Wine Metabolites Research Articles

Comparison of volatile and non-volatile metabolites in rice wine fermented by Koji inoculated with Saccharomycopsis fibuligera and Aspergillus oryzae

This study investigated volatile and nonvolatile metabolite profiles of makgeolli (a traditional rice wine in Korea) fermented by koji inoculated with Saccharomycopsis fibuligera and/or Aspergillus oryzae. The enzyme activities in koji were also examined to determine their effects on the formation of metabolites. The contents of all 18 amino acids detected were the highest in makgeolli fermented by S. fibuligera CN2601-09, and increased after combining with A. oryzae CN1102-08, unlike the contents of most fatty acids. On the other hand, major volatile metabolites were fusel alcohols, acetate esters, and ethyl esters. The contents of most fusel alcohols and acetate esters were the highest in makgeolli fermented by S. fibuligera CN2601-09, for which the protease activity was the highest, leading to the largest amounts of amino acods. The makgeolli samples fermented only by koji inoculated with S. fibuligera could be discriminated on PCA plots from the makgeolli samples fermented in combination with A. oryzae. In the case of nonvolatile metabolites, all amino acids and some metabolites such as xylose, 2-methylbenzoic acid, and oxalic acid contributed mainly to the characteristics of makgeolli fermented by koji inoculated with S. fibuligera and A. oryzae. These results showed that the formations of volatile and nonvolatile metabolites in makgeolli can be significantly affected by microbial strains with different enzyme activities in koji. To our knowledge, this study is the first report on the effects of S. fibuligera strains on the formation of volatile and non-volatile metabolites in rice wine, facilitating their use in brewing rice wine.

The impact of SO2 on wine flavanols and indoles in relation to wine style and age

Wine has one of the broadest chemical profiles, and the common oenological practice of adding the antioxidant and antimicrobial sulfur dioxide has a major impact on its metabolomic fingerprint. In this study, we investigated novel discovered oenological reactions primarily occurring between wine metabolites and sulfur dioxide. The sulfonated derivatives of epicatechin, procyanidin B2, indole acetic acid, indole lactic acid and tryptophol were synthesized and for the first time quantified in wine. Analysis of 32 metabolites in 195 commercial wines (1986–2016 vintages) suggested that sulfonation of tryptophan metabolites characterised white wines, in contrast to red wines, where sulfonation of flavanols was preferred. The chemical profile of the oldest wines was strongly characterised by sulfonated flavanols and indoles, indicating that could be fundamental metabolites in explaining quality in both red and white aged wines. These findings offer new prospects for more precise use of sulfur dioxide in winemaking.

Effects of area, year and climatic factors on Barbera wine characteristics studied by the combination of 1H-NMR metabolomics and chemometrics

ABSTRACTBarbera is one of the most common red grape in Italy, found in almost every region of the country. The widespread market of the corresponding wine deserves studies on its traceability and authenticity. To this end, the 1H-NMR database of several wine metabolites was built using spectra recorded in a wine-like matrix containing ethanol and tartaric acid with pH precisely adjusted. The combination of 1H-NMR metabolomics study and chemometrics (multivariate data analysis) is able to discriminate between samples of such a wine produced in three different areas of Piedmont. The influence of climatic factors on the wine characteristics has also been studied.

Determination of amines and phenolic acids in wine with benzoyl chloride derivatization and liquid chromatography–mass spectrometry

Amine and phenolic metabolites are important contributors to the flavor and health effects of many foods, including wine. Determination of these metabolites often involves UV detection following separation by liquid chromatography. While this is sufficient for some applications, chemical derivatization with LC–MS provides greater sensitivity and selectivity relative to LC-UV.We have developed an assay for 56 amine and phenolic metabolites in wine using benzoyl chloride derivatization and LC–MS. Isotopically labeled benzoyl chloride was used to prepare internal standards for each metabolite. Nanomolar limits of detection were achieved for all metabolites. To demonstrate the application of this assay, we compared metabolite profiles from Merlot and Cabernet Sauvignon wines from California and Australia. We found five metabolites which were significantly different when grouped by varietal, while twenty-four were different when grouped by location of production. This shows that the method can identify differences between various wines.

Direct residue analysis of systemic insecticides and some of their relevant metabolites in wines by liquid chromatography – mass spectrometry

A direct large volume injection (DI-LVI) high performance liquid chromatography – tandem mass spectrometry (HPLC–MS/MS) method was developed and validated for the quantitative determination of 16 systemic insecticides and their main plant metabolites. The assays were conducted on commercial red and white wines made from grapes grown in major wine-producing regions nationally and internationally. Using a 1:20 dilution and an injection volume of 800μL, a limit of quantitation (LOQ) of 1μgL−1 for all analytes was achieved. Matrix-matched standards (MM) were used for accurate quantitation. Imidacloprid (IMI) and methoxyfenozide (MET) were the most frequently detected parent insecticides in the wines reaching concentrations of 1–132μgL−1. Two important plant metabolites imidacloprid-olefin (IMI-OLE) and spirotetramat-enol (SPT-EN) were found at higher concentrations. In five samples SPT-EN was detected in the mgL−1 range with a maximum concentration of 16.3mgL−1 measured in a conventional white wine sample. Most “organic” wines contained no detectable or low insecticide residues, except for one sample, which showed the highest IMI (14.7μgL−1) and IMI-OLE (331μgL−1) concentrations. Considering the maximum residue limit (MRL) definition for the different insecticides, three “conventional” wine samples were non-compliant for SPT. This study highlights the importance to determine both parent and metabolite forms of systemic insecticides in the finished product.

A Comparative Study of Phenols in Apulian Italian Wines.

Nutraceutics is a growing research field in which researchers study and attempt to improve the biological properties of metabolites in food. Wine is one of the most consumed products in the world and contains a plethora of molecules biologically relevant to human health. In this article, several polyphenols with potential antioxidant activity were measured in wines from Apulia, in Southeast Italy. Hydroxytyrosol, gallic and syringic acids, luteolin, quercetin, and trans-resveratrol were identified and quantified by HPLC. The amount of the analyzed metabolites in wines were largely dependent on their color, with red ones being the richest compared to white and rose wines. Gallic acid was the most abundant polyphenol, followed by syringic acid and luteolin. Nevertheless, significant amounts of hydroxytyrosol, quercetin, and trans-resveratrol were also found. The average concentration of polyphenols found in these wines could have potential health-promoting effects, especially if consumed in moderate quantities on a regular basis.

Changes in grape polyphenols (V. vinifera L.) as a consequence of post-harvest withering by high-resolution mass spectrometry: Raboso Piave versus Corvina.

Grape dehydration is an oenological process used for the production of high-quality reinforced and sweet wines. Corvina and Raboso Piave are two red grape varieties used for production of high-quality Italian wines, such as Recioto, Amarone di Valpolicella and Raboso Passito. Changes of polyphenolic composition of the grapes as a consequence of the withering were studied by ultra-high performance liquid chromatography-quadrupole time of flight mass spectrometry (UHPLC/QTOF); for identification of compounds a homemade HR-MS database of grape and wine metabolites, was used. Concomitant with trans-resveratrol and viniferins, relevant increases of other stilbenes (piceatannol, resveratrol trimers and tetramers) and antioxidant compounds (quercetin, syringetin and tamarixetin) were observed. These compounds are part of the induced metabolism occurring during the withering process and in general improve the nutraceutical properties of grapes and wines. On the other hand, longer processes showed to decrease flavan-3-ols and glycoside flavonols. Constant increase of E/Z ε-viniferin ratio was observed in all samples, and this parameter can be used to monitor the process. Copyright © 2016 John Wiley & Sons, Ltd.

