1H nuclear magnetic resonance-based metabolomic characterization of wines by grape varieties and production areas

(1)H NMR spectroscopy was used to investigate the metabolic differences in wines produced from different grape varieties and different regions. A significant separation among wines from Campbell Early, Cabernet Sauvignon, and Shiraz grapes was observed using principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA). The metabolites contributing to the separation were assigned to be 2,3-butanediol, lactate, acetate, proline, succinate, malate, glycerol, tartarate, glucose, and phenolic compounds by PCA and PLS-DA loading plots. Wines produced from Cabernet Sauvignon grapes harvested in the continental areas of Australia, France, and California were also separated. PLS-DA loading plots revealed that the level of proline in Californian Cabernet Sauvignon wines was higher than that in Australian and French Cabernet Sauvignon, Australian Shiraz, and Korean Campbell Early wines, showing that the chemical composition of the grape berries varies with the variety and growing area. This study highlights the applicability of NMR-based metabolomics with multivariate statistical data sets in determining wine quality and product origin.     

Model aging and oxidation effects on varietal, fermentative, and sulfur compounds in a dry botrytized red wine

From harvest until wine arrives to the consumer, oxygen plays a crucial role in the definition of the final aroma. In the present research, the effect of the model oxidative aging on a dry red Botrytis wine, such as Italian Amarone, was considered. Amarone wine was submitted to model oxidative aging and then analyzed with two different approaches (SPE-GC-MS and HS-SPME/GC-MS). The same sampling plan was adopted to study the model aging of the same Amarone wine in anaerobic conditions. The HS-SPME/GC-MS method was applied to investigate for the first time the effect of the oxidative aging on a vast number of fermentative sulfur compounds. This research highlighted peculiar evolutions for several volatile compounds. In particular, benzaldehyde showed a sensitive increment during the oxidative aging, with a rate much higher than that reported for non-Botrytis red wines. On the other hand, several sulfides (dimethyl sulfide, 3-(methylthio)-1-propanol, etc.) disappeared after just 15 days of oxidative aging. A wine oxidation marker such as 3-(methylthio)-propanal was not found in any of the oxidized wines; conversely methionol-S-oxide was tentatively identified. This evidence has not been mentioned in the literature. A possible involvement of grape withering process and Botrytis in these mechanisms was supposed: a dry red wine, produced from the same but without any grape withering process and Botrytis infection (e.g., Bardolino wine), was submitted to oxidative aging and analysis. This red wine showed an evolution similar to those reported in the literature for dry red wines but significantly different from the Amarone wine.     

Evidence for protein degradation by Botrytis cinerea and relationships with alteration of synthetic wine foaming properties

Botrytis cinerea is an important fungal pathogen particularly dreaded in the cool climate vineyard. It is responsible for important damage, especially the decrease in foamability of sparkling wines, such as Champagne. Different studies have shown that proteins are largely involved in the stabilization of Champagne foam despite their low concentration. Other works demonstrated changes in the electrophoretic characteristics of must proteins originating from botrytized grapes, although the cause of such alterations was never explained. In the first part of this study, results showed the release by B. cinerea of 3.5 mg/L total proteins in a synthetic liquid medium. Among these proteins, the presence of a protease activity on bovine serum albumin (BSA) and must proteins was demonstrated by using a colorimetric method and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the model wine, the Bradford method showed a BSA loss of 66% after 24 h and a loss of 96% after 120 h. In the same model wine, the soluble must protein concentration decreased by 35% after 1 week and by 53% after 2 weeks while the control showed no protein loss. B. cinerea proteases were then able to degrade BSA and must proteins and were above all active at must and wine pH and in the presence of ethanol and SO(2). The second part of this work was dedicated to the relationship between the presence of B. cinerea proteases and its effects on the synthetic wine foaming properties. The addition of a B. cinerea culture medium (1/33 v/v) to the synthetic wine containing 21 mg/L soluble grape proteins induced a decrease in foamability by 60% after 1 week. For BSA in the model wine, the foamability decreased by 32% after 24 h and by 95% after 120 h, as shown by the colorimetric method. These experiments demonstrate for the first time the relationship between B. cinerea protease activity and the decrease in wine foaming properties.     

