Discovery and Validation of a Volatile Signature of Eosinophilic Airway Inflammation in Asthma.

RATIONALE: Volatile organic compounds (VOCs) in asthmatic breath may be associated with sputum eosinophilia. We developed a volatile biomarker-signature to predict sputum eosinophilia in asthma. METHODS: VOCs emitted into the space above sputum samples (headspace) from severe asthmatics (n=36) were collected onto sorbent tubes and analysed using thermal desorption gas chromatography-mass spectrometry (TD-GC-MS). Elastic net regression identified stable VOCs associated with sputum eosinophilia ≥3% and generated a volatile biomarker signature. This VOC signature was validated in breath samples from: (I) acute asthmatics according to blood eosinophilia ≥0.3x109cells/L or sputum eosinophilia of ≥ 3% in the UK EMBER consortium (n=65) and U-BIOPRED-IMI consortium (n=42). Breath samples were collected onto sorbent tubes (EMBER) or Tedlar bags (U-BIOPRED) and analysed by gas-chromatography-mass spectrometry (GC×GC-MS -EMBER or GC-MS -U-BIOPRED). MAIN RESULTS: The in vitro headspace identified 19 VOCs associated with sputum eosinophilia and the derived VOC signature yielded good diagnostic accuracy for sputum eosinophilia ≥ 3% in headspace (AUROC (95% CI) 0.90(0.80-0.99), p<0.0001), correlated inversely with sputum eosinophil % (rs= -0.71, p<0.0001) and outperformed FeNO (AUROC (95% CI) 0.61(0.35-0.86). Analysis of exhaled breath in replication cohorts yielded a VOC signature AUROC (95% CI) for acute asthma exacerbations of 0.89(0.76-1.0) (EMBER cohort) with sputum eosinophilia and 0.90(0.75-1.0) in U-BIOPRED - again outperforming FeNO in U-BIOPRED 0.62 (0.33-0.90). CONCLUSIONS: We have discovered and provided early-stage clinical validation of a volatile biomarker signature associated with eosinophilic airway inflammation. Further work is needed to translate our discovery using point of care clinical sensors.

Tile-Based Random Forest Analysis for Analyte Discovery in Balanced and Unbalanced GC × GC-TOFMS Data Sets.

In this study, we introduce a new nontargeted tile-based supervised analysis method that combines the four-grid tiling scheme previously established for the Fisher ratio (F-ratio) analysis (FRA) with the estimation of tile hit importance using the machine learning (ML) algorithm Random Forest (RF). This approach is termed tile-based RF analysis. As opposed to the standard tile-based F-ratio analysis, the RF approach can be extended to the analysis of unbalanced data sets, i.e., different numbers of samples per class. Tile-based RF computes out-of-bag (oob) tile hit importance estimates for every summed chromatographic signal within each tile on a per-mass channel basis (m/z). These estimates are then used to rank tile hits in a descending order of importance. In the present investigation, the RF approach was applied for a two-class comparison of stool samples collected from omnivore (O) subjects and stored using two different storage conditions: liquid (Liq) and lyophilized (Lyo). Two final hit lists were generated using balanced (8 vs Eight comparison) and unbalanced (8 vs Nine comparison) data sets and compared to the hit list generated by the standard F-ratio analysis. Similar class-distinguishing analytes (p < 0.01) were discovered by both methods. However, while the FRA discovered a more comprehensive hit list (65 hits), the RF approach strictly discovered hits (31 hits for the balanced data set comparison and 29 hits for the unbalanced data set comparison) with concentration ratios, [OLiq]/[OLyo], greater than 2 (or less than 0.5). This difference is attributed to the more stringent feature selection process used by the RF algorithm. Moreover, our findings suggest that the RF approach is a promising method for identifying class-distinguishing analytes in settings characterized by both high between-class variance and high within-class variance, making it an advantageous method in the study of complex biological matrices.

Comprehensive Insight into Colorectal Cancer Metabolites and Lipids for Human Serum: A Proof-of-Concept Study.

