High Gravity Brewing by Continuous Process Using Immobilised Yeast: Effect of Wort Original Gravity on Fermentation Performance

The present work evaluated the influence of all‐malt wort original gravity on fermentative parameters and flavour‐active compound formation during primary fermentation of high gravity brewing by a continuous process using a lager yeast immobilised on a natural carrier obtained from brewer's spent grain (the main brewery by‐product). The all‐malt worts with original gravity (OG) ranging from 13.4 to 18.5°Plato were prepared by diluting a very‐high‐gravity wort (20°Plato) with sterile brewery water. The continuous assay was carried out in a bubble column bioreactor with a total working volume of 5.2 litres, at 15°C, using a constant gas flow rate of 250 mL/min (200 mL/min of CO₂ and 50 mL/min of air) and a dilution rate of 0.04 h^?1 (residence time of 25 h). The results indicated that as the wort OG was increased, the ethanol concentration of the outflowing beer increased. On the other hand, the continuous fermentation of the most concentrated worts (16.6 and 18.5°Plato) resulted in beers with unbalanced flavour profiles due to excessive ethyl acetate formation. The immobilised cell concentration appeared to be nearly independent from increasing wort OG.     

High gravity primary continuous beer fermentation using flocculent yeast biomass

The current work assessed a new immobilized cell reactor system throughout a long‐term (54 days) continuous primary fermentation of lager‐type wort of high specific gravity. The experiment was performed in a 4 L airlift bioreactor and immobilization of biomass was attained solely by flocculation. Despite the constant liquid agitation and washout of biomass, up to 53 g dry wt/L of yeast remained immobilized in the system. Two types of beer were produced without interrupting the reactor, based on two types of wort: a Pilsener type with high specific gravity of 15.6 ± 0.3°P; and a dark lager wort with specific gravity of 14.4 ± 0.03°P. Even during the inlet of high gravity wort, the desired attenuation was achieved without the need for either recirculation or an auxiliary second stage bioreactor. The specific saccharide consumption rate was kept around 7.9 ± 0.4 g/L/h and ethanol productivity oscillated at 3.36 ± 0.2 g/L/h for nearly a month. During this period the volumetric productivity of the current bioreactor reached 1.6 L beer/L/day. The green beers produced from the Pilsener and dark lager worts met the standards of regular finished primary beer fermentation. The productivity of diacetyl through the entire experiment could be correlated to the free amino nitrogen consumption rate. Copyright © 2014 The Institute of Brewing & Distilling     

IDENTIFICATION OF LAMBIC SUPERATTENUATING MICRO-ORGANISMS BY THE USE OF SELECTIVE ANTIBIOTICS

The microbial population present in Iambic beer after one year of spontaneous fermentation consists mainly of Brettanomyces yeasts, lactic acid bacteria and acetic acid bacteria. The density of the wort by that time has decreased to around 3.5° Plato. At that time a period of superattentuation is initiated, resulting in a Iambic with sometimes less than 1° Plato. Such old Iambics are used in the production of gueuze. In order to find out which organisms are really necessary for this process. Iambic attenuated to around 3.5° Plato was pasteurized and re-inoculated with a mixed microbial population obtained from fermenting Iambic. By the addition of the antibiotics actidione, pimaricin, gentamycin, oxytetracycline and nisin it was found that Brettanomyces was the main organism responsible for superattenuation, although this was less pronounced when Pediococcus was absent. Acetic acid bacteria were not involved. Bacteria alone were not really superattenuating. The process with Iambic wort which had reached the 3.5° Plato value by a natural spontaneous fermentation was slower than with a Iambic wort pre-fermented to 3.5° Plato with S. cerevisiae. It was found that Brettanomyces but not Saccharomyces survives well under the conditions normally found for a 1 year old Iambic.     

PEPTIDE REMOVAL FROM ALL-MALT AND ADJUNCT WORTS BY SACCHAROMYCES CEREVISIAE NCYC 240

A simple method for following the disappearance of wort peptides during fermentations is described. This method has been used to analyse the effects on peptide removal of supplementing worts with linoleic acid. Supplementation of brewer's all-malt and adjunct worts with linoleic acid caused Saccharomyces cerevisiae NCYC 240 to become enriched with residues of this fatty acid. The rate and extent of fermentation of all-malt wort were unaffected by this supplementation, whereas linoleic acid increased both the rate and extent of fermentation of adjunct wort containing low concentrations of assimilable nitrogen. Removal of peptides by the yeast from all-malt wort occurred at the same rate irrespective of the presence of linoleic acid. However, linoleic acid increased uptake of peptides from adjunct wort.     

