Inoculated Yeast Strain Research Articles

Evaluation and selection of yeasts as potential aroma enhancers for the production of dry-cured ham

Yeasts play a critical role in the flavor formation of dry-cured ham. In this study, 41 yeast isolates from the dry-cured ham at different processing stages were evaluated for their technological properties. Debaryomyces hansenii was the most dominant yeast and has been detected at each phase of dry-cured ham, followed by Candida zeylanoides which was mainly detected in salting phase. Yarrowia bubula and Yarrowia alimentaria were found at the first two-phase of dry-cured ham. All isolates of yeast showed enzymatic activities against milk protein and tributyrin, while only 4 strains displayed proteolytic activity on meat protein. Yeast strains were grown in a meat model medium and volatile compounds were identified. The result showed that inoculated yeast strains could promote the production of volatiles and there were significant differences among strains. D. hansenii S25 showed the highest production of volatile compounds, followed by the strain C. zeylanoides C4. D. hansenii S25 was the highest producer of alcohols showing the highest production of benzeneethanol and 3-(methylthio)-1-propanol. Based on OAV and PLS analysis, D. hansenii S25 was strongly correlated with overall flavor and key volatile compounds of dry-cured ham, which could be selected as potential starter cultures.

Wine chemical, sensory, aroma compound and protein analysis of wines produced from chemical and biological fungicide treated Chenin blanc grapes

Fungal diseases in vineyards are one of the main causes leading to economic losses within the viticulture sector and are continuously increasing with time. The most common of these fungal diseases are powdery mildew, downy mildew and grey mould which are controlled by the use of fungicides. However, fungicide residues can alter the fermentation process and prevent some biochemical pathways of yeast metabolism involved in phenolic and/or aroma compound production that are critical for sensory quality. In this study, Chenin blanc grapes treated with chemical and natural fungicides were used to produce small-scale wines and laboratory-scale fermentations. Treatments consisted of a control (an approved chemical used in industry), a 1x and 2x treatment (new formulations). Laboratory-scale fermentations were conducted in duplicate using the commercial Saccharomyces cerevisiae (S. cerevisiae) Active Dry Wine Yeast (ADWY) strains VIN 13 and VIN 7. The fermentations were monitored by weighing frequently until they stabilised (CO2 weight loss). Small-scale wines were made according to the standard Nietvoorbij experimental wine making procedure. At the end of fermentation, lees samples were subjected to CHEF gel electrophoreses to confirm that the inoculated yeast strain completed the fermentations. Moreover, fermenting wine samples, collected at the start and end of the fermentation, were subjected to protein extraction, quantification and characterisation the yeast proteins expressed at the various stages of fermentation. Resultant wines were subject to chemical and sensory evaluations five months after bottling. From the results obtained in the above study, it was concluded that the inoculated yeast strains used for wine making completed the fermentations at a similar rate to their respective controls (registered fungicide). In addition, small-scale cellar fermentations showed that the fungicide treatments (1x the recommended dosage [Treatment 1] and 2x the recommended dosage [Treatment 2]) compared to the controls had no notable negative effects on wine aroma and sensory profiles although differences were observed in the proteins expressed after the fermentation. The chemical and biological fungicides had no notable effects on the wine making procedure and quality of resulting wines compared to the registered fungicide.

Anthocyanins enhance yeast’s adsorption of Ochratoxin A during the alcoholic fermentation

Ochratoxin A (OTA) is a mycotoxin that can be present in food products and beverages such as wine, in which the European Union has regulated its maximum concentration. Since yeasts can adsorb OTA during the alcoholic fermentation (AF), we have analyzed whether anthocyanins influence the reduction of OTA in wine during AF carried out by two Saccharomyces cerevisiae strains. Laboratory experiments were conducted with one white must and two red musts that were obtained by adding to the white must two different concentrations of anthocyanins extract. Musts were supplemented with OTA and tested at two pHs. A reduction of OTA was observed in all wines, being higher in red wines. These data point that a chemical bond between anthocyanins and OTA is probably responsible for enhanced adsorption of OTA by yeast. On the other hand, the removal of OTA between the two inoculated yeast strains was different. This result cannot be only attributed to differences in the yeasts’ cell wall composition, because differences were not detected in white wines.

Specific Phenotypic Traits of Starmerella bacillaris Related to Nitrogen Source Consumption and Central Carbon Metabolite Production during Wine Fermentation.

