Diagnostic Key Classes of Off-Character Compounds S compounds

The S-Containing Volatiles:

Sulfur-containing volatile compounds are perhaps the most challenging of the off-aromas that can form during fermentation. These compounds are reactive and can exist in different chemical states making both detection and removal difficult. The off-aroma may seem to have disappeared only to return later in processing as chemical conditions change. More oxidized forms may be non-volatile or non-aromatic, but if the environment becomes more reduced will be restored to the aromatic reduced form and reappear as a sensory defect.

Hydrogen sulfide: The formation of volatile sulfur compounds in wine is a chronic problem that occurs globally. Many factors have been associated with the appearance of hydrogen sulfide in wine and it has been difficult to design fermentation management strategies that guarantee the absence of sulfide during production.  This is in large part due to the fact that there is a significant influence of strain genetic background on both the level of sulfide produced and the conditions under which it appears. There is also a strong interactive impact of juice composition that exacerbates the underlying genetic differences across strains. Hydrogen sulfide can be produced in yeast as a consequence of the reduction of sulfate that is needed in order to synthesize the sulfur containing amino acids methionine and cysteine.  In this case, it has been hypothesized that more sulfide is reduced than can be incorporated such that the excess sulfide is released from the cell as hydrogen sulfide.

However, H2S has recently been shown to perform an important signaling function. H2S will arrest respiration and signal onset of fermentation. This allows the population of cells to coordinate metabolic activity. H2S is deliberately made and released to coordinate rapid fermentation onset. This serves to accelerate strain control of an environment by rapid generation of ethanol by the population, and signals the availability of sugar to the culture. In this case H2S formation would not be responsive to nitrogen availability as it is not being released under conditions of starvation.  Bacterial respiration will likewise be inhibited by the presence of H2S and this metabolite may play an underappreciated role in allowing ecosystem dominance by Saccharomyces. In this case there would be a strong selective advantage to the production of H2S under certain environmental conditions. In contrast it is difficult to imagine the benefit if being metabolically sloppy and reducing more sulfate than can be consumed by cellular metabolism. The strain variation in H2S formation and high level production found particularly in vineyard isolates indicates that formation of H2S is beneficial to the strains.

H2S can also be formed from the degradation of sulfur containing amino acids. If the levels of methionine and cysteine are high relative to other nitrogen containing compounds they can be used as nitrogen sources. In this case the nitrogen is what is of value to the cell, not the sulfur. The sulfide can then be released either as H2S or as a thiol or some other aromatic waste form of organic sulfur.  Some rootstocks have been shown to yield fruit with relatively high concentrations of the sulfur-containing amino acids. Inorganic sulfur used as an antifungal treatment on the grapes can also be reduced to H2S during fermentation.  This process requires the reducing conditions established by yeast metabolism, but is basically a chemical reduction that is thought for the most part to be strain independent.

Yeast strains can produce sulfide early in the fermentation, typically as cell biomass is expanding and fermentation rates are at their highest or it can be produced late in fermentation as the cells remain in contact with the wine. This latter formation of hydrogen sulfide is thought to be due to the degradation of sulfur containing amino acids, and does show a genetic influence, meaning that some strains appear to have more of a tendency to produce sulfides under these conditions than others.

During yeast lees aging it is also thought that H2S can be produced directly from SO2.  If sulfite reductase is active and there is a source of electrons to generate reduced cofactors, the sulfite can be reduced to sulfide and released.

Higher Sulfides: More complex sulfides can also be formed in wine. The commonly found sulfides are shown in the following table.

Volatile Sulfides Found in Wine

Compound

Aroma Description

Source

Concentration in Wine (µg/L)

Putative Thresholda (µg/L)

Hydrogen sulfide

Rotten eggs, fecal

Sulfate reduction, S-amino acids

nd-370

50-80

Dimethyl sulfide

Cooked corn, cooked asparagus, cabbage, molasses, canned vegetable, skunk, clam

S- amino acids

ndb-480

10/25/60

Diethyl sulfide

Garlic, onion, strong garlic

S- amino acids

nd-10

0.9

Dimethyl disulfide

Cooked vegetable, strong onion

Cabbage

S- amino acids

nd-22

29

Diethyl disulfide

Strong onion, burnt rubber

S- amino acids

nd-80

4.3

Dimethyl trisulfide

Onion, garlic, meat, green, cabbage

S- amino acids

nd- 0.25

0.2

1Thresholds vary by the matrix of the solution tested. The variation in number indicates different matrix conditions were evaluated.  Thresholds should always be viewed with caution.

2 nd= not detected

The higher sulfide compounds are believed to largely generate from the degradation of the S-containing amino acids. Spiking wines with methionine, cysteine or the cysteine-containing tripeptide glutathione lead to the formation of these compounds in juices and wines. Some of these compounds appear to continue to increase during storage of the wine after yeast activity has ceased suggesting that there are precursor forms present in wine that as the reductive conditions of the wine change generate S-volatiles. In beer for example dimethyl sulfide can form from reduction of Dimethyl sulfoxide derived from S-methylmethionine. This pathway has not been shown to exist in wine and dimethyl sulfide is believed to come from the degradation of cysteine, glutathione, methionine or S-adenosyl–L-methionine. Some higher sulfides may also come from degradation of S-containing pesticides, but this is a rare occurrence and more often these characters derived from catabolism of S-amino acids and their derivatives glutathione and S-adenosyl-L-methionine.

Thiols and Thioalcohols: Mercaptans and thioalcohols can also be found in wine. These components likewise are believed to derive from degradation products of S-containing amino acids and their derivatives or from the interaction of H2S with acetaldehyde, which forms the reactive 1,1-ethanedithiol, and other reactive components in wine. Some of these products are reactive themselves leading to even more diverse S-volatiles. The chemical reactivity of these compounds in combination with a host of potential reactants in wine and very low thresholds of detection has made it challenging to delineate the true pathways by which they are formed.

Thiols and Thioalcohols Found in Wine

Compound

Aroma Description

Source

Concentration in Wine (µg/L)

Putative Thresholda (µg/L)

Methanethiol (Methyl mercaptan)

Cooked cabbage, rotten eggs

S- amino acids

ndb-16

0.2/2/12

Ethanethiol (Ethyl mercaptan)

Onion, rubber, natural gas, fecal

S- amino acids

nd-12

1.1

2-Mercaptoethanol

Poultry, barnyard, solvent

S- amino acids

nd-180

1000

3-(Methylthio)-1-ethanol

Green beans

S- amino acids

nd-70

Methionol (3-(methylthio)-1-propanol)

Potato, cauliflower

S- amino acids

nd- 6300

10-50

4-(Methylthio)-1-butanol

Onion, garlic, earthy

S-amino acids

nd-180

100

2-Methyl-3-furanthiol

Cooked meat

Thiamine degradation

nd-0.300

Furfurylthiol

Roasted coffee, meat, popcorn

Toasted barrel wood extractives and H2S

nd-0.050

0.0004

Benzylthiol

Smoky, flintstone

H2S; Formed during aging

nd-0.015

0.0003

1Thresholds vary by the matrix of the solution tested. The variation in number indicates different matrix conditions were evaluated.  Thresholds should always be viewed with caution.

2 nd= not detected

The mercaptans can form disulfides in wine that can be reduced back to the mercaptan by sulfite or ascorbic acid treatment. The disulfide form does not react with copper and if mercaptans are present in the wine it may be necessary to treat the wine with ascorbate prior to addition of copper to remove the mercaptans along with H2S.

Mercaptoethanol is generally not found in wines above an odor threshold. However this compound is highly reactive and is formed in high levels in synthetic fermentation conditions. Given its reactivity it is not surprising that it is seldom detected as such in wines.

