Richard Marchal

The ancestral method of sparkling wine production requires no sugar addition and is based on a single alcoholic fermentation that finishes in the bottle. It has recently gained popularity among both natural and conventional producers due to its capacity to expand sparkling wine portfolios and its advantages for sparkling wine production in warm regions such as the Mediterranean basin. However, some technological and other key aspects of the process remain poorly defined. This study investigates how must settling (using pectolytic enzymes) and pre-bottling filtration influence the physicochemical composition, colloidal structure, foaming properties, volatile profile, and sensory attributes of ancestral sparkling wines. Macabeo grapes (2023 vintage) were vinified under four treatment combinations of these two operations, and the wines were analysed after eighteen months of lees ageing. Significant differences were observed in total polyphenol index, titratable acidity, macromolecular composition, and colloidal particle concentration. Non-settled wines exhibited higher colloidal loads and lower foamability, whereas settling and filtration improved foam height and stability. Volatile (GC-Headspace) analysis showed that settled wines contained higher concentrations of fruity and floral esters, while non-settled and non-filtered wines had greater levels of volatile phenols and volatile acids associated with unpleasant aromas. Sensory assessment confirmed perceptible differences between treatments, particularly in phenolic, fruity/floral, and Maillard reaction aromaticity. These findings provide insights into the impact of two critical oenological practices on wine stability and aroma and provide technologically relevant guidance for optimizing quality in this emerging sparkling-wine category.

Botrytis cinerea is a fungal pathogen responsible for the decrease in foamability of sparkling wines. The proteolysis of must proteins originating from botrytized grapes is well known, but far less information is available concerning the effect of grape juice contamination by Botrytis. The impact from Botrytis on the biochemical and physico-chemical characteristics of proteins released from Saccharomyces during alcoholic fermentation remains elusive. To address this lack of knowledge, a model grape juice was inoculated with three enological yeasts with or without the Botrytis culture supernatant. Size exclusion chromatography coupled to multi-angle light scattering (SEC-MALLS) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) techniques (AgNO₃ and periodic acid Schiff staining) was used in the study. When Botrytis enzymes were present, a significant degradation of the higher and medium MW molecules released by Saccharomyces was observed during alcoholic fermentation whilst the lower MW fraction increased. For the three yeast strains studied, the results clearly showed a strong decrease in the wine foamability when synthetic musts were inoculated with 5% (v/v) of Botrytis culture due to fungus proteases.

In sparkling wine cool-climate regions like Champagne, it is sometimes necessary to pick the healthy grape clusters that have a relatively low maturity level to avoid the deleterious effects of Botrytis cinerea. In such conditions, we know that classical oenological parameters (sugars, pH, total acidity) may change but there is little information concerning the impact of grape berry maturity on wine proteins and foaming properties. Therefore, healthy grapes (Chardonnay and Pinot meunier) in 2015 and 2016 were picked at different maturity levels within the range of common industrial maturity for potential alcohol content 8–11% v/v in the Champagne region. Base wine protein content and foamability, and oenological parameters in grape juice and their corresponding base wines, were investigated. The results showed that base wine protein contents (analyzed by the Bradford method and by electrophoresis) and foamability were higher when the grapes were riper. The Pearson’s correlation test found significant positive correlations (r = 0.890–0.997, p < 0.05) between Chardonnay grape berry maturity degree (MD) and base wine foamability in both vintages. Strong correlations between MD and most of the oenological parameters in grape juice and base wine were also found for the two cultivars. Under the premise of guaranteed grape health, delaying harvest date is an oenological decision capable of improving base wine protein content and foamability.

Botrytis cinerea is an important fungal pathogen particularly dreaded in the cool climate vineyard. It is responsible for important damage, especially the decrease in foamability of sparkling wines, such as Champagne. Different studies have shown that proteins are largely involved in the stabilization of Champagne foam despite their low concentration. Other works demonstrated changes in the electrophoretic characteristics of must proteins originating from botrytized grapes, although the cause of such alterations was never explained. In the first part of this study, results showed the release by B. cinerea of 3.5 mg/L total proteins in a synthetic liquid medium. Among these proteins, the presence of a protease activity on bovine serum albumin (BSA) and must proteins was demonstrated by using a colorimetric method and sodium dodecyl sulfate−polyacrylamide gel electrophoresis. In the model wine, the Bradford method showed a BSA loss of 66% after 24 h and a loss of 96% after 120 h. In the same model wine, the soluble must protein concentration decreased by 35% after 1 week and by 53% after 2 weeks while the control showed no protein loss. B. cinerea proteases were then able to degrade BSA and must proteins and were above all active at must and wine pH and in the presence of ethanol and SO₂. The second part of this work was dedicated to the relationship between the presence of B. cinerea proteases and its effects on the synthetic wine foaming properties. The addition of a B. cinerea culture medium (1/33 v/v) to the synthetic wine containing 21 mg/L soluble grape proteins induced a decrease in foamability by 60% after 1 week. For BSA in the model wine, the foamability decreased by 32% after 24 h and by 95% after 120 h, as shown by the colorimetric method. These experiments demonstrate for the first time the relationship between B. cinerea protease activity and the decrease in wine foaming properties. Keywords: Botrytis cinerea; protease; foam; grape proteins; BSA; Bradford method; SDS−PAGE; grape juice

