Yeast Starter Research Articles

Modulation of the volatile and non-volatile profiles of coffee fermented with Yarrowia lipolytica: I. Green coffee

Yeast starter cultures have been used in the fermentation of different coffee substrates to modulate the volatile and aroma profiles of coffees with the exception of green coffee beans. This two-part study aimed to modulate the volatile profiles of roasted coffee via the fermentation of green coffee beans with a non-conventional aerobic yeast Yarrowia lipolytica. The objective of part I of this study was to evaluate the effects of Y. lipolytica fermentation on the volatile and non-volatile profiles of green coffee beans. After fermentation, sugars concentration decreased by 1.2-fold while the amino and phenolic acids concentrations decreased by 1.3-fold. The decrease in phenolic acid concentration could account for the 1.5-fold increase in the volatile phenols levels resulting from phenolic acid catabolism. The degradation of l-phenylalanine via the Ehrlich pathway by the yeast led to a 1.9-fold increase in 2-phenylethanol levels, whereas the decreases in alkanes, acids and aldehydes levels were attributed to the hydrophobic substrate metabolizing pathways that were unique to Y. lipolytica. Hence, this work showed that Y. lipolytica fermentation of green coffee beans resulted in the modification of the aroma precursors and volatile composition of green coffees.

Presence of Candida zemplinina in Sicilian Musts and Selection of a Strain for Wine Mixed Fermentations

The purpose of this work was to investigate the presence of C. zemplinina yeasts in Sicilian musts and grapes and to identify strains of oenological interest. We report on the taxonomical reclassification of Candida yeast isolates from Sicilian musts and on the selection of one strain of oenological interest (Cz3), based on mixed micro-fermentation experiments in sterile Nero d’Avola musts. Our results show that Candida zemplinina is abundant in Sicilian grapes and musts, and that the Cz3 strain is suitable for Candida zemplinina/Saccharomyces cerevisiae mixed fermentations. The higher glycerol content and the lower ethanol level stood out as the most promising features of the wines obtained upon sequential inoculation of the Cz3 and (S. cerevisiae) NDA21 yeast starters. We therefore have isolated a Sicilian Cz strain endowed with very promising features for the future development of mixed fermentation protocols.

Implementing principles of traditional concentrated grape must fermentation to the production of new generation balsamic vinegars. Starter selection and effectiveness.

In an effort to implement principles of traditional concentrated grape must fermentation to the production of new generation balsamic vinegars (BVs), the specific goals of the study were the isolation and molecular identification of the predominant yeasts in concentrated grape must (cv. Xinomavro), their technological characterization and the evaluation of the fermentative aptitude of the selected strains. Tolerance against 5-hydroxymethyl-furfural (HMF) and furfural, acetic acid and glucose concentration was examined by appropriate methods and tests. The enological characteristics studied were acetic acid and H2S production, foaming and flocculation ability and key enzymatic activity. PCR-RFLP analysis revealed only the presence of Saccharomyces cerevisiae and Hanseniaspora uvarum among the 14 predominant osmophilic yeast isolates. Tolerance to both HMF and furfural was found strain- and dose-dependent and was suggested as a critical factor in the pre-selection of yeast starters. The most tolerant yeasts to these stress factors, a S. cerevisiae and a non-Saccharomyces strains, showed satisfactory growth in the presence of high glucose and acetic acid content (up to 600g/L and 2% w/w, respectively) and desirable enological characteristics. Results from the comparative evaluation of the fermentative aptitude of these strains with a commercial wine strain highlighted that the isolates had glucophilic behaviour and ability to produce desirable amounts of ethanol (100-120g/kg) in short time (~20d). The key volatiles useful for varietal discrimination and differentiation between the BVs and the traditional ones were also evaluated.

Simultaneous Alcoholic and Malolactic Fermentations by Saccharomyces cerevisiae and Oenococcus oeni Cells Co-immobilized in Alginate Beads.

