Wine preservative SO2 levels: review of SO2 binders and the important effect of malolactic fermentation

Niveles de SO2 como conservante: revisión de los compuestos ligados a SO2 y papel relevante de la fermentación maloláctica
Nick Jackowetz,1 Erhu Li1 y Ramón Mira de Orduña2
1Graduate Research Assistant. 2 Associate Professor
Dept. of Food Science, NYS Agricultural Experiment Station
College of Agriculture and Life Science
Cornell University, NY, USA

Except for alcohol, sulphur dioxide (SO2) is the only component that prompts a warning statement on wine labels, although fining proteins potentially present in wine are under regulatory review as allergens in some countries. SO2 has many useful functions in winemaking. It can serve as enzyme inhibitor in musts to prevent juice browning and oxidation, as a microbiological control agent in musts and wines, and to prevent the oxidation of finished wines1. It also has the ability to bind to undesirable volatile compounds, such as acetaldehyde, thus reducing their sensory impact.

Unfortunately, SO2 also is an irritant and can have negative health effects on sensitive consumers. It is estimated that up to 1% of the population has an increased sensitivity to sulfites2, and other studies suggest that 5% of asthmatics may risk adverse reactions upon ingestion of SO23,4. However, it could be argued that considering its historic utilization in a number of foods the amount of health related data available is scarce.

Our laboratory has studied the microbial metabolism of carbonyl compounds since 2000 with the aim of controlling and reducing the use of SO2 5-12. In this article, we provide an overview of the results and information for winemakers interested in minimizing the production of these SO2 binding compounds and final SO2 levels.


Legal limits for SO2 and consumer perceptions

The importance placed on SO2 may lie in the overall negative connotations associated with its use, even though it has a long record of utilization as a preservative in a number of foods and is perfectly safe for the majority of consumers. The remarkable increase in the number of organic wineries as well as “sustainable” or “fresh” wines with lower SO2 concentrations may also be indicative of the increasing consumer interest in minimally processed wines.

How do legal SO2 limits compare internationally? The European Union (EU) has further reduced maximum limits for total SO2 in red and white wines to 150 and 200 mg/l, respectively (EC 606/2009, Annex I B). However, these limits apply only to dry wines (<5 g/l of combined glucose and fructose).  Numerous exemptions are listed in an additional four pages in this regulation, permitting up to 400 mg/l of total SO2 in some wines—which is greater than the maximum of 350 mg/l allowed in any wine according to U.S. regulations (27 CFR 4.22(b)(1). Among the laws of the countries surveyed, lowest limits exist in South Africa (see Table 1).

Table 1: Legal limits for total SO2 in major winemaking nations(in mg/l)1

Country / Zone

Wine type, RS


Legal Reference / Description




27 CFR 4.22(b)(1)


<35 g/l sugars


ANZFSC 4.5.1: Clause 5(5)(a)

>35 g/l sugars



New Zealand

<35 g/l sugars



>35 g/l sugars



White / Rosé, < 5 g/l
EC Nº 606/2009, Annex I B
Red, < 5 g/l sugars
White / Rosé > 5 g/L
Red > 5 g/l sugars
Specific wines
Eg., Spätlese (can be dry), Bordeaux Sup., Côtes de Bordeaux, C. de Bergerac, Navarra, Penedès, several French VdP and Hungarian and some Greek sweets
Specific wines
Eg.: Auslese (can be dry), sweet wines from Romania, Czech Rep., Slovakia and Slovenia
Specific wines
Eg. : Beerenauslese, TBA Eiswein, French sweet wines such Sauternes, Barsac, etc., sweet Greek with > 45 g/l sugars, sweet Eastern European wines
Canadian Food & Drug Reg. B.02.100
Prevention of Foof Adulteration Act & Rules, Appendic C, Table 3
All (>1%)
Japan's Specifications and Standards for Foord Additives
White, < 5 g/l sugars
Liquor Products Act 60 of 1989 Regulations. Regulation 32 (Table 8)
Red, < 5 g/l sugars
All, > 5 g/l sugars
Specific wines
Eg.: noble late harvest and naturally dried

1) Information retrieved from FIVS-Abridge database (
2) Unit is mg/kg.
3) Canada prescribes a maximum of 70 mg/l free or 350 mg/l combined SO2.

Certainly, it is not particularly useful to consider SO2 levels across regions with different climatic conditions. Cool climate wines may require some residual sugar for balance - also, growing seasons may suffer from humid summers, and adverse weather conditions during harvest can lead to an additional SO2 burden. For example, the 2006 harvest in Eastern France and Germany was characterized by rains and fruit with a high degree of rot. In the final wines, bound SO2 levels (especially due to pyruvic acid and acetaldehyde) were significant and, consequently, Germany and Alsace requested a 40 mg/l increase in total SO2 maxima for the 2006 vintage, which was granted (EC 423/2008 Article 23(4) and Annex XV) as it had been for the 2000 vintage.