Odorous impact of volatile thiols on the aroma of young botrytized sweet wines: identification and quantification of new sulfanyl alcohols

Specific extraction of volatile thiols using sodium p-hydroxymercuribenzoate revealed the presence of three new sulfanylalcohols in wines made from Botrytis-infected grapes: 3-sulfanylpentan-1-ol (II), 3-sulfanylheptan-1-ol (III), and 2-methyl-3-sulfanylbutan-1-ol (IV). The first two have citrus aromas, whereas the third is reminiscent of raw onion. In addition, 2-methyl-3-sulfanylpentan-1-ol, which has a raw onion odor, was tentatively identified. Like 3-sulfanylhexan-1-ol (I), already reported in Sauternes wines, compounds II, III, and IV were absent from must. They were found in wine after alcoholic fermentation, and their concentrations were drastically higher when Botrytis cinerea had developed on the grapes. In the commercial botrytized wines analyzed, the mean levels of II, III, and IV were 209, 51, and 103 ng/L, respectively. Despite their low odor activity values, sensory tests showed additive effects among I, II, and III, thus confirming their olfactory impact on the overall aroma of botrytized wines.     

Determination of the grape invertase content (using PTA-ELISA) following various fining treatments versus changes in the total protein content of wine. relationships with wine foamability

Proteins have proven to play a major role in the stabilization of foam in Champagne wines despite their low concentration that ranges from 4 to 20 mg/L. The aim of this study was to evaluate the effect of fining on total protein and grape invertase contents of champenois base wines and their foaming properties. Data showed that fining and especially the use of bentonite at doses ranging from 10 to 50 g/hL leads to a significant decrease in the total protein content of wines together with that of the grape invertase content, with such a decrease being very detrimental to the foaming properties of the treated wines in terms of foam height (HM) and foam stability (HS). Only a slight decrease in the total protein content, in the grape invertase concentration, and in the foam quality of wines was observed when using casein (10 and 20 g/hL) or bentonite combined with casein (both at 20 g/hL). Our study thus clearly establishes the good correlation existing between the wine protein concentration and its foaming properties. A remarkable correlation was observed between the decrease in the grape invertase content and the total protein content of wines, following bentonite treatments, suggesting that the grape invertase (which represents at least 10-20% of the wine proteins) follows a similar behavior upon fining to other proteins of Champagne wines, despite the high molecular mass and the highly glycosylated structure of this particular protein. Moreover, the decrease in total protein and grape invertase contents of wine after fining with bentonite was found to be correlated with a decrease in the foaming properties of the corresponding wines (with respectively R(2) = 0.89 and 0.95).     

3-Sulfanylhexanol precursor biogenesis in grapevine cells: the stimulating effect of Botrytis cinerea

Volatile thiols, compounds that contribute strongly to the varietal aroma, are present in much higher concentrations in sweet wines than in dry wines. This positive effect, due to the presence of Botrytis cinerea on the berries, in fact results from a strong enrichment of cysteine S-conjugate precursors in botrytized berries. In the present study, a convenient model was investigated to reproduce and therefore study this phenomenon. A Vitis vinifera cell culture was used as a simple model, and we focused on S-3-(hexan-1-ol)-l-cysteine (P-3SH), the cysteinylated precursor of 3-sulfanylhexanol. We demonstrated that grapevine cells were able to produce P-3SH and that the presence of B. cinerea considerably increased the precursor level (up to 1000-fold). This positive result was determined to be due to metabolites secreted by the fungus. These molecules were temperature sensitive, unstable over time, and their production was activated in the presence of grapevine cells. Moreover, part of the pathway leading to P-3SH was deciphered: it was directly derived from the cleavage of S-3-(hexan-1-ol)-l-glutathione, which itself was generated after a conjugation of glutathione on (E)-2-hexenal.     

Levels of stilbene oligomers and astilbin in French varietal wines and in grapes during noble rot development

Phenolics from grapes and wines can play a role against oxidation and development of atherosclerosis. Stilbenes have been shown to have cancer chemopreventive activity and to protect lipoproteins from oxidative damage. A method for the direct determination of stilbene oligomers (viniferin and pallidol) as well as astilbin in different types of wine using high-performance liquid chromatography with UV detection is described. In a survey of 21 commercial wines from the south of France, levels of pallidol and viniferin are reported for the first time in different types of wines. Viniferin was found to be present only in red and botrytized sweet white wines with levels between 0.1 and 1.63 mg/L; pallidol was not found in dry and sweet white wines but only in wines made by maceration with stems, with levels between 0.38 and 2.22 mg/L. Highest levels of astilbin were found in Egiodola (15.13 mg/L), Merlot (11.61 mg/L), and Cabernet Sauvignon (8.24 mg/L) for red wines and in Sauvignon (5.04 mg/L) for white varietal wines. Astilbin levels are highest for recent vintages, but pallidol is not found in older vintages. During noble rot development in Sauvignon or Sémillon grapes from the Sauternes area, levels of trans-astringin, trans-resveratrol, trans-piceid, and pallidol are quite low (<0.5 mg/kg of grapes). Viniferin and astilbin levels become optimum at 2 and 30 mg/kg, respectively, during spot grape and speckle grape stages.     