Effect of heat treatment of skim milk and final fermentation pH on graininess and roughness of stirred yogurt

The effects of heat treatment of skim milk (95°C/80 s, 95°C/256 s, 110°C/40 s, 110°C/180 s, 130°C/20 s and 130°C/80 s) and final fermentation pH of yogurt (4.8 and 4.4) on physical characteristics of stirred yogurt were investigated. Physical properties, including graininess and roughness, of stirred yogurt were determined during storage at 4°C for 15 days. Number of grains, perimeter of grains, visual roughness, storage modulus and yield stress decreased, when heating temperature or final fermentation pH increased. For practical applications, processing parameters such as heat treatment and fermentation pH can be optimized to improve quality or modified to create fermented milk products with different physical properties.     

Investigation of foam‐active polypeptides during beer fermentation

During ale fermentation there was an accumulation of total and hydrophobic polypeptides in the foam relative to the wort. Comparisons were made not only of the total and hydrophobic polypeptide contents but also of the molecular weights of these polypeptides present in wort, partially fermented wort and its concomitant foam. Wort, fermented wort and foam fractions had very similar polypeptide compositions with a major group having molecular weights of 40–43 kDa. Material of molecular weight in the range of 5–17 kDa and at 66 kDa was also detected. The polypeptides accumulated in foam displayed both hydrophobic and non‐hydrophobic character. The presence of yeast polypeptides in foam was confirmed. Comparison was also made between the fermentations of 10°Plato and 15°Plato wort. The results of the work may contribute to a better understanding of the mechanism of foam formation during beer fermentation, leading to reduced foaming and enabling an increase in the working capacities of fermenters. Copyright © 2004 Society of Chemical Industry     

Impact of full-scale brewing processes on lager beer nitrogen compounds

This paper analyses in a full-scale industrial process firstly the nitrogen compounds in all-malt and in maize-adjunct worts, then their fate during the main brewing steps and finally the influence on them of two different separation technologies: mash filter (Meura 2001) and lauter tun (Steinecker FVAS 26). Data showed that (1) maize-adjunct worts have a lower total nitrogen compounds than all-malt worts; (2) assimilable nitrogen represents 20–24% out of the total nitrogen in both all-malt and in adjunct worts; (3) free amino nitrogen nearly doubles in all-malt compared with adjunct worts; (4) proline and asparagine are the most abundant amino acids in both worts; (5) ammonium disappears during fermentation in wort with the lowest nitrogen content, i.e. in maize-adjunct wort. Moreover, the total nitrogen is reduced in all-malt by 80% with the Steinecker FVAS 26 lauter tun and 25% with the Meura 2001 filter, while in adjunct worts by 87% with the Steinecker FVAS 26 lauter tun and 29% with the Meura 2001 filter. After mash filtration, the content of assimilable nitrogen remains to be adequate for an efficient fermentation, but after lauter tun separation, the assimilable nitrogen reaches values that may compromise the regular fermentation process in both all-malt and adjunct worts. Therefore, when using lauter tun, we have to intervene to reduce its impact on nitrogen compounds and/or plan the wort nitrogen supplementation to overcome the stuck and sluggish fermentations.     

The Yeast Vacuole ‐A Scanning Electron Microscopy Study During High Gravity Wort Fermentations

The yeast vacuole has been shown to exhibit morphological responses to environmental conditions when exposed to worts of different gravity during fermentation. Marked effects of high gravity wort (20° Plato) on yeast morphology compared to more conventional wort gravity (12° Plato) were observed. High gravity worts caused vacuolar enlargement compared to conventional gravity wort. These results suggested that yeast cells experienced severe alterations with the vacuolar tonoplast when exposed to high osmotic pressure and elevated levels of ethanol.     