Malolactic fermentation (MLF) usually takes place after the end of alcoholic fermentation (AF). However, the inoculation of lactic acid bacteria together with yeast starter cultures is a promising system to enhance the quality and safety of wine. In recent years, the use of immobilized cell systems has been investigated, with interesting results, for the production of different fermented foods and beverages. In this study we have carried out the simultaneous immobilization of Saccharomyces cerevisiae and Oenococcus oeni in alginate beads and used them in microvinifications tests to produce Negroamaro wine. The process was monitored by chemical and sensorial analyses and dominance of starters and cell leaking from beads were also checked. Co-immobilization of S. cerevisiae and O. oeni allowed to perform an efficient fermentation process, producing low volatile acidity levels and ethanol and glycerol concentrations comparable with those obtained by cell sequential inoculum and co-inoculum of yeast and bacteria cells in free form. More importantly, co-immobilization strategy produced a significant decrease of the time requested to complete AF and MLF. The immobilized cells could be efficiently reused for the wine fermentation at least three times without any apparent loss of cell metabolic activities. This integrated biocatalytic system is able to perform simultaneously AF and MLF, producing wines similar in organoleptic traits in comparison with wines fermented following traditional sequential AF and MLF with free cell starters. The immobilized-cell system, that we here describe for the first time in our knowledge, offers many advantages over conventional free cell fermentations, including: (i) elimination of non-productive cell growth phases; (ii) feasibility of continuous processing; (iii) re-use of the biocatalyst.

Metagenomic sequencing reveals the relationship between microbiota composition and quality of Chinese Rice Wine.

Chinese Rice Wine (CRW) is a common alcoholic beverage in China. To investigate the influence of microbial composition on the quality of CRW, high throughput sequencing was performed for 110 wine samples on bacterial 16S rRNA gene and fungal Internal Transcribed Spacer II (ITS2). Bioinformatic analyses demonstrated that the quality of yeast starter and final wine correlated with microbial taxonomic composition, which was exemplified by our finding that wine spoilage resulted from a high proportion of genus Lactobacillus. Subsequently, based on Lactobacillus abundance of an early stage, a model was constructed to predict final wine quality. In addition, three batches of 20 representative wine samples selected from a pool of 110 samples were further analyzed in metagenomics. The results revealed that wine spoilage was due to rapid growth of Lactobacillus brevis at the early stage of fermentation. Gene functional analysis indicated the importance of some pathways such as synthesis of biotin, malolactic fermentation and production of short-chain fatty acid. These results led to a conclusion that metabolisms of microbes influence the wine quality. Thus, nurturing of beneficial microbes and inhibition of undesired ones are both important for the mechanized brewery.

Non-conventional Yeast Species for Lowering Ethanol Content of Wines

Rising sugar content in grape must, and the concomitant increase in alcohol levels in wine, are some of the main challenges affecting the winemaking industry nowadays. Among the several alternative solutions currently under study, the use of non-conventional yeasts during fermentation holds good promise for contributing to relieve this problem. Non-Saccharomyces wine yeast species comprise a high number or species, so encompassing a wider physiological diversity than Saccharomyces cerevisiae. Indeed, the current oenological interest of these microorganisms was initially triggered by their potential positive contribution to the sensorial complexity of quality wines, through the production of aroma and other sensory-active compounds. This diversity also involves ethanol yield on sugar, one of the most invariant metabolic traits of S. cerevisiae. This review gathers recent research on non-Saccharomyces yeasts, aiming to produce wines with lower alcohol content than those from pure Saccharomyces starters. Critical aspects discussed include the selection of suitable yeast strains (considering there is a noticeable intra-species diversity for ethanol yield, as shown for other fermentation traits), identification of key environmental parameters influencing ethanol yields (including the use of controlled oxygenation conditions), and managing mixed fermentations, by either the sequential or simultaneous inoculation of S. cerevisiae and non-Saccharomyces starter cultures. The feasibility, at the industrial level, of using non-Saccharomyces yeasts for reducing alcohol levels in wine will require an improved understanding of the metabolism of these alternative yeast species, as well as of the interactions between different yeast starters during the fermentation of grape must.