Colorectal cancer (CRC) ranks as the third most frequently diagnosed cancer and the second leading cause of cancer-related deaths. The current endoscopic-based or stool-based diagnostic techniques are either highly invasive or lack sufficient sensitivity. Thus, there is a need for less invasive and more sensitive screening approaches. We, therefore, conducted a study on 64 human serum samples representing three different groups (adenocarcinoma, adenoma, and control) using cutting-edge GC×GC-LR/HR-TOFMS (comprehensive two-dimensional gas chromatography coupled with low/high-resolution time-of-flight mass spectrometry). We analyzed samples with two different specifically tailored sample preparation approaches for lipidomics (fatty acids) (25 µL serum) and metabolomics (50 µL serum). In-depth chemometric screening with supervised and unsupervised approaches and metabolic pathway analysis were applied to both datasets. A lipidomics study revealed that specific PUFA (ω-3) molecules are inversely associated with increased odds of CRC, while some PUFA (ω-6) analytes show a positive correlation. The metabolomics approach revealed downregulation of amino acids (alanine, glutamate, methionine, threonine, tyrosine, and valine) and myo-inositol in CRC, while 3-hydroxybutyrate levels were increased. This unique study provides comprehensive insight into molecular-level changes associated with CRC and allows for a comparison of the efficiency of two different analytical approaches for CRC screening using same serum samples and single instrumentation.

Top-Down Approach to Retention Time Prediction in Comprehensive Two-Dimensional Gas Chromatography-Mass Spectrometry.

In this contribution, we describe a novel modeling approach to predicting retention times (tr) in comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC × GC-ToF-MS) with a particular emphasis on the second-dimension (2D) retention time predictions (2tr). This approach is referred to as a "top-down" approach in that it breaks down the complete GC × GC separation into two independent one-dimensional gas chromatography separations (1D-GC). In this regard, both dimensions, that is, first dimension (1D) and second dimension (2D) are treated separately, and the cryogenic modulator is simply considered as a second consecutive injection device. Separate 1D-GC tr predictions are performed on both dimensions using the same flow rate as the one deployed in the conventional GC × GC system. The separate tr predictions are then combined to account for the two-dimensional separation. This model was applied to 24 analytes from 2 standard mixtures (Grob Test Mix and Fragrance Materials Test Mix) and assessed across 9 GC × GC chromatographic conditions. The experimental and predicted chromatographic retention space occupations were assessed by using the convex hull approach defined by the Delaunay triangulation. The predicted percentage of space occupation corresponded favorably with the experimental values. Furthermore, the top-down approach enabled an accurate prediction of the 2tr of all investigated analytes, providing an average 2tr modeling error of 0.26 ± 0.01 s.

Comprehensive two-dimensional gas chromatographic platforms comparison for exhaled breath metabolites analysis.

The high potential of exhaled breath for disease diagnosis has been highlighted in numerous studies. However, exhaled breath analysis is suffering from a lack of standardized sampling and analysis procedures, impacting the robustness of inter-laboratory results, and thus hampering proper external validation. The aim of this work was to verify compliance and validate the performance of two different comprehensive two-dimensional gas chromatography coupled to mass spectrometry platforms in different laboratories by monitoring probe metabolites in exhaled breath following the Peppermint Initiative guidelines. An initial assessment of the exhaled breath sampling conditions was performed, selecting the most suitable sampling bag material and volume. Then, a single sampling was performed using Tedlar bags, followed by the trapping of the volatile organic compounds into thermal desorption tubes for the subsequent analysis using two different analytical platforms. The thermal desorption tubes were first analyzed by a (cryogenically modulated) comprehensive two-dimensional gas chromatography system coupled to high-resolution time-of-flight mass spectrometry. The desorption was performed in split mode and the split part was recollected in the same tube and further analyzed by a different (flow modulated) comprehensive two-dimensional gas chromatography system with a parallel detection, specifically using a quadrupole mass spectrometer and a vacuum ultraviolet detector. Both the comprehensive two-dimensional gas chromatography platforms enabled the longitudinal tracking of the peppermint oil metabolites in exhaled breath. The increased sensitivity of comprehensive two-dimensional gas chromatography enabled to successfully monitor over a 6.5 h period a total of 10 target compounds, namely α-pinene, camphene, ß-pinene, limonene, cymene, eucalyptol, menthofuran, menthone, isomenthone, and neomenthol.

