Growth and metabolism of halophilic Candida versatilis and Tetragenococcus halophilus in simultaneous and sequential fermentation of salted soy whey.

This study investigated various strategies: mono-, simultaneous and sequential fermentation of halophilic Candida versatilis and Tetragenococcus halophilus to valorize salted whey, a side stream of salted tofu (pressed beancurd) production, with an ultimate goal of creating a soy sauce-like condiment. Growth, glucose, organic acids were monitored throughout fermentation, while free amino acids and volatile compounds were analyzed on the final days. In monoculture fermentation, both C. versatilis and T. halophilus thrived in salted soy whey. However, in co-culture fermentation, an antagonistic relationship was observed, wherein C. versatilis growth was slightly suppressed and T. halophilus was significantly inhibited. In C. versatilis-involved fermentations, no significant (p > 0.05) differences in key volatile and non-volatile chemical components were found among various fermentation modes. Key soy sauce-like volatile compounds, such as 4-ethylguaiacol and 4-ethylphenol, were detected in all C. versatilis-fermented salted soy whey, while T. halophilus primarily functioned as a lactic and acetic acids producer. This study highlights the potential of mixed culture fermentation involving soy sauce yeast and lactic acid bacteria for eventually developing a soy sauce-like condiment from salted soy whey, with C. versatilis playing a crucial role in flavour development. The findings suggest that fermenting of a single culture of C. versatilis in lactic acid-adjusted salted soy whey could be a viable and efficient choice for future production of soy sauce-like condiment.

Enhancing the flavour quality of Laiyang pear wine by screening sorbitol-utilizing yeasts and co-fermentation strategies.

This study took a novel approach to address the dual challenges of enhancing the ethanol content and aroma complexity in Laiyang pear wine. It focused on sorbitol as a pivotal element in the strategic selection of yeasts with specific sorbitol-utilization capabilities and their application in co-fermentation strategies. We selected two Saccharomyces cerevisiae strains (coded as Sc1, Sc2), two Metschnikowia pulcherrima (coded as Mp1, Mp2), and one Pichia terricola (coded as Tp) due to their efficacy as starter cultures. Notably, the Sc2 strain, alone or with Mp2, significantly increased the ethanol content (30% and 16%). Mixed Saccharomyces cerevisiae and Pichia terricola fermentation improved the ester profiles and beta-damascenone levels (maximum of 150%), while Metschnikowia pulcherrima addition enriched the phenethyl alcohol content (maximum of 330%), diversifying the aroma. This study investigated the efficacy of strategic yeast selection based on sorbitol utilization and co-fermentation methods in enhancing Laiyang pear wine quality and aroma.

Fermentation of coffee fruit with sequential inoculation of Lactiplantibacillus plantarum and Saccharomyces cerevisiae: effect on sensory attributes and chemical composition of the beans.

This study has innovative aspects related to the use of sequential inoculation technique in the coffee bean fermentation process: the inoculation of Lactiplantibacillus plantarum followed by Saccharomyces cerevisiae, in the fermentation of coffee fruit for the production of specialty natural coffees. The objective was to evaluate the effect of this technique and of the total fermentation time on the sensory attributes of the coffee beverage and on the organic acid profile, bioactive compounds, and fatty acid profile of the beans. The fermentation of coffee fruit with sequential inoculation resulted in greater acidity of the beverage and contributed to increases of up to 2 points in coffee fermented. The total fermentation time was directly related to the organic acid content, and the longer the total fermentation time was, the greater the organic acid content. The fatty acid content and bioactive compound content showed little variation among treatments.

Fermentation of coffee fruit with sequential inoculation of Lactiplantibacillus plantarum and Saccharomyces cerevisiae: Effects on volatile composition and sensory characteristics.

