Factors, Considerations and Recommendations to Produce a Healthy Fermentation

Factors/Considerations Recommendations/Comments
Vineyard       Fermentation problems are often vineyard specific. Nitrogen deficiency in apparently heathy grapes can be very severe. Drought, grape vine nutrient deficiencies, high incidences of fungal degradation and fruit maturity all influence must nitrogen and vitamins, as do cultivar, rootstock, crop load, season and winemaking practices. For example, some varieties have a greater tendency towards deficiency such as Chardonnay. Some rootstocks produce more nutritious grapes than others with regard to total nitrogen. For example, grapes on St. George are much higher than those on AXR1.
      The concentration of alpha amino nitrogen in grapes changes as a function of maturity and crop load. The concentration increases with fruit maturation and decreases with crop load.
      Table 2 from Henick-Kling et al. (1996) shows the concentrations of the two important sources of assimilable N in six cultivars at fruit maturity for two seasons. This data illustrates several important points. There are large variations from one season to the next in both free ammonia and free amino nitrogen. There is also a significant difference in the concentrations of both sources of N among cultivars. The amount of assimilable N required for fermentation is greater than 200 mg/L. As can be seen in Table 2, none of the varieties listed has an adequate concentration.
      As reported previously, there is a simple, effective method of determining assimilable N in juice and wines. This procedure requires only a few reagents and a pH meter and is certainly within the capability of all wine producers. The specifics of this analysis are available on this website or by contacting my office.
Yeast Preparation       Hydration of fresh culture in warm water at exactly the suppliers stated temperature is critical for maximum viability. A large percentage of the cells die if rehydration is done at cooler or warmer temperatures, resulting in a significant loss of cell viability. After rehydration the yeast should be added to the must within 20-30 minutes or a source of sugar added to the culture. If this is not done, cells go into a premature decline phase resulting in an inoculum of low cell concentration.
      It is imperative that one avoids temperature shock (no more than 5-7C differential between culture and must temperature). Temperature shock kills great numbers of yeasts. For example, adding a yeast culture at 104F/40C to a must at 60F/15C kills about half the cell population.
Yeast Strains       There are large differences among strains in ability to ferment to dryness and their nitrogen requirements.
Yeast population       Yeast population should be large enough to overwhelm indigenous micro-flora and grow to 2 to 5 x 106 yeast cells/mL of must (1 to 3% vol/vol of an active starter). These concentrations apply when the Brix is below 24, the pH is above 3.1, and the temperature is above 55F.
      Increases in the inoculum volume should be made when the must is outside these parameters.
Nutrient Addition       Many musts lack sufficient N, vitamins and other ingredients needed by yeasts during their growth phase for healthy fermentations. As suggested, levels of greater than 200 mg/L fermentable nitrogen are required for heathy fermentations. Supplementation should be made using a balanced source of DAP (diammonium phosphate), amino acids, minerals, and vitamins.
      DAP addition of 1 g/L (8.3 lb/1,000 gal) provides about 258 mg/L fermentable N. This is greater than the suppliers recommended level. In the US the legal limit of DAP is 960 mg/L which is equal to 208 mg N/L. Fermaid K at 2 lb/1000 gal (25 g/hL) = 25 mg N/L while DAP at 2 lb/1000 gal (25 g/hL) = 50 mg N/L. As indicated, fermentable nitrogen concentration in juice or wine can be easily estimated (Zoecklein et al., 1995).
