Indigenous Saccharomyces cerevisiae Could Better Adapt to the Physicochemical Conditions and Natural Microbial Ecology of Prince Grape Must Compared with Commercial Saccharomyces cerevisiae FX10

Indigenous Saccharomyces cerevisiae, as a new and useful tool, can be used in fermentation to enhance the aroma characteristic qualities of the wine-production region. In this study, we used indigenous S. cerevisiae L59 and commercial S. cerevisiae FX10 to ferment Prince (a new hybrid variety from Lion Winery) wine, detected the basic physicochemical parameters and the dynamic changes of fungal communities during fermentation, and analyzed the correlations between fungal communities and volatile compounds. The results showed that the indigenous S. cerevisiae L59 could quickly adapt to the specific physicochemical conditions and microbial ecology of the grape must, showing a strong potential for winemaking. Compared with commercial S. cerevisiae FX10, the wine fermented by indigenous S. cerevisiae L59 contained more glycerol and less organic acids, contributing to a rounder taste. The results of volatile compounds indicated that the indigenous S. cerevisiae L59 had a positive effect on adding rosy, honey, pineapple and other sweet aroma characteristics to the wine. Overall, the study we performed showed that selection of indigenous S. cerevisiae from the wine-producing region as a starter for wine fermentation is conducive to improving the aroma profile of wine and preserving the aroma of the grape variety.     

Effect of Hanseniaspora uvarum–Saccharomyces cerevisiae Mixed Fermentation on Aroma Characteristics of Rosa roxburghii Tratt,Blueberry, and Plum Wines

Hanseniaspora uvarum, a non-Saccharomyces cerevisiae species, has a crucial effect on the aroma characteristics of fruit wines, thus, attracting significant research interest in recent years. In this study, H. uvarum–Saccharomyces cerevisiae mixed fermentation was used to ferment Rosa roxburghii Tratt, blueberry fruit wine, and plum fruit wines using either a co-inoculated or a sequentially inoculated approach. The three fruit wines’ volatile aroma characteristics were analyzed by headspace–solid-phase microextraction–gas chromatography-mass spectrometry (HS–SPME–GC–MS). The results showed that the mixed inoculation of H. uvarum and S. cerevisiae reduced the alcoholic content of Kongxinli fruit wine. Moreover, H. uvarum–S. cerevisiae fermented Rosa roxburghii Tratt, blueberry, and plum fruit wines and further enriched their flavor compounds. The overall flavor characteristics of sequentially inoculated fruit wines differed significantly from those fermented with S. cerevisiae alone, although several similarities were also observed. Sequential inoculation of H. uvarum and S. cerevisiae positively affected the mellowness of the wine and achieved a better harmony of the overall wine flavors. Therefore, H. uvarum–Saccharomyces cerevisiae mixed fermentation can improve the complexity of the wines’ aromatic composition and empower them with a unique identity. In particular, H. uvarum–Saccharomyces cerevisiae blueberry wine produced by mixed fermentation had the widest variety and content of aroma compounds among the fermented wines. Therefore, H. uvarum–Saccharomyces cerevisiae mixed-fermentation inoculation in the three fermented fruit wines significantly increased the aroma compound variety and content, thus, enriching their aroma richness and complexity. This study is the first comparative evaluation of the aroma characteristics of different fruit wines fermented with a mixed inoculation of H. uvarum and S. cerevisiae and provides a preliminary guide for these fruit wines produced with non-Saccharomyces yeast.     

Adaptive Evolution of Saccharomyces cerevisiae with Enhanced Ethanol Tolerance for Chinese Rice Wine Fermentation

