Formation of α-ketoglutaric acid by wine yeasts and its oenological significance

α-Ketoglutaric acid was measured enzymically in wines made in the laboratory from three grape varieties by pure cultures of 12 wine yeasts of the genus Saccharomyces. The results were confirmed with the same juices and 4 yeasts on pilot-plant scale in replicated 30 gallon lots. Mean values for the 12 yeasts ranged from 9 to 117 ppm (overall mean 53). In any one juice the yeasts differed by at least 10-fold in the amounts produced, and certain yeasts produced consistently high or low yields in all juices. The amounts of α-ketoglutaric acid produced depended somewhat on the grape juices used, even though these had comparable pH values, and a significant yeast-juice interaction occurred. The amount of α-ketoglutaric acid formed during fermentation at 15° was 60 per cent of that formed at 25°, and over twice as much was formed at pH 4.2 as at pH 3.0, using four yeast strains. Formation of α-ketoglutaric and pyruvic acids were not significantly correlated. The α-ketoglutaric acid content of 18 white table wines made under comparable conditions on pilot-plant scale from different grape varieties using the same yeast strain ranged from 38 to 152 ppm (mean 90). The significance of the results is discussed, particularly in relation to the binding of sulphur dioxide in wine, and recommendations are given on how to make wines which are low in α-ketoglutaric acid. Formation of α-ketoglutaric acid by three yeasts in a chemically defined medium was lower with increased amounts of nitrogen as ammonium sulphate and higher in the presence of L-glutamic acid, both being used separately as sole nitrogen sources. These findings are discussed in relation to the rǒle of α-ketoglutaric acid in nitrogen metabolism of yeasts.     

Fermentation of grape must by sequential association of yeasts: Accumulation of pyruvic and α-ketoglutaric acid

Fermentations of grape (cv. Malvar) musts from two consecutive vintages were carried out using the autochthonous microflora, a sequential association of yeasts and conventional fermentations with addition of sulfur dioxide to the must. The pyruvic and α-ketoglutaric acid content over the course of fermentation was measured and showed that for both vintages tested the maximum accumulation of the ketoacid pyruvic acid took place several days earlier in fermentations using a sequential association of yeasts than in conventional fermentations. The accumulation of pyruvic acid was higher in the must made from grapes with a higher degree of ripening and the lowest level of added SO₂. In the fermentations using either a sequential association of yeasts or the autochthonous microflora with no added SO₂, accumulation of α-ketoglutaric acid was higher in the must with the higher nitrogen content when the species making the greatest percentage contributions at the start of fermentation presented high levels of proteolytic activity.     

Influence of yeast strain and pH on pyruvic acid content of wines

Pyruvic acid content of wines was investigated on laboratory and pilot-plant scale and found to be strongly influenced by the strain of yeast used to conduct the fermentation, and pH of the medium, with high pH correlating with high pyruvic acid content. Results obtained with artificial medium confirmed those obtained with grape juice. Yeast strain and pH appear to be the two most important factors influencing pyruvic acid content of wines, and their biochemical and oenological significance is discussed.     

Fermentation of sulphite-free white musts with added lysozyme and oenological tannins: Nitrogen consumption and biogenic amines composition of final wines

The influence on amino acid consumption and biogenic amines composition of white wines obtained by replacing SO₂ during fermentation with lysozyme and tannins was studied. At the same time, the fermentative performance of two low SO₂ producing selected yeasts strains was evaluated. For this purpose, a series of laboratory-scale fermentations of fresh white must and a HPLC-DAD method for the analysis of amino acids, biogenic amines and ammonium ion were undertaken. The presence of SO₂ or lysozyme affected the consumption of nitrogen as a function of the yeast strain, while oenological tannin had no substantial influence. Strain 1042 increased the consumption of total YAN in the presence of SO₂, as a consequence of the enhanced utilization of ammonium ion and a number of amino acids. By contrast, strain 333 tended not to change the total YAN uptake, whatever the juice treatment, and reduced the consumption of aspartic and glutamic acids, GABA and other compounds in the case of samples added with SO₂. When compared with lysozyme addition, for both strains, SO₂ increased the consumption of alanine and glutamine, the latter being a major contributor to the assimilable nitrogen of the must. No influence of must treatments was found on the content of biogenic amines in the final wines.     

