Different Stages Of Fermentation Research Articles

Impact of assimilable nitrogen availability in glucose uptake kinetics in Saccharomyces cerevisiae during alcoholic fermentation

BackgroundThe expression and activity of the different Saccharomyces cerevisiae hexose uptake systems (Hxt) and the kinetics of glucose uptake are considered essential to industrial alcoholic fermentation performance. However, the dynamics of glucose uptake kinetics during the different stages of fermentation, depending on glucose and nitrogen availability, is very poorly characterized. The objective of the present work was to examine thoroughly the alterations occurring in glucose uptake kinetics during alcoholic fermentation, by the wine strain S. cerevisiae PYCC 4072, of a synthetic grape juice basal medium with either a limiting or non-limiting initial nitrogen concentration and following nitrogen supplementation of the nitrogen-depleted sluggish fermentation.ResultsIndependently of the initial concentration of the nitrogen source, glucose transport capacity is maximal during the early stages of fermentation and presumably sustained by the low-affinity and high-capacity glucose transporter Hxt1p. During nitrogen-limited sluggish fermentation, glucose uptake capacity was reduced to approximately 20% of its initial values (Vmax = 4.9 ± 0.8 compared to 21.9 ± 1.2 μmol h-1 10-8 cells), being presumably sustained by the low-affinity glucose transporter Hxt3p (considering the calculated Km = 39.2 ± 8.6 mM). The supplementation of the sluggish fermentation broth with ammonium led to the increase of glucose transport capacity associated to the expression of different glucose uptake systems with low and high affinities for glucose (Km = 58.2 ± 9.1 and 2.7 ± 0.4 mM). A biclustering analysis carried out using microarray data, previously obtained for this yeast strain transcriptional response to equivalent fermentation conditions, indicates that the activation of the expression of genes encoding the glucose transporters Hxt2p (during the transition period to active fermentation) and Hxt3p, Hxt4p, Hxt6p and Hxt7p (during the period of active fermentation) may have a major role in the recovery of glucose uptake rate following ammonium supplementation. These results suggest a general derepression of the glucose-repressible HXT genes and are consistent with the downregulation of Mig1p and Rgt1p.ConclusionsAlthough reduced, glucose uptake rate during nitrogen-limited fermentation is not abrogated. Following ammonium supplementation, sluggish fermentation recovery is associated to the increase of glucose uptake capacity, related to the de novo synthesis of glucose transporters with different affinity for glucose and capacity, presumably of Hxt2p, Hxt3p, Hxt4p, Hxt6p and Hxt7p. This study is a contribution to the understanding of yeast response to different stages of alcoholic fermentation at the level of glucose uptake kinetics, in particular under nitrogen limitation or replenish, which is useful knowledge to guide fermentation practices.

Bacterial diversity in Philippine fermented mustard (burong mustasa) as revealed by 16S rRNA gene analysis

Previous studies on the bacterial profile of burong mustasa, a traditional Philippine fermented food, had been conducted using culture-dependent techniques. Since these methods may underestimate the total microbiota of a sample, a culture-independent study was done to determine the bacterial diversity in burong mustasa through molecular biology techniques. Bacterial DNA was isolated from fermented mustard samples at different stages of fermentation. The isolated genomic DNA was amplified by PCR using specific primers for the 16S ribosomal RNA gene (16S rDNA). The 1.5 kb amplicons obtained were subjected to nested PCR using primers for the internal variable region of the 16S rDNA. The 585 bp nested PCR amplicons were then subjected to denaturing gradient gel electrophoresis (DGGE) to separate the different bacteria present in each sample. Distinct and unique bands in the DGGE profile were excised, reamplified, purified and sequenced for bacterial identification. Molecular cloning of the 1.5 kb 16S rDNA was also performed using the pGEM-T Easy Vector System. The cloned gene was sequenced for bacterial identification. The identified microbiota in burong mustasa at different stages of fermentation include lactic acid bacteria and several uncultured bacteria (initial up to the final stages); acetic acid bacteria (middle stage); and Streptobacillus and Fusobacterium species (initial stage). The potential probiotic bacteria found in burong mustasa are Weissella and Lactobacillus.

Oral administration of Lactobacillus plantarum K68 ameliorates DSS-induced ulcerative colitis in BALB/c mice via the anti-inflammatory and immunomodulatory activities

Many different kinds of fermented food are consumed daily in Taiwan, such as stinky tofu, suan-tsai, and fu-tsai. We have previously reported the diversity of lactic acid bacteria (LAB) at different stages of fermentation in the production of suan-tsai and fu-tsai. In this study, the anti-inflammatory and immunomodulatory activities of Lactobacillus plantarum K68 (K68) isolated from fu-tsai were evaluated. K68 significantly inhibited the production of tumor necrosis factor-α (TNF-α) and prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-induced murine macrophage RAW 264.7 cells and stimulated interferon-γ (IFN-γ) production in human peripheral blood mononuclear cells (hPBMCs). Additionally, orally administered K68 ameliorated dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) in BALB/c mice. Both the disease activity index (DAI) and histological scores (HIS) showed that the severity of UC was significantly reduced by oral administration of K68. Furthermore, the production of pro inflammatory cytokines TNF-α, interleukin-1β (IL-1β), and interleukin-6 (IL-6) was significantly reduced in K68-administered group. Colonic mRNA expression levels of TNF-α, cyclooxygenase-2 (COX-2), forkhead box P3 (Foxp3), suppressors of cytokine signaling 3 (SOCS3), and toll like receptor 4 (TLR4), were also reduced in the K68-administered group. These results suggest that K68 exhibits anti-inflammatory and immunomodulatory activities that ameliorate DSS-induced experimental colitis.

Detection of abnormal fermentations in wine process by multivariate statistics and pattern recognition techniques

Three multivariate statistical techniques (Multiway Principal Component Analysis, Multiway Partial Least Squares, and Stepwise Linear Discriminant Analysis) and one artificial intelligence method (Artificial Neural Networks) were evaluated to detect and predict early abnormal behaviors of wine fermentations. The techniques were tested with data of thirty-two variables at different stages of fermentation from industrial wine fermentations of Cabernet Sauvignon. All the techniques studied considered a pre-treatment to obtain a homogeneous space and reduce the overfitting. The results were encouraging; it was possible to classify at 72h 100% of the fermentation correctly with three variables using Multiway Partial Least Squares and Artificial Neural Networks. Additional and complementary results were obtained with Stepwise Linear Discriminant Analysis, which found that ethanol, sugars and density measurements are able to discriminate abnormal behavior.

Survival of Escherichia coli O157:H7 in Cucumber Fermentation Brines

Bacterial pathogens have been reported on fresh cucumbers and other vegetables used for commercial fermentation. The Food and Drug Administration currently has a 5-log reduction standard for E. coli O157:H7 and other vegetative pathogens in acidified pickle products. For fermented vegetables, which are acid foods, there is little data documenting the conditions needed to kill acid resistant pathogens. To address this knowledge gap, we obtained 10 different cucumber fermentation brines at different stages of fermentation from 5 domestic commercial plants. Cucumber brines were used to represent vegetable fermentations because cabbage and other vegetables may have inhibitory compounds that may affect survival. The 5-log reduction times for E. coli O157:H7 strains in the commercial brines were found to be positively correlated with brine pH, and ranged from 3 to 24 d for pH values of 3.2 to 4.6, respectively. In a laboratory cucumber juice medium that had been previously fermented with Lactobacillus plantarum or Leuconostoc mesenteroides (pH 3.9), a 5-log reduction was achieved within 1 to 16 d depending on pH, acid concentration, and temperature. During competitive growth at 30 °C in the presence of L. plantarum or L. mesenteroides in cucumber juice, E. coli O157:H7 cell numbers were reduced to below the level of detection within 2 to 3 d. These data may be used to aid manufacturers of fermented vegetable products determine safe production practices based on fermentation pH and temperature. Disease causing strains of the bacterium E. coli may be present on fresh vegetables. Our investigation determined the time needed to kill E. coli in cucumber fermentation brines and how E. coli strains are killed in competition with naturally present lactic acid bacteria. Our results showed how brine pH and other brine conditions affected the killing of E. coli strains. These data can be used by producers of fermented vegetable products to help assure consumer safety.

