Effect of mixed species alcoholic fermentation on growth and malolactic activity of lactic acid bacteria.

In recent years, there is an increasing interest from the winemaking industry for the use of mixed fermentations with Starmerella bacillaris (synonym Candida zemplinina) and Saccharomyces cerevisiae, due to their ability to modulate metabolites of oenological interest. The current study was carried out to elucidate the effect of this fermentation protocol on the growth and malolactic activity of lactic acid bacteria (LAB) used for malolactic fermentation (MLF) and on the chemical and volatile profile of Nebbiolo wines and their chromatic characteristics. To this end, two LAB species, namely Lactobacillus plantarum and Oenococcus oeni, were inoculated at the beginning and at the end of the alcoholic fermentation (AF) performed by pure and mixed yeast using the abovementioned yeasts. The different yeast inoculation protocols and the combination of species tested influenced greatly the interactions and behavior of the inoculated yeasts and LAB during AF and MLF. For both LAB species, inoculation timing was critical to how rapidly MLF started and finished. Fermentation inoculated with L. plantarum, at the beginning of the AF, completed MLF faster than those inoculated with O. oeni. The presence of Starm. bacillaris in mixed fermentation promoted LAB growth and activity, in particular, O. oeni. Furthermore, LAB species choice had a greater impact on the volatile and chromatic profile of the wines than inoculation time. These findings reveal new knowledge about the importance of LAB species choice and inoculation time to ensure fast MLF completion and to improve wine characteristics in mixed fermentation with Starm. bacillaris and S. cerevisiae.

Aromatic Profiling of New Table Grape Varieties Using Gas Chromatography/Mass Spectrometry and Olfactometry.

The aim of this study is the aromatic characterization of new table grape varieties, namely Guzun (V. vinifera), Melona (V. vinifera), Cotton Candy (V. vinifera), IVC SA3 (V. labrusca), and IVC SB1 (V. labrusca). The qualitative and quantitative analysis of odorant molecules present in the berries allows for the definition of the aroma profile of the grape. This analysis benefits from the progress of analytical techniques and sensory methodologies. Gas chromatography/mass detection enable the efficient detection of the substances present and their concentrations. Through the coupling of gas chromatography with sensory detection (gas chromatography-olfactometry), it is possible to correlate the compounds detected by gas chromatography with olfactory stimuli, exploiting the human olfactory system. Aroma, a significant flavor component, is an important attribute of table grape that contributes to defining their quality. This characteristic is highly valued by consumers, and consequently, the market asks for table grapes with a particular or new aroma. Aromatic characterization is a crucial step in the study of the table grape varieties to evaluate their potential at the commercial level or, for instance, in breeding programs focusing on organoleptic properties.

Aroma precursors of Grignolino grapes (Vitis vinifera L.) and their modulation by vintage in a climate change scenario.

Current climatic conditions may cause significant changes in grapevine phenology and maturity dynamics linked often with changes to ecoclimatic indicators. The influence exerted by different meteorological conditions during four consecutive years on the aromatic potential of Grignolino grapes was investigated for the first time. The samples were collected from three vineyards characterized by different microclimatic conditions mainly related to the vineyard exposure and by a different age of the plants. Important differences as far as temperature and rainfall patterns are concerned during ripening were observed among the 4 years. Grape responses to abiotic stress, with particular emphasis on aromatic precursors, were evaluated using gas chromatography coupled to mass spectrometry. The results highlighted significant differences among the vintages for each vineyard in terms of the berry weight and the aromatic precursor concentration. For the grapes of the younger-vine vineyard, the content of aroma compounds showed a different variability among the vintages if compared to the old-vine vineyards. Optimal conditions in terms of temperature and rainfall during the green phase followed by a warm and dry post-veraison period until harvest favored all classes of compounds especially terpenoids mainly in the grapes of the old vines. High-temperature (>30°C) and low-rainfall pattern before veraison led to high benzenoid contents and increased differences among vineyards such as berry weight, whereas cooler conditions favored the terpenoid levels in grapes from southeast-oriented vineyards. In a hilly environment, lack of rainfall and high temperature that lately characterize the second part of berry development seem to favor the grape quality of Grignolino, a cultivar of medium-late ripening, by limiting the differences on bunch ripening, allowing a greater accumulation of secondary metabolites but maintaining at the same time an optimum balance sugar/acidity.

Tasting the Italian Terroir through Craft Beer: Quality and Sensory Assessment of Cascade Hops Grown in Central Italy and Derived Monovarietal Beers.

The present study aimed to chemically and sensorially characterize hop samples, cv Cascade, grown in two different Italian regions (Latium and Tuscany) as well as their derived beers by a multi analytical approach. Significant differences in bitter acid, oil and polyphenol content were observed for hop samples according to their origin. Gas chromatography-olfactometry analysis pointed out floral notes for Tuscany samples, where hops from Latium were characterized by spicy and resinous notes, correlated to the presence of sesquiterpenes. Differences in the molecular fingerprinting were also highlighted by Fourier-Transform Infrared Spectroscopy. The differences found in the hops were reflected in the beers, which were clearly recognized as distinct by a sensory panel. Both beer samples were mainly characterized by six aroma compounds (linalool, geraniol and ß-damascenone, citronellol, 2-phenylethyl acetate, and 2-phenylethanol), three of which were potentially responsible for the geographic origin of the hops given their significantly different concentrations.

Direct quantification of sulfur dioxide in wine by Surface Enhanced Raman Spectroscopy.

A rapid Surface Enhanced Raman Spectroscopy (SERS) method to detect SO2 in wine is presented, exploiting the preferential binding of silver nanoparticles (AgNPs) with sulfur-containing species. This interaction promotes the agglomeration of the AgNPs and inducing the formation of SERS "hot spots" responsible for SO2 signals enhancement. For increasing SO2 concentrations from 0 to100 mg/l in wine simulant, SERS intensity showed an increasing trend, following a Langmuir absorption function (R2 = 0.94). Due to the wine matrix variability, a standard additions method was then employed for quantitative analysis in red and white wines. This method does not require the SO2 separation but only a matrix pre-cleaning by solid phase extraction. The limit of detection (LOD) was defined for each wine tested, ranging from 0.6 mg/l to 9.6 mg/l. The results obtained were validated by comparison with the International Organization of Vine and Wine method (OIV-MA-AS323-04A).

Key norisoprenoid compounds in wines from early-harvested grapes in view of climate change.

In view of climate change, the scheduling of an early harvest may be an agronomic option to limit wine alcohol, provided that a satisfactory content of secondary metabolites can be ensured in grapes. To better understand the link between grape ripening, seasonal trend and wine aroma, the aromatic expression of Barbera and Pinot Noir wines produced with early harvested grapes was assessed. Attention was focused on C13 norisoprenoids during both alcoholic fermentation and after three months of storage. At the end of fermentation, the highest ß-damascenone content was detected in wines obtained from less ripe grapes, the content subsequently increased significantly after three months of storage; however, the levels of ß-ionone decreased significantly during the same period. The reduction of wine alcohol as a result of harvesting earlier, especially for Barbera, was associated with optimal aromatic levels as well as good technological parameters.

Impact of Increasing Levels of Oxygen Consumption on the Evolution of Color, Phenolic, and Volatile Compounds of Nebbiolo Wines.

Since the end of the last century, many works have been carried out to verify the effect of controlled oxygen intake on the chemical and organoleptic characteristics of red wines. In spite of the large number of studies on this subject, oxygen remains a cutting-edge research topic in oenology. Oxygen consumption leads to complex and not univocal changes in wine composition, sometimes positive such as color stabilization, softening of mouthfeel, increase of aroma complexity. However, the variability of these effects, which depend both on the oxygenation conditions and the composition of the wine, require more efforts in this research field to effectively manage wine oxygen exposure. The present study is focused on the evolution of the chemical composition of four different Nebbiolo wines, each of them added with 4 different doses of oxygen (7, 14, 21, and 28 mg/L total intake) during the first month of storage. In this perspective, the evolution over time of wine color and polyphenols was studied. Acetaldehyde, glyceraldehyde and glyoxylic acid were quantified by HPLC. These compounds can play a role in wine aging creating condensed colored and stable products involving anthocyanins with or without tannins. Moreover, some volatile aldehydes correlated with oxidized olfactory notes, including methional and (E)-2-alkenals, have been quantified by GC-MS. Overall, during storage a decrease of color intensity, total and free anthocyanins and an increase in polymeric pigments (in particular the contribution to the red color of pigments not-bleachable by SO2 or dTAT%) and some minor aldehydes was observed. Nevertheless, the differences in color parameters between the samples with different doses of oxygen were modest. These evidences were in contrast with an evident and detectable increase of free acetaldehyde content at increasing doses of oxygen measured after 60 days of storage. The effect of oxygen on color and production of SO2 non-bleachable pigments during aging varies with wine composition, with Nebbiolo wines appearing not very reactive in this respect, probably due to their low content in anthocyanins and high content in tannins.

Comparison of the physicochemical and volatile composition of wine fractions obtained by two different dealcoholization techniques.

Three wines (a Rosé wine, a Pelaverga and a Barbera red wines) were dealcoholized at 5% v/v ethanol with two different techniques: membrane contactor (MC) and distillation under vacuum (D). The influence of the treatments on the physicochemical composition and aromatic profile of the dealcoholized fractions was investigated. The main difference between the two techniques was the concentration grade reached by the dealcoholized fractions, which was 5-6times higher with D than with MC due to the concomitant loss of water. The main fixed compounds (organic acids, cations, polyphenols, anthocyanins) were not lost during the dealcoholization, while the losses of volatile compounds were relevant. Finally, the δ18O isotopic ratio was studied. The δ18O isotope ratio increased with D and decreased with MC. At the same time the δ18O isotope ratio fell within legal limits when the final ethanol content was adjusted to 2% by blending with dealcoholized fractions.

Influence of solvents on the composition of condensed tannins in grape pomace seed extracts.

The extracting effectiveness of some solvents (water, ethanol, acetone, ethyl acetate), used as pure or in binary and ternary mixtures, was studied for the extraction of seeds from the fermented pomace of Nebbiolo cv. The aqueous mixtures of acetone provided the highest extraction yields in total polyphenols and total flavonoids. Polyphenolic extracts were also obtained with aqueous mixtures of ethanol or acetone at variable concentrations, and the condensed tannins were quantified with the phloroglucinolysis method. The aqueous mixtures of acetone were more effective than the corresponding aqueous mixtures of ethanol. The solvent influenced the extraction yield and the composition of the extracts: with the increase of the yield, the mean degree of polymerization (mDP) of the condensed tannins increased. A significant correlation was noticed between mDP and the molar percentages of (+)-catechin as terminal unit (negative correlation), and of (-)-epicatechin and (-)-epicatechin-3O-gallate as extension units (positive correlation).

Grape aroma precursors in cv. Nebbiolo as affected by vine microclimate.

The influence exerted by bunch microclimate on some C13-norisoprenoid precursors content was investigated for the first time in Nebbiolo grapes during ripening. Samples were collected, during two consecutive seasons, from two vineyards, which are characterized by different microclimatic conditions caused by vine vigour heterogeneity and different vineyard aspects. Enzymatic hydrolysis of the glycosides extracted from the grapes, and subsequent GC-MS determination of the aglycones, highlighted that the majority of norisoprenoid glycosides accumulated in Nebbiolo berries from pre-veraison until 3-4weeks post-veraison. Vineyard aspect and vine vigour affected the timing of the maximum concentration of norisoprenoid precursors and their subsequent decrease at harvest. Low light in the vigorous blocks penalized norisoprenoids peak concentration. In the south less vigorous blocks, a decline of total norisoprenoids content during the pre-harvest period was observed. This decline appeared mainly regulated by the temperature. Vintage and/or microclimatic conditions affected the final content of some important norisoprenoids.

Validation of a rapid conductimetric test for the measurement of wine tartaric stability.

This work was aimed at optimizing a rapid and reproducible conductivity test for the evaluation of wine tartaric stability, in order to improve the practices for the prevention of tartaric precipitations during bottle aging. The test consists in measuring the drop of conductivity in wines kept under stirring for a fixed time, at low temperature, after the addition of micronized potassium bitartrate crystals (KHT). An experimental design was planned to study three factors affecting the test: temperature, duration and dose of added potassium bitartrate. A standard protocol was defined to produce a micronized potassium bitartrate starting from available commercial products, since the dimensions of the crystals can affect the final conductivity values. After the choice of the best conditions the method was validated. Two different stability thresholds were defined for white wines and for red/rosé wines by comparing the results of the mini-contact test with those of the cold test.

