Variation in the molecular mass of exopolysaccharides during the time course of extended fermentations of skimmed milk by lactic acid bacteria

The weight average molecular mass (M_w) of the exopolysaccharide (EPS) secreted by Lactobacillus acidophilus 5e2 when grown on skimmed milk supplemented by glucose was monitored during extended fermentation times. During the exponential growth phase, the increase in M_w closely followed the increase in yield of EPS. Under the fermentation conditions applied in this study, few if any new polysaccharide chains were formed during this growth phase despite a twenty five-fold increase in the cell count; almost the entire increase in yield can be accounted for by an increase in chain length. These results suggest that synthesis of new EPS chains is switched off during the exponential and stationary phase of fermentation. The increase in yield observed in this period is a consequence of the bacteria's ability to extend existing chains right up to the mid-stationary phase. These results raise questions about the factors that control EPS production and chain length.     

SOME EFFECTS OF AERATION ON THE GROWTH AND METABOLISM OF SACCHAROMYCES CEREVISIAE IN CONTINUOUS CULTURE

In continuous fermentation of a complex, synthetic medium by Saccharomyces cerevisiae, a steady state was maintained for prolonged periods in the absence of oxygen. Growth reached a maximum at a low rate of aeration and under these conditions ester formation was markedly inhibited. At higher levels of aeration growth was reduced, the cells formed chains and aggregated, the rate of fermentation decreased and large quantities of acetoin and acetaldehyde were produced. Under fully aerobic conditions, ethanol production continued to account for the major part of the glucose consumed. The amount of diacetyl produced was insignificant at all rates of aeration. The extent of growth and the nitrogen content of the yeast were inversely related.     

MICROBIOLOGICAL ASPECTS OF A MIXED YEAST—BACTERIAL FERMENTATION IN THE PRODUCTION OF A SPECIAL BELGIAN ACIDIC ALE

The evolution of the microbial population during the industrial fermentation of a Belgian acidic ale was followed, and the most important micro-organisms were identified. In essence, the process can be considered to be a tandem fermentation, as a period of ethanolic fermentation is followed by a period of lactic acid fermentation. The process consists of three stages. The first stage is a seven day fermentation mainly by Saccharomyces; the second is a four to five week fermentation in which Lactobacilli become important; in the third stage a twenty to twenty-four month fermentation in cask, Brettanomyces species, Lactobacilli, Pediococcus parvulus and Acetic Acid bacteria are all present. The evolution of the Brettanomyces-Pediococcus coculture during the tertiary fermentation shows a remarkable similarity with the evolution during a gueuze fermentation.     

MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE

Lambic and Gueuze are special Belgian beers obtained by spontaneous fermentation. Micro-organisms involved in this fermentation were counted and differentiated using several selective growth media. Micro-organisms were also isolated from samples of Lambic of different age and originating from different casks and brews and identified by classical tests. The following general pattern of microbial development was observed. After 3 to 7 days the fermentation started with the development of wort Enterobacteriaceae and strains of Kloeckera apiculata. These organisms were overgrown after 3 to 4 weeks by strains of Saccharomyces cerevisiae and S. bayanus. These were responsible for the main fermentation, lasting for 3 to 4 months. This was followed by a strong bacterial activity. This period was dominated by the growth of strains of Pediococcus cerevisiae. These reached their maximal numbers during the summer months and were responsible for a fivefold increase in lactic acid concentration. In some casks they caused ropiness. After the main fermentation period Lambic is very sensitive to spoilage by acetic acid bacteria of the genus Acetomonas. The presence of air may be the determining factor for their development. After 8 months a new increase in yeast cells was noted. These belonged now mainly to the genus Brettano-myces bruxellensis and Br. lambicus. They caused a further slow decrease in residual extract and the appearance of special flavours. Oxidative yeasts of the genera Candida, Cryptococcus, Torulopsis and Pichia were also detected and may be responsible for the formation of a flim on the beer surface after the main fermentation.     