Intraspecific Diversity of Yeast Associated to Vitis vinifera Albariño Must from Different Vineyard Ecosystems

The genetic diversity of Saccharomyces cerevisiae associated to the must (Vitis vinifera Albarino) from different geographic areas and spontaneous fermentation was studied by analysis of mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP). Colonies were isolated in musts at different stages of fermentation from three different geographic areas, O Val do Salnes, O Rosal and O Condado do Tea from Galicia (NW Spain). A total of 17 different patterns out of 273 S. cerevisiae colonies were identified using mitochondrial DNA restriction analysis. Pattern I from O Rosal, VI from O Val do Salnes and XII from O Condado do Tea were the predominant ones. Only pattern II was common to all fermentations. This molecular study provides useful information about distribution and genetic relationships among several S. cerevisiae strains associated to Vitis vinifera Albarino from different vineyard ecosystems and their influence on certain oenological properties.

Risk assessment for the occurrence of <i>Escherichia coli</i> 0157:H7 in indigenous fermented milk (Lee Naga a Agbora) produced in Uganda

Different types of indigenous fermented milks are widely consumed in pastoralist communities of Africa, including Uganda. Local processing of milk generally takes place under uncontrolled hygienic conditions. Lee naga a agbora, a locally fermented milk product in northern Uganda, was evaluated for microbial contamination. The changes in pH, Total Plate Counts (TPC), Total Coliforms (TC), yeasts and moulds, and Escherichia coli 0157:H7, particularly its survival during different stages of fermentation were studied. The pH was found to decrease with fermentation time (6.54 to 4.54). E. coli was detected in most samples of Lee naga a agbora and its occurrence was observed throughout the fermentation period. The mean value of TPC (6.60± 0.10 log cfu ml-1) at the start of fermentation was significantly different (p0.05) between 48 and 72 hours. The rate of decrease in TC counts in the initial fermentation period (0 to 48 hours) was lower than the later stages of fermentation (48 to 144 hours). Yeast and mould counts increased with fermentation time (6.20 to 7.42 log cfu ml-1). On the whole, there is risk associated with consumption of Lee naga a agbora given the occurrence of E. Coli.

Genetic and phenotypic diversity of autochthonous cider yeasts in a cellar from Asturias

This paper analyses yeast diversity and dynamics during the production of Asturian cider. Yeasts were isolated from apple juice and at different stages of fermentation in a cellar in Villaviciosa during two Asturian cider-apple harvests. The species identified by ITS-RFLP corresponded to Hanseniaspora valbyensis, Hanseniaspora uvarum, Metschnikowia pulcherrima, Pichia guilliermondii, Candida parapsilosis, Saccharomyces cerevisiae and Saccharomyces bayanus/Saccharomyces pastorianus/Saccharomyces kudriavzevii/Saccharomyces mikatae. The species C. parapsilosis is reported here for the first time in cider. The analysis of Saccharomyces mtDNA patterns showed great diversity, sequential substitution and the presence of a small number of yeast patterns (up to 8), present in both harvests. Killer (patterns nos. 22′ and 47), sensitive (patterns nos. 12, 15, 33 and 61) and neutral phenotypes were found among the S. cerevisiae isolates. The detection of β-glucosidase activity, with arbutin as the sole carbon source, allowed two S. cerevisiae strains (patterns nos. 3′ and 19′) to be differentiated by means of this enzymatic activity. Yeast strains producing the killer toxin or with β-glucosidase activity are reported for the first time in autochthonous cider yeasts.