Timing       Amino acids are not taken up equally by the yeast cell - some are utilized at beginning of the growth cycle, some later, and some not at all. Ammonia, on the other hand is consumed preferentially to amino acids. Therefore, timing of DAP (25.8% ammonia, 74.2% phosphate) addition is important. One large addition of DAP at the beginning may delay/inhibit uptake of amino acids. Multiple additions are preferred. Adding nutrient supplements all at once can lead to too fast of a fermentation rate and an imbalance in uptake and usage of nitrogen compounds. Supplements added too late (after half the fermentation ) may not be used by the yeasts , in part because the alcohol prevents their uptake. For the same reason, adding nutrients to a stuck fermentation seldom does any good at all. Do not wait until you have a sluggish or stuck fermentation to add nutrients.
Vitamin Addition       Musts can be deficient in vitamins and often will be when there is a high incidence of microorganisms (mold, yeast and/or bacteria). Addition of sulfur dioxide tends to inactivate thiamine which is necessary for yeast growth. It is usually desirable to add a mixed vitamin supplement along with a nitrogen supplement.
      If grapes are degraded by Botrytis and/ or Kloeckera add extra thiamine.
Fermentation Rate       The rate of fermentations should be monitored by the use of Brix hydrometers and/ or an analysis of residual sugar. What is desired is a steady fermentation that gradually declines.
Oxygen/SO2       Oxygen should be considered an essential yeast nutrient. Slight aeration during yeast stationary and growth phases increases the production of lipids (principally oleanoloic acid) and sterols( ergosterol, and zymosterol) which are important cell membrane constituents. It has been shown that yeast propagated aerobically contain a higher proportion of unsaturated fatty acids and up three time the steroid level of anaerobically grown yeast. This correlates positively with improved yeast viability and fermentation. Because yeasts are not able to synthesize membrane components in the absence of oxygen, existing steroids must be distributed within the growing populations. Without initial oxygen, yeast multiplication is usually restricted to 4 to 5 generations, due largely to diminished levels of steroids, lipids and unsaturated fatty acids. CO2 , nitrogen gas and ascorbic acid reduce molecular oxygen.
      Additionally, it should be noted that SO2 inhibits the enzyme polyphenol-oxidase. In the complete absence of sulfur dioxide, this common plant enzyme system converts diphenols to quinones using a large concentration of available oxygen.
      As indicated, sulfur dioxide also inactivates thiamine. If additions of more than 50 mg/L SO2 occur, extra thiamine should be added to the fermenter.
pH       Maintain pH as high above 3.1 as wine style permits. Musts which have a pH below 3.1 should receive an increased yeast inoculum.
Non-soluble solids Reduction of the non-soluble solids content to below 0.5% prior to white wine fermentation can result in nutrient deficiencies. Too high a level may cause fermentation rates to proceed too quickly. Fermentation in contact with bentonite is occasionally done to help obtain white wine protein stability. Bentonite additions in the fermenter can reduce must N and should only be done in conjunction with supplemental nutrient additions.
Sedimentation       Yeast cells at the bottom of a fermenter can die prematurely. To help avoid this problem, large tanks should be mixed.
CO2 toxicity       Carbon dioxide in concentrations of up to 0.2 atm stimulates yeast growth. Above this level carbon dioxide becomes toxic to the yeasts. Agitation to prevent supersaturation of CO2 can minimize this problem.
Sugar toxicity
      High sugar concentrations can inhibit yeast growth due to osmotic pressure. Saccharomyces sp. are more tolerant than most others.
      High sugar musts start fermentation slowly and are likely to stick.
      There is a synergism between alcohol and sugar concentration.
      Inoculation with greater than 5 x 106 yeast cells/mL should occur if the must is 25-30 Brix. Inoculate with an additional 1 x 106 yeast cells for each degree increase in a Brix above 30.
Alcohol toxicity       Alcohol is toxic to all yeasts, including Sacccharomyces sp. Alcohol has a profound effect on all aspects of yeast metabolism from membrane integrity to nitrogen uptake and sugar transport. There are many factors which are synergistic with alcohol including pH, high temperature, acetic acid, sugar, short chained fatty acids, nitrogen depletion, and deficiency of sterols and vitamins.
      As indicated, light aeration during growth phase of the yeast helps to produce lipids needed by the yeast cell wall. Nitrogen supplementation is helpful in reducing the affects of alcohol toxicity.