CONTROL OF AMMONIUM UPTAKE FROM MALT EXTRACT MEDIUM BY SACCHAROMYCES CEREVISIAE

When glucose was used as the energy source in the assay, the perceived ammonium permease activity of Saccharomyces cerevisiae NCYC 1108 remained relatively unchanged in yeast samples taken throughout the main period of fermentation of a malt extract medium. This was in contrast to the earlier work on the fermentation of a defined glucose/salts medium where the specific ammonium permease activity rose rapidly as fermentation proceeded. The present data suggest that the level of ammonium permease activity of cells fermenting a malt extract medium was low due to nitrogen catabolite repression and that inhibition of the carrier system was important in determining the actual rate of uptake from the medium. When ammonium permease in cells from a malt extract medium was assayed using maltose, essentially similar results were obtained. However, the actual values were lower and the results suggested that at some points during the fermentation the ability of the cells to take up maltose during the assay was an important factor in determination of the perceived ammonium permease activity.     

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 .     

Texture improvement of fermented minced pepper under vacuum impregnation with pectin methylesterase and CaCl2 during fermentation

Fermented minced pepper (FMP) usually suffers from the deterioration of texture quality during fermentation, which can affect sensory and consumer acceptance. In this study, vacuum impregnation (VI) with CaCl₂, pectin methylesterase (PME) and CaCl₂ and PME (PME + CaCl₂ + VI) were compared to improve the texture quality of FMP. FMP treated with PME + CaCl₂ + VI showed the relatively intact cells structure after fermentation. In that case, its firmness maintained high level, while water-soluble pectin (WSP) content was minimum after fermentation. Meanwhile, the molar ratio of most monosaccharides of WSP in PME + CaCl₂ + VI treated FMP decreased, while rhamnose (Rha) molar ratio significantly (p < 0.05) increased after fermentation. The high Rha content represents the stability of rhamnogalacturonan-I linear skeleton of WSP. The negative effect on molecular weight of WSP was delayed by PME + CaCl₂ + VI treatment, and its peak area and value increased after fermentation. Atomic force microscope images indicated that PME + CaCl₂ + VI treatment could retain the long chain and branch structures, and inhibit the degradation of WSP net-like structure at some extent. Hence, PME + CaCl₂ + VI treatment was effective to improve the texture of FMP and inhibit the solubilisation of WSP via the formation of cross-linked pectin chains between Ca²⁺ and demethylesterified pectin.     

Variation in maltose in sweet wort from barley malt and rice adjuncts with differences in amylose structure

Starch from malt and solid adjuncts provides the majority of fermentable sugars for fermentation. However, there is no current data on the variation in starch structure (particularly long chained amylose) and its impact on the final wort composition of fermentable sugars, specifically maltose. This is the first study to report variation in amylose structure from barley malt and rice used as an adjunct and how this impacts the production of maltose. We compared four commercial malts with two rice adjuncts mashes, in solid and liquid additions, with an all‐malt mash used as a control. All combinations of malt and rice adjuncts were tested under two grist‐to‐liquor (G:L) ratios (1:3 and 1:4) in a 65°C ramped mash. After mashing, the wort original gravity and maltose concentration were measured. The commercial malts had different malt quality but very similar gelatinisation temperatures (~65°C). The malts varied in starch and amylose contents but had only minor variations in average amylose chain lengths. The two rice adjuncts also had similar average amylose chains lengths, but quite different amylose contents, and hence different gelatinisation temperatures. The results showed that liquid adjunct mashes had higher original gravity and maltose concentration for both G:L ratios. However, there was no consistent result in original gravity or maltose between G:L ratio or adjunct type, suggesting interactions between each malt and rice adjunct. Knowing amylose chain length could improve understanding of the potential maltose levels of the sweet wort prior to fermentation. © 2018 The Institute of Brewing & Distilling     