Whole Genome Comparison Reveals High Levels of Inbreeding and Strain Redundancy Across the Spectrum of Commercial Wine Strains ofSaccharomyces cerevisiae

Humans have been consuming wines for more than 7000 yr . For most of this time, fermentations were presumably performed by strains of Saccharomyces cerevisiae that naturally found their way into the fermenting must . In contrast, most commercial wines are now produced by inoculation with pure yeast monocultures, ensuring consistent, reliable and reproducible fermentations, and there are now hundreds of these yeast starter cultures commercially available. In order to thoroughly investigate the genetic diversity that has been captured by over 50 yr of commercial wine yeast development and domestication, whole genome sequencing has been performed on 212 strains of S. cerevisiae, including 119 commercial wine and brewing starter strains, and wine isolates from across seven decades. Comparative genomic analysis indicates that, despite their large numbers, commercial strains, and wine strains in general, are extremely similar genetically, possessing all of the hallmarks of a population bottle-neck, and high levels of inbreeding. In addition, many commercial strains from multiple suppliers are nearly genetically identical, suggesting that the limits of effective genetic variation within this genetically narrow group may be approaching saturation.

Cellar-Associated Saccharomyces cerevisiae Population Structure Revealed High-Level Diversity and Perennial Persistence at Sauternes Wine Estates.

Three wine estates (designated A, B, and C) were sampled in Sauternes, a typical appellation of the Bordeaux wine area producing sweet white wine. From those wine estates, 551 yeast strains were collected between 2012 and 2014, added to 102 older strains from 1992 to 2011 from wine estate C. All the strains were analyzed through 15 microsatellite markers, resulting in 503 unique Saccharomyces cerevisiae genotypes, revealing high genetic diversity and a low presence of commercial yeast starters. Population analysis performed using Fst genetic distance or ancestry profiles revealed that the two closest wine estates, B and C, which have juxtaposed vineyard plots and common seasonal staff, share more related isolates with each other than with wine estate A, indicating exchange between estates. The characterization of isolates collected 23 years ago at wine estate C in relation to recent isolates obtained at wine estate B revealed the long-term persistence of isolates. Last, during the 2014 harvest period, a temporal succession of ancestral subpopulations related to the different batches associated with the selective picking of noble rotted grapes was highlighted. High genetic diversity of S. cerevisiae isolates from spontaneous fermentation on wine estates in the Sauternes appellation of Bordeaux was revealed. Only 7% of all Sauternes strains were considered genetically related to specific commercial strains. The long-term persistence (over 20 years) of S. cerevisiae profiles on a given wine estate is highlighted.

It's Gettin’ Hot in Here: Breeding Robust Yeast Starter Cultures for Cocoa Fermentation

Cocoa beans have to undergo post-harvest fermentation and drying to develop the typical 'cocoa flavor' associated with chocolate. Yeasts play a pivotal role during the fermentation but are generally outcompeted early in the process. Meersman and colleagues describe an elegant breeding-based approach to generate robust yeast starter cultures for cocoa fermentation.

Effect of Maize/Millet Malt Hydrolysis and Baker’s Yeast/Burukutu Starter Fermentation on the Yield of Ethanol from Dioscorea Spp (Yam)

The ethanol production ability of baker’s yeast (Saccharomyces cerevisiae) and burukutu starter culture were examined on yam starch hydrolyzed with malts of maize and millet. Four substrate treatments which include yam starch (as control), yam starch hydrolyzed with millet malt (B), yam starch hydrolyzed with maize malt (C) and yam starch hydrolyzed with a mixture both millet and maize malt (D) were prepared in duplicate and made up to 600 ml. Baker’s yeast (4.40 g) was inoculated on one set of four while burukutu starter was inoculated on the other set of four and fermented for a period of four days at a temperature of 26°C. The pH was observed to drop as fermentation period increased, the highest ethanol yield of up to 74.5 g/Kg was obtained from starch hydrolyzed with maize malt and inoculated with baker’s yeast. Yam is a potential source of industrial scale ethanol production, there was a significant difference (p<0.05) between maize and millet malts in ethanol yield but between baker’s yeast and burukutu starter culture fermentation there was no significant difference (p<0.05), the interaction between malts and the inoculums was significant (p<0.05). Ethanol yield strongly depends on the hydrolysis of yam starch into fermentable sugar.

