Mixed starter yeasts as a biotechnological tool to produce wine with low alcoholic content

Recently due to climatic changes a progressive increase in the percentage of ethanol in wines was observed. In fact, the ripening phase of the grapes tends to coincide with hot periods resulting in high accumulations of sugars and production of wines characterized by excessive alcoholic content, with negative notes on the quality of the final product. At the same time, national and international markets request well-structured and full-bodied wines, characterized by high aromatic complexity and moderate alcohol content. Alternative approaches have been proposed for the reduction of alcohol content, including viticultural, pre-fermentation, and post-fermentation practices. The microbiological strategies proposed for reducing the alcohol content include the use of genetically modified yeasts or the use of non-conventional wine yeasts belonging to non-Saccharomyces group, selected from the microflora usually present on the grapes and/or in the cellar. This technique, appreciated by producers and consumers, is based on the development of controlled multi-starter fermentations that combine the use of these non-Saccharomyces yeasts with S. cerevisiae. The enological interest in this group of yeasts is correlated to lower yield in ethanol than Saccharomyces as the non-Saccharomyces yeasts utilize alternative metabolic pathways during the fermentation of the grape must sugars, and to their ability to release aromas, proteins and other active sensory compounds that are not produced by single fermentation with Saccharomyces cerevisiae. These characteristics allow improving the aromatic profile of the wine, reducing at the same time its alcohol content. However, some species can produce high concentrations of some secondary compounds which can have undesirable effects on the aromatic quality of the wine, such as ethyl acetate, and high levels of volatile acidity. Therefore, it is necessary to make a careful selection of the species and the inoculation methods to be used. In this thesis we proposed the use of non-Saccharomyces wine yeasts in mixed fermentations to select the most promising combination of strains to be used as a tool to reduce the ethanol content in wines. In the first phase, 29 non-Saccharomyces strains belonging to the species Hanseniaspora guilliermondii/osmophila, Torulaspora delbrueckii, Metschnikowia pulcherrima, Pichia fermentans, Saccharomycodes ludwigii, were tested for parameters of technological interest, such as production level of hydrogen sulphide (H2S), resistance to ethanol, SO2 and copper, evaluation of β-glucosidase activity. The strains selected in this first screening phase were tested in mixed fermentations at laboratory scale, using different inoculation protocols (simultaneous and sequential inoculation). The mixed starter culture, characterized by suitable oenological properties and able of reducing the ethanol content in the produced wine, was tested in different vinification conditions at laboratory scale using yeast as dried form and as immobilized cells in microcapsules. The aim of this step was to evaluate the influence of the starter formulation on the yeast behavior during the fermentation process. In the last research phase, the mixed starter culture selected during laboratory trials was validated at pilot scale in the cellar to identify the optimal fermentation conditions to be proposed to the winemakers for production of wine with a reduced alcohol content and good aromatic complexity