Malolactic conversion (also known as malolactic fermentation or MLF) is a process in

winemaking Winemaking or vinification is the production of wine, starting with the selection of the fruit, its fermentation into alcohol, and the bottling of the finished liquid. The history of wine-making stretches over millennia. The science of wine and ...

in which

tart A tart is a baked dish consisting of a filling over a pastry base with an open top not covered with pastry. The pastry is usually shortcrust pastry; the filling may be sweet or savoury, though modern tarts are usually fruit-based, sometimes wit ...

-tasting malic acid, naturally present in grape must, is converted to softer-tasting

lactic acid Lactic acid is an organic acid. It has a molecular formula . It is white in the solid state and it is miscible with water. When in the dissolved state, it forms a colorless solution. Production includes both artificial synthesis as well as nat ...

. Malolactic fermentation is most often performed as a secondary fermentation shortly after the end of the primary

fermentation Fermentation is a metabolic process that produces chemical changes in organic substrates through the action of enzymes. In biochemistry, it is narrowly defined as the extraction of energy from carbohydrates in the absence of oxygen. In food p ...

, but can sometimes run concurrently with it. The process is standard for most red wine production and common for some white

grape varieties This list of grape varieties includes cultivated grapes, whether used for wine, or eating as a table grape, fresh or dried (raisin, currant, sultana). For a complete list of all grape species including those unimportant to agriculture, see V ...

such as

Chardonnay Chardonnay (, , ) is a green-skinned grape variety used in the production of white wine. The variety originated in the Burgundy wine region of eastern France, but is now grown wherever wine is produced, from England to New Zealand. For new ...

, where it can impart a "buttery" flavor from

diacetyl Diacetyl (IUPAC systematic name: butanedione or butane-2,3-dione) is an organic compound with the chemical formula (CH3CO)2. It is a yellow liquid with an intensely buttery flavor. It is a vicinal diketone (two C=O groups, side-by-side). Diacet ...

, a byproduct of the reaction. The fermentation reaction is undertaken by the family of lactic acid bacteria (LAB); '' Oenococcus oeni'', and various species of ''

Lactobacillus ''Lactobacillus'' is a genus of Gram-positive, aerotolerant anaerobes or microaerophilic, rod-shaped, non-spore-forming bacteria. Until 2020, the genus ''Lactobacillus'' comprised over 260 phylogenetically, ecologically, and metabolically div ...

'' and '' Pediococcus''. Chemically, malolactic fermentation is a

decarboxylation Decarboxylation is a chemical reaction that removes a carboxyl group and releases carbon dioxide (CO2). Usually, decarboxylation refers to a reaction of carboxylic acids, removing a carbon atom from a carbon chain. The reverse process, which is ...

, which means

carbon dioxide Carbon dioxide ( chemical formula ) is a chemical compound made up of molecules that each have one carbon atom covalently double bonded to two oxygen atoms. It is found in the gas state at room temperature. In the air, carbon dioxide is t ...

is liberated in the process.K. Fugelsang, C. Edwards ''Wine Microbiology'' Second Edition pgs 29-44, 88-91, 130-135, 168-179 Springer Science and Business Media, New York (2010) Jean Jacobson ''"Introduction to Wine Laboratory Practices and Procedures"'' pgs 188-191, Springer Science and Business Media, New York (2010) The primary function of all these bacteria is to convert L-malic acid, one of the two major grape acids found in wine, to another type of acid, L+ lactic acid. This can occur naturally. However, in commercial winemaking, malolactic conversion typically is initiated by an

inoculation Inoculation is the act of implanting a pathogen or other microorganism. It may refer to methods of artificially inducing immunity against various infectious diseases, or it may be used to describe the spreading of disease, as in "self-inoculati ...

of desirable bacteria, usually ''O. oeni''. This prevents undesirable bacterial strains from producing "off" flavors. Conversely, commercial winemakers actively prevent malolactic conversion when it is not desired, such as with fruity and floral white grape varieties such as

Riesling Riesling (, ; ) is a white grape variety that originated in the Rhine region. Riesling is an aromatic grape variety displaying flowery, almost perfumed, aromas as well as high acidity. It is used to make dry, semi-sweet, sweet, and sparkling wh ...

and Gewürztraminer, to maintain a more tart or acidic profile in the finished wine.Dr. Yair Margalit, ''Winery Technology & Operations A Handbook for Small Wineries'' pgs 75-78, 103 & 183-184 The Wine Appreciation Guild (1996) B. Zoecklein, K. Fugelsang, B. Gump, F. Nury ''Wine Analysis and Production'' pgs 160-165, 292-302 & 434-447 Kluwer Academic Publishers, New York (1999) Malolactic fermentation tends to create a rounder, fuller

mouthfeel Mouthfeel refers to the physical sensations in the mouth caused by food or drink, making it distinct from taste. It is a fundamental sensory attribute which, along with taste and smell, determines the overall flavor of a food item. Mouthfeel ...

. Malic acid is typically associated with the taste of green

apple An apple is an edible fruit produced by an apple tree (''Malus domestica''). Apple trees are cultivated worldwide and are the most widely grown species in the genus '' Malus''. The tree originated in Central Asia, where its wild ancest ...

s, while lactic acid is richer and more buttery tasting. Grapes produced in cool regions tend to be high in acidity, much of which comes from the contribution of malic acid. Malolactic fermentation generally enhances the body and flavor persistence of

wine Wine is an alcoholic drink typically made from Fermentation in winemaking, fermented grapes. Yeast in winemaking, Yeast consumes the sugar in the grapes and converts it to ethanol and carbon dioxide, releasing heat in the process. Different ...

, producing wines of greater palate softness. Many winemakers also feel that better integration of fruit and oak character can be achieved if malolactic fermentation occurs during the time the wine is in barrel.J. Robinson (ed) ''"The Oxford Companion to Wine"'' Third Edition pgs 422 & 508 Oxford University Press 2006 A wine undergoing malolactic conversion will be cloudy because of the presence of bacteria, and may have the smell of

butter Butter is a dairy product made from the fat and protein components of churned cream. It is a semi-solid emulsion at room temperature, consisting of approximately 80% butterfat. It is used at room temperature as a spread, melted as a condim ...

popcorn Popcorn (also called popped corn, popcorns or pop-corn) is a variety of corn kernel which expands and puffs up when heated; the same names also refer to the foodstuff produced by the expansion. A popcorn kernel's strong hull contains the se ...

, the result of the production of diacetyl. The onset of malolactic fermentation in the bottle is usually considered a wine fault, as the wine will appear to the consumer to still be fermenting (as a result of CO₂ being produced).John Hudelson ''"Wine Faults-Causes, Effects, Cures"'' pgs 46-53, The Wine Appreciation Guild (2011) However, for early

Vinho Verde Vinho Verde () (literally 'green wine') refers to Portuguese wine that originated in the historic Minho province in the far north of the country. The modern-day 'Vinho Verde' region, originally designated in 1908, includes the old Minho provi ...

production, this slight effervesce was considered a distinguishing trait, though Portuguese wine producers had to market the wine in

opaque Opacity or opaque may refer to: * Impediments to (especially, visible) light: ** Opacities, absorption coefficients ** Opacity (optics), property or degree of blocking the transmission of light * Metaphors derived from literal optics: ** In lingui ...

bottles because of the increase in

turbidity Turbidity is the cloudiness or haziness of a fluid caused by large numbers of individual particles that are generally invisible to the naked eye, similar to smoke in air. The measurement of turbidity is a key test of water quality. Fluids ...

and sediment that the "in-bottle MLF" produced. Today, most Vinho Verde producers no longer follow this practice and instead complete malolactic fermentation prior to bottling with the slight sparkle being added by artificial

carbonation Carbonation is the chemical reaction of carbon dioxide to give carbonates, bicarbonates, and carbonic acid. In chemistry, the term is sometimes used in place of carboxylation, which refers to the formation of carboxylic acids. In inorganic ch ...

.R. Boulton, V. Singleton, L. Bisson, R. Kunkee ''Principles and Practices of Winemaking'' pgs 244-273 & 369-374 Springer 1996 New York

History

Malolactic fermentation is possibly as old as the

history of wine Wine has been produced for thousands of years, with evidence of ancient wine production in Georgia from BC (the earliest known traces of wine), West Azerbaijan province of Iran from BC, Armenia from BC (large-scale production), and Si ...

, but scientific understanding of the positive benefits of MLF and control of the process is a relatively recent development. For many centuries, winemakers noticed an "activity" that would happen in their wines stored in barrel during the warm spring months following

harvest Harvesting is the process of gathering a ripe crop from the fields. Reaping is the cutting of grain or pulse for harvest, typically using a scythe, sickle, or reaper. On smaller farms with minimal mechanization, harvesting is the most l ...

. Like primary alcoholic fermentation, this phenomenon would release carbon dioxide gas and seem to have a profound change on the wine that was not always welcomed. It was described as a "second fermentation" in 1837 by the German enologist Freiherr von Babo and the cause for increased

in the wine. Von Babo encouraged winemakers to quickly respond at the first sight of this activity by

racking Racking, often referred to as Soutirage or Soutirage traditionnel (meaning racking in French), also filtering or fining, is the process of moving wine or beer from one container to another using gravity rather than a pump, which can be disruptiv ...

the wine into a new barrel, adding

sulfur dioxide Sulfur dioxide (IUPAC-recommended spelling) or sulphur dioxide (traditional Commonwealth English) is the chemical compound with the formula . It is a toxic gas responsible for the odor of burnt matches. It is released naturally by volcanic a ...

, and then following up with another set of racking and sulfuring to stabilize the wine.Sibylle Krieger '
''The History of Malolactic Bacteria in Wine
'' pgs 15-21. Accessed: 14 May 2013 In 1866,

Louis Pasteur Louis Pasteur (, ; 27 December 1822 – 28 September 1895) was a French chemist and microbiologist renowned for his discoveries of the principles of vaccination, microbial fermentation and pasteurization, the latter of which was named afte ...

, one of the pioneers of modern

microbiology Microbiology () is the scientific study of microorganisms, those being unicellular (single cell), multicellular (cell colony), or acellular (lacking cells). Microbiology encompasses numerous sub-disciplines including virology, bacteriology, ...

, isolated the first bacteria from wine and determined that all bacteria in wine were a cause for wine spoilage. While Pasteur did notice an acid reduction in wine with the lactic bacteria, he did not link that process to a consumption of malic acid by the bacteria, but rather assumed it was just tartrate precipitation. In 1891, the Swiss enologist Hermann Müller theorized that bacteria may be the cause of this reduction. With the aid of peers, Müller explained his theory of "biological deacidication" in 1913 to be caused by wine bacterium ''Bacterium gracile''. In the 1930s, the French enologist Jean Ribéreau-Gayon published papers stating the benefits of this bacterial transformation in wine. During the 1950s, advances in

enzymatic analysis Enzyme assays are laboratory methods for measuring enzymatic activity. They are vital for the study of enzyme kinetics and enzyme inhibition. Enzyme units The quantity or concentration of an enzyme can be expressed in molar amounts, as with any ...

allowed enologists to better understand the chemical processes behind malolactic fermentation.

Émile Peynaud Émile Peynaud (June 29, 1912 – July 18, 2004) was a French oenologist and researcher who has been credited with revolutionizing winemaking in the latter half of the 20th century, and has been called "the forefather of modern oenology". Biogra ...

furthered

enology Oenology (also enology; ) is the science and study of wine and winemaking. Oenology is distinct from viticulture, which is the science of the growing, cultivation, and harvesting of grapes. The English word oenology derives from the Greek word ...

understanding of the process and soon cultured stock of beneficial lactic acid bacteria was available for winemakers to use.

Role in winemaking

The primary role of malolactic fermentation is to deacidify wine. It can also affect the sensory aspects of a wine, making the

seem smoother and adding potential complexity in the flavor and aroma of the wine. For these other reasons, most red wines throughout the world (as well as many

sparkling wine Sparkling wine is a wine with significant levels of carbon dioxide in it, making it fizzy. While the phrase commonly refers to champagne, European Union countries legally reserve that term for products exclusively produced in the Champagne regi ...

s and nearly 20% of the world's white wines) today go through malolactic fermentation. Malolactic fermentation deacidifies the wine by converting the "harsher"

diprotic In computer science, ACID ( atomicity, consistency, isolation, durability) is a set of properties of database transactions intended to guarantee data validity despite errors, power failures, and other mishaps. In the context of databases, a sequ ...

malic acid to the softer monoprotic lactic acid. The different structures of malic and lactic acids leads to a reduction of titratable acidity (TA) in the wine by 1 to 3 g/L and an increase in pH by 0.3 units. Malic acid is present in the grape throughout the

growing season A season is a division of the year marked by changes in weather, ecology, and the amount of daylight. The growing season is that portion of the year in which local conditions (i.e. rainfall, temperature, daylight) permit normal plant growth. Whi ...

, reaching its peak at '' veraison'' and gradually decreasing throughout the ripening process. Grapes harvested from cooler climates usually have the highest malic content and have the most dramatic changes in TA and pH levels after malolactic fermentation.

Malolactic fermentation can aid in making a wine "microbiologically stable" in that the lactic acid bacteria consume many of the leftover nutrients that other spoilage microbes could use to develop wine faults. However, it can also make the wine slightly "unstable" due to the rise in pH, especially if the wine already was at the high end of wine pH. It is not unusual for wines to be "deacidified" by malolactic fermentation only to have the winemaker later add acidity (usually in the form of

tartaric acid Tartaric acid is a white, crystalline organic acid that occurs naturally in many fruits, most notably in grapes, but also in bananas, tamarinds, and citrus. Its salt, potassium bitartrate, commonly known as cream of tartar, develops naturally ...

) to lower the pH to more stable levels.

Conversion of malic into lactic

Lactic acid bacteria convert malic acid into lactic acid as an indirect means of creating energy for the bacteria by

chemiosmosis Chemiosmosis is the movement of ions across a semipermeable membrane bound structure, down their electrochemical gradient. An important example is the formation of adenosine triphosphate (ATP) by the movement of hydrogen ions (H+) across a memb ...

which uses the difference in pH gradient between inside the cell and outside in the wine to produce ATP. One model on how this is accomplished notes that the form of L-malate most present at the low pH of wine is its negatively charged monoanionic form. When the bacteria move this anion from the wine into higher pH level of its cellular plasma membrane, it causes a net-negative charge that creates

electrical potential The electric potential (also called the ''electric field potential'', potential drop, the electrostatic potential) is defined as the amount of work energy needed to move a unit of electric charge from a reference point to the specific point in ...

. The decarboxylation of malate into L-lactic acid releases not only carbon dioxide but also consumes a proton, which generates the pH gradient which can produce ATP. Lactic acid bacteria convert L-malic acid found naturally in wine grapes. Most commercial malic acid additives are a mixture of the

enantiomer In chemistry, an enantiomer ( /ɪˈnænti.əmər, ɛ-, -oʊ-/ ''ih-NAN-tee-ə-mər''; from Ancient Greek ἐνάντιος ''(enántios)'' 'opposite', and μέρος ''(méros)'' 'part') – also called optical isomer, antipode, or optical anti ...

s D+ and L-malic acid.

Sensory influences

Many different studies have been conducted on the sensory changes that occur in wines that have gone through malolactic fermentation. The most common descriptor is that acidity in the wine feels "softer" due to the change of the "harsher" malic acid to the softer lactic acid. The perception of sourness comes from the titratable acidity in the wine, so the reduction in TA that follows MLF leads to a reduction in perceived sour or "tartness" in the wine. The change in mouthfeel is related to the increase in pH, but may also be due to the production of polyols, particularly the

sugar alcohol Sugar alcohols (also called polyhydric alcohols, polyalcohols, alditols or glycitols) are organic compounds, typically derived from sugars, containing one hydroxyl group (–OH) attached to each carbon atom. They are white, water-soluble solids ...

erythritol Erythritol is an organic compound, a four-carbon sugar alcohol (or polyol) with no optical activity, used as a food additive and sugar substitute. It is naturally occurring. It can be made from corn using enzymes and fermentation. Its formula is ...

and

glycerol Glycerol (), also called glycerine in British English and glycerin in American English, is a simple triol compound. It is a colorless, odorless, viscous liquid that is sweet-tasting and non-toxic. The glycerol backbone is found in lipids known ...

. Another factor that may enhance the mouthfeel of wines that have gone through malolactic fermentation is the presence of ethyl lactate which can be as high as 110 mg/L after MLF. The potential influence on the aroma of the wine is more complex and difficult to predict with different strains of ''Oenococcus oeni'' (the bacterium most commonly used in MLF) having the potential to create different aroma compounds. In Chardonnay, wines that have gone through MLF are often described as having "

hazelnut The hazelnut is the fruit of the hazel tree and therefore includes any of the nuts deriving from species of the genus '' Corylus'', especially the nuts of the species ''Corylus avellana''. They are also known as cobnuts or filberts according ...

" and "

dried fruit Dried fruit is fruit from which the majority of the original water content has been removed either naturally, through sun drying, or through the use of specialized dryers or dehydrators. Dried fruit has a long tradition of use dating back to th ...

" notes, as well as the aroma of freshly baked

bread Bread is a staple food prepared from a dough of flour (usually wheat) and water, usually by baking. Throughout recorded history and around the world, it has been an important part of many cultures' diet. It is one of the oldest human-made f ...

