Wine Musts Research Articles

Impact of high lipid contents on the production of fermentative aromas during white wine fermentation.

In Cognac, the musts are rich in grape solids and fermentations are usually run with turbidities ranging between 500 and 1500 NTU (nephelometric turbidity unit). These conditions, considered favourable for generating the desired organoleptic profiles of the final Eaux-de-vies, are unusual in winemaking, and, consequently, their impact on yeast metabolism is poorly understood. This study aims to better describe and understand the synthesis of fermentative aromas in such lipid-excess conditions, while integrating the effect of two other very important parameters: the initial concentration of assimilable nitrogen and the temperature of fermentation. To reach this objective, a Box-Behnken design was implemented to describe and model the simple effects of these factors as well as their interactions. Although the lipid concentration was very high, impacts on the production of fermentative aromas were observed. Indeed, high lipid levels promoted the synthesis of higher alcohols. Observing this effect was surprising because there is no metabolic connection between the anabolic pathways of production of these alcohols and the lipid pathway. This effect may be partly explained by impairment in the activity of alcohol acetyl transferases in the presence of lipids, which catalyse the conversion of higher alcohols into the corresponding esters. Therefore, in this study, the negative impact of turbidity was very significant on acetate esters related to the production of acetyl-CoA, which was the main molecule disturbed by the strong presence of lipids. Finally, and more surprisingly, lipid intake did not impact the synthesis of ethyl esters, which depended on the concentration of exogenous lipids. KEY POINTS: • Innovative work on the fermentation of white wine musts with very high lipid contents. • Precise fermentation management and monitoring in Cognac-making conditions. • Experimental design to study the impact of lipids, assimilable nitrogen and temperature on fermentative aroma synthesis.

Influence of Autochthonous and Commercial Yeast Strains on Fermentation and Quality of Wines Produced from Vranec and Cabernet Sauvignon Grape Varieties from Tikveš Wine-Growing Region, Republic of North Macedonia

Two autochthonous yeast strains called F-8 and F-78 (isolated and selected from the Tikveš wine-producing region) were inoculated in wine musts from Vranec and Cabernet Sauvignon grape varieties. The fermentation process and quality of the produced wines were compared to the wines produced from the same grape varieties, but with a commercial yeast strain (D-80). The fermentation process was undertaken at 23–25 °C for 16 days. The highest alcohol content was detected in Vranec and Cabernet Sauvignon wines fermented with autochthonous F-8 yeast strain. Conversely, Vranec wine fermented by the F-78 yeast strain contained the lowest alcohol content (14.11%). The low sugar amount (2–3 g/L) in both grape varieties is indicated the production of dry wines. The total amount of phenolic compounds was significantly higher in Vranec wines. Unlike the alcohol content, Vranec wine produced by commercial yeast strain D-80 indicated the highest concentration of total phenolic compounds (1450 mg/L) and total anthocyanins (572 mg/L), while the lowest concentrations were observed in wine fermented by autochthonous yeast strain F-78 (1612 mg/L and 470 mg/L, respectively). The maximum color intensity was measured in the Vranec wine fermented by autochthonous yeast strain F-8, while wines from Cabernet Sauvignon grape variety fermented with the same yeast strain F-8 and commercial yeast strain D-80 had comparable values in terms of color intensity.

Influence of Autochthonous and Commercial Yeast Strains on Fermentation and Quality of Wines Produced from Vranec and Cabernet Sauvignon Grape Varieties from Tikveš Wine-Growing Region, Republic of North Macedonia

Two autochthonous yeast strains called F-8 and F-78 (isolated and selected from the Tikves wine-producing region) were inoculated in wine musts from Vranec and Cabernet Sauvignon grape varieties. The fermentation process and quality of the produced wines were compared to the wines produced from the same grape varieties, but with a commercial yeast strain (D-80). The fermentation process was undertaken at 23–25 °C for 16 days. The highest alcohol content was detected in Vranec and Cabernet Sauvignon wines fermented with autochthonous F-8 yeast strain. Conversely, Vranec wine fermented by the F-78 yeast strain contained the lowest alcohol content (14.11%). The low sugar amount (2–3 g/L) in both grape varieties is indicated the production of dry wines. The total amount of phenolic compounds was significantly higher in Vranec wines. Unlike the alcohol content, Vranec wine produced by commercial yeast strain D-80 indicated the highest concentration of total phenolic compounds (1450 mg/L) and total anthocyanins (572 mg/L), while the lowest concentrations were observed in wine fermented by autochthonous yeast strain F-78 (1612 mg/L and 470 mg/L, respectively). The maximum color intensity was measured in the Vranec wine fermented by autochthonous yeast strain F-8, while wines from Cabernet Sauvignon grape variety fermented with the same yeast strain F-8 and commercial yeast strain D-80 had comparable values in terms of color intensity.

Biocontrol Ability and Action Mechanism of Starmerella bacillaris (Synonym Candida zemplinina) Isolated from Wine Musts against Gray Mold Disease Agent Botrytis cinerea on Grape and Their Effects on Alcoholic Fermentation.

Gray mold is one of the most important diseases of grapevine in temperate climates. This plant pathogen affects plant growth and reduces wine quality. The use of yeasts as biocontrol agents to apply in the vineyard have been investigated in recent years as an alternative to agrochemicals. In this work, fermenting musts obtained from overripe grape berries, therefore more susceptible to infection by fungal pathogens such as Botrytis cinerea, were considered for the selection of yeasts carrying antifungal activity. Thirty-six isolates were identified as Starmerella bacillaris, a species recently proven to be of enological interest. Among them 14 different strains were studied and antifungal activity against B. cinerea was demonstrated, for the first time, to be present in S. bacillaris species. The production of volatile organic compounds (VOCs), tested in vitro, was found to be the main responsible of S. bacillaris antifungal effects. All the strains were able to reduce B. cinerea decay on wounded grape berries artificially inoculated with gray mold. The colonization level of wound was very high reaching, after 5 days, a concentration of 106 cells per ml of grape juice obtained after berry crushing. At this cell concentration S. bacillaris strains were used to ferment synthetic and natural musts. The sequential yeast inoculation, performed by adding S. cerevisiae 48 h after S. bacillaris, was needed to complete sugar consumption and determined a significant increase in glicerol content and a reduction of ethanol and acetic acid concentrations. The high wound colonization ability, found in this work, together with the propensity to colonize grape berry and the interesting enological traits possessed by the selected S. bacillaris strains allow the use of this yeast as biocontrol agent on vine and grape berries with possible positive effects on must fermentation, although the presence of S. cerevisiae is needed to complete the fermentation process. This work introduces new possibilities in wine yeast selection programs in order to identify innovative wine yeasts that are simultaneously antifungal agents in vineyards and alternative wine starters for grape must fermentation and open new perspective to a more integrated strategy for increasing wine quality.

Wine fermentation microbiome: a landscape from different Portuguese wine appellations.

Grapes and wine musts harbor a complex microbiome, which plays a crucial role in wine fermentation as it impacts on wine flavour and, consequently, on its final quality and value. Unveiling the microbiome and its dynamics, and understanding the ecological factors that explain such biodiversity, has been a challenge to oenology. In this work, we tackle this using a metagenomics approach to describe the natural microbial communities, both fungal and bacterial microorganisms, associated with spontaneous wine fermentations. For this, the wine microbiome, from six Portuguese wine appellations, was fully characterized as regards to three stages of fermentation – Initial Musts (IM), and Start and End of alcoholic fermentations (SF and EF, respectively). The wine fermentation process revealed a higher impact on fungal populations when compared with bacterial communities, and the fermentation evolution clearly caused a loss of the environmental microorganisms. Furthermore, significant differences (p < 0.05) were found in the fungal populations between IM, SF, and EF, and in the bacterial population between IM and SF. Fungal communities were characterized by either the presence of environmental microorganisms and phytopathogens in the IM, or yeasts associated with alcoholic fermentations in wine must samples as Saccharomyces and non-Saccharomyces yeasts (as Lachancea, Metschnikowia, Hanseniaspora, Hyphopichia, Sporothrix, Candida, and Schizosaccharomyces). Among bacterial communities, the most abundant family was Enterobacteriaceae; though families of species associated with the production of lactic acid (Lactobacillaceae, Leuconostocaceae) and acetic acid (Acetobacteriaceae) were also detected. Interestingly, a biogeographical correlation for both fungal and bacterial communities was identified between wine appellations at IM suggesting that each wine region contains specific and embedded microbial communities which may contribute to the uniqueness of regional wines.

