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UPDATE

Number 2 – 2005

FLASH NEWS

❖ Ochratoxin Removal by Strains SPECIAL EDITION OF WINEMAKING UPDATE Ochratoxin A (OTA), a toxin produced by some fungi, is now being carefully Yeast Inoculation Strategies – regulated in the and in other countries to limit its concentration in Getting The Best such food products as cereal and coffee. OTA has also been found in and , and appears to be more Possible! predominant in Mediterranean-type . ood alcoholic fermentation steady and complete fermentation is the It was found that different oenological management is the key to avoid- presence of an adequate cell population can reduce OTA at different levels Ging stuck or sluggish fermenta- when the yeast has finished growing, via an adsorption mechanism (Bejaoui et tions while making quality wine. which is normally when 30% to 50% of al., 2004). Higher yeast biomass Selecting the most appropriate yeast the remain to be fermented. To concentration was also found to reduce strains based on the conditions and complete fermentation and minimize more OTA. Another study carried out by the type of wine style desired is an the risk of fermentation problems, at this the ICV in collaboration with Lallemand has important winemaking decision. The stage you need a minimum yeast cell shown that OTA reduction was also related proper strain, carefully rehydrated and at population of approximately 120-150 x to the yeast strains. For example, ICV D80 the correct inoculation dose for ferment- 106 cells/mL. At inoculation densities had the highest ing the must, should be foremost in the below 5 x106 cell/mL, the likelihood of OTA reduction of all six yeast strains tested. ’s decision to help avoid prob- achieving a successful fermentation More studies are underway to verify if OTA lems with stuck or sluggish fermentation decreases significantly. In part, this is can be potentially released back into the and aromatic deviations due to the demand placed on the initial wine during aging. The malolactic bacteria yeast cell population to divide for more This special edition of Winemaking Oenococcus oeni can also play a role in than five or six generations. Update will describe the basics behind degrading OTA. The Lalvin 31 strain was determining the inoculation rate to min- The suggested 25 g/hL rate of yeast inoc- shown to do this very successfully. imize such problems, will define the con- ulation is based on the fact that a proper ditions that will get you the most out of initial cell density is required for the fermentation under high potential alco- onset and the completion of fermenta- WINEMAKING UPDATE hol conditions, will explain the use of tion. Before the onset of fermentation, the winemaker should keep the lag WINEMAKING UPDATE is a Lallemand Inc. sequential inoculation and dynamic syn- phase as short as possible to avoid micro- publication. Its goal is to inform oenolo- ergy, and will provide a brief description gists and winemaking staff about news of nutrient strategies to obtain a smooth bial spoilage and the production of and indications arising from research. To and complete fermentation. undesirable compounds. The length of the lag phase is significantly influenced request previous issues, send your ques- 1. Why use a minimum of 25 g/hL? tions or comments contact us at: by the initial yeast cell density or inocu- Lallemand During wine fermentation, yeast cells lation rate, as well as factors like juice Gordon Specht will usually undergo at least five cell clarity, and SO2 concentrations, P.O. Box 5512, divisions (doubling the yeast population which will also impact on the relation Petaluma, CA 94955, U.S.A. with every cell division). One of the between lag phase duration and inocula- [email protected] most important factors in obtaining a tion rate. At 25 g/hL (or 2 pounds per The WINEMAKING UPDATE technical infor- Continued mation is true and accurate; due to the great diversity of operative situations all FLASH NEWS the advices and warnings are given with- ❖ H S Production by Wine Yeasts In a study done in collaboration with Dr. Seung Park at the out any guarantee and formal engage- 2 ment. The Lallemand products are widely University of California – Davis, the relative hydrogen sulphide (H2S) production of different yeast available through a distribution network. strains was studied under different levels of nitrogen in the must. The study found that under To find your local distributor contact us at low nitrogen conditions (60 ppm), yeast strains had a tendency to produce more H2S compared the above address. to nitrogen levels of 170 ppm. The yeast strains could be classified as low, medium or high producers. This information will help determine the proper yeast and yeast nutrition under www.lallemandwine.com@ specific must conditions. 1,000 gallons), the inoculation will pro- point due to the crowding effect. With require higher inoculation rates (30-50 vide approximately 5 x 106 cells/mL with that amount of cells at the beginning, g/hL) to complete fermentation because the majority of selected and dried natural you significantly increase your chance to their metabolism is very different. yeast preparations. Increasing the inocu- obtain the required 120-150 x106 The following results (Figure 1) illustrate lum density will result in decreased lag cells/mL to successfully complete fermen- the importance of an inoculum of time, but is ineffective beyond a certain tation. Note that some yeast strains will 25 g/hL versus 10 g/hL. A study done in