Over the last few years, the potential of non-Saccharomyces yeasts to improve the sensory quality of wine has been well recognized. In particular, the use of Starmerella bacillaris in mixed fermentations with Saccharomyces cerevisiae was reported as an appropriate way to enhance glycerol formation and reduce ethanol production. However, during sequential fermentation, many factors, such as the inoculation timing, strain combination, and physical and biochemical interactions, can affect yeast growth, the fermentation process, and/or metabolite synthesis. Among them, the availability of yeast-assimilable nitrogen (YAN), due to its role in the control of growth and fermentation, has been identified as a key parameter. Consequently, a comprehensive understanding of the metabolic specificities and the nitrogen requirements would be valuable to better exploit the potential of Starm. bacillaris during wine fermentation. In this study, marked differences in the consumption of the total and individual nitrogen sources were registered between the two species, while the two Starm. bacillaris strains generally behaved uniformly. Starm. bacillaris strains are differentiated by their preferential uptake of ammonium compared with amino acids that are poorly assimilated or even produced (alanine). Otherwise, the non-Saccharomyces yeast exhibits low activity through the acetaldehyde pathway, which triggers an important redistribution of fluxes through the central carbon metabolic network. In particular, the formation of metabolites deriving from the two glycolytic intermediates glyceraldehyde-3-phosphate and pyruvate is substantially increased during fermentations by Starm. bacillaris This knowledge will be useful to better control the fermentation process in mixed fermentation with Starm. bacillaris and S. cerevisiaeIMPORTANCE Mixed fermentations using a controlled inoculation of Starmerella bacillaris and Saccharomyces cerevisiae starter cultures represent a feasible way to modulate wine composition that takes advantage of both the phenotypic specificities of the non-Saccharomyces strain and the ability of S. cerevisiae to complete wine fermentation. However, according to the composition of grape juices, the consumption by Starm. bacillaris of nutrients, in particular of nitrogen sources, during the first stages of the process may result in depletions that further limit the growth of S. cerevisiae and lead to stuck or sluggish fermentations. Consequently, understanding the preferences of non-Saccharomyces yeasts for the nitrogen sources available in grape must together with their phenotypic specificities is essential for an efficient implementation of sequential wine fermentations with Starm. bacillaris and S. cerevisiae species. The results of our study demonstrate a clear preference for ammonium compared to amino acids for the non-Saccharomyces species. This finding underlines the importance of nitrogen sources, which modulate the functional characteristics of inoculated yeast strains to better control the fermentation process and product quality.

Correlation between Low Temperature Adaptation and Oxidative Stress in Saccharomyces cerevisiae.

Many factors, such as must composition, juice clarification, fermentation temperature, or inoculated yeast strain, strongly affect the alcoholic fermentation and aromatic profile of wine. As fermentation temperature is effectively controlled by the wine industry, low-temperature fermentation (10–15°C) is becoming more prevalent in order to produce white and “rosé” wines with more pronounced aromatic profiles. Elucidating the response to cold in Saccharomyces cerevisiae is of paramount importance for the selection or genetic improvement of wine strains. Previous research has shown the strong implication of oxidative stress response in adaptation to low temperature during the fermentation process. Here we aimed first to quantify the correlation between recovery after shock with different oxidants and cold, and then to detect the key genes involved in cold adaptation that belong to sulfur assimilation, peroxiredoxins, glutathione-glutaredoxins, and thioredoxins pathways. To do so, we analyzed the growth of knockouts from the EUROSCARF collection S. cerevisiae BY4743 strain at low and optimal temperatures. The growth rate of these knockouts, compared with the control, enabled us to identify the genes involved, which were also deleted and validated as key genes in the background of two commercial wine strains with a divergent phenotype in their low-temperature growth. We identified three genes, AHP1, MUP1, and URM1, whose deletion strongly impaired low-temperature growth.

Study of Saccharomyces cerevisiae wine strains for breeding through fermentation efficiency and tetrad analysis.