Efficacy of ultraviolet radiation as an alternative technology to inactivate microorganisms in grape juices and wines

Since sulphur dioxide (SO₂) is associated with health risks, the wine industry endeavours to reduce SO₂ levels in wines with new innovative techniques. The aim of this study was, therefore, to investigate the efficacy of ultraviolet radiation (UV)-C (254 nm) as an alternative technology to inactivate microorganisms in grape juices and wines.A pilot-scale UV-C technology (SurePure, South Africa) consisting of an UV-C germicidal lamp (100 W output; 30 W UV-C output) was used to apply UV-C dosages ranging from 0 to 3672 J l⁻¹, at a constant flow rate of 4000 l h⁻¹ (Re > 7500). Yeasts, lactic and acetic acid bacteria were singly and co-inoculated into 20 l batches of Chenin blanc juice, Shiraz juice, Chardonnay wine and Pinotage wine, respectively. A dosage of 3672 J l⁻¹, resulted in an average log₁₀ microbial reduction of 4.97 and 4.89 in Chardonnay and Pinotage, respectively. In Chenin blanc and Shiraz juice, an average log₁₀ reduction of 4.48 and 4.25 was obtained, respectively.UV-C efficacy may be influenced by liquid properties such as colour and turbidity. These results had clearly indicated significant (p < 0.05) germicidal effect against wine-specific microorganisms; hence, UV-C radiation may stabilize grape juice and wine microbiologically in conjunction with reduced SO₂ levels.     

Effect of the Winemaking Practices and Aging on Phenolic Content of Smederevka and Chardonnay Wines

Changes of the polyphenolic content, including total phenolics (TP), total flavonoids (TF), and total flavan-3-ols (TF_3-ols) determined by spectrophotometric methods, have been studied in Macedonian white wines from Vitis vinifera L. cv, Smederevka and Chardonnay, obtained under different vinifications and analyzed during 16 months of aging in bottles. Winemaking treatments for both varieties included an addition of two doses of SO₂ (50 and 100 mg/L) and two commercial yeasts for fermentation, Vinalco and Levuline, both Saccharomyces cerevisiae species, in order to study the influence of SO₂ and yeasts on the phenolic content of the wines. Comparing the two varieties, Chardonnay wines contained higher levels of total phenolics, flavonoids, and flavan-3-ols, probably as a result of the cultivar differences. Sulfur dioxide protected the phenolic oxidation, resulting in higher phenolic content in the wines with higher SO₂ dose, while the yeast did not show significant influence on the total phenolics. During aging, the TP, TF, and TF_3-ols in the wines from both varieties decreased significantly up to the second month of storage and remained stable till the sixth month. The Student–Newman–Keuls test has been applied to ascertain possible significant differences between the studied wines, and a principal component analysis has been employed, showing separation and grouping of the wines according to the cultivar and time of aging.     

New perspectives in safety and quality enhancement of wine through selection of yeasts based on the parietal adsorption activity