High tolerance towards ethanol is a desirable property for the Saccharomyces cerevisiae strains used in the alcoholic beverage industry. To improve the ethanol tolerance of an industrial Chinese rice wine yeast, a sequential batch fermentation strategy was used to adaptively evolve a chemically mutagenized Chinese rice wine G85 strain. The high level of ethanol produced under Chinese rice wine-like fermentation conditions was used as the selective pressure. After adaptive evolution of approximately 200 generations, mutant G85X-8 was isolated and shown to have markedly increased ethanol tolerance. The evolved strain also showed higher osmotic and temperature tolerances than the parental strain. Laboratory Chinese rice wine fermentation showed that the evolved G85X-8 strain was able to catabolize sugars more completely than the parental G85 strain. A higher level of yeast cell activity was found in the fermentation mash produced by the evolved strain, but the aroma profiles were similar between the evolved and parental strains. The improved ethanol tolerance in the evolved strain might be ascribed to the altered fatty acids composition of the cell membrane and higher intracellular trehalose concentrations. These results suggest that adaptive evolution is an efficient approach for the non-recombinant modification of industrial yeast strains.     

Color Stabilization of Apulian Red Wines through the Sequential Inoculation of Starmerella bacillaris and Saccharomyces cerevisiae

Mixed fermentation using Starmerella bacillaris and Saccharomyces cerevisiae has gained attention in recent years due to their ability to modulate the qualitative parameters of enological interest, such as the color intensity and stability of wine. In this study, three of the most important red Apulian varieties were fermented through two pure inoculations of Saccharomyces cerevisiae strains or the sequential inoculation of Saccharomyces cerevisiae after 48 h from Starmerella bacillaris. The evolution of anthocyanin profiles and chromatic characteristics were determined in the produced wines at draining off and after 18 months of bottle aging in order to assess the impact of the different fermentation protocols on the potential color stabilization and shelf-life. The chemical composition analysis showed titratable acidity and ethanol content exhibiting marked differences among wines after fermentation and aging. The 48 h inoculation delay produced wines with higher values of color intensity and color stability. This was ascribed to the increased presence of compounds, such as stable A-type vitisins and reddish/violet ethylidene-bridge flavonol-anthocyanin adducts, in the mixed fermentation. Our results proved that the sequential fermentation of Starmerella bacillaris and Saccharomyces cerevisiae could enhance the chromatic profile as well as the stability of the red wines, thus improving their organoleptic quality.     

Dry Red Wine Making Using Yeast Immobilized on Cork Pieces

The commercial Saccharomyces cerevisiae Uvaferme 299 wine yeast was immobilized on cork pieces to produce a biocatalyst for use in dry red wine making. The immobilized biocatalyst was suitable for clarified and non-clarified grape must fermentation at ambient and low temperatures (0–25 °C). Fermentation times were low and very low amounts of residual sugar were detected, showing suitability for dry wine production. The presence of suspended solids in the non-clarified must did not affect the activity of the immobilized cells. Complete fermentation of sugars at 0 °C was possible with immobilized Uvaferme 299, although not a cryotolerant strain, indicating a cryoprotective effect of cork. The presence of cork did not affect the composition of major volatiles with methanol and acetaldehyde kept at low levels. Reduction of amyl alcohols on total volatiles was also observed in wines fermented at low temperatures. Differences in the headspace aroma profile in wines produced by immobilized and free cells were found by GC–MS analysis, but no cork taint compounds were detected.     

Influence of Triazole Pesticides on Wine Flavor and Quality Based on Multidimensional Analysis Technology

Triazole pesticides are widely used to control grapevine diseases. In this study, we investigated the impact of three triazole pesticides—triadimefon, tebuconazole, and paclobutrazol—on the concentrations of wine aroma compounds. All three triazole pesticides significantly affected the ester and acid aroma components. Among them, paclobutrazol exhibited the greatest negative influence on the wine aroma quality through its effect on the ester and acid aroma substances, followed by tebuconazole and triadimefon. Qualitative and quantitative analysis by solid-phase micro-extraction gas chromatography coupled with mass spectrometry revealed that the triazole pesticides also changed the flower and fruit flavor component contents of the wines. This was attributed to changes in the yeast fermentation activity caused by the pesticide residues. The study reveals that triazole pesticides negatively impact on the volatile composition of wines with a potential undesirable effect on wine quality, underlining the desirability of stricter control by the food industry over pesticide residues in winemaking.     

