Comprehensive two-dimensional gas chromatography— A discussion on recent innovations

Although comprehensive 2-D GC is an established and often applied analytical method, the field is still highly dynamic thanks to a remarkable number of innovations. In this review, we discuss a number of recent developments in comprehensive 2-D GC technology. A variety of modulation methods are still being actively investigated and many exciting improvements are discussed in this review. We also review interesting developments in detection methods, retention modeling, and data analysis.     

Comprehensive two-dimensional gas chromatography (GC×GC) in environmental analysis and monitoring

Compared to conventional one-dimensional gas chromatography (1D-GC), comprehensive two-dimensional gas chromatography (GC×GC) offers increased peak capacity, improved resolution and enhanced mass sensitivity. In addition, it generates structured two-dimensional (2-D) chromatograms, which aids in the identification of compound classes. Sample preparation procedures can often be minimized, or even eliminated in some cases, due to the superior separating power offered by the technique. All of these advantages make GC×GC a very powerful tool in environmental analysis involving the determination of trace levels of toxic compounds in complex matrices. This review paper summarizes and examines the historical and recent GC×GC applications in environmental analysis and monitoring.     

Development of a certified reference material for cachaça: an effective material for quality assurance

Cachaça is the typical and genuine denomination for the sugar cane beverage produced in Brazil. It has an alcoholic content between 38 and 48 % by volume, at 20 °C, and is obtained from the distillation of fermented sugar cane juice with the possible addition of up to 6 g/L of sugar, expressed by sucrose. This paper aims to outline the approach taken for the certification of the mass fraction of individual alcohols and their associated uncertainties in a cachaça certified reference material (CRM) produced by the Organic Analysis Laboratory of the Chemical Metrology Division of Inmetro. The requirements of ISO Guide 31, ISO Guide 34 and ISO Guide 35 were followed for the certification. This included the assessment of the degree of homogeneity of the material, the short-term stability study to determine the transport conditions, the long-term stability study to establish the shelf life and storage conditions of the CRM and the characterization process. The certified values were determined by two analytical techniques: gas chromatography coupled to mass spectrometry and gas chromatography with flame ionization detection. In order to support the routine testing of cachaça, in helping analytical laboratories validate and standardize their methods, the certified reference material (CRM 02.1/10.003b) was developed and certified for the mass fractions of methanol (9.18 mg/100 g ± 0.82 mg/100 g), 2-butanol (4.15 mg/100 g ± 0.40 mg/100 g), 1-butanol (1.30 mg/100 g ± 0.18 mg/100 g), isobutanol (24.0 mg/100 g ± 1.0 mg/100 g) and 1-propanol (29.4 mg/100 g ± 1.3 mg/100 g) in a spiked cachaça.     

Development and validation of a comprehensive two-dimensional gas chromatography–mass spectrometry method for the analysis of phytosterol oxidation products in human plasma

Phytosterol oxidation products (POPs) have been suggested to exert adverse biological effects similar to, although less severe than, their cholesterol counterparts. For that reason, their analysis in human plasma is highly relevant. Comprehensive two-dimensional gas chromatography (GC×GC) coupled with time-of-flight mass spectrometry (TOF-MS) has been proven to be an extremely powerful separation technique for the analysis of very low levels of target compounds in complex mixtures including human plasma. Thus, a GC×GC/TOF-MS method was developed and successfully validated for the simultaneous quantification of ten POPs in human plasma. The calibration curves for each compound showed correlation coefficients (R ²) better than 0.99. The detection limits were below 0.1 ng mL⁻¹. The recovery data were between 71.0% and 98.6% (RSDs <10% for all compounds validated). Good results were obtained for within- and between-day repeatability, with most values being below 10%. In addition, non-targeted sterol metabolites were also identified with the method. The concentrations of POPs found in human plasma in the current study are between 0.3 and 4.5 ng mL⁻¹, i.e., 10–100 times lower than the typical values found for cholesterol oxidation products.     

Quantitative determination of jatrophone in "cachaça" prepared with Jatropha elliptica

A method for sample preparation and analysis by high performance liquid chromatography with UV detection (HPLC-UV) was developed for analysis of jatrophone in "cacha?a" prepared with Jatropha elliptica, administered orally, employed in Brazil for the treatment of venomous snake bites. The linearity, accuracy, precision of the procedure was evaluated. Analytical curve for jatrophone was linear in the range of 16.24-81.20 microg ml(-1). The recovery of the jatrophone in the samples analyzed was 98.99-99.89%. The percent coefficient of variation for the quantitative analysis of the "cacha?a" in the analyses was under 2%.     

Comprehensive two-dimensional liquid chromatography — practical impacts of theoretical considerations. A review

A theory of comprehensive two-dimensional separations by liquid chromatographic techniques is overviewed. It includes heart-cutting and comprehensive two-dimensional separation modes, with attention to basic concepts of two-dimensional separations: resolution, peak capacity, efficiency, orthogonality and selectivity. Particular attention is paid to the effects of sample structure on the retention and advantages of a multi-dimensional HPLC for separation of complex samples according to structural correlations. Optimization of 2D separation systems, including correct selection of columns, flow-rate, fraction volumes and mobile phase, is discussed. Benefits of simultaneous programmed elution in both dimensions of LCxLC comprehensive separations are shown.     

Optimized use of a 50 μm ID secondary column in comprehensive two-dimensional gas chromatography–mass spectrometry

The objective of the present research is directed towards the optimized use of a 50 μm ID secondary column, in a comprehensive two-dimensional gas chromatography–quadrupole mass spectrometry (GC × GC–qMS) system. The analytical aim was achieved by exploiting a split-flow GC × GC approach, and a rapid-scanning qMS instrument. The stationary phase combination consisted of an apolar (silphenylene polymer) 30 m × 0.25 mm ID column, linked by means of a Y-union, to an MS-connected 1 m × 0.05 mm ID polar one [poly(ethyleneglycol)], and to a 0.20 m × 0.05 mm ID uncoated capillary segment; the latter was connected to a manually operated split-valve. It will be herein demonstrated that the split-flow GC × GC approach, successfully employed in previous H₂-based, flame ionization detection experiments, provides equally satisfactory results using mass spectrometric detection and helium as carrier gas. An optimized split-flow GC × GC–qMS method was developed and exploited for the analysis of a perfume sample. The results attained were compared with those observed using the same analytical column combination, but with no flow-splitting. It was found that it is not convenient to employ a 50 μm ID secondary column in a conventional GC × GC–MS instrument. On the contrary, the use a 50 μm ID secondary column, in a split-flow, twin-oven system, provided a good performance. A recently developed comprehensive chromatography software was used for data processing.     

A modified simplex method for multifactor optimization of selectivity in gas chromatography

A computer-assisted advanced simplex method is presented for the simultaneous optimization of multifactor ( stationary phase loading, carrier gas flow rate and column temperature ) for separation of ten compounds in gas chromatography. A three factors factorial design was used. The method was based on a special polynomial established from fifteen preliminary runs, using resolution as the selection criterion, with connection to a general simplex method. Excellent agreement is found between the predicted data and the experimental results, and most of experiments required in the general simplex method can be omitted.     

Permethylated β-cyclodextrin in liquid poly(oxyethylene) as a stationary phase for capillary gas chromatography

This paper reports the study of poly(oxyethylene) as a solvent for heptakis(2,3,6-tri-O-methyl)-β-cyclodextrin (PM-β-CD) and the potential of these mixtures for constructing enantioselective gas chromatography columns. Enantioseparations of volatile racemic mixtures using capillaries coated with 10–50% PM-β-CD diluted in Carbowax 20M were evaluated. The influence of the polymer type on retention, separation and reproducibility over a period of time is critically discussed. The poly(oxyethylene)-based columns were also compared to columns prepared by dilution of PM-β-CD in two polysiloxanes, SE-30 and SE-54, at several temperatures. Finally, we evaluated a new stationary phase prepared by crosslinking the chiral selector to the poly(oxyethylene) matrix.     

New way to quantify multiple steroidal compounds in wastewater by comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry

The applicability of comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry to the screening of steroidal compounds in wastewater is demonstrated. Advanced software was utilized to identify unknown compounds in complex two-dimensional chromatograms exploiting retention indices and two different mass spectral databases. Response factors calculated as a function of the individual mass spectra of six commercial standards at different concentrations were used to develop a model allowing the quantitation of all steroidal compounds identified in the sample. The model, based on partial least squares regression equations, provided good accuracy (prediction error < 16%) in the quantitation of steroidal compounds, so offering a valuable alternative to conventional quantitation methods where reference compounds are required for the verification of analytical measurements. Special attention was paid to the development of an exhaustive sample preparation method for the separate analysis of conjugated and free steroids in both water phase and suspended solid particles. The method, including the exploitation of chemometrics, was successfully applied to the determination of steroidal compounds in effluent and influent waters collected at a local wastewater treatment plant.     

