Improving analytical and annotation robustness in small molecule metabolomics using GCxGC-HRTOFMS

editorial reviewedThis presentation focus on the enhanced annotation and detectability capacity provided by high resolution information contained in multidimensional chromatography data. The presentation will focus on the combination of untargeted screening of small metabolites and small lipids in human blood serum samples from different form of colorectal cancers

Profiling and comparison of five bottled refermented handmade beers analyzed by comprehensive twodimensional gas chromatography time-of-flight mass spectrometry

Fermented beverage, such as beer, are considered as complex mixtures by flavor and food chemists. The beer volatolome contains hundreds of compounds, which affect the taste and the mouthfeel. In this highly competitive market, it is crucial for brewers to master each step of the process to understand and control the taste and to guarantee aroma quality and stability. Aroma compounds are essentially derived from the raw ingredients and the conditions used to produce beers, including the fermenting yeast strain. This study aims to determine the impact of the yeast on beer’s volatolome. More specifically, the impact of yeast used for bottle fermentation. On microbrewery set-up, we brewed 5 different beers with the same feedstock and fermenting yeast. During the bottling, we used 5 different yeasts generally used by brewers. After an aging period of 8 weeks, all samples were analyzed using solid-phase microextraction couple with comprehensive two-dimensional gas chromatography coupled to mass spectrometry (SPME-GC×GC-TOFMS). A dedicated data processing workflow was then optimized. Principal component analysis (PCA) was used to visualize the clustering between sample types. The resulting clusters were compared with classical beer quality metrics, such as isoamyl acetate and polyphenols concentrations. After data curation, peaks area of VOCs founds in each beer were used to carry out unsupervised statistical analyses. Two distinct groups were observed on PCA; two beers are considered very similar regarding the VOCs while the remaining three constitute the second cluster. Based on these results, the aroma profiles of the different beers could be dressed. While the group of two beers was dominated by ethyl esters giving fruity aromas, less associated fruity fragrance esters were expressed in the other group. The beer aroma profile was impacted by the yeast strain used for the bottle fermentation. Specific esters were produced, resulting in beer-type classification on PCA

Development of an analytical workflow for bronchoalveolar lavage fluids analysis using GCxGC-TOFMS

editorial reviewe

Identification d'un profil de composés organiques volatils pour la détection de l'odeur de verra dans les échantillons de graisse par GC×GC-TOFMS de l'espace de tête.

peer reviewe

La TD-GC×GC-HRTOFMS pour investiguer la fibrose pulmonaire chez des patients

Comprehensive two-dimensional gas chromatography has a great potential for exhaled breath analysis. The increased peak capacity and sensitivity, provided by the combination of two capillary columns of different stationary phases by means of a modulator, enable the chromatographic separation and detection of thousands of compounds from a complex matrix. For this reason, we carried out an exploratory study on SSc. Basically, breath samples were collected in 5L Tedlar® bags. Volatiles contained in the sampling bag were then transferred onto Tenax®GR/Carbopack™B thermal desorption tubes and finally released and separated into a Pegasus GC-HRT 4D through a mid-polar Rxi-624SilMS as first column (dimension) and a polar Stabilwax as second dimension. The exhaled breath of 32 patients and 30 healthy subjects was therefore analyzed. The high resolving power of this approach and the use of statistical models enabled the identification of 16 compounds discriminating SSC patients from healthy ones. However, further investigations had to be held to reach a better disease classification. In fact, the biomarkers highlighted here could be related to the scarring of the lungs making these non-specific to SSCs. The second phase of the study aims to go deeper in patient stratification. Three groups were investigated: 50 SSC patients, 50 SSC-fibrosis patients and 50 ILD ones. The samples were collected at Maastricht medical center and CHU of Liège. All samples were then analyzed in the OBiACHem lab. Currently, a classification model is under construction to stratify patients based on their fibrosis status

BREATHOMICS APPROACH TO INVESTIGATE SYSTEMIC SCLEROSIS USING THERMAL DESORPTION AND COMPREHENSIVE TWO-DIMENSIONAL GAS CHROMATOGRAPHY HIGH-RESOLUTION TIME-OF-FLIGHT MASS SPECTROMETRY

