Microbial Fermentation Process Research Articles

Environmental pollutant-mediated disruption of gut microbial metabolism of the prebiotic inulin

Exposure to environmental pollutants is associated with a greater risk for metabolic diseases including cardiovascular disease. Pollutant exposure can also alter gut microbial populations that may contribute to metabolic effects and progression of inflammatory diseases. Short-chain fatty acids (SCFAs), produced from gut fermentation of dietary carbohydrates, such as inulin, exert numerous effects on host energy metabolism and are linked to a reduced risk of diseases. The hypothesis was that exposure to dioxin-like pollutants modulate gut microbial viability and/or fermentation processes. An inulin-utilizing isolate was collected from murine feces, characterized and used in subsequent experiments. Exposure to polychlorinated biphenyl, PCB 126 impeded bacterial viability of the isolate at concentrations of 20 and 200 μM. PCB 126 exposure also resulted in a significant loss of intracellular potassium following exposure, indicating cell membrane disruption of the isolate. Furthermore, total fecal microbe samples from mice were harvested, resuspended and incubated for 24 h in anaerobic media containing inulin with or without PCB 126. HPLC analysis of supernatants revealed that PCB 126 exposure reduced succinic acid production, but increased propionate production, both of which can influence host glucose and lipid metabolism. Overall, the presented evidence supports the idea that pollutant exposure may contribute to alterations in host metabolism through gut microbiota-dependent mechanisms, specifically through bacterial fermentation processes or membrane disruption.

Analytical Enzymatic Saccharification of Lignocellulosic Biomass for Conversion to Biofuels and Bio-Based Chemicals

Lignocellulosic feedstocks are an important resource for biorefining of renewables to bio-based fuels, chemicals, and materials. Relevant feedstocks include energy crops, residues from agriculture and forestry, and agro-industrial and forest-industrial residues. The feedstocks differ with respect to their recalcitrance to bioconversion through pretreatment and enzymatic saccharification, which will produce sugars that can be further converted to advanced biofuels and other products through microbial fermentation processes. In analytical enzymatic saccharification, the susceptibility of lignocellulosic samples to pretreatment and enzymatic saccharification is assessed in analytical scale using high-throughput or semi-automated techniques. This type of analysis is particularly relevant for screening of large collections of natural or transgenic varieties of plants that are dedicated to production of biofuels or other bio-based chemicals. In combination with studies of plant physiology and cell wall chemistry, analytical enzymatic saccharification can provide information about the fundamental reasons behind lignocellulose recalcitrance as well as about the potential of collections of plants or different fractions of plants for industrial biorefining. This review is focused on techniques used by researchers for screening the susceptibility of plants to pretreatment and enzymatic saccharification, and advantages and disadvantages that are associated with different approaches.

FUNGAL CITRIC ACID PRODUCTION USING WASTE MATERIALS: A MINI-REVIEW

Citric acid is one of the extensively used organic acids in many industries. There is a worldwide demand for citric acid consumption due to their many industrial applications. It is also considered as a Generally Recognized As Safe (GRAS) compound. In this review, current developments in microbial fermentation processes for citric acid production have been discussed. In food, beverages, milk and sugar processing several biodegradable organic waste materials are generated in large quantities. These include sugarcane bagasse, grape pomace, apple pomace, pineapple pomace, vegetables, tapioca, coconut husk, banana peels, citrus peels, whey and decaying fruits are found to be potential substrates for citric acid production. A significant effect of substrate concentration, inorganic salts, initial sugar concentration, moisture and additives etc on citric acid production have been highlighted for further improvement in fermentation process. Recent developments in upstream and downstream processes for citric acid production are also deeply discussed. This review gives insights for future possibilities of cost effective fermentation process for citric acid production from several cheap raw materials. Formation of citric acid as an industrial byproduct will help to tackle waste disposal issue and also reduce the dependency of industry over other citric acid producers. Thus, the industry would be benefitted ecologically and economically.