ACCELERATED PROTEOLYSIS OF SOY PROTEINS DURING FERMENTATION OF THUA-NAO INOCULATED WITH BACILLUS SUBTILIS

Thua‐nao is a traditional Thai‐fermented soy product. It was prepared by the conventional method and also with starter inoculation. Inoculated soybean exhibited a higher rate of fermentation than did the natural fermentation as indicated by greater rate of pH increase, higher extent of proteolysis, ammonia‐nitrogen content and nitrogen solubility. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis patterns indicated the extensive degradation of soy proteins during fermentation. Activity staining of Thua‐nao extract suggested that proteolysis of soybean was mediated by various proteinases. Active proteinases with estimated molecular weight (MW) of 29,000, 27,000 and 19,000 Da were present during natural fermentation of soybeans. However, proteinases with MW of 40,000 and 29,000 Da were predominant in inoculated soybeans and like the activity bands observed with spent culture broth of Bacillus subtilis BIOTEC 7123. The results suggested that proteinases released by the dominant species in the inoculum, especially B. subtilis, play an important role in proteolysis of soy proteins during fermentation. Inoculation with B. subtilis resulted in an increased proteolysis, which is useful for subsequent reactions leading to development of Thua‐nao characteristics.     

THE ROLE OF DIMETHYL SULPHOXIDE REDUCTASE IN THE FORMATION OF DIMETHYL SULPHIDE DURING FERMENTATIONS

The ability to produce dimethyl sulphide (DMS) during fermentation of wort is apparently a general characteristic of Saccheromyces cerevisiae and Saccharomyces uvarum. Washed suspensions of these yeasts reduce dimethyl sulphoxide (DMSO) but there is no correlation between DMS formation in fermentation and DMSO reduction in whole cells. Although different strains vary widely in their ability to reduce DMSO they all contain similar levels of NADPH—dependent DMSO reductase. This enzyme is a multi-protein system which closely resembles methionine sulphoxide reductase. Spoilage bacteria including Enterobacter cloacae can reduce DMSO more efficiently than can yeast probably because a different enzyme system is involved.     

Quality control method based on quartz crystal microbalance and WGA for flour milled from germinated wheat

Plasma membrane is the initial sensor of different stress conditions and its composition is modified with response to environmental changes. In the present study, we have modified the lipid composition of the membrane by growing Saccharomyces cerevisiae in the presence of different fatty acids and ergosterol. All supplemented fatty acids were incorporated into the cell and this incorporation produced significant changes in the lipid composition. The incubation with ergosterol also modified the lipid composition of the cells; however, these cells presented a strong reduction in the content of this sterol. The different cellular lipid composition has been related to viability and fermentation performance at low temperature (13 °C). The cells incubated with palmitoleic acid (C16:1) showed higher viability and significant reduction in the fermentation length. These cells presented higher C16:1 and ergosterol content, shorter chain length of the fatty acids and higher ratio of sterols/phospholipids. Therefore redesigning the composition of cellular membranes during industrial yeast propagation seems to be a promising strategy for improving fermentation performance in the winery.     

Changes in texture qualities and pectin characteristics of fermented minced pepper during natural and inoculated fermentation process

Texture qualities and pectin characteristics in fermented minced pepper (FMP) prepared by natural fermentation (NF) and inoculated fermentation (IF) were analysed during fermentation. The results showed variation in texture qualities and pectin characteristics was similar during NF and IF process. The hardness, cell wall material, sodium carbonate-soluble pectin (SSP) and chelate-soluble pectin (CSP) content, and CSP esterification degree decreased, while water-soluble pectin (WSP) content significantly (P < 0.05) increased after fermentation. The rhamnose (Rha) molar ratio in three pectins increased, but arabinose (Ara) and galactose (Gal) molar ratios in most pectins decreased after fermentation. Changes in Ara/Gal and (Gal + Ara)/Rha ratios represented the backbone and branched chains of rhamnogalacturonan-I in three pectins depolymerised during fermentation. The decrease of molecular weight (M_w) in CSP was more obvious than that in WSP and SSP, and it was extensively depolymerised into low-M_w pectin after fermentation. Pearson's correlation analysis showed FMP hardness was extremely (P < 0.01) positively correlated with CSP content and significantly (P < 0.05) positively correlated with SSP content and CSP M_w. Hence, CSP was the main pectin to affect texture compared with WSP and SSP, and its characteristics played a crucial role for regulating FMP texture during NF and IF process.     