Distinguishing between Decaffeinated and Regular Coffee by HS-SPME-GC×GC-TOFMS, Chemometrics, and Machine Learning.

Coffee, one of the most popular beverages in the world, attracts consumers by its rich aroma and the stimulating effect of caffeine. Increasing consumers prefer decaffeinated coffee to regular coffee due to health concerns. There are some main decaffeination methods commonly used by commercial coffee producers for decades. However, a certain amount of the aroma precursors can be removed together with caffeine, which could cause a thin taste of decaffeinated coffee. To understand the difference between regular and decaffeinated coffee from the volatile composition point of view, headspace solid-phase microextraction two-dimensional gas chromatography time-of-flight mass spectrometry (HS-SPME-GC×GC-TOFMS) was employed to examine the headspace volatiles of eight pairs of regular and decaffeinated coffees in this study. Using the key aroma-related volatiles, decaffeinated coffee was significantly separated from regular coffee by principal component analysis (PCA). Using feature-selection tools (univariate analysis: t-test and multivariate analysis: partial least squares-discriminant analysis (PLS-DA)), a group of pyrazines was observed to be significantly different between regular coffee and decaffeinated coffee. Pyrazines were more enriched in the regular coffee, which was due to the reduction of sucrose during the decaffeination process. The reduction of pyrazines led to a lack of nutty, roasted, chocolate, earthy, and musty aroma in the decaffeinated coffee. For the non-targeted analysis, the random forest (RF) classification algorithm was used to select the most important features that could enable a distinct classification between the two coffee types. In total, 20 discriminatory features were identified. The results suggested that pyrazine-derived compounds were a strong marker for the regular coffee group whereas furan-derived compounds were a strong marker for the decaffeinated coffee samples.

Breathomics to diagnose systemic sclerosis using thermal desorption and comprehensive two-dimensional gas chromatography high-resolution time-of-flight mass spectrometry.

Systemic sclerosis is a rare autoimmune disease associated with rapidly evolving interstitial lung disease, responsible for the disease severity and mortality. Specific biomarkers enabling the early diagnosis and prognosis associated with the disease progression are highly needed. Volatile organic compounds in exhaled breath are widely available and non-invasive and have the potential to reflect metabolic processes occurring within the body. Comprehensive two-dimensional gas chromatography coupled to high-resolution mass spectrometry was used to investigate the potential of exhaled breath to diagnose systemic sclerosis. The exhaled breath of 32 patients and 30 healthy subjects was analyzed. The high resolving power of this approach enabled the detection of 356 compounds in the breath of systemic sclerosis patients, which was characterized by an increase of mainly terpenoids and hydrocarbons. In addition, the use of 4 complementary statistical approaches (two-tailed equal variance t-test, fold change, partial least squares discriminant analysis, and random forest) resulted in the identification of 16 compounds that can be used to discriminate systemic sclerosis patients from healthy subjects. Receiver operating curves were generated that provided an accuracy of 90%, a sensitivity of 92%, and a specificity of 89%. The chemical identification of eight compounds predictive of systemic sclerosis was validated using commercially available standards. The analytical variations together with the volatile composition of room air were carefully monitored during the timeframe of the study to ensure the robustness of the technique. This study represents the first reported evaluation of exhaled breath analysis for systemic sclerosis diagnosis and provides surrogate markers for such disease.

Advanced mono- and multi-dimensional gas chromatography-mass spectrometry techniques for oxygen-containing compound characterization in biomass and biofuel samples.

A wide variety of biomass, from triglycerides to lignocellulosic-based feedstock, are among promising candidates to possibly fulfill requirements as a substitute for crude oils as primary sources of chemical energy feedstock. During the feedstock processing carried out to increase the H:C ratio of the products, heteroatom-containing compounds can promote corrosion, thus limiting and/or deactivating catalytic processes needed to transform the biomass into fuel. The use of advanced gas chromatography techniques, in particular multi-dimensional gas chromatography, both heart-cutting and comprehensive coupled to mass spectrometry, has been widely exploited in the field of petroleomics over the past 30 years and has also been successfully applied to the characterization of volatile and semi-volatile compounds during the processing of biomass feedstock. This review intends to describe advanced gas chromatography-mass spectrometry-based techniques, mainly focusing in the period 2011-early 2020. Particular emphasis has been devoted to the multi-dimensional gas chromatography-mass spectrometry techniques, for the isolation and characterization of the oxygen-containing compounds in biomass feedstock. Within this context, the most recent advances to sample preparation, derivatization, as well as gas chromatography instrumentation, mass spectrometry ionization, identification, and data handling in the biomass industry, are described.