Protection of originality of Tokaji Aszú: amines and organic acids in botrytized wines by high-performance liquid chromatography

Amine and organic acid composition of Aszú wines from the Tokaj region of Hungary, nonbotrytized Hungarian wines from different regions, and foreign botrytized wines were analyzed by high-performance liquid chromatography. Hungarian and foreign wines (36 Hungarian and seven foreign botrytized wines) were compared in different ways by calculation of ratios of given amine compounds, analyses of variance, principal component, and discriminant analysis. In wines, putrescine and in some samples 3-methyl-butylamine and/or phenyl ethylamine were found in remarkable concentrations, while in botrytized wines four other amines were verified in high concentration. Good separation between Aszú and foreign botrytized wines was found by calculation of the amine component's ratio. The first two principal components of the principal component analysis accounted for 77 and 84% of the total variance in the data of amines and acids, respectively. The component scores of samples grouped according to Aszú, foreign botrytized, and nonbotrytized wines. Linear discriminant analysis was used for differentiation of Aszú, foreign botrytized, and normal wines. Using nine amines and two acids as variables, the correct classification was 97.6%. On the basis of results, an objective evaluation method can be elaborated for quality control in order to protect the authenticity and origin of wine specialties made from botrytized grapes.     

Content of the flavonols myricetin, quercetin, and kaempferol in finnish berry wines

The amounts of myricetin, quercetin, and kaempferol were analyzed in 16 red and 2 white berry and grape wines after acid hydrolysis using an RP-HPLC method with diode array detection. The red berry wines analyzed were made mainly from black currant, crowberry, and bog whortleberry, i.e., berries rich in flavonols. The red grape wines were made mainly from Cabernet Sauvignon or Merlot grapes in several countries. The white wines studied were gooseberry and white currant wines and Chardonnay and Riesling wines. The amount of myricetin ranged from 3.8 to 22.6 mg L(-1) in red berry wines and from 0 to 14.6 mg L(-1) in red grape wines. The amount of quercetin was from 2.2 to 24.3 mg L(-1) red berry wines and from <1.2 to 19.4 mg L(-1) in red grape wines. Low levels of kaempferol were found in all red berry wines and in 9 red grape wines. The total concentration of these flavonols was from 6 to 46 mg L(-1) (mean 20 mg L(-1)) in red berry wines and from 4 to 31 mg L(-1) (mean 15 mg L(-1)) in red grape wines. Small amounts of quercetin were found in white currant and gooseberry wines, whereas no flavonols were detected in white grape wines. These results demonstrate that the contents of flavonols in red     

Champagne wine polyphenols protect primary cortical neurons against peroxynitrite-induced injury

White wines are generally low in polyphenol content as compared to red wines. However, Champagne wines have been shown to contain relatively high amounts of phenolic acids that may exert protective cellular actions in vivo. In this study, we have investigated the potential neuroprotective effects of Champagne wine extracts, and individual phenolics present in these extracts, against peroxynitrite-induced injury. Organic and aqueous Champagne wine extracts exhibited potent neuroprotective activity against peroxynitrite-induced injury at low concentrations (0.1 microg/mL). This protection appeared to be in part due to the cellular actions of individual components found in the organic extracts, notably tyrosol, caffeic acid, and gallic acid. These phenolics were observed to exert potent neuroprotection at concentrations between 0.1 and 10 microM. Together, these data suggest that polyphenols present in Champagne wine may induce a neuroprotective effect against oxidative neuronal injury.     

Gas chromatographic quantification of major volatile compounds and polyols in wine by direct injection

Methanol, propanol, isobutanol, isoamyl alcohol, 2-phenylethanol, acetaldehyde, 1,1-diethoxyethane, acetoin, ethyl acetate, ethyl lactate, and ethyl succinate and the polyols 2,3-butanediol (levo and meso forms) and glycerol were quantified by direct injection of wine samples. Linear responses over the usual concentration ranges for these compounds and r2 values from 0.9932 to 0.9998 were obtained. The confidence limits for the mean values ranged from 2.34% for diethyl succinate to 8.52% for 1,1-diethoxyethane, both at a probability level of 0.05. Relative errors ranged from 8 to 10% for the polyols and 1,1-diethoxyethane and were all less than 5% for alcohols and acetaldehyde. The proposed method is useful with a view to identifying relationships between alcoholic fermentation byproducts and controlling biological or chemical aging in wines.     