Analysis of non-Saccharomycesyeast populations isolated from grape musts from Sicily (Italy)

The aim of this study was to identify the non-Saccharomyces yeast populations present in the grape must microflora from wineries from different areas around the island of Sicily. Yeasts identification was conducted on 2575 colonies isolated from six musts, characterized using Wallerstein Laboratory (WL) nutrient agar, restriction analysis of the amplified 5.8S-internal transcribed spacer region and restriction profiles of amplified 26S rDNA. In those colonies, we identified 11 different yeast species originating from wine musts from two different geographical areas of the island of Sicily. We isolated non-Saccharomyces yeasts and described the microflora in grape musts from different areas of Sicily. Moreover, we discovered two new colony morphologies for yeasts on WL agar never previously described. This investigation is a first step in understanding the distribution of non-Saccharomyces yeasts in grape musts from Sicily. The contribution is important as a tool for monitoring the microflora in grape musts and for establishing a new non-Saccharomyces yeast collection; in the future, this collection will be used for understanding the significance of these yeasts in oenology.

Microbiological and Sensory Effects of Musts Treated by High-pressure Homogenization

The winemaking sector needs to ensure the microbiological quality of its products to guarantee acceptance and wide commercialization. There are treatments such as high pressure homogenization (HPH) that, apparently, do not affect the final nutritional and sensory food properties. This study is presented to evaluate possibilities of utilizing HPH treatments to reduce the indigenous flora accompanying wine musts, as well as their effects on the fermentative process, oenological parameters, color, aroma, and taste properties. Two different must varieties were used: a white must (Parellada variety) and a red one (Trepat variety). Results showed that the use of HPH at 200 MPa is capable of reducing the microbial load of musts. Residual populations of total bacteria were detected, but neither fungi, nor yeasts, nor lactic acid bacteria were detected after the treatment in either musts. Furthermore, as a result of the decrease of the wild microbiota of the musts, the implantation of the selected yeast for alcoholic fermentation was improved. Sensory assessments of the must and wines showed that there were no significant changes caused by the treatment.

Recovery of Wine-Must Aroma Compounds by Supercritical CO2

The fractionation of wine musts (the fermentation broth that leads from grape juice to wine) of highly aromatic grape varieties was performed with supercritical carbon dioxide at pressures between 11 and 18 MPa and temperatures between 308 and 318 K. Different configurations of the separation devices were tested after the extraction step to improve the fractionation process. Headspace chromatography and Karl Fisher analyses were used to measure the composition of the extracts. A significant enrichment of the supercritical extracts on aromatic compounds was observed. The recovery of the aromatic compounds from the high pressure solvent stream poses technical problems that must be solved before the process can be feasible in an industrial scale.

Application of fluorescence in situ hybridisation (FISH) to the analysis of yeast population dynamics in winery and laboratory grape must fermentations

To analyse the yeast population diversity during wine fermentations, specific fluorescein-labelled oligonucleotide probes targeted to the D1/D2 region of the 26S rRNA of different yeast species known to occur frequently in this environment were designed and tested with reference strains. The probes were then used to identify wine must isolates and to follow, in combination with plate counts, the evolution of yeast populations in two winery fermentations of white and red grape musts. In both cases, a high diversity of non- Saccharomyces yeast species was detected, including Candida stellata, Hanseniaspora uvarum, H. guilliermondii, Kluyveromyces marxianus, K. thermotolerans and Torulaspora delbrueckii. Some of these species (e.g., K. marxianus, K. thermotolerans and T. delbrueckii) were present in significant amounts during the tumultuous fermentation stage, despite the predominance of Saccharomyces cerevisiae cells following the inoculation of the wine musts with a starter strain. To further clarify the yeast population dynamics at the late phase of the fermentations, and because winery conditions do not allow a reliable control of experimental variables, strains isolated from the industrial musts were used to conduct two laboratory microvinifications in synthetic grape juice, using different ratios of S. cerevisiae/non- Saccharomyces in the inocula. Under these conditions, the results were similar to those obtained in the winery, showing a yeast profile with mixed species throughout the first fermentation stage, i.e. until about 40–50% of the total sugar was consumed. Non- Saccharomyces yeasts were outgrown by S. cerevisiae only after ethanol reached concentrations around 4–5% ( v / v), which argues in favour of a potential important role of non- Saccharomyces in the final organoleptic characteristics of the wine.