SO2 binders, their occurrence in wines, and their analysis

For any given amount of free SO2, the final SO2 concentrations depend on the concentration of SO2 binders. Compounds with carbonyl functions bind to SO2. Relevant SO2 binders in wine include glucose, acetoin, diacetyl, galacturonic, a-ketoglutaric and pyruvic acids, and especially acetaldehyde. A number of methods to quantify SO2 binding compounds in wines are available 13-17. Individual analysis typically is tedious and cumbersome. In our laboratory, major SO2 binders are determined simultaneously using High Performance Liquid Chromatography (HPLC). However, the most important SO2 binders—acetaldehyde, pyruvic and a-ketoglutaric acids—can also be determined with relatively cheap enzymatic tests using a standard spectrophotometer.

In recent years, we have analyzed numerous wines for major SO2 binders. In reds, galacturonic and a-ketoglutaric acids were found in higher concentrations than in whites. In contrast, white wines contained more pyruvic acid and acetaldehyde than reds (Table 2).

Table 2: Average concentrations (mg/l) of several SO2-binding compounds in white and red wines from New York State. A total of 237 wines were analyzed.

Wine Type


Galacturonic Acid








Using this data we could calculate that the most relevant SO2 binding compounds were acetaldehyde, pyruvic, a-ketoglutaric and galacturonic acids because of their binding properties and their concentrations in wines. In white wines, acetaldehyde typically accounts for over 70% of the bound SO2, followed by pyruvic (17%) and a-ketoglutaric (8%) acids. In reds, acetaldehyde was found to account for over 50% of bound SO2 followed by a-ketoglutaric (>20%), pyruvic (12%) and galacturonic (10%) acids. In sweet wines, glucose may also be relevant for SO2 binding.

Because of its importance for bound SO2, the formation and degradation of acetaldehyde is particularly relevant.

Acetaldehyde in wines, its role, formation, and degradation

Acetaldehyde is the most important volatile wine carbonyl and can be formed both biologically (through yeast activity) and chemically (by wine oxidation). It is a small and highly reactive molecule with a green grass, apple-like or nutty aroma.


A common misconception is that the risk of acetaldehyde formation only begins with the end of alcoholic fermentation. However,  to a large extent, acetaldehyde found in wine stems from yeast activity. Enological yeast, including commercial Saccharomyces cerevisiae, excrete acetaldehyde during the initial phases of alcoholic fermentation10,11,18. After reaching a peak value, acetaldehyde is then re-utilized to a certain degree (Figure 1). Typical residues found  in Gewürztraminer and Riesling after alcoholic fermentation ranged from 22-49 mg/l of acetaldehyde 11.


Figure 1

Figure 1: Formation of acetaldehyde (ACHO in mg/l) during alcoholic fermentation by Saccharomyces cerevisiae. ∀:, acetaldehyde; -, growth; 8: glucose; Χ: fructose.


Chemical formation of acetaldehyde relies on exposure to oxygen, the presence of transition metals such as copper and iron, and phenolics19,20. In fact, if wines are exposed to atmospheric oxygen after alcoholic and malolactic fermentations and removal of yeast lees, significant amounts of acetaldehyde may be formed from the oxidation of ethanol. Figure 2 shows the average changes in acetaldehyde concentrations during vinifications of 16 wines in 8 wineries of the 2009 vintage. Post-fermentative ageing, rackings, transfer, filtration and bottling operations contributed to acetaldehyde increases.

Figure 2

Figure 2: Average changes of acetaldehyde concentrations between winemaking steps during vinifications in eight New York State wineries in 2009. Averages from eight wineries ±SE displayed. Following the end of AF sample, all other values were obtained from samples taken after the indicated winemaking step.



In addition to the partial re-utilization of acetaldehyde by yeast in the second half of the alcoholic fermentation (Figure 1), acetaldehyde is also degraded by lactic acid bacteria7,9. Typically, acetaldehyde is depleted during malolactic fermentation, either simultaneously with the degradation of L-malic acid, or some days later (Figure 3).

Figure 3

Figure 3: Degradation of malic acid (g/l) and acetaldehyde (mg/l)during malolactic fermentation by Oenococcus oeni at pH 3.3 and 3.6 in Chardonnay. : acetaldehyde -, growth; 8, malic acid.


Accordingly, if a complete degradation of acetaldehyde is desired, wines should not be stabilized until five days after malic acid depletion. More importantly, malolactic fermentation has been shown to cause a substantial reduction of pyruvic acid, and the partial reduction of a-ketoglutaric acid, as well (Table 3). Hence, malolactic fermentation offers a very significant contribution towards achieving lower bound and total SO2 levels.

Table 3: Percent degradation of SO2 binding compounds by 12 strains of wine lactic acid bacteria (Oenococcus oeni) during malolactic fermentation (average values represented)

SO2 Binding Compounds

% degradation









Galacturonic Acid

Major factors influencing acetaldehyde residues in wines

Both biological and chemical factors are important in determining the level of acetaldehyde in wines.