Production of ethanol from waste paper using immobilized yeasts

This work is aimed at a selection of yeast strains suitable for simultaneous saccharification and fermentation of waste paper. The waste paper, as a lignocellulosic material, represents an unconventional source for the production of ethanol which is a promising alternative fuel. The yeast strains Saccharomyces cerevisiae and Pichia kudriavzevii produced the highest amounts of ethanol at 30 °C and were also resistant at 40 °C during the first 92 h of fermentation. These two strains were immobilized by entrapment into poly(vinyl alcohol) hydrogel lens-shaped particles LentiKats^®. The immobilized S. cerevisiae was a better ethanol producer and retained higher metabolic activity in repeated batch fermentations than P. kudriavzevii. The immobilized S. cerevisiae was also suitable for a long-term storage, with 23% decrease in the ethanol production ability after 1-year storage of yeast cells.     

Monitoring of yeast population isolated during spontaneous fermentation of Moravian wine

In enology, yeasts play an important role in the characteristics of the final product. They are predominant in the biochemical interaction with components of must. Rapid identification of the yeast population is necessary for fermentation process monitoring and for obtaining a good quality wine. The main goal of this study was the isolation and characterisation of the yeast microbial community naturally present on grape berries, leaves and occurring during the spontaneous fermentation process of the white wine Veltlin green from the South Moravian region, Czech Republic. The results, based on PCR-RFLP of the 5.8S-ITS region of rDNA, PCR-fingerprinting using microsatellite oligonucleotide primers (GAG)₅, (GTG)₅, (GAC)₅, and M13 primer, showed great diversity of the yeast population. Including grape berries and fermented must, the following yeast species were identified: Hanseniaspora uvarum, Aureobasidium pullulans, Metschnikowia pulcherrima, Torulaspora delbrueckii, a number of Pichia species such as P. fermentans, P. membranifaciens, P. kluyveri, also Sporidiobolus salmonicolor, Rhodosporidium toruloides, Rhodotorula mucilaginosa, Rhodotorula glutinis as well as Saccharomyces cerevisiae and Saccharomyces bayanus. Monitoring of the yeast strains during the wine fermentation process of traditional Moravian wine can contribute to the improvement of wine quality.     

Enhanced biotransformation of furfural and hydroxymethylfurfural by newly developed ethanologenic yeast strains

Furfural and hydroxymethylfurfural (HMF) are representative inhibitors among many inhibitive compounds derived from biomass degradation and saccharification for bioethanol fermentation. Most yeasts, including industrial strains, are susceptible to these inhibitory compounds, especially when multiple inhibitors are present. Additional detoxification steps add cost and complexity to the process and generate additional waste products. To promote efficient bioethanol production, we studied the mechanisms of stress tolerance, particularly to fermentation inhibitors such as furfural and HMF. We recently reported a metabolite of 2,5-bis-hydroxymethylfuran as a conversion product of HMF and characterized a dose-dependent response of ethanologenic yeasts to inhibitors. In this study, we present newly adapted strains that demonstrated higher levels of tolerance to furfural and HMF. Saccharomyces cerevisiae 307-12H60 and 307-12H120 and Pichia stipitis 307 10H60 showed enhanced biotransformation ability to reduce HMF to 2,5-bis-hydroxymethylfuran at 30 and 60 mM, and S. cerevisiae 307-12-F40 converted furfural into furfuryl alcohol at significantly higher rates compared to the parental strains. Strains of S. cerevisiae converted 100% of HMF at 60 mM and S. cerevisiae 307-12-F40 converted 100% of furfural into furfuryl alcohol at 30 mM. The results of this study suggest a possible in situ detoxification of the inhibitors by using more inhibitor-tolerant yeast strains for bioethanol fermentation. The development of such tolerant strains provided a basis and useful materials for further studies on the mechanisms of stress tolerance.     