Utilization of capillary electrophoresis/mass spectrometry (CE/MSn) for the study of anthocyanin dyes

Hyphenation of capillary electrophoresis with electrospray ionization mass spectrometry was utilized for the monitoring of anthocyanins in wine and wine musts. CE/MS was performed in two electrolytes: 1) an acidic one (chloroacetate-ammonium, pH 2) and 2) a basic one with high selectivity towards derivatives containing vicinal hydroxy groups (borate-ammonium, pH 9). The setup of MS was optimized and the fragmentation of common anthocyanins was studied in detail. Attention was also focused on the fragmentation of anthocyanidin skeleton. The anthocyanidins substituted with hydroxy groups fragment via a cascade of neutral losses of water and carbon monoxide. Fragmentation of anthocyanidins containing a methoxy group on their B-ring starts with the cleavage of methane and/or methyl radical. The optimized method was utilized for the monitoring of changes in anthocyanin profile in red wines as well as the process of release of anthocyanins to wine must.

Biomass content governs fermentation rate in nitrogen-deficient wine musts.

Problematic fermentations are common in the wine industry. Assimilable nitrogen deficiency is the most prevalent cause of sluggish fermentations and can reduce fermentation rates significantly. A lack of nitrogen diminishes a yeast's metabolic activity, as well as the biomass yield, although it has not been clear which of these two interdependent factors is more significant in sluggish fermentations. Under winemaking conditions with different initial nitrogen concentrations, metabolic flux analysis was used to isolate the effects. We quantified yeast physiology and identified key metabolic fluxes. We also performed cell concentration experiments to establish how biomass yield affects the fermentation rate. Intracellular analysis showed that trehalose accumulation, which is highly correlated with ethanol production, could be responsible for sustaining cell viability in nitrogen-poor musts independent of the initial assimilable nitrogen content. Other than the higher initial maintenance costs in sluggish fermentations, the main difference between normal and sluggish fermentations was that the metabolic flux distributions in nitrogen-deficient cultures revealed that the specific sugar uptake rate was substantially lower. The results of cell concentration experiments, however, showed that in spite of lower sugar uptake, adding biomass from sluggish cultures not only reduced the time to finish a problematic fermentation but also was less likely to affect the quality of the resulting wine as it did not alter the chemistry of the must.

Bioelectrochemical determination of lactic and malic acids in wine

Electroanalytical methods based on biosensor technology and differential pH measurement for the determination of l-lactic and l-malic acids in wine were developed. Lactic acid was determined according to two procedures consisting of the use of two lactate electrochemical biosensors, one based on a Clark O 2 probe and the other based on the H 2O 2 probe. The two probes were assembled employing a polymeric membrane where the enzyme lactate oxidase was previously immobilized. Both sensors exhibited high storage and operational stabilities, and good reproducibility when used in wine matrix. Measurement of lactate was carried out using two different flow-through cells assembled with the lactate probes. The low detection limit of these probes and the relatively high concentration of lactate in wine samples required a serial dilution in the range 1:100 to 1:200, thus eliminating all potential electrochemical or enzyme interferences present in the sample. Malic acid was determined using a differential commercially available pH-meter and an enzymatic procedure. For this analysis, we employed the malic enzyme (EC 1.1.1.40) and the NADP + cofactor. The pH variation due to the action of this enzyme on malic acid was found to be proportional to the malic acid present in the sample. Experimental parameters such as pH, temperature, and co-factor concentration were optimized, resulting in malate determination in less than 1 min with good reproducibility. Various samples of wine and wine musts were assayed for lactate with the two biosensor procedures, and for malate with differential pH-metry. The results when compared with those obtained with the commonly used spectrophotometric procedure correlated well.