One of the issues that most concerns to both winemakers and producers of active dry yeasts is the stuck and sluggish fermentations of grape musts with high levels of sugar, reflecting the inability of inoculated yeast strain to complete the fermentation process. It is difficult to obtain a wine strain that possesses both adequate oenological and technological properties; thus, the correct approach to solving these problems is the application of breeding programs primarily focused on both properties. The first step toward this process is to characterize the phenotypic diversity between potential parental strains. In the present study, we have analyzed the fermentative behavior of 26 Saccharomyces cerevisiae wine strains in high-sugar conditions at 20°C, using a range of tests, such as sporulation ability, spore viability, and tetrad analysis to determine the tolerance of these yeasts to several stress conditions. Most tested strains were homothallic and heterozygous for more than one character. Two auxotrophic derivatives with defects in amino acid or nucleic acid metabolism were obtained, and these strains could potentially be used for the development of hybridization techniques without using laboratory strains.

Effect of fat and salt reduction on the sensory quality of slow fermented sausages inoculated with Debaryomyces hansenii yeast

The inoculation of a Debaryomyces hansenii strain in dry fermented sausages with reduced fat and salt contents was evaluated in terms of chemical, microbial and consumer acceptability. The implantation of the inoculated yeast strain was confirmed by RAPDs of M13 minisatellite. A reduction of 17–20% salt and 10–16% fat content was achieved. These reductions affected the sausage quality by producing an increase in aw, hardness and chewiness values and a decrease of staphylococci growth. However, D. hansenii inoculation compensated these changes although it was not able to modify neither the hardness of reduced fat batches nor the staphylococci growth decrease. In terms of sensory acceptability, different preferences patterns of consumers were found. Yeast inoculation improved the aroma and taste quality when fat or salt reductions were carried out in dry fermented sausages.

Impact of Debaryomyces hansenii strains inoculation on the quality of slow dry-cured fermented sausages

Debaryomyces hansenii strains, M4 and P2, isolated from natural fermented sausages were inoculated in slow fermented sausages to study their effect on processing parameters, microbial population, volatile compound and sensory characteristics. The inoculation of D. hansenii strains, M4 and P2, did not affect the ripening process as no differences in pH and Aw were detected. The dominance of the inoculated yeast strains along the process was followed by RAPDs of M13 minisatellite. The inoculated yeasts, P2 and M4, were recovered at the end of the ripening process although P2 appeared in higher counts than M4. The sausages inoculated with P2 resulted in a decrease in lipid oxidation values (TBARS) and a reduction of lipid-oxidation derived aldehydes in addition to a highest acid compound abundance. M4 inoculated sausages resulted in highest sulphur containing compound abundance. However, no differences in consumer acceptance were detected. Moreover, both yeast strains were responsible for the generation of ethyl methyl-branched ester compounds in the dry-cured sausages.

Generation of aroma compounds in a fermented sausage meat model system by Debaryomyces hansenii strains

The ability of seven Debaryomyces hansenii strains to generate aroma compounds in a fermented sausage model system was evaluated. The presence of the yeast, in the inoculated models, was confirmed by PCR amplification of M13 minisatellite. Volatile compounds production was analysed using Solid Phase Micro-Extraction and gas chromatography/mass spectrometry. Forty volatile compounds were detected, quantified and their odour activity values (OAVs) calculated. All volatile compounds increased during time in the inoculated models although significant differences were found amongst them. Ester and sulphur production was strongly dependent on the strain inoculated. D. hansenii P2 and M6 strains were the highest producers of sulphur compounds where dimethyl disulphide and dimethyl trisulfide were the most prominent aroma components identified by their OAVs whereas, M4 showed the highest OAVs for ester compounds followed by the P2 strain. The meat model system has been useful to show the real ability of yeast strains to produce aroma compounds.

Malolactic fermentation of Tempranillo wine: contribution of the lactic acid bacteria inoculation to sensory quality and chemical composition

Malolactic fermentation (MLF) of Tempranillo Rioja wines (Spain) inoculated with two lactic acid bacteria (LAB) strains were studied and compared with spontaneous MLF. Inoculation with selected Oenococcus oeni lyophila shortened MLF duration up to 19days and lead to wines with more fresh and fruity characters, especially when implantation was 100%. We demonstrated modifications in the concentration of volatile and nitrogenous compounds and a good correlation between analytical and sensory attributes was also noted. In addition, the low initial amino acid concentration and the consumption of these compounds by the inoculated yeast strain during alcoholic fermentation resulted in wines with very low biogenic amines levels (under 3.75mgL -1) after MLF and 3month storage period in all cases. The results showed the significance of choose the most suitable starter to elaborate quality wines and suggest the control of amino acid content in must and wine to prevent the formation of biogenic amines. © 2011 The Authors. International Journal of Food Science and Technology © 2011 Institute of Food Science and Technology.