Comparing the Impact of Environmental Factors During Very High Gravity Brewing Fermentations

The impact of the initial dissolved oxygen, fermentation temperature, wort concentration and yeast pitching rate on the major fermentation process responses were evaluated by full factorial design and statistical analysis by JMP 5.01 (SAS software) software. Fermentation trials were carried out in 2L‐EBC tall tubes using an industrial lager brewing yeast strain. The yeast viability, ethanol production, apparent extract and real degree of fermentation were monitored. The results obtained demonstrate that very high gravity worts at 22°P can be fermented in the same period of time as a 15°P wort, by raising the temperature to 18°C, the oxygen level to about 22 ppm, and increasing the pitching rate to 22 × 10⁶ cell/mL. When diluting to obtain an 11.5°P beer extract, the volumetric brewing capacity increased 91% for the 22°P wort fermentation and 30% using the 15°P wort. After dilution, the fermentation of the 22°P wort resulted in a beer with higher esters levels, primarily the compound ethyl acetate.     

Acid Washing and Serial Repitching a Brewing Ale Strain of Saccharomyces cerevisiae in High Gravity Wort and the Role of Wort Oxygenation Conditions†

Acid washing pitching yeast is an effective method for removing bacterial contamination, but if the yeast is washed incorrectly decreased fermentation performance and beer quality problems may result. Various factors can affect the acid resistance of yeast strains during brewery fermentations. Yeast from shaking flask experiments was more resistant to the combination of high gravity and acid washing conditions than yeast cropped from static fermentations. Yeast harvested from static high gravity wort (20° Plato; 1.083 OG) fermentations was more adversely affected by acid washing than yeast from standard gravity (12° Plato; 1.048 OG) wort. Wort oxygenation resulted in enhanced yeast fermentation performance and healthier yeast crops when yeast was serially repitched into 20° Plato wort. Yeast cropped from fermentations with air saturated high gravity wort responded poorly when acid washed. These results suggest that the structure of the plasma membrane particularly the sterol and fatty acid composition, may have an important role in tolerating high gravity wort and acid washing conditions.     

Production and Characterization of Wine with Sugarcane Piece Immobilized Yeast Biocatalyst

In the present study, an economically cheap and sustainable food-grade yeast biocatalyst for wine production was successfully prepared. It was prepared through absorption technique by using fresh sugarcane (Saccharum officinarum L.) pieces as immobilizing support for yeast, Saccharomyces cerevisiae CFTRI 101. Immobilization was confirmed by scanning electron microscopy. Suitability of biocatalyst was investigated at low and room temperatures by using it in repeated batch. The fermentation rate and other parameters were compared with free yeast cells at different temperatures. In all cases, fermentation time was short (24 h at 30 °C and 98 h at 10 °C) and ethanol productivities were high as 4.2 g/l/h (3.33-fold at 30 °C and 2.31-fold at 10 °C). The volatile compounds methanol, ethyl acetate, propanol-1, isobutanol (2-methyl-1-propanol), and amyl alcohols (2-methyl-1-butanol and 3-methyl-1-butanol) that formed during fermentation were analyzed with the help of a gas chromatograph–flame ionization detector. The concentrations of ethyl acetate and methanol were not more than 100 mg/l in all cases, indicating an improvement in the product quality. Cell metabolism of immobilized yeast was not negatively affected by immobilization process. It was found that the immobilization of yeast cells on sugar pieces increased the fermentation rate and viability of yeast cells. Preliminary sensory tests recognized the fruity aroma, fine taste, and the overall improved quality of the produced wines.     

The Loss of Hydrophobic Polypeptides during Fermentation and Conditioning of High Gravity and Low Gravity Brewed Beer

The object of this study was to investigate the loss of hydrophobic polypeptides, which are important for foam quality and stability in finished beer. Loss of hydrophobic polypeptide due to fermenter foaming occurs during transfer of fermented wort since a gradient of hydrophobic polypeptides towards the surface is created during fermentation. Due to higher polyphenol levels in high gravity (20°Plato) wort, more hydrophobic polypeptides are lost due to cold break (cold trub) precipitation compared to low gravity (12°Plato) wort. Another important factor affecting the loss of hydrophobic polypeptides could be proteinase A activity during fermentation, especially in high gravity fermentation where the yeast is exposed the higher stress. During high gravity fermentation, where osmotic pressures are higher, ethanol levels become greater, and nitrogen‐carbohydrate ratios are lower, more proteinase A is released by the yeast. This release of proteinase A into fermenting wort could have implications for the foam stability of the finished product.     