Mixed starter cultures of lactic acid bacteria and yeasts used in the production of fermented foods, including coffee, can improve the sensory quality and food safety. The objective of this study was to evaluate the effects of fermentation of coffee with inoculation of Lactiplantibacillus plantarum followed by Saccharomyces cerevisiae and the effects of fermentation time on the aroma and flavor of the coffee beverage and on the volatile composition of the roasted coffee beans. The coffee was fermented for 48 h or 96 h after inoculation of Lactiplantibacillus plantarum followed by inoculation of Saccharomyces cerevisiae or the respective controls. The aroma and flavor of the coffee beverage fermented with sequential inoculation showed complexity, with a predominance of fruity and fermented sensory notes. Forty-seven volatile compounds were identified. In addition, the sequentially inoculated coffees had greater formation of volatiles and led to greater perception of fruity and fermented flavor and aroma.

Effect of Sequential Inoculation of Tetragenococcus halophilus and Wickerhamomyces anomalus on the Flavour Formation of Early-Stage Moromi Fermented at a Lower Temperature.

Microbial inoculation in moromi fermentation has a great influence on the physicochemical and flavour properties of soy sauces. This work investigated the effect of inoculating Tetragenococcus halophilus and Wickerhamomyces anomalus on the flavour formation of early-stage moromi (30 days) fermented at a lower temperature (22 °C) by determining their physicochemical and aroma changes. The results showed that single yeast or LAB inoculation increased the production of amino nitrogen, lactic acid and acetic acid, as well as free amino acids and key flavour components. Particularly, the sequential inoculation of T. halophilus and W. anomalus produced more free amino acids and aromatic compounds, and there might be synergistic effects between these two strains. More characteristic soy sauce flavour compounds, such as benzaldehyde, HEMF, guaiacol and methyl maltol were detected in the sequentially inoculated moromi, and this sample showed higher scores in savoury, roasted and caramel intensities. These results confirmed that sequential inoculation of T. halophilus and W. anomalus could be a choice for the future production of moromi with good flavour and quality under a lower temperature.

Impact of Candida railenensis during fermentation on the aromatic profile of Vidal blanc icewine.

Mixed culture fermentation with non-Saccharomyces yeasts and Saccharomyces cerevisiae as multi-starters has more advantages than spontaneous fermentation, and wine products with distinctive and pleasant aromas can fulfill the diverse demands of consumers. This study was carried out to illuminate the effect of sequential inoculation of indigenous Candida railenensis and S. cerevisiae on alcoholic fermentation behavior and chemical and aromatic characteristics of Vidal blanc icewine. During the mixed culture fermentation, C. railenensis was present in the initial and middle stages but was absent after 14 days. The results of basic chemical parameters showed that the glycerol content in the mixed culture-fermented icewine was higher than that of the pure fermented icewine, but the acetic acid content was the opposite. In terms of volatile aroma compounds, C. railenensis in the mixed culture fermentation reduced some metabolites such as lower alcohols, 1-hexanol, 3-methylthiopropanol, and their unpleasant notes and increased the production of some desired volatile aroma compounds such as benzaldehyde, ß-damascenone, 2-furanmethanol, and 5-methyl furfural associated with rose, honey, nut, and caramel characteristics. Furthermore, C. railenensis also changed the sensory performance of icewine by participating in the fermentation of S. cerevisiae. These findings suggest that C. railenensis with positive enological properties has the ability to be used in icewine production, which has never been reported before.

Transforming Spent Coffee Grounds' Hydrolysates with Yeast Lachancea thermotolerans and Lactic Acid Bacterium Lactiplantibacillus plantarum to Develop Potential Novel Alcoholic Beverages.

In the present work, the modification of spent coffee grounds (SCG) hydrolysate composition by mixed cultures of a non-Saccharomyces yeast, Lachancea thermotolerans, and a lactic acid bacterium, Lactiplantibacillus plantarum, as well as their interactions, were evaluated. It was found that L. plantarum inhibited the growth and survival of L. thermotolerans as compared with that in the yeast alone. On the other hand, the growth and survival of L. plantarum was slowed in sequential fermentation, but not in co-culture. Compared with co-culture, higher ethanol content, less residual sugars, and less acetic and succinic acids were found in sequential fermentation. In addition, lower amounts of caffeine and phenolic acids (e.g., ferulic, caffeic, and p-coumaric acids) were obtained in mixed (co- and sequential) cultures with corresponding levels of volatile phenols relative to the yeast monoculture. Moreover, co-culturing resulted in the highest contents of total alcohols (ethanol excluded) and total esters. Therefore, mixed culturing of L. plantarum and L. thermotolerans presented positive effects on the chemical constituents of fermented SCG hydrolysates, which might be a new alternative approach to valorizing the SCG into novel alcoholic drinks with different ethanol and flavor constituents.

