Rapid fermentation of beer using an immobilized yeast multistage bioreactor system

A multistage bioreactor system for rapid beer fermentation was developed. The main fermentation process, which conventionally requires 7 d, could be shortened to 2 d by this system. The concentration of esters and higher alcohols are major factors in brewery fermentation, their production being closely related to the yeast growth phase. Yeast metabolism was successfully subdivided into a growth and a restricted phase through a combination of a continuous stirredtank reactor (CSTR) and an immobilized yeast packed-bed reactor (PBR). Production of higher alcohols was high in the CSTR because of its association with the level of biosynthetic activityde novo. A small amount was also produced in the PBR, however, possibly a result of an overflow in carbohydrate metabolism. Ester formation mainly occurred in the PBR, a linear increase in the level of ester being observed with flow through the PBR. The reactor system control strategy was to maximize the level of both higher alcohol and ester formation. The CSTR/PBR control range, based on extract consumption, was varied between 1:1 and 1:2. A ratio of 1:1 tended to create a flat beer, whereas a ratio 1:2 gave a beer of richer quality. Amino acid uptake by the yeast directly contributed to a reduction in the wort pH, whereas no relation could be observed between the level of organic acid production and pH.     

Continuous Production of Isomalto-oligosaccharides by Thermo-inactivated Cells of <Emphasis Type="Italic">Aspergillus niger</Emphasis> J2 with Coarse Perlite as an Immobilizing Material

The coarse perlite 40–80 mesh was selected as an immobilizing material and put into a packed bed reactor (PBR) to continuously convert maltose to isomalto-oligosaccharides (IMOs). The PBR was prepared by mixing the thermo-inactivated cells (TIC) from Aspergillus niger J2 strain with the coarse perlite, then the mixture was put into an overpressure-resistant column. Compared with diatomite 40–80 mesh and thin perlite 80–120 mesh in PBR, coarse perlite was chosen as the best filtration aid, when the ratio of coarse perlite versus TIC was 1:1. The thermal and pH stability of the free and immobilized TIC and the optimum conditions for the transglycosylation reactions were determined. The results show that approximately 75 and 82% and 87 and 91% of α-glucosidase activity were reserved for free and immobilized TIC at temperatures from 30 to 60 °C and pH from 3.00 to 7.00 for 12 h, respectively. With 30% malt syrup under the conditions of 50 °C and pH 4.00, a mini-scale packed bed reactor (Mi-PBR) and medium-scale packed bed reactor (Me-PBR) could continuously produce IMO over 25 and 34 days with the yield of effective IMO (eIMO) ≥?35% and total IMO (tIMO) ≥?50%, respectively. The strategy of mixing the coarse perlite with TIC in PBR is a novel approach to continuously produce IMO and has great application potential in industry.     

Enabling Heterologous Synthesis of Lupulones in the Yeast Saccharomyces cerevisiae

Lupulones, naturally produced by glandular trichomes of hop (Humulus lupulus), are prenylated phloroglucinol derivatives that contribute the bitter flavor of beer and demonstrate antimicrobial and anticancer activities. It is appealing to develop microbial cell factories such that lupulones may be produced via fermentation technology in lieu of extraction from limited plant resources. In this study, the yeast Saccharomyces cerevisiae transformants harboring a synthetic lupulone pathway that consisted of five genes from hop were constructed. The transformants accumulated several precursors but failed to accumulate lupulones. Overexpression of 3-hydroxy-3-methyl glutaryl co-enzyme A reductase, the key enzyme in precursor formation in the mevalonate pathway, also failed to achieve a detectable level of lupulones. To decrease the consumption of the precursors, the ergosterol biosynthesis pathway was chemically downregulated by a small molecule ketoconazole, leading to successful production of lupulones. Our study demonstrated a combination of molecular biology and chemical biology to regulate the metabolism for heterologous production of lupulones. The strategy may be valuable for future engineering microbial process for other prenylated natural products.

