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A Technical Guide for Wine Producers
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How to ensure and improve wine fermentation by first protecting the yeast
Antonio Palacios García1, Carlos Suárez Martínez1; José Heras1; Anne Ortiz-Julien2, Jean-Michel Salmon3
1 Lallemand Spain, Penˇnsula Ibérica, Ctra Logroño-Vitoria, No 14, 26360 Fuenmayor, La Rioja, Spain, email: apalacios@lallemand.com
Introduction
One of the primary concerns of the oenologist during winemaking is to ensure steady and complete fermentation so all the sugars in the must are converted into alcohol. This should be done in order to avoid any unpleasant surprises that could arise with stuck fermentation or problems due to secondary metabolisms detrimental to the aroma and taste of the wine.
Nutrition and protection: two different concepts
Nutrition, in the form of assimilable nitrogen, membrane lipids, vitamins and minerals, is vital for yeast during fermentation, but protection is also important -two concepts that need to be differentiated and treated separately. Before feeding selected yeasts, first ensure they are well protected and in good physiological condition, in order to improve viability in a stressful medium not ideal for growth. For proper fermentation, good oenological practices include both nutrition and protection.
How to manage and optimize yeast protection
Nourished with the micronutrients (the vitamins and minerals available in the must), sterols are responsible for maintaining the integrity of the yeast cell membrane and increasing its resistance to ethanol. The protection and nutrition of yeast are consecutive and complementary steps. Their benefits should be combined to ensure good alcoholic fermentation, with no problems with stuck fermentation or secondary metabolisms compromising the quality of the wine.
In recent years, the use of selected yeasts in winemaking has become widespread, allowing greater control of the alcoholic fermentation and reducing the risk of unwanted sensory effects resulting from the growth and metabolism of contaminating indigenous yeast. However, a lack of nutrients in the fermentation medium, as is the case in very mature harvests (rich in sugars and polyphenols), harvests with Botrytis, highly clarified musts with low turbidity, and in the presence of inhibiting substances, can lead to slow or stuck fermentations.
Numerous oenological studies and research projects have focused on defining the best supplements and correct levels of nutrients in the fermentation medium. The means used in winemaking are mainly assimilable nitrogen, vitamins, minerals, anaerobic growth factors and oxygen. Contrary to assimilable nitrogen, must deficiencies in terms of sterols, vitamins and minerals are rarely foreseeable when the selected yeasts are inoculated, and therefore in adverse fermentation conditions general oenological practices should include protecting the yeast for fermentation.
Protection during the rehydration of the yeasts
During rehydration in an aqueous medium, active dry yeast can be supplemented with protective nutritional elements derived from inactive yeast rich in certain cellular components, to increase fermentation and metabolic capacity in the must. Lallemand, in collaboration with INRA (Montpellier) has discovered that the membrane lipids and the sterols from the inactive yeast form in situ hydrophobic micellar colloids in water during rehydration of selected active yeast. These colloidal particles move through the rehydration water until they reach the membrane of the active yeast, and integrate into its structure, which is particularly effective at stimulating efficient alcoholic fermentation. The spontaneous formation of these sterol-rich micellae occurs through the formation of colloids that stay in suspension in a aqueous medium and are made up of membrane phospholipids, specific polysaccharides of cell walls and sterols, all deriving from the inactive yeast prepared for that purpose. During the rehydration of the active yeast, these micellae are capable of interacting with the cellular membranes, modifying the original structure of the plasmatic membrane and significantly increasing sterol and unsaturated fatty acid content - very useful in maintaining the fluidity of the membrane and also making it more resistant to ethanol (see Figures 1 and 2).
Figure 1: Normal yeast and yeast protected with Natstep during rehydration, where the sterol-enriched membrane can be observed.
The selected natural yeast strains used to ferment wine contain only 8% moisture in their dry form, thereby guaranteeing ideal conservation until they are used in winemaking. The yeast cell membrane is made up of two flexible and permeable lipid layers, performing a vital function which consists of controlling the entry and exit of cellular metabolites. The yeast protector acts specifically during the rehydration phase, releasing specific micronutrients and micro-protectors that move from the inactive yeast to the active yeast in the warm rehydration water, becoming available to the fermentation yeast and enhancing its effectiveness during alcoholic fermentation.
