Vintner's Corner


Vol.16, No. 3 May - June, 2001

Bruce W. Zoecklein

Department of Food Science and Technology

VPI & SU - 0418

Blacksburg, VA 24061, E-mail:

Table of Contents

I. Sulfur Containing Compounds 1

II. Control of Hydrogen Sulfide and Mercaptans in Wine 3

III. Oxygen and Yeast Metabolism 3

IV. Up-Coming Events

Winery Planning and Design Workshop 4

Vineyard Equipment Demonstration and Field Day 4

Crop Estimation, Crop Sampling and Basic Fruit Chemistry Workshop 4

V. Enology Notes 5

I. Sulfur-Containing Compounds

The formation of sulfur containing compounds has been a winemaking problem for as long as wines have been produced. The problem remains, although our knowledge of the nature of the compounds, and the mechanisms influencing their control, are increasing. The following is a general review of reductive tone formation.

The number of factors which can influence the production of sulfur containing compounds like H2S seems unlimited, and include:

Low nitrogen Pantothenate

High nitrogen limitation

Low methionine High fermentation

High methionine rates

High cysteine Low fermentation

High threonine rates

Low gluthathione High elemental

High glutathione sulfur

High sulfite High temperature

Low temperature High metal ion


Source: Bisson (2000)

Volatile sulfur-containing compounds are known to impart distinctive aromas to wines such as rubbery, skunky, or like onion, garlic, cabbage, kerosene, etc. The objectionable odor of hydrogen sulfide, generally described as rotten-egg-like, also has been observed. If no correction is made, hydrogen sulfide may undergo reactions with other wine components to yield mercaptans, which can have detrimental effects on wine palatability and may be difficult to remove.

Hydrogen sulfide contains sulfur in its most reduced, negatively charged form (S-2). Other sulfur containing compounds of interest to winemakers include oxidized forms such as S+4O2 or copper sulfate (CuS+4O4).

Mercaptans are the other principle group of sulfur containing compounds. They all contain this (-SH) group. Ethyl mercaptan possesses a burnt rubber, skunk or garlic-like character. Methyl mercaptan has a sensory characteristic of cooked cabbage. The sensory threshold of both mercaptans is approximately 1 ppb (part per billion).

Mercaptans can oxidize to disulfides when exposed to air. This oxidation not only influences the sensory attributes but influences the ability to bind with copper sulfate (see below). The sensory threshold of disulfide is around 30 ppm.

Sulfur, an essential element for yeast growth, is utilized in the formation of cell components such as protein and vitamins. Available as sulfate (SO42-) in grape juice, it can be reduced to hydrogen sulfide (H2S). As H2S is an integral part of yeast metabolism, it is not possible to completely prevent its formation. However, vineyard management including selection and timing of spray applications and wine processing techniques may effectively minimize its detrimental effects. The following is a review of some of the causes and solutions to the production of sulfur containing compounds.

Elemental Sulfur: Elemental sulfur is used as a fungicide in vineyards throughout the world. Because of increasing awareness of the problems associated with sulfur in winemaking, most viticulturists are using micronized sulfur, which consists of very small particles, ranging from 6 to 8 Ám in size, which are readily miscible in water. An advantage of micronized sulfur is that the application rate is less than one-third the normal dusting sulfur rate for the same measure of fungal control. Only 5 mg/L of elemental sulfur in the must is enough to produce H2S concentrations which cannot be removed. Therefore, sulfur sprays should not occur less than 35 days prior to harvest.

An additional source of elemental sulfur in juice is sulfur candles, used by some vintners to disinfect barrels. These candles may not burn completely, so that unburned sulfur enters the wine or juice. The use of dripless sulfur sticks and/or sulfur cups may effectively overcome this problem.

Redox State and Temperature: Hydrogen sulfide formation also is a function of the oxidation-reduction (redox) state of the must during fermentation. Higher levels of H2S are produced from fermentations carried out in tall (height to diameter) tanks. The design of such fermentors is conducive to a rapid drop in redox potential. The fermentation temperature also affects the overall formation of H2S; generally, less H2S is produced at lower temperatures. However, at lower temperatures, less H2S is lost through entrainment with carbon dioxide.