The present article aims to review research papers that focus on the parietal adsorption activity of wine yeast and on its contribution to the enhancement of wine safety and quality. There is a great diversity among yeasts for their parietal adsorption activity: the outermost layer of the cell wall has a chemical composition that notably varies from yeast to yeast. Parietal mannoproteins can contain phosphate, pyruvate, or glucuronic acid, which impart negative charges, modifying the electrostatic and ionic interactions with wine components. The following could give a good reason to propose a specific selection of wine yeasts based on their parietal adsorption activity to improve wine safety and quality: (a) ochratoxin A content of wines is greatly reduced by expressly selected yeasts, sequestering the toxin during winemaking; (b) yeast influences concentration and composition of phenolic compounds in wine, above all by adsorbing them on cell wall; (c) among grape pigments, anthocyanins, in particular, may be adsorbed by yeast cell wall; and (d) yeast can also interact with wine colour producing anthocyanin-beta-d-glucosidase, pyruvic acid, and acetaldehyde or releasing mannoproteins and different polysaccharides. Genomic strategies could also be used to obtain a further enhancement of the adsorption/non-adsorption activity of wine yeasts. Based on winemaking requirements and on parietal adsorption activity, a specific selection of yeasts might be performed: (a) to protect wine colour during red winemaking, (b) to remove residual wine colour during white winemaking, (c) to selectively remove ochratoxin A, and (d) to protect phenolic compounds responsible for antioxidant activity.     

Metabolism of SO₂ binding compounds by Oenococcus oeni during and after malolactic fermentation in white wine

Sulfur dioxide SO? is the key additive for the preservation of wines. Carbonyl and keto compounds in wine can bind to SO? and decrease its efficacy, resulting in higher total SO? requirements. Increased consumer demand for low sulfite and organic wines pose production challenges if SO? binders have not been properly managed during vinification. Malolactic fermentation (MLF) has been known to reduce bound SO? levels but detailed time course studies are not available. In this work, the kinetics of major SO? binding compounds and malic acid were followed during MLF in wine with 12 commercially available strains of Oenococcus oeni. Pyruvic acid, acetaldehyde and α-ketoglutaric acid were degraded to various degrees by O. oeni, but galacturonic acid was not. At the time of malic acid depletion, percent degradation of pyruvate, α-ketoglutaric acid and acetaldehyde was 49%, 14% and 30%, respectively. During MLF, the decrease in average bound SO? levels, as calculated from carbonyl metabolism, was 22%. The largest reduction in wine carbonyl content occurred in the week after completion of MLF and was 53% (107 mg/L to 34 mg/L) calculated as bound SO?. Prolonged activity of bacteria in the wines (up to 3 weeks post malic acid depletion) resulted only in reduced additional reductions in bound SO? levels. The results suggest that microbiological wine stabilization one week after malic acid depletion is an effective strategy for maximum removal of SO? binders while reducing the risk of possible post-ML spoilage by O. oeni leading to the production acetic acid and biogenic amines.     

Reducing the sulfur-dioxide binding power of sweet white wines by solid-phase extraction

The high sulfur-dioxide binding power of sweet white wines may be reduced by extracting the naturally present carbonyl compounds from wine that are responsible for carbonyl bisulphites formation. The carbonyl compounds mainly responsible for trapping SO₂ are acetaldehyde, pyruvic acid, and 2-oxoglutaric acid. The method employed was selective solid phase extraction, using phenylsulfonylhydrazine as a scavenging agent. The scavenging function was grafted onto a support prepared from raw materials derived from lignin. This approach is more acceptable to winemakers than the polymer media previously reported, as it reduces the possible contamination of wine to molecules already present in the wine making process.     

Influence of different yeasts on the growth of lactic acid bacteria in wine

The influence of various yeasts on the growth of lactic acid bacteria in wine was tested by inoculating Lactobacillus hilgardii, L. brevis and two strains of Leuconostoc mesenteroides into experimental wines made with twelve different yeasts of the genus Saccharomyces. Wines made from juice which had been infected with several spoilage yeasts and then fermented with a wine yeast were also tested in this way. It was found that the yeasts differed considerably in their effects on bacterial growth. In some of the experimental wines bacterial growth was delayed or failed altogether. Generally, the unfavourable influence of any yeast on bacterial growth was much reduced if the wines were left in contact with the yeast cells for some weeks after the fermentation. The significance of these results in relation to the occurrence of malo-lactic fermentation in commercial wineries is discussed.     

