Abstract:
During fermentation Saccharomyces cerevisiae metabolises the nutrients present in the surrounding environment and secretes metabolic products. Therefore, its metabolism together with the substrate composition play an important role in determining the characteristics of the final fermented products. For example, during Sauvignon blanc wine making, a polyunsaturated fatty acid present in the grape juice, linoleic acid, affects the development of aroma compounds and other properties of the wine fermented by S. cerevisiae. However, only one single S. cerevisiae strain was used to ferment the wine and it is not known if linoleic acid has a similar effect on other strains. Moreover, previous research focused on the impact of linoleic acid on the wine profile rather than on the cellular pathways. For these reasons, the effect of linoleic acid on the overall cell metabolism is still not clear. This project focused on unlocking the metabolic response of S. cerevisiae to linoleic acid during wine making and in a laboratory-controlled environment using metabolomics and lipidomics platforms. These approaches provided an overall idea of how linoleic acid affects the wine fermented by different wine yeast strains (S. cerevisiae EC1118, AWRI796 and VIN13) and which cellular pathways were involved. Firstly, I investigated the effect of linoleic acid on the development of aroma compounds and other metabolites of Sauvignon blanc wines. Linoleic acid clearly affected the levels of acetylated aroma compounds, several amino acids, and antioxidant molecules, independent of the yeast strain used for fermentation. The analysis of the resulting wines provided an indirect evidence of the linoleic acid effect on S. cerevisiae without clarifying which yeast metabolic pathways were affected. In order to investigate this, the yeast cells were cultured on glucose supplementing them linoleic acid, and intracellular and extracellular profiles were determined using mass spectrometry. The transport of linoleic acid into the cells had an impact on primary carbon metabolism increasing glucose consumption and ethanol production, thus accelerating the fermentation rate. The energetic state of the cells was therefore affected and the glycolytic pathway, the TCA cycle and the amino acid production were up-regulated. Moreover, since the S. cerevisiae fatty acid profile was altered, an experiment in parallel was performed supplementing the medium with a labelled isotope of linoleic acid to follow its metabolic fate. The finding showed that linoleic acid was metabolised into longer and shorter chain fatty acids. Since fatty acids strongly influence the cellular lipids, an analysis of the lipidome was also performed showing a general reduction of the lipid content in response to linoleic acid. Lipid biosynthesis requires ATP, therefore its reduction is due to the fact that available energy is diverted to linoleic acid uptake and compartmentalisation. The effect of linoleic acid on S. cerevisiae metabolism could be used by several industries. For example, during wine making, the production of aroma compounds can be manipulated through linoleic acid supplementation in the juice in order to reach a desired profile. Moreover, the biofuel industry could test the supplementation of a chosen substrate with this fatty acid, since linoleic acid improves ethanol production by S. cerevisiae.