     

Microrobots in Brewery: Dual Magnetic/Light-Powered Hybrid Microrobots for Preventing Microbial Contamination in Beer

Yeasts play a key role in the production of alcoholic beverages by fermentation processes. However, because of their continuous growth, they commonly cause spoilage of the final product. Herein, we introduce dual magnetic/light-responsive self-propelled microrobots that can actively move in a beer sample and capture yeast cells. The presence of magnetic nanoparticles on the surface of the microrobots enables their magnetic actuation under fuel-free conditions. In addition, their photoactivity under visible-light irradiation leads to an overall enhancement of their swimming and yeast removal capabilities. It was found that after the application of the microrobots into a real unfiltered beer sample, these micromachines were able to remove almost 100 % of residual yeasts. In addition, these microrobots could also be added at the initial step of the fermentation process without altering the final beer properties, such as alcohol level, color, and pH. This work demonstrates the potential of using externally actuated microrobots as an innovative and low-cost solution for avoiding yeast spoilage in complex liquid environments, such as alcoholic beverages. Therefore, these autonomous self-propelled microrobots open new avenues for future applications in the food industry.     

On-line dialysis of organic acids from a Propionibacterium freudenreichii fermentation

The efficiency of an ion exchange system coupled to a bioreactor to extract on-line inhibitory organic acids produced was evaluated. Batch fermentations without dialysis and fed-batch experiments with and without dialysis of Propio nibacterium freudenreichii were conducted. It was possible to keep the propionic acid concentration in the reactor at a low level to avoid complete growth inhibition. Improvements in biomass and propionic acid productivities were achieved when the dialysis system was used. The performance of the dialysis system was improved when a new pH control strategy evaluated in this study was used.     

The Effect of Sound Frequency and Intensity on Yeast Growth,Fermentation Performance and Volatile Composition of Beer

This study investigated the impact of varying sound conditions (frequency and intensity) on yeast growth, fermentation performance and production of volatile organic compounds (VOCs) in beer. Fermentations were carried out in plastic bags suspended in large water-filled containers fitted with underwater speakers. Ferments were subjected to either 200–800 or 800–2000 Hz at 124 and 140 dB @ 20 µPa. Headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) was used to identify and measure the relative abundance of the VOCs produced. Sound treatment had significant effects on the number of viable yeast cells in suspension at 10 and 24 h (p < 0.05), with control (silence) samples having the highest cell numbers. For wort gravity, there were significant differences between treatments at 24 and 48 h, with the silence control showing the lowest density before all ferments converged to the same final gravity at 140 h. A total of 33 VOCs were identified in the beer samples, including twelve esters, nine alcohols, three acids, three aldehydes, and six hop-derived compounds. Only the abundance of some alcohols showed any consistent response to the sound treatments. These results show that the application of audible sound via underwater transmission to a beer fermentation elicited limited changes to wort gravity and VOCs during fermentation.     

Magnetic resonance imaging of flow-distributed oscillations

The formation of stationary concentration patterns in a packed-bed reactor (PBR), using a manganese-catalyzed Belousov-Zhabotinsky (BZ) reaction in a mixed sulfuric-phosphoric acid medium, was studied using magnetic resonance imaging (MRI). The PBR was composed of a column filled with glass beads, which was fed by a continuous stirred tank reactor (CSTR). As the reactor is optically opaque, investigation of the three-dimensional (3D) structure of these reaction-diffusion-advection waves is not possible using conventional image capture techniques. MRI has been used to probe this system and the formation, 3D structure, and development of these waves has been studied. At reactor startup, traveling waves were observed. After this initial period the waves stabilized and became stationary. Once fixed, they were found to be remarkably stable. There was significant heterogeneity of the reaction fronts, which were not flat, as would be expected from a plug-flow reactor. Instead, the reaction wave fronts were observed to be conical in shape due to the local hydrodynamics of the bed and specifically the higher velocities and therefore lower residence times close to the wall of the reactor.     

Development of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> Producing Higher Levels of Sulfur Dioxide and Glutathione to Improve Beer Flavor Stability

Sulfur compounds, such as sulfite (SO₂), hydrogen sulfide (H₂S), and glutathione (GSH), play different roles in beer flavor stability. SO₂ and GSH have antiaging effects which are helpful to improve the flavor stability of beer, whereas H₂S is undesirable to beer flavor because of its unpleasant aroma. Here, we report the development of Saccharomyces cerevisiae which produces higher levels of SO₂ and GSH but lower level of H₂S to improve beer flavor stability by nongenetic engineering approaches. After two rounds of UV mutagenesis coupled with specific plate screening methods, one promising mutant named MV16 was obtained. Compared with the original strain, the SO₂ and GSH production of MV16 in fermenting liquor increased by 31% and 30.2%, respectively, while H₂S content decreased by 74.9%, and the DPPH radical clearance and the resistance staling value of beer fermented by MV16 increased by 24.6% and 33.0%, respectively. The antioxidizability of the mutant was improved significantly. The strategy adopted in our study could be used to obtain S. cerevisiae of improved antiaging properties, and the mutant would be safe for public use.     