Development of a gas chromatography — mass spectrometry method for the determination of carbon disulfide in the atmosphere

Carbon disulfide (CS₂), a relevant reduced sulfur compound in air, is well-known for its malodor and its significant effect on global atmospheric chemistry. Therefore, a reliable method for determining CS₂ in atmospheric samples has been developed based on solid-phase sampling and gas chromatography–mass spectrometry (GC–MS). Two types of solid-phase sampling supports (Orbo-32 and SKC) and the elution with organic solvents — hexane and toluene — were evaluated for low-volume outdoor sampling. Recovery studies and the standard addition method were carried out to demonstrate the proper determination of CS₂ in the absence of the influence of interferences such as ozone, hydrogen sulfide or water — important atmospheric pollutants —. The proposed methodology was validated by performing experiments in a high-volume smog chamber and by comparison with two reference optical methods, Fourier Transform Infrared (FTIR) and Differential Optical Absorption Spectroscopy (DOAS) installed in these facilities. Satisfactory analytical parameters were reported: fast analysis, a correct repeatability of 6 ± 1% and reproducibility of 14 ± 3%, and low detection limits of 0.3–0.9 pg m^? 3. Finally, the method was successfully applied to industrial samples near a pulp factory area, where a high correlation between industrial emissions and reported carbon disulfide concentrations were observed.     

Implementation of comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry for the simultaneous determination of halogenated contaminants and polycyclic aromatic hydrocarbons in fish

In the presented study, comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC?×?GC-TOFMS) was shown to be a powerful tool for the simultaneous determination of various groups of contaminants including 18 polychlorinated biphenyls (PCBs), seven polybrominated diphenyl ethers (PBDEs), and 16 polycyclic aromatic hydrocarbons (PAHs). Since different groups of analytes (traditionally analyzed separately) were included into one instrumental method, significant time savings were achieved. Following the development of an integrated sample preparation procedure for an effective and rapid isolation of several groups of contaminants from fish tissue, the GC?×?GC-TOFMS instrumental method was optimized to obtain the best chromatographic resolution and low quantification limits (LOQs) of all target analytes in a complex mixture. Using large-volume programmable temperature vaporization, the following LOQs were achieved-PCBs, 0.01-0.25 μg/kg; PBDEs, 0.025-5 μg/kg; PAHs 0.025-0.5 μg/kg. Furthermore, several capillary column combinations (BPX5, BPX50, and Rxi-17Sil-ms in the first dimension and BPX5, BPX50, Rt-LC35, and HT8 in the second dimension) were tested during the experiments, and the optimal separation of all target analytes even of critical groups of PAHs (group (a): benz[a]anthracene, cyclopenta[cd]pyrene and chrysene; group (b): benzo[b]fluoranthene, benzo[j]fluoranthene and benzo[k]fluoranthene; group (c): dibenz[ah]anthracene, indeno[1,2,3-cd]pyrene and benzo[ghi]perylene) was observed on BPX5?×?BPX50 column setup. Moreover, since the determination of target analytes was performed using TOFMS detector, further identification of other non-target compounds in real life samples was also feasible.     

A β-cyclodextrin covalent organic framework used as a chiral stationary phase for chiral separation in gas chromatography

Chiral covalent organic frameworks(CCOFs) featuring chirality, stability, and good porosity have attracted a considerable amount of attention due to their important applications, such as asymmetric catalysis, chiral separation, and chiral recognition. In this study, a β-cyclodextrin(β-CD) covalent organic framework(β-CD-COF) diluted with polysiloxane OV-1701 was explored as a novel chiral stationary phase(CSP) for gas chromatography(GC) separation of racemates. The β-CD-COF coated capillary colu...     

Evaluation of World Anti-Doping Agency criteria for anabolic agent analysis by using comprehensive two-dimensional gas chromatography–mass spectrometry

This work presents the validation study of the comprehensive two-dimensional gas chromatography (GC×GC)–time-of-flight mass spectrometry method performance in the analysis of the key World Anti-Doping Agency (WADA) anabolic agents in doping control. The relative abundance ratio, retention time, identification and other method performance criteria have been tested in the GC×GC format to confirm that they comply with those set by WADA. Furthermore, tens of other components were identified with an average similarity of >920 (on the 0–999 scale), including 10 other endogenous sterols, and full mass spectra of 5,000+ compounds were retained. The testosterone/epitestosterone ratio was obtained from the same run. A new dimension in doping analysis has been implemented by addressing separation improvement. Instead of increasing the method sensitivity, which is accompanied by making the detector increasingly “blind” to the matrix (as represented by selected ion monitoring mode, high-resolution mass spectrometry (MS) and tandem MS), the method capabilities have been improved by adding a new “separation” dimension while retaining full mass spectral scan information. Apart from the requirement for the mass spectral domain that a minimum of three diagnostic ions with relative abundance of 5% or higher in the MS spectra, all other WADA criteria are satisfied by GC×GC operation. The minimum of three diagnostic ions arises from the need to add some degree of specificity to the acquired mass spectrometry data; however, under the proposed full MS scan method, the high MS similarity to the reference compounds offers more than the required three diagnostic ions for an unambiguous identification. This should be viewed as an extension of the present criteria to a full-scan MS method.     