Systemic sclerosis (SSc) is a chronic and heterogenous auto-immune disease of unknown origin characterized by fibrosis, inflammation, vascular damages, and involvement of internal organs. Organ involvement appears at the early stage of the disease[1,2]. Interstitial lung disease (ILD) is one of the most common types of pulmonary involvement, responsible for the disease severity, and leading to high morbidity and mortality. One of the challenges in SSc remains the early diagnosis of patients with a high risk of disease progression driving mortality[3]. There is an unmet need for biological markers enabling SSc early diagnosis, prognosis, disease progression monitoring, and improving patients’ classification for more targeted therapies. Ideally, new diagnostic methods for SSc should be simple, fast, accurate, and cost-effective. Comprehensive two-dimensional gas chromatography (GC×GC) has a great potential for exhaled breath analysis. The increased peak capacity and sensitivity of GC×GC, provided by the combination of two capillary columns of different stationary phases by means of a modulator, enable the chromatographic separation and detection of thousands of compounds from a complex matrix[4]. For this reason, we carried out an exploratory study on SSc[5]. Basically, breath samples were collected in 5L Tedlar® bags. Volatiles contained in the sampling bag were then transferred onto Tenax®GR/Carbopack™B thermal desorption tubes (Markes International Ltd., Llantrisant, UK) and finally released and separated into a Pegasus GC-HRT 4D (LECO Corporation, St Joseph, MI, USA) through a mid-polar Rxi-624SilMS (30 m × 0.25 mm × 1.4 μm) as first column (dimension) and a polar Stabilwax (2 m × 0.25 mm ×0.5μm) as second dimension. The exhaled breath of 32 patients and 30 healthy subjects was therefore analyzed. The high resolving power of this approach and the use of statistical models enabled the identification of 16 compounds discriminating SSC patients from healthy ones[5]. However, further investigations had to be held to reach a better disease classification. In fact, the biomarkers highlighted here could be related to the scarring (fibrosis) of the lungs making these non-specific to SSCs. The second phase of the study aims to go deeper in patient stratification. Three groups were investigated: 50 SSC patients, 50 SSC-fibrosis patients and 50 ILD ones. The samples were collected at Maastricht medical center and CHU of Liège. All samples were then analyzed in the OBiACHem lab. Currently, a classification model is under construction to stratify patients based on their fibrosis status. [1] E. Zanatta, V. Codullo, J. Avouac, Y.A.-J. bone spine, undefined 2020, Elsevier (2019). [2] O. Bonhomme, B. André, F. Gester, … D. de S.-, undefined 2019, Academic.Oup.Com (n.d.). [3] J. Guiot, M. Henket, B. Andre, M. Herzog, N. Hardat, M.S. Njock, C. Moermans, M. Malaise, R. Louis, Clin. Epigenetics 12 (2020). [4] D. Zanella, J. Focant, F.A. Franchina, Anal. Sci. Adv. 2 (2021) 213–224. [5] D. Zanella, J. Guiot, P.-H. Stefanuto, L. Giltay, M. Henket, F. Guissard, B. André, M. Malaise, J. Potjewijd, F. Schleich, R. Louis, J.-F. Focant, Anal. Bioanal. Chem. 2021 41314 413 (2021) 3813–3822

Modeling spinal muscular atrophy in Drosophila links Smn to FGF signaling

FGF signaling in neurons is regulated by Survival Motor Neuron, a component of a complex that regulates snRNP biogenesis and FGF receptor expression

Modeling Spinal Muscular Atrophy in Drosophila

Spinal Muscular Atrophy (SMA), a recessive hereditary neurodegenerative disease in humans, has been linked to mutations in the survival motor neuron (SMN) gene. SMA patients display early onset lethality coupled with motor neuron loss and skeletal muscle atrophy. We used Drosophila, which encodes a single SMN ortholog, survival motor neuron (Smn), to model SMA, since reduction of Smn function leads to defects that mimic the SMA pathology in humans. Here we show that a normal neuromuscular junction (NMJ) structure depends on SMN expression and that SMN concentrates in the post-synaptic NMJ regions. We conducted a screen for genetic modifiers of an Smn phenotype using the Exelixis collection of transposon-induced mutations, which affects approximately 50% of the Drosophila genome. This screen resulted in the recovery of 27 modifiers, thereby expanding the genetic circuitry of Smn to include several genes not previously known to be associated with this locus. Among the identified modifiers was wishful thinking (wit), a type II BMP receptor, which was shown to alter the Smn NMJ phenotype. Further characterization of two additional members of the BMP signaling pathway, Mothers against dpp (Mad) and Daughters against dpp (Dad), also modify the Smn NMJ phenotype. The NMJ defects caused by loss of Smn function can be ameliorated by increasing BMP signals, suggesting that increased BMP activity in SMA patients may help to alleviate symptoms of the disease. These results confirm that our genetic approach is likely to identify bona fide modulators of SMN activity, especially regarding its role at the neuromuscular junction, and as a consequence, may identify putative SMA therapeutic targets

THE BEER’S VOLATOLOME, A COMPARATIVE STUDY BY COMPREHENSIVE TWO-DIMENSIONAL GAS CHROMATOGRAPHY TIME-OF-FLIGHT MASS SPECTROMETRY

Fermented beverages, such as beer, are considered as complex mixtures by flavor and food chemists. The beer volatolome contains hundreds of compounds, which affect the taste and the mouthfeel. In this highly competitive market, it is crucial for brewers to master each step of the process to understand and control the taste and to guarantee aroma quality and stability. Aroma compounds are essentially derived from the raw ingredients and the conditions used to produce beers. This study aims to determine the impact of the yeast used for bottle fermentation on beer’s volatolome. We brewed 5 different beers with the same feedstock. During the bottling, we used 5 distinct yeasts generally used by brewers. After an aging period of 8 weeks, all samples were analyzed using solid-phase microextraction couple with comprehensive two-dimensional gas chromatography coupled to mass spectrometry (SPME-GC×GC-TOFMS). A dedicated data processing workflow was then optimized. Principal component analysis (PCA) was used to visualize the clustering between samples. After data curation, peaks area of VOCs founds in each beer were used to carry out unsupervised statistical analyses. Three distinct groups were observed on PCA; two beers are considered very different regarding the VOCs while the remaining three constitute another cluster. Based on these results, the aroma profile of the different beers could be dressed. While the two first beers were dominated by specific ethyl esters giving fruity aromas, less associated fruity fragrance esters were expressed in the other group