Comparison of the effect of chemically and biologically induced inflammation on the volatile metabolite production of lung epithelial cells by GC×GC-TOFMS.

Exhaled breath analysis has a high potential for early non-invasive diagnosis of lung inflammatory diseases, such as asthma. The characterization and understanding of the inflammatory metabolic pathways involved into volatile organic compounds (VOCs) production could bring exhaled breath analysis into clinical practice and thus open new therapeutic routes for inflammatory diseases. In this study, lung inflammation was simulated in vitro using A549 epithelial cells. We compared the VOC production from A549 epithelial cells after a chemically induced oxidative stress in vitro, exposing the cells to H2O2, and a biological stress, exposing the cells to an inflammatory pool of sputum supernatants. Special attention was devoted to define proper negative and positive controls (8 different types) for our in vitro models, including healthy sputum co-culture. Sputum from 25 asthmatic and 8 healthy patients were collected to create each pool of supernatants. Each sample type was analyzed in 4 replicates using solid-phase microextraction (SPME) comprehensive two-dimensional gas chromatography hyphenated to time-of-flight mass spectrometry (GC×GC-TOFMS). This approach offers high resolving power for complex VOC mixtures. According to the type of inflammation induced, significantly different VOCs were produced by the epithelial cells compared to all controls. For both chemical and biological challenges, an increase of carbonyl compounds (54%) and hydrocarbons (31%) was observed. Interestingly, only the biological inflammation model showed a significant cell proliferation together with an increased VOC production linked to asthma airway inflammation. This study presents a complete GC×GC-TOFMS workflow for in vitro VOC analysis, and its potential to characterize complex lung inflammatory mechanisms.

Investigating aroma diversity combining purge-and-trap, comprehensive two-dimensional gas chromatography, and mass spectrometry.

Headspace gas chromatography is frequently used for aroma profiling thanks to its ability to naturally exploit the volatility of aroma compounds, and also to provide chemical information on sample composition. Its main advantages rely on simplicity, no use of solvent, amenability to automation, and the cleanliness of the extract. In the present contribution, the most effective sampling (dynamic extraction), separation (multidimensional gas chromatography), and detection (mass spectrometry) techniques for untargeted analysis are exploited in combination, showing their potential in unraveling aroma profiles in fruit beers. To complete the overall analytical process, a neat workflow for data analysis is discussed and used for the successful characterization and identification of five different beer flavors (berries, cherry, banana, apple, and peach). From the technical viewpoint, the coupling of purge-and-trap, comprehensive two-dimensional gas chromatography, and mass spectrometry makes the global methodology unique, and it is for the first time discussed. A (low-)flow modulation approach allowed for the full transfer into the second dimension with mass-spectrometry compatible flow (< 7 mL/min), avoiding the need of splitting before detection and making the overall method sensitive (1.2-5.2-fold higher signal to noise ratio compared to unmodulated gas chromatography conditions) and selective.

Exhaled Volatile Organic Compounds Are Able to Discriminate between Neutrophilic and Eosinophilic Asthma.