A Sauvignon white wine was fined with a dry bentonite (BS), e.g directly incorporated in the wine without swelling treatment, or with the same bentonite swelled in water 24 hours before utilization (BG). The volumes of lees generated by the fining were situated between 0,27 and 1,51 % (v/v) for BS, and between 1,07 and 4,59 % (v/v) for BG. The relation between the quantity of bentonite introduced in the wine (g/hl) and the volume of lees (v/v) follows a power law. The volume of lees only increased by 70 % when the dose of swelled bentonite was doubled; for higher doses of bentonite (rarely used), one can observe a packing of the lees. For BS, lees were approximately twice more volumetric when the dose was doubled. We also observe very good relationship between the clarifying efficiency and the volume of lees. When the clarifying efficiency of BG increased by 10 % the volume of lees increased by 74 %. For BS, when the clarifying efficiency increased by 10 % the volume of lees increased by 86 % because of its low efficiency during clarification. The decrease of the natural proteic haze risk and the volume of lees generated by bentonite fining were also closely correlated by an exponential law. For BG, the mathematical law showed that when the volume of lees increased by 0,1 % (v/v), colloidal haze risk decreased only by 22 %. On the contrary, for BS, when the volume of lees increased by 0,1 %, proteic haze risk decreased by 44 %. Finally, the estimation of the cost of fining showed that the use of a non swelled bentonite was economically more interesting than the utilization of a swelled bentonite. This was true when this Sauvignon wine is sold both in bottle or in bulk. For the studied wine, the estimated winning was 115 euros for 10 hl sold in bulk.

The bovine spongiform encephalopathy (BSE, mad cow) crisis has led some wine-makers to question gelatin as a fining agent and to reject the use of these animal proteins. The search for a substitute for gelatin was begun by comparing vegetable proteins (particularly gluten) with gelatin fining treatments. Clairette de Die (French-controlled appellation) is a sparkling sweet wine. The alcoholic fermentation begins in a tank, continues in a crown-cap bottle, and stops before the complete consumption of sugar. All particles present in the bottle have to be removed before corking, and it is important to have a must as clear as possible. This study concerned the clarifying of a Muscat must, treated with pectinases, using a very efficient flotation technique. The must is fined with bentonite/silica/protein (fish gelatin, wheat gluten, or lupin isolate) to induce flocculation and then pressurized (6 bar). After depressurization, microbubbles cling to flocculates and climb to the top of the flotation tank (this flotation foam is clarified using a rotary filter). At laboratory scale, gluten (20 g/hL) and gelatin (10 g/hL) (each combined with bentonite and silica gel) gave turbidities of 50 and 35 NTU, respectively (6.5 and 4.1% of that of the nontreated must). The Muscat must was also clarified by static settling. The turbidity decreased by 86% for the gluten/bentonite fining and by 60% for the gelatin/bentonite fining. Visually, gluten flocculation takes longer to occur and flocculates sedimentation is longer than with gelati, but the removal of insoluble particles is more complete and leads to lower turbidities. At an industrial scale, gluten (20 g/hL) and gelatin (10 g/hL) (each combined with bentonite and silica gel) gave turbidities of 60 and 48 NTU, respectively. Turbidities measured in the tanks 14 h after the flotation showed a better efficiency for the wheat gluten (24 NTU) compared to gelatin (28 NTU). This is explained by the static settling that completed the clarifying effect of the flotation. The last experiment showed comparable efficiencies for wheat gluten and lupin proteins. BSE caused a situation of crisis (in Europe particularly) leading the public and wine-makers to lose their confidence in the use of gelatin as fining agent and to reject animal proteins in general. It is proposed here that vegetable proteins could efficiently replace gelatin. Keywords: Flotation; vegetable proteins; wheat gluten; must; Muscat; bentonite; gelatin