Biomaterials from beer manufacture waste for bone growth scaffolds

Abstract

Agricultural wastes are a source of renewable raw materials (RRM), with structures that can be tailored for the use envisaged. Here, they have proved to be good replacement candidates for use as biomaterials for the growth of osteoblasts in bone replacement therapies. Their preparation is more cost effective than that of materials presently in use with the added bonus of converting a low-cost waste into a value-added product. Due to their origin these solids are ecomaterials.

In this study, several techniques, including X-ray diffraction (XRD), chemical analysis, mercury intrusion porosimetry (MIP), scanning electron microscopy (SEM), and bioassays, were used to compare the biocompatibility and cell growth of scaffolds produced from beer bagasse, a waste material from beer production, with a control sample used in bone and dental regenerative processes.     

Highly improved electrocatalytic behavior of sulfite at carbon ionic liquid electrode: application to the analysis of some real samples

The electrocatalytic oxidation of sulfite was investigated at carbon ionic liquid electrode (CILE). This electrode is a very good alternative to previously described electrodes because the electrocatalytic effect is achieved without any electrode modification. Comparative experiments were carried out using carbon paste electrode (CPE) and glassy carbon electrode (GCE). At CILE, highly reproducible and well-defined cyclic voltammograms were obtained for sulfite with a peak potential of 0.55 V vs. Ag/AgCl. Sulfite oxidation at CILE does not result in deactivation of the electrode surface. The kinetic parameters for this irreversible heterogeneous electron transfer process were determined. Under optimal experimental conditions, the peak current response increased linearly with sulfite concentration over the range of 6-1000 μM. The detection limit of the method was 4 μM. The method was applied to the determination of sulfite in mineral water, grape juice and non-alcoholic beer samples.     

Production of polyhydroxyalkanoate from sesame seed wastewater by sequencing batch reactor cultivation process of Haloferax mediterranei

Due to the high cost of bioplastic production, sesame wastewater, generated from the sesame seed hulling process, was investigated to be used as inexpensive and renewable carbon source for the production of biodegradable polyhydroxyalkanoate (PHA) by extreme Haloferax mediterranei. The sesame wastewater (SWW) was hydrolyzed using different concentrations of hydrochloric acid (0.4. 1.00 and 2.00 M) at different period of times (15, 60 and 90 min). The concentration of salt (NaCl) and nitrogen source (NH₄Cl and yeast) required for H. mediterranei cells growth and the accumulation of PHA biopolymer was optimized. A maximum 0.53 g/L concentration of PHA was achieved when the SWW extract media was supplemented with 100 g/L NaCl and 6.0 g/L yeast extract. The cultivation was scaled-up using sequencing batch reactor (SBR) fermentation under non-sterile conditions. The SBR results showed that SWW needs an auxiliary carbon source to obtain high PHA production. Consequently, the system fed with SWW and glucose produced higher PHA (20.9 g/L) than the system fed with SWW.     

Experimental techniques,data treatment for studying the dynamics of ethanol production/consumption in baker's yeast

The dynamics of ethanol production/consumption in baker's yeast were studied under feed- rate controlled conditions. The yeast was grown on molasses in an 8-l fed-batch reactor and experiments were done at cell concentrations between 5 and 65 g l^?1. Small changes in the feed rate were made around a feed rate corresponding to the critical growth rate, at which the yeast cell metabolism switches between ethanol consumption and production. A membrane gas sensor was used for on-line measurement of the ethanol concentration in the broth. The measured ethanol signal was used for control and the system was excited through changes in the regulator set-point. The closed-loop experiments ensured that feed variations were within the critical range, and thus facilitated reproducible experiments. Data were fitted to a second-order difference equation by statistical methods. Results were compared with a theoretically derived model. The process gain could be understood in terms of the underlying stoichiometry by using the “bottleneck” view of yeast glucose metabolism. The process time constant was found to be longer than is implied by a simple Monod relation between glucose uptake rate and concentration.     

Electrodeposition of coatings from sulfite gold-plating electrolyte and their properties

Mechanism of the electrolytic gold-plating and some properties of coatings deposited from a sulfite gold-plating electrolyte were studied. It was shown that soft coatings with low internal stresses can be produced from sulfite electrolytes. The mechanism of the electrolytic gold-plating process was described in terms of the nucleation–growth model in which the metal is deposited at growth points via a single-electron reaction in which sulfite complexes of gold(I) are reduced.     

