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Chemical Reactions and Wine Production

CH COCOOH CH CHO + CO→3 3 2CH CH OH CH CHO CH COOH→ →3 2 3 3

When we open the wine but we don’t finish it and we reopen the bottle after 2/3 days, we can feel some oxidation, this sensory perception in Italian is called “svanito”. Some of the alcohol was oxidised in acetaldehyde.

Acetic acid, main constituent of the volatile acidity in physiological fermentations it should not exceed 0,2-0,3 g/L. if we have 0,2-0,3 g/L of acetic acid it means the fermentation process went very well.

At the end of the fermentation we have grape pomace, different types according to the type of wine produced:

What can we do with the pomace? We can send them to distilleries, for example to make grappa (from the distillation of fermented pomace that come as a collateral product of red wine production.

- →Sweet pomace (white wines) the fermented pomace is done by the distillery. It’s a very good substrate for micro-organism.

The distilleries have to start alcoholic

fermentation to make them fermented pomace. But sweetpomace is a very nice broth for any other kind of microorganisms. At the end of autumn there are ahuge amount of pomace that are transferred to distilleries, this pomace have to be storedsomewhere, or during the way to distilleries, in order to allow alcoholic fermentation and inhibit anyother kind of fermentation. Malolactic and Acetic microorganisms should be inhibited. Molds tooshould not be allowed to grow. How to facilitate alcoholic fermentation in the pomace that are indistilleries? Normally the pomace are processed after one month, but sometimes they are processedafter several monthsYeast starters could be used. But acetic bacteria would grow to produce acetic acid. We could useSO2? No, because it is volatile, and we would have it in the grappa and we don’t want it.We can add an acid, like sulphuric acid, now days phosphoric acid is most used. In this way we lowthe pH under 3 and we inhibit malolactic fermentation, butacetic bacteria grow very well in acidic environment so also no, well not only. What can we do is to also remove oxygen, to avoid the growth of acetic bacteria? So first we add phosphoric acid, then we add yeasts, then we press the pomace in these bags that could be sausages (sometimes in Veneto in distilleries there are backyards full of pomace sausage bags), in this way oxygen is removed, so we have an anaerobic environment. In this way they can wait until spring. There are studies that evaluated the grappa, the quality of grappa produced in autumn and the grappa produced in April, and grappa produced with pomace pressed within one month always resulted better. - Fermented pomace (red wines) directly to the distillery to make grappa. - Semi-fermented pomace, from rosé wines. Other minor compounds: - Succinct acid (rough taste, acidic) - Lactic acid - Citric acid (Krebs cycle) - Piratic acid - Superior alcohols such as propanol, butanol, amyl alcohol (very important sensory compounds)

deamination process of aminoacids). All these compounds are not really sensorially appreciated. This does not happen only in wine, also in beers and distillates (for example distillates from young wines, as these compounds are very volatile, they are easily transferred in the distillate as well, this is called "fusel alcohols") The superior alcohols in the distillation process are called fuel alcohols.

Why wine is good even if we have these compounds formed after fermentation? The storage process, the maturation. During maturation some chemical reactions take place, the most important among these molecules is the esterification process. All of those (most of them) are organic acids. Esters (combination between an alcohol and an acid), can be ethyl-esters, acetic-esters, for example we may have diethylsuccinate, succinic acid can react with ethanol. Esters have note of fruit, so fruity esters are produced by this process during the maturation. At the same time, we have a decrease of

volatile acidity, and increase in some fruity sensory property. (acetic acid + amyl alcohol, decrease of volatile acidity and increase of the banana flavour) (for ex. ethyl lactate is common during malolactic fermentation because lactic acid is produced from malolactic bacteria and it can find ethanol in wine and combine to be an ester)

The process of alcoholic fermentation is not as easy as it seems. If we use only the yeasts naturally present on the grapes, in a first phase we can have apiculate yeasts (not always so desirable yeasts actually) that start the fermentation, they are present on the grape skin, they produce max 5% of ethanol, then ethanol inhibits them and they die. After saccharomyces cerevisiae prevails and produces up to 18% of alcohol volume. All the yeasts are inhibited by a content of 18% of alcohol. Normally yeasts do not arrive to that amount of alcohol, that means that alcohol was added, for example if we want to stabilise for example Port wine or marsala law allows to