Diversity of lactic acid bacteria in suan-tsai and fu-tsai, traditional fermented mustard products of Taiwan

Fu-tsai and suan-tsai are spontaneously fermented mustard products traditionally prepared by the Hakka tribe of Taiwan. We chose 5 different processing stages of these products for analysis of the microbial community of lactic acid bacteria (LAB) by 16S rRNA gene sequencing. From 500 LAB isolates we identified 119 representative strains belonging to 5 genera and 18 species, including Enterococcus (1 species), Lactobacillus (11 species), Leuconostoc (3 species), Pediococcus (1 species), and Weissella (2 species). The LAB composition of mustard fermented for 3 days, known as the Mu sample, was the most diverse, with 11 different LAB species being isolated. We used sequence analysis of the 16S rRNA gene to identify the LAB strains and analysis of the dnaA, pheS, and rpoA genes to identify 13 LAB strains for which identification by 16S rRNA gene sequences was not possible. These 13 strains were found to belong to 5 validated known species: Lactobacillus farciminis, Leuconostoc mesenteroides, Leuconostoc pseudomesenteroides, Weissella cibaria, and Weissella paramesenteroides, and 5 possibly novel Lactobacillus species. These results revealed that there is a high level of diversity in LAB at the different stages of fermentation in the production of suan-tsai and fu-tsai.

Ecological study of wine yeast in inoculated vats from La Mancha region

Use of certain selected commercial yeast cultures has become widespread throughout the wine-making sector, but there was not any study about their effectiveness in Castilla-La Mancha region. This study sought to determine, whether the commercial yeast successfully displaced native yeasts during the fermentation process, or whether it was replaced by them. A total of 41 vats containing different grape varieties were sampled at different stages of fermentation, selecting 1640 isolated yeasts, which were characterized using molecular biology techniques (RFLP mtDNA). From results obtained, it can be concluded that inoculation of fermentation tanks with active dry yeasts does not guarantee their implantation during alcoholic fermentation. Indeed, these added strains may be scarce or even non-existent during the process.

Variability of resveratrol (3, 5, 4′-trihydroxystilbene) content in relation to the fermentation processes by Saccharomyces cerevisiae strains

The variability of the four monomeric forms of resveratrol (3, 5, 4′-trihydroxystilbene) caused by 14 strains of fermentative yeasts belonging to the Saccharomyces cerevisiae species in Cannonau grape musts during three different stages of fermentation was assessed by HPLC. During the process a decrease in the glucoside forms (cis and trans-β-D-glucopyranoside) was observed in all the musts independently of the yeast strain used. As regards the activity of the free forms (cis and trans-resveratrol), the greatest differences were found in the first two stages of fermentation when some strains appeared to cause an increase in either one or both of the free forms while others produced a decrease in the two compounds.

Note. Histamine production by some lactic acid bacteria isolated from ciders / Nota. Producción de histamina por algunas bacterias lácticas aisladas a partir de sidras

Histidine decarboxylase activity has been investigated in 23 strains of lactic acid bacteria, belonging to Oenococcus oeni, Lactobacillus brevis and Lactobacillus spp., isolated during the cidermaking process at different stages of fermentation. After 14 days at 25 °C on a semisynthetic medium supplemented with L-histidine, 13 strains were histamine producers and six of these strains were selected to study the kinetics of growth and histamine production at different temperatures. The results showed that histamine accumulation was maximal at 25 °C after 10-15 days of growth. Comparison of growth and histamine level curves demonstrated a delay time of eight days between maximal growth and the highest histamine content of the cultures.

Effect of pH on physiology of Metarhizium anisopliae for production of swainsonine

The effect of pH on the production of swainsonine and fungal morphology at different stages of fermentation of Metarhizium anisopliae was investigated. When no control was applied, the pH of the culture dropped from 6.5 to 3.8 within the first 72 hours and the concentration of swainsonine reached 43.3 mg l −1. When the pH was held constant either at the beginning or throughout the fermentation, the maximum recorded swainsonine level was only 8.4 mg l −1 corresponding with an increase in the formation of pellets. A late pH control applied after 72 hours, resulted in a swainsonine titer of 45.5 mg l −1.