An approach to the simultaneous quantitative analysis of metabolites in table wines by 1H NMR self-constructed three-dimensional spectra

Wine consists of several hundred components with different concentrations, including water, ethanol, glycerol, organic acids and sugars. Accurate quantification of target compounds in such complex samples is a difficult task based on conventional 1H NMR spectra due to some challenges. In this paper, the three-dimensional spectrum was constructed firstly by simply repeating 1H NMR spectrum itself so as to extract the features of target compounds by Tchebichef moment method. A proof-of-concept model system, the determination of five metabolites in wines was utilized to evaluate the performance of the proposed strategy. The results indicate that the proposed approach can provide accurate and reliable concentration predictions, probably the best results ever achieved using PLS and interval-PLS methods. Our novel strategy has not only good performance but also does not require laborious multi-step and subjective pretreatments. Therefore, it is expected that the proposed method could extend the application of conventional 1H NMR.

Associations among Wine Grape Microbiome, Metabolome, and Fermentation Behavior Suggest Microbial Contribution to Regional Wine Characteristics.

ABSTRACTRegionally distinct wine characteristics (terroir) are an important aspect of wine production and consumer appreciation. Microbial activity is an integral part of wine production, and grape and wine microbiota present regionally defined patterns associated with vineyard and climatic conditions, but the degree to which these microbial patterns associate with the chemical composition of wine is unclear. Through a longitudinal survey of over 200 commercial wine fermentations, we demonstrate that both grape microbiota and wine metabolite profiles distinguish viticultural area designations and individual vineyards within Napa and Sonoma Counties, California. Associations among wine microbiota and fermentation characteristics suggest new links between microbiota, fermentation performance, and wine properties. The bacterial and fungal consortia of wine fermentations, composed from vineyard and winery sources, correlate with the chemical composition of the finished wines and predict metabolite abundances in finished wines using machine learning models. The use of postharvest microbiota as an early predictor of wine chemical composition is unprecedented and potentially poses a new paradigm for quality control of agricultural products. These findings add further evidence that microbial activity is associated with wine terroir.

Polyphenol metabolomics of twenty Italian red grape varieties

“Suspect screening analysis”method to study grape metabolomics, was performed. This method is a middle-way “targeted” and “untargeted”approach aiming at identifying the largest number of metabolites in grape samples. A new database of putative grape and wine metabolites (GrapeMetabolomics ), which currently contains around 1,100 compounds, was constructed by CREA at Conegliano. By performing high-resolution mass spectrometry analysis of the grape extract in both positive and negative ionization mode, averaging 320-450 putative compounds are identified. Most of them are grape polyphenols, such as anthocyanins, flavonols and stilbene derivatives. By performing PCA and Cluster Analysis the composition in anthocyanins and flavonols of 20 Italian red grape varieties, was studied.

Anionic metabolite profiling by capillary electrophoresis–mass spectrometry using a noncovalent polymeric coating. Orange juice and wine as case studies

In several metabolomic studies, it has already been demonstrated that capillary electrophoresis hyphenated to mass spectrometry (CE–MS) can detect an important group of highly polar and ionized metabolites that are overseen by techniques such as NMR, LC–MS and GC–MS, providing complementary information. In this work, we present a strategy for anionic metabolite profiling by CE–MS using a cationic capillary coating. The polymer, abbreviated as PTH, is composed of a poly-(N,N,N’,N’-tetraethyldiethylenetriamine, N-(2-hydroxypropyl) methacrylamide, TEDETAMA-co-HPMA (50:50) copolymer. A CE–MS method based on PTH-coating was optimized for the analysis of a group of 16 standard anionic metabolites. Separation was achieved within 12min, with high separation efficiency (up to 92,000 theoretical plates per meter), and good repeatability, namely, relative standard deviation values for migration times and peak areas were below 0.2 and 2.1%, respectively. The optimized method allowed the detection of 87 metabolites in orange juice and 142 metabolites in red wine, demonstrating the good possibilities of this strategy for metabolomic applications.