THE EFFECT OF YEAST TREHALOSE CONTENT AT PITCHING ON FERMENTATION PERFORMANCE DURING BREWING FERMENTATIONS

The effect of yeast trehalose content at pitching on the fermentation performance during brewing fermentations was studied using a commercial strain of lager yeast, Saccharomyces cerevisiae (AJL 2155). Pitching yeasts with different trehalose contents were obtained by collecting cells in suspension after 96 h and 144 h of fermentation in EBC tubes in 10.8°P brewers wort at 14°C. The trehalose content of the pitching yeast had no effect on growth, specific gravity and ethanol production during the subsequent fermentation. A high trehalose content of the pitching yeast, however, sustained cell viability during the initial stage of fermentation, increased the carbohydrate utilisation rate and increased the production of isoamyl alcohol and isobutanol. For these aspects of fermentation performance, the trehalose content of the pitching yeast may prove useful in evaluating the vitality of pitching yeasts within the brewery .     

Brewing with Rice Malt — A Gluten‐free Alternative

The preparation of beer‐like beverages with rice malt as the only raw material is reported. Several tests were performed on a laboratory scale and in a 25 L‐capacity pilot plant. Both the decoction and the infusion procedure were tested; malt and water were mixed in a ratio 1:3.5 for both methods and the mash was brewed without adding exogenous enzymes. The obtained worts were fermented using bottom fermenting yeasts, while “beers” were re‐fermented utilizing top fermenting yeasts and adding either sterile wort or sugar. A maximum ethanol of 4.5% vol. was obtained after the primary fermentation from an initial wort with an original gravity of 11.8°Plato. All parameters of the beer were found to be acceptable using a standard beer analysis. Owing to a suitable hop addition, an aroma very similar to that of a normal beer was obtained.     

Low‐temperature wine‐making using yeast immobilized on pear pieces

A biocatalyst was prepared by the immobilization of Saccharomyces cerevisiae AXAZ‐1 yeast cells on pear pieces and tested for grape must fermentation in both batch and continuous conditions. The immobilized yeast cells were stable and active even at low temperatures (<10 °C). Wine production under batch fermentation at 8 °C was completed within 15 days while at 3 °C it took 36 days. In continuous fermentation, the bioreactor was operated for 33 days, then stored for 12 days at 10 °C, and re‐run for another 15 days without any diminution of the ethanol productivity. Total acidity of the produced wines remained within the ranges usually observed in dry wines, while volatile acidity was found in rather increased levels. The concentrations of higher alcohols (1‐propanol, isobutyl alcohol and amyl alcohols) were relatively low, while ethyl acetate was detected at up to 118 mg l^?1, contributing to the fruity aroma of the wines produced. Preliminary sensory evaluations carried out in the laboratory indicated the fine quality of the produced wines. Copyright © 2004 Society of Chemical Industry     

Freeze-dried wheat supported biocatalyst for low temperature wine making

A new biocatalyst was prepared by immobilization of Saccharomyces cerevisiae AXAZ-1 yeast cells on whole wheat grains. This biocatalyst was subjected to freeze-drying and the effects of several protective agents and storage at 5 °C on viability and fermentative activity of yeasts cells were studied. Glycerol was the best protective agent that preserved the viability of immobilized yeast cells even after 9 month storage. After freeze-drying the biocatalyst was used for repeated batch wine making at extremely low temperatures until 5 °C. The produced wines were analyzed for volatile byproducts by GC and GC/MS and the results showed that the freeze-dried immobilized biocatalysts, with glycerol as protectant, produced wines with higher formation of esters than free cells, having at least similar aromatic profile to those produced by wet biocatalysts.     

