Development of an Untargeted Method for the Analysis of the Volatile Profile of Onions via Solid-Phase Microextraction Arrow-Headspace-Gas Chromatography-Mass Spectrometry Using Design of Experiments.

The distinctive aroma of onions, consisting primarily of sulfur-containing compounds, is one of the reasons for the popularity of the vegetable. The rapid enzymatic and chemical reactions that occur after the destruction of onion bulb tissue render the analysis of the volatile profile challenging. Therefore, sample preparation is a crucial step in the analysis of the onion volatilome, but it often does not receive the necessary attention in the literature. In this work, we focused on two aspects: Firstly, we compared different sample preparation approaches to maximize the volatile yield and to enable the description of the onion volatile profile as it would emerge after a solid-phase microextraction (SPME) Arrow sampling procedure. For headspace (HS)-gas chromatography-mass spectrometry analysis, onion juice with the addition of an ammonium sulfate solution proved to be the best option. Secondly, we optimized the HS sampling and desorption parameters (enrichment time, enrichment temperature, agitator speed, desorption time) for onion volatiles using the efficient design of experiments (DoE) approach. The optimal conditions for the analysis with HS-SPME Arrow were an enrichment time of 75 min at 60°C with an agitator speed of 713 rpm and a desorption time of 120 s.

Alterations of meat quality, lipid composition and flavor in breast meat of laying hens with fatty liver hemorrhagic syndrome.

Fatty liver hemorrhagic syndrome (FLHS) has a high incidence rate in laying hens, and lots of FLHS-affected meat products enter the market every year. At the same time, the meat of laying hens is an important component of the human diet. However, the impact of FLHS on meat of laying hens is unknown, which could have a negative impact on consumers. To explore the effects of FLHS on chicken breast meat, a total of 36 twenty-five-wk-old Jingfen laying hens were used in the experiment. The hens were randomly divided into Control group and Model group, with 6 replicates per group and 3 hens per replicate. All chickens were raised in double-story step cages with individual pens. After a 3-wk acclimation period, the formal experiment began at 28 wk of age and lasted for 8 wk. Blood, liver, and breast meat samples were collected for the study of FLHS effects on breast meat. The impact of FLHS on meat quality was assessed by measuring indicators such as water-holding capacity and tenderness of the breast meat. Absolute quantitative lipidomics was employed to reveal the impact of FLHS on the lipid composition of chicken breast meat, and then validated by using RT-qPCR. Moreover, the volatilomics was utilized to detect changes in volatile organic compounds (VOCs) in chicken breast meat and to elucidate the resulting flavor changes. This research results showed that the meat quality of chicken breast meat decreased under FLHS, mainly manifested as reduced water holding capacity and decreased tenderness. The lipid content in the breast meat of FLHS-affected hens was significantly increased (P < 0.05). Among the affected lipids, 38 triglycerides exhibited notable elevation, possibly linked to heightened gene expression, such as lysophosphatidylcholine acyltransferase 3. The breast meat of laying hens with FLHS demonstrated an increased presence of VOCs, with 20 differential VOCs identified. Notably, 14 VOCs, particularly in 2-Undecenal, trans-Geranylacetone and ethyl nonanoate, exhibited substantial increases. These 3 VOCs had been identified as playing an important role in the formation of flavor in the breast meat of FLHS-affected laying hens. Correlation analysis suggested that the increase in these 3 VOCs might be related to the increase in lipid molecules such as phosphatidylethanolamine (38;3e) and acyl carnitine (10:3). In summary, FLHS reduced the breast meat quality of laying hens, altered its lipid profiles, and enhanced its flavor. These findings underscore the profound impact of FLHS on lipid and VOC profiles in chicken breast meat, offering valuable insights for chicken meat quality affected by FLHS.

SPME arrow-based extraction for enhanced targeted and untargeted urinary volatilomics.

