Yeast Vitality Research Articles

Optimizing the production of wheat beer by recycling the yeast biomass: perspective for cost reduction with quality maintenance

A considerable investment is being experienced worldwide in the production of artisanal beers of different styles, such as wheat-based Weizen beer. The challenge is to reduce production costs while maintaining the commercially accepted quality. Thus, the improvement of the production process involves the possibility of shortening the fermentation time by reusing the yeast biomass without affecting the yeast vitality and the quality of the end product. In this study, the fermentation parameters of a Weizen beer were measured directly from a scaled-up process in a microbrewing in order to identify the optimum point at which the process might be ended without compromising the final characteristics. The results were confirmed by laboratory studies that compared three different yeast strains in a wheat-based industrial substrate. The results led to the conclusion that the fermentation time was reduced by more than 50% from 7 to 3 days, with less variation in the concentration of the investigated metabolites. By reducing the time of fermentation, the yeast biomass was maintained active to be used up to six consecutive cycles of fermentation. All these improvements might help in reducing the cost of beer processing, which is of paramount interest to microbreweries worldwide.

Differences in yeast behaviour during ageing of sparkling wines made with Charmat and Traditional methods

In this study, we followed the yeast and wine behaviour during the second fermentation and subsequent lees ageing of sparkling wines produced by Traditional and Charmat methods at an industrial scale. During this period, we conducted physicochemical, microbiological, gene expression, and marker analyses of characteristics related to wine ageing. Our results show that the yeast behaviour during the fermentation is similar in both methods. However, after fermentation, there is a faster decrease in yeast vitality and viability in the Charmat method, together with an increase in the expression of autophagy-related genes (AMS1, APE1, and ATG8). We relate these factors to ageing with the continuous homogenization of the liquid practised in the Charmat method, and static ageing with the lees concentrated at the bottom of the bottle performed in the Traditional method. Despite the variation in yeast viability during ageing, there are no differences in soluble proteins, free amino nitrogen, total phenols, antioxidant activity, and colour evolution between the wines produced by the two methods, assuming few differences over the time that monitoring was conducted.

Provocation: all yeast cells are born equal, but some grow to be more equal than others

Yeast is central to the brewing process and as such deserves great care and attention to ensure it is fit-for-purpose. It is widely accepted that assessment based on simple viability testing is useful but far too blunt an instrument to give much useful information regarding the physiological state of the live yeast cells within the population. The need to address this issue has spawned a plethora of procedures collectively termed ‘vitality tests’. The concept of yeast vitality is nebulous, and this goes some way to explain why no single test has been universally adopted. There is, however, another major flaw in the underlying assumptions of the majority of vitality tests. This is, most are based on bulk samples which ignore heterogeneities within the population of cells being examined. The asymmetric mode of proliferation of Saccharomyces yeasts means that mother and daughter cells are always unequal. Individual cells have finite lifespans, old cells die and must be replaced with new fitter progeny. We are fortunate in that budding yeasts are widely used as a model cell for studies into ageing in higher eukaryotes and as a result there is a huge and ever-expanding body of literature devoted to this subject. This has shown that yeast populations comprise several distinct sub-populations which respond differently to external messengers. These messengers may be generated by one sub-set of the population and received and acted upon by another indicating a degree of cooperation designed to ensure survival of the whole. Death of individuals within populations can be as a result of the application of lethal external stresses but more usually is a programmed process in which targeted cells commit suicide in an expression of altruism for the greater good of the whole. On reflection, perhaps this is unexpected but perfectly logical behaviour for single celled organisms to adopt. Most yeast population studies have been performed using haploid laboratory strains cultivated under relatively defined conditions. The industrial scale model is, of course, far removed from this. Brewing yeast strains are very different, and the complexities of serial fermentations and associated yeast handling processes introduce added layers of complexity. Nevertheless, there is every reason to believe that similar population heterogeneities and cooperative behaviour exist within the context of brewing. The thrust of the arguments presented in this article is that we should embrace this and consider individual cells within yeast populations within a fermenter, for example, as being individual free-living components of a multicellular organism. Further, intercellular cooperation is managed by a network of messenger molecules many of which are likely to trigger the global shifts associated in metabolic activities which accompany the reversible passage from growth to quiescence and which coincidentally are likely to be of importance in beer flavour. The purpose of this article is to argue that the majority of current yeast vitality tests are not fit for their intended purpose. They do not provide outputs that are predictive of fermentation performance or an indication of the metabolic status of the population as a whole. Simple viability tests based on differential responses to biological stains, including methylene blue, are of equal value in commercial brewing. This situation can be improved by the introduction of tests which give information of how sub-populations develop and interact with each other. Probably dual staining techniques which probe membrane competence and detect cells in the early stages of programmed cell death, as discussed in this article, are promising candidates. © 2021 The Institute of Brewing & Distilling