CHARACTERIZATION OF SOME ITALIAN V. VINIFERA RED GRAPE CULTIVARS ON THE BASIS OF THEIR ANTHOCYANIN PROFILES

Suspect screening analysis? is a middle-way approach in metabolomics between ?targeted? and ?untargeted? analyses. To develop a new method of grape metabolomics, a new database of putative grape and wine metabolites (GrapeMetabolomics) has been expressly constructed. Currently, this database contains more than a thousand compounds. By performing high-resolution mass spectrometric analyses of grape extracts in positive and negative ionization mode, and using GrapeMetabolomics, 320-450 putative compounds have been identified. Most of them are grape metabolites very important in determining the organoleptic characteristics of wine or with biological proprieties, such as anthocyanins, stilbene derivatives, and flavonols. In hybrid grapes, this method allowed the identification of 30 anthocyanins. In the present study, this method was used to study the anthocyanin composition of ten autochthonous Italian V. vinifera red grape cultivars. By performing statistical analyses (PCA and cluster analysis), the effect of cultivar on the anthocyanin composition of the grape was studied.

Integrating analytical resolutions in non-targeted wine metabolomics

Direct injection Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), and ultra-high performance liquid chromatography coupled to mass spectrometry (UPLC/MS) were combined for the non-targeted analysis of wine metabolites. The unrivalled resolution on mass measurement allowed by the former and the separation ability of isomeric and isobaric substances by the latter, clearly increases the scope of detectable unknown metabolites in wines. Such methodology is illustrated through the comparison of chemical spaces of a young and an older Pinot noir wine. RP and HILIC chromatography could reveal up to five isomers for a given mass, throughout the explored mass range. CHO, CHOS and CHONS chemical spaces exhibited higher diversities in the older wine, illustrating a molecular oeno-diagenesis process during wine ageing, whereas nitrogen-containing compounds (CHON chemical space) appeared to be fewer in the older wine, likely due to precipitation. This methodological combination is a promising contribution to the wine metabolomics toolkit.

Untargeted metabolomic analysis using liquid chromatography quadrupole time-of-flight mass spectrometry for non-volatile profiling of wines

The current study presents a method for comprehensive untargeted metabolomic fingerprinting of the non-volatile profile of the Graciano Vitis vinifera wine variety, using liquid chromatography/electrospray ionization time of flight mass spectrometry (LC–ESI-QTOF). Pre-treatment of samples, chromatographic columns, mobile phases, elution gradients and ionization sources, were evaluated for the extraction of the maximum number of metabolites in red wine. Putative compounds were extracted from the raw data using the extraction algorithm, molecular feature extractor (MFE). For the metabolite identification the WinMet database was designed based on electronic databases and literature research and includes only the putative metabolites reported to be present in oenological matrices. The results from WinMet were compared with those in the METLIN database to evaluate how much the databases overlap for performing identifications. The reproducibility of the analysis was assessed using manual processing following replicate injections of Vitis vinifera cv. Graciano wine spiked with external standards. In the present work, 411 different metabolites in Graciano Vitis vinifera red wine were identified, including primary wine metabolites such as sugars (4%), amino acids (23%), biogenic amines (4%), fatty acids (2%), and organic acids (32%) and secondary metabolites such as phenols (27%) and esters (8%). Significant differences between varieties Tempranillo and Graciano were related to the presence of fifteen specific compounds.

Characterization of some ItalianV. viniferaL. grape varieties on the basis of their flavonol profile

“Suspect screening metabolomics” is a mid-way approach between “targeted” and “untargeted” analysis. For this aim, a new database of putative grape and wine metabolites (GrapeMetabolomics) was expressly constructed. Currently, this database contains around 1,100 compounds. By performing UHPLC/QTOF mass spectrometry analysis in both positive and negative ionization mode, in a grape extract averaging 320–450 putative compounds are identified. Most of these compounds are important grape metabolites, including flavonols, anthocyanins, and stilbene derivatives. In the present study, this approach was focalized on the characterization of flavonols of 18 important Italian red and white grape varieties and the method provided the identification of 15 flavonols. By performing statistical analysis (Principal Component Analysis and Cluster Analysis), the effect of the variety on the flavonol composition of the grapes was studied. Both the red and white samples fell into three different groups, respectively, on the basis of their flavonol profiles. Because the samples were cultivated in the same vineyard, their profile potentially was not affected by cultural or environmental factors. Anyway, these preliminary results will have to be confirmed by the study of grape samples collected in different years and from different vineyards.

