate the practical limitations of PARAFAC for the deconvolution of isomers at varying degrees of chromatographic resolution and mass spectral selectivity. In this report, multivariate selectivity was tested as a metric for evaluating GC x GC-TOFMS data that is subjected to PARAFAC peak deconvolution. It was found that deconvolution results were best with multivariate selectivities over 0.18. Furthermore, the application of GC x GC-TOFMS followed by TLD/PARAFAC is demonstrated for a plant metabolite sample. A region of GC x GC-TOFMS data from a complex natural sample of a derivatized metabolic plant extract from Huilmo (Sisyrinchium striatum) was analyzed using TLD/PARAFAC, demonstrating the utility of this analytical technique on a natural sample containing overlapped analytes without selective ions or peak shape assumptions.     

Improved enantiomer resolution and quantification of free D-amino acids in serum and urine by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

The potential of comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) in the quantitative analysis of amino acid enantiomers (AAEs) as their methyl chloroformate (MCF) derivatives in physiological fluids was investigated. Of the two column sets tested, the combination of an Rt-γDEXsa chiral column with a polar ZB-AAA column provided superior selectivity. Twenty AAEs were baseline resolved including L-Leu and D-Ile, which had failed separation by one-dimensional chiral GC-quadrupole-MS (GC-qMS). Lower limits of quantification (LLOQ) were in the range of 0.03-2 μM. Reproducibility of the analysis of a serum specimen in octaplicate ranged from 1.3 to 16.6%. The GC×GC-TOFMS method was validated by analyzing AAEs in 48 urine and 43 serum specimens, respectively, and by comparing the results with data obtained by a previously validated GC-qMS method. Mean recoveries ranged from 78.4% for D-Leu to 116.4% for D-Pro in urine and 72.2% for L-Thr to 129.4% for L-Ile in serum. The method was applied to the comparison of AAE serum levels in patients suffering from liver cirrhosis to a control group. Significantly increased D-AA concentrations were found for the patient group, whereas L-AA levels were slightly decreased.     

Comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry for the identification of organobrominated compounds in bluefin tuna

This study evaluates comprehensive two-dimensional gas chromatography (GC×GC) coupled to time-of-flight mass spectrometry (GC×GC-ToF MS) for the simultaneous analysis of several classes of organobromines (OBs), including polybrominated diphenyl ethers (PBDEs), polybrominated biphenyls (PBBs), methoxylated PBDEs (MeO-PBDEs), several halogenated naturally produced compounds (HNPs) and eight novel brominated flame retardants (NBFRs), polybrominated hexahydroxanthene derivates (PBHDs), 2,4,6-tribromoanisole and a mixed halogenated compound (MHC-1), in bluefin tuna muscles. The proposed methodology maximised separation of both within and among OB families, and among these and other halogenated micropollutants detected in these samples and co-extracted matrix components. Special attention has been paid to solve co-elution problems observed during the analysis of OBs with one-dimensional GC-based techniques. Satisfactory separation among several relevant PBDEs and MeO-PBDEs has been obtained allowing their unambiguous determination in a single run. Additional studies were conducted to identify selected NBFRs and HNPs. 2,4-Dibromoanisole, a dibromophenol isomer and hexabromobenzene were identified in the investigated samples. Several new tri- and tetra-BHD derivates were also identified, indicating that these compounds could apparently exist as structured families in nature. In addition, a tetrabrominated diMeO-biphenyl and two tetrabrominated diMeO-BDEs were also tentatively identified.     

Qualitative evaluation of thermal desorption-programmable temperature vaporization-comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry for the analysis of selected halogenated contaminants

The separation of 38 toxic and predominant polychlorinated biphenyl (PCB) congeners, 11 persistent halogenated pesticides, 1 brominated biphenyl (BB), and 8 polybrominated diphenyl ethers (PBDEs) has been optimized using comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC x GC-TOFMS). A thermal desorption-programmable temperature vaporization (TD-PTV) step was used for the injection. Different column sets were investigated, and a 100% dimethylpolysiloxane (15 m x 0.25 mm i.d. x 0.25 microm film thickness) narrowbore capillary column coupled to a high temperature (8% phenyl)-polycarborane-siloxane (2 m x 0.10 mm i.d. x 0.10 microm film thickness) microbore column set was selected. Of the 58 compounds investigated, only one pair of PCBs was not resolved. All other analytes were either baseline separated into the chromatographic plane or were virtually separated using the deconvolution capability of the TOFMS.     

