Livio Lencioni

Selected non-Saccharomyces wine yeasts in controlled multistarter fermentations with Saccharomyces cerevisiae

Non-Saccharomyces yeasts are metabolically active during spontaneous and inoculated must fermentations, and by producing a plethora of by-products, they can contribute to the definition of the wine aroma. Thus, use of Saccharomyces and non-Saccharomyces yeasts as mixed starter cultures for inoculation of wine fermentations is of increasing interest for quality enhancement and improved complexity of wines. We initially characterized 34 non-Saccharomyces yeasts of the genera Candida, Lachancea (Kluyveromyces), Metschnikowia and Torulaspora, and evaluated their enological potential. This confirmed that non-Saccharomyces yeasts from wine-related environments represent a rich sink of unexplored biodiversity for the winemaking industry. From these, we selected four non-Saccharomyces yeasts to combine with starter cultures of Saccharomyces cerevisiae in mixed fermentation trials. The kinetics of growth and fermentation, and the analytical profiles of the wines produced indicate that these non-Saccharomyces strains can be used with S. cerevisiae starter cultures to increase polysaccharide, glycerol and volatile compound production, to reduce volatile acidity, and to increase or reduce the total acidity of the final wines, depending on yeast species and inoculum ratio used. The overall effects of the non-Saccharomyces yeasts on fermentation and wine quality were strictly dependent on the Saccharomyces/non-Saccharomyces inoculum ratio that mimicked the differences of fermentation conditions (natural or simultaneous inoculated fermentation).

Outlining a future for non-Saccharomyces yeasts: selection of putative spoilage wine strains to be used in association with Saccharomyces cerevisiae for grape juice fermentation.

The use of non-Saccharomyces yeasts that are generally considered as spoilage yeasts, in association with Saccharomyces cerevisiae for grape must fermentation was here evaluated. Analysis of the main oenological characteristics of pure cultures of 55 yeasts belonging to the genera Hanseniaspora, Pichia, Saccharomycodes and Zygosaccharomyces revealed wide biodiversity within each genus. Moreover, many of these non-Saccharomyces strains had interesting oenological properties in terms of fermentation purity, and ethanol and secondary metabolite production. The use of four non-Saccharomyces yeasts (one per genus) in mixed cultures with a commercial S. cerevisiae strain at different S. cerevisiae/non-Saccharomyces inoculum ratios was investigated. This revealed that most of the compounds normally produced at high concentrations by pure cultures of non-Saccharomyces, and which are considered detrimental to wine quality, do not reach threshold taste levels in these mixed fermentations. On the other hand, the analytical profiles of the wines produced by these mixed cultures indicated that depending on the yeast species and the S. cerevisiae/non-Saccharomyces inoculum ratio, these non-Saccharomyces yeasts can be used to increase production of polysaccharides and to modulate the final concentrations of acetic acid and volatile compounds, such as ethyl acetate, phenyl-ethyl acetate, 2-phenyl ethanol, and 2-methyl 1-butanol.

Potential spoilage non-Saccharomyces yeasts in mixed cultures with Saccharomyces cerevisiae

With the aim of exploring the possibility to improve wine quality through the utilization of wine-related yeasts generally considered as spoilage, mixed cultures of Saccharomyces cerevisiae with Hanseniaspora osmophila, Pichia fermentans, Saccharomycodes ludwigii and Zygosaccharomyces bailii were inoculated in grape juice. All the fermentations got to completion and most of the compounds normally produced at high concentrations by pure cultures of non-Saccharomyces yeasts, and considered detrimental for wine quality, did not reach the threshold taste level in mixed fermentations with S. cerevisiae. Interestingly, the association of S. cerevisiae with P. fermentans, S. ludwigii and Z. bailii produced significant increases in the production of polysaccharides as compared to pure cultures of S. cerevisiae. Since polysaccharides improve wine taste and body, and exert positive effects on aroma persistence and protein and tartrate stability, a possible use for these yeasts can be envisaged in mixed starter cultures with S. cerevisiae for the enhancement of the final quality of wine.

Controlled mixed fermentation at winery scale using Zygotorulaspora florentina and Saccharomyces cerevisiae.