Sulphite-binding power of wines and ciders. I. Equilibrium constants for the dissociation of carbonyl bisulphite compounds

The apparent Equilibrium Constants were determined at pH3 and 4 for the dissociation of the bisulphite compounds of acetaldehyde, pyruvic acid, 2-ketoglutaric acid, L -xylosone, D -threo-2,5-hexodiulose, 2,5-diketogluconic acid, 2-ketogluconic acid and galacturonic acid. Determinations were also made over a wider pH range for acetaldehyde (pH 1 to 13) and pyruvic acid (pH 1.5 to 8).     

Sweet Wine Production by Two Osmotolerant Saccharomyces cerevisiae Strains

The use of Saccharomyces cerevisiae to produce sweet wine is difficult because yeast is affected by a hyperosmotic stress due to the high sugar concentrations in the fermenting must. One possible alternative could be the coimmobilization of the osmotolerant yeast strains S. cerevisiae X4 and X5 on Penicillium chrysogenum strain H3 (GRAS) for the partial fermentation of raisin musts. This immobilized has been, namely, as yeast biocapsules. Traditional sweet wine (that is, without fermentation of the must) and must partially fermented by free yeast cells were also used for comparison. Partially fermented sweet wines showed higher concentration of the volatile compounds than traditionally produced wines. The wines obtained by immobilized yeast cells reached minor concentrations of major alcohols than wines by free cells. The consumption of specific nitrogen compounds was dependent on yeast strain and the cellular immobilization. A principal component analysis shows that the compounds related to the response to osmotic stress (glycerol, acetaldehyde, acetoin, and butanediol) clearly differentiate the wines obtained with free yeasts but not the wines obtained with immobilized yeasts.     

Multifactorial analysis of acetaldehyde kinetics during alcoholic fermentation by Saccharomyces cerevisiae

Acetaldehyde is the terminal electron acceptor in the alcoholic fermentation by Saccharomyces cerevisiae. Quantitatively the most important carbonyl by-product, it has relevance for ethanol production yields as well as product stabilization and toxicology. The aim of this study was to investigate the effect of various enological parameters on acetaldehyde kinetics during alcoholic fermentations. Two commercial yeast strains were tested in two grape musts and the pH, temperature, SO₂ and nutrient addition were varied. All incubations had uniform kinetics where acetaldehyde reached an initial peak value followed by partial reutilization. Peak acetaldehyde concentrations and residual concentrations after 15 days of fermentations ranged from 62 to 119 mg l^− 1 and 22 to 49 mg l^− 1, respectively. A positive linear relationship was found between peak and final acetaldehyde levels in Gewürztraminer, but not Sauvignon Blanc fermentations, where sluggish fermentations were observed. Several factors had a significant effect on peak and/or final acetaldehyde levels. SO₂ addition, grape cultivar and fermentation nutrition were important regulators of peak acetaldehyde production, while final acetaldehyde concentrations were correlated with SO₂ addition, grape cultivar and temperature. The results allowed to estimate the acetaldehyde increase caused by SO₂ addition to 366 ??g of acetaldehyde per mg of SO₂ added to the must. The course of the final fermentation phase was shown to determine acetaldehyde residues. Comparison of acetaldehyde and hexose kinetics revealed a possible relationship between the time of occurrence of peak acetaldehyde concentrations and the divergence of glucose and fructose degradation rates.     

Influence of SO<Subscript>2</Subscript> on the evolution of volatile compounds through alcoholic fermentation of must stabilized by pulsed electric fields

The aim of this work was to study the influence of sulphur dioxide (SO₂) on the formation of volatile compounds by yeast through wine alcoholic fermentation. Thus Parellada must was microbiologically stabilized using a pulsed electric field (PEF) treatment and inoculated with Saccharomyces cerevisiae Na33 strain. Fermentation was carried out with or without SO₂ and the results showed that the evolution of the volatile compounds profile throughout the process was similar. The content of volatile acids in wine obtained by using sulphur dioxide was not significantly different from that fermented without adding the compound. However, the final content of total alcohols and esters was significantly different even thought the differences were small. Consequently, when grape must is treated by PEF the sulphur dioxide concentration usually used in winemaking could be reduced to safer levels or even eliminated without an important effect on the volatile compounds content of the final product. Therefore, the absence of sulphur dioxide should not have a negative impact on the sensory characteristics of wine.     