Characterization of yeast strains for wine production

Sixteen yeast strains isolated from grapefruit (Citrus paradis), orange (Citrus sinensis) and pineapple (Ananas comosus) were characterized using standard microbiological procedures. The species were identified as Saccharomyces uvarum, S. cerevisiae, S. carlbergensis, and S. ellipsoideus. Their abilities for wine production were tested by using sugar and ethanol tolerance tests. The best biochemically active strain, S. ellipsoideus, was used along with commercially available baker's yeast (S. cerevisiae) to produce wine from grapefruit, orange, and pineapple juices. After fermentation for 14 d with S. cerevisiae and 21 d with S. ellipsoideus, wines produced were compared with Baron de Valls (standard). The highest (10.47% [v/v]) and lowest (7.68% [v/v]) alcohol concentrations with corresponding residual sugar concentrations of 1.88% (w/v) and 7.7% (w/v) were produced from orange after fermentation with S. cerevisiae and S. ellipsoideus, respectively. S. ellipsoideus was found to be the best yeast strain producing wine with the highest acceptable score of 7.41 from orange. The study revealed the possibility of producing wine from our locally available fruits using simple, cheap, and adaptable technology with biochemically characterized yeast strains.     

Relationship between sensory and instrumental analysis of 2,4,6-trichloroanisole in wine and cork stoppers

The effect of different corks stoppers on the sensory and instrumental determination of 2,4,6-trichloroanisole (TCA) was studied in wine and cork. A relationship between both measurements was also established.Four types of cork were used to seal white wine bottles for 8 months. The stoppers were from different raw material: a high quality commercial batch (C), slabs with yellow stain (YS) and slabs with a high musty and mouldy taint (T). Spiked samples (S) were prepared from C batch by injecting 1002 ng of TCA into the cork stoppers.TCA was determined by gas chromatography with electron capture detection after headspace solid phase microextraction in bottled wine (12 per group) and after extraction with pentane in the case of corks (six per group). Seven semi-trained assessors evaluated the different samples using a ranking test in 12 and six sessions for wine and cork stoppers, respectively.Wines and corks from S samples showed the highest TCA values in both sensory and instrumental measurements, the lowest values being for C samples. YS and T corks had intermediate values, although in general TCA concentration was higher in T. A slight tendency to increase the TCA content in stoppers with yellow stain compared to C samples was observed in wine. A high correlation coefficient (r=0.82) was found between sensory and instrumental analysis for wine, whereas this coefficient was much lower (r=0.56) for cork stoppers. Some hypotheses are given in order to explain these differences.     

Effect of Bentonite Addition to Pedro Ximénez White Grape Musts before Their Fermentation with Selected Yeasts on the Major Volatile Compounds and Polyols of Wines and Tentative Relationships with the Sensorial Evaluation

In this work, we study the effect of bentonite addition to the grape must before alcoholic fermentation on the chemical composition and sensorial profile of the obtained wines. Fermentations were carried out with two Saccharomyces cerevisiae commercial active dry yeasts treated or not with bentonite and were compared with a control wine obtained by spontaneous fermentation (using the grape must microbiota). Several significant effects on the chemical and sensorial attributes were established by statistical treatments. The selection by multiple variable analysis of seven volatile molecules (ethyl acetate; methanol; 1-propanol; isobutanol; 2-methyl-1-butanol; 3-metyl-1-butanol and 2-phenylethanol) provided several footprints that provide an easy visualization of bentonite effects on wine volatile compounds. A Principal Component Analysis carried out with all the compounds quantified by Gas-Chromatography revealed that the first two Principal Components explain 60.15 and 25.91%, respectively, of the total variance and established five groups that match with the five wines analyzed. Lastly, predictive models at p ≤ 0.05 level for the attributes sight, smell and taste were obtained by Partial Least Squared regression analysis of selected chemical variables.     

Glycosidically-Bound Volatile Phenols Linked to Smoke Taint: Stability during Fermentation with Different Yeasts and in Finished Wine