Indigenous Georgian Wine-Associated Yeasts and Grape Cultivars to Edit the Wine Quality in a Precision Oenology Perspective.

In Georgia, one of the most ancient vine-growing environment, the homemade production of wine is still very popular in every rural family and spontaneous fermentation of must, without addition of chemical preservatives, is the norm. The present work investigated the yeast biodiversity in five Georgian areas (Guria, Imereti, Kakheti, Kartli, Ratcha-Lechkhumi) sampling grapes and wines from 22 different native cultivars, in 26 vineyards and 19 family cellars. One hundred and eighty-two isolates were ascribed to 15 different species by PCR-ITS and RFLP, and partial sequencing of D1/D2 domain 26S rDNA gene. Metschnikowia pulcherrima (F' = 0.56, I' = 0.32), Hanseniaspora guilliermondii (F' = 0.49, I' = 0.27), and Cryptococcus flavescens (F' = 0.31, I' = 0.11) were the dominant yeasts found on grapes, whereas Saccharomyces cerevisiae showed the highest prevalence into wine samples. Seventy four isolates with fermentative potential were screened for oenological traits such as ethanol production, resistance to SO2, and acetic acid, glycerol and H2S production. Three yeast strains (Kluyveromyces marxianus UMY207, S. cerevisiae UMY255, Torulaspora delbrueckii UMY196) were selected and separately inoculated in vinifications experiments at a Georgian cellar. Musts were prepared from healthy grapes of local varieties, Goruli Mtsvane (white berry cultivar) and Saperavi (black berry cultivar). Physical (°Brix) and microbial analyses (plate counts) were performed to monitor the fermentative process. The isolation of indigenous S. cerevisiae yeasts beyond the inoculated strains indicated that a co-presence occurred during the vinification tests. Results from quantitative GC-FID analysis of volatile compounds revealed that the highest amount of fermentation flavors, such as 4-ethoxy-4-oxobutanoic acid (monoethyl succinate), 2-methylpropan-1-ol, ethyl 2-hydroxypropanoate, and 2-phenylethanol, were significantly more produced in fermentation conducted in Saperavi variety inoculated with K. marxianus, whereas other aromatic compounds like 3-methylbutyl acetate, ethyl hexanoate and dihydrofuran-2(3H)-one (γ- butyrolactone) showed a higher content in Goruli Mtsvane variety samples fermented by S. cerevisiae. The selected yeast strains have proved to be promising for enhancing the flavor potential in low aromatic Georgian cultivars. This work intends to be a knowledge contribution for a precision oenology toward the strategic concept of "one grape variety-one yeast".

Use of polyaspartate as inhibitor of tartaric precipitations in wines.

All additives used to stabilize wines against the precipitations of potassium bitartrate have some limits: metatartaric acid (MTA) is effective but very unstable, carboxymethylcellulose is stable and effective in white wines but affects color stability in red wines, mannoproteins have a variable effectiveness depending on wine composition. This work was aimed at testing the effect of new stabilizing products on tartaric precipitations, focusing on the use of Na and K polyaspartate salts (PASPs). The effectiveness of 4 different PASPs and 1 MTA added to red and white wines was compared using the mini-contact test and cold test. The dose effect and the stability of the products over time were also studied. The PASPs showed a similar stabilizing effect and a longer stability over time compared to MTA. PASPs can be considered interesting as additives for wine tartaric stabilization. Further work is in progress to better characterize their enological properties.

Influence of the matrix composition on the volatility and sensory perception of 4-ethylphenol and 4-ethylguaiacol in model wine solutions.