Selectable one-dimensional or two-dimensional gas chromatography–mass spectrometry with simultaneous olfactometry or element-specific detection

A novel selectable one-dimensional (¹D) or two-dimensional (²D) gas chromatography–mass spectrometry (selectable ¹D/²D GC–MS) system with selective detection was developed by using capillary flow technology and low thermal mass GC (LTM-GC). The main advantages of this system are the simple and fast selection of ¹D GC–MS or ²D GC–MS operation without any instrumental set-up change (e.g.²D GC–MS can be run just after ¹D GC–MS run), and simultaneous mass spectrometric and olfactometry or element-specific detection for both ¹D and ²D separation to assure selection of a heart-cut region and correct identification of compounds of interest. The feasibility and benefit of the proposed system with selective detection, e.g. olfactometry, nitrogen phosphorus detection (NPD), and pulsed flame photometric detection (PFPD), was demonstrated with an identification of trace amounts of aroma components in beverages (beer and coffee). Using stir bar sorptive extraction (SBSE) and selectable ¹D/²D GC–Olfactometry/MS on a beer sample, β-damascenone could be determined at 1.9 ng mL⁻¹ (RSD 3.1%, n = 6) as a potent aroma compound. In a coffee sample, two odor active compounds were clearly resolved from a 4.2 s heart-cut and were assigned probable identifications as 4,5-dimethyl thiazole and dimethyl trisulfide based on a NIST library search, dual linear retention indices (dual LRI) and elemental information obtained by SBSE in combination with selectable ¹D/²D GC–NPD/PFPD/MS.     

Solving chromatographic challenges in comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry using multivariate curve resolution–alternating least squares

Multivariate curve resolution–alternating least squares (MCR–ALS) analysis is proposed to solve chromatographic challenges during two-dimensional gas chromatography–time-of-flight mass spectrometry (GC?×?GC–TOFMS) analysis of complex samples, such as crude oil extract. In view of the fact that the MCR–ALS method is based on the fulfillment of the bilinear model assumption, three-way and four-way GC?×?GC–TOFMS data are preferably arranged in a column-wise superaugmented data matrix in which mass-to-charge ratios (m/z) are in its columns and the elution times in the second and first chromatographic columns are in its rows. Since m/z values are common for all measured spectra in all second-column modulations, unavoidable chromatographic challenges such as retention time shifts within and between GC?×?GC–TOFMS experiments are properly handled. In addition, baseline/background contributions can be modeled by adding extra components to the MCR–ALS model. Another outstanding aspect of MCR–ALS analysis is its extreme flexibility to consider all samples (standards, unknowns, and replicates) in a single superaugmented data matrix, allowing joint analysis. In this way, resolution, identification, and quantification results can be simultaneously obtained in a very fast and reliable way. The potential of MCR–ALS analysis is demonstrated in GC?×?GC–TOFMS analysis of a North Sea crude oil extract sample with relative errors in estimated concentrations of target compounds below 6.0 % and relative standard deviations lower than 7.0 %. The results obtained, along with reasonable values for the lack of fit of the MCR–ALS model and high values of the reversed match factor in mass spectra similarity searches, confirm the reliability of the proposed strategy for GC?×?GC–TOFMS data analysis.      

Optimization of a method for the determination of a mustard gas biomarker in human blood plasma by liquid chromatography–high-resolution mass spectrometry

A method for the determination of a mustard gas biomarker (an S-hydroxyethylthioethyl adduct with albumin) in blood plasma was optimized with the use of HPLC with high-resolution tandem mass-spectrometric detection. This method is based on the hydrolysis of this adduct by the proteinase K enzyme with the formation of the following stable tripeptide with cysteine, proline, and phenylalanine: S-[HETE]-Cys- Pro-Phe. The conditions of the sample preparation of human plasma artificially contaminated by mustard gas (the selection of an aliquot portion volume and an enzyme for the hydrolysis), the mass-spectrometric detection (the selection of optimum pairs of ion reactions and high-resolution detection modes), and the gradient elution program in the HPLC separation of an analyzed mixture were optimized. The detection limit of mustard gas in human blood plasma was 1 ng/mL. The approach proposed was tested in the analysis of human blood plasma samples by the standard addition technique and also within the framework of the first official biomedical test carried out by the Organization for the Prohibition of Chemical Weapons (OPCW) in 2016, and it exhibited a good accuracy, reproducibility, and specificity of determination.