Design Process of Hydrolysis and Fermentation Bioethanol Production from Seaweed Eucheuma cottonii to Renewable Energy Sovereignity

&lt;p&gt;The aim of this research was study of E. cottonii to produce bioethanol fermentation substrate with a high reduction sugar content and low Hidroxymethilfurfural (HMF). Fermentation done by instant yeast and Saccharomyces cerevisiae culture of FNCC 3012.The best treatment was obtained in the combination of 2% of H2SO4 by time reaction of 120 minutes in 80°C produced 15.61 g/l reducing sugar and 5.03 g/l HMF. In fermented process, the hydrolysate with instant yeast starter delivered much more efficiency in 3.63 ml CO2 volume, 87.53% in fermentation efficiency, and 1.96 g/l reducing sugar on fifth day of fermentation. &lt;br /&gt;&lt;strong&gt;Keywords&lt;/strong&gt;: bioethanol, Eucheuma cottonii, fermentation, hydrolysis, process design&lt;/p&gt;

The impact of yeast starter cultures on the microbial communities and volatile compounds in cocoa fermentation and the resulting sensory attributes of chocolate.

Theobroma cacao seeds are the main raw material for chocolate production. During their fermentation, a succession of microorganisms are responsible for the physicochemical changes occurring in the pulp and inside the beans. The aim of this study was to investigate the effects of yeast inoculation (Saccharomyces cerevisiae UFLA CA11, Pichia kluivery CCMA0237, and Hanseniaspora uvarum CCMA0236) on the profile of the volatile compounds and microbial communities in cocoa fermentation. The resulting chocolate was also evaluated by temporal dominance of sensations (TDS) analyses. The dominant microorganisms during spontaneous fermentation were S. cerevisiae, H. uvarum, H. guilliermondii, Lactobacillus fermentum, Pediococcus sp., and Acetobacter pasteurianus. Similarly, S. cerevisiae, P. kluyveri, Candida sp., Pediococcus sp., and A. pasteurianus were the predominant microorganisms assessed by Denaturing Gradient Gel Electrophoresis (DGGE) in inoculated fermentation. Sixty-seven volatile compounds were detected and quantified by gas chromatography/mass spectrometry (GC/MS) at the end of fermentation and chocolates. The main group of volatile compound found after the inoculated and spontaneous fermentations was esters (41 and 39%, respectively). In the chocolates, the main group was acids (73 and 44% from the inoculated and spontaneous fermentations, respectively). The TDS analyses showed a dominance of bitter and cocoa attributes in both chocolates. However, in the inoculated chocolate, lingering fruity notes were more intense, while the chocolate produced by spontaneous fermentation was more astringent. Thus, the inoculation of yeast influenced the microbial profile, which likely affected the volatile compounds that affect sensory characteristics, resulting in chocolate with dominant bitter, cocoa, and fruity attributes.

Production, reactivation and nutrient requirements of active dried yeast in winemaking: theory and practice

Wine yeast, from production to application, traverse an incredible range of environments. Central to their ability to not only survive but to thrive throughout this journey is their capacity to transform themselves in response to the varied conditions encountered. Wine yeast resilience alone, however, is insufficient. Producers and winemakers must be assiduous in their use of wine yeast with an understanding of yeast requirements throughout the process to achieve optimal results. Active dried yeast (ADY) starter cultures for initiating grape must fermentations are convenient, reliable and available for a wide range of strains suited to different wine types. These benefits cannot be fully realised without optimisation of: their production; reactivation of the dried product prior to inoculation; and appropriate nutrient management during fermentation. This review discusses current knowledge on production of Saccharomyces ADY for greatest biological stability and activity, and factors that affect their activity following rehydration and inoculation into grape must. The efficacy of various types of commercial additives used in reactivation of ADY and, winemaking interventions and nutrients that are used to optimise fermentation are also discussed.

Ongoing domestication of wine yeast: past, present and future

Yeast starter cultures for winemaking have traditionally been developed from successful fermentation isolates. Over the past 10–20 years there has been a move to harness genetic techniques to improve production strains. The approaches to achieve this have been similar to those applied in the agricultural sector for crop and livestock development. Thus, domestication of wine yeasts now involves hybridisation, mutagenesis, selection and screening. There is also a growing interest in tapping into isolates of non-Saccharomyces yeasts to develop novel starter cultures that bring complexity and potentially enhance regionality of wines. This review will cover these developments and will look to the future opportunities arising from the application of genetic engineering and synthetic biology approaches for the introduction of novel phenotypes that are outside of the reach of traditional methods.

Simultaneous inoculation of yeasts and lactic acid bacteria: Effects on fermentation dynamics and chemical composition of Negroamaro wine

Traditional vinification process is undertaken with the inoculation of the lactic acid bacteria (LAB) at the end of alcoholic fermentation (AF) to induce malolactic fermentation (MLF). MLF is an important phase during winemaking and the LAB co-inoculation with yeast starter represents a promising approach to enhance the quality and safety of wine. In this investigation we have studied: i) the effect of timing of LAB inoculation on the vinification dynamics and chemical features of Negroamaro wines; ii) the interactions between two commercial yeast and two commercial Oenococcus oeni strains. The fermentations dynamics were monitored by microbial counts, quantifying l-malic acid concentration and analyzing the volatile compounds contents in the obtained wines. Our results indicate that simultaneous yeasts/bacteria inoculation at the beginning of vinification reduces the processes duration and simultaneously lowers of volatile acidity. Wine obtained after co-inoculum showed a profile dominated by red and ripe fruits notes associated to esters and to buttery and creamy notes linked to diethyl succinate and ethyl lactate. Furthermore, compatibility specification between commercial yeasts and LAB strains were observed, suggesting the importance of the assessment of microbial-compatibility before their utilization in large-scale vinification.

New process for production of fermented black table olives using selected autochthonous microbial resources.

Table olives represent one important fermented product in Europe and, in the world, their demand is constantly increasing. At the present time, no systems are available to control black table olives spontaneous fermentation by the Greek method. During this study, a new protocol for the production of black table olives belonging to two Italian (Cellina di Nardò and Leccino) and two Greek (Kalamàta and Conservolea) cultivars has been developed: for each table olive cultivar, starter-driven fermentations were performed inoculating, firstly, one selected autochthonous yeast starter and, subsequently, one selected autochthonous LAB starter. All starters formulation were able to dominate fermentation process. The olive fermentation was monitored using specific chemical descriptors able to identify a first stage (30 days) mainly characterized by aldehydes; a second period (60 days) mainly characterized by higher alcohols, styrene and terpenes; a third fermentation stage represented by acetate esters, esters and acids. A significant decrease of fermentation time (from 8 to 12 months to a maximum of 3 months) and an significant improvement in organoleptic characteristics of the final product were obtained. This study, for the first time, describes the employment of selected autochthonous microbial resources optimized to mimic the microbial evolution already recorded during spontaneous fermentations.

Genomic expression program of Saccharomyces cerevisiae along a mixed-culture wine fermentation with Hanseniaspora guilliermondii.

BackgroundThe introduction of yeast starter cultures consisting in a blend of Saccharomyces cerevisiae and non-Saccharomyces yeast strains is emerging for production of wines with improved complexity of flavor. The rational use of this approach is, however, dependent on knowing the impact that co-inoculation has in the physiology of S. cerevisiae. In this work the transcriptome of S.cerevisiae was monitored throughout a wine fermentation, carried out in single culture or in a consortium with Hanseniasporaguilliermondii, this being the first time that this relevant yeast–yeast interaction is examined at a genomic scale.ResultsCo-inoculation with H. guilliermondii reduced the overall genome-wide transcriptional response of S. cerevisiae throughout the fermentation, which was attributable to a lower fermentative activity of S. cerevisiae while in the mixed-fermentation. Approximately 350 genes S. cerevisiae genes were found to be differently expressed (FDR < 0.05) in response to the presence of H. guilliermondii in the fermentation medium. Genes involved in biosynthesis of vitamins were enriched among those up-regulated in the mixed-culture fermentation, while genes related with the uptake and biosynthesis of amino acids were enriched among those more expressed in the single-culture. The differences in the aromatic profiles of wines obtained in the single and in the mixed-fermentations correlated with the differential expression of S. cerevisiae genes encoding enzymes required for formation of aroma compounds.ConclusionsBy integrating results obtained in the transcriptomic analysis performed with physiological data our study provided, for the first time, an integrated view into the adaptive responses of S. cerevisiae to the challenging environment of mixed culture fermentation. The availability of nutrients, in particular, of nitrogen and vitamins, stands out as a factor that may determine population dynamics, fermentative activity and by-product formation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0318-1) contains supplementary material, which is available to authorized users.