Red wine making by immobilized cells and influence on volatile composition

Red wine making using yeast cells immobilized in two types of raisin berries, at various temperatures (6-30 degrees C), was studied. A modification of the batch bioreactor was used to separate the grape skins used for color extraction from the biocatalyst and the fermenting grape must. The evaluation of the immobilized biocatalysts was made on terms of productivity and organoleptic quality, including color intensity and formation of volatiles. The immobilized cells were found capable of low-temperature wine making, producing red wines containing more than 11% v/v alcohol in 8 days at 6 degrees C. The quality of wines was examined by gas chromatography (GC) and GC-MS analysis and sensory evaluation. Higher alcohol concentrations were decreased, and ethyl acetate concentrations increased by the drop of temperature. Many esters, alcohols, carbonyls, and miscellaneous compounds were identified in wines produced by immobilized cells, revealing no significant qualitative differences as compared to wines produced by free cells. The sensory evaluation showed that the best red wine was produced at 6 degrees C.     

Vineyard and fermentation studies to elucidate the origin of 1,8-cineole in Australian red wine

Preliminary investigations revealed that the proximity of Eucalyptus trees to grapevines can directly influence the concentration of the aroma compound 1,8-cineole present in the corresponding red wines. For two different vineyards, the closer the grapevines were to the trees, the greater was the amount of 1,8-cineole in the wines elaborated from those grapes. This led us to carry out further studies to quantify the levels of 1,8-cineole found in grape berries, leaves, and stems at set distances from Eucalyptus trees over multiple vintages. Generally, the highest concentration of 1,8-cineole was found in the grapevine leaves, followed by grape stems and then grapes. In each sample type, we observed greater concentrations of 1,8-cineole in samples closer to the trees. Various fermentation treatments carried out with Shiraz grapes showed that matter other than grapes (MOG, e.g., Eucalyptus or grape leaves) could contribute significant amounts of 1,8-cineole to the finished wines. These studies confirmed that vineyard position and winemaking conditions can determine the 1,8-cineole concentration in red wine. The fermentation study also showed for the first time that the concentration of rotundone in red wine can be strongly influenced by grapevine leaves and stems in the ferment.     

Gluconic acid consumption in wines by Schizosaccharomyces pombe and its effect on the concentrations of major volatile compounds and polyols

Schizosaccharomyces pombe 1379 (ATCC 26760) yeast strain in wine substantially increases acetaldehyde and 1,1-diethoxyethane concentrations and to decreases levo-2,3-butanediol, glycerol, acetoin, and gluconic acid concentrations. In this study, S. pombe has been used for the first time to reduce gluconic acid in wine under aerobic conditions. Only acetaldehyde and acetoin exhibited significantly higher levels in the wines containing gluconic acid. The high in vitro specific activity of alcohol dehydrogenase observed may be directly related to the high production of acetaldehyde by the studied fission yeast.     

Associations between the sensory attributes and volatile composition of Cabernet Sauvignon wines and the volatile composition of the grapes used for their production

The sensory properties of wine are influenced by the chemical composition of the grapes used to produce them. Identification of grape and wine chemical markers associated with the attributes perceived by the consumer of the wine will enable better prediction of the potential of a parcel of grapes to produce wine of a certain flavor. This study explores the relationships between Cabernet Sauvignon grape volatile composition and wine volatile profiles with the sensory properties of wines. Twenty grape samples were obtained from nine vineyard sites across three vintages and wines vinified from these parcels using controlled winemaking methods. The volatile composition of the grapes were analyzed by SBSE-GCMS, the wines were analyzed by SPME-GCMS, and these data sets were compared to that obtained from the sensory analysis of the wines. Statistical treatment of the data to account for vintage and region effects allowed underlying relationships to be seen between wine sensory attributes and wine or grape volatile components. The observed associations between grape or wine volatile compounds and wine sensory attributes has revealed target compounds and pathways whose levels may reflect the biochemical effects on grape composition by differing growth conditions during berry development and ripening. The compounds identified in this study may be useful grape or wine markers for potential wine sensory characteristics.     