Barriers and opportunities in bio-based production of hydrocarbons

Global climate change caused by the accumulation of greenhouse gases (GHGs) has caused concerns regarding the continued reliance on fossil fuels as our primary energy source. Hydrocarbons produced from biomass using microbial fermentation processes can serve as high-quality liquid transportation fuels and may contribute to a reduction in GHG emissions. Here, we discuss the barriers and opportunities for bio-based production of hydrocarbons to be used as diesel and jet fuels and review recent advances in engineering microbes for production of these chemicals. There are two main challenges associated with establishing bio-based hydrocarbon production from cheap feedstocks; lowering the cost of developing efficient and robust microbial cell factories and establishing more efficient routes for biomass hydrolysis to sugars for fermentation. We discuss how to develop novel systems and synthetic biology tools that can enable faster and cheaper construction of microbial cell factories and thereby address the first challenge, as well as recent advances in biomass processing that will likely lead to overcoming the second challenge in the near future.

Optimal control of nonlinear switched system in an uncoupled microbial fed-batch fermentation process

The bio-dissimilation of glycerol to 1,3-propanediol (1,3-PD) is a complex bioprocess due to the multiple inhibitions of substrate and products onto the cell growth. In consideration of both the inhibition mechanisms of 3-hydroxypropionaldehyde (3-HPA) and the transport modes of glycerol and 1,3-PD across the cell membrane, we establish a novel switched system which is represented by a ten-dimensional nonlinear dynamical equation containing both extracellular and intracellular environments. The uncoupled microbial fed-batch fermentation process are modeled using the switched system which the glycerol and alkali are respectively poured into. Taking the feeding rates of glycerol and alkali, the switching times and the mode sequence as the control variables, an optimal control model is proposed with the concentration of the terminal time 1,3-PD as performance index. In order to maximize the yield of 1,3-PD, the control parameterization technique and the exact penalty function method are used to solve the considered problem. Numerical results show that under the obtained optimal feeding rates of glycerol and alkali, switching times and mode sequence, the productivity of 1,3-PD at the terminal time is increased significantly compared with previous results.

Necrotic cell death induced by dithianon on Saccharomyces cerevisiae

Dithianon is a broad-spectrum anthraquinone fungicide used to control several diseases of grapes, apples, and other fruits and vegetables. Its mode of action is described as multi-site and associated to thiol-reactivity. As other fungicides can affect non-phytopathogenic organisms as yeasts and bacteria, with impact on microbial population, diversity, and fermentation processes. In this context, we study the effect of dithianon on the model organism and fermentative yeast Saccharomyces cerevisiae in order to elucidate the mechanisms involved in yeast cell death., and explain its interference on wine fermentation kinetics. Thus for, we analyzed cellular protein and non-protein thiols, membrane and cell wall integrity, reactive oxygen species accumulation, mitochondrial membrane potential, and phosphatidylserine externalization. The results showed that when exponentially aerobic growing cells of S. cerevisiae are submitted to acute dithianon treatment they loss cell wall and membrane integrity, dying by necrosis, and this behavior is associated to a depletion of reduced proteic and non-proteic thiol groups. We also detected an important increase of cellular reactive oxygen species (ROS) associated to mitochondrial membrane potential modifications on dithianon treated cells. ROS accumulation was not associated to apoptotic cell death, but can be responsible for intracellular damages. Moreover, necrotic cell death induced by dithianon explains its effect on the kinetics of wine fermentations.

Simultaneous biosynthesis of (R)-acetoin and ethylene glycol from D-xylose through in vitro metabolic engineering

(R)-acetoin is a four-carbon platform compound used as the precursor for synthesizing novel optically active materials. Ethylene glycol (EG) is a large-volume two-carbon commodity chemical used as the anti-freezing agent and building-block molecule for various polymers. Currently established microbial fermentation processes for converting monosaccharides to either (R)-acetoin or EG are plagued by the formation of undesirable by-products. We show here that a cell-free bioreaction scheme can generate enantiomerically pure acetoin and EG as co-products from biomass-derived D-xylose. The seven-step, ATP-free system included in situ cofactor regeneration and recruited enzymes from Escherichia coli W3110, Bacillus subtilis shaijiu 32 and Caulobacter crescentus CB 2. Optimized in vitro biocatalytic conditions generated 3.2 mM (R)-acetoin with stereoisomeric purity of 99.5% from 10 mM D-xylose at 30 °C and pH 7.5 after 24 h, with an initial (R)-acetoin productivity of 1.0 mM/h. Concomitantly, EG was produced at 5.5 mM, with an initial productivity of 1.7 mM/h. This in vitro biocatalytic platform illustrates the potential for production of multiple value-added biomolecules from biomass-based sugars with no ATP requirement.