these sweet wines to have an addition of alcohol. At the end of fermentation S. Cerevisiae die because they are inhibited by ethanol, the cells fall broken at the bottom of the tank and release DNA, RNA, proteins, etc. These residues can be used as nutrients for malolactic bacteria. In bad hygienic conditions, especially in wood barrels, which have not been sanitised, the presence of Brettanomyces (better to use stainless steel) is not likeable because they can use the products of the autolysis of the S. Cerevisiae to create undesirable compounds. Importance of hygienic conditions in wine making. Brettanomyces can take some components of the wine, like cinnamic acids, caffeic acids and transform them in volatile phenols, that have a taste that can be pharmaceutical, horse sweat, asphalt, rubber etc, those strange tastes are called "Bret". The bret character may occur in wine due to the contamination of wood barrels. It can give a very bad taste especially in red wines. In orderto avoid bad fermentation, we have to know the conditions that inhibit yeasts: - The sugar content is important, higher than 60 Brix inhibit yeasts, for instance concentrated must, it is stable. - Tannins, at a concentration higher than >5 g/L - in sparkling wines, like champagne, an overpressure higher that 8 atm will inhibit yeasts. (No natural sparkling wines with a pressure higher than 8 atm) Selective yeasts are produced with a freeze drying process, so usually there is no cellular death under 0°. Nitrogen compounds and the winemaking process During alcoholic fermentation not only sugars are useful for the yeast development but also the nitrogen content in order to grow: they need nitrogen compounds in order to build their own proteins and enzymes, for their metabolism. The must should have a good nitrogen content, the fraction of nitrogen useful for yeasts is called YAN (Yeast Available Nitrogen) and it's formed from: NH (inorganic form) + free aa + amines.4 + The idea of

Fermentation is reported in the figure which shows a cell of yeast, above there's the must. The yeasts have to absorb amino acids (aa) present in the must. They need a transporter and they need to use energy, so they hydrolyse ATP - ADP to get the energy and use it to transport the aa inside the cell.

If the right amino acids are not present, they'll take the proteins present in the must. They will have to hydrolyse the proteins outside the cell in order to transport the aa. Once done, they absorb the free aa through the membrane. The enzyme transaminase will convert the amino groups from one amino acid to another in order to get different amino acids (such as glutamate) needed for the yeasts to produce their structural proteins and enzymes. They will use the sugar present in the must to get energy.

The figure shows yeast preferences towards N compounds: on the top ammonium groups and free aa. In the top part is formed by YAN meanwhile the peptide and proteins are the least favourable because they...

have to hydrolyse them, so they have to spend energy to get free aa.

YAN is the sum of the concentration of the ammonium + free aa - proline (simple aa not consumed by yeasts during fermentation). The nitrogen concentration in grape must can vary according to the weather, time of harvest (if hot, the grape will grow fast and that means they use all the proteins and the N available will be low because the grapes will be overripe) and also to the type of grapes. It can range from 100 to 600 mg/l. Low N means less than 150 mg/l and in that case the yeasts will have big difficulties: they will produce off-odours during the alcoholic fermentation.

Mechanism of off-odours production: lack of free aa so the proteins are taken by the yeasts from the environment (the must in this case). These are hydrolysed and they can create aa that can release sulphuric compounds which are responsible for the reductive off-flavours. It's called reductive because the sulphuric compounds are eliminated by oxygen.

but during the alcoholic fermentation the oxygen is consumed so they are still present can give the aroma of rotten egg which is a big defect in wines. The rotten egg odour (H2S) comes from cysteine and methionine, the two mains aa containing sulphur. To prevent this mechanism we should give the yeasts the fraction of nitrogen compounds that they need before or during the alcoholic fermentation. The wineries together with the inoculation of yeasts, they'll add yeasts nutrients. These compounds should be added just if there's a lack of nitrogen compounds but the analysis is time consuming so most of the time they're added in any case. If the N is in excess there shouldn't be any disadvantages. Yeast nutrients are vitamins (group B, especially thiamin) and DAP (diammonium phosphate). The quantities of these nutrients depend on the quantity of sugars present in the grape must: if the sugar content is high that means that the yeasts can develop intensively, so they will need more.YAN.[Innovative technique: autolysed yeasts. Yeasts are put into a reactor with N and sugars and after that the cells will die leading to the autolysis. After the autolysis the content of the cells will be released and it will be used as a nutrient in the grape must]

Nitrogen compounds are related to some volatile compounds that are sensory active so practically the amino acids absorbed by the yeast are inter-converted into some others amino acids. One of metabolic activity of the yeasts is the reduction and decarboxylation of amino acids. They can be decarboxylated and reduced to get alcohols. Phenyl-ethanol coming from phenylalanine, has a strong odour of rose. Superior alcohols are the alcohols which have a molecular weight higher than ethanol. Other alcohol that come from other shorter aa, may form amyl-alcohols (5 carbons atoms alcohols). When they're concentrated their odour is of nail polish solvent and an odour of cut grass as well.

These compounds can evolve in wine. All these

Compounds formed during fermentation evolve during the storage of wine.