FIGURE 1. (215 g/L sugars, pH 3.17, T.A. 6.5 g/L) fermentation at 16°C comparing inoculation rates of 10 g/hL and 25 g/hL of EC-1118 yeast strain with Fermaid added at mid-fermentation (30 g/hL). Lag Phase Fermentation length

50 800 45 700 40 600 35 30 500 25 400 Time (h) 20 Time (h) 300 15 200 10 5 100 0 0 10 g/hL 25 g/hL 10 g/hL 25 g/hL EC-1118+Fermaid EC-1118+Fermaid EC-1118+Fermaid EC-1118+Fermaid

Volatile acidity

0,55 ) 4

SO 0,50 2 0,45

0,40

0,35

0,30

0,25

Volatile acidity (g/L H Volatile 0,20 10 g/hL 25 g/hL EC-1118+Fermaid EC-1118+Fermaid

FIGURE 2. Residual sugar of a Bourboulenc- (14°C and 28°C, 15.4% vol., 258 g/L sugar, 220 NTU, 70 mg/L assimilable N) fermentation with 10 and 25 g/hL of EC-1118 yeast and with the pied de cuve technique.

14°C 28°C

7 16

6 14

5 12 10 4 8 3 6 2 4 Residual sugar (g/L) Residual Residual sugar (g/L) Residual 1 2 0 0 10 g/hL 25 g/hL 10 g/hL 25 g/hL EC-1118+Fermaid EC-1118+Fermaid EC-1118+Fermaid EC-1118+Fermaid

Pied de Cuve (14°C & 28°C)