BACKGROUND: Volatile organic compounds (VOCs) present in human urine are promising biomarkers for various health conditions and environmental exposures. However, their reliable detection is challenging due to the complexity of urinary matrices and the low concentrations of VOCs. Moreover, untargeted approaches present considerable challenges in terms of data interpretation, increasing the complexity of method development. Here we address these challenges by developing a new method that combines solid-phase microextraction (SPME) Arrow with gas chromatography-high resolution mass spectrometry (GC-HRMS), using a design of experiments (DOE) approach for targeted and untargeted compounds. This methodology, specifically tailored for SPME Arrow, represents a significant advancement in untargeted urinary analysis. RESULTS: The method was developed based on targeted and untargeted outcomes, were ranking results focus on the highest response area of 11 spiked target VOCs representative of urinary volatilomics, and on identifying the maximum untargeted number of VOCs. The method was developed focusing on the highest response area of 11 spiked target VOCs representative of urinary volatilomics and identifying the maximum number of VOCs. A univariate method determined the optimal coating type, urine volume, and salt addition. Subsequently, a central composite design (CCD) DOE was used to determine ideal temperature, extraction, and incubation times. The best method obtained has an extraction time of 60 min at a temperature of 53 °C, with an SPME Arrow CAR/PDMS using 2 mL of urine, with 0.25 % w/v of NaCl and a pH of 2. Compared to conventional SPME fibers, the SPME Arrow showed improved extraction efficiency, detecting more VOCs. Finally, the enhanced method was successfully applied to urine samples from children exposed and non-exposed to tobacco smoke, identifying specific VOCs, like p-cymene and p-isopropenyl toluene related to tobacco exposure. SIGNIFICANCE: By integrating both targeted and untargeted approaches, the developed method comprehensively captures the complexity of urinary metabolomics. This dual strategy ensures the precise identification of known compounds and the discovery of novel biomarkers, thereby providing a more complete metabolic profile. Such an approach is crucial for advancing in non-invasive diagnostics and environmental health studies, as it offers deeper insights into the intricate relationships between metabolic processes and various health conditions.

Non-Invasive Diagnostic Approaches for Kidney Disease: The Role of Electronic Nose Systems.

Kidney diseases are a group of conditions related to the functioning of kidneys, which are in turn unable to properly filter waste and excessive fluids from the blood, resulting in the presence of dangerous levels of electrolytes, fluids, and waste substances in the human body, possibly leading to significant health effects. At the same time, the toxins amassing in the organism can lead to significant changes in breath composition, resulting in halitosis with peculiar features like the popular ammonia breath. Starting from this evidence, scientists have started to work on systems that can detect the presence of kidney diseases using a minimally invasive approach, minimizing the burden to the individuals, albeit providing clinicians with useful information about the disease's presence or its main related features. The electronic nose (e-nose) is one of such tools, and its applications in this specific domain represent the core of the present review, performed on articles published in the last 20 years on humans to stay updated with the latest technological advancements, and conducted under the PRISMA guidelines. This review focuses not only on the chemical and physical principles of detection of such compounds (mainly ammonia), but also on the most popular data processing approaches adopted by the research community (mainly those relying on Machine Learning), to draw exhaustive conclusions about the state of the art and to figure out possible cues for future developments in the field.

Widely targeted volatilomics and transcriptome analyses reveal the differences in volatile organic components in differently shaped Amomum tsao-ko fruits.

BACKGROUND: Amomum tsao-ko is an important aromatic crop used in medicines and food. It can be categorized into three main types based on the fruit shape: long (L), oval (O), and round (R). However, limited information is available on the volatile substances present in differently shaped A. tsao-ko fruits. This study investigated the characteristics and biosynthesis of volatile organic compounds (VOCs) in fresh and dried A. tsao-ko fruits of different shapes using widely targeted volatilomics and transcriptome analyses. RESULTS: In total, 978 VOCs, primarily terpenoids, esters, and heterocyclic compounds, were detected. The number of differentially accumulated volatile organic compounds (DAVOCs) in dried fruits of various shapes was significantly higher than that in fresh fruits, with terpenoids, esters, and heterocyclic compounds accounting for approximately 50% of the total DAVOCs. Notably, α-phellandrene, identified as a shared differential accumulated terpenoid across various fruit shapes, was detected in both fresh and dried fruits. Through transcriptome analysis, 40 candidate genes implicated in the terpenoid biosynthesis pathway were screened. An integrated analysis of the metabolome and transcriptome revealed that the structural genes HMGR-2, TPS7, TPS5-10, TPS21-3, TPS21-5, TPS21-6, TPS21-7, and TPS21-9, along with 81 transcription factors (including 17 NACs, 16 MYBs, 16 AP2/ERFs, 13 WRKYs, 13 bHLHs, and 6 bZIPs), co-regulate the biosynthesis of volatile terpenoids. CONCLUSIONS: This study expands our understanding of the volatile metabolism profile of A. tsao-ko and provides a solid foundation for future investigations of the mechanisms governing fruit quality.