When wine grapes are exposed to smoke, there is a risk that the resulting wines may possess smoky, ashy, or burnt aromas, a wine flaw known as smoke taint. Smoke taint occurs when the volatile phenols (VPs) largely responsible for the aroma of smoke are transformed in grape into a range of glycosides that are imperceptible by smell. The majority of VP-glycosides described to date are disaccharides possessing a reducing β-d-glucopyranosyl moiety. Here, a two-part experiment was performed to (1) assess the stability of 11 synthesized VP-glycosides towards general acid-catalyzed hydrolysis during aging, and (2) to examine whether yeast strains differed in their capacity to produce free VPs both from these model glycosides as well as from grapes that had been deliberately exposed to smoke. When fortified into both model and real wine matrices at 200 ng/g, all VP-disaccharides were stable over 12 weeks, while (42–50 ng/g) increases in free 4-ethylphenol and p-cresol were detected when these were added to wine as their monoglucosides. Guaiacol and phenol were the most abundantly produced VPs during fermentation, whether originating from natural VP-precursors in smoked-exposed Pinot Noir must, or due to fortification with synthetic VP-glycosides. Significant yeast strain-specific differences in glycolytic activities were observed for phenyl-β-d-glycopyranoside, with two strains (RC212 and BM45) being unable to hydrolyze this model VP, albeit both were active on the guaiacyl analogue. Thus, differences in Saccharomyces cerevisiae β-glucosidase activity appear to be influenced by the VP moiety.     

A Robust Fermentation Process for Natural Chocolate-like Flavor Production with Mycetinis scorodonius

Submerged fermentation of green tea with the basidiomycete Mycetinis scorodonius resulted in a pleasant chocolate-like and malty aroma, which could be a promising chocolate flavor alternative to current synthetic aroma mixtures in demand of consumer preferences towards healthy natural and ‘clean label’ ingredients. To understand the sensorial molecular base on the chocolate-like aroma formation, key aroma compounds of the fermented green tea were elucidated using a direct immersion stir bar sorptive extraction combined with gas chromatography–mass spectrometry–olfactometry (DI-SBSE-GC-MS-O) followed by semi-quantification with internal standard. Fifteen key aroma compounds were determined, the most important of which were dihydroactinidiolide (odor activity value OAV 345), isovaleraldehyde (OAV 79), and coumarin (OAV 24), which were also confirmed by a recombination study. Furthermore, effects of the fermentation parameters (medium volume, light protection, agitation rate, pH, temperature, and aeration) on the aroma profile were investigated in a lab-scale bioreactor at batch fermentation. Variation of the fermentation parameters resulted in similar sensory perception of the broth, where up-scaling in volume evoked longer growth cycles and aeration significantly boosted the concentrations yet added a green note to the overall flavor impression. All findings prove the robustness of the established fermentation process with M. scorodonius for natural chocolate-like flavor production.     

Aromatic components produced by non-Saccharomyces Cerevisiae derived from natural fermentation of grape

Aromatic components are important functional products during the wine fermentation process. In the current study, nine strains (Y10, Y5, Y21, Y2, Y19, Y16, Y3, Y13 and Y4) of non-Saccharomyces were isolated from Cabernet Sauvignon grape wine. Aromatic components from Cabernet sauvignon-fermented wine were determined the phylogenetic evolution status of different non-Saccharomyces based on 26S rDNA and D1/D2 sequence analysis and analysed by a gas chromatography–mass spectrometry analysis, and they were grouped into one category with four different yeast genus which were Meyerozyma guilliermondii, Brettanomyces naardenensis, Pichia guilliermondi and Candida fermentati. A total of 102 kinds of aroma components were detected, including 39 kinds of esters, 31 kinds of alcohols, 8 kinds of ketones, 10 kinds of alkanes, 15 kinds of acids and 4 kinds of other aroma substances.     

Butanol Tolerance in a Selection of Microorganisms

Butanol tolerance is a critical factor affecting the ability of microorganisms to generate economically viable quantities of butanol. Current Clostridium strains are unable to tolerate greater than 2% 1-butanol thus membrane or gas stripping technologies to actively remove butanol during fermentation are advantageous. To evaluate the potential of alternative hosts for butanol production, we screened 24 different microorganisms for their tolerance to butanol. We found that in general, a barrier to growth exists between 1% and 2% butanol and few microorganisms can tolerate 2% butanol. Strains of Escherichia coli, Zymomonas mobilis, and non-Saccharomyces yeasts were unable to surmount the 2% butanol growth barrier. Several strains of Saccharomyces cerevisiae exhibit limited growth in 2% butanol, while two strains of Lactobacillus were able to tolerate and grow in up to 3% butanol.     

