Krieger, S.

Tosi, E.

Zapparoli, G.

English

JKI Open Journal Systems (Julius Kühn-Institut)

Vitis 45 (4), 197–198 (2006)plicate using a volume of 100 l for each trial. Commer-cial strains of Saccharomyces cerevisiae Lalvin Rhône 4600 and of Oenococcus oeni, Lalvin 31, were prepared according to the manufacture instructions. Bacteria were inoculated 16 h after the yeast inoculation when total and free SO2 were 28.0 and 6.4 mg·l-1, respectively. The fer-mentations were monitored by analysis of ethanol produc-tion, L-malic acid consumption, yeast and bacteria cell concentrations. After AF, commercial MLF nutrient (Opti Malo Plus, Lallemand) was added according to the manu-facture instructions. Total acidity, sugars, ethanol, sulfite and pH were determined using standard methods, organic acids were quantified using enzyme kits (Roche). Eight biogenic amine (BA) (histamine, cadaverine, putrescine, phenylethylalanine, amylamine, isobutylamine, methyl-amine and isopropylamine) were determined as previously described (TORREA and ANCIN 2001). Data are average of two determinations. Cell counts were carried out plating on WL agar (Oxoid) medium for yeasts and MRS (Fluka) added 10 % tomato juice and 0.01 % actidione (Fluka) for bacteria. Primary classification of isolated LAB was car-ried out by morphological and biochemical tests as gram staining, catalase and sugar fermentation. Results and Discussion: The effect of pH of Char-donnay must on AF and MLF is reported in the Figure. In the trials at pH 3.18, ethanol production was slower and the viability of the yeast population was longer than in the other trials. ROSI et al. (2003) reported a similar behaviour of yeast population coinoculated with bacteria in white grape juice adjusted to different pH, but sugar fermentation was faster at pH 3.2 than at 3.4. The bacteria population, after the initial decline, was less pronounced at pH 3.18, and the cell viability recovered in all trials. At the end of AF, bacteria cell concentration was about 1 log CFU·ml-1 higher in wine at low pH than in the other wines because of the better adaptability of cells. This result confirms that strain Lalvin 31 is correctly characterized by tolerance to low pH and temperature, stressful conditions when it was originally selected. The Table reports the wine composi-tion after AF and MLF. At the end of AF the content of free and total SO2 were similar for all the wines, which were less than 10 and 60 mg·ml-1, respectively. Sugar ca-tabolism was performed by yeasts and not by O. oeni con-firming that MLF can occur in the presence of fermentable sugars without significant increase of acetic and D-lactic acid (KRIEGER 2002). After the wine lees were removed, in concomitance with the reduction of the winery tempera-ture, bacteria populations underwent a different fate. At the lowest pH, a clear increase of cell concentration was ob-served and MLF finished in about 15 d. In the other wines, an initial loss of cell viability was seen, followed by cell growth but malic acid depletion ceased. The high acetic acid content of wine which had undergone a partial MLF, especially at the highest pH, is the result of microbiologi-cal instability caused by the failure of the starter coloniza-tion. In fact, Lactobacillus spp. were isolated from wines that had undergone a partial malolactic conversion at the concentration of 5.0 x 103 and 1.5 x 106 CFU·ml-1 in trials at initial pH 3.45 and 3.91, respectively. Moreover, in the Research NoteInfluence of the pH of Chardonnay must on malolactic fermentation in-duced by bacteria co-inoculated with yeastsG. ZAPPAROLI1), E. TOSI2) and S. KRIEGER3)1) Dipartimento Scientifico e Tecnologico, Università degli Studi di Verona, Verona, Italy2) Centro per la Sperimentazione in Vitivinicoltura, Provincia di Verona, Servizio Agricoltura, San Floriano, Verona, Italy3) Lallemand, office Korntal-Münchingen, GermanyK e y   w o r d s :  malolactic fermentation, yeast-bacteria co-inoculation, Chardonnay wine.Introduction: In wine, the success of the malolactic fermentation (MFL) can be unsure when unfavorable con-ditions for the malolactic activity of lactic acid bacteria (LAB) occur. Slow or incomplete MLF is especially prom-inent in white wine due to the presence of concomitant fac-tors such as nutritional deficiencies, less than optimal pH and temperature, high SO2 and high ethanol content (PI-LATTE and NIELSEN 1999). Previous investigations focused on the effects of the timing of bacteria inoculation in wine and the co-inoculation of yeast and bacteria is proposed as alternative technique for wines that often have difficulty to undergo MLF (KRIEGER 2002, JUSSIER et al. 2006). With this technique, the interaction between yeasts and bacteria is stricter and more complex than with traditional inocula-tion. The interest to understand the mechanisms that govern yeast-bacteria interaction in wine is growing (ALEXANDRE et al. 2004). ARNINK and HENICK KLING (2005) studied dif-ferent yeast-bacteria combinations to predict the success or failure of MLF in wine. Under winemaking conditions the result of one specific combination seems to be influenced by several factors that make MLF prediction difficult.In