Contribution of Starmerella bacillaris and Oak Chips to Trebbiano d'Abruzzo Wine Volatile and Sensory Diversity.

In this study, six fermentation trials were carried out: co-inoculation and sequential inoculation of Saccharomyces cerevisiae and Starmerella bacillaris in the presence and absence of oak chips. Moreover, Starm. bacillaris strain was attached to the oak chips and co-inoculated or sequentially inoculated with S. cerevisiae. Wines fermented with Starm. bacillaris adhered to oak chips showed a higher concentration of glycerol (more than 6 g/L) than the others (about 5 g/L). These wines also showed a higher content of polyphenols (more than 300 g/L) than the others (about 200 g/L). The addition of oak chips induced an increase of yellow color (b* value of about 3). Oak-treated wines were characterized by a higher concentration of higher alcohols, esters and terpenes. Aldehydes, phenols and lactones were detected only in these wines, independently from the inoculation strategy. Significant differences (p < 0.05) were also observed in the sensory profiles. The fruity, toasty, astringency, and vanilla sensations were perceived as more intense in wines treated with oak chips. The white flower descriptor showed a higher score in wines fermented without chips. Oak surface-adhered Starm. bacillaris cells could be a good strategy to improve the volatile and sensory profile of Trebbiano d'Abruzzo wines.

Diversity of Volatile Aroma Compound Composition Produced by Non-Saccharomyces Yeasts in the Early Phase of Grape Must Fermentation.

There is a lack of studies evaluating the metabolic contribution of non-Saccharomyces yeasts in early fermentation phases. This study aimed to investigate the volatile aroma profiles produced by various non-Saccharomyces yeasts just before sequential inoculation with Saccharomyces cerevisiae to provide an insight into the particular effects they induce at this stage. The grape must of Malvazija istarska was inoculated with monocultures of Torulaspora delbrueckii, Metschnikowia pulcherrima, Pichia kluyveri, Lachancea thermotolerans, and Schizosaccharomyces pombe, alongside a S. cerevisiae control. Eighty volatile compounds were quantified via headspace solid-phase microextraction and gas chromatography-mass spectrometry, and the data were statistically elaborated. Volatile profiles of non-Saccharomyces yeasts differed significantly from the S. cerevisiae control. Most treatments caused increases in linalool and ß-damascenone, decreases in higher alcohols and fatty acids, and improved synthesis of odoriferous esters. Torulaspora delbrueckii and M. pulcherrima produced compounds not commonly found in S. cerevisiae fermented wines. Multivariate statistical analysis linked the investigated yeasts to specific, particularly abundant compounds. Future studies should explore to what degree these contributions persist after sequential inoculation with S. cerevisiae in diverse grape must matrices.

Effects of sequential inoculation of Lachancea thermotolerans and Oenococcus oeni on chemical composition of spent coffee grounds hydrolysates.

Spent coffee grounds (SCG) disposal is an environmental problem. These residues from coffee brewing and instant coffee production have potential to produce novel alcoholic beverages. SCG valorization through sequential alcoholic and malolactic fermentation was investigated using a yeast, Lachancea thermotolerans Concerto and a lactic acid bacterium (LAB), Oenococcus oeni Lalvin 31 in this study. Our results showed that sequential inoculation prevented early death of yeast confronted when simultaneous inoculation was adopted, allowing for growth and persistence of both yeast and LAB till the end of fermentation. Adequate ethanol production (4.91 ± 0.13 %, v/v) with low residual sugar content was also attained. In addition, relatively lower levels of acetic, lactic, and succinic acids were produced by sequential inoculation than that of simultaneous inoculation. Furthermore, SCG hydrolysates fermented via sequential inoculation had the widest variety of volatiles (e.g. esters and ketones). Overall, our results indicated that sequential inoculation of L. thermotolerans and O. oeni in SCG hydrolysates might be a way to develop novel beverages with pleasant flavor profiles.

