1. Sulfur Compounds in Wine
Linda Bisson
Department of Viticulture and Enology

2. Introduction to S-Containing Faults

3. Why Are Sulfur Compounds a Problem?
Low thresholds of detection
Negatively-associated aromas
Chemical reactivity
Difficulty in removal
Difficulty in masking

4. The Classic Sulfur Fault Descriptors
Rotten egg
Fecal
Rubber/Plastic tubing
Burnt match
Burnt molasses
Burnt rubber
Rotten vegetable: cauliflower, cabbage, potato,
asparagus, corn
Onion/Garlic
Clam/Tide pool
Butane/Fuel/Chemical

5. The Sulfur Taints Hydrogen sulfide Higher sulfides Mercaptans
Dimethyl (Diethyl) sulfide
Dimethyl disulfide
Mercaptans
Methyl (Ethyl) mercaptan
Thioesters
Methyl (ethyl) thioacetate
Other S-amino acid metabolites
Thioethers
Cyclic and heterocyclic compounds

6. Sources of Sulfur Compounds
Non-biological
Elemental sulfur
S-containing pesticides
Biological
Sulfate/Sulfite reduction and reduced sulfide reactions
S-containing amino acid metabolism
S-containing vitamins and cofactors degradation
Glutathione metabolism and degradation
S-containing pesticides degradation

7. Timing of Sulfur Fault Formation
Primary Fermentation Early: Hydrogen Sulfide
Primary Fermentation Late: Hydrogen Sulfide
Post Fermentation: Hydrogen Sulfide or Sur Lie Faults
Bottling: S-fault development

8. Hypotheses to Explain S-Taint Formation
Correlated with H2S formation during the primary fermentation
Correlated with late H2S formation (peak 2) but not with H2S formation during primary fermentation
Associated with S-containing amino acid levels during primary fermentation
Due to degradation of S-containing metabolites during yeast lees aging, but not related to levels of these compounds present in the initial juice
Yeast strain most important
Juice composition most important

9. Problems with Previous Studies
Lack of control of all variables
Invalid comparisons (too many variables)
Confounding factors not considered to be important
Differences in strains and conditions used
Driving reactions by having an excess of precursors, beyond anything found in juices or wines

10. Hydrogen Sulfide

11. Why is H2S formed? Off-shoot of metabolism Reductive environment
Signaling molecule

12. Hydrogen Sulfide Formation: Off-Shoot of Metabolism
Due to release of reduced sulfide from the enzyme complex sulfite reductase
Reduction of sulfate decoupled from amino acid synthesis
Sulfate reduction regulated by nitrogen availability
Lack of nitrogenous reduced sulfur acceptors leads to excessive production of reduced sulfate and release as H2S
Also a stress response
Strain variation

13. Stress Response: Reduction Pathway Remains Operational
Need cysteine for glutathione (tripeptide cytoplasmic redox (electron) buffer
Need methionine for S-adenosylmethionine and one carbon transfers needed for ethanol tolerance

14. Sulfate Reduction Pathway
SO4
SUL1, SUL2
SO4
MET3
Adenylylsulfate
MET14
H2S
Phosphoadenylylsulfate
MET16 (1,8,20,22)
Sulfite
MET10 (1,5?,8,20)
Sulfide
MET17/25/15
Cysteine Cystathionine Homocysteine Methionine
CYS3
CYS4
MET6

15. Hydrogen Sulfide Formation: Reductive Environment
Biological energy is obtained from recapture of light (carbon bond) energy, from proton movements and from electron movements
Cell is dealing with an excess of electrons that exceeds buffering capacity
Many electrons can be used to reduce a single sulfate molecule restoring the proper balance of cytoplasmic electrons

16. Hydrogen Sulfide Formation: Reductive Environment
Tank dimensions leading to stratification of electron gradients
Settling of yeast cells
Chemical composition of juice
Oxygen level and content of juice

17. Hydrogen Sulfide Formation: Signaling Molecule
Hydrogen sulfide coordinates population metabolic activities: shuts down respiration in favor of fermentation, coordinating population of cells in fermentation
Hydrogen sulfide inhibits respiration of a variety of organisms: allows more rapid domination of fermentation
Explains selective pressure for high sulfide producers in the wild

18. Current Understanding of H2S Formation
Nitrogen levels not well-correlated with H2S formation, but generally see increased H2S at lower nitrogen
Tremendous strain variation in H2S production
Can get H2S with high nitrogen
Get more H2S with higher solids content
Get more H2S with unsound fruit

19. Factors Impacting H2S Formation
Level of total nitrogen
Level of methionine relative to total nitrogen
Fermentation rate
Use of SO2
Vitamin deficiency
Presence of metal ions
Inorganic sulfur in vineyard
Use of pesticides/fungicides
Strain genetic background

20. Timing of Formation of H2S
Brix
Time

21. Timing of Formation of H2S
Early (first 2-4 days): due to N/vitamin shortage, electron imbalance, signaling
Late (end of fermentation): due to degradation of S-containing compounds
Sur lie (post-fermentation aging): due to autolysis
In Bottle: screw cap closures: return from an altered chemical form