50 45 40 35 30 25 20 15 10 Residual sugar (g/L) Residual 5 0 14°C 28°C by INRA Pech Rouge in collabo- or sweet wines (vins liquoreux), higher mentation in difficult conditions, such as ration with Lallemand (Ortiz-Julien, inoculation rates are required. In the case a high sugar situation. For example, a 2003) clearly showed the benefits of of very high sugar content , the yeast strain that is particularly interesting inoculating a “normal” Chardonnay must hyperosmotic stress placed on the yeast in terms of sensory contribution is often with an potential of 12.7% with can reduce yeast biomass and ultimately desired by . However, this 25 g/hL of the EC-1118 yeast strain. This generate a stuck or sluggish fermentation, yeast strain is very sensitive to difficult resulted in a significant reduction of lag as well as increase the production of winemaking conditions and may end up phase time, fermentation length and volatile acidity. Therefore, when the having difficulty finishing the fermenta- volatile acidity. level is above 30° Brix, an inoculation tion. This yeast is thus associated with In another study, under more difficult rate of 50 g/hL is required. For example, another yeast strain that is quite resistant conditions, the must of Bourboulenc- in a study carried out on icewine (35 to to difficult winemaking conditions and Grenache (15.4% vol., 258 g/L sugar, 42° Brix) at the Cool Climate reliable for completing the fermentation. 220 NTU, 70 mg/L assimilable N), was and Institute at Brock The result is a wine that possesses the fermented with 10 g and 25 g/hL of the University in Canada, it was shown that sensory characteristics that were wished EC-1118 yeast strain and compared to a higher inoculation rate (50 g/hL vs. 20 for by using the first yeast strain, but the use of the pied de cuve technique at g/hL) along with the utilisation of Go- without any worries regarding fermenta- both 14°C and 28°C. The levels of resid- Ferm in the rehydration water signifi- tion kinetics or completion, thanks to ual sugars were shown to be significantly cantly reduced the fermentation time as the second yeast strain. Such mixtures for higher with the pied de cuve inoculation well as the rate of volatile acid produced use in a single inoculum are in develop- (up to 48 g/L) compared to the inocula- as a function of sugar consumed ment and will soon be available to wine- tion rate of 25 g/hL as shown in Figures (Kontkanen et al, 2004). makers. 2a and 2b. Therefore, working with 25 2.2 Other strategies: sequential inocu- 3. Good fermentation practices – g/hL of inoculum is a good strategy to lation and dynamic synergy a quick review avoid sluggish or , A novel approach to ensure a steady and The followings are other good fermenta- even in difficult situations (i.e., low complete fermentation in over-mature tion practice considerations when deal- nitrogen, high sugar). (28° Brix) is based on a study of ing with high-Brix , and Table 1, on Although recent results also suggest that red wines conducted by Professor page 4, summarizes the different inocula- a high inoculation rate may allow a Edmundo Bordeu at the Universidad tion strategies. reduction in the amount of SO2 used in Católica de in 2004 on the use of In reds: fermentation, it is too early to give an sequential inoculation. The principle is • Aeration or oxygen additions when the accurate estimation of the SO2 dosage as the following: the selected yeast strain is a function of inoculation rate. inoculated in the must at 30 g/hL in two cap forms (usually when 15 g/L sugar is stages, first at the beginning of fermenta- fermented,) as well as nutrient addition 2. What to do in high potential at one third sugar depletion. alcohol (high sugar) situations? tion and then when the brix has reached a certain point, but before symptoms of • Temperature management during yeast 2.1 High sugar musts require more stuck or sluggish fermentation are appar- rehydration, the initial phase of fermen- yeast inoculum ent. At first, 15 g/hL of the selected yeast tation and at the peak of fermentation. High sugar musts are increasingly com- is rehydrated in water supplemented with • Regular movement of the yeast near mon, especially for New World winemak- Go-Ferm, then used for inoculation. the end of fermentation and into the ers and in hot climates, and are often When the alcohol level reaches about yeast cell decline phase. coupled with nitrogen deficiencies. In 4%, the other 15 g/hL of yeast (also such situations, fermentation manage- rehydrated in water and supplemented In whites: ment requires particular attention. Not with Go-Ferm) is used for inoculation. • Initial juice turbidity level optimum only is proper nutrition during fermenta- This method works well as it is always between 80 and 100 NTU. tion and rehydration essential for a very difficult to restart a stuck fermenta- • Aeration or oxygen addition as soon as steady and complete event, the appropri- tion. When reinoculated early, there was the fermentation is active (usually ate inoculation rate should receive con- a clear advantage in terms of lower resid- when 15 g/L sugar is fermented), as sideration. As we saw in Figure 2 A, the ual sugars and lower volatile acidity. well as nutrient addition at one third levels of residual sugars are lower when A dynamic synergy is also a good alterna- sugar depletion. 25 g/hL were used in a must containing tive. This new concept is based on the 258 g/L of sugar (approximately 25° • Temperature management during yeast use of two or more selected yeast strains rehydration and yeast inoculation Brix). The volatile acidity was also signif- particularly well adapted to each other in icantly lower, the lag phase duration was that they complement sensory properties • Regular movement of the yeast near reduced and the fermentation rate was and fermentation kinetics in perfect syn- the end of fermentation and into the faster (results not shown) in the ergy. After many months of research at yeast cell decline phase. Bourboulenc-Grenache wine fermented Lallemand, our studies have shown that REFERENCES with 25 g/hL of EC-1118 yeast. the proper combination of certain strains Bejaoui, H., P. Mathieu, P. Taillandier In an even higher sugar must for icewine in the proper ratio can help carry out fer- and A. Lebrihi, 2004. OTA removal in TABLE 1. Different inoculation strategies in different must conditions