Comparison of fungal communities and nonvolatile flavor components in black Huangjiu formed using different inoculation fermentation methods.

Black Huangjiu (BH) is a traditional alcoholic beverage in China, which is very popular among people. The different methods (simultaneous inoculation, sequential inoculation), were applied to ferment BH in this study, which were investigated the changes in the composition of fungal communities and non-volatile flavor compounds (nVFCs) by high-throughput sequencing (HTS) and ultraperformance liquid chromatography-tandem mass spectrometer (UPLC MS/MS). The results showed that Rhizopus and Saccharomyces were the predominant fungal genera throughout fermentation, and 471 nVFCs were detected in BH after fermentation. Compared to that observed simultaneous inoculation, Rhizopus increased at the end of sequential fermentation, and the contents of the organic acids and their derivatives increased significantly [variable importance in the projection (VIP) > 1.0, p < 0.05, fold change (FC) > 2], while that of lipids and lipid-like molecules decreased significantly (VIP > 1.0, p < 0.05, FC < 0.5). Through the correlation analysis of 32 nVFCs with significant differences (VIP > 1.0, p < 0.05, FC >32 or < 0.03) and the community, it was found that lipids and lipid-like molecules (12) and organic acids and their derivatives (10) were significantly (p < 0.05) negatively correlated with Saccharomyces, but they were significantly (p < 0.05) positively correlated with Rhizopus. Compared with simultaneous inoculation, BH fermented by sequential inoculation, the taste was stronger, sweeter, mellow, and softer. Our findings provide information on nVFC dynamics and will aid in the selection of beneficial strains to improve BH quality.

Different approaches to supplement polysaccharide-degrading enzymes in vinification: Effects on color extraction, phenolic composition, antioxidant activity and sensory profiles of Malbec wines.

Enzymes play an essential role in the winemaking process. Besides an improvement in the color of the wine it also enhances the quality, which makes it well worth to develop new strategies of application. The present study examined two approaches to apply microbial enzymes during different winemaking techniques using Malbec grapes from the San Rafael wine region, Mendoza, Argentina. i) Torulaspora delbrueckii was used in sequential inoculation with an autochthonous Saccharomyces cerevisiae strain. The former strain was first inoculated to produce in situ an enzyme complex during winemaking for four days, and subsequently the latter strain was added. The process was carried out under traditional winemaking conditions at 22 °C (Td-Sc). ii) A second approach was through inoculation of a multi-enzyme extract produced by Aureobasidium pullulans m11-2 during pre-fermentative cold maceration (PCM) for four days at 8 °C (PCM-E), and S. cerevisiae was inoculated at the end of the PCM. Controls were carried out without enzymatic treatments (Sc and PMC, respectively). T. delbrueckii started the fermentation and remained viable throughout the process. The fermentation kinetics were not affected in the mixed culture compared with a pure culture of S. cerevisiae. Monitoring of the pectinolytic activity of Td-Sc showed maximum activity (14.2 U mg-1) on day one of the inoculation, while in PCM-E the must activity increased during the PCM until day one of fermentation, reaching a maximum of 16.85 U mg-1. Both enzyme-treated wines maintained their red hue, regardless of the duration of the macerations, while the control wines were characterized by a faster rate of color loss and more yellow tones. In the sensory evaluation, enzymatically treated wines presented greater color intensity, redder and more violet tones, floral and fruit aromas, which are also the distinctive features of the Malbec variety. Anthocyanin values obtained in PCM and in PCM-E were higher than in vinifications without cold maceration (Td-Sc and Sc), which could be attributed to the effectiveness of the maceration technique. Similarly, the composition of stilbenes, especially trans-resveratrol, showed an increase in PCM techniques with or without enzymatic treatment, and both techniques were comparable to one other and statistically superior to traditional winemaking.