Contribution of dimethyl sulfide to the aroma of Syrah and Grenache Noir wines and estimation of its potential in grapes of these varieties

The contribution of dimethyl sulfide (DMS) to the aroma of Syrah and Grenache Noir wines from the Rhone Valley of France was investigated by sensory analysis, and its levels in these wines were measured. The potential DMS in the corresponding grapes and wines, susceptible to release during wine aging, was evaluated. Free DMS and potential DMS assessed by a heat-alkaline treatment were measured in grape juices and wines by SPME-GC-MS using methods previously reported and slightly modified. A relationship between potential DMS from grapes and the total DMS levels in wine was demonstrated. Furthermore, a linear regression between the ratio of free DMS levels to these total DMS levels in wine and time of storage was found. Free and potential DMS levels in grapes and wines depended on grape variety, vintage, and vine location. DMS imparted a noticeable and complex contribution to the aroma of the wines investigated, depending on the mode of sensory perception used, either before or after glass swirling. It significantly enhanced the fruity notes of the wines, and additional truffle and black olive notes.     

Polysaccharide profile and content during the vinification and aging of Tempranillo red wines

Passing from must to wine produced a loss of low-molecular-weight grape structural glucosyl polysaccharides, and an important gain in yeast mannoproteins (MP) and grape-derived arabinogalactan proteins (AGP), and rhamnogalacturonans-II (RG-II). AGP were more easily extracted than RG-II, and small quantities of RG-II monomers and galacturonans were detected. Postmaceration produced a reduction in all grape polysaccharide families, particularly acute in AGP. The reduction of polysaccharides during malolactic fermentation only affected grape AGP, and MP were continuously liberated during the entire vinification process. Wine oak and bottle aging was associated with a relative stability of the polysaccharide families. AGP were thus the majority polysaccharides in young wines but, contrary to what may be thought, structural glucosyl oligosaccharides dominated in musts and MP in aged wines. Precipitation of polysaccharides was noticeable during vinification, and it mainly affected high-molecular-weight AGP and MP. Hydrolytic phenomena affected the balance of wine polysaccharides during late maceration-fermentation.     

Evolution and occurrence of 1,8-cineole (eucalyptol) in Australian wine

A new method has been developed for the quantitation of 1,8-cineole in red and white wines using headspace solid-phase microextraction (SPME) combined with stable isotope dilution analysis (SIDA) and gas chromatography-mass spectrometry (GC-MS). An extensive survey of Australian wines (44 white and 146 red) highlighted that only red wines contained significant amounts of 1,8-cineole (up to 20 μg/L). Hydrolytic studies with limonene and α-terpineol, putative precursors to 1,8-cineole, showed a very low conversion into 1,8-cineole (< 0.6%) over a 2 year period, which does not account for the difference between white and red wines. 1,8-Cineole was chemically stable in model wine solution over 2 years, and absorption from a Shiraz wine by bottle closures was most evident for a synthetic closure only (14% absorption after 1 year). Two commercial ferments at two different locations were monitored daily to investigate the evolution of 1,8-cineole throughout fermentation. Both ferments showed daily increases in 1,8-cineole concentration while in contact with grape solids, but this accumulation ceased immediately after pressing. This observation is consistent with the extraction of 1,8-cineole into the ferment from the solid portions of the grape berries.     

Phenolic and volatile composition of wines made from Vitis vinifera cv. Muscat lefko grapes from the island of Samos

A study of the phenolic and volatile composition of wines produced from the white cultivar Muscat lefko from the island of Samos was conducted. Dry, fortified, naturally sweet wines and mistelles (aged and nonaged) have been studied. The phenolic components (flavan-3-ols, hydroxycinnamates, and flavonols) were measured by high-performance liquid chromatography after solid phase extraction (SPE). The terpenes (free and glycosidically linked) were determined by the use of gas chromatography-mass spectrometry (GC-MS) after SPE. The fermentation aroma components were analyzed by GC-MS after liquid-liquid extraction. It was found that the dry wines contained lower amounts of most of the phenolics and higher quantities of terpenes and fermentation aroma compounds than the sweet wines. The aged mistelle wines contained lower levels of coutaric and caftaric acids, higher concentrations of the free acids, and markedly fewer free and bound terpenes and fermentation aroma components compared to the other sweet wines. The naturally sweet wine contained relatively increased amounts of phenolics, 2,3-butanediol, and glycosidically linked terpenes.