Breeding Strategy To Generate Robust Yeast Starter Cultures for Cocoa Pulp Fermentations.

Cocoa pulp fermentation is a spontaneous process during which the natural microbiota present at cocoa farms is allowed to ferment the pulp surrounding cocoa beans. Because such spontaneous fermentations are inconsistent and contribute to product variability, there is growing interest in a microbial starter culture that could be used to inoculate cocoa pulp fermentations. Previous studies have revealed that many different fungi are recovered from different batches of spontaneous cocoa pulp fermentations, whereas the variation in the prokaryotic microbiome is much more limited. In this study, therefore, we aimed to develop a suitable yeast starter culture that is able to outcompete wild contaminants and consistently produce high-quality chocolate. Starting from specifically selected Saccharomyces cerevisiae strains, we developed robust hybrids with characteristics that allow them to efficiently ferment cocoa pulp, including improved temperature tolerance and fermentation capacity. We conducted several laboratory and field trials to show that these new hybrids often outperform their parental strains and are able to dominate spontaneous pilot scale fermentations, which results in much more consistent microbial profiles. Moreover, analysis of the resulting chocolate showed that some of the cocoa batches that were fermented with specific starter cultures yielded superior chocolate. Taken together, these results describe the development of robust yeast starter cultures for cocoa pulp fermentations that can contribute to improving the consistency and quality of commercial chocolate production.

Part I: the influence of serial repitching ofSaccharomyces pastorianuson the uptake dynamics of metal ions and fermentable carbohydrates during the fermentation of barley and gluten-free buckwheat and quinoa wort

Recently, research has been focusing on the use of alternative raw materials for brewing purposes and gluten-free beer-like beverages from malted buckwheat and quinoa are of commercial interest. A common commercial process involves the serial repitching of the yeast biomass, but this has not been described using buckwheat and quinoa wort fermentations. Our research studies (Parts I–III) explored the serial repitching of the yeast strain Saccharomyces pastorianus TUM 34/70 on the composition of a barley, buckwheat and quinoa fermentation medium. The present paper focuses on the fermentation performance and the uptake dynamics of metal ions and fermentable carbohydrates. Both pseudocereals showed high variations in all of the attributes examined during successive fermentations. In buckwheat the differences between successive fermentations were similar to those observed with barley, whereas differences in quinoa varied quite significantly from those observed with barley and showed a directional trend, suggesting a general weakening of the yeast from the sixth successive fermentation onward. In particular, the assimilation of the fermentable carbohydrates lessened and metal ion uptake appeared poorly controlled. It was concluded that buckwheat showed good potential for serial repitching of S. pastorianus TUM 34/70, whereas serial repitching of a quinoa wort appeared to be limited to five or six fermentations. Copyright © 2015 The Institute of Brewing & Distilling

Effects of indigenous yeasts on physicochemical and microbial properties of Korean soy sauce prepared by low-salt fermentation

This study deals with understanding the effects of salt reduction on both the physicochemical and microbiological properties of soy sauce fermentation and also the application of indigenous yeast starters to compensate for undesirable changes occurring in salt-reduced processes. Fermentation was tested in situ at a Korean commercial soy sauce processing unit. Salt reduction resulted in higher acidity as well as lower pH and contents of residual sugar and ethanol. Moreover, undesired flavor characteristics, due to a lack of distinctive compounds, was observed. In addition, putrefactive Staphylococcus and Enterococcus spp. were present only during salt-reduced fermentation. To control these adverse effects, a single or mixed culture of two indigenous yeasts, Torulaspora delbrueckii and Pichia guilliermondii, producing high ethanol and 3-methyl-1-butanol, respectively, were tested. Overall, all types of yeast applications inhibited undesirable bacterial growth despite salt reduction. Of the starter cultures tested, the mixed culture resulted in a balance of more complex and richer flavors with an identical flavor profile pattern to that obtained from high salt soy sauce. Hence, this strategy using functional yeast cultures offers a technological option to manufacture salt-reduced soy sauce while preserving its typical sensory characteristics without affecting safety.