Rationale: Analysis of exhaled breath for asthma phenotyping using endogenously generated volatile organic compounds (VOCs) offers the possibility of noninvasive diagnosis and therapeutic monitoring. Induced sputum is indeed not widely available and markers of neutrophilic asthma are still lacking.Objectives: To determine whether analysis of exhaled breath using endogenously generated VOCs can be a surrogate marker for recognition of sputum inflammatory phenotypes.Methods: We conducted a prospective study on 521 patients with asthma recruited from the University Asthma Clinic of Liege. Patients underwent VOC measurement, fraction of exhaled nitric oxide (FeNO) spirometry, sputum induction, and gave a blood sample. Subjects with asthma were classified in three inflammatory phenotypes according to their sputum granulocytic cell count.Measurements and Main Results: In the discovery study, seven potential biomarkers were highlighted by gas chromatography-mass spectrometry in a training cohort of 276 patients with asthma. In the replication study (n = 245), we confirmed four VOCs of interest to discriminate among asthma inflammatory phenotypes using comprehensive two-dimensional gas chromatography coupled to high-resolution time-of-flight mass spectrometry. Hexane and 2-hexanone were identified as compounds with the highest classification performance in eosinophilic asthma with accuracy comparable to that of blood eosinophils and FeNO. Moreover, the combination of FeNO, blood eosinophils, and VOCs gave a very good prediction of eosinophilic asthma (area under the receiver operating characteristic curve, 0.9). For neutrophilic asthma, the combination of nonanal, 1-propanol, and hexane had a classification performance similar to FeNO or blood eosinophils in eosinophilic asthma. Those compounds were found in higher levels in neutrophilic asthma.Conclusions: Our study is the first attempt to characterize VOCs according to sputum granulocytic profile in a large population of patients with asthma and provide surrogate markers for neutrophilic asthma.

A minimally-invasive method for profiling volatile organic compounds within postmortem internal gas reservoirs.

In forensic casework, non-invasive and minimally-invasive methods for postmortem examinations are extremely valuable. Whole body postmortem computed tomography (PMCT) is often used to provide visualization of the internal characteristics of a body prior to more invasive procedures and has also been used to locate gas reservoirs inside the body to assist in determining cause of death. Preliminary studies have demonstrated that exploiting the volatile organic compounds (VOCs) located in these gas reservoirs by comprehensive two-dimensional gas chromatography-high-resolution time-of-flight mass spectrometry (GC×GC-HRTOF-MS) may assist in providing information regarding the postmortem interval. The aim of the current study was to further develop the procedures related to solid-phase microextraction (SPME) and GC×GC-HRTOF-MS analysis of gas reservoirs collected from deceased individuals. SPME fiber extraction parameters, internal standard approach, and sample stability were investigated. Altering the SPME parameters increased the selectivity and sensitivity for the VOC profile, and the use of a mixed deuterated internal standard contributed to data quality. Samples were found to be stable up to 6 weeks but were recommended to be analyzed within 4 weeks due to higher variation observed beyond this point. In addition, 29 VOC markers of interest were identified, and heart and/or abdominal cavity samples were suggested as a possible standardized sampling location for future studies. The data presented in this study will contribute to the long-term goal of producing a routine, accredited method for minimally-invasive VOC analysis in postmortem examinations.

GC × GC-TOFMS and supervised multivariate approaches to study human cadaveric decomposition olfactive signatures.

In forensic thanato-chemistry, the understanding of the process of soft tissue decomposition is still limited. A better understanding of the decomposition process and the characterization of the associated volatile organic compounds (VOC) can help to improve the training of victim recovery (VR) canines, which are used to search for trapped victims in natural disasters or to locate corpses during criminal investigations. The complexity of matrices and the dynamic nature of this process require the use of comprehensive analytical methods for investigation. Moreover, the variability of the environment and between individuals creates additional difficulties in terms of normalization. The resolution of the complex mixture of VOCs emitted by a decaying corpse can be improved using comprehensive two-dimensional gas chromatography (GC × GC), compared to classical single-dimensional gas chromatography (1DGC). This study combines the analytical advantages of GC × GC coupled to time-of-flight mass spectrometry (TOFMS) with the data handling robustness of supervised multivariate statistics to investigate the VOC profile of human remains during early stages of decomposition. Various supervised multivariate approaches are compared to interpret the large data set. Moreover, early decomposition stages of pig carcasses (typically used as human surrogates in field studies) are also monitored to obtain a direct comparison of the two VOC profiles and estimate the robustness of this human decomposition analog model. In this research, we demonstrate that pig and human decomposition processes can be described by the same trends for the major compounds produced during the early stages of soft tissue decomposition.

Reducing variation in decomposition odour profiling using comprehensive two-dimensional gas chromatography.