Characteristics of Bacterial Strains with Desirable Flavor Compounds from Korean Traditional Fermented Soybean Paste (Doenjang)

To identify and analyze the characteristics of the microorganisms involved in the formation of the desirable flavor of Doenjang, a total of 179 strains were isolated from ninety-four Doenjang collected from six regions in South Korea, and fourteen strains were selected through a sensory evaluation of the aroma of each culture. The enzyme activities of amylase, protease and lipase was shown in the various strains. Bacillus sp.-K3, Bacillus sp.-K4 and Bacillus amyloliquefaciens-J2 showed relatively high protease activity, at 317.1 U, 317.3 U and 319.5 U, respectively. The Bacillus sp.-K1 showed the highest lipase activity at 2453.6 U. In the case of amylase, Bacillus subtilis-H6 showed the highest activity at 4105.5 U. The results of the PCA showed that Bacillus subtilis-H2, Bacillus subtilis-H3, and Bacillus sp.-K2 were closely related to the production of 3-hydroxy-2-butanone (23.51%~43.37%), and that Bacillus subtilis-H5 and Bacillus amyloliquefaciens-J2 were significantly associated with the production of phenethyl alcohol (0.39% and 0.37%). The production of peptides was observed to vary among the Bacillus cultures such as Val-Val-Pro-Pro-Phe-Leu and Pro-Ala-Glu-Val-Leu-Asp-Ile. These peptides are precursors of related volatile flavor compounds created in Doenjang via the enzymatic or non-enzymatic route; it is expected that these strains could be used to enhance the flavor of Doenjang.     

Improvements of Tolerance to Stress Conditions by Genetic Engineering in Saccharomyces Cerevisiae during Ethanol Production

Saccharomyces cerevisiae, industrial yeast isolate, has been of great interest in recent years for fuel ethanol production. The ethanol yield and productivity depend on many inhibitory factors during the fermentation process such as temperature, ethanol, compounds released as the result of pretreatment procedures, and osmotic stress. An ideal strain should be able to grow under different stress conditions occurred at different fermentation steps. Development of tolerant yeast strains can be achieved by reprogramming pathways supporting the ethanol metabolism by regulating the energy balance and detoxicification processes. Complex gene interactions should be solved for an in-depth comprehension of the yeast stress tolerance mechanism. Genetic engineering as a powerful biotechnological tool is required to design new strategies for increasing the ethanol fermentation performance. Upregulation of stress tolerance genes by recombinant DNA technology can be a useful approach to overcome inhibitory situations. This review presents the application of several genetic engineering strategies to increase ethanol yield under different stress conditions including inhibitor tolerance, ethanol tolerance, thermotolerance, and osmotolerance.     

Bioethanol Production from Soybean Residue via Separate Hydrolysis and Fermentation

Bioethanol was produced using polysaccharide from soybean residue as biomass by separate hydrolysis and fermentation (SHF). This study focused on pretreatment, enzyme saccharification, and fermentation. Pretreatment to obtain monosaccharide was carried out with 20% (w/v) soybean residue slurry and 270 mmol/L H₂SO₄ at 121 °C for 60 min. More monosaccharide was obtained from enzymatic hydrolysis with a 16 U/mL mixture of commercial enzymes C-Tec 2 and Viscozyme L at 45 °C for 48 h. Ethanol fermentation with 20% (w/v) soybean residue hydrolysate was performed using wild-type and Saccharomyces cerevisiae KCCM 1129 adapted to high concentrations of galactose, using a flask and 5-L fermenter. When the wild type of S. cerevisiae was used, an ethanol production of 20.8 g/L with an ethanol yield of 0.31 g/g consumed glucose was obtained. Ethanol productions of 33.9 and 31.6 g/L with ethanol yield of 0.49 g/g consumed glucose and 0.47 g/g consumed glucose were obtained in a flask and a 5-L fermenter, respectively, using S. cerevisiae adapted to a high concentration of galactose. Therefore, adapted S. cerevisiae to galactose could enhance the overall ethanol fermentation yields compared to the wild-type one.