Science behind wine fermentation

Science behind wine fermentation

Science behind wine fermentationSara Belchik28 June 2014Wine fermentation

OverviewHistoryYeastSugar metabolismEthanol fermentationOther productsSpontaneous fermentationInoculated fermentationHow to get the perfect yeast

Wine predates written records

But we did not understand it6000 BCEEarliest evidence of wine production1680 ADDescription of small spheres during fermentation

But we did not understand it6000 BCEEarliest evidence of wine production1680 ADDescription of small spheres during fermentation1830 ADSmall spheres identified as living organisms

Pasteur experimentSterilizedNot sterilizedPasteur experimentSterilizedNot sterilized

Yeast are responsible agents6000 BCEEarliest evidence of wine production1680 ADDescription of small spheres during fermentation1830 ADSmall spheres identified as living organisms1863 ADYeast identified as microbe responsible for fermentation of grapes into wine1890 ADAddition of pure yeast culture to help with fermentation 1965 ADCommercially available dehydrated yeast for sale


What are yeast?

Related to humans11Yeast are eukaryotic cellsUnicellular organismsKingdom FungiComplex interior allows for complex reactions to occurYeast is umbrella termAlmost 2000 speciesEarliest domesticated organism

Semi-permeable membrane

Water-loving parts on exteriorWater-hating portions on interiorDifficult for many molecules to get throughRequire transport proteins to help get across membrane

Amphipathic nature of membrane and semi-permeablityTransport proteins required to get molecules like glucose into the cell13Many yeast species

Not all yeast are created equal14Wine yeast generaPredominant species on surface of grape berriesKloeckera Also found on grapes but to lesser extentBrettanomycesCandidaCrytococcusKluyveromycesMetschnikowiaPichiaRhodotorulaFound in vineyards or on winemaking equipmentDebaryomycesSaccharomycesSaccharomycodesSchizosaccharomycesZygosaccharomyces


Early earth had no oxygen

How did early microbes generate energy?Need to generate energy in absence of oxygenLife evolved to utilize glucoseGlucose molecule has energy within its bondsMultiple enzymes adapted to produce maximum energy out of these bondsFirst part of glucose metabolism is glycolysisAll life on earth uses glycolysis

ENERGYGlucose is a sugar/carbohydrate18Glycolysis generates pyruvate and NADHGlucose 2 pyruvate

2 ATP generatedEnergy currency of the cellNeeded for most cellular processes

2 NADH generatedElectron carrier of the cellNeeds to be oxidized to NAD+ for glycolysis to continueFate of NADH determined by environment

Glycolysis generates pyruvate and NADHGlucose 2 pyruvate

2 ATP generatedEnergy currency of the cellNeeded for most cellular processes

2 NADH generatedElectron carrier of the cellNeeds to be oxidized to NAD+ for glycolysis to continueFate of NADH determined by environment

NADH and NAD+

Need to regenerate this to continue using glucoseRegenerate NAD+ to keep using glucose

This pathway uses oxygen and generates lots of ATPOur muscles sometimes do thisGet better version from book websiteSince early life 22OverviewHistoryYeastSugar metabolismEthanol fermentationOther productsSpontaneous fermentationInoculated fermentationHow to get the perfect yeast

Yeast use ethanol fermentation

To generate NAD+24From glucose to ethanol

Glucose comes from the grape


Butwine isnt just ethanol

Ethanol, glycerol, and organic acids come from microorganisms28Glycerol in wine

Glycerol does not affect aroma, but it has a slightly sweet taste and is viscousMore common in red wines than white wines29Glycerol production by yeast

Helps to balance redox and help with osmotic stressIncreased glycerol production results in decreased ethanol content30Organic acidsTartaric, malic, and citric acid present in grapesTartaric and citric acid mostly stable throughout fermentationMalic acid is tart and considered undesirable in a red wineLuckily, bacteria (not yeast) can convert malic acid to more palatable lactic acid

Oenococcus oeniCertain bacteria are preferred over others based on byproducts of fermentation

Products of malolactic acid fermentationLactic acidAcetoin and diacetylAcetic acidVarious estersHigher alcohols


What about before 1863?Microorganisms are ubiquitous and live in various environmentsGrapes are just one example of a habitatYeast and certain bacteria live on grapesWhen grapes are crushed, the natural microbes mix with grape juiceCalled spontaneous fermentation

Some wineries still do this34Spontaneous fermentationLow concentrations of microbes on grapesFermentation takes longer to startEntire process requires more time

Wild yeast may not have favorable traitsLow alcohol toleranceResults in high sugar content

Other microbes presentUncertainty of final productOff-aromas and weird esters


Yeast added to control end product

Inoculated fermentationAdded at high concentrations to grape mustInoculum outcompetes other microorganisms presentPreferred yeast is Saccharomyces cerevisiaeMultiple strains existDifferent strains for different wines

Other yeast can also be usedKloeckeraZygosaccharomyces


Breeding to generate perfect yeastYeast can be haploid (one copy of chromosomes) or diploid (two copies)

Take two yeast with desirable traits and breed them

Result is diploid cell containing genetic information from both parents

Not a perfect process

Target specific genes in yeast

Lets target glycolysisProcess involves a series of enzymesWe can increase number of enzymes in the processIncreased flux through glycolysis results in more pyruvateMore pyruvate results in more ethanolNot much successRate limiting step is getting glucose into the cell

Lets target sugar transportIncrease amount of glucose getting into cellIncreased glucose in cell ultimately results in more ethanolTarget glucose transportersSuccess!

Lets target sugar transportIncrease amount of glucose getting into cellIncreased glucose in cell ultimately results in more ethanolTarget glucose transportersSuccess!

Lets improve ethanol toleranceDifferent yeast strains have different ethanol tolerance limitsEthanol destabilizes the membraneTarget membrane fluidity or detoxification enzymesMany GMOs with increased tolerance

Other modificationsSulfur dioxide resistanceSO2 used to decrease unwanted microbesHigher SO2 resistance in yeast means we can increase concentration and kill off even more unwanted microbes

Nitrogen assimilationYeast require carbon and nitrogenNitrogen not abundant in grapesIncrease nitrogen uptake or utilization to increase robustness of yeastYeast GMOs result in more wine varieties

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