The micronutrients (vitamins and minerals) are absorbed by the yeast cells through the rehydration water, with the dry yeast cells acting like sponges absorbing the water, causing cell volume to increase 1.5 times. This allows the yeast cells to reactivate their internal metabolism.
The micro-protectors (specific sterols and polyunsaturated fatty acids [PUFA]) gradually integrate into the yeast's cellular membrane, strengthening it and facilitating exchanges with the exterior, thereby preventing the loss of internal cell material (20-30% of the dry weight of the yeast), which the yeast undergoes when solubilized and dispersed in the rehydration water (see Figure 2).
Figure 2: Sterol incorporation during an ADY (1 g) rehydration in 10 mL of water containing 0.5 g of glucose in the presence of (solid line) or absence of (broken line) of 25 mg of lipid colloids at 37°C.
When rehydration with yeast protectors is complete, the yeast is ready to act efficiently and resist the multiple stress factors present in the alcoholic fermentation of a must with technical difficulties.
The protection strategy assists the yeast at the following critical points:
During incorporation into the must, the yeast suffers osmotic shock due to the high sugar concentration in the must. The specific micro-protectors protect the yeast, helping it resist the osmotic shock during incorporation into the must. The specific micronutrients allow the yeast to acclimatize better and develop an ideal metabolism. As a result, the yeast produces less volatile acidity and hydrogen sulphide (H2S), etc., that affects the final quality of the wine.
During the multiplication phase, the inoculated yeast will transfer part of its own cellular material to the following generations, gradually reducing the thickness of the cell membrane in each successive generation. The yeast protector offers it a supplement of reserves in micronutrients and micro-protectors that can be transferred to the future generations, increasing the resistance and viability of the final population, which would usually be in poor condition and facing worse fermentation conditions, given that nutrients are practically non-existent in this phase, and the degree of alcohol is higher, with the possibility of toxins that could inhibit their activity. Maintaining a membrane rich in micro-protectors increases the resistance of future generations and enhances the viability of the total yeast population (see Figure 3).
Figure 3: Fermentation kinetics of selected yeast without protection (blue) and with protection (red) during rehydration.
At the end of the fermentation process, the yeast suffers a high degree of stress due to the high alcohol concentration and the difficulties in transporting residual sugars: the alcohol diffuses through the membrane and kills the cell. During the rehydration phase, protected yeast maintains a very high viability rate. Its membrane, rich in sterols and PUFA is able to withstand high concentrations of alcohol and prevent the alcohol from entering the cells. With this protection, the yeast can finish consuming all the sugars in the must. Healthy and stress-free yeast cells do not produce excessive amounts of undesirable compounds or volatile acidity. Protecting the yeast guarantees that the end of the fermentation is faster and safer (see Figure 4).
Figure 4: Viable yeast cells on completion of alcoholic fermentation in white and red wines.
Conclusion
In conclusion, we may state that protecting the yeast cells during rehydration guarantees the following principal parameters:
During fermentation, the protected yeast also indirectly assists in:
Availability in South Africa
This recently developed concept is known as Natstep (NATural STErols Protection), and a patented product is now on the market. This product is called Go-Ferm Protect© and is available in South Africa through Protea Chemicals. Please contact them at tel: (021) 550 8100 to place an order. If you do need more technical information, please contact Piet Loubser from Lallemand South Africa at tel: (021) 913 7555 or e-mail: - ploubser@lallemand.com
Bibliography
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Saulite, V. A., A. I. Rapoport, and M. E. Beker. 1986. Lipid inclusions in cells and changes in them during dehydration and reactivation of yeasts. Microbiology. 55:99-104.
Soubeyrand, V., V. Luparia, P. Williams, T. Doco, A. Vernhet, A. Ortiz-Julien, and J. M. Salmon. 2005. Formation of micella containing solubilized sterols during rehydration of active dry yeasts improves their fermenting capacity. Journal of Agriculture and Food Chemistry. August:33-45.
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