Yeast and Yeast Physiology: Yeasts differ significantly in their ability to form hydrogen sulfide. However, due to the complexity of factors influencing their production, no strain can be said to be problem free. Some yeasts are known to have deficiencies in their sulfur metabolism that promote increased production of H2S. Such yeasts appear to have an absolute requirement for the vitamins pantothenate and/or pyridoxine (vitamin B6). Although grape juices normally are not deficient in these two vitamins, must treatment, seasonal variations, rot, etc. may result in the depletion of one or both.

Free amino nitrogen (FAN) components of must, therefore, play a role in subsequent H2S formation. Specifically, assimilable free amino nitrogen content is inversely related to H2S levels. Deficiencies in total yeast assimilable nitrogen are not always correlated with the formation of H2S.

Yeast Autolysis: Upon yeast cell death, degradation and rupture of cell membranes release cytoplasmic components including free amino acids, peptides, and polypepetides. Other degradation products include fatty acids, as well as components of the yeast nucleic acids, and vitamins. Yeast autolysate may play a role in the character and complexity of wine. However, the process of sur lie with heavy lees (particularly in the absence of stirring or oxygen) can occasionally result in the production of 'off' flavors and aromas, including H2S and mercaptans. However, if reductive tones were not present at the completion of fermentation, they rarely occur later.

Proper utilization of lees is an important quality and stylistic tool (see Enology Notes #6, The Power of Macro-Molecules).

Sulfur Compounds and Metals: Copper, manganese, and zinc are components of many vineyard fungicides. Late-season application of metal-containing fungicides to the grapes is known to increase the production of H2S and possibly other sulfur containing compounds. Because of the nature of our growing seasons, some are inclined to apply Bordeaux mix fairly late in an attempt to help minimize downy mildew. There is a definite relationship between the use of copper-containing fungicides, like Bordeaux mix, and increased incidences of H2S formation in wines.

Questions regarding how late Bordeaux mix can be applied and how much copper stimulates H2S formation are not resolved. Copper ions are constituents of both certain enzyme systems as well as known inhibitors of respiration. Yeast grown in the presence of copper adopt a protective mechanism of H2S formation and consequently copper sulfide formation. The advantage of late season copper sprays for mildew control must be balanced with concerns for the production of sulfur containing compounds. If late season fungicides are applied, juice settling of whites and the addition of a yeast nutrient is advisable.

The pre-fermentation addition of sulfur dioxide can impact H2S production. High initial levels of added sulfur dioxide bind acetaldehyde, which is normally reduced to form ethanol. If not enough acetaldehyde is present, juice sulfates may be instead reduced, forming H2S. Additionally, SO2 can convert H2S to elemental sulfur which may later be reduced back to H2S.


II. Control of Hydrogen Sulfide and Mercaptans in Wine

A key to minimizing H2S formation is the maintenance of assimilable nitrogen in the fermentor and avoiding yeast stress. It is recommended that the N status of the juice be tested prior to fermentation (see Formol Analysis on my web site at If supplementation is required it is best to do it in two stages - first an addition of a nutrient such as Fermard K, Superfood, etc., followed by the addition of DAP after the fermentation has begun. (see Vintner Corner Vol. 14, No. 4 posted on our web site.)

Formation of excessive H2S in white wines often can be minimized by settling, centrifuging, or filtering the juice prior to fermentation. These practices all accomplish the same goal, that is, removal of high-density solids along with associated elemental sulfur and possibly metals when present. In the case of red wine fermentation, some winemakers deal with excessive H2S by aeration at first racking, thus volatilizing the H2S. Increased H2S production will occur, however, if aeration is carried out during or too soon after the completion of alcoholic fermentation. In these cases, elemental sulfur is believed to act as a hydrogen acceptor, forming H2S.

Coincidental with H2S formation are increases in yeast populations arising as a result of transient exposure to oxygen.

Additional techniques for controlling H2S include sparging problem wines with nitrogen gas shortly after the completion of alcoholic fermentation. This practice may be relatively effective in eliminating minor quantities of H2S, but desirable volatile wine components also may be swept away during excessive sparging. In cases where methyl mercaptan appears to be the problem carefully controlled aeration may bring about oxidation of methyl mercaptan to the less objectionable compound dimethyl disulfide.