FORMATION OF HIGH CONCENTRATIONS OF ALCOHOL BY VARIOUS YEASTS*

The presence of Aspergillus oryzae-proteolipid (PL) at the early stage off fermentation was essential to obtain yeast cells capable of producing high concentrations of alcohol, when the fermentation tests were carried out by stepwise addition of sucrose to the synthetic media. High concentrations of alcohol were produced by strains of Saccharomyces cerevisiae Hansen and Saccharomyces uvarum (carlsbergensis) as well as Saccharomyces sake which produced more than 20 percent alcohol in the PL-supplemented medium at 20°C.     

SYNTHESIS OF HIGHER ALCOHOLS IN THE GENUS ZYMOMONAS

Several strains of bacteria of the genus Zymomonas were examined with respect to their potential for higher alcohol synthesis. All strains studied were able to produce higher alcohols during growth in a simple medium containing glucose and yeast extract. The higher alcohols produced were mainly n-propanol and iso-amyl alcohol. In contrast to Saccharomyces cerevisiae, only trace amounts of higher alcohols were produced from glucose by resting cells. When amino acids or other precursors were added to the fermentation medium, the resting cells formed higher alcohols. The stimulation of n-propanol synthesis by precursors was the most pronounced. The results obtained indicate that, with minor differences, the mechanisms of higher alcohol synthesis are comparable to these used by yeasts.     

LACTIC ACID BACTERIA AND MALOLACTIC FERMENTATION IN THE MANUFACTURE OF SPANISH CIDER

Numeration, isolation and identification of the lactic acid bacteria found in Asturian cider during processing and storage has been carried out. The species Leuconostoc oenos was predominant during both alcoholic and malolactic fermentations. In most of the ciders studied the malolactic fermentation and the alcoholic fermentation started at the same time; production of D-lactate as well as a decrease of L-lactate were to be found once the malolactic fermentation was complete .     

WORT ENTEROBACTERIA AND OTHER MICROBIAL POPULATIONS INVOLVED DURING THE FIRST MONTH OF LAMBIC FERMENTATION

Lambic is a special type of Belgian beer obtained by spontaneous fermentation. The micro-organisms occurring during the first month of lambic fermentation were counted using several selective media. During this period a mixed flora is present in which the Enterobacteriaceae predominate. They were isolated from samples of lambic of different ages originating from different breweries and identified using the minitek identification system. Much variation may occur in types of Enterobacteriaceae: Klebsiella aerogenes, Enterobacter cloacae, E. aerogenes, Escherichia coli, Citrobacter freundii, A-D group bacteria and Hafnia alvei are found during the first month of fermentation. E. cloacae is found most frequently followed by K. aerogenes. After 30 to 40 days all Enterobacteria die off because of the activities of yeasts. Lactic acid and acetic acid bacteria are found in very low numbers during the first month of fermentation; Staphylococci and Streptococci are not present.     

Bacterial Community Dynamics of Salted and Fermented Shrimp based on Denaturing Gradient Gel Electrophoresis

The Korean traditional seafood jeotgal is consumed directly or as an additive in other foods to improve flavor or fermentation efficiency. Saeujot, made from salted and fermented tiny shrimp (SFS; Acetes japonicus), is the best‐selling jeotgal in Korea. In this study, we reveal the microbial diversity and dynamics in naturally fermented shrimp by denaturing gradient gel electrophoresis (DGGE). The population fingerprints of the predominant microbiota and its succession were generated by DGGE analysis of universal V3 16S rDNA polymerase chain reaction (PCR) amplicons. Overall, 17 strains were identified from sequencing of 30 DGGE bands. The DGGE profiles showed diverse bacterial populations in the sample, throughout the fermentation of SFS. Staphylococcus equorum, Halanaerobium saccharolyticum, Salimicrobium luteum, and Halomonas jeotgali were the dominant bacteria, and their levels steadily increased during the fermentation process. Certain other bacteria, such as Psychrobacter jeotgali and Halomonas alimentaria appeared during the early‐fermentation process, while Alkalibacterium putridalgicola, Tetragenococcus muriaticus, and Salinicoccus jeotgali appeared during the late‐fermentation process. The members of the order Bacillales were found to be predominant during the fermentation of SFS. Furthermore, S. equorum was identified as the dominant bacterial isolate by the traditional method of culturing under aerobic and facultative anaerobic conditions. We expect that this information will facilitate the design of autochthonous starter cultures for the production of SFS with desired characteristic sensory profiles and shorter ripening times.     