Indigenous Non-Saccharomyces Yeasts With ß-Glucosidase Activity in Sequential Fermentation With Saccharomyces cerevisiae: A Strategy to Improve the Volatile Composition and Sensory Characteristics of Wines.

Non-Saccharomyces (NS) yeasts with high ß-glucosidase activity play a vital role in improving the aroma complexity of wines by releasing aroma compounds from glycosidic precursors during fermentation. In this study, the effect of sequential inoculation fermentation of Meyerozyma guilliermondii NM218 and Hanseniaspora uvarum BF345 with two Saccharomyces cerevisiae strains [Vintage Red™ (VR) and Aroma White™ (AW)] on volatile compounds and sensory characteristics of wines was investigated. Prior to winemaking trials, the sequential inoculation times of the two NS yeasts were evaluated in synthetic must, based on changes in strain population and enzyme activity. The intervals for inoculation of NM218 and BF345 with the S. cerevisiae strains were 48 and 24 h, respectively. In the main experiment, sequential inoculation fermentations of the two strains with S. cerevisiae were carried out in Cabernet Sauvignon (CS) and Chardonnay (CH) grape must. The oenological parameters, volatile composition, and sensory characteristics of the final wines were assessed. No clear differences were observed in the oenological parameters of the sequentially fermented CH wines compared with the control, except for residual sugar and alcohol. However, in CS wines, the total acid contents were significantly lower in the wines fermented by sequential inoculation compared to the control. Both NM218 and BF345 improved the aroma complexity of wines by increasing esters and terpenes when inoculated with S. cerevisiae strains compared to inoculation with S. cerevisiae strains alone. NM218 resulted in a more positive effect on CS wine aroma, with higher levels of citronellol and trans-nerolidol. BF345 significantly enhanced the floral and fruity aromas of CH wine by producing higher concentrations of geranyl acetone, ß-damascenone, trans-nerolidol, and nerol. Both NM218 and BF345 yeasts could potentially be used to improve wine aroma and overall quality, especially wine floral and fruity aromas, when used in sequential inoculation with S. cerevisiae.

Technological Improvements on FML in the Chianti Classico Wine Production: Co-Inoculation or Sequential Inoculation?

Winemaking variables and techniques are known to affect the composition of wines. To obtain a rapid and safe fermentation course, with a reduced risk of proliferation of unwanted microbial species, frequent recourse is made to the use of selected microorganisms, which can greatly simplify the complex management of the fermentation process. In particular, selected strains of lactic acid bacteria are used, which are much more sensitive than yeasts to the operating conditions of the medium. In this regard, the overall aim of this research was to verify whether the early inoculation of homolactic acid bacteria for hexoses (Lactobacillus plantarum) carried out after 24 h, compared with that of saccharomycetes operating alcoholic fermentation, could be advantageous compared with a traditional innoculation with a different heterolactic bacterial strain for hexoses (Oenococcus oeni) operated at the end of alcoholic fermentation. The grape variety chosen was Sangiovese, the protagonist of Tuscan oenology. The evaluation focused on different aspects such as the management of winery operations, and the quality and longevity of the product; was carried out in all phases of winemaking; and analysed both from a chemical and sensory point of view.

The content of biogenic amines in Rondo and Zweigelt wines and correlations between selected wine parameters.

The purpose of this study was to evaluate the content of biogenic amines (BAs) in wines using dispersive liquid-liquid microextraction-gas chromatography-mass spectrometry (DLLME-GC-MS). An additional objective was to assess the correlations between selected parameters characterizing the samples such as the content of BAs, sugars, and organic acids, pH, and total acidity. Wines produced from the same grape variety in which alcoholic fermentation (AF) was carried out by different yeast strains and in which malolactic fermentation (MLF) was spontaneous, differed in the content of biogenic amines. The concentrations of putrescine, cadaverine and tryptamine were higher in the Rondo wines (237-405, 34.04-61.11,

Combinatorial analysis of population dynamics, metabolite levels and malolactic fermentation in Saccharomyces cerevisiae/ Lachancea thermotolerans mixed fermentations.

The outcome of co- or sequential inoculation of Lachancea thermotolerans in winemaking remains unpredictable due to a lack of integrated data regarding the impact of grape juice composition on L. thermotolerans fermentation behaviour. Here, we investigate the impact of nitrogen composition on fermentation characteristics and aroma compound production in grape juice sequentially inoculated with commercial L. thermotolerans and S. cerevisiae strains. Subsequently, all treatments were subjected to malolactic fermentation (MLF) using two commercial strains of Oenococcus oeni. Addition of amino acids led to faster growth for S. cerevisiae fermentations, compared to the nitrogen-equivalent addition of diammonium phosphate (DAP). L. thermotolerans persistence in the mixed fermentations was significantly higher following DAP addition, with higher glycerol and lactic acid production. Interestingly, the lower total Nitrogen content in DAP-treated musts compared to other treatments did not alter the subsequent growth of S. cerevisiae. MLF was more similar between musts fermented with L. thermotolerans, regardless of nutrient regime, whereas significant differences in MLF completion times were observed for different nitrogen treatments in S. cerevisiae fermentations. Collectively, the data present an integrated view of the impact of nitrogen treatment on multispecies co-inoculation (growth kinetics and aromatic outcomes) and the downstream impact on MLF.

Impact of Sequential Inoculation with the Non-Saccharomyces T. delbrueckii and M. pulcherrima Combined with Saccharomyces cerevisiae Strains on Chemicals and Sensory Profile of Rosé Wines.

Controlled inoculations of non-Saccharomyces yeasts are becoming increasingly used to produce high-quality wines due to their enological potential. In this study, we evaluated the impact of sequential inoculation with the commercial non-Saccharomyces yeasts (Torulaspora delbrueckii and Metschnikowia pulcherrima) in combination with Saccharomyces cerevisiae on the chemical and sensory profile of rosé wines. Sequential inoculation with T. delbrueckii produced wines with an overall reduction in esters, mainly explained by the lower concentrations of ethyl esters of medium-chain fatty acids and isoamyl acetate. The lower ester concentrations of these wines were related to a reduction in fruity descriptors. An increase was observed, however, in other minor esters such as cinnamates and ethyl esters of branched acids. Zinc, ethyl isobutyrate, and ethyl dihydrocinnamate were selected as potential markers for this fermentation strategy. Sequential inoculation with M. pulcherrima resulted in rosé wines with an enhanced ester profile, reduced acetaldehyde, and increased anthocyans and tannins. Compared to the control wines fermented with S. cerevisiae, the changes observed in these wines were far subtler, especially for the volatile profile, sensory characteristics, and color parameters, with isobutyl hexanoate and isoamyl butyrate being selected as potential markers.

A Rapid Method for Selecting Non-Saccharomyces Strains with a Low Ethanol Yield.

The alcohol content in wine has increased due to external factors in recent decades. In recent reports, some non-Saccharomyces yeast species have been confirmed to reduce ethanol during the alcoholic fermentation process. Thus, an efficient screening of non-Saccharomyces yeasts with low ethanol yield is required due to the broad diversity of these yeasts. In this study, we proposed a rapid method for selecting strains with a low ethanol yield from forty-five non-Saccharomyces yeasts belonging to eighteen species. Single fermentations were carried out for this rapid selection. Then, sequential fermentations in synthetic and natural must were conducted with the selected strains to confirm their capacity to reduce ethanol compared with that of Saccharomyces cerevisiae. The results showed that ten non-Saccharomyces strains were able to reduce the ethanol content, namely, Hanseniaspora uvarum (2), Issatchenkia terricola (1), Metschnikowia pulcherrima (2), Lachancea thermotolerans (1), Saccharomycodes ludwigii (1), Torulaspora delbrueckii (2), and Zygosaccharomyces bailii (1). Compared with S. cerevisiae, the ethanol reduction of the selected strains ranged from 0.29 to 1.39% (v/v). Sequential inoculations of M. pulcherrima (Mp51 and Mp FA) and S. cerevisiae reduced the highest concentration of ethanol by 1.17 to 1.39% (v/v) in synthetic or natural must. Second, sequential fermentations with Z. bailii (Zb43) and T. delbrueckii (Td Pt) performed in natural must yielded ethanol reductions of 1.02 and 0.84% (v/v), respectively.