Winemakers can remove objectionable H2S and mercaptans from a 'still' (non-fermenting wine) wine by direct contact with copper. The addition of 4 g copper (II) sulfate (CuSO4 5H20) per 1000 gallons raises the copper content by 0.2 mg/L. Although governmental regulations permit additions of up to 0.5 mg/L (as copper), residual levels in the wine cannot exceed 0.2 mg/L (as copper).

It should be noted that although mercaptans react with copper, dimethyl disulfide does not. Thus if the wine in question has undergone any oxidation, it may be necessary to reduce dimethyl disulfide back to the reactive species, methyl mercaptan. This can be accomplished by addition of ascorbic acid (see Zoecklein et al., 1995). Generally, addition levels of 50 mg/L or more of ascorbic acid are used, and such additions usually are made several days prior to the addition of copper. (The sulfur dioxide analysis by Ripper titration cannot be performed accurately in wines containing ascorbic acid, as the latter also reacts with the iodine titrant). Copper should not be added until the fermentation is complete and the yeast titer reduced by racking, filtration, and so on. Yeast cells will bind copper ions to cell surfaces and may reduce reactivity with H2S.

Lastly, the addition of SO2 to still wines may reduce H2S levels. The addition results in an SO2-induced oxidation of H2S to yield elemental sulfur, which after precipitation, may be removed by centrifugation or filtration.

SO2 + 2H2S - 3SO + 2H2O


III. Oxygen and Yeast Metabolism

Oxygen plays important roles in the physiological status of yeast. Molecular oxygen is required for the synthesis of lipids and steroids needed for functional yeast cell membranes. Steroids play a structural role in membrane organization, interacting with and stabilizing the phospholipid component of the membrane which helps to protect the membrane against alcohol. It has been shown that yeasts propagated aerobically contain a higher proportion of unsaturated fatty acids and up to three times the steroid level of conventionally prepared cultures. This increase correlates well with improved yeast viability during the fermentative phase.

As fermentation begins, oxygen present in must is rapidly consumed, usually within several hours. After utilization of initial oxygen present, fermentations become anaerobic. Because yeasts are not able to synthesize membrane components in the absence of oxygen, existing steroids must be redistributed within the growing population. Under such conditions, yeast multiplication is usually restricted to 4 or 5 generations, due largely to diminished levels of steroids, lipids and unsaturated fatty acids. This can result in stuck fermentation.

Methodology of starter propagation is important with respect to subsequent requirements for oxygen. Aerobic propagation has been demonstrated to significantly enhance subsequent fermentative activity. Yeast populations reach higher final cell numbers and fermentations proceed at a faster rate and are more likely to complete.

The group of enzymes responsible for catalyzing oxidative reactions in juice are the polyphenol- oxidases, also referred to as phenolases or tyrosinases. Polyphenoloxidases catalyze the oxidation of dihydroxyphenols to their corresponding quinones (brown products shown below).

Traditionally, winemakers added sulfur dioxide to inhibit this reaction. Currently, sulfur dioxide is generally not added until long after the fermentation is completed. As the above reaction proceeds, phenols polymerize and precipitate from solution. The result is a wine with a lower tannin level and, frequently in the case of whites, a finish that is less harsh and less astringent.

The current popularity of not adding sulfur dioxide prior to fermentation may be contributing to sluggish and/or stuck fermentations. As stated, unsaturated fatty acids and sterols help to protect the yeast cell membrane from the effects of alcohol. These can only be produced if molecular oxygen is present. Therefore, winemakers should have some air present during the first 30-72 hours of the fermentation. If no SO2 is present, the grape's polyphenyloxidase will greatly reduce the oxygen available for the yeast (see reaction above). Sulfur dioxide will inactivate the enzyme and thus allow a greater concentration of oxygen to be available for yeast lipid production. As a reducing or antioxidizing agent, sulfur dioxide can react or bind with oxygen. This reaction, however, is relatively slow and much slower than polyphenoloxidase utilization of available oxygen.