The effect of leaf area to yield ratio on secondary metabolites in grapes and wines of <em>Vitis vinifera</em> L. cv. Sauvignon Blanc

<p style="text-align: justify;"><strong>Aim</strong>: To investigate the effect of reducing leaf area by shoot hedging in combination with bunch thinning on metabolite concentration and sensorial quality of Sauvignon blanc grapes and wines.</p><p style="text-align: justify;"><strong>Methods and results</strong>: Four vine treatments were conducted: shoot hedging/bunch thinning (SH/BT), shoot hedging/no bunch thinning (SH/NBT), full canopy/bunch thinning (FC/BT) and full canopy/no bunch thinning (FC/NBT). Shoot hedging delayed total soluble solids accumulation at the beginning of the grape maturation in SH/BT and SH/NBT treatments. At harvest there were no significant differences in the concentration of hydroxycinnamoyl tartaric acids, glutathione, total soluble solids, titratable acidity and pH value in grape juice between all treatments and methoxypyrazines were below the limit of detection. Lutein concentration in grape berry was higher in treatments without bunch thinning, while there was no significant difference in the concentration of β-carotene and neoxanthin. The highest leaf area to yield ratio (FC/BT) resulted in higher concentration of glutathione in must and higher concentration of thiols in Sauvignon blanc wines. Upon sensory evaluation, the FC/BT wine was best scored for overall quality and heavier tropical aroma, whereas the FC/NBT wine was best scored for fresh tropical aroma and second best for overall quality.</p><p style="text-align: justify;"><strong>Conclusion</strong>: Leaf area to yield ratio impacted berry ripening kinetics, grape and wine metabolite composition, and sensorial properties of Sauvignon blanc wine.</p><p style="text-align: justify;"><strong>Significance and impact of the study</strong>: The study showed that the highest leaf area to yield ratio resulted in the best overall sensorial quality of wine.</p>

Characterisation of the flavour profile from Graciano Vitis vinifera wine variety by a novel dual stir bar sorptive extraction methodology coupled to thermal desorption and gas chromatography–mass spectrometry

The aim of this work was to develop a new analytical technique for the study of the organoleptic compounds (flavour profile) of the Graciano Vitis vinifera wine variety. The cv. Graciano is a singular variety of red grapes with its origins in La Rioja and Navarra (northern Spain). This variety transfers an intense red colour, aroma and high acidity to musts and provides greater longevity and, consequently, a better capacity for ageing wine. A new dual-stir bar sorptive extraction approach coupled with thermal desorption (TD) and GC–MS has been used to extract the volatile and semivolatile compounds. In this extraction step, the optimal values for the experimental variables were obtained through the Response Surface Methodology (RSM). Full scan chromatogram data were evaluated with two deconvolution software tools, and the results were compared. The volatile and semivolatile components were identified with an MS match ≥80%. As a result, the flavour metabolome of the Graciano Vitis vinifera wine variety was obtained, and 205 metabolites were identified using different databases. These metabolites were grouped into esters, acids, alcohols, nitrogen compounds, furans, lactones, ketones, aldehydes, phenols, terpenes, norisoprenoids, sulphur compounds, acetals and pyrans. The majority of the metabolites observed had already been reported in the literature; however, this work also identified new, previously unreported metabolites in red wines, which may be characteristic of the Graciano variety.