Basal catalase activity and high glutathione levels influence the performance of non-Saccharomyces active dry wine yeasts

Non-Saccharomyces wine yeasts are useful tools for producing wines with complex aromas or low ethanol content. Their use in wine would benefit from their production as active dry yeast (ADY) starters to be used as co-inocula alongside S. cerevisiae. Oxidative stress during biomass propagation and dehydration is a key factor in determining ADY performance, as it affects yeast vitality and viability. Several studies have analysed the response of S. cerevisiae to oxidative stress under dehydration conditions, but not so many deal with non-conventional yeasts. In this work, we analysed eight non-Saccharomyces wine yeasts under biomass production conditions and studied oxidative stress parameters and lipid composition. The results revealed wide variability among species in their technological performance during ADY production. Also, for Metschnikowia pulcherrima and Starmerella bacillaris, better performance correlates with high catalase activity and glutathione levels. Our data suggest that non-Saccharomyces wine yeasts with an enhanced oxidative stress response are better suited to grow under ADY production conditions.

Optimisation of yeast vitality measurement to better predict fermentation performance

Brewers are increasingly aware of the importance of yeast health and need appropriate methods to measure its vitality. Many of the current methods are time consuming, expensive or impractical at an industrial scale. Here, the measurement of CO2 production using the ANKOM RF Gas Production Measurement System was optimised. Using response surface methodology, the optimal temperature, yeast biomass and media composition were established. The performance of the method was assessed by measuring the vitality of yeast from laboratory (20 L) and brewery (400-600 hL) fermentations. The drop in pH in early fermentation, diacetyl reduction and fermentation rate were correlated to measurement of yeast vitality. It was shown that the method can be used to assess yeast health and to predict its performance in brewery fermentation. The measurement time was 20 minutes and a minimum vitality of approximately 200 mbar was found to be sufficient for brewery fermentations. The method can be used to optimise yeast propagation processes or to determine the optimal harvesting/cropping time to improve yeast quality, fermentation consistency and beer quality together with reducing cycle time and time in fermenter. © 2020 The Institute of Brewing & Distilling

Influence of Glutathione on Yeast Fermentation Efficiency under Copper Stress.

Copper in grape musts can influence the fermentation efficiency of Saccharomyces cerevisiae during winemaking. The present study revealed the impact of glutathione addition on yeast strains with variable copper sensitivity. The antioxidant glutathione increased yeast vitality and fastened sugar metabolism at copper concentrations up to 0.39 mM. A significant accumulation of acetaldehyde at high copper concentrations was mitigated by the addition of 20 mg L-1 glutathione. Low recovery of glutathione added implicated a complexation of both compounds. Specific alcohol dehydrogenase (ADH) activity was inhibited or reduced in the enzyme extracts of the copper-stressed yeast cells. The activity was restored in fermentations with glutathione at a copper concentration of 0.16 mM. At low copper concentrations, glutathione decreased ADH activity presumably due to complexation of essential copper amounts. Results provide important information on the use of glutathione as an antioxidant in winemaking to counteract negative effects of copper-rich musts on copper-sensitive yeast strains.

Blending studies using wheat and lentil cotyledon flour—Effects on rheology and bread quality

AbstractBackground and objectivesLentil (Lens Culinaris. Medik) is a highly nutritious food staple widely consumed within India subcontinent and the Mediterranean region. Although gaining popularity in western diets, wheat will continue to be a major crop as it can be used to manufacture a wide range of products. The nutritional benefits of lentils are acknowledged, particularly as a source of high protein so the incorporation of lentil flour into wheat‐based foods has the potential to improve the nutritive value of range food products. Twelve blended flours were made using different concentrations of red lentil cotyledon, wheat, and additional gluten. A blending study was undertaken to access yeast vitality, rheological properties of dough and baking characteristic of resulting bread.FindingsHigh ratio blends of lentil flour had no negative effect on yeast vitality even at the highest concentration of 40%. Increasing substitution of lentil flour was highly correlated to increased protein (r = 0.98) and ash (r = 0.95) and a concomitant decrease in dough strength but not extensibility. Loaf volume and baking quality were also compromised at higher concentrations. At a concentration of 5% lentil flour, there were no deleterious effects on dough quality traits or on baking quality. The addition of bakers’ gluten 0.1 g/gram flour had a restorative effect on the rheological and baking characteristics of wheat–lentil composites at higher concentrations of up to 20%.ConclusionsOur results show that optimal baking quality of wheat–lentil flour can be achieved using either low concentrations of up to 5% lentil flour or up to 20% lentil flour with the addition of gluten which maintained a superior loaf and crumb quality.Significance and noveltyThe protein and ash content of baked breads significantly increased when wheat was partially substituted with lentil flour. The concentration of lentil flour decreased dough strength and dough development time and decreased loaf volume while increasing crumb firmness in resulting bread. The addition of gluten improved the rheological and product quality of bread which allowed higher concentrations of lentil flour to be used in bread making. Balancing the ratio of lentil flour and gluten to optimize the rheological properties will result in a composite wheat–lentil bread with acceptable baking performance and enhanced nutritional benefits for consumers.