Some wineries are adding a small amount of SO2 (15-20 mg/L total) before fermentation of white wines to help avoid oxygen depletion. Oxygen should be considered an essential 'nutrient' required for proper yeast cell growth. Its management in the initial stages of fermentation may be an important factor in determining if a wine will complete fermentation.


IV. Up-Coming Events

Winery Planning and Design Workshop

Saturday, July 21, 2001, in Winston Salem, NC

The North Carolina Winegrowers Association and the Enology-Grape Chemistry Group at Virginia Tech is sponsoring this workshop. It will include winery business planning and economics, winery design considerations including gravity flow and caves, equipment, selection, refrigeration and winery utilities, government compliance issue, etc.

There is limited capacity and pre-registration is required. Registration fee is $115. Make checks payable to North Carolina Winegrowers Association, P.O. Box 27647, Raleigh, NC 27611. For details check or email/call Chanel McIntyre at or (919) 733-7136.

Vineyard Equipment Demonstration and Field Day

Saturday, July 28, 2001 at Indian Springs Farm and Vineyard, Woodstock, VA. See Dr. Wolf's newsletter for additional information.

Crop Estimation, Crop Sampling and Basic Fruit Chemistry Workshop

Thursday, August 2, 2001; 10:00 a.m. until 3:00 p.m., Ivy Creek Farms, Ivy, VA

Friday, August 3, 2001; 10:00 a.m. until 3:00 p.m. AHS Jr. Agricultural Research and Extension Center, Winchester, VA

The objective of this program is to provide growers with the skills to estimate crop and to do basic lab and sensory analyses to judge crop maturity. The morning program (10:00 a.m. until noon) is open to all and has no registration cost associated with it. Dr. Tony Wolf will lead a discussion of grape crop estimation procedures. The afternoon program (1:00 to 3:00 p.m.) is strictly limited to 16 participants, and pre-registration is required to participate. Dr. Bruce Zoecklein will conduct a hands-on lab analysis of fruit pH, titratable acidity, and Brix, with sensory assessments of tannin maturity and juice aroma. The restriction to 16 participants each day is required due to a limitation of lab apparatus and space.

See Dr. Tony Wolf's newsletter for registration information.


V. Enology Notes

Enology notes are periodic email communications discussing various topics. The subjects reviewed are generally not the same as those covered in the Vintner's Corner newsjournals. If you are not receiving the Enology Notes electronic mailing and would like to be added, send me your email address. Enology notes and the Vintner's Corner newsjournals are also posted on my web site at



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Bisson, L. 2000. Proceeding of Taint Necessary 50: Detecting and Indentifying Off-Flavors in Wine.

Dukes, B.C. and Butske, C.E. 1998. Rapid determination of primary amino acids in grape juice using a o-pthaldi-aldehyde/N-acetyl-L-cysteine spectophotometric assay. Am. J. Enol. Vitic. 49:125-134.

Henschke, P.A. and Jiranek, V. 1993. Yeasts - Metabolism of Nitrogen Compounds. In: Wine Microbiology and Biotechnology. Graham H. Fleet (ed.). Harwood Academic Publishers. pp. 77-164.

Monk, P. 1994. Nutrient requirements of wine yeast. Practical Winery and Vineyard. July/August p. 24.

Ough, C.S. 1969. Substances extracted during skin contact with white must. I. General wine composition and quality changes with contact time. Am. J. Enol. and Vitic. 20:93-100.

Radler, F. 1965. The main constituents of the surface waxes of varieties and species of the genus Vitis. Am. J. Enol. and Vitic. 16:159-167.

Sponholz, W.R. 1991. Nitrogen compounds in grapes must and wine. In: Proceedings of the International Symposium on Nitrogen in Grapes and Wine. Seattle, WA. Am. J. Enol. Vitic. J. Rantz (ed.) pp. 67-77.

Vos, P.J.A., Crous, E., and Swart, L. 1980. Fermentation and the optimal nitrogen balance of musts. Wynboer. 582:58-62.

Zoecklein, B.W., Fugelsang, K.C., Gump, B.H. and Nury, F.S. 1995. Wine Analysis and Production. Chapman & Hall. Thomson Publishing.