7种酵母酿造摩尔多瓦葡萄酒的发酵特性比较

在常用的商业酵母中筛选一株酵母,应用于摩尔多瓦葡萄酒酿造,从而实现鲜食摩尔多瓦葡萄的增值。以摩尔多瓦葡萄为材料,选用VL1、F15、NS-D、P、HXD29、LE28、LD1015 7种酵母进行葡萄酒酿造试验,以自然发酵作为对照,对不同酵母发酵酒的总酸、总酚等理化性质进行检测和感官评价,比较不同酵母对摩尔多瓦葡萄酒理化性质及品质的影响。结果表明,酵母LD1015发酵的摩尔多瓦酒干浸物含量最高,为17.96 g/L;总糖含量为3.54 g/L,总酚含量为1.17 g/L,略低于酵母VL1、HXD29、LE28,感官评分最高,为91分。综合判定酵母LD1015具有较强的糖转化和干物质浸提的能力,有利于产出口感醇厚、香气馥郁的葡萄酒,是7种酵母中最适合鲜食葡萄摩尔多瓦酿酒的酵母菌种。     

Selection and modification of yeasts and lactic acid bacteria for wine fermentation

Historically, the fermentation of grape juice to wine has been carried out by indigenous yeasts found on the berry. However, in newer wine regions, e.g. the USA, inoculation with selected wine yeast strains is employed. Grape juice is high in nutritional factors and difficulties in fermentation usually arise from the inhibitory effects of the high concentration of sugar initially present and the ethanol produced. A secondary fermentation, brought about by indigenous or added lactic acid bacteria, converts malic acid to lactic acid and carbon dioxide and often occurs. This ‘malolactic’ fermentation is usually slow. For both yeast and bacterial fermentations strain selection is based more on fermentation performance than on sensory characteristics of the wine, with increased tolerance of the yeast to ethanol and of the bacteria to low pH being emphasized. Attempts to increase the malolactic fermentation rate have been made by cloning and transferring the malolactic gene from Lactobacillus to wine yeast. In early attempts to produce wines with enhanced or novel sensory characteristics a leucine-less mutant of a homothallic wine yeast has been obtained which does not produce isoamyl alcohol.     

Formation of higher alcohols by wine yeasts,and relationship to taste thresholds

The amounts of n-propanol, iso-butanol and iso-amyl plus active amyl alcohol produced during fermentation of grape juice were found to vary considerably according to the yeast used. The yeasts studied included 11 wine yeasts and one brewery yeast, all belonging to the genus Saccharomyces, and 4 yeasts belonging to genera which sometimes cause wine spoilage. Experiments were carried out in the laboratory and confirmed on pilot-plant scale. The average production of n-propanol in juices from 4 grape varieties varied from 13 to 106 ppm depending on the wine yeast used. The corresponding variations in the production of iso-butanol and isoamyl plus active amyl alcohol were 9 to 37 ppm and 115 to 262 ppm respectively,. Juices from different varieties' of grapes (Vitis vinifera) differed somewhat in the amounts of higher alcohols formed, irrespective of the yeast strain used. An increase in temperature of fermentation from 15° to 25° with four yeasts produced an average of 24% more iso-amyl plus active alcohol and 39% more iso-butanol, and 17% less n-propanol. A yeast/temperature interaction occurred. An increase in pH from 3.0 to 4.2 with four yeasts produced 28% more iso-amyl plus active amyl alcohol, 85% more iso-butanol, and 11% more n-propanol. A yeast /pH interaction occurred. Spoilage yeasts examined also formed higher alcohols and the amounts were not related to sugar consumed during the fermentation. Taste thresholds of the higher alcohols, with the exception of active amyl alcohol, were measured in dry white wine with 7 tasters. Iso-amyl alcohol thresholds ranged from 100 to 900 ppm (mean 300 ppm) and iso-butanol and n-propanol both exceeded 500 ppm. The average threshold of iso-amyl alcohol in distilled water was 4 ppm. For certain tasters strains of yeast can produce sufficient differences in iso-amyl alcohol to be detectable organoleptically, but differences in iso-butanol and n-propanol are not large enough to be detectable.     