Resveratrol suppresses ethanol stress in winery and bottom brewery yeast by affecting superoxide dismutase, lipid peroxidation and fatty acid profile.

Mid-exponential cultures of two traditional biotechnological yeast species, winery Saccharomyces cerevisiae and the less ethanol tolerant bottom-fermenting brewery Saccharomyces pastorianus, were exposed to different concentrations of added ethanol (3, 5 and 8%) The degree of ethanol-induced cell stress was assessed by measuring the cellular activity of superoxide dismutase (SOD), level of lipid peroxidation products, changes in cell lipid content and fatty acid profile. The resveratrol as an antioxidant was found to decrease the ethanol-induced rise of SOD activity and suppress the ethanol-induced decrease in cell lipids. A lower resveratrol concentration (0.5mg/l) even reduced the extent of lipid peroxidation in cells. Resveratrol also alleviated ethanol-induced changes in cell lipid composition in both species by strongly enhancing the proportion of saturated fatty acids and contributing thereby to membrane stabilization. Lower resveratrol concentrations could thus diminish the negative effects of ethanol stress on yeast cells and improve their physiological state. These effects may be utilized to enhance yeast vitality in high-ethanol-producing fermentations or to increase the number of yeast generations in brewery.

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

The effect of pitching rate on fermentation, maturation and flavour compounds of beer produced on an industrial scale

The aim of the study was to determine the effect of the initial number of yeast cells in the wort on the process of fermentation, maturation and the content of the volatile components of beer, as well as the viability and vitality of the yeast biomass. The experiments were performed on an industrial scale, with fermentation and maturation in cylindro-conical fermentation tanks with a capacity of 3800 hL. Yeast for pitching was collected after secondary fermentation (third passage) and wort pitching levels were 5 × 106, 7 × 106 and 9 × 106 cells/mL. During fermentation and maturation, the changes in the content of the extract, yeast growth, yeast vitality and selected volatile components were investigated. Experiments showed that the yeast inoculum had a significant impact on the course of the fermentation and metabolic changes. With increasing numbers of cells introduced into the wort, the content of the esters and fusel alcohols increased, while the acetaldehyde concentration decreased. These changes affected the final quality of the beer. Copyright © 2015 The Institute of Brewing & Distilling

Image-Based Cytometric Analysis of Fluorescent Viability and Vitality Staining Methods for Ale and Lager Fermentation Yeast

Saccharomyces cerevisiae have been an essential component of beer production for centuries. The viability and vitality of yeast during a fermentation brewing process is an especially important consideration for proper cell growth, consistent flavor, and optimal production yield. The current definition of viability refers to yeast with intact membranes, while vitality refers to yeast with quantifiable metabolic activity and an ability to proliferate. Yeast may be viable, but may not be actively proliferating, which can affect the fermentation process. The traditional method for measuring yeast viability utilizes manual counting of methylene blue stained yeast cells in a hemacytometer. However, this method can be time consuming and has user-dependent variations. In this work, we demonstrate the capability of Cellometer Vision image cytometry for multi-fluorescent yeast viability and vitality measurements. Nine fluorescent stains were tested with this image cytometry system, including nucleic acid stains (PI, EB, 7-AAD, and DAPI), membrane potential, intracellular, and enzymatic stains (oxonol, MgANS, and CFDA-AM), as well as dual-fluorescent stains (AO/PI and CFDA-AM/PI). Each combination was validated against the traditional methylene blue method. Most importantly, we performed a time-course study to compare the viability and vitality of lager and ale yeast at Avery Brewing Company, which was used to better understand the physical and metabolic characteristics of yeast throughout the fermentation process. The results provide baseline knowledge for monitoring yeast health that may improve quality assurance procedures and produce more consistent beverage products (quality, flavor, alcohol content, etc.).

Effect of yeast physiological indexes on proteinase A secretion detected by resonance light scattering technique

A simple and sensitive method was developed for the determination of protease A activity (PAA) using the resonance light scattering (RLS) technique. The PAA was measured by a spectrofluorimeter (468 nm, 40 °C, pH 3.0). This method was rapid and convenient with low detection limit of detection (0.662 µg/ml), wide linear range (1–500 µg/ml), and long-term stability (at least 150 min). Using the established RLS method, impact factors of proteinase A secretion were investigated from the angle of yeast strains, physiological state, the generation time (GT), yeast vitality (VT) and yeast mortality. The results showed that there were great differences among the PAA from different yeast strains. For the yeast in the stationary stage, the PAA ascended significantly. The PAA in the yeast with long GT was larger than that with short GT. Early in the fermentation process, the yeast did not secrete proteinase A, but during the final fermentation process, the PAA increased gradually. Meanwhile, The PAA was proportional to yeast mortality, and inversely proportional to yeast VT. Therefore, it is important to select a favorable yeast strains with a shorter GT, higher VT, and lower mortality to produce beers with good foam performance.

Intracellular Temperature Measurement for the Evaluation of Yeast Vitality

Intracellular Temperature Measurement for the Evaluation of Yeast Vitality

EFFECT OF XANTHOHUMOL ON BREWING YEAST CELLS

Xanthohumol, a prenylated chalcone from hops and beer, is a phenolic compound that has received considerable attention in recent years. This compound has a range of interesting biological properties that may have therapeutic utility: anti-inflammatory, antioxidant, antilipoperoxidative activities as well as antiangiogenic, antiproliferative and apoptotic effects, mainly assessed in vitro studies that reasonably suggest a potential chemopreventive activity. Phenomenon how xanthohumol affects brewing yeast’s metabolism, yeast viability and vitality were studied during the production of a xanthohumol enriched beer (10 mg/L xanthohumol). The results showed that yeast viability was slightly decreased by xanthohumol, but on the other hand yeast vitality in the xanthohumol enriched brewing trials was slightly better. The content of higher alcohols and esters was similar to the control in all the xanthohumol enriched brewing trials. In laboratory scale experiment about effect of xanthohumol on brewing yeast growing was done. No effect of xanthohumol on yeast growth was detected. Due to the fact that yeast viability was slightly decreased by xanthohumol, experiment with zymolyase was done. Yeast cells cultivated with xanthohumol are significantly more sensitive to zymolyase treatment.

Impact ofSaccharomyces Cerevisiaeon Viability of ProbioticLactobacillus Rhamnosusin Fermented Milk under Ambient Conditions

The viability of probiotic Lactobacillus rhamnosus HN001 was examined in fermented milk with and without Saccharomyces cerevisiae var. bayanus EC-1118. The yeast enhanced the viability of L. rhamnosus at 30C but not at 12C. High temperature (40C) was detrimental to yeast vitality and diminished its viability-enhancing effect. The yeast inoculated at 103 cfu/g multiplied to 106 cfu/g and enhanced the viability of L. rhamnosus. Viable yeast exerted maximum beneficial effects, although the cell-free yeast supernatant also enhanced the survival of L. rhamnosus. The addition of yeast after fermentation decreased its viability-enhancing effect. Removal of oxygen achieved the viability of L. rhamnosus that was comparable with that achieved in the presence of viable yeast. The results suggest that the improved viability of L. rhamnosus might involve yeast viability, oxygen removal and yeast metabolite(s). The findings would help develop novel microbial starter technology for ambient-stable fermented milks with live probiotics. Practical Applications Maintaining cell viability of probiotics is a scientific challenge, especially under ambient conditions. The findings of this study would lay the foundation for developing novel microbial starter technology for ambient-stable fermented foods with high cell counts of live probiotics, which entails stringent process and engineering control to ensure hygienic conditions and product safety. Benefits of such products would include: reducing distribution and storage costs by saving capital cost involved in chilled-chain distribution, storage and retailing; market penetration and growth where it was not possible due to a lack of or poor refrigeration; extending shelf life and increasing consumer convenience.

A Novel and Rapid Method for Yeast Vitality Evaluation Based on the Methylene Blue Dye Reduction Test

Variation in the quality of yeast is one of the major factors that causes irregularity in fermentation. Yeast quality needs to be estimated accurately to maintain product quality. In the past, several methods for measuring yeast vitality have been proposed. However, these methods have not been widely used in spiked programs for yeast vitality because they show low sensitivity and little correlation with fermentation performance. In this paper, a novel method of evaluating yeast vitality based on the methylene blue dye reduction test (MBRT) is studied. The method is further developed and standardized by measuring varied yeast vitality, and a T½ value is used to estimate yeast vitality. The data demonstrate that MBRT can be used to rapidly and accurately evaluate yeast vitality. T½ values of 0 to 100 sec signify a highly active yeast with good physiological state, while T½ values of 100 to 250 sec indicate reduced metabolic activity and T½ values of more than 250 sec indicate low metabolic competence that could result in sluggish fermentation. This is the first report of the use of quantification of the decoloration rate to measure yeast vitality. The assay could be extended to other applications that involve cell quality.