ETHANOL FERMENTATION OF VARIOUS RED RICES WITHOUT COOKING

With various red rices as raw material, fermentation without cooking was allowed to proceed at pH 4.0 and 30°C for 4 d for production of novel alcoholic beverages. Ethanol fermentation was achieved with various rice grains. In particular, uncooked wild-rice (Zizania aquatica) is reported now for the first time as being used for ethanol fermentation. The final concentration of ethanol achieved during fermentation was 8.8–9.3% (v/v). Rice wine made from aromatic red rice (Oryza sativa var. Indica, Tapol) had a wine-like red color. It was rich in a fruity aroma and had a characteristic sour taste, as revealed by organoleptic testing. Volatile substances formed have been analysed by gas chromatography. Rice wine made from black rice (Oryza sativa var. Indica) also had a wine-like red color and a sour taste. By contrast, rice wines made from red rice (Oryza sativa var. Japonica) and wild-rice were faintly yellow in color and had a sake-like and a somewhat green tea-like flavor, respectively. Although the red pigments of the aromatic red rice and black rice were solubilized during uncooked ethanol fermentation, the pigments of red rice and wild-rice were insoluble and not available as material for brewing of red-colored alcoholic beverages. Aromatic red rice appears to be a suitable material for the production of a wine-like alcoholic beverage.     

DETERMINATION OF ULTRASONIC‐BASED RHEOLOGICAL PROPERTIES OF DOUGH DURING FERMENTATION1

An ultrasonic technique was used to study the changes of the rheological properties of dough during fermentation at 37C and compared with the extensional properties of fermented dough obtained from tensile tests carried out in a Universal Testing Maching. The velocity and attenuation of a longitudinal wave (P‐wave) propagated through the dough samples were measured and analyzed to obtain the viscoelastic moduli of the dough; the storage modulus M' and the loss modulus M''. These moduli include both the bulk and the shear moduli. A wavelet analysis also was used to determine the effect of frequency on the ultrasonic‐based viscoelastic moduli and the effect of the fermentation process on the ultrasonic velocity dispersion. A decrease in ultrasonic velocity was observed with increasing fermentation times. Ultrasonic waves were strongly attenuated in the dough subjected to long fermentation times and fermentation had a large influence on the viscoelastic moduli of the dough. The ultrasonic velocity increased with increasing frequency, clearly showing the viscoelastic nature of the fermented dough. The analysis also showed significant ultrasonic velocity dispersion upon fermentation. Ultrasonic measurements yielded results that agreed with those obtained from conventional rheology commonly used to characterize the extensional properties of dough. Both tests clearly showed the loss of elasticity by the dough samples upon fermentation.     

Application of Plackett–Burman experimental design for investigating the effect of wort amino acids on flavour‐active compounds production during lager yeast fermentation

Aroma‐active higher alcohols and esters are produced intracellularly in the cytosol by fermenting lager yeast cells, which are of major industrial interest because they determine aroma and taste characteristics of the fermented beer. Wort amino acid composition and their utilization by yeast during brewer's wort fermentation influence both the yeast fermentation performance and the flavour profile of the finished product. To better understand the relationship between the yeast cell and wort amino acid composition, Plackett–Burman screening design was applied to measure the changes in nitrogen composition associated with yeast amino acids uptake and flavour formation during fermentation. Here, using an industrial lager brewing strain of Saccharomyces pastorianus , we investigated the effect of amino acid composition on the accumulation of higher alcohols and volatile esters. The objective of this study was to identify the significant amino acids involved in the flavour production during beer fermentation. Our results showed that even though different flavour substances were produced with different amino acid composition in the fermentation experiments, the discrepancies were not related to the total amount of amino acids in the synthetic medium. The most significant effect on higher alcohol production was exercised by the content of glutamic acid, aromatic amino acids and branch chain amino acids. Leucine, valine, glutamic acid, phenylalanine, serine and lysine were identified as important determinants for the formation of esters. The future applications of this information could drastically improve the current regime of selecting malt and adjunct or their formula with desired amino acids in wort. Copyright © 2017 The Institute of Brewing & Distilling