Effect of pH on physiology of Metarhizium anisopliae for production of swainsonine.

The effect of pH on the production of swainsonine and fungal morphology at different stages of fermentation of Metarhizium anisopliae was investigated. When no control was applied, the pH of the culture dropped from 6.5 to 3.8 within the first 72 hours and the concentration of swainsonine reached 43.3 mg 1(-1). When the pH was held constant either at the beginning or throughout the fermentation, the maximum recorded swainsonine level was only 8.4 mg 1(-1) corresponding with an increase in the formation of pellets. A late pH control applied after 72 hours, resulted in a swainsonine titer of 45.5 mg 1(-1).

Multiple strains of Saccharomyces cerevisiae on a single grape vine.

On the basis of the levels of secondary product formation four different phenotypes were represented among the 28 strains of Saccharomyces cerevisiae isolated during the spontaneous fermentation of grape juice. The genetic analysis indicated that four different strains, representing each phenotypic class, were derived, one from the other, by mutation. The spontaneous fermentation of a Malvasia must was dominated by different strains of Saccharomyces cerevisiae at different stages of fermentation.

Modeling of heat and mass transfer for solid state fermentation process in tray bioreactor

A mathematical model is developed to simulate oxygen consumption, heat generation and cell growth in solid state fermentation (SSF). The fungal growth on the solid substrate particles results in the increase of the cell film thickness around the particles. The model incorporates this increase in the biofilm size which leads to decrease in the porosity of the substrate bed and diffusivity of oxygen in the bed. The model also takes into account the effect of steric hindrance limitations in SSF. The growth of cells around single particle and resulting expansion of biofilm around the particle is analyzed for simplified zero and first order oxygen consumption kinetics. Under conditions of zero order kinetics, the model predicts upper limit on cell density. The model simulations for packed bed of solid particles in tray bioreactor show distinct limitations on growth due to simultaneous heat and mass transport phenomena accompanying solid state fermentation process. The extent of limitation due to heat and/or mass transport phenomena is analyzed during different stages of fermentation. It is expected that the model will lead to better understanding of the transport processes in SSF, and therefore, will assist in optimal design of bioreactors for SSF.

Intracellular Nicotinamide Adenine Dinucleotide Content of Brewer's Yeast during Different Stages of Fermentation

A method for extraction and estimation of nicotinamide adenine dinucleotide (NAD) and reduced NAD (NADH) in yeast during different stages of fermentation was developed. Yeast cells were washed in n...

Intracellular Nicotinamide Adenine Dinucleotide Content of Brewer�s Yeast During Different Stages of Fermentation

Intracellular Nicotinamide Adenine Dinucleotide Content of Brewer�s Yeast During Different Stages of Fermentation

Intracellular accumulation of AMP as a cause for the decline in rate of ethanol production by Saccharomyces cerevisiae during batch fermentation.

A general hypothesis is presented for the decline in the rate of ethanol production (per unit of cell protein) during batch fermentation. Inhibition of ethanol production is proposed to result from the intracellular accumulation of AMP during the transition from growth to the stationary phase. AMP acts as a competitive inhibitor of hexokinase with respect to ATP. When assayed in vitro in the presence of ATP and AMP concentrations equivalent to those within cells at different stages of fermentation, hexokinase activity declined in parallel with the in vivo decline in the rate of ethanol production. The coupling of glycolytic flux and fermentation to cell growth via degradation products of RNA may be of evolutionary advantage for Saccharomyces cerevisiae. Such a coupling would reduce the exposure of nongrowing cells to potentially harmful concentrations of waste products from metabolism and would conserve nutrients for future growth under more favorable conditions.