Study of some factors involved in ethanal production during alcoholic fermentation

The influence of temperature, turbidity and oxygen uptakes to the musts on the production of ethanal by yeasts, during the alcoholic fermentation (AF), has been studied. The aim was to evaluate how increasing and controlling the production of this molecule, in order to anticipate the reactions between anthocyanins and flavans from the second half of the AF. This concerns, in particular, those winemaking techniques consisting in a temporarily separate fermentation of a part of the must (35–40% of the total mass), that is drained before the start of the AF. The experimental design was a complete factorial plan with three factors upon two levels (T = 18 °C and 22 °C; with or without oxygen; turbidity of 30 and 120 NTU); data were elaborated with 3-ways ANOVA. Temperature proved to be the main factor influencing ethanal production during AF: higher temperatures (22 °C versus 18 °C) cause an increase of the ethanal content during the first days of fermentation; after having reached 6–7% of ethanal, the effect is inverted. The influence of turbidity is less important, and it's synergic with the effect of temperature. Oxygen has a detrimental effect on the production of ethanal only at the beginning of the AF, while later, in presence of ethanol, it promotes it through a chemical way.     

Carbohydrate content and structure during malting and brewing: a mass balance study

A holistic view of the fate of barley starch, arabinoxylan and β-glucan throughout malting and brewing is largely missing. Here, an industrial scale malting trial and pilot brewing trial were performed, and the concentration and structural characteristics of carbohydrates were analysed at 28 key points in the process. The barley starch content decreased during malting from 75.0% to 69.7%. During mashing, malt starch was converted to fermentable sugars (75.3%), dextrin (22.8%) or was retained in spent grains (1.8%). Arabinoxylan was partially hydrolysed during malting. Despite mashing-in at 45°C, no further solubilisation of arabinoxylan was observed during mashing. However, the average degree of polymerisation of the soluble arabinoxylan fraction decreased slightly. During fermentation, the arabinoxylan content decreased to 2.5 g/L. The amount of barley β-glucan decreased gradually in time during malting. Of the solubilised β-glucan, 31% was retained in the spent grains during wort filtration, slightly lowering the β-glucan content in the wort. The β-glucan content remained at 0.5 g/L during fermentation. Sucrose was hydrolysed during mashing, probably by barley invertases. From the total amount of malt used, 41.0% was converted to fermentable sugars. This mashing yield could have been improved by the full hydrolysis to fermentable sugars of the present β-glucan (to 41.1%), the remaining starch in spent grains (to 42.0%) and dextrin in wort (to 50.3%). These results provide more insight into the carbohydrate conversions during malting and brewing and can act as a baseline measurement for future work. © 2020 The Institute of Brewing & Distilling     

Fermented Frozen Dough: Impact of Pre-fermentation Time and of Freezing Rate for a Pre-fermented Frozen Dough on Final Volume of the Bread

This paper proposes an original study on the impact of the process condition using pre-fermented frozen dough on the final bread volume. The impact of the degree of pre-fermentation before freezing and the impact of the freezing conditions have been considered with an experimental design procedure. The first fermentation step (pre-fermentation) was between 60 and 120 min and the corresponding second (or final) fermentation was between 60 and 0 min, respectively, resulting in a total fermentation time of 120 min. Freezing was performed between the first and the second fermentation at −20°C, −30°C or −40°C in blast air tunnel. The results show that the faster the freezing rate, the higher the final bread volume. Further work is needed to fully optimise this process, which is minimally present at the industrial bread market, whereas the technology is now adopted by many viennoiseries such as croissant and laminated puffing pastries.     

Evaluation of the fermentative potential of Candida zemplinina yeasts for craft beer fermentation

The fermentative potential of Candida zemplinina Y.01667 and Y.01670 was evaluated to explore the potential use of these yeasts for craft beer fermentations. Fermentation experiments were carried out at different temperatures and soluble solid concentrations, using synthetic media with glucose syrup as a sugar source and with a laboratory malt wort plus different adjuncts. Results showed that both strains fermented well at 14 °C and had improved fermentative activity at 20 °C. The fermentative kinetics of C. zemplinina Y.01667 and Y.01670 were not affected when experiments at higher concentrations of soluble solids were conducted. Furthermore, C. zemplinina strains had better growth, higher viable cells counts, less free amino nitrogen consumption, lower sedimentation rates and slighter changes in pH values, when compared with results of the lager beer yeast Saccharomyces cerevisiae S‐23 in the synthetic medium tested. Fermentations in a malt wort with different adjuncts indicated that C. zemplinina Y.01670 could possibly be used as a yeast in craft beer production. Copyright © 2016 The Institute of Brewing & Distilling