Effect of alterations in phospholipids and free fatty acids on aroma-active compounds in instant-boiled chuck tender, sirloin and silverside beef.

Beef flavor profiles are strongly influenced by cooking methods and lipid composition. However, the effect of instant-boiling on the aroma of different beef slices was unclear. This study investigated the lipid profiles and instant-boiling volatile profiles of chuck tender (M. Supraspinatus), sirloin (M. Longissimus dorsi) and silverside (M. Biceps femoris). Quantitative lipidomics identified 336 lipid molecular species, of which 84-112 were quantitatively different among the three beef slices. Sirloin had lower phosphatidylcholine, phosphatidylinositol, phosphatidylglycerol and free fatty acids than chuck tender and silverside. The unsaturated fatty acid acyl chains in phosphatidylethanolamine differed significantly. Solid phase microextraction-gas chromatography-olfactometry-mass spectrometry (SPME-GC-O-MS) identified hexanal, octanal, nonanal, decanal, (E)-2-octenal, (E)-2-nonenal, (E)-2-undecenal, (E,E)-2,4-nonadienal, (E,E)-2,4-decadienal, 1-octen-3-ol, 2-pentylfuran and acetoin as the aroma-active compounds of instant-boiled beef. Unsaturated free fatty acids and phosphatidylglycerols with unsaturated fatty acid residues positively correlated with the aroma-active compounds and might be crucial in flavor differences among the three beef slices. These findings provide greater understanding of the lipid and instant-boiling aroma-active compound profiles in chuck tender, sirloin and silverside, and reflect the suitability of different beef slices for instant-boiling from the aroma perspective.

Flavoromic analysis of wines using gas chromatography, mass spectrometry and sensory techniques.

Various sensory perceptions drive the quality and typicality of wines, with the volatile profile playing a fundamental role in the characteristics of odor, aroma and consequently flavor, which combines the smell (odor and aroma), taste, and trigeminal sensations. Efforts have been made in both the field of instrumental and sensory analysis to understand the relationship of volatile compounds with sensory attributes in omics approaches. Gas chromatography (monodimensional and two-dimensional (heartcutting and comprehensive)) associated with mass spectrometry (GC/MS, GC-GC/MS and GCxGC/MS) and chemometric tools have contributed to foodomics analyses, specifically those linked to metabolomics/volatilomics. These tools, along with the elucidation of sensory properties (sensomics), lead to advanced results in the field of flavoromics. They also help to define the best practices in both vineyard management and winemaking that enable the production of high-quality wines. The objective of this review is to report the challenges of determining the volatile profile of wines, pointing out the ways that can be followed in successful identification and quantification of volatile compounds. The state of the art of sensory evaluation methods is also addressed, providing information that helps in choosing the most appropriate sensory method to be conducted with chromatographic analysis to achieve more in-depth results in the field of flavoromics.

Infestation of Rice Striped Stem Borer (Chilo suppressalis) Larvae Induces Emission of Volatile Organic Compounds in Rice and Repels Female Adult Oviposition.

Plants regulate the biosynthesis and emission of metabolic compounds to manage herbivorous stresses. In this study, as a destructive pest, the pre-infestation of rice striped stem borer (SSB, Chilo suppressalis) larvae on rice (Oryza sativa) reduced the subsequent SSB female adult oviposition preference. Widely targeted volatilomics and transcriptome sequencing were used to identify released volatile metabolic profiles and differentially expressed genes in SSB-infested and uninfested rice plants. SSB infestation significantly altered the accumulation of 71 volatile organic compounds (VOCs), including 13 terpenoids. A total of 7897 significantly differentially expressed genes were identified, and genes involved in the terpenoid and phenylpropanoid metabolic pathways were highly enriched. Correlation analysis revealed that DEGs in terpenoid metabolism-related pathways were likely involved in the regulation of VOC biosynthesis in SSB-infested rice plants. Furthermore, two terpenoids, (-)-carvone and cedrol, were selected to analyse the behaviour of SSB and predators. Y-tube olfactometer tests demonstrated that both (-)-carvone and cedrol could repel SSB adults at higher concentrations; (-)-carvone could simultaneously attract the natural enemies of SSB, Cotesia chilonis and Trichogramma japonicum, and cedrol could only attract T. japonicum at lower concentrations. These findings provide a better understanding of the response of rice plants to SSB and contribute to the development of new strategies to control herbivorous pests.

A lipidomic and volatilomic approach to map the lipid profile and related volatile compounds in roasted quail meat using circulating non-fried roast technology.

Lipids play a significant role in aroma formation. However, lipid variations and their impact on aroma during the processing of quail meat remain unknown. Therefore, a comprehensive analysis of lipids and aroma compounds was conducted in circulating non-fried roasted quail meat. Nineteen odorants were identified as key aroma compounds in the roasted quail meat at 40 min with OAVs of >1. The concentrations of most key odorants significantly increased in circulating non-fried roasted (CNR) quail meat within the first 30 min of roasting, reaching maximum values at 40 min. Phospholipids, neutral lipids, and sphingolipids emerged as potential markers for distinguishing different samples. Neutral lipids had the highest peak areas and significantly contributed to the aroma retention. Phospholipids and neutral lipids with unsaturated fatty acids, particularly C18 acyl groups, played a crucial role in aroma formation. This study provides valuable insights into the role of lipids in determining aroma quality.

Innovative Approach for Human Semen Quality Assessment Based on Volatilomics.

The volatilome profile of some biofluids (blood, urine, and human semen) identified by Solid-Phase Microextraction-Gas Chromatography/Mass Spectrometry (SPME-GC/MS) and collected from young men living in two high-pollution areas in Italy, i.e., Land of Fires and Valley of Sacco River, have been coupled to sperm parameters obtained by spermiogram analysis to build general multiple regression models. Panels of volatile organic compounds (VOCs) have been selected to optimize the models and used as predictive variables to estimate the different sperm quality parameters (sperm cell concentration, total and progressive motility/immotile cells, total/head/neck/tail morphology anomalies, semen round cell concentration). The results of the multiple linear regression models based on the different subgroups of data joining VOCs from one/two or three biofluids have been compared. Surprisingly, the models based on blood and urine VOCs have allowed an excellent estimate of spermiogram values, paving the way towards a new method of indirect evaluation of semen quality and preventive screening. The significance of VOCs in terms of toxicity and dangerousness was discussed with the support of chemical databases available online.

Combining Targeted Metabolomics with Untargeted Volatilomics for Unraveling the Impact of Sprouting on the Volatiles and Aroma of False Flax (Camelina sativa) Cold-Pressed Oil.

Sprouting of stored oilseeds due to improper storage can lead to quality defects of cold-pressed oils obtained from them. This study aimed to evaluate the effect of seed sprouting on volatile organic compounds (VOCs), aroma-active compounds, and the content of nonvolatile metabolites in cold-pressed false flax oil obtained from sprouted seeds. In this study, 88 unique VOCs were detected in sprouted oils, whereas only 42 were found in the control oils. The control oils were characterized by a higher abundance of alcohols, while all other groups of compounds were associated with sprouted seeds. The formation of many VOCs was reflected in changes in the nonvolatile precursors. Fifteen aroma-active compounds were identified in sprouted oil, with five compounds playing a significant role (FD ≥ 128) in aroma formation. The presented approach allowed identification of differences caused by seed sprouting, resulting in oils with a much stronger aroma and a richer profile of VOCs due to intensive metabolic changes. The origin of many VOCs can be explained by alterations in the content of nonvolatile metabolites.

