Comprehensive study of the dynamic interaction between SO2 and acetaldehyde during alcoholic fermentation.

In this work, we focused on the effect of the initial content of SO2 in synthetic grape juice on yeast metabolism linked to the production of acetaldehyde. Lengthening of the lag phase duration was observed with an increase in the initial SO2 content. Nevertheless, an interesting finding was a threshold value of an initial SO2 content of 30 mg L-1 in the juice led to equilibrium between intracellular SO2 diffusion and SO2 production from the sulfate pool by yeast. The ratios of free and bound acetaldehydes were measured during fermentation, and the maximum accumulation of free acetaldehyde was observed when SO2 concentration equilibrium between diffusion and production was reached in the fermenting juice. Moreover, it was observed that SO2 addition resulted in significant changes in the synthesis of aroma compounds. Production of volatile molecules related to sulfur metabolism (methionol) was changed. But, more surprisingly, synthesis of some volatile carbon compounds (diacetyl, isoamyl alcohol, isobutyl alcohol, phenyl ethanol and their corresponding esters) was also altered because of major disruptions in the NADPH/NADP+ redox equilibrium. Finally, we demonstrated that acetaldehyde bound to SO2 could not be metabolized by the yeast during the time course of fermentation and that only free acetaldehyde could impact metabolism.

Vicinal diketones and their precursors in wine alcoholic fermentation: Quantification and dynamics of production.

Vicinal diketones produced during wine fermentation influence the organoleptic qualities of wine. Diacetyl and 2,3-pentanedione are well known for their contribution to butter or butterscotch-like flavours. We developed an analysis method to quantify vicinal diketones and their precursors, α-acetolactate and α-acetohydroxybutyrate, under oenological conditions. Five-fold dilution of the sample in a phosphate-citrate buffer (pH7.0) strongly attenuated matrix effects between the beginning and end of alcoholic fermentation and protected the sample from spontaneous precursor decarboxylation. The use of diacetyl-d6 as an internal reference improved precision by eliminating differences in the derivatization and extraction yields between the internal standard and the analytes. We obtained unexpected results for alcoholic fermentation by Saccharomyces cerevisiae using this approach. Indeed, the level of diacetyl and 2,3-pentanedione throughout fermentation were very low. However, we observed a large quantity of both precursors. The production dynamics of α-acetolactate were unconventional and there were two distinct phases of accumulation. The first corresponded to the growth phase, and the second to glucose depletion. There was a rapid decrease of precursor levels at the end of fermentation, but there was still a significant amount of α-acetolactate. The amount of precursor remaining at the end of fermentation constitutes a potential source of diacetyl during wine maturation. α-Acetohydroxybutyrate accumulated during the growth phase followed by a continuous decrease of its concentration during the stationary phase. Residual quantities of α-acetohydroxybutyrate found in wine at the end of fermentation does not constitute a sufficient source of 2,3-pentanedione to affect the aromatic profile.

Impact of initial lipid content and oxygen supply on alcoholic fermentation in champagne-like musts.

Available nitrogen, lipids, or oxygen are nutrients with major impact on the kinetics of winemaking fermentation. Assimilable nitrogen is usually the growth-limiting nutrient which availability determines the fermentation rate and therefore the fermentation duration. In some particular cases, as in Champagne, grape musts have high available nitrogen content and low turbidity, i.e., below 50 Nephelometric Turbidity Unit (NTU). In the case of low turbidity, the availability of lipids, particularly phytosterols, becomes limiting. In this situation, control of oxygenation, which is necessary for lipid synthesis by yeast, is particularly crucial during fermentation. To mimic and understand these situations, a synthetic medium simulating the average composition of a Champagne must was used. This medium contained phytosterol (mainly ß-sitosterol) concentrations ranging from 0 to 8mg/L corresponding to turbidity between 10 and 90 NTU. Population reached during the stationary phase and the maximum fermentation rate are conditioned by the initial phytosterol concentration determining the amount of nitrogen consumption. An early loss of viability was observed when the lipid concentrations were very low. For example, the viability continuously decreased during the stationary phase to a final value of 50% for an initial phytosterol concentration of 1mg/L. In some fermentations, 10mg/L oxygen were added at the end of the growth phase to combine the effects of initial content of phytosterols in the musts and the de novo synthesis of ergosterol and unsaturated fatty acids induced by oxygen addition. Effect of oxygen supply on the fermentation kinetics was particularly significant for media with low phytosterol contents. For example, the maximum fermentation rate was increased by 1.4-fold and the fermentation time was 70h shorter with oxygen addition in the medium containing 2mg/L of phytosterols. As a consequence of the oxygen supply, for the media containing 3, 5 and 8mg/L of phytosterols, the assimilable nitrogen was completely exhausted and the fermentation kinetics, as well as the final populations and viabilities (greater than 90%), were identical for the 3 conditions. The impacts of the lipid content and additional oxygen on acetate, glycerol and succinate synthesis were also studied. The phytosterols decreased the acetate and increased the succinate synthesis, and oxygenation resulted in a decrease in succinate formation. This work highlights the similarities and differences between the effects of lipids and oxygen on fermentation kinetics and yeast metabolism. This research highlights the need for an optimal combined management of lipid content in the must via turbidity and oxygenation, particularly in nitrogen-rich musts.