22. Higher SulfideS

23. Higher Sulfides Emerge late in fermentation and during sur lie aging
Release of compounds during entry into stationary phase by metabolically active yeast
Come from degradation of sulfur containing amino acids
Biological
Chemical
From reaction of reduced sulfur intermediates with other cellular metabolites?
Formed chemically due to reduced conditions?
Degradation of cellular components: autolysis

24. Volatile Sulfur Compounds
Methanethiol: CH3-SH
Ethanethiol: C2H5-SH
Dimethyl sulfide: CH3-S-CH3
Dimethyl disulfide: CH3-S-S-CH3
Dimethyl trisulfide: CH3-S-S-S-CH3
Diethyl sulfide: C2H5-S-C2H5
Diethyl disulfide: C2H5-S-S-C2H5

25. Sources of Higher Sulfides
S-Containing Amino Acids
S-Containing Vitamins and Co-factors
Glutathione (Cysteine-containing tripeptide involved in redox buffering)

26. Defining Metabolic Behaviors Resulting in Taint Formation
S-amino acid catabolism
Vitamin/Co-factor interactions and metabolism
Glutathione turnover and reactions
Metabolic roles of sulfate reduction

27. Defining Metabolic Behaviors Resulting in Taint Formation
S-amino acid catabolism
Degradation of methionine and cysteine: methional and methionol
Chemical reaction products of methionine and cysteine: stress resistance
Influence of wine composition and chemistry on yeast behavior
Vitamin/Co-factor interactions and metabolism
Glutathione turnover and reactions
Metabolic roles of sulfate reduction

28. Defining Metabolic Behaviors Resulting in Taint Formation
S-amino acid catabolism
Vitamin/Co-factor interactions and metabolism
Role of thiamin
Role of S-adenosylmethionine
Glutathione turnover and reactions
Metabolic roles of sulfate reduction

29. Defining Metabolic Behaviors Resulting in Taint Formation
S-amino acid catabolism
Vitamin/Co-factor interactions and metabolism
Glutathione turnover and reactions
Role in stress response: prevention of oxidative damage
Impact of nitrogen level on metabolism
Biological turnover of ‘reacted’ glutathione
Metabolic roles of sulfate reduction

30. Defining Metabolic Behaviors Resulting in Taint Formation
S-amino acid catabolism
Vitamin/Co-factor interactions and metabolism
Glutathione turnover and reactions
Metabolic roles of sulfate reduction
Stress response:
Prevention of oxidative damage
Role in ethanol tolerance
Environmental/metabolic detoxification
Banking on reactivity to inactivate a toxic substance
Metabolic demands

31. Understanding the Interface between Metabolite Production and Wine Chemistry and Composition
What environmental conditions impact S-compound metabolic activities?
Separating a biological response from a chemical one
Control the metabolites
Control the chemistry

32. Sulfur Compound Flight #1 Spiked Compounds
Glass 1: Control Wine (Cabernet Sauvignon) Glass 2: Hydrogen sulfide H2S Glass 3: Dimethyl sulfide CH3-S-CH3 Glass 4: Dimethyl trisulfide: CH3-S-S-CH3 Glass 5: Diethyl sulfide: C2H5-S-C2H5 Glass 6: Diethyl disulfide: C2H5-S-S-C2H5

33. Sulfur Compound Flight #1 Spiked Compounds
G 1: Control Wine (Cabernet Sauvignon)
G2: Hydrogen sulfide: rotten egg
G 3: Dimethyl sulfide: cabbage, cooked corn, asparagus, canned vegetable
G 4: Dimethyl trisulfide: meaty, fishy, clams, green, onion, garlic, cabbage
G 5: Diethyl sulfide: garlic, onion
G 6: Diethyl disulfide: overripe onion, greasy, garlic, burnt rubber, manure

34. Sulfur Compound Flight #2: Taints produced late in fermentation
Glass 1: Control Wine (Cabernet Sauvignon) Glass 2: Ethanethiol Glass 3: Mercapto -2- methyl propanol (Methionol) Glass 4: Methyl thiopropionaldehyde (Methional) Glass 5: Mercapto-3-methyl butanol Glass 6: BM45 French Colombard

35. Sulfur Compound Flight #2 Spiked Compounds
G 1: Control Wine (Cabernet Sauvignon)
G 2: Ethanethiol: onion, rubber, natural gas
G 3: Methionol: cauliflower, cabbage, potato
G 4: Methional: musty, potato, onion, meaty
G 5: Mercapto-3-methyl butanol: meaty
G 6: French Colombard: reduced

36. BM 45: Isolated in Montalcino
Produces high polyphenol reactive polysaccharides = mouth feel
Has high nitrogen requirements and can produce H2S
Aroma characteristics: fruit jam, rose, cherry, spice, anise, cedar and earthy