On the biochemical conversion technology of edible fungi cultivation waste based on microbial fermentation process

Considering the biochemical conversion parameters of cultivation waste, the secondary development of product substrates and the standardized assessment after conversion of culture substrate waste, this paper takes the cultivation waste of edible fungi as the test materials and studies the microbial conversion of such waste, and then applies the converted culture substrates in the culture of cucumber seedlings. The results show that, when the cultivation waste substrate was treated through microbial fermentation, the temperature inside the substrate changed periodically. The rapeseed cake could promote the growth of the fermenting bacteria and effectively increase their metabolism; and the mixed pearlite in the waste had a positive effect in improving the porosity and water absorption of the culture substrate. The pH value decreased during the initial microbial transformation, because large amounts of organic acids were produced through the metabolism of aerobic bacteria, and then, when the temperature rose, the thermophilic microorganisms began to decompose organic acids, resulting in the continuous rise of the pH value. The higher content of substrate there was in the cultivation waste, the smaller the germination rate would be and the lower the healthy index would be, indicating that the waste substrate contained substances that inhibited seed germination. When the seedlings became adapted to the environment of the cultivation waste substrate, the nutrients in the substrate would facilitate the late-stage growth and development of seedlings.

Preparation of fish waste silage and its effect on the growth performance and meat quality of broiler chickens.

This experiment was conducted to evaluate the effects of replacing soybean meal with fish-waste silage (FWS) in diets on growth performance and meat quality in broiler chickens. Fish waste (FW) was fermented with molasses and Lactobacillus plantarum and Aspergillus oryzae for 15 days. A total of 240 day-old male Cobb 500 broiler chickens were used in a 42-day experiment in which the birds were randomly allotted to one of five dietary treatments (containing 0, 30, 60, 90, and 120 gkg-1 FWS) in a completely randomized design. Results indicated that the fermentation process significantly reduced pH and ether extract, but it increased crude protein content and lactic acid bacteria count in FW. The use of FWS significantly improved daily weight gain and feed conversion ratio of broilers. The amount of thiobarbiturate-reducing substances and lipid content in chicken's meat fed diet containing FWS was significantly lower than in the group fed a control diet. In conclusion, the microbial fermentation process is effective in improving nutritive values of FW. Feeding up to 120 gkg-1 FWS also improves the performance and meat quality of birds and can be used as a suitable protein source in broiler chickens' diet. © 2018 Society of Chemical Industry.

Optimization of kojic acid production conditions from cane molasses using Plackett-Burman design

Fungal synthesis of kojic acid has gained more interest in these days as an alternative way to chemical synthetic. The aspect of the microbial fermentation process is to develop a suitable culture medium to obtain the maximum amount of kojic acid using statistical methods. In this study; different selected three isolates of Aspergillus flavus (No 1, 2 and 3) were screened for their ability to produced kojic acid and the isolate No 3 was the highest kojic acid producer one. The capability of A. flavus No 3 to produce kojic acid was improved using Plackett-Burman design. From ten different agro-industrial wastes cane molasses recorded the highest kojic acid productivity with 2.24 g/l-1 day-1 and was the most effective parameter plays a crucial role in Plackett-Burman design. Maximum kojic acid production (24.65 g/l) by A. flavus (No. 3) obtained under the fermentation conditions: incubation temperature at 25oC, incubation time 9 days, pH 3, inoculum size 0.5%, shaking rate at 150 rpm and medium constituents: Cane molasses 60 g/l, yeast extract 7 g/l, KH2PO4 2 g/l, ZnSO4·7H2O 100 µg/l and MgSO4·7H2O 1 g/l with regression analysis (R2) 99.45% and 2.33-fold increase in comparison to the production of the original level (10.6 g/l). DOI: http://dx.doi.org/10.5281/zenodo.1211517