Using RFLP-mtDNA for the rapid monitoring of the dominant inoculated yeast strain in industrial wine fermentations

The analysis of restriction fragment length polymorphism of mitochondrial DNA (mtDNA-RFLP) has been applied as a test to monitor the abundance of the starter yeast strain during industrial wine fermentations without previous isolation of yeast colonies. For white wine fermentations, we performed a rapid assay consisting in taking a sample of fermenting must, purifying the DNA from harvested cells, and obtaining the restriction patterns by digestion with the endonuclease HinfI. The same protocol, but adding an overnight cultivation step before DNA purification, was also applied to red wine fermentations. The results were compared with those obtained from the subsequent characterisation of strains, for the same samples, by analysis of the electrophoretic karyotype of isolated yeast colonies. In all cases, when the inoculated strain was dominant within the yeast population, the rapid assay anticipated the result by showing the coincidence between the restriction profiles obtained from both total cells and the inoculated strain. The results were obtained at 11 or 23 h after sampling for white- or red-wine fermentations respectively. This method allows a rapid intervention of the wine-producer if the presence of the inoculated yeasts has suffered a sudden decrease in any phase of the fermentation process.

Features ofSaccharomyces cerevisiaeas a culture starter for the production of the distilled sugar cane beverage, cachaça in Brazil

To evaluate the dominance and persistence of strains of Saccharomyces cerevisiae during the process of sugar cane fermentation for the production of cachaça and to analyse the microbial compounds produced in each fermentative process. Three S. cerevisiae strains were evaluated during seven consecutive 24-h fermentation batches using recycled inocula. The UFLA CA 116 strain had the largest population of viable organisms, and the maximum population was achieved in the fourth batch after 96 h of fermentation. The UFLA CA 1162 and UFLA CA 1183 strains grew more slowly, and the maximum population was reached in the seventh batch. Molecular characterization of isolated yeast cells using PFGE (pulse field gel electrophoresis) revealed that more than 86% of the isolates corresponded to the initially inoculated yeast strain. The concentration of aldehydes, esters, methanol, alcohol and volatile acids in the final-aged beverages were within the legal limits. Cachaça produced by select yeast strains exhibits analytical differences. UFLA CA 1162 and UFLA CA 116 S. cerevisiae isolates can be considered the ideal strains for the artisanal production of cachaça in Brazil. The use of select yeast strains can improve the quality and productivity of cachaça production. Our findings are important for the appropriate monitoring of yeast during sugar cane fermentation. In addition, we demonstrate that UFLA CA 116 and UFLA CA 1162, the ideal yeast strains for cachaça production, are maintained at a high population density. The persistence of these yeast strains in the fermentation of sugar cane juice promotes environmental conditions that prevent or decrease bacterial contamination. Thus, the use of select yeast strains for the production of cachaça is a viable economic alternative to standardize the production of this beverage.

Influence of Saccharomyces cerevisiae yeast strain on the major volatile compounds of wine

As must fermentation is often conducted with selected yeasts, it is important to determine the influence of the inoculated yeast strain on the wine organoleptic properties. We analysed the production of major volatile compounds (acetaldehyde, ethyl acetate and some fusel alcohols) during fermentations of musts from different grape cultivars and conducted with selected Saccharomyces cerevisiae yeast strains. The production of these volatile compounds was variable, and depended mostly on the must composition and the fermentation conditions. Occasionally, the amount of certain volatiles compounds in the wine depended on the yeast strain. We observed inverse correlations between acetaldehyde and isobutanol production, and, between ethyl acetate and total fusel alcohol production. Also, a direct correlation was found between the organoleptic wine quality and the amount of ethyl acetate. The most appreciated wines were made with yeast strains that did not produce high amounts of any of the analysed compounds, while the lowest quality wines were made with yeast strains that produced high amounts of acetaldehyde and fusel alcohols.

Yeast starter cultures affecting wine fermentation and volatiles

The influence of the addition of different concentrated inocula of various strains of Saccahromyces cerevisiae to high sugar content must on both evolution of sugar concentration and the volatile composition of the wines were studied. Evolution of sugar content was characteristic of the yeast strain and one was incapable of finishing fermentation. Volatile compounds were analyzed by using fused silica capillary GC column. Application of cluster analysis to the data showed that differences in the volatile fraction are mainly due to the inoculated yeast strain, although concentration of higher alcohols increases directly as the inoculum concentration.