Identification and quantification of flavor compounds in smoked tuna fish based on GC-Orbitrap volatolomics approach.

Cold smoking enhances the appeal of fish products, offering consumers a smooth texture and a delicate smoky flavor. This study aims to explore variations in the volatile profile from different exposure times during cold smoking processing (light, moderate, and full-cure) in tune samples. An innovative untargeted analytical approach, headspace solid-phase microextraction combined with gas chromatography and a hybrid quadrupole-orbitrap mass analyzer, was employed to identify 86 volatiles associated with the cold smoking process. Most of these compounds, including phenols, furan derivates, aldehydes, cyclic ketones, and different aromatic species, were found to contribute to the smoke odor. The development of a QuEChERS-based extraction and clean-up method facilitated the quantification of 25 relevant smoky markers across all smoking degrees, revealing significant concentration differences after 15 h of smoking. This research sheds light on the dynamics of cold smoking impact and its on the flavor profile and safety quality of processed fish products.

LC-IMS-HRMS for identification of biomarkers in untargeted metabolomics: The effects of pterostilbene and resveratrol consumption in liver steatosis, animal model.

Untargeted metabolomics with the combination of ion mobility separation coupled to high resolution mass spectrometry (IMS-HRMS) was applied to investigate the impact of resveratrol and pterostilbene supplementation on the metabolic fingerprint of the Wistar rats liver with induced liver steatosis. RP-LC and HILIC in both ionisation modes were employed to analyse the liver samples (n = 40) from Wistar rats fed with a high-fat and high-fructose diet, supplemented or not with resveratrol and pterostilbene. After univariate and multivariate statistical analysis, 34 metabolites were highlighted in the different diets and elucidated. Despite the structural similarity, different alterations in liver metabolism were observed by the supplementations. Resveratrol treatment was characterised by the alteration in metabolism of 17 lysophospholipids, while pterostilbene affected some vitamins and derivatives, among others. IMS has demonstrated great potential in the elucidation process thanks to the additional structural descriptor the CCS (Å2), providing more confidence in the identification.

Untargeted metabolomics approach using UHPLC-IMS-QTOF MS for surface body samples to identify low-volatility chemosignals related to maternal care in mice.

The present study is focused on the determination of low-volatile chemosignals excreted or secreted by mouse pups in their early days of life involved in maternal care induction in mice adult females. Untargeted metabolomics was employed to differentiate between samples collected with swabs from facial and anogenital area from neonatal mouse pups receiving maternal care (first two weeks of life) and the elder mouse pups in the weaning period (4th week old). The sample extracts were analysed by ultra-high pressure liquid chromatography (UHPLC) coupled to ion mobility separation (IMS) in combination with high resolution mass spectrometry (HRMS). After data processing with Progenesis QI and multivariate statistical analysis, five markers present in the first two weeks of mouse pups life and putatively involved in materno-filial chemical communication were tentatively identified: arginine, urocanic acid, erythro-sphingosine (d17:1), sphingosine (d18:1) and sphinganine. The four-dimensional data and the tools associated to the additional structural descriptor obtained by IMS separation were of great help in the compound identification. The results demonstrated the great potential of UHPLC-IMS-HRMS based untargeted metabolomics to identity putative pheromones in mammals.

Discovery of Food Intake Biomarkers Using Metabolomics.

Due to the high impact of diet exposure on health, it is crucial the generation of robust data of regular dietary intake, hence improving the accuracy of dietary assessment. The metabolites derived from individual food or group of food have great potential to become biomarkers of food intake (BFIs) and provide more objective food consumption measurements.Herein, it is presented an untargeted metabolomic workflow for the discovery BFIs in blood and urine samples, from the study design to the biomarker identification. Samples are analyzed by liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS). A wide variety of compounds are covered by separate analyses of medium to nonpolar molecules and polar metabolites based on two LC separations as well as both positive and negative electrospray ionization. The main steps of data treatment of the comprehensive data sets and statistical analysis are described, as well as the principal considerations for the BFI identification.

Benefits of Ion Mobility Separation in GC-APCI-HRMS Screening: From the Construction of a CCS Library to the Application to Real-World Samples.

The performance of gas chromatography (GC) combined with the improved identification properties of ion mobility separation coupled to high-resolution mass spectrometry (IMS-HRMS) is presented as a promising approach for the monitoring of (semi)volatile compounds in complex matrices. The soft ionization promoted by an atmospheric pressure chemical ionization (APCI) source designed for GC preserves the molecular and/or quasi-molecular ion information enabling a rapid, sensitive, and efficient wide-scope screening. Additionally, ion mobility separation (IMS) separates species of interest from coeluting matrix interferences and/or resolves isomers based on their charge, shape, and size, making IMS-derived collision cross section (CCS) a robust and matrix-independent parameter comparable between instruments. In this way, GC-APCI-IMS-HRMS becomes a powerful approach for both target and suspect screening due to the improvements in (tentative) identifications. In this work, mobility data for 264 relevant multiclass organic pollutants in environmental and food-safety fields were collected by coupling GC-APCI with IMS-HRMS, generating CCS information for molecular ion and/or protonated molecules and some in-source fragments. The identification power of GC-APCI-IMS-HRMS for the studied compounds was assessed in complex-matrix samples, including fish feed extracts, surface waters, and different fruit and vegetable samples.