Wild yeast/bacteria, infected grapes, poor sanitation, long setting, and late inoculation
      Wild yeast/bacteria, infected grapes, poor sanitation, long setting, and late inoculation deplete must nutrients and may produce toxins. In such cases the level of yeast inoculum should be increased along with the addition of 250-500 mg/L N.
      Acetic acid bacteria, Lactobacillus sp., Leuconstoc sp., and 'wild' yeast can produce inhibitors and deplete must N and vitamins. Acetic acid is a potent inhibitor of Saccharomyces sp. especially in combination with other negative influences such a high alcohol late in the fermentation. A stuck wine with more than about 0.8 g/L acetic acid may need to go through a R.O. to reduce the acetic acid content before attempting refermentation.
      Some Saccharomyces sp. and strains and some non-Saccharomyces yeasts can produce killer toxins that inhibit sensitive stains. These killer toxins can play a roll in stuck fermentations. It is suggested that vigorous strains be used for high risk fermentations.
Uninoculated Musts       Usually non-Saccharomyces from the vineyard and Saccharomyces from the winery dominate the initial fermentation of uninoculated musts, possibly resulting in a significant depletion of N and vitamins such as thiamine. Kloeckera sp. which may dominate the early portion of uninoculated fermentations are cold and sulfur dioxide tolerant and can produce high levels of ethyl acetate. Kloeckera can also significantly deplete N and thiamine. It is desirable to supplement uninoculated fermentations with nitrogen and vitamins.
Temperature       Increase inoculum when fermenting at low temperature. Decrease inoculum slightly for uncontrolled high temperature and select a slower fermenting strain of yeast. Add yeast nutrient to protect the yeast at each end of the temperature range.
Fructose       Grape juice is usually composed of equal concentrations of glucose and fructose sugars. Stress can affect the yeast's ability to metabolize the last residual fructose. Add small amounts of glucose to a small portion of the wine to determine if this is the cause of a stuck fermentation. This problem seems to occur more with the S. bayanus strains which are more glucophilic and, therefore, unable to ferment fructose.
      Use fructose syrup as last choice for amelioration. Fermentation rate can be easily measured by the use of Durham or fermentation tubes.
Yeast Hulls       Yeast hulls additions (0.2 g/L) stimulate fermentation not simply by detoxification as was previously believed, but by supplying unsaturated fatty acids (C-16, C-18) as an oxygen substitute and preventing deficiencies of this nutrient. Also, yeast hulls add some amino acids and facilitate the release of CO2.
Pesticides       Pesticides can influence fermentation by causing production of stress metabolites such as reductive compounds, as well as inhibiting and/or preventing fermentation. Not all yeasts and bacteria are affected the same way by pesticides. There is a significant difference between systemic and contact fungicides in regard to residues. Vinification style influences pesticide residue concentrations. For example, contact pesticide residues are influenced by preclarification of whites and the addition of bentonite. To help prevent the problem of residual pesticides be aware of spray schedules, use less than the maximum permitted when possible and avoid late season spraying. Late season copper sulfate sprays (Bordeaux mix) can significantly increase the production of hydrogen sulfide and mercaptans.

Adapted from: Dr. Clayton Cone, Lailemand, Inc., Montreal, Quebec, Canada ;Wine Analysis and Production, Zoecklein et al., 1995; Wine Microbiology Fugelsang, 1996; Die Wein Wissenschaft, 1996; and Van de Water, personnel communication.

Table 2:

Survey results from 1993 and 1994.
Mean content of free ammonia and free amino nitrogen (less free ammonia)
 Free Ammonia (mg/L)Free Amino Nitrogen (mg/L)
Cayuga White68    32    74      197      
Chardonnay46    55    151      177      
Riesling52     56    102      123      
Seyval Blanc19    14    82      156      
Pinot Noir52    88    135      116      
Cabernet Sauvignon49    69    74      142      
TOTAL(all cultivars)45    56    120      149      

Source: Henick-Kling et al. (1996)