The Impact of Xanthohumol on a Brewing Yeast's Viability, Vitality and Metabolite Formation

In order to understand how xanthohumol affects a brewing yeast's metabolism, yeast viability and vitality were studied during the production of a xanthohumol enriched beer (10 mg/L xanthohumol) on a 50 L pilot plant scale. The results showed that yeast viability was not significantly affected, but yeast vitality in the xanthohumol enriched brewing trials was slightly better. The content of higher alcohols, esters and organic acids was similar to the control in all the xanthohumol enriched brewing trials, however the content of sulphur dioxide, acetaldehyde and saturated fatty acids was lower in the xanthohumol enriched brewing trials. To the authors' knowledge, this is the first time brewing trials have been carried out showing that xanthohumol has a positive effect on a yeast's physiological condition.

Application of a Short Intracellular pH Method to Flow Cytometry for Determining Saccharomyces cerevisiae Vitality

The measurement of yeast's intracellular pH (ICP) is a proven method for determining yeast vitality. Vitality describes the condition or health of viable cells as opposed to viability, which defines living versus dead cells. In contrast to fluorescence photometric measurements, which show only average ICP values of a population, flow cytometry allows the presentation of an ICP distribution. By examining six repeated propagations with three separate growth phases (lag, exponential, and stationary), the ICP method previously established for photometry was transferred successfully to flow cytometry by using the pH-dependent fluorescent probe 5,6-carboxyfluorescein. The correlation between the two methods was good (r(2) = 0.898, n = 18). With both methods it is possible to track the course of growth phases. Although photometry did not yield significant differences between exponentially and stationary phases (P = 0.433), ICP via flow cytometry did (P = 0.012). Yeast in an exponential phase has a unimodal ICP distribution, reflective of a homogeneous population; however, yeast in a stationary phase displays a broader ICP distribution, and subpopulations could be defined by using the flow cytometry method. In conclusion, flow cytometry yielded specific evidence of the heterogeneity in vitality of a yeast population as measured via ICP. In contrast to photometry, flow cytometry increases information about the yeast population's vitality via a short measurement, which is suitable for routine analysis.

Net effect of wort osmotic pressure on fermentation course, yeast vitality, beer flavor, and haze

The net effect of increased wort osmolarity on fermentation time, bottom yeast vitality and sedimentation, beer flavor compounds, and haze was determined in fermentations with 12 degrees all-malt wort supplemented with sorbitol to reach osmolarity equal to 16 degrees and 20 degrees. Three pitchings were performed in 12 degrees/12 degrees/12 degrees, 16 degrees/16 degrees/12 degrees, and 20 degrees/20 degrees/12 degrees worts. Fermentations in 16 degrees and 20 degrees worts decreased yeast vitality measured as acidification power (AP) by a maximum of 10%, lowered yeast proliferation, and increased fermentation time. Repitching aggravated these effects. The 3rd "back to normal" pitching into 12 degrees wort restored the yeast AP and reproductive abilities while the extended fermentation time remained. Yeast sedimentation in 16 degrees and 20 degrees worts was delayed but increased about two times at fermentation end relative to that in 12 degrees wort. Third "back-to-normal" pitching abolished the delay in sedimentation and reduced its extent, which became nearly equal in all variants. Beer brewed at increased osmolarity was characterized by increased levels of diacetyl and pentanedione and lower levels of dimethylsulfide and acetaldehyde. Esters and higher alcohols displayed small variations irrespective of wort osmolarity or repitching. Increased wort osmolarity had no appreciable effect on the haze of green beer and accelerated beer clarification during maturation. In all variants, chill haze increased with repitching.

Yeast vitality determination based on intracellular NAD(P)H fluorescence measurement during aerobic-anaerobic transition

This work presents a yeast-cell vitality-assessment method based on on-line intracellular fluorescence measurement. The intracellular NAD(P)H fluorescence of a cell suspension is recorded during transition from aerobic to anaerobic conditions and the output signal is evaluated as a measure of yeast vitality (quality). This fluorescence method showed a highly satisfactory correlation with even low dead cell numbers where the acidification power test could not be applied.