1H‐NMR‐based metabolomic analysis of the effect of moderate wine consumption on subjects with cardiovascular risk factors

Moderate wine consumption is associated with health-promoting activities. An H-NMR-based metabolomic approach was used to identify urinary metabolomic differences of moderate wine intake in the setting of a prospective, randomized, crossover, and controlled trial. Sixty-one male volunteers with high cardiovascular risk factors followed three dietary interventions (28 days): dealcoholized red wine (RWD) (272mL/day, polyphenol control), alcoholized red wine (RWA) (272mL/day) and gin (GIN) (100mL/day, alcohol control). After each period, 24-h urine samples were collected and analyzed by (1) H-NMR. According to the results of a one-way ANOVA, significant markers were grouped in four categories: alcohol-related markers (ethanol); gin-related markers; wine-related markers; and gut microbiota markers (hippurate and 4-hydroxphenylacetic acid). Wine metabolites were classified into two groups; first, metabolites of food metabolome: tartrate (RWA and RWD), ethanol, and mannitol (RWA); and second, biomarkers that relates to endogenous modifications after wine consumption, comprising branched-chain amino acid (BCAA) metabolite (3-methyl-oxovalerate). Additionally, a possible interaction between alcohol and gut-related biomarkers has been identified. To our knowledge, this is the first time that this approach has been applied in a nutritional intervention with red wine. The results show the capacity of this approach to obtain a comprehensive metabolome picture including food metabolome and endogenous biomarkers of moderate wine intake.

Characterization of microbial metabolism of Syrah grape products in an in vitro colon model using targeted and non-targeted analytical approaches.

PurposeSyrah red grapes are used in the production of tannin-rich red wines. Tannins are high molecular weight molecules, proanthocyanidins (PAs), and poorly absorbed in the upper intestine. In this study, gut microbial metabolism of Syrah grape phenolic compounds was investigated.MethodsSyrah grape pericarp was subjected to an enzymatic in vitro digestion model, and red wine and grape skin PA fraction were prepared. Microbial conversion was screened using an in vitro colon model with faecal microbiota, by measurement of short-chain fatty acids by gas chromatography (GC) and microbial phenolic metabolites using GC with mass detection (GC–MS). Red wine metabolites were further profiled using two-dimensional GC mass spectrometry (GCxGC-TOFMS). In addition, the effect of PA structure and dose on conversion efficiency was investigated by GC–MS.ResultsRed wine exhibited a higher degree of C1–C3 phenolic acid formation than PA fraction or grape pericarp powders. Hydroxyphenyl valeric acid (flavanols and PAs as precursors) and 3,5-dimethoxy-4-hydroxybenzoic acid (anthocyanin as a precursor) were identified from the red wine metabolite profile. In the absence of native grape pericarp or red wine matrix, the isolated PAs were found to be effective in the dose-dependent inhibition of microbial conversions and short-chain fatty acid formation.ConclusionsMetabolite profiling was complementary to targeted analysis. The identified metabolites had biological relevance, because the structures of the metabolites resembled fragments of their grape phenolic precursors or were in agreement with literature data.Electronic supplementary materialThe online version of this article (doi:10.1007/s00394-012-0391-8) contains supplementary material, which is available to authorized users.

Anthocyanins and Their Variation in Red Wines I. Monomeric Anthocyanins and Their Color Expression

Originating in the grapes, monomeric anthocyanins in young red wines contribute the majority of color and the supposed beneficial health effects related to their consumption, and as such they are recognized as one of the most important groups of phenolic metabolites in red wines. In recent years, our increasing knowledge of the chemical complexity of the monomeric anthocyanins, their stability, together with the phenomena such as self-association and copigmentation that can stabilize and enhance their color has helped to explain their color representation in red wine making and aging. A series of new enological practices were developed to improve the anthocyanin extraction, as well as their color expression and maintenance. This paper summarizes the most recent advances in the studies of the monomeric anthocyanins in red wines, emphasizing their origin, occurrence, color enhancing effects, their degradation and the effect of various enological practices on them.