Over the last few years the use of multi-starter inocula has become an attractive biotechnological practice in the search for wine with high flavour complexity or distinctive characters. This has been possible through exploiting the particular oenological features of some non-Saccharomyces yeast strains, and the effects that derive from their specific interactions with Saccharomyces. In the present study, we evaluated the selected strain Zygotorulaspora florentina (formerly Zygosaccharomyces florentinus) in mixed culture fermentations with Saccharomyces cerevisiae, from the laboratory scale to the winery scale. The scale-up fermentation and substrate composition (i.e., white or red musts) influenced the analytical composition of the mixed fermentation. At the laboratory scale, mixed fermentation with Z. florentina exhibited an enhancement of polysaccharides and 2-phenylethanol content and a reduction of volatile acidity. At the winery scale, different fermentation characteristics of Z. florentina were observed. Using Sangiovese red grape juice, sequential fermentation trials showed a significantly higher concentration of glycerol and esters while the sensorial analysis of the resulting wines showed higher floral notes and lower perception of astringency. To our knowledge, this is the first time that this yeasts association has been evaluated at the winery scale indicating the potential use of this mixed culture in red grape varieties.

Zygotorulaspora florentina and Starmerella bacillaris in multistarter fermentation with Saccharomyces cerevisiae to reduce volatile acidity of high sugar musts

Background and Aims The possibility to decrease wine volatile acidity (VA) is an important aspect in wine production. This applies in particular to wines that are produced from musts with high sugar concentration, where the osmotic pressure promotes an increase in acetic acid production by Saccharomyces cerevisiae. This study aimed to identify suitable yeast strains and fermentation temperature to undertake the alcoholic fermentation of high sugar musts. Methods and Results To lower VA during fermentation of high sugar musts, two non‐Saccharomyces yeast strains, Zygotorulaspora florentina and Starmerella bacillaris, were used in multistarter fermentations with S. cerevisiae at a fermentation temperature of 14 and 20°C. The fermentation temperature influenced the yeast behaviour and the composition of the two mixed fermentations. Conclusions Independent of fermentation temperature, the mixed fermentations with Z. florentina performed best to reduce VA. Significance of the Study Mixed fermentations with the non‐Saccharomyces yeast strains Z. florentina and S. bacillaris may represent a valuable approach for the fermentation of high sugar musts.

Evaluation of the Yeast Schizosaccharomyces japonicus for Use in Wine Production

Schizosaccharomyces japonicus UCD2489 was evaluated for potential use as a starter culture in wine-making. Laboratory-scale fermentations of Trebbiano grape juice were set up to compare fermentation kinetics of pure cultures of Sch. japonicus, immobilized Sch. japonicus cells, a commercial strain of Saccharomyces cerevisiae (EC1118), and mixed cultures of both species. The fermentation kinetics of the sequential and coinoculated fermentations were largely driven by the presence of S. cerevisiae. UCD2489 cell immobilization resulted in a significant reduction in ethanol levels in mixed fermentations compared with EC1118. Acetic acid levels similar to those of EC1118 in pure culture were produced when fermentations were coinoculated. The ability of UCD2489 to consume malic acid was adversely affected by EC1118, particularly in the coinoculated fermentation, suggesting that acid levels could be manipulated by adjusting the relative ratios of the two yeasts and the timing of inoculation with S. cerevisiae. Depending upon the inoculation conditions used, Sch. japonicus produced a quantity of glycerol ~2-fold higher than those released by S. cerevisiae. The analyses of volatile compounds showed increases in aroma-impacting compounds such as ethyl acetate in all Sch. japonicus wines, and acetaldehyde in the free-cell coinoculated fermentation that exceed reported sensory thresholds for these compounds, and for other important aroma compounds such as isoamyl acetate, hexyl acetate, phenyl ethyl acetate, ethyl isobutyrate, and ethyl butyrate. Polysaccharide release by UCD2489 was ~4.7-fold greater than that of S. cerevisiae alone. Reduction of induced wine protein haze was correlated with the concentration of polysaccharides. Our findings suggest that Sch. japonicus could be useful in wine production to reduce acidity and final ethanol levels and to increase glycerol, volatile compounds, and active polysaccharides with potential beneficial enhancement of protein stability.