Characteristics of New England India Pale Ale Beer Produced with the Use of Norwegian KVEIK Yeast

The aim of this research was to determine the potential of four unconventional Norwegian yeasts of the KVEIK type to produce NEIPA beer. The influence of yeast strains on fermentation process, physicochemical properties, antioxidant potential, volatile compounds, and sensory properties was investigated. The KVEIK-fermented beer did not differ in terms of physicochemical parameters from the beer produced with the commercial variants of US-05 yeast. The yeast strain influenced the sensory quality (taste and aroma) of the beers, with KVEIK-fermented beer rating significantly higher. The antioxidant activity of the tested beers also significantly depended on the yeast strain applied. The beers fermented with KVEIK had a significantly higher antioxidant potential (ABTS^•+) than those fermented with US-05. The strongest antioxidant activity was found in the beer brewed with the Lida KVEIK yeast. The use of KVEIK to produce NEIPA beer allowed enrichment of the finished products with volatile compounds isobutanol, 2-pentanol, 3-methylobutanol, ethyl octanoate, and ethyl decanoate.     

Effect of the variation of the level of lactose conversion in an immobilized lactase reactor upon operating costs for the production of baker’s yeast from hydrolyzed cottage cheese whey permeate

Operating costs for the production of Baker’s yeast from hydrolyzed permeate from the ultrafiltration of cottage cheese whey were calculated as a function of the level of lactose conversion in the immobilized lactase reactor. These costs were calculated for the case of 90% conversion of lactose in the reactor and compared to those that result when running the reactor at lower conversions with recycle of unreacted lactose. Total operating costs were estimated by combining individual operating costs for the immobilized enzyme reactor, costs associated with processing a lactose recycle stream, and energy costs associated with cooling the reactor feed stream and sterilizing the hydrolysate stream. It was determined that operating costs are minimized at about 9.9 ￠/lb. of lactose when the reactor is run at approx. 72% conversion. This represents a savings of 2.4 ￠/lb. of lactose over the case of a once-through 90% conversion of lactose in the reactor.     

Beer clarification by microfiltration — product quality control and fractionation of particles and macromolecules

Beer clarification by microfiltration demands a finely balanced retention of colloidal particulates (yeast cells, chill haze flocs, etc.) and transmission of soluble macromolecules including carbohydrates, proteins, flavour, and colour compounds which give the “whole some” quality of a beer. The required porous transmission of these macromolecular species led to an unavoidable, complex and dynamic in-pore membrane fouling in terms of fouling constituents, formation, structure and kinetics, which are the main obstacles in obtaining an economically viable flux and consistency in permeate quality.This experimental study was carried out with the aims of understanding the dynamic inter-relation between flux, fouling and system selectivity during a cross-flow beer microfiltration process so that an effective operating strategy for flux optimisation could be formulated in conjunction with the parallel objective of good product (permeate) quality control. Tubular ceramic membranes (Ceramem) with nominal pore diameters of 0.2, 0.5, and 1.3 μm were used. Simultaneous measurement of flux and permeate qualities, such as specific gravity and chill haze level enabled identification of the effect of anti-fouling techniques, such as backflushing on transmission of essential beer components and on the filtered beer quality. The experimental evidence lead to an understanding that the drastic flux enhancement achieved by employing backflushing at reversed membrane morphology was associated with enhanced solute transmission which could, without careful control, upset a balanced transmission of essential beer components and the retention of unwanted “chill haze” components. Further operating parameters and varying system configurations were investigated over their effect on both flux performance and system selectivity. These include membrane pore size, filtration temperature, and the addition of an amorphous silica particles as coagulation agent for hydrophilic proteins.     