. In red wines, some strains metabolize the amino acid

methionine Methionine (symbol Met or M) () is an essential amino acid in humans. As the precursor of other amino acids such as cysteine and taurine, versatile compounds such as SAM-e, and the important antioxidant glutathione, methionine plays a critical ...

into a derivative of

propionic acid Propionic acid (, from the Greek words πρῶτος : ''prōtos'', meaning "first", and πίων : ''píōn'', meaning "fat"; also known as propanoic acid) is a naturally occurring carboxylic acid with chemical formula CH3CH2CO2H. It is a li ...

that tends to produce roasted aroma and

chocolate Chocolate is a food made from roasted and ground cacao seed kernels that is available as a liquid, solid, or paste, either on its own or as a flavoring agent in other foods. Cacao has been consumed in some form since at least the Olmec ci ...

notes. Red wines that go through malolactic fermentation in the barrel can have enhanced spice or smoke aromas. However, some studies have also shown that malolactic fermentation may diminish primary fruit aromas such as

Pinot noir Pinot Noir () is a red-wine grape variety of the species ''Vitis vinifera''. The name may also refer to wines created predominantly from pinot noir grapes. The name is derived from the French words for ''pine'' and ''black.'' The word ''pine ...

, often losing

raspberry The raspberry is the edible fruit of a multitude of plant species in the genus ''Rubus'' of the rose family, most of which are in the subgenus '' Idaeobatus''. The name also applies to these plants themselves. Raspberries are perennial with ...

and

strawberry The garden strawberry (or simply strawberry; ''Fragaria × ananassa'') is a widely grown hybrid species of the genus '' Fragaria'', collectively known as the strawberries, which are cultivated worldwide for their fruit. The fruit is widely ap ...

notes after MLF. Additionally, red wines may endure a loss of color after MLF due to pH changes that causes a shift in the equilibrium of the anthocyanins which contribute to the stability of color in wine.

Lactic acid bacteria

All lactic acid bacteria (LAB) involved in winemaking, whether as a positive contributor or as a source for potential faults, have the ability to produce lactic acid through the metabolism of a sugar source, as well as the metabolism of L-malic acid. Species differ in how they metabolise the available

sugars in wine Sugars in wine are at the heart of what makes winemaking possible. During the process of fermentation, sugars from wine grapes are broken down and converted by yeast into alcohol (ethanol) and carbon dioxide. Grapes accumulate sugars as they gro ...

(both

glucose Glucose is a simple sugar with the molecular formula . Glucose is overall the most abundant monosaccharide, a subcategory of carbohydrates. Glucose is mainly made by plants and most algae during photosynthesis from water and carbon dioxide, u ...

and

fructose Fructose, or fruit sugar, is a ketonic simple sugar found in many plants, where it is often bonded to glucose to form the disaccharide sucrose. It is one of the three dietary monosaccharides, along with glucose and galactose, that are absorb ...

, as well as the unfermentable

pentose In chemistry, a pentose is a monosaccharide (simple sugar) with five carbon atoms. The chemical formula of many pentoses is , and their molecular weight is 150.13 g/mol.wine yeasts do not consume). Some bacteria species use the sugars through a homofermentative pathway, meaning only one main end product (usually lactate) is produced, while others use heterofermentative pathways that can create multiple end products such as carbon dioxide,

ethanol Ethanol (abbr. EtOH; also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound. It is an alcohol with the chemical formula . Its formula can be also written as or (an ethyl group linked to a ...

, and

acetate An acetate is a salt formed by the combination of acetic acid with a base (e.g. alkaline, earthy, metallic, nonmetallic or radical base). "Acetate" also describes the conjugate base or ion (specifically, the negatively charged ion called ...

. While only the L-

isomer In chemistry, isomers are molecules or polyatomic ions with identical molecular formulae – that is, same number of atoms of each element – but distinct arrangements of atoms in space. Isomerism is existence or possibility of isomers. Is ...

of lactate is produced by LAB in the conversion of malic acid, both hetero- and homofermenters can produce D-, L- and DL-isomers of lactic from glucose which may contribute to slightly different sensory properties in the wine. While ''O. oeni'' is often the LAB most desired by winemakers to complete malolactic fermentation, the process is most often carried out by a variety of LAB species that dominate the must at different points during fermentations. Several factors influence which species will be dominant, including fermentation temperature, nutritional resources, the presence of

, interaction with yeast and other bacteria, pH, and alcohol levels (''Lactobacillus'' species, for example, tend to prefer higher pH and can tolerate higher alcohol levels than ''O. oeni''), as well as initial inoculation (such as "wild" ferments versus an inoculation of cultured ''O. oeni'').

''Oenococcus''

The genus '' Oenococcus'' has one main member involved in winemaking, ''O. oeni'', once known as ''Leuconostoc oeni''. Despite having the name ''Oeno

coccus A coccus (plural cocci) is any bacterium or archaeon that has a spherical, ovoid, or generally round shape. Bacteria are categorized based on their shapes into three classes: cocci (spherical-shaped), bacillus (rod-shaped) and spiral ( of whi ...

'', under the microscope, the bacterium has a

bacillus (shape) A bacillus (), also called a bacilliform bacterium or often just a rod (when the context makes the sense clear), is a rod-shaped bacterium or archaeon. Bacilli are found in many different taxonomic groups of bacteria. However, the name ''Bacill ...

rod shape. The bacteria is a

Gram-positive In bacteriology, gram-positive bacteria are bacteria that give a positive result in the Gram stain test, which is traditionally used to quickly classify bacteria into two broad categories according to their type of cell wall. Gram-positive bact ...

facultative anaerobe A facultative anaerobic organism is an organism that makes ATP by aerobic respiration if oxygen is present, but is capable of switching to fermentation if oxygen is absent. Some examples of facultatively anaerobic bacteria are '' Staphylococ ...

that can utilize some

oxygen Oxygen is the chemical element with the symbol O and atomic number 8. It is a member of the chalcogen group in the periodic table, a highly reactive nonmetal, and an oxidizing agent that readily forms oxides with most elements ...

for

aerobic respiration Cellular respiration is the process by which biological fuels are oxidised in the presence of an inorganic electron acceptor such as oxygen to produce large amounts of energy, to drive the bulk production of ATP. Cellular respiration may be des ...

but usually produces cellular energy through fermentation. ''O. oeni'' is a heterofermenter that creates multiple end products from the use of glucose with D-lactic acid and carbon dioxide being produced in roughly equal amounts to either ethanol or acetate. In reductive conditions (such as near the end of alcoholic fermentation), the third end product is usually ethanol while in slightly

oxidative Redox (reduction–oxidation, , ) is a type of chemical reaction in which the oxidation states of substrate change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is the gain of electrons or a ...

(such as early in alcohol fermentation or in an untopped barrel), the bacteria are more likely to produce acetate. Some ''O. oeni'' strains can use fructose to create

mannitol Mannitol is a type of sugar alcohol used as a sweetener and medication. It is used as a low calorie sweetener as it is poorly absorbed by the intestines. As a medication, it is used to decrease pressure in the eyes, as in glaucoma, and to lo ...

(which can lead to wine fault known as mannitol taint), while many other strains can break down the

amino acid Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha ...

arginine Arginine is the amino acid with the formula (H2N)(HN)CN(H)(CH2)3CH(NH2)CO2H. The molecule features a guanidino group appended to a standard amino acid framework. At physiological pH, the carboxylic acid is deprotonated (−CO2−) and both the am ...

(which can be present in the wine that is resting on the lees after fermentation from the autolysis of dead yeast cells) into

ammonia Ammonia is an inorganic compound of nitrogen and hydrogen with the formula . A stable binary hydride, and the simplest pnictogen hydride, ammonia is a colourless gas with a distinct pungent smell. Biologically, it is a common nitrogenous ...

. In addition to the

hexose In chemistry, a hexose is a monosaccharide (simple sugar) with six carbon atoms. The chemical formula for all hexoses is C6H12O6, and their molecular weight is 180.156 g/mol. Hexoses exist in two forms, open-chain or cyclic, that easily convert ...

glucose and fructose sugars, most strains of ''O. oeni'' can use the residual pentose sugars left behind from yeast fermentation including L-arabinose and

ribose Ribose is a simple sugar and carbohydrate with molecular formula C5H10O5 and the linear-form composition H−(C=O)−(CHOH)4−H. The naturally-occurring form, , is a component of the ribonucleotides from which RNA is built, and so this com ...

. Only around 45% of ''O. oeni'' strains can ferment

sucrose Sucrose, a disaccharide, is a sugar composed of glucose and fructose subunits. It is produced naturally in plants and is the main constituent of white sugar. It has the molecular formula . For human consumption, sucrose is extracted and refine ...