Challenges in decomposition odour profiling have led to variation in the documented odour profile by different research groups worldwide. Background subtraction and use of controls are important considerations given the variation introduced by decomposition studies conducted in different geographical environments. The collection of volatile organic compounds (VOCs) from soil beneath decomposing remains is challenging due to the high levels of inherent soil VOCs, further confounded by the use of highly sensitive instrumentation. This study presents a method that provides suitable chromatographic resolution for profiling decomposition odour in soil by comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry using appropriate controls and field blanks. Logarithmic transformation and t-testing of compounds permitted the generation of a compound list of decomposition VOCs in soil. Principal component analysis demonstrated the improved discrimination between experimental and control soil, verifying the value of the data handling method. Data handling procedures have not been well documented in this field and standardisation would thereby reduce misidentification of VOCs present in the surrounding environment as decomposition byproducts. Uniformity of data handling and instrumental procedures will reduce analytical variation, increasing confidence in the future when investigating the effect of taphonomic variables on the decomposition VOC profile.

Detection of decomposition volatile organic compounds in soil following removal of remains from a surface deposition site.

PURPOSE: Cadaver-detection dogs use volatile organic compounds (VOCs) to search for human remains including those deposited on or beneath soil. Soil can act as a sink for VOCs, causing loading of decomposition VOCs in the soil following soft tissue decomposition. The objective of this study was to chemically profile decomposition VOCs from surface decomposition sites after remains were removed from their primary location. METHODS: Pig carcasses were used as human analogues and were deposited on a soil surface to decompose for 3 months. The remains were then removed from each site and VOCs were collected from the soil for 7 months thereafter and analyzed by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS). RESULTS: Decomposition VOCs diminished within 6 weeks and hydrocarbons were the most persistent compound class. Decomposition VOCs could still be detected in the soil after 7 months using Principal Component Analysis. CONCLUSIONS: This study demonstrated that the decomposition VOC profile, while detectable by GC×GC-TOFMS in the soil, was considerably reduced and altered in composition upon removal of remains. Chemical reference data is provided by this study for future investigations of canine alert behavior in scenarios involving scattered or scavenged remains.

Development and validation of a classification model for boar taint detection in pork fat samples.

This study aims to characterize a complete volatile organic compound profile of pork neck fat for boar taint prediction. The objectives are to identify specific compounds related to boar taint and to develop a classification model. In addition to the well-known androstenone, skatole and indole, 10 other features were found to be discriminant according to untargeted volatolomic analyses were conducted on 129 samples using HS-SPME-GC×GC-TOFMS. To select the odor-positive samples among the 129 analyzed, the selection was made by combining human nose evaluations with the skatole and androstenone concentrations determined using UHPLC-MS/MS. A comparison of the data of the two populations was performed and a statistical model analysis was built on 70 samples out of the total of 129 samples fully positive or fully negative through these two orthogonal methods for tainted prediction. Then, the model was applied to the 59 remaining samples. Finally, 7 samples were classified as tainted.

Characterization of volatile organic compounds from human analogue decomposition using thermal desorption coupled to comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry.

Complex processes of decomposition produce a variety of chemicals as soft tissues, and their component parts are broken down. Among others, these decomposition byproducts include volatile organic compounds (VOCs) responsible for the odor of decomposition. Human remains detection (HRD) canines utilize this odor signature to locate human remains during police investigations and recovery missions in the event of a mass disaster. Currently, it is unknown what compounds or combinations of compounds are recognized by the HRD canines. Furthermore, a comprehensive decomposition VOC profile remains elusive. This is likely due to difficulties associated with the nontarget analysis of complex samples. In this study, cadaveric VOCs were collected from the decomposition headspace of pig carcasses and were further analyzed using thermal desorption coupled to comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (TD-GC × GC-TOFMS). Along with an advanced data handling methodology, this approach allowed for enhanced characterization of these complex samples. The additional peak capacity of GC × GC, the spectral deconvolution algorithms applied to unskewed mass spectral data, and the use of a robust data mining strategy generated a characteristic profile of decomposition VOCs across the various stages of soft-tissue decomposition. The profile was comprised of numerous chemical families, particularly alcohols, carboxylic acids, aromatics, and sulfides. Characteristic compounds identified in this study, e.g., 1-butanol, 1-octen-3-ol, 2-and 3-methyl butanoic acid, hexanoic acid, octanal, indole, phenol, benzaldehyde, dimethyl disulfide, and trisulfide, are potential target compounds of decomposition odor. This approach will facilitate the comparison of complex odor profiles and produce a comprehensive VOC profile for decomposition.

