Winetech Technical Yearbook 2022

yeasts, such as Metschnikowia pulcherrima and Torulaspora delbrueckii , can also produce fatty acids with more double bonds (known as polyunsaturated fatty acids). The yeast uses all these fatty acids to produce phospholipids and sphingolipids. Together with ergosterol, they form the backbone of cellular membranes (figure 1). Proteins such as nutrient transporters are then embedded into the membranes. It should be emphasised that the production of ergosterol and unsaturated fatty acids requires oxygen, which is in minimal supply during fermentations. While yeasts can produce their own lipids in the presence of oxygen, they can also take up lipids from their environment (i.e., grape juice). This is especially important in the absence of oxygen when the presence of lipids, mainly unsaturated fatty acids and phytosterols, in grape juice becomes essential. HOW DO LIPIDS IMPACT YEAST STRESS SURVIVAL DURING WINE FERMENTATION? As fermentation progresses, oxygen is rapidly depleted, and the yeasts must rely on the presence of sterols and unsaturated fatty acids in grape juice. These can be taken up to compensate for the lack of oxygen-dependent production in the cells. These lipids ensure higher viability and the optimal transport of nutrients as the environment becomes increasingly unfavourable due to the rise in ethanol levels. Indeed, ethanol negatively affects yeast viability by disrupting membrane structure and function. If yeasts are unable to adjust their membrane composition, the accumulation of ethanol will result in an inhibition of sugar uptake and metabolism, ultimately leading to a stuck fermentation. Adjustments in the lipid composition to counteract the detrimental effects of ethanol include the enhanced incorporation of sterols and monounsaturated fatty acids in S. cerevisiae to maintain membrane fluidity (figure 2) . On the other hand, the presence of lower concentrations of ergosterol and higher amounts of polyunsaturated fatty acids has been linked to ethanol sensitivity in wine-related non- Saccharomyces yeasts indicating variability in the lipid metabolism of wine yeasts. In addition to playing a role in ethanol tolerance, the presence of unsaturated fatty acids is necessary to increase plasma membrane Acetic acid production can be influenced by the presence of lipids in grape juice. Indeed, if the concentration of lipids in the juice is sub-optimal, yeasts are forced to produce their own lipids as a compensatory mechanism. To achieve this, the yeasts increase their production of acetyl-CoA (which derives from sugar metabolism), which serves as a substrate for lipid production. This over-production of acetyl-CoA frequently results in the production of acetic acid and, subsequently, ethyl acetate, thereby increasing the wine’s volatile acidity. Medium chain fatty acid levels tend to increase, because of this overproduction. On the other hand, a higher concentration of lipids (or oxygen) has been connected to enhanced production of ethyl esters. Moreover, the addition of lipids has also been shown to increase the production of the sought-after thiols 4-mercapto-4-methyl-2-pentanone (4MMP) and 3-mercapto 1-hexanol (3MH) responsible for box tree or fruity aromas in a S. cerevisiae x Saccharomyces kudriavzevii hybrid, but to decrease 3-mercaptohexyl acetate (3MHA) responsible for fruity aromas . The latter is due to the repression of the enzyme that catalyses the conversion of higher alcohols to acetate esters. RECOMMENDATIONS TO WINEMAKERS Although it can be achieved in a well-equipped analytical laboratory, quantifying lipids is not a straightforward affair. However, the quantity of grape solids (grape juice turbidity) is a satisfactory proxy for lipid availability in white or rosé grape juice. Turbidity is typically measured by a turbidity meter and expressed in nephelometric turbidity units (NTU). To ensure enough lipids fluidity during fermentations at a low temperature. LIPIDS’ IMPACT ON THE WINE BOUQUET

are available for utilisation by yeasts, turbidities of 50 - 150 NTU are recommended, with 100 NTUs being the best compromise to prevent stuck fermentations, whilst minimising the production of off-flavours. Commercial lipid-rich nutrients can also be used during rehydration to increase yeast viability, especially if fermentation conditions are predicted to be difficult (e.g., cold temperature or high sugar content). Micro-oxygenation/aeration can also be used to induce lipid biosynthesis and increase yeast viability, but should be implemented with caution during white winemaking to prevent oxidation and the appearance of off flavours. CURRENT RESEARCH It was recently discovered that wine-related non- Saccharomyces yeasts, such as Lachancea thermotolerans , T. delbrueckii and M. pulcherrima , cannot take up sterols. Thus, the poor fermentation performance of non- Saccharomyces yeasts could, in part, be due to an inability of these yeasts to adjust their membrane composition. Indeed, whereas S. cerevisiae assimilates sterols in addition to fatty acids to reinforce the plasma membrane as ethanol accumulates, it appears that selected non Saccharomyces yeasts are limited to only utilising fatty acids during fermentation, but this hypothesis remains to be assessed. Overall, there is still much that we do not fully understand about the role of sterols and fatty acids on yeast tolerance to ethanol and cold temperatures, as well as on aroma production. These aspects require more research. ABSTRACT Lipids are essential for yeast cell structure, functioning and energy storage. These compounds are removed from grape juice during clarification, and lipid deficiency may cause stuck fermentations. Free fatty acids and phytosterols present in grape must are utilised by S. cerevisiae for fermentation performance, ethanol stress resistance and viability. Furthermore, these compounds are used to produce volatile compounds which impact the organoleptic properties of wine. Therefore, winemakers should ensure that the grape juices they work with have sufficient lipids for successful fermentation. Grape juice turbidities of 50 - 150 nephelometric turbidity units (NTU) are recommended to ensure successful fermentation. In case of over-clarification, the lipid content of grape juice can be increased by adding commercial lipid-containing nutrients. Alternatively, micro-oxygenation/ aeration could be applied to increase lipid biosynthesis and viability in yeast. HIGHLIGHTS • Lipids are vital components of yeast cell membranes. • During fermentation, yeasts must take up lipids from grape juice to handle ethanol and cold temperature stress. • Lipid deficiency can occur, because of over-clarification in white grape juice. • Turbidities of 50 - 150 NTU are recommended to avoid deficiency and possible stuck fermentations. • Alternatively, lipid-rich nutrients can be added, or micro oxygenation/aeration implemented with caution. REFERENCES https://www.wineland.co.za/lipids-in-white-winemaking-the too-oft-ignored-essential-nutrients/

For more information, contact Prof Benoit Divol at divol@sun.ac.za.

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WINETECH TECHNICAL YEARBOOK 2022