Comprehensive insights into the organoleptic characteristics attributes of "HuangYuXiang": Integration of volatilomics, sensomics, macrogenomics, lipidomics, and metabolomics.

HuangYuXiang (HYX) is a colorful and flavorful traditional cuisine in China, which development of organoleptic attributes is a complex process. Flavor sensory attributes was explored through volatilomics, sensomics, macrogenomics, lipidomics, and metabolomics in seven HYXs. Group B demonstrated the highest sensory scores. A total of 41 volatiles were detected, of which 7 were identified as key volatiles. Caulobacteraceae sp., Psychrobacter faecalis, Ralstonia pickettii, Carnobacterium divergens, and Psychrobacter cibarius were representative bacteria in HYXs. A total of 679 lipids (251 differential lipids) and 329 (113 differential metabolites) metabolites were identified. The differential compounds were the main contributors to flavor differences. L-homocitrulline, arg-ser, 4-aminobenzoic acid, arg-gly, sucrose, pyridoxine, D-cyclohexylglycine, PC 21:4/22:6, PC O-15:0/22:5, PC O-20:2/20:5, and FA 18:2 were heavily accumulated under the microbial action, which in turn promoted the formation of aroma and taste substances. The results of this study provide a theoretical basis for the standardized processing of high-quality HYX.

Progress and challenges of developing volatile metabolites from exhaled breath as a biomarker platform.

BACKGROUND: The multitude of metabolites generated by physiological processes in the body can serve as valuable biomarkers for many clinical purposes. They can provide a window into relevant metabolic pathways for health and disease, as well as be candidate therapeutic targets. A subset of these metabolites generated in the human body are volatile, known as volatile organic compounds (VOCs), which can be detected in exhaled breath. These can diffuse from their point of origin throughout the body into the bloodstream and exchange into the air in the lungs. For this reason, breath VOC analysis has become a focus of biomedical research hoping to translate new useful biomarkers by taking advantage of the non-invasive nature of breath sampling, as well as the rapid rate of collection over short periods of time that can occur. Despite the promise of breath analysis as an additional platform for metabolomic analysis, no VOC breath biomarkers have successfully been implemented into a clinical setting as of the time of this review. AIM OF REVIEW: This review aims to summarize the progress made to address the major methodological challenges, including standardization, that have historically limited the translation of breath VOC biomarkers into the clinic. We highlight what steps can be taken to improve these issues within new and ongoing breath research to promote the successful development of the VOCs in breath as a robust source of candidate biomarkers. We also highlight key recent papers across select fields, critically reviewing the progress made in the past few years to advance breath research. KEY SCIENTIFIC CONCEPTS OF REVIEW: VOCs are a set of metabolites that can be sampled in exhaled breath to act as advantageous biomarkers in a variety of clinical contexts.

Paired metabolomics and volatilomics provides insight into transient high light stress response mechanisms of the coral Montipora mollis.

The coral holobiont is underpinned by complex metabolic exchanges between different symbiotic partners, which are impacted by environmental stressors. The chemical diversity of the compounds produced by the holobiont is high and includes primary and secondary metabolites, as well as volatiles. However, metabolites and volatiles have only been characterised in isolation so far. Here, we applied a paired metabolomic-volatilomic approach to characterise holistically the chemical response of the holobiont under stress. Montipora mollis fragments were subjected to high-light stress (8-fold higher than the controls) for 30 min. Photosystem II (PSII) photochemical efficiency values were 7-fold higher in control versus treatment corals immediately following high-light exposure, but returned to pre-stress levels after 30 min of recovery. Under high-light stress, we identified an increase in carbohydrates (> 5-fold increase in arabinose and fructose) and saturated fatty acids (7-fold increase in myristic and oleic acid), together with a decrease in fatty acid derivatives in both metabolites and volatiles (e.g., 80% decrease in oleamide and nonanal), and other antioxidants (~ 85% decrease in sorbitol and galactitol). These changes suggest short-term light stress induces oxidative stress. Correlation analysis between volatiles and metabolites identified positive links between sorbitol, galactitol, six other metabolites and 11 volatiles, with four of these compounds previously identified as antioxidants. This suggests that these 19 compounds may be related and share similar functions. Taken together, our findings demonstrate how paired metabolomics-volatilomics may illuminate broader metabolic shifts occurring under stress and identify linkages between uncharacterised compounds to putatively determine their functions.

Unravelling the importance of seed roasting for oil quality by the non-targeted volatilomics and targeted metabolomics of cold-pressed false flax (Camelina sativa L.) oil and press cakes.

False flax (Camelina sativa L.), known as camelina, is an ancient oil plant that has gathered renewed interest. In this study, a comprehensive analysis encompassing nontargeted volatilomics and targeted, quantitative metabolomics performed for cold-pressed oil and press cake and was integrated with sensory analysis of cold-pressed camelina oil and the effect of seed roasting was evaluated. Roasting in general resulted in the formation of 22 new volatile organic compounds (VOCs) in oil, while roasting at 140 and 180 °C resulted in the formation of 12 and 124 unique VOCs, respectively. Roasting notably influenced the profile of primary and secondary metabolites in both oil and press cakes, as well as volatilome and aroma of cold-pressed camelina oil. Many VOCs can be attributed to thermal degradation of primary and secondary metabolites. Roasting intensified the flavour of cold-pressed camelina oil, enhancing the perception of notes formed through the Maillard reaction.

Comprehensive insights into the mechanism of flavor formation driven via inoculation with mixed starter cultures in dry-fermented tilapia sausages: Integration of macrogenomics, volatilomics, and lipidomics.

To investigate the mechanisms of flavor formation in dry-fermented tilapia sausages, the volatiles, bacterial community, and lipid composition during fermentation were analyzed using gas chromatography-ion mobility spectrometry, 16S high throughput sequencing, and ultra-performance liquid chromatography-mass spectrometer. Pediococcus pentosaceus, Staphylococcus xylosus, and Staphylococcus carnosus became dominant bacteria during the fermentation. A total of 66 volatiles and 293 lipids (48 differential lipids) were identified. PC and PE content decreased. Aldehyde and 1-octen-3-ol content decreased. Most esters and ketones content increased during fermentation. Six metabolic pathways associated with differential lipids were identified by enrichment analysis. Glycerophospholipid metabolism was the main metabolic pathway. Correlation analysis revealed that PC and PE were precursors for volatiles, including PC 16:0/18:2 and PE 18:0/22:6. The dominant bacteria facilitate the hydrolysis of PC and PE, leading to the formation of esters and ketones. This study provides a theoretical basis for the targeted regulation of fermented sausage flavors.

Volatilomics of Fruit Wines.

Volatilomics is a scientific field concerned with the evaluation of volatile compounds in the food matrix and methods for their identification. This review discusses the main groups of compounds that shape the aroma of wines, their origin, precursors, and selected metabolic pathways. The paper classifies fruit wines into several categories, including ciders and apple wines, cherry wines, plum wines, berry wines, citrus wines, and exotic wines. The following article discusses the characteristics of volatiles that shape the aroma of each group of wine and the concentrations at which they occur. It also discusses how the strain and species of yeast and lactic acid bacteria can influence the aroma of fruit wines. The article also covers techniques for evaluating the volatile compound profile of fruit wines, including modern analytical techniques.

Volatilomics as a tool to ascertain food adulteration, authenticity, and origin.

Over recent years, there has been an increase in the number of reported cases of food fraud incidents, whereas at the same time, consumers demand authentic products of high quality. The emerging volatilomics technology could be the key to the analysis and characterization of the quality of different foodstuffs. This field of omics has aroused the interest of scientists due to its noninvasive, rapid, and cost-profitable nature. This review aims to monitor the available scientific information on the use of volatilomics technology, correlate it to the relevant food categories, and demonstrate its importance in the food adulteration, authenticity, and origin areas. A comprehensive literature search was performed using various scientific search engines and "volatilomics," "volatiles," "food authenticity," "adulteration," "origin," "fingerprint," "chemometrics," and variations thereof as keywords, without chronological restriction. One hundred thirty-seven relevant publications were retrieved, covering 11 different food categories (meat and meat products, fruits and fruit products, honey, coffee, tea, herbal products, olive oil, dairy products, spices, cereals, and others), the majority of which focused on the food geographical origin. The findings show that volatilomics typically involves various methods responsible for the extraction and consequential identification of volatile compounds, whereas, with the aid of data analysis, it can handle large amounts of data, enabling the origin classification of samples or even the detection of adulteration practices. Nonetheless, a greater number of specific research studies are needed to unlock the full potential of volatilomics.