Theoretical modeling and machine learning-based data processing workflows in comprehensive two-dimensional gas chromatography-A review.

In recent years, comprehensive two-dimensional gas chromatography (GC × GC) has been gradually gaining prominence as a preferred method for the analysis of complex samples due to its higher peak capacity and resolution power compared to conventional gas chromatography (GC). Nonetheless, to fully benefit from the capabilities of GC × GC, a holistic approach to method development and data processing is essential for a successful and informative analysis. Method development enables the fine-tuning of the chromatographic separation, resulting in high-quality data. While generating such data is pivotal, it does not necessarily guarantee that meaningful information will be extracted from it. To this end, the first part of this manuscript reviews the importance of theoretical modeling in achieving good optimization of the separation conditions, ultimately improving the quality of the chromatographic separation. Multiple theoretical modeling approaches are discussed, with a special focus on thermodynamic-based modeling. The second part of this review highlights the importance of establishing robust data processing workflows, with a special emphasis on the use of advanced data processing tools such as, Machine Learning (ML) algorithms. Three widely used ML algorithms are discussed: Random Forest (RF), Support Vector Machine (SVM), and Partial Least Square-Discriminate Analysis (PLS-DA), highlighting their role in discovery-based analysis.

Analysis of mixed plastic pyrolysis oil by comprehensive two-dimensional gas chromatography coupled with low- and high-resolution time-of-flight mass spectrometry with the support of soft ionization.

According to the annual production of plastics worldwide, in 2020 about 370 million tons of plastic were produced in the world. Chemical recycling, particularly pyrolysis of plastic wastes, could be a valuable solution to resolve these problems and provide an alternative pathway to produce "recycled" chemical products for the petrochemical industry. Nevertheless, the pyrolysis oils need a detailed characterization before the upgrading test to re-use them to generate new recycled products. Multidimensional gas chromatography coupled with both low- and high-resolution time-of-flight mass spectrometers was employed for a detailed investigation among and within different chemical classes present in bio-plastic oil. The presence of several isomeric species as well as homologs series did not allow a reliable molecular identification, except for a few compounds that showed both MS similarity >800/1000 and retention index within ±20. Indeed, the identification of several isomeric species was assessed by high-resolution mass spectrometry equipped with photoionization interface. This soft ionization mode was an additional filter in the identification step allowing unambiguous identification of analytes not identified by the standard electron ionization mode at 70 eV. The injection method was also optimized using a central composite design to successfully introduce a wide range of carbon number compounds without discrimination of low/high boiling points.

Lipid Serum Profiling of Boar-Tainted and Untainted Pigs Using GC×GC-TOFMS: An Exploratory Study.

Mass spectrometry (MS)-based techniques, including liquid chromatography coupling, shotgun lipidomics, MS imaging, and ion mobility, are widely used to analyze lipids. However, with enhanced separation capacity and an optimized chemical derivatization approach, comprehensive two-dimensional gas chromatography (GC×GC) can be a powerful tool to investigate some groups of small lipids in the framework of lipidomics. This study describes the optimization of a dedicated two-stage derivatization and extraction process to analyze different saturated and unsaturated fatty acids in plasma by two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) using a full factorial design. The optimized condition has a composite desirability of 0.9159. This optimized sample preparation and chromatographic condition were implemented to differentiate between positive (BT) and negative (UT) boar-tainted pigs based on fatty acid profiling in pig serum using GC×GC-TOFMS. A chemometric screening, including unsupervised (PCA, HCA) and supervised analysis (PLS-DA), as well as univariate analysis (volcano plot), was performed. The results suggested that the concentration of PUFA ω-6 and cholesterol derivatives were significantly increased in BT pigs, whereas SFA and PUFA ω-3 concentrations were increased in UT pigs. The metabolic pathway and quantitative enrichment analysis suggest the significant involvement of linolenic acid metabolism.