Novel sampling strategy for alive animal volatolome extraction combined with GC-MS based untargeted metabolomics: Identifying mouse pup pheromones.

In this study, we identify 11 mouse pup volatiles putatively involved in maternal care induction in adult females. For this purpose, we have adapted the dynamic headspace methodology to extract the volatolome of whole alive animals. Untargeted metabolomic methodology was used to compare the volatolome of neonatal (4-6 days) with elder pups until the age of weaning (21-23 days old). Pup volatolome was analyzed by gas chromatography (GC) coupled to single quadrupole mass spectrometry (MS) using automated thermal desorption for sample introduction. After data processing and multivariate statistical analysis, comparison with NIST spectral library allowed identifying compounds secreted preferentially by neonatal pups: di(propylen glycol) methyl ether, 4-nonenal, di(ethylene glycol) monobutyl ether, 2-phenoxyethanol, isomethyl ionone, tridecanal, 1,3-diethylbenzene, 1,2,4,5-tetramethylbenzene, 2-ethyl-p-xylene and tri(propylene glycol) methyl ether. Palmitic acid was enriched in the volatolome of fourth week youngsters compared to neonatal pups. The results demonstrated the great potential of the new sampling procedure combined with GC-MS based untargeted volatolomics to identify volatile pheromones in mammals.

Gas chromatography-mass spectrometry based untargeted volatolomics for smoked seafood classification.

With the increase of the demand of low flavouring smoked seafood products, there is a need of methodologies able to distinguish between different seafood treatments, as not all of them are allowed in all markers. Following this objective, in the present work an untargeted volatolomics approach was applied to identify volatile markers that demonstrate that Cold smoked products can be distinguished from Tasteless smoke neither Carbon monoxide treated seafood, which are prohibited in the European Union. The use of dynamic headspace for the volatile extraction followed by thermal desorption in combination with Gas Chromatography (GC) coupled to single quadrupole Mass Spectrometry (MS) has been employed for the determination of volatile composition of smoked fish. Data processing consisted on the use of PARADISe software, applied for GC/MS data treatment, followed by the multivariate analysis with PLS_Toolbox (MATLAB), and finally the creation and validation of statistical classification model. All 107 variables obtained allowed the construction of a model reaching the correct classification of 97% of the blind samples, while a simplified model with only 11 variables correctly classified up to 93% of the blind samples. These 11 compounds were elucidated to develop subsequent target volatolomics approaches, if needed. Ordered according to the importance in the classification model, the elucidated compounds were: 3-methyl-cyclopentanone, ethylbenzene, 2-methyl-2-cyclopenten-1-one, 2-methyl-benzofuran, furfuryl alcohol, 2-acetylfuran, acetophenone, guaiacol, 1-hydroxy-2-butanone, 4-vinylguaicol and acetoin. The results demonstrated the great potential of untargeted volatolomics for smoked seafood treatments classification.

Ultra-Performance Liquid Chromatography-Ion Mobility Separation-Quadruple Time-of-Flight MS (UHPLC-IMS-QTOF MS) Metabolomics for Short-Term Biomarker Discovery of Orange Intake: A Randomized, Controlled Crossover Study.

A major problem with dietary assessments is their subjective nature. Untargeted metabolomics and new technologies can shed light on this issue and provide a more complete picture of dietary intake by measuring the profile of metabolites in biological samples. Oranges are one of the most consumed fruits in the world, and therefore one of the most studied for their properties. The aim of this work was the application of untargeted metabolomics approach with the novel combination of ion mobility separation coupled to high resolution mass spectrometry (IMS-HRMS) and study the advantages that this technique can bring to the area of dietary biomarker discovery, with the specific case of biomarkers associated with orange consumption (Citrus reticulata) in plasma samples taken during an acute intervention study (consisting of a randomized, controlled crossover trial in healthy individuals). A total of six markers of acute orange consumption, including betonicines and conjugated flavonoids, were identified with the experimental data and previous literature, demonstrating the advantages of ion mobility in the identification of dietary biomarkers and the benefits that an additional structural descriptor, as the collision cross section value (CCS), can provide in this area.

Untargeted metabolomics approach for unraveling robust biomarkers of nutritional status in fasted gilthead sea bream (Sparus aurata).

A metabolomic study has been performed to identify sensitive and robust biomarkers of malnutrition in farmed fish, using gilthead sea bream (Sparus aurata) as a model. The metabolomic fingerprinting of serum from fasted fish was assessed by means of ultra-high performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry. More than 15,000 different m/z ions were detected and Partial Least Squares-Discriminant analysis allowed a clear differentiation between the two experimental groups (fed and 10-day fasted fish) with more than 90% of total variance explained by the two first components. The most significant metabolites (up to 45) were elucidated on the basis of their tandem mass spectra with a broad representation of amino acids, oligopeptides, urea cycle metabolites, L-carnitine-related metabolites, glutathione-related metabolites, fatty acids, lysophosphatidic acids, phosphatidylcholines as well as biotin- and noradrenaline-related metabolites. This untargeted approach highlighted important adaptive responses in energy and oxidative metabolism, contributing to identify robust and nutritionally-regulated biomarkers of health and metabolic condition that will serve to assess the welfare status of farmed fish.