this study, a yeast-bacteria combination was evalu-ated for the malolactic conversion in Chardonnay wine; the bacteria were inoculated with yeasts in must at different pH, one of the most important factors influencing MLF in wine. Using real winery conditions, this study is an appli-cative and useful contribution to the science of winemak-ing.Material and Methods: Chardonnay must was composed as follows: pH 3.45, total acidity 8.58 g·l-1 (as tartaric acid), reducing sugars 185.0 g·l-1, L-malic acid 5.70 g·l-1, citric acid 0.42 g·l-1. The must was divided in three aliquots and in two of them pH was adjusted to 3.18 and 3.91. Microvinifications were performed in du-Correspondence to: Dr. G. ZAPPAROLI, Dipartimento Scientifico e Tecnologico, Università degli Studi di Verona, Strada Le Gra-zie,15, 37134 Verona, Italy, Fax: +39045802729 E-mail: giaco-mo.zapparoli@univr.it 198 G. ZAPPAROLI et al.In conclusion, this study highlights the importance of a rapid degradation of malic acid in wine in order to avoid stuck malolactic fermentation. The results confirm that the success or failure of MLF strongly depends on the occur-rence of favorable or unfavorable winemaking conditions specifically for the malolactic strain inoculated. The yeast-bacteria combination used here was successful in must at the lowest pH. Under this condition, the success of MLF is attributed to the full adaptation of the starter bacteria that allowed fast colonization in wine. Further investigations are necessary to evaluate to which degree the cohabitation of bacteria with yeasts during FA influences MLF and, in particular, the contribution of yeasts to malolactic bacteria performance.ALEXANDRE, H.; COSTELLO, P., J.; REMIZE, F.; GUZZO, J.; GUILLOUX-BENA-TIER, M.; 2004: Saccharomyces cerevisiae-Oenococcus oeni inter-action in wine: Current knowledge and perspectives. Int. J. Food Microbiol. 93, 141-154.ARNINK, K.; HENICK KLING, T.; 2005: Influence of Saccharomyces cerevi-siae and Oenococcus oeni strains on successful malolactic conver-sion in wine. Am. J. Enol. Vitic. 56, 228-237.JUSSIER, D.; DUBÉ MORNEAU, A.; MIRA DE ORDUÑA, R.; 2006 Effect of si-multaneous inoculation with yeast and bacteria on fermentation ki-netics and key wine parameters of cool-climate Chardonnay. Appl. Environ. Microbiol. 72, 221-227.KRIEGER, S.; 2002: Starter cultures for malolactic fermentation – time of inoculation. In: H. TROGUS (Ed.): Proc. 13th Int. Enol. Symp., 77-91. Int. Association of Enology, 9-12 June 2002, Montpellier, France.LONVAUD-FUNEL, A.; 1999: Lactic acid bacteria in the quality improvement and depreciation of wine. Antonie van Leeuwenhoek 76, 317-331.PILATTE, E.; NIELSEN, J. C.; 1999: Mise au point d’un nouveau ferment malolactique à ensemencement direct, pour vins blancs. Rev. Oenol. 93, 15-16.ROSI, I.; FIA, G.; CANUTI, V.; 2003: Influence of different pH values and inoculation time on malolactic activity of a strain of Oenococcus oeni. Aust. J. Grape Wine Res. 9, 194-199.TORREA, D.; ANCIN, C.; 2001: Influence of yeast strain on biogenic amines content in wines: relationship with the utilization of amino acids during fermentation. Am. J. Enol. Vitic. 52, 185-190.Received March 29, 2006latter, Pediococcus spp. was isolated at the concentration of 5.0 x 105 CFU·ml-1. These LAB are frequently predomi-nant in wine with high pH, where SO2 is ineffective, induc-ing spoilage and they are mainly responsible for high BA production in wine (LONVAUD-FUNEL 1999). Nevertheless, in these wines the BA content, determined after MLF, was low and similar (the total amount of 8 identified BA was 4.63, 4.60 and 4.73 mg·ml-1 in trials at pH 3.18, 3.45 and 3.91, respectively) and no significant differences were ob-served among the wines for each BA analysed (data not shown). T a b l e   Composition of wines produced from Chardonnay must at different pH co-inoculated with yeasts and bacteriaWine compositionafter AF at after MLFa atpH 3.18 3.45 3.91 3.18 3.45 3.91pHtotal acidityacetic acidL-malic acidL-lactic acidD-lactic acidcitric acidg·l-1bg·l-1g·l-1g·l-1g·l-1g·l-13.207.400.302.301.680.160.253.556.770.292.721.530.190.324.065.540.283.800.710.210.443.315.750.400.023.110.180.143.636.290.601.872.010.200.274.184.851.161.792.210.210.30 a completed for pH 3.18 and partial for pH 3.45 and 3.91. b as tartaric acid. 3456789CFU/mLCell concentration (log 10 CFU l–1 ) 0 2 4 6 8 10 12 14 16 18 20 0 7 14 21 28 35 42 49 56 63 70 Time (d) 0 1 2 3 4 5 6 7 --   Ethanol (% vol), Temperature (°C) Malic acid (g l-1 ) Figure: Cell concentration (open symbols: yeasts, closed sym-bols: bacteria), ethanol (open symbols) and malic acid content (closed symbols) measured during the alcoholic and malolactic fermentation in the trials at initial pH 3.18 (circle), 3.45 (triangle) and 3.91 (square). Winery temperature is indicated by crosses. The arrow indicates the end of alcoholic fermentation.

Influence of the pH of Chardonnay must on malolactic fermentation induced by bacteria co-inoculated with yeasts

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Influence of the pH of Chardonnay must on malolactic fermentation induced by bacteria co-inoculated with yeasts

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