Must conditions Inoculation rate Go-Ferm Fermaid

High N > 250 mg/L: 30 g/hL No addition “Normal” conditions N 150 < N < 250 mg/L: 30 g/hL (red, white, rosé) 25 g/hL Add 30 g/hL at 1/3 through fermentation below 25°Brix Low N < 150 mg/L: 30 g/hL Add 15 g/hL at inoculum and 15 g/hL at 1/3 through fermentation

Sweet wines/ Regardless of nitrogen content, add 30 g/hL at Icewine 50 g/hL 60 g/hL inoculum and 20 g/hL at 1/3 through fermenta- 30-42° Brix tion

High N > 250 mg/L : 30 g/hL at 1/3 through fermentation 25-50 g/hL. (The higher the N 150 < N < 250 mg/L: sugar content, 30-60 g/hL Add 50 g/hL at 1/3 through fermentation the higher the inoc- Low N < 150 mg/L: ulation rate) High sugar must/ Add 30 g/hL at inoculum and 20 g/hL at 1/3 over mature grapes through fermention over 25° Brix 30 g/hL in sequen- tial inoculation : 15g/hL at the 20 g/hL for each Regardless of nitrogen content, add 20 g/hL at beginning and inoculation each inoculation 15 g/hL at approxi- mately 4% alcohol synthetic and natural grape juice by and nutrient addition on icewine fer- Ortiz-Julien, A., 2002. Inoculation rate selected oenological Saccharomyces mentation. Am J Enol Vitic. 55(4):363- study. INRA Montpellier. Internal strains. J App Microbiol. 97:1038-1044. 370. report. Bordeu, E., 2005. Inoculation of yeast Monk, P., 1997. Optimum usage of active Park, S., 2005. UC Davis. Internal during alcoholic fermentation to face dry wine yeast. ASVO Seminar. report. problems of stuck . Internal Advances in Juice Clarification and Yeast Querol, A., E. Barrio, T. Huerta and D. report. Inoculation. 22-23 Ramón, 1992. Molecular monitoring of Kontkanen, D., D.L. Inglis, G.J. Ortiz-Julien, A., 2003. Inoculation rate wine fermentations conducted by active Pickering and A. Reynolds, 2004. Effect study. INRA Pech-Rouge. Internal dry yeast. App Environ Microbiol. of yeast inoculation rate, acclimatization report. 58 (9):2948-2953.

TO SUMMARIZE … Every must requires particular attention, and as every brings its own variability, reducing the risks of fermentation devia- tion is important. Keep in mind that in a very short time a naturally selected yeast in a dry form has to become an actively func- tioning unit in the wine environment, which can be hostile (high sugar, low pH, high SO2), and one of the keys to good fermen- tation management is the proper inoculation rate. The points to remember are the following: 1. A minimum of 25 g/hL is necessary to obtain the proper cell density for a smooth and complete fermentation. 2. The inoculation dosage has to be adapted to the must conditions and the wine type: higher sugar musts requires higher inocula- tion, and sweet-wine types require a much higher inoculation rate. 3. The nutritional status of the must, as well as the nitrogen/oxygen requirements of the yeast strains have to be considered. Rehydrating the dried yeast in warm water and Go-Ferm, as well as supplying the yeast with Fermaid during fermentation, are part of good fermentation management. 4. Other strategies can be investigated with over-mature grapes, such as sequential inoculation or the use of dynamic synergy prod- ucts. Consult your Lallemand representative for the best conditions adapted to your particular situation.