Evaluation of two-dimensional gas chromatography-time-of-flight mass spectrometry for the determination of multiple pesticide residues in fruit

In recent years, comprehensive two-dimensional gas chromatography (GC x GC) has attained increasing attention for its outstanding separation potential and capability to solve demanding analytical tasks. Trace level analysis of pesticides residues in complex food matrices represents such a demanding task. For some commodities, such as baby food, the requirements on method detection limits are very strict and the unambiguous confirmation of the pesticide presence based on mass spectrometric detection is required. In this work, GC x GC coupled to time-of-flight mass spectrometry (TOF MS) has been evaluated for the determination of pesticides residues in fruit samples. Twenty modern pesticides with a broad range of physico-chemical properties were analysed in apple and peach samples. It has been demonstrated that the application of comprehensive two-dimensional gas chromatography brings distinct advantages such as enhanced separation of target pesticides from matrix co-extracts as well as their improved detectability. The limits of detection of the pesticides comprised in the study (determined at S/N = 5) ranged from 0.2 to 30 pg, injected with the exception of the last eluted deltamethrin, for which 100 pg could be detected. When compared to one-dimentional GC-TOF MS analysis under essentially the same conditions the detectability enhancement was 1.5-50-fold. Full mass spectral information by time-of-flight mass spectrometry and the deconvolution capability of the dedicated software allowed for reliable identification of most pesticides at levels below 0.01 mg/kg (< 10 pg injected) in fruit. Performance characteristics of the GC x GC-TOF MS method, such as linearity of calibration curves, repeatability of (summed) peak areas, as well as repeatability of first and second dimension retention times, were shown to fully satisfy the requirements for trace level analysis of the pesticide residues in food.     

Odour fingerprint acquisition by means of comprehensive two-dimensional gas chromatography-olfactometry and comprehensive two-dimensional gas chromatography/mass spectrometry

The analysis of complex matrices, such as perfumes, by means of gas chromatography-olfactometry (GC-O) can be rather imprecise due to the co-elutions, leading to a possible masking of odour-active trace-level compounds by major interferences or agglomeration of olfactive impressions resulting in unreliable olfactive characterization. To overcome these limits an innovative technique, comprehensive two-dimensional gas chromatography-olfactometry (GC x GC-O), was applied, revealing several relevant co-elutions, as in the linalool and linalyl acetate zones. A total of 177 compounds, out of these 135 odour-active, were detected by GC-O, while about 481 out of 818 compounds presented odour-activity through GC x GC-O analyses. In addition, GC/mass spectrometry (GC/MS) and GC x GC/MS analyses were also performed. Peak assignment was achieved by means of different information sources, such as GC/MS, GC x GC/MS, LRI, injection of standards and olfactive impressions.     

Determination of Teucrium chamaedrys volatiles by using direct thermal desorption-comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry

The direct qualification and quantification of the volatile components of Teucrium chamaedrys was studied using a direct thermal desorption (DTD) technique with comprehensive two-dimensional (2D) gas chromatography-time-of-flight mass spectrometry (GC x GC-TOF/MS). The GC x GC separation chromatographically resolved hundreds of components within this sample, and with the separation coupled with TOF/MS for detection, high probability identifications were made for 68 compounds. The quantitative results were determined through the use of internal standards and the desorption of differing amounts of raw material in the injector. The highest yield of volatile compounds (0.39%, w/w) was obtained at 150 degrees C thermal desorption temperature using 1.0mg of dried sample placed in a glass injector liner when studied over the range 1.0-7.0mg. Lowest yield of 0.33% (w/w) was found for the largest sample size of 7.0mg. Relative standard deviation (RSD) for 10 replicates at each size sample were in the range 3.9-21.6%. The major compounds identified were beta-pinene, germacrene D, alpha-pinene, alpha-farnesene, alpha-gurjunene, gamma-elemene and gamma-cadinene. All identified compounds were quantified using total ion chromatogram (TIC) peak areas. DTD is a promising method for quantitative analysis of complex mixtures, and in particular for quantitative analysis of plant samples, which can yield data without the traditional obligation for costly and time-consuming extraction techniques.     

Comparison of comprehensive two-dimensional gas chromatography/time-of-flight mass spectrometry and gas chromatography-mass spectrometry for the analysis of tobacco essential oils

A sample of tobacco essential oil was analyzed using gas chromatography-mass spectrometry (GC/MS) and comprehensive two-dimensional gas chromatography coupled to a time-of-flight mass spectrometry (GC × GC/TOFMS), respectively. In the GC/MS analysis, serially coupled columns were used. By comparing the GC/MS results with GC × GC/TOFMS results, many more components in the essential oil could be found within the two-dimensional separation space of GC × GC. The quantitative determination of components in the essential oil was performed by GC × GC with flame ionization detection (FID), using a method of multiple internal standards calibration.     

Application of head-space solid-phase microextraction coupled to comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry for the determination of multiple pesticide residues in tea samples

A new method has been developed to detect 36 pesticides that may contaminate tea samples (green, black and fruit tea). The hyphenation of solid-phase microextraction in head-space mode with a comprehensive two-dimensional gas chromatography coupled with high-speed time-of-flight mass spectrometry (HS-SPME-GC × GC/TOF MS) proved to be a quick alternative to conventional GC/MS methodology which employs solvent-based extraction. The key parameters for controlling HS-SPME performance were optimized, including fiber coating type, temperature and absorption time settings and tea matrix modification by adding water. Quantification was carried out using matrix-matched calibration. The repeatability of measurements, expressed as relative standard deviation (R.S.D.), was less than 24% for all analytes. The limits of quantification ranged from 1 to 28 μg kg⁻¹. The optimized method was applied to analyze real life samples obtained from a retail market. Results generated by the new SPME procedure and those obtained by using a conventional one involving ethyl acetate extraction and high-performance gel permeation chromatography (HPGPC) clean up agreed with each other for positive (containing residue) samples.