超高效液相色谱-串联质谱法检测麦汁和啤酒中的麦香风味物质

应用超高效液相色谱-串联质谱(UPLC-MS/MS)技术定量检测了啤酒和麦汁中的2,5-二甲基-4-羟基-3(2H)-呋喃酮(DMHF)、2(或5)-乙基-5(或2)-甲基-4-羟基-3(2H)-呋喃酮(EMHF)和2-乙酰吡咯(2-AP)3种麦香风味物质。使用C₁₈固相萃取柱净化样品。采用ACQUITY UPLC HSS T3色谱柱(50 mm×2.1 mm, 1.8 μm)分离,以0.1%(v/v)甲酸水溶液和0.01%(v/v)甲酸乙腈溶液为流动相进行梯度洗脱。结合这3种化合物的保留时间,在正离子模式下,采用多反应监测(MRM)技术进行定量检测。当质量浓度低于1000 μ g/L时,校准曲线的线性良好(R²>0.999)。方法的加标回收率在74.3%~86.7%之间,相对标准偏差(RSD,n=6)在4.8%~7.3%之间。由于酵母发酵时会生成DMHF和EMHF,导致啤酒中这两种物质的含量明显高于麦汁。某些品类啤酒如印度淡色爱尔啤酒(IPA),通常含有较高的麦香风味物质。该法样品处理简单,选择性好,且灵敏、准确、重现性好,可用于啤酒生产的过程控制。     

The Elemental Profile of Beer Available on Polish Market: Analysis of the Potential Impact of Type of Packaging Material and Risk Assessment of Consumption

Twenty-five elements, including the most essential and toxic metals, were determined in fifty beer samples stored in cans and bottles by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) and Cold Vapor Atomic Absorption Spectroscopy (CVAAS) techniques. The packaging material was analyzed using the Scanning Electron Microscopy with Energy Dispersive Spectroscopy (SEM-EDS) technique. The control of the level of individual metals is necessary, not only to maintain the organoleptic properties of the product, but also to fulfill the standards regarding the permissible maximum concentrations. Metals can originate from different sources, including the brewing water, malt grains, hops, adjuncts, fruits, and spices. They may also come from contamination from the brewery equipment, i.e., vessels and tanks, including beer packing, storing and transporting (kegs, casks, cans). Discriminant analysis revealed that the differentiation of three types of beer (Lager, Ale, Craft) was possible, based on elemental concentrations, for the reduced data set after their selection using the Kruskal-Wallis test. The analysis of the impact of the packaging material (can or bottle) proved that when this parameter was used as a differentiating criterion, the difference in the content of Na, Al, Cu and Mn can be indicated. The risk assessment analysis showed that the consumption of beer in a moderate quantity did not have any adverse effect in terms of the selected element concentrations, besides Al. However, in the case of Al, the risk related to consumption can be considered, but only for the beer stored in cans produced from aluminum.     

Banana as Adjunct in Beer Production: Applicability and Performance of Fermentative Parameters

Traditionally, the raw materials for beer production are barley, hops, water, and yeast, but most brewers use also different adjuncts. During the alcoholic fermentation, the contribution of aroma compounds from other ingredients to the final beer flavor depends on the wort composition, on the yeast strain, and mainly on the process conditions. In this context, banana can also be a raw material favorable to alcoholic fermentation being rich in carbohydrates and minerals and providing low acidity. In this work, the objective was to evaluate the performance of wort adjusted with banana juice in different concentrations. For this, static fermentations were conducted at 15 °C at pilot scale (140 L of medium). The addition of banana that changed the concentration of all-malt wort from 10 °P to 12 and 15 °P were evaluated (°P is the weight of the extract or the sugar equivalent in 100 g solution, at 20 °C). The results showed an increase in ethanol production, with approximately 0.4 g/g ethanol yield and 0.6 g/L h volumetric productivity after 84 h of processing when concentrated wort was used. Thus, it was concluded that banana can be used as an adjunct in brewing methods, helping in the development of new products as well as in obtaining concentrated worts.     

Characterization of winemaking yeast by cell number-size distribution analysis through flow field-flow fractionation with multi-wavelength turbidimetric detection

Yeasts are widely used in several areas of food industry, e.g. baking, beer brewing, and wine production. Interest in new analytical methods for quality control and characterization of yeast cells is thus increasing. The biophysical properties of yeast cells, among which cell size, are related to yeast cell capabilities to produce primary and secondary metabolites during the fermentation process. Biophysical properties of winemaking yeast strains can be screened by field-flow fractionation (FFF). In this work we present the use of flow FFF (FlFFF) with turbidimetric multi-wavelength detection for the number-size distribution analysis of different commercial winemaking yeast varieties. The use of a diode-array detector allows to apply to dispersed samples like yeast cells the recently developed method for number-size (or mass-size) analysis in flow-assisted separation techniques. Results for six commercial winemaking yeast strains are compared with data obtained by a standard method for cell sizing (Coulter counter). The method here proposed gives, at short analysis time, accurate information on the number of cells of a given size, and information on the total number of cells.