Biological factors

Acetaldehyde residues will be higher, if (1) the initial formation is increased and/or (2) its re-utilization is reduced. A factor that affects initial formation is the yeast strain. Saccharomyces cerevisiae strains that predominate in alcoholic fermentation, regardless of whether inoculated or not, tend to produce more acetaldehyde than most non-Saccharomyces cerevisiae yeast, with some exceptions: For example, a strain of Schizosaccharomyces pombe was found to excrete large amounts. Among Saccharomyces cerevisiae strains, the acetaldehyde excretion and re-utilization was not identical, but fairly uniform compared to non-Saccharomyces cerevisiae10.

The most important factor for the biological formation of acetaldehyde is, however, the addition of SO2 to musts (Figure 4). Yeast produce more acetaldehyde in response to SO2 additions. Across several studies with more than 20 yeast strains, we found that the addition of 1 mg/l of SO2 increases the final acetaldehyde residue after alcoholic fermentation by 0.2-0.5 mg/l. Hence, a must SO2addition of 50 mg/l will increase final acetaldehyde levels by 10-25 mg/l and, thus, increase bound SO2 levels by 15-37 mg/l 10.

Figure 4

Figure 4: Acetaldehyde formation and re-utilization during AF by Saccharomyces cerevisiae with 50 mg l-1 of SO2 added to the must before AF (-) and during AF with no SO2 added (∀)

The re-utilization of acetaldehyde by yeast in the late phase of alcoholic fermentation was found to be improved by factors that maintain a large number of viable yeast. Accordingly, adding yeast nutrients and maintaining moderate temperatures (20°C) led to reduced acetaldehyde residues, while maintaining cool temperatures (12°C) throughout fermentations and not adding any nutrients led to larger residues 11. Early racking of yeast lees also reduces acetaldehyde re-utilization and leads to higher bound SO2 levels 11.

The potential of malolactic fermentation to reduce acetaldehyde and other SO2 binders is significant. However, malolactic fermentation can be difficult to achieve in some wines, especially those, which would most benefit from it. Among the inhibiting factors for malolactic bacteria, the alcohol and free SO2 concentrations are know to be crucial, as well as the temperature, the pH and the availability of nutrients. Less attention has been given to bound SO2 levels. Recent results obtained by our group clearly demonstrate that malolactic bacteria are inhibited by acetaldehyde-bound SO2 (acetaldehyde hydroxysulfonate) even in the complete absence of free SO2 (Table 4).


Tabla 4: Duration of lag-phase, and depletion of acetaldehyde-bound SO2 and malic acid during malolactic fermentation by a commercial strain of Oenococcus oeni in the presence of various concentrations of bound SO2

Acetaldehyde-bound SO2 concentration [mgl1]


30 60 90

Duration of lag phase and time-point of malic acid/acetaldehyde depletion [days]



8,2 12,8 18,7

Malic acid depletion


10.6 14.9 27
Bound SO2 depletion
10.4 27 30.8

This observation is critical for a winemaking approach interested in reducing SO2. Large must SO2 additions will result in large final bound SO2 levels after alcoholic fermentation. These, may then prevent any further reduction of SO2 binders by malolactic fermentation.

Currently, malolactic fermentations remains the single most relevant tool to reduce bound SO2 concentrations in wines. In the future, new methods for the reduction of SO2 binding compounds may be available. Bordeaux researchers have patented a method that would allow the reduction of SO2 binding carbonyls in wines using an insoluble resin. The method was especially targeted at Sauternes, which tend to suffer from high SO2 binder concentrations. So far, the method has not been scrutinized by the scientrific community, and it is not currently permitted 21-23.

Main Points

  • For a given concentration of free SO2, the total SO2 content depends on bound SO2 and hence, the concentration of SO2 binders. Acetaldehyde and pyruvate are the most important SO2 binders in most wines.
  • The most important SO2 binder in wines is acetaldehyde. Pyruvate, a-ketoglutarate and galacturonic acid (in reds) may also be relevant.
  • Acetaldehyde may be formed biologically (by yeast at the start of alcoholic fermentation) and chemically (mainly after alcoholic fermentation when wines are not protected from oxidation by atmospheric oxygen).
  • Yeast will degrade acetaldehyde during later stages of alcoholic fermentation if they are still viable and remain in contact with the wine.
  • Malolactic bacteria will degrade acetaldehyde and pyruvate significantly during malolactic fermentation.
  • All winemaking steps (racking, pumping, filtration, bottling) after the end of alcoholic or malolactic fermentation can  lead to increased acetaldehyde levels.
  • Addition of SO2 in the presence of active yeast will lead to the formation of SO2 binders. For every 10 mg/l of SO2 added to the must, bound SO2 levels in the final wine will increase by 3-7 mg/l.



Funding for these projects was provided by the New York Wine and Grape Foundation, Nolan and Canandaigua Wine Co. Endowment Funds, USDA Federal Formula Funds, Lallemand Inc,. and the China Scholarship Council (State Scholarship Fund No. 2008630061).



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