This study was aimed at investigating the influence of ethanol, wine polyphenols and yeast extract on the volatility of 4-ethylphenols in red wines. The use of solid-phase microextraction with short extraction times for the study of the headspace composition, revealed a strong influence of ethanol and polyphenols on reducing the volatility of 4-ethylphenols in a model wine. At the same time, the influence of ethanol and polyphenols on the perception of the "Brett character" (stable, manure, horse sweat and phenolic notes) due to 4-ethylphenols was evaluated by sensory analysis. Polyphenols and ethanol were found to have a significant influence on the olfactory perception of the tainted wines. On the contrary, a direct influence of yeast extract on the volatility of 4-ethylphenols was not observed, although some results suggested that the effectiveness of polyphenols in reducing the volatility of 4-ethylphenols could be limited by the presence of yeast proteins.

Effect of the closure type on the evolution of the physical-chemical and sensory characteristics of a Montepulciano d'Abruzzo Rosé wine.

The present work studied the effect of the kind of closure (a screw cap, a natural cork, and 2 synthetic closures) on the evolution of the oxygen content and on the physical-chemical and sensory characteristics of a Montepulciano d'Abruzzo rosé wine during the 1st 12 mo of bottle aging. The chemical analyses concerned the parameters more involved in the oxidative reactions (SO(2) , acetaldehyde, phenols, wine color), as well as the main fermentative volatile compounds. The kind of closure influenced the oxygen content in wines, free and total SO(2) concentration, and wine color (color intensity and hue). During bottle aging, free and total SO(2) concentration was significantly and negatively correlated with absorbance at 420 nm (A420), whereas the correlations with A520 were weak. Probably, the limited extent of the variations in red color (A520), when varying SO(2) concentration, were due to the low pH of this rosé wine. No effect of the kind of closure on phenols and the main fermentative volatile compounds was observed. The wines bottled with cork closures (N trials), after 12 mo of storage, had higher color intensity and hue, measured by spectrophotometry, and were visually distinguished from the other trials for the more intense pink reflections. On the whole, under the conditions of this work, all the used closures guaranteed a good preservability to the rosè wines during the 1st year of bottle aging, and the changes in composition did not significantly affect wine sensory characteristics. Therefore, these synthetic closures can represent an alternative to the cork closures for a medium to long term bottle aging of these wines.

Quantification by solid phase micro extraction and stable isotope dilution assay of norisoprenoid compounds in red wines obtained from Piedmont rare varieties.

A method to identify and quantify megastigmane norisoprenoid compounds in wines was developed using headspace solid phase micro extraction (SPME) coupled with gas-chromatography/mass-spectrometry (GC-MS). Three different compounds were quantified by stable isotope dilution assay (SIDA): ß-damascenone, ß-ionone and α-ionone. Particular attention was paid to maximising the method's sensitivity while reducing the extraction time. To optimise the extraction conditions, a statistically designed experiment was performed using extraction time, extraction temperature and ethanol content as operating variables. Five different SPME fibres suitable for the analysis of volatile compounds were compared. This study confirmed that the PDMS/DVB coating performs best for the quantification of ß-damascenone and ß-ionone, and the crucial influence of ethanol content of the sample on extraction effectiveness. Finally, the optimised method was applied to the study of various wines derived from rare and autochthonous grape varieties of north-western Italy.

Analytical and sensory characterization of the aroma of "Langhe D.O.C. Nebbiolo" wines: influence of the prefermentative cold maceration with dry ice.

The present work concerns the determination of the volatile compounds, as well as the description of the olfactory characteristics, of "Langhe DOC Nebbiolo" wines produced through the prefermentative cold maceration with dry ice, on full scale (30 quintals) in private cellars. The free volatile compounds and those obtained by enzymatic and chemical hydrolysis of the glycosilated precursors were analyzed by gas chromatography-mass spectrometry. Among free volatile compounds, a loss of alcohols with 6 carbon atoms in the cryo-macerated trials was observed. The cryo-maceration technique, when correctly applied, increased the floral and fruity notes (blackberry, cherry, and plum). This increase is related to the extraction of polyphenolic compounds (maceration effect), but no volatile compounds, neither in free form, nor produced by enzymatic hydrolysis of the glycosilated precursors, can discriminate the wines according to their content of polyphenols. Only some compounds produced by chemical hydrolysis of the precursors can discriminate the wines (statistically significant differences): 3-hydroxy-ß-damascone, methylvanillate, and blumenol C. These compounds, however, form during aging and do not have any influence on young wines aroma.