Methionol is the end product of the deamination of methionine via the Ehrlich pathway. Methionol comes from the α-keto acid 3-(methylthio)-propanoic acid. The acid can be decaboxylated and then reduced to an alcohol. In some wines methionol is the major S-containing volatile compound. Detection of its tuber note may be masked by other grape components. 4-methylthio-1-butanol and 2-mercaptoethanol can be formed similarly from homocysteine and cysteine via the Ehrlich pathway.

2-Methyl-3-furanthiol and its disulfide, bis(2-methyl-3-furyl)disulfide are thought to form from the degradation of thiamine. Other thiol compounds arise during barrel aging and seem to be present in higher concentrations if H2S is also present, but the exact mechanisms of synthesis are unknown.

Thioacetates and Thiazoles: Other forms of S-volatiles can also be found in wine. These compounds are derived indirectly or directly from S-containing amino acids or from thiamine.

Thioacetates and Thiazoles Found in Wine

Compound

Aroma Description

Source

Concentration in Wine (µg/L)

Putative Thresholda (µg/L)

Methyl thioacetate

Cheese, rotten vegetables, sulfurous

Methionine, methanethiol + acetylCoA

ndb-20

300 (beer)

Ethyl thioacetate

Cheese, onion, meaty, coffee, burnt, sulfurous

Ethanethiol + acetylCoA

nd-56

40

Benzothiazole

Rubber

Thermal degradation of cysteine, thiamine

nd-14

50/200

2-Methyltetrahydro-thiophene-3-one

Metallic, butane

Thermal degradation of cysteine, methionine, thiamine

nd-167

1Thresholds vary by the matrix of the solution tested. The variation in number indicates different matrix conditions were evaluated.  Thresholds should always be viewed with caution.

2 nd= not detected

Methyl and ethyl thioacetate appear to derive from the reaction of methanethiol and ethanethiol, respectively, with acetyl-CoA. The reaction appears to be catalyzed by alcohol acetyltransferases the same enzymes used for the generation of the family of acetate esters. This acetylation may be a detoxification mechanism for the thiols. High thiol formation has been correlated with release of high levels of H2S by some strains so the appearance of these compounds may indicate conditions of breakdown of S-containing amino acid pools. Their production is also strain dependent. The thioacetates have much higher thresholds of detection than their thiol precursors. Generally the thioacetates are not over their thresholds of detection in most wines. However like all esters they will hydrolyze over time during wine aging leading to the generation of the original thiol and acetate. These compounds can serve as a reservoir of off-characters in wine.

The azoles, benzothiazole and 2-methyltetrahydro-thiophene-3-one, are formed during the thermal degradation of S-containing amino acids and thiamine in the presence of carbonyls. These compounds have also been found in grapes. Thus, they can form from yeast metabolites in heated wines or juices. Benzothiazole is generally found below its threshold of detection with the exception of mousy wines where it can contribute to the off-odor of the wines. The mechanism of formation under these conditions is not known but it may be accelerated due to the production of carbonyls by the lactic acid bacteria.  2-Methyltetrahydro-thiophene-3-one has been found following heating of model solutions containing either methionine, cysteine or thiamine in the presence of dicarbonyls. The exact mechanism of formation in wine is unknown.

Sulfur compound chemistry is complex. Several studies have tried to correlate appearance of these compounds with the formation of H2S. Direct sulfylation by H2S of precursor compounds has not been demonstrated in wine or juice conditions. The appearance of H2S simultaneously with compounds that are degradation products of the S-containing amino acids may simply indicate that degradation is occurring and H2S is being made along with the other derivatives.

Yeast can also degrade S-containing cysteinylated compounds found in grapes forming thiol that are important to the character of several varietals. These compounds are not considered to be off-characters in the wine although if present at very high concentrations they may be. Typically these compounds are desired at the levels found in wines.

Sulfur compounds used as antifungal agents in the vineyard can lead to the formation of S-off-characters in wine. In many regions these compounds are no longer used and more effective and targeted treatments are available. However if an odd off-odor develops in a wine the vineyard should be evaluated as a potential source of the problem.