Effect of refermentation conditions and micro-oxygenation on the reduction of volatile acidity by commercial S. cerevisiae strains and their impact on the aromatic profile of wines

Herein, we evaluate the applicability of previously characterized commercial and indigenous Saccharomyces cerevisiae strains and non-S. cerevisiae species for the deacidification of white and red wines at a pilot scale. The effect of the refermentation process (mixture of acidic wine with musts from freshly crushed grapes or with residual marc) as well as micro-oxygenation (MO) on acetic acid removal efficiency and wine aromatic composition was also assessed in a red wine. The commercial strains S26 and S29 efficiently reduced both acetic acid (43 and 47%, respectively) and sugar (100%) after 264 h of refermentation of an acidic white wine that was supplemented with grape must. Similar results (60-66% of acetic acid removal) were observed for red wine deacidification using grape must, independently of MO. When residual marc was used for deacidification, strain S26 removed 40% of acetic acid, whereas strain S29 did not initiate refermentation with or without MO. Wines obtained by refermentation with the must had significantly lower acetic acid and a higher total SO₂ concentration in comparison to the wines deacidified by the grape marcs. The volatile aroma compound's composition of deacidified red wines was dependent on the refermentation process used, rather than on MO. The marc-deacidified wine obtained by the use of strain S26 and without MO achieved the best sensory classification. When data from all analytical and sensory evaluation were combined, Principal Component Analysis (PCA) separated the wines into three distinct groups according to the strain and the refermentation process independently of MO. We successfully established an efficient and cheap enological solution for the rectification of volatile acidity of wines.     

Replacement of sulfur dioxide by lysozyme and oenological tannins during fermentation: influence on volatile composition of white wines

BACKGROUND: In recent years the use of sulfur dioxide, a commonly used additive in winemaking, has been questioned because of its toxic effects on human health. Studies have been conducted to find alternatives that can effectively substitute for this additive in all its various technological functions. In previous work, lysozyme and oenological tannins were found as possible substitutes in controlling bacterial undesirable fermentations and phenolic oxidation. However, data on the volatile composition of wines obtained by that protocol are lacking. In this work, the effects on volatile composition of white wines by the substitution of SO₂ during fermentation with lysozyme and tannin were studied. At the same time, the technological performance of two strains of yeast that produce low amounts of SO₂ were evaluated. RESULTS: The results showed that both SO₂ and lysozyme prevented the development of undesirable bacterial fermentations. The study of volatile compounds shows differences in the alcohol, acid and ester contents among the final products: wines fermented with strain 1042 and lysozyme had higher total alcohol concentration, while the addition of SO₂ promoted higher production of 3‐methyl‐1‐butanol, 3‐methylthio‐1‐propanol, phenylethyl alcohol and 4‐hydroxy‐benzenethanol. Esters, as a total, were influenced by the different strain and tannins addition, while amounts of medium‐chain fatty acid ethyl esters and their corresponding fatty acids were found in higher amounts in wines coming from fermentations with lysozyme. The sensory analysis revealed a preference for wines to which lysozyme and tannins had been added. CONCLUSION: The data suggest that the addition of lysozyme and oenological tannins during alcoholic fermentation could represent a promising alternative to the use of SO₂ and for the production of wines with reduced content of SO₂. The composition of the volatiles in the final wines was affected by the different vinification protocols (mainly with regards to alcohols and ethyl esters). Copyright © 2009 Society of Chemical Industry