Dynamic microbial and metabolic changes during Apulian Caciocavallo cheesemaking and ripening produced according to a standardized protocol.

The microbiota of a cheese play a critical role in influencing its sensory and physicochemical properties. In this study, traditional Apulian Caciocavallo cheeses coming from 4 different dairies in the same area and produced following standardized procedures were examined, as well as the different bulk milks and natural whey starter (NWS) cultures used. Moreover, considering the cheese wheels as the blocks of Caciocavallo cheeses as whole, these were characterized at different layers (i.e., core, under-rind, and rind) of the block using a multi-omics approach. In addition to physical-chemical characterization, culturomics, quantitative PCR, metagenomics, and metabolomics analysis were carried out after salting and throughout the ripening time (2 mo) to investigate major shifts in the succession of the microbiota and flavor development. Culture-dependent and 16S rRNA metataxonomics results clearly clustered samples based on microbiota biodiversity related to the production dairy plant as a result of the use of different NWS or the intrinsic conditions of each production site. At the beginning of the ripening, cheeses were dominated by Lactobacillus, and in 2 dairies (Art and SdC), Streptococcus genera were associated with the NWS. The analysis allowed us to show that although the diversity of identified genera did not change significantly between the rind, under-rind, and core fractions of the same samples, there was an evolution in the relative abundance and absolute quantification, modifying and differentiating profiles during ripening. The real-time PCR, also known as quantitative or qPCR, mainly differentiated the temporal adaptation of those species originating from bulk milks and those provided by NWS. The primary starters detected in NWS and cheeses contributed to the high relative concentration of 1-butanol, 2-butanol, 2-heptanol, 2-butanone, acetoin, delta-dodecalactone, hexanoic acid ethyl ester, octanoic acid ethyl ester, and volatile free fatty acids during ripening, whereas cheeses displaying low abundances of Streptococcus and Lactococcus (dairy Del) had a lower total concentration of acetoin compared with Art and SdC. However, the subdominant strains and nonstarter lactic acid bacteria present in cheeses are responsible for the production of secondary metabolites belonging to the chemical classes of ketones, alcohols, and organic acids, reaffirming the importance and relevance of autochthonous strains of each dairy plant although only considering a delimited production area.

WTV2.0: A high-coverage plant volatilomics method with a comprehensive selective ion monitoring acquisition mode.

Volatilomics is essential for understanding the biological functions and fragrance contributions of plant volatiles. However, the annotation coverage achieved using current untargeted and widely targeted volatomics (WTV) methods has been limited by low sensitivity and/or low acquisition coverage. Here, we introduce WTV 2.0, which enabled the construction of a high-coverage library containing 2111 plant volatiles, and report the development of a comprehensive selective ion monitoring (cSIM) acquisition method, including the selection of characteristic qualitative ions with the minimal ion number for each compound and an optimized segmentation method, that can acquire the smallest but sufficient number of ions for most plant volatiles, as well as the automatic qualitative and semi-quantitative analysis of cSIM data. Importantly, the library and acquisition method we developed can be self-expanded by incorporating compounds not present in the library, utilizing the obtained cSIM data. We showed that WTV 2.0 increases the median signal-to-noise ratio by 7.6-fold compared with the untargeted method, doubled the annotation coverage compared with the untargeted and WTV 1.0 methods in tomato fruit, and led to the discovery of menthofuran as a novel flavor compound in passion fruit. WTV 2.0 is a Python library with a user-friendly interface and is applicable to profiling of volatiles and primary metabolites in any species.