(the form of sugar usually added for chaptalization that gets converted by yeast into glucose and fructose). Winemakers tend to prefer ''O. oeni'' for several reasons. First, the species is compatible with the main wine yeast ''

Saccharomyces cerevisiae ''Saccharomyces cerevisiae'' () (brewer's yeast or baker's yeast) is a species of yeast (single-celled fungus microorganisms). The species has been instrumental in winemaking, baking, and brewing since ancient times. It is believed to have b ...

'', though in cases where both MLF and alcoholic fermentation are started together, the yeast most often outcompetes the bacterium for nutritional resources which may cause a delay in the onset of malolactic fermentation. Second, most strains of ''O. oeni'' are tolerant to the low pH levels of wine and can usually deal with the standard alcohol levels that most wines reach by the end of fermentation. Additionally, while sulfur dioxide levels above 0.8 molecular SO₂ (pH dependent but roughly 35-50 ppm) will inhibit the bacteria, ''O. oeni'' is relatively resistant compared to other LAB. Finally, ''O. oeni'' tends to produce the least amount of

biogenic amines A biogenic amine is a biogenic substance with one or more amine groups. They are basic nitrogenous compounds formed mainly by decarboxylation of amino acids or by amination and transamination of aldehydes and ketones. Biogenic amines are organic ...

(and most lactic acid) among the lactic acid bacteria encountered in winemaking.

''Lactobacillus''

Within the genus ''Lactobacillus'' are both heterofermentative and homofermentative species. All lactobacilli involved in winemaking are Gram-positive and microaerophilic, with most species lacking the enzyme

catalase Catalase is a common enzyme found in nearly all living organisms exposed to oxygen (such as bacteria, plants, and animals) which catalyzes the decomposition of hydrogen peroxide to water and oxygen. It is a very important enzyme in protecting t ...

needed to protect themselves from

oxidative stress Oxidative stress reflects an imbalance between the systemic manifestation of reactive oxygen species and a biological system's ability to readily detoxify the reactive intermediates or to repair the resulting damage. Disturbances in the normal ...

. Species of ''Lactobacillus'' that have been isolated from wine and grape must samples across the globe include '' L. brevis, L. buchneri, L. casei, L. curvatus'', ''L. delbrueckii'' subsp. ''lactis'', '' L. diolivorans, L. fermentum, L. fructivorans, L. hilgardii, L. jensenii, L. kunkeei, L. leichmannii, L. nagelii, L. paracasei, L. plantarum'', and '' L. yamanashiensis''. Most ''Lactobacillus'' species are undesirable in winemaking with the potential of producing high levels of

volatile acidity A wine fault or defect is an unpleasant characteristic of a wine often resulting from poor winemaking practices or storage conditions, and leading to wine spoilage. Many of the compounds that cause wine faults are already naturally present in win ...

, off odors, wine haze, gassiness, and

sediment Sediment is a naturally occurring material that is broken down by processes of weathering and erosion, and is subsequently transported by the action of wind, water, or ice or by the force of gravity acting on the particles. For example, sand ...

that can be deposited in the bottle, especially if the wine had not been filtered. These bacteria also have the potential to create excessive amounts of lactic acid which can further influence the flavor and sensory perception of the wine. Some species, such as the so-called "ferocious ''Lactobacillus''", have been implicated in causing sluggish or

stuck fermentation A stuck fermentation occurs in brewing beer or winemaking when the yeast become dormant before the fermentation has completed. Unlike an "arrested fermentation" where the winemaker intentionally stops fermentation (such as in the production of ...

s, while other species, such as ''L. fructivorans'', have been known to create a cottony

mycelium Mycelium (plural mycelia) is a root-like structure of a fungus consisting of a mass of branching, thread-like hyphae. Fungal colonies composed of mycelium are found in and on soil and many other substrates. A typical single spore germinates ...

-like growth on the surface of wines, nicknamed "Fresno mold" after the wine region where it was discovered.

''Pediococcus''

So far, four species from the genus ''Pediococcus'' have been isolated in wines and grape must, '' P. inopinatus, P. pentosaceus, P. parvulus'', and '' P. damnosus'', with the last two being the species most commonly found in wine. All ''Pediococcus'' species are Gram-positive with some species being micro-aerophilic while others utilizing mostly aerobic respiration. Under the microscope, ''Pediococcus'' often appear in pairs of pairs or tetrads which can make them identifiable. Pediococci are homofermenters, metabolizing glucose into a

racemic mixture In chemistry, a racemic mixture, or racemate (), is one that has equal amounts of left- and right-handed enantiomers of a chiral molecule or salt. Racemic mixtures are rare in nature, but many compounds are produced industrially as racemates. ...

of both L- and D-lactate by

glycolysis Glycolysis is the metabolic pathway that converts glucose () into pyruvate (). The free energy released in this process is used to form the high-energy molecules adenosine triphosphate (ATP) and reduced nicotinamide adenine dinucleotide (NADH ...

. However, in the absence of glucose, some species, such as ''P. pentosaceus'', begin using

, degrading it into

pyruvate Pyruvic acid (CH3COCOOH) is the simplest of the alpha-keto acids, with a carboxylic acid and a ketone functional group. Pyruvate, the conjugate base, CH3COCOO−, is an intermediate in several metabolic pathways throughout the cell. Pyruvic a ...

which later can be converted to diacetyl, acetate,

2,3-butanediol 2,3-Butanediol is the organic compound with the formula (CH3CHOH)2. It is classified as a ''vic''-diol ( glycol). It exists as three stereoisomers, a chiral pair and the meso isomer. All are colorless liquids. Applications include precursors ...

and other compounds that can impart unfavorable characteristics to the wine. Most ''Pediococcus'' species are undesirable in winemaking due to the high levels of diacetyl that can be produced, as well as increased production of biogenic amines that has been implicated as one potential cause for

red wine headache Red wine headache ("RWH") is a headache often accompanied by nausea and flushing that occurs after consuming red wine in susceptible individuals. White wine headaches have been less commonly reported. Sulfites Many wines contain a warni ...

s. Many species of ''Pediococcus'' also have the potential to introduce off odors or other wine faults to the wine such as the bitter-tasting "acrolein taint" that comes from degradation of glycerol into

acrolein Acrolein (systematic name: propenal) is the simplest unsaturated aldehyde. It is a colorless liquid with a piercing, acrid smell. The smell of burnt fat (as when cooking oil is heated to its smoke point) is caused by glycerol in the burning fat ...

which then reacts with phenolic compounds in the wine to produce a bitter-tasting compound. One species, ''P. parvulus'', has been found in wines that have not gone through MLF (meaning malic acid is still present in the wine), but has still had its bouquet altered in a way that enologist have described as "not spoiled" or flaw. Other studies have isolated ''P. parvulus'' from wines that have gone through malolactic fermentation without the development of off odors or wine faults.

Nutritional requirements

Lactic acid bacteria are fastidious organisms that cannot synthesize on their own all of their complex nutritional requirements. For LAB to grow and complete malolactic fermentation, the constitution of the wine medium must provide for their nutritional needs. Like wine yeast, LAB require a carbon source for energy metabolism (usually sugar and malic acid), nitrogen source (such as amino acids and

purine Purine is a heterocyclic aromatic organic compound that consists of two rings ( pyrimidine and imidazole) fused together. It is water-soluble. Purine also gives its name to the wider class of molecules, purines, which include substituted purines ...

s) for

protein synthesis Protein biosynthesis (or protein synthesis) is a core biological process, occurring inside cells, balancing the loss of cellular proteins (via degradation or export) through the production of new proteins. Proteins perform a number of critical ...

, and various vitamins (such as

niacin Niacin, also known as nicotinic acid, is an organic compound and a form of vitamin B3, an essential human nutrient. It can be manufactured by plants and animals from the amino acid tryptophan. Niacin is obtained in the diet from a variet ...

riboflavin Riboflavin, also known as vitamin B2, is a vitamin found in food and sold as a dietary supplement. It is essential to the formation of two major coenzymes, flavin mononucleotide and flavin adenine dinucleotide. These coenzymes are involved i ...

, and

thiamine Thiamine, also known as thiamin and vitamin B1, is a vitamin, an essential micronutrient, that cannot be made in the body. It is found in food and commercially synthesized to be a dietary supplement or medication. Phosphorylated forms of thi ...

) and minerals to assist in the synthesis of enzymes and other cellular components. The source for these nutrients is often found in the grape must itself, though MLF inoculations that run concurrent with alcoholic fermentation risk the yeast outcompeting the bacteria for these nutrients. Towards the end of fermentation, while most of the original grape must resources have been consumed, the

lysis Lysis ( ) is the breaking down of the membrane of a cell, often by viral, enzymic, or osmotic (that is, "lytic" ) mechanisms that compromise its integrity. A fluid containing the contents of lysed cells is called a ''lysate''. In molecular b ...

of dead yeast cells (the "lees") can be a source for some nutrients, particularly amino acids. Plus, even "dry" wines that have been fermented to dryness still have unfermentable pentose sugars (such as arabinose, ribose and

xylose Xylose ( grc, ξύλον, , "wood") is a sugar first isolated from wood, and named for it. Xylose is classified as a monosaccharide of the aldopentose type, which means that it contains five carbon atoms and includes an aldehyde functional g ...

) left behind that can be used by both positive and spoilage bacteria. As with wine yeast, manufacturers of cultured LAB inoculum usually offer specially prepared nutritional additives that be used as a supplement. However, unlike wine yeast, lactic acid bacteria can not use the supplement

diammonium phosphate Diammonium phosphate (DAP; IUPAC name diammonium hydrogen phosphate; chemical formula (NH4)2(HPO4) is one of a series of water- soluble ammonium phosphate salts that can be produced when ammonia reacts with phosphoric acid. Solid diammonium ph ...

as a nitrogen source. Before the introduction of complex nutritional supplements and advances in freeze-dried cultures of LAB, winemakers would cultivate their inoculum of lactic acid bacteria from culture slants provided by laboratories. In the 1960s, these winemakers found it easier to create starter cultures in media that contained

or tomato juice. This "tomato juice factor" was discovered to be a derivative of

pantothenic acid Pantothenic acid, also called vitamin B5 is a water-soluble B vitamin and therefore an essential nutrient. All animals require pantothenic acid in order to synthesize coenzyme A (CoA) – essential for fatty acid metabolism – as well as to, ...

, an important growth factor for the bacteria. As with yeast, oxygen can be considered a nutrient for LAB, but only in very small amount and only for microaerophilic species such as ''O. oeni''. However, no evidence exists currently to suggest that malolactic fermentation runs more smoothly in aerobic conditions than in complete anaerobic conditions, and in fact, excessive amounts of oxygen can retard growth of LAB by favoring conditions of competing microbes (such as ''Acetobacter'').

Native LAB species in the vineyard and the winery

Harvested_grapes_being_loaded_into_crusher_destemmer

''Oenococcus oeni'', the LAB species most often desired by winemakers to carry out malolactic fermentation, can be found in the vineyard, but often at very low levels. While moldy, damaged fruit has the potential to carry a diverse flora of microbes, the LAB most often found on clean, healthy grapes after

are species from the ''Lactobacillus'' and ''Pediococcus'' genera. After crushing, microbiologists usually find populations under 10³

colony forming units In microbiology, colony-forming unit (CFU, cfu or Cfu) is a unit which estimates the number of microbial cells ( bacteria, fungi, viruses etc.) in a sample that are viable, able to multiply via binary fission under the controlled conditions. Coun ...

/mL containing a mix of '' P. damnosus, L. casei, L. hilgardii,'' and '' L. plantarum'', as well as ''O. oeni''. For musts that do not receive an early dose of sulfur dioxide to "knock back" these wild populations of LAB, this flora of bacteria compete with each other (and the wine yeasts) for nutrients early in fermentation. In the winery, multiple contact points can be home to native population of LAB including

oak barrels Oak is used in winemaking to vary the color, flavor, tannin profile and texture of wine. It can be introduced in the form of a barrel during the fermentation or aging periods, or as free-floating chips or staves added to wine fermented in a vess ...

, pumps, hoses, and bottling lines. For wines where malolactic fermentation is undesirable (such as fruity white wines), the lack of proper

sanitation Sanitation refers to public health conditions related to clean drinking water and treatment and disposal of human excreta and sewage. Preventing human contact with feces is part of sanitation, as is hand washing with soap. Sanitation syste ...

of wine equipment can lead to the development of unwanted MLF and result in wine faults. In cases of oak barrels where full and complete sanitation is almost impossible, wineries often mark barrels that have contained wines going through MLF and keep them isolated from "clean" or brand new barrels that they can use for wines that are not destined to go through MLF.

''Schizosaccharomyces'' yeast

Several species in the genus '' Schizosaccharomyces'' use L-malic acid, and enologists have been exploring the potential of using this wine yeast for deacidifying wines instead of the traditional route of malolactic fermentation with bacteria. However, early results with '' Schizosaccharomyces pombe'' have shown a tendency of the yeast to produce off odors and unpleasant sensory characteristics in the wine. In recent years, enologists have been experimenting with a

mutant In biology, and especially in genetics, a mutant is an organism or a new genetic character arising or resulting from an instance of mutation, which is generally an alteration of the DNA sequence of the genome or chromosome of an organism. It ...

strain of '' Schizosaccharomyces malidevorans'' that has so far been shown to produce less potential wine flaws and off odors.

Influence of inoculation timing

Winemakers differ in when they choose to inoculate their must with LAB, with some winemakers pitching the bacteria at the same time as the yeast, allowing both alcoholic and malolactic fermentations to run concurrently, while some wait till the end of fermentation when the wine is racked off its lees and into barrel, and others doing it somewhere between. For practitioners of minimalist or " natural winemaking" who choose not to inoculate with cultured LAB, malolactic fermentation can happen at any time depending on several factors such as the microbiological flora of the winery and the competing influences of these other microbes. All options have potential benefits and disadvantages. The benefits of inoculating for MLF during alcoholic fermentation include: *More potential nutrients from the grape must (though the bacteria will be competing with the yeast for these) *Lower sulfur dioxide and ethanol levels which can otherwise inhibit the LAB *Higher fermentation temperatures which are more conducive to LAB growth and an earlier completion of MLF: The optimal temperatures for malolactic fermentation are between , while the process is significantly inhibited at temperatures below . Wine stored in the barrels in the cellar during the winter following fermentation will often have a very prolonged malolactic fermentation due to the cool cellar temperatures. *Early completion of malolactic fermentation means the winemaker can make a postfermentation SO₂ earlier to protect the wine from

oxidation Redox (reduction–oxidation, , ) is a type of chemical reaction in which the oxidation states of substrate change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is the gain of electrons or ...

and spoilage microbes (such as '' Acetobacter''). Since sulfur dioxide can inhibit MLF, delaying LAB inoculation till after alcoholic fermentation may mean a delay in sulfur addition till early spring when cellar temperatures warm up enough to encourage the completion of MLF. *Less diacetyl production The disadvantages for early inoculation include: *Wine yeast and LAB competing for resources (including glucose) and potential antagonism between the microbes *Heterofermenters such as ''O. oeni'' metabolizing the glucose still present in the must and potentially creating undesirable byproducts such as acetic acid Many of the advantages for postalcoholic fermentation answer the disadvantages of early inoculation (namely less antagonism and potential for undesirable byproducts). Also, the advantage is seen of the lees being a nutrient source through the autolysis of the dead yeast cells, though that nutrient source may not always be enough to ensure MLF runs successfully to completion. Conversely, many of the disadvantages of late inoculation are the absence of the advantages that come from early inoculation (higher temperatures, potentially quicker completion, etc.).

Preventing MLF

For some wine styles, such as light, fruity wines or for low-acid wines from warm climates, malolactic fermentation is not desired. Winemakers can take several steps to prevent MLF from taking place, including: *Limited maceration, early pressing, and early

to limit contact time of the LAB with potential nutrient sources *Maintain sulfur dioxide levels to at least 25 ppm of "free" (unbound) SO₂, depending on the pH of the wine, this may mean an addition of 50–100 mg/L of SO₂ *Maintain pH levels below 3.3 *Keep the wine cool at temperatures between 10 and 14 °C (50. 0 to 57.2 °F) *Filter the wine at bottling with at least a 0.45-

micron The micrometre ( international spelling as used by the International Bureau of Weights and Measures; SI symbol: μm) or micrometer (American spelling), also commonly known as a micron, is a unit of length in the International System of Un ...

membrane filter to prevent any bacteria from making it into the bottle In addition, winemakers can use chemical and biological inhibitors such as

lysozyme Lysozyme (EC 3.2.1.17, muramidase, ''N''-acetylmuramide glycanhydrolase; systematic name peptidoglycan ''N''-acetylmuramoylhydrolase) is an antimicrobial enzyme produced by animals that forms part of the innate immune system. It is a glycoside ...

, nisin,

dimethyl dicarbonate Dimethyl dicarbonate (DMDC) is a colorless liquid and a pungent odor at high concentration at room temperature. It is primarily used as a beverage preservative, processing aid, or sterilant ( INS No. 242) being highly active against typical bever ...

(Velcorin), and fumaric acid, though some (like Verlcorin) are restricted in winemaking countries outside the United States. Fining agents, such as

bentonite Bentonite () is an absorbent swelling clay consisting mostly of montmorillonite (a type of smectite) which can either be Na-montmorillonite or Ca-montmorillonite. Na-montmorillonite has a considerably greater swelling capacity than Ca-m ...

, and putting the wine through cold stabilization will also remove potential nutrients for LAB, thus inhibiting malolactic fermentation. Some experimentation with the use of

bacteriophage A bacteriophage (), also known informally as a ''phage'' (), is a duplodnaviria virus that infects and replicates within bacteria and archaea. The term was derived from "bacteria" and the Greek φαγεῖν ('), meaning "to devour". Bac ...

s (

virus A virus is a submicroscopic infectious agent that replicates only inside the living cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, including bacteria and archaea. Since Dmitri Ivanovsk ...

es that infect bacteria) has been conducted to limit malolactic fermentations, but disappointing results in the cheesemaking industry have led to skepticism about the practical use of bacteriophages in winemaking.

Measuring malic content

Winemakers can track the progression of malolactic fermentation by paper chromatography or with a spectrophotometer. The paper chromatography method involves using capillary tubes to add small samples of the wine to chromatograph paper. The paper is then rolled and placed in a jar filled with a

butanol Butanol (also called butyl alcohol) is a four-carbon alcohol with a formula of C4 H9 O H, which occurs in five isomeric structures (four structural isomers), from a straight-chain primary alcohol to a branched-chain tertiary alcohol; all are a bu ...

solution containing bromocresol green indicator dye for several hours. After the paper is pulled out and dried, the distance of yellow-colored "splotches" from the base line denotes the presence of various acids, with tartaric being closest to the baseline followed by citric, malic, and finally lactic acids near the top of the paper. A significant limitation to paper chromatography is that it will not show exactly how much malic is still remaining in the wine, with the size of the "splotch" on the paper having no correlation to a quantitative figure. The sensitivity of the paper is also limited to a detection threshold of 100–200 mg/L while most measurements of "MLF stability" target a malic level of less than 0.03 g/L (30 mg/L). The enzymatic method allows for a quantitative measurement of both malic and lactic acids, but requires the expense of reagent kits and a spectrophotometer that can measure

absorbance Absorbance is defined as "the logarithm of the ratio of incident to transmitted radiant power through a sample (excluding the effects on cell walls)". Alternatively, for samples which scatter light, absorbance may be defined as "the negative lo ...

values at 334, 340, or 365 nm.

Other products produced

The main products of malolactic fermentation are lactic acid, diacetyl, acetic acid,

acetoin Acetoin, also known as 3-hydroxybutanone or acetyl methyl carbinol, is an organic compound with the formula CH3CH(OH)C(O)CH3. It is a colorless liquid with a pleasant, buttery odor. It is chiral. The form produced by bacteria is (''R'')-acetoin. ...

, and various

ester In chemistry, an ester is a compound derived from an oxoacid (organic or inorganic) in which at least one hydroxyl group () is replaced by an alkoxy group (), as in the substitution reaction of a carboxylic acid and an alcohol. Glycerides ...

s. The amount and exact nature of these products depends on the species/strain of LAB conducting the malolactic fermentation and the condition influencing that wine (pH, available nutrients, oxygen levels, etc.). Some strains of ''O. oeni'' can synthesize higher alcohols which can contribute to fruity notes in the aroma of the wine. Additionally, some strains of the bacterium have beta-glucosidase enzymes that can break down monoglucosides which are aroma compounds attached to a sugar molecule. When the sugar component is cleaved, the rest of the compound becomes

volatilized In chemistry, volatility is a material quality which describes how readily a substance vaporizes. At a given temperature and pressure, a substance with high volatility is more likely to exist as a vapour, while a substance with low volatility ...

, meaning it can potentially be detected in the aroma bouquet of the wine. In the early 21st century, some strains of ''O. oeni'' were shown to use

acetaldehyde Acetaldehyde (IUPAC systematic name ethanal) is an organic chemical compound with the formula CH3 CHO, sometimes abbreviated by chemists as MeCHO (Me = methyl). It is a colorless liquid or gas, boiling near room temperature. It is one of the ...

by breaking it down into ethanol or acetic acid. While this may help for wines with excessive levels of acetaldehyde, for red wines, it can also destabilize the color of the wine by interfering with acetaldehyde's reaction with anthocyanins to create polymeric pigments that help create a wine's color.

Diacetyl

Unstirred and stirred barrels of chardonnay on lees

Diacetyl (or 2,3-butanedione) is the compound associated with the "buttery" aromas of Chardonnays, but it can affect any wine that has gone through malolactic fermentation. At an odor detection threshold of 0.2 mg/L in white wines and 2.8 mg/L in red wines, it can be perceived as slightly buttery or "nutty" while at concentrations greater than 5 to 7 mg/L (5-7 ppm) can overwhelm other

aroma notes The aromas of wine are more diverse than its flavours. The human tongue is limited to the primary tastes perceived by taste receptors on the tongue – sourness, bitterness, saltiness, sweetness and savouriness. The wide array of fruit ...

in the wine. Diacetyl can be produced by the LAB through metabolism of sugar or of

citric acid Citric acid is an organic compound with the chemical formula HOC(CO2H)(CH2CO2H)2. It is a colorless weak organic acid. It occurs naturally in citrus fruits. In biochemistry, it is an intermediate in the citric acid cycle, which occurs in ...

. While citric acid is naturally present in grapes, it is in a very small amount with most of it coming from deliberate addition by the winemaker to acidify the wine. In the presence of both malic and citric acids, the LAB use both, but use the malic much more quickly, with the rate of citric use/diacetyl formation influenced by the particular bacterial strain (with most strains of ''O. oeni'' producing less diacetyl than ''Lactobacillus'' and ''Pediococcis'' species), as well as the

redox Redox (reduction–oxidation, , ) is a type of chemical reaction in which the oxidation states of substrate change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is the gain of electrons or ...

potential of the wine. In wine conditions that have a low redox potential (meaning it is more oxidative such as in a barrel that is not fully topped up), more citric acid will be consumed and diacetyl formed. In more reductive conditions, such as in alcoholic fermentations where yeast populations are at their peak and the wine is heavily saturated with carbon dioxide, the formation of diacetyl is much slower. The yeasts also help keep levels low by consuming diacetyl and reducing it to acetoin and

butylene glycol Butanediol, also called butylene glycol, may refer to any one of four stable structural isomer In chemistry, a structural isomer (or constitutional isomer in the IUPAC nomenclature) of a compound is another compound whose molecule has the same ...

. Diacetyl production is favored in fermentations that run warm with temperatures between . It also tends to be produced at higher levels in wines with lower pH levels (under 3.5), though at levels below 3.2, most strains of LAB desirable for MLF tend to be inhibited. "Wild" (as in uninoculated) malolactic ferments have the potential to produce more diacetyl than inoculated ferments due to the lower initial populations during the lag phase with inoculated ferments usually having an initial inoculum of 10⁶ CFU/mL. Late MLF inoculations, after alcoholic fermentation, also tend to produce higher levels of diacetyl. Chardonnay producers desiring to make the high-diacetyl "buttery style" will often do late or "wild" inoculation in the barrel after primary fermentation, allowing the wine to spend several weeks or even months ''

sur lie Lees are deposits of dead yeast or residual yeast and other particles that precipitate, or are carried by the action of " fining", to the bottom of a vat of wine after fermentation and aging. The same while brewing beer at a brewery is known as ...

'' in reductive conditions that promote diacetyl production. Some sources point out that diacetyl is actually decreased by sur lie, due to surviving yeast metabolizing diacetyl, and therefore malolactic fermentation is best performed apart from lees. With wines that have excessive levels of diacetyl, some winemakers use sulfur dioxide to bind with the compound and reduce the perception of diacetyl by 30 to 60%. This binding is a reversible process and after only a few weeks

aging Ageing ( BE) or aging ( AE) is the process of becoming older. The term refers mainly to humans, many other animals, and fungi, whereas for example, bacteria, perennial plants and some simple animals are potentially biologically immortal. In ...

in the bottle or tank, the high levels of diacetyl return. However, sulfur dioxide added earlier in the malolactic fermentation process limits diacetyl production by inhibiting the bacteria and limiting their activity in its entirety, including the conversion of malic to lactic acid.

Wine faults

Bottles being filled in a bottling truck

The most common fault associated with malolactic fermentation is its occurrence when it is not desired. This could be for a wine that is meant to be acidic and fruity (such as Riesling) or it could be a wine that was previously thought to have gone through MLF and bottled only to have malolactic fermentation commence in the bottle. The outcome of this "in-bottle" fermentation is often gassy, hazy wine that can be unpalatable to consumers. Improvement in sanitation and control of lactic acid bacteria in the winery can limit the occurrence of these faults. For early Vinho Verde producers, the slight effervesce that came from in-bottle malolactic fermentation was considered a distinguishing trait that consumers enjoyed in the wine. However, wineries had to market the wine in opaque bottles to mask the turbidity and sediment that the "in-bottle MLF" produced. Today, most Vinho Verde producers no longer follow this practice and instead complete malolactic fermentation prior to bottle with the slight sparkle being added by artificial carbonation. While not necessarily a fault, malolactic fermentation does have the potential of making a wine "protein unstable" due to the resulting change in pH which affects the solubility of

proteins in wine Proteins are present in wine. The most common proteins include thaumatin-like proteins and chitinases and have a role in the formation of turbidity (haze) especially visible in white wine. The quantity of haze forming is dependent on the quantity ...

. For this reason, protein fining and heat stability tests on wine usually take place after malolactic fermentation has run to completion.

Volatile acidity

While

(VA) is usually measured in terms of acetic acid content, its sensory perception is a combination of acetic (

vinegar Vinegar is an aqueous solution of acetic acid and trace compounds that may include flavorings. Vinegar typically contains 5–8% acetic acid by volume. Usually, the acetic acid is produced by a double fermentation, converting simple sugars to eth ...

y aromas) and

ethyl acetate Ethyl acetate ( systematically ethyl ethanoate, commonly abbreviated EtOAc, ETAC or EA) is the organic compound with the formula , simplified to . This colorless liquid has a characteristic sweet smell (similar to pear drops) and is used in glues ...

(

nail polish remover Nail polish (also known as nail varnish or nail enamel) is a lacquer that can be applied to the human fingernail or toenails to decorate and protect the nail plates. The formula has been revised repeatedly to enhance its decorative properties ...

and

model airplane A model aircraft is a small unmanned aircraft. Many are replicas of real aircraft. Model aircraft are divided into two basic groups: flying and non-flying. Non-flying models are also termed static, display, or shelf models. Aircraft manufactur ...

glue Adhesive, also known as glue, cement, mucilage, or paste, is any non-metallic substance applied to one or both surfaces of two separate items that binds them together and resists their separation. The use of adhesives offers certain advant ...

aromas). High levels of VA can inhibit wine yeast and may lead to a sluggish or stuck fermentation. Several microbes can be a source for VA, including ''Acetobacter'', ''

Brettanomyces ''Brettanomyces'' is a non-spore forming genus of yeast in the family Saccharomycetaceae, and is often colloquially referred to as "Brett". The genus name ''Dekkera'' is used interchangeably with ''Brettanomyces'', as it describes the teleomorp ...

'', and film yeast such as '' Candida'', as well as LAB. However, while LAB usually only produce acetic acid, these other microbes often produce ethyl acetate, as well as acetic acid. Most wine-producing countries have laws regulating the amount volatile acidity permitted for wine available for sale and consumption. In the

United States The United States of America (U.S.A. or USA), commonly known as the United States (U.S. or US) or America, is a country Continental United States, primarily located in North America. It consists of 50 U.S. state, states, a Washington, D.C., ...

, the legal limit is 0.9 g/L for foreign wine exported to the United States, 1.2 g/L for white table wine, 1.4 g/L for red wine, 1.5 g/L for white

dessert wine Dessert wines, sometimes called pudding wines in the United Kingdom, are sweet wines typically served with dessert. There is no simple definition of a dessert wine. In the UK, a dessert wine is considered to be any sweet wine drunk with a meal ...

, and 1.7 g/L for red dessert wine.

European Union wine regulations European Union wine regulations are common legislation related to wine existing within the European Union (EU), Heterofermenting species of ''Oenococcus'' and ''Lactobacillus'' have the potential to produce high levels of acetic acid through the metabolism of glucose, though with most strains of ''O. oeni'', the amount is usually only 0.1 to 0.2 g/L. Several species of ''Pediococcus'' can also produce acetic acid through other pathways. Wines starting out with a high pH levels (above 3.5) stand the greatest risk of excessive acetic acid production due to the more favorable conditions for ''Lactobacillus'' and ''Pediococcus'' species.Wibowo, D., Eschenbruch, R., Davis, C.R., Fleet, G.H., and Lee, T.H. '
Occurrence and Growth of Lactic Acid Bacteria in Wine
'' American Journal for Enology and Viticulture. Vol. 36 No. 4 302-313 (1985) ''L. Kunkeei'', one of the so-called "ferocious ''Lactobacillus''" species, has been known to produce 3 to 5 g/L of acetic acid in wines—levels which can easily lead to stuck fermentations.

"Ferocious" ''Lactobacillus''

In the late 20th century, among American winemakers, seemingly healthy fermentation were reported becoming rapidly inundated with high levels of acetic acid that overcame wine yeasts and led to stuck fermentations. While a novel species of ''Acetobacter'' or wine spoilage yeast was initially thought to be the culprit, it was eventually discovered to be several species of ''Lactobacillus'', ''L. kunkeei'', ''L. nagelii'', and ''L. hilgardii'', collectively nicknamed "ferocious" ''Lactobacillus'' for their aggressive acetic acid production, how quickly they multiply, and their high tolerance to sulfur dioxides and other microbiological controls. Ferments of high-pH wines (greater than 3.5) that spent time

cold soak Cold is the presence of low temperature, especially in the atmosphere. In common usage, cold is often a subjective perception. A lower bound to temperature is absolute zero, defined as 0.00K on the Kelvin scale, an absolute thermodynamic ...

ing prior to yeast inoculations and received little to no sulfur dioxide during crushing seem to be at the most risk for "ferocious" ''Lactobacillus''. While infection seems to be vineyard-specific, currently, none of any of the implicated lactobacilli has been reported as being found on the surface of freshly harvested wine grapes.

Acrolein and mannitol taint

The degradation of glycerol by some strains of LAB can yield the compound

. Glycerol is a sweet-tasting

polyol In organic chemistry, a polyol is an organic compound containing multiple hydroxyl groups (). The term "polyol" can have slightly different meanings depending on whether it is used in food science or polymer chemistry. Polyols containing two, thr ...

present in all wines, but at higher levels in wines that have been infected with ''

Botrytis cinerea ''Botrytis cinerea'' is a necrotrophic fungus that affects many plant species, although its most notable hosts may be wine grapes. In viticulture, it is commonly known as "botrytis bunch rot"; in horticulture, it is usually called "grey mould" or ...

''. An "active-

aldehyde In organic chemistry, an aldehyde () is an organic compound containing a functional group with the structure . The functional group itself (without the "R" side chain) can be referred to as an aldehyde but can also be classified as a formyl gro ...

", acrolein can interact with some phenolic compounds in wine to create highly bitter-tasting wines, described as ''amertume'' by Pasteur. While at least one strain of ''O. oeni'' has been shown to produce acrolein, it is more commonly found in wines that have been infected by strains of ''Lactobacillus'' and ''Pediococcus'' species such as ''L. brevis, L. buchneri'', and ''P. parvulus''. Acrolein taint has also shown to be more common in wines that have been fermented at high temperatures and/or made from grapes that have been harvested at high

Brix Degrees Brix (symbol °Bx) is a measure of the dissolved solids in a liquid, and is commonly used to measure dissolved sugar content of an aqueous solution. One degree Brix is 1 gram of sucrose in 100 grams of solution and represents the strength ...

levels. Heterofermenting species from the genus ''Lactobacillus'', as well as some wild strains of ''O. oeni'', have the potential to metabolize fructose (one of the main sugars in wine) into the sugar alcohols

and (less commonly)

. These are sweet-tasting compounds can add sweetness to a wine where it is not desired (such as

Cabernet Sauvignon Cabernet Sauvignon () is one of the world's most widely recognized red wine grape varieties. It is grown in nearly every major wine producing country among a diverse spectrum of climates from Australia and British Columbia, Canada to Lebanon ...

). Mannitol taint, described as ''mannite'' by Pasteur, in wines is often accompanied by other wine faults, including the presence of excessive levels of acetic acid, diacetyl, lactic acid, and

2-butanol 2-Butanol, or ''sec''-butanol, is an organic compound with formula C H3CH( OH)CH2CH3. Its structural isomers are 1-butanol. isobutanol, and ''tert''-butanol. 2-Butanol is chiral and thus can be obtained as either of two stereoisomers designa ...

, which can contribute to a "vinegary-

y" aroma. The wine may also have a slimy sheen on the surface.

Fresno mold and ropiness

In the mid-20th century, a cottony

-like growth began appearing in the bottles of some sweet

fortified wines Fortified wine is a wine to which a distilled spirit, usually brandy, has been added. In the course of some centuries, winemakers have developed many different styles of fortified wine, including port, sherry, madeira, Marsala, Commandar ...

produced in

California California is a state in the Western United States, located along the Pacific Coast. With nearly 39.2million residents across a total area of approximately , it is the most populous U.S. state and the 3rd largest by area. It is also the m ...

's Central Valley. Being fortified, these wines often had alcohol levels in excess of 20% which is usually a level that discourages growth of most spoilage organisms associated with winemaking. Nicknamed "Fresno mold" due to where it was first discovered, the culprit of this growth was determined to be ''L. fructivorans'', a species which can be controlled by sanitation and maintaining adequate sulfur dioxide levels. Some ''Lactobacillus'' and ''Pediococcus'' species (particularly ''P. damnosus'' and ''P. pentosaceus'') have the potential to synthesize

polysaccharides Polysaccharides (), or polycarbohydrates, are the most abundant carbohydrates found in food. They are long chain polymeric carbohydrates composed of monosaccharide units bound together by glycosidic linkages. This carbohydrate can react with w ...

that add an oily

viscosity The viscosity of a fluid is a measure of its resistance to deformation at a given rate. For liquids, it corresponds to the informal concept of "thickness": for example, syrup has a higher viscosity than water. Viscosity quantifies the int ...

to the wine. In the case of ''Lactobacillus'', some of these saccharides may be

glucan A glucan is a polysaccharide derived from D-glucose, linked by glycosidic bonds. Glucans are noted in two forms: alpha glucans and beta glucans. Many beta-glucans are medically important. They represent a drug target for antifungal medications o ...

s that can be synthesized from glucose present in the wine as low as 50–100 mg/L (0.005 to 0.01% residual sugar) and afflict seemingly "dry" wines. While "ropiness" can occur in the barrel or tank, it is often observed in the wines several months after they are bottled. Wines with pH levels above 3.5 and low sulfur dioxide levels are at most risk for developing this fault. Called ''graisse'' (or "grease") by the French and ''les vins filant'' by Pasteur, this fault has been observed in apple wines and

cider Cider ( ) is an alcoholic beverage made from the fermented juice of apples. Cider is widely available in the United Kingdom (particularly in the West Country) and the Republic of Ireland. The UK has the world's highest per capita consumption, ...

. It can also be potentially be caused by other spoilage microbes such as '' Streptococcus mucilaginous'', ''

Candida krusei ''Candida krusei'' is a budding yeast (a species of fungus) involved in chocolate production. ''Candida krusei'' is an emerging fungal nosocomial pathogen primarily found in the immunocompromised and those with hematological malignancies. It h ...

'', and '' Acetobacter rancens''.

Mousiness and geranium taint

Wines infected with ''L. brevis, L. hilgardii'', and ''L. fermentum'' have been known to occasionally develop an aroma reminiscent of rodent droppings. The aroma becomes more pronounced when the wine is rubbed between the fingers and, if consumed, can leave a long, unpleasant finish. The aroma can be very potent, detectable at a sensory threshold as low as 1.6 parts per billion (

μg In the metric system, a microgram or microgramme is a unit of mass equal to one millionth () of a gram. The unit symbol is μg according to the International System of Units (SI); the recommended symbol in the United States and United Kingdom whe ...

/l). The exact compound behind this is derivatives of the amino acid

lysine Lysine (symbol Lys or K) is an α-amino acid that is a precursor to many proteins. It contains an α-amino group (which is in the protonated form under biological conditions), an α-carboxylic acid group (which is in the deprotonated − ...

created through an oxidation reaction with ethanol. While undesirable LAB species have been most commonly associated with this fault, wine infected by ''Brettanomyces'' yeast in the presence of

ammonium phosphate Ammonium phosphate is the inorganic compound with the formula (NH4)3PO4. It is the ammonium salt of orthophosphoric acid. A related "double salt", (NH4)3PO4.(NH4)2HPO4 is also recognized but is impractical use. Both triammonium salts evolve ammoni ...

and lysine have also been known to exhibit this fault. Sorbate is often used as a yeast-inhibitor by home winemakers to stop alcoholic fermentation in the production of sweet wines. Most species of lactic acid bacteria can synthesize sorbate to produce 2-ethoxyhexa-3,5-diene which has the aroma of crushed

geranium ''Geranium'' is a genus of 422 species of annual, biennial, and perennial plants that are commonly known as geraniums or cranesbills. They are found throughout the temperate regions of the world and the mountains of the tropics, but mostly i ...

leaves.

Tourne

Compared to malic and citric acids, tartaric acid is usually considered microbiologically stable. However, some species of ''Lactobacillus'' (particularly ''L. brevis'' and ''L. plantarum'') have the potential to degrade tartaric acid in wine, reducing a wine's total acidity by 3-50%. French winemakers had long observed this phenomenon and called it ''tourne'' (meaning "turn to brown") in reference to the color change that can occur in the wine at the same time likely due to other processes at work in addition to the tartaric loss. While ''Lactobacillus'' is the most common culprit of ''tourne'', some species of the spoilage film yeast '' Candida'' can also metabolize tartaric acid.

Health-related faults

While the presence of ethyl carbamate is not a sensory wine fault, the compound is a suspected

carcinogen A carcinogen is any substance, radionuclide, or radiation that promotes carcinogenesis (the formation of cancer). This may be due to the ability to damage the genome or to the disruption of cellular metabolic processes. Several radioactive sub ...

which is subjected to regulation in many countries. The compound is produced from the degradation of the amino acid

which is present in both grape must and released in the wine through the autolysis of dead yeast cells. While the use of

urea Urea, also known as carbamide, is an organic compound with chemical formula . This amide has two amino groups (–) joined by a carbonyl functional group (–C(=O)–). It is thus the simplest amide of carbamic acid. Urea serves an important ...

as a source of yeast assimilable nitrogen (no longer legal in most countries) was the most common cause of ethyl carbamate in wine, both ''O. oeni'' and ''L. buchneri'' have been known to produce carbamyl phosphate and

citrulline The organic compound citrulline is an α-amino acid. Its name is derived from '' citrullus'', the Latin word for watermelon. Although named and described by gastroenterologists since the late 19th century, it was first isolated from watermelon in ...

which can be

precursor Precursor or Precursors may refer to: * Precursor (religion), a forerunner, predecessor ** The Precursor, John the Baptist Science and technology * Precursor (bird), a hypothesized genus of fossil birds that was composed of fossilized parts of u ...

s to ethyl carbamate formation. ''L. hilgardii'', one of the "ferocious ''Lactobacillus''" species, has also been suspected of contributing to ethyl carbamate production. In the United States, the

Alcohol and Tobacco Tax and Trade Bureau The Alcohol and Tobacco Tax and Trade Bureau, statutorily named the Tax and Trade Bureau and frequently shortened to TTB, is a bureau of the United States Department of the Treasury, which regulates and collects taxes on trade and imports of alcoh ...

has established a voluntary target limit of ethyl carbamate in wine to less than 15 μg/L for table wines and less than 60 μg/L for dessert wines. Biogenic amines have been implicated as a potential cause of

s. In wine,

histamine Histamine is an organic nitrogenous compound involved in local immune responses, as well as regulating physiological functions in the gut and acting as a neurotransmitter for the brain, spinal cord, and uterus. Since histamine was discover ...

cadaverine Cadaverine is an organic compound with the formula (CH2)5(NH2)2. Classified as diamine, it is a colorless liquid with an unpleasant odor. It is present in small quantities in living organisms but is often associated with the putrefaction of an ...

, phenylethylamine,

putrescine Putrescine is an organic compound with the formula (CH2)4(NH2)2. It is a colorless solid that melts near room temperature. It is classified as a diamine. Together with cadaverine, it is largely responsible for the foul odor of putrefying flesh ...

, and

tyramine Tyramine ( ) (also spelled tyramin), also known under several other names, is a naturally occurring trace amine derived from the amino acid tyrosine. Tyramine acts as a catecholamine releasing agent. Notably, it is unable to cross the blood ...

have all been detected. These amines are created by the degradation of amino acids found in grape must and left over from the breakdown of dead yeast cells after fermentation. Most LAB have the potential to create biogenic amines, even some strains of ''O. oeni'', but high levels of biogenic amines are most often associated with species from the ''Lactobacillus'' and ''Pediococcus'' genera. In the

European Union The European Union (EU) is a supranational union, supranational political union, political and economic union of Member state of the European Union, member states that are located primarily in Europe, Europe. The union has a total area of ...

, the concentration of biogenic amines in wine is beginning to be monitored, while the United States currently does not have any regulations.

References

External links

*Purdue University -
The Joy of Malolactic Fermentation
Accessed 27 Dec. 2007 *Vintessential Articles for Winemakers

{{Winemaking Winemaking