The use of a coulter counter to study the alcoholic fermentation in enological conditions

A Coulter counter was used to rapidly determine the exact stage of alcoholic fermentation of wine musts on a growth and mortality curve. By this technique it was also possible to check the yeast population involved in this fermentation and detect assimilable nitrogen deficiencies of musts.

Rheology of wine musts during fermentation

The rheological behaviour of white grape must during fermentation has been studied at 14, 18 and 22°C and is described by an exponential function relating the coefficient of viscosity with the temperature and density of the must during fermentation (and, by implication, with the residual content of fermentable sugars).

Ethanol tolerance in yeasts.

It is now certain that the inherent ethanol tolerance of the Saccharomyces strain used is not the prime factor regulating the level of ethanol that can be produced in a high sugar brewing, wine, sake, or distillery fermentation. In fact, in terms of the maximum concentration that these yeasts can produce under batch (16 to 17% [v/v]) or fed-batch conditions, there is clearly no difference in ethanol tolerance. This is not to say, however, that under defined conditions there is no difference in ethanol tolerance among different Saccharomyces yeasts. This property, although a genetic determinant, is clearly influenced by many factors (carbohydrate level, wort nutrition, temperature, osmotic pressure/water activity, and substrate concentration), and each yeast strain reacts to each factor differently. This will indeed lead to differences in measured tolerance. Thus, it is extremely important that each of these be taken into consideration when determining "tolerance" for a particular set of fermentation conditions. The manner in which each alcohol-related industry has evolved is now known to have played a major role in determining traditional thinking on ethanol tolerance in Saccharomyces yeasts. It is interesting to speculate on how different our thinking on ethanol tolerance would be today if sake fermentations had not evolved with successive mashing and simultaneous saccharification and fermentation of rice carbohydrate, if distillers' worts were clarified prior to fermentation but brewers' wort were not, and if grape skins with their associated unsaturated lipids had not been an integral part of red wine musts. The time is now ripe for ethanol-related industries to take advantage of these findings to improve the economies of production. In the authors' opinion, breweries could produce higher alcohol beers if oxygenation (leading to unsaturated lipids) and "usable" nitrogen source levels were increased in high gravity worts. White wine fermentations could also, if desired, match the higher ethanol levels in red wines if oxygenation (to provide the unsaturated lipids deleted in part by the removal of the grape skins) were practiced and if care were given to assimilable nitrogen concentrations. This would hold true even at 10 to 14 degrees C, and the more rapid fermentations would maximize utilization of winery tankage.(ABSTRACT TRUNCATED AT 400 WORDS)

Dansyl Amino-Acids Behavior on a Radial Pak C18 Column. Derivatization of Grape Wine Musts, Wines, and Wine Vinegars

Abstract Investigation was carried out into the retention behaviour of dansyl amino acids with the concentration of the organic modifier, the pH and the ionic strenght of the eluting buffer. The quantitative result precision obtained using gradients of polarity and the stability of dansyl amino acids have been calculated. Finally, the dansylation conditions of grape wine musts, wines and wine vinegars are fixed.

The Influence of Malolactic Strain on the Fermentation on Wine Quality of Three Eastern Red Wine Grape Cultivars

Malolactic fermentation was induced in red wine musts of the grape cultivars Marechal Foch, deChaunac and Chancellor by using the <i>Leuconostoc oenos</i> strains ML-34, PSU-1 and LS-5A. The total acidity of the wines subjected to the malolactic fermentation was lower and the pH slightly higher than in the control wines. Among the 12 nonvolatile acids quantitatively determined, malic acid practically disappeared in the wines which had undergone malolactic fermentation, while an equivalent amount of lactic acid appeared. ML-34 was the slowest among the three bacterial strains in completing this fermentation, while the cultivar Chancellor was the easiest to be fermented by all malolactic strains. Malolactic strains also influenced sensory ranking of the wines. LS-5A and PSU-1 treated wines were generally ranked superior to ML-34 or control treated wines.

Acid Treatment of Red Table Wine Musts for Color Retention

Color retention during aging was improved by adding acid to musts. Quality was also improved at certain levels of added acid. Of the three acids used, tartaric was best from the quality standpoint, sulfuric best for color retention, and fumaric undesirable in this instance because of its biological instability and undesirable formation of off odors. Other chemical changes were noted.