The aptitude of commercial yeast strains for lowering the ethanol content of wine.

The high alcohol content in wine usually has a negative impact on its sensory properties, but can also affect the general health of the consumers. The possibility to reduce ethanol production in wines during fermentation involves the use of different yeast strains characterized by the increased production of fermentation by-products (glycerol, 2,3-butanediol, etc.) from the available sugar. The activity of these strains should not impair the sensory properties of the wine. In general, the use of genetically and evolutionarily (non-GM) engineered Saccharomyces cerevisiae strains is still not close enough to commercial application, and therefore, it is unavailable for wine producers. Thus, the aim of this study was to examine the possibility of reducing the production of ethanol in wines using different selected yeast strains (S. cerevisiae, Saccharomyces bayanus, Torulaspora delbrueckii, and Metschnikowia pulcherrima) available at the market. The application of individual yeast and sequential inoculation for wine alcoholic fermentation was examined. The achieved effects were evaluated by determining the content of ethanol, as well as fermentation by-products (glycerol and volatile acids) and aromatic components in wine samples. Depending on the strain/s used, a decrease in ethanol content of up to 0.9% v/v was recorded in comparison with fermentation by S. cerevisiae alone. The sensory analysis of produced wine showed significant differences in taste and flavor. The results of the experiment conducted at the laboratory level and with the use of sterile must were compared to the ones from the scale-up experiment in real vinification conditions. The observed differences in the alcohol content of produced wines were significantly lower.

Impact of Nutrient Availability on the Fermentation and Production of Aroma Compounds Under Sequential Inoculation With M. pulcherrima and S. cerevisiae.

Non-Saccharomyces yeasts are currently widely used in winemaking to enhance aroma profile diversity among wines. The use of Metschnikowia pulcherrima in sequential inoculation with S. cerevisiae was compared to the inoculation of a pure culture of S. cerevisiae. Moreover, various concentrations of sugar, nitrogen and lipids were tested in synthetic must to assess their impact on fermentation and its outcomes using a Box-Behnken design. Due to its phenotypic specificities, early inoculation with M. pulcherrima led to important modifications, first altering the fermentation kinetics. This may relate, at least in part, to the depletion of some nitrogen sources by M. pulcherrima during the first part of fermentation. Beyond these negative interactions on fermentation performance, comparisons between pure cultures and sequentially inoculated cultures revealed changes in the distribution of carbon fluxes during fermentation in presence of M. pulcherrima, resulting in a positive impact on the production of central carbon metabolites and aromas. Furthermore, the expression of varietal thiols was strongly increased as a consequence of positive interactions between the two species. The mechanism of this release still needs to be investigated. Significant differences in the final concentrations of fermentative and varietal aromas depending on the initial must composition were obtained under both inoculation strategies. Interestingly, the response to changes in nutrient availability varied according to the inoculation modality. In particular, a greater incidence of lipids on the production of fatty acids and their ethyl esters derivatives was found during sequential fermentation compared with pure culture, to be viewed in combination with the metabolic characteristics of M. pulcherrima regarding the production of volatile compounds from acetyl-CoA. Overall, the importance of managing nutrient availability under M. pulcherrima/S. cerevisiae sequential inoculation in order to derive the maximum benefit from the potentialities of the non-Saccharomyces species while carrying out fermentation to dryness was highlighted.