Biochemical activities of lactose-derived prebiotics — a review

In the dairy industry different types of prebiotics, such galacto-oligosaccharide, lactulose, lactosucrose, tagatose, lactitol, lactobiono- and glucono-δ-lactone are synthesized through different chemical and biochemical reactions (hydrolysis, transgalactosylation, isomerization, fructosyl-transfer, reduction, and oxidation) as well as microbial fermentation processes using raw whey or isolated lactose as feedstock. Lactose-derived prebiotics have several functional and nutritional values. The biochemical activities of lactose-based prebiotics are expressed in the presence of probiotics (lactic acid bacteria, yeasts, Bacillus spp.). Galacto-oligosaccharide and lactosucrose reduce the risk of bowel disorder (diarrhea), inflammatory bowel disease (ulcerative colitis and crohn’s disease), and colon cancer. Galacto-oligosaccharide helps colonic absorption of minerals (iron, magnesium and calcium) and prevents osteoporosis. Lactulose, galacto-oligosaccharide, and lactitol promote laxative activity. Furthermore, lactulose may decrease the risk of hepatic encephalopathy. Prebiotics have low calorific value and glycemic index. Galacto-oligosaccharide and tagatose reduce the risks of hyperglycemia (Type 2 diabetes) and low density lipid (lipoprotein) accumulation in blood stream. Moreover, prebiotics improve immunomodulation, which reduces the risk of respiratory infection and allergies. This review describes unique biochemical mechanisms of several types of lactose-derived prebiotics.

High Substrate Uptake Rates Empower Vibrio natriegens as Production Host for Industrial Biotechnology.

The productivity of industrial fermentation processes is essentially limited by the biomass-specific substrate consumption rate (qS ) of the applied microbial production system. Since qS depends on the growth rate (μ), we highlight the potential of the fastest-growing nonpathogenic bacterium, Vibrio natriegens, as a novel candidate for future biotechnological processes. V. natriegens grows rapidly in BHIN complex medium with a μ of up to 4.43 h-1 (doubling time of 9.4 min) as well as in minimal medium supplemented with various industrially relevant substrates. Bioreactor cultivations in minimal medium with glucose showed that V. natriegens possesses an exceptionally high qS under aerobic (3.90 ± 0.08 g g-1 h-1) and anaerobic (7.81 ± 0.71 g g-1 h-1) conditions. Fermentations with resting cells of genetically engineered V. natriegens under anaerobic conditions yielded an overall volumetric productivity of 0.56 ± 0.10 g alanine liter-1 min-1 (i.e., 34 g liter-1 h-1). These inherent properties render V. natriegens a promising new microbial platform for future industrial fermentation processes operating with high productivity.IMPORTANCE Low conversion rates are one major challenge to realizing microbial fermentation processes for the production of commodities operating competitively with existing petrochemical approaches. For this reason, we screened for a novel platform organism possessing characteristics superior to those of traditionally employed microbial systems. We identified the fast-growing V. natriegens, which exhibits a versatile metabolism and shows striking growth and conversion rates, as a solid candidate to reach outstanding productivities. Due to these inherent characteristics, V. natriegens can speed up common laboratory routines, is suitable for already existing production procedures, and forms an excellent foundation for engineering next-generation bioprocesses.

Effects of soybean meal replacement with fermented soybean meal on growth, serum biochemistry and morphological condition of liver and distal intestine of Florida pompanoTrachinotus carolinus

This study evaluated the suitability of commercially produced fermented soybean meal (FSBM) known as PepSoyGen™, in a plant-based diet for Florida pompano, Trachinotus carolinus fingerlings. An 8-week growth trial was conducted to evaluate the effect of four isonitrogenous and isolipidic diets containing 0, 206, 309 and 410 g/kg FSBM, replacing approximately 0%, 50%, 75% and 100% SBM (designated as Basal, FSBM 50, FSBM 75 and FSBM 100, respectively) on growth performance, body composition, serum biochemistry and morphological condition of liver and distal intestine of Florida pompano. There were no significant differences in final mean weight, percentage weight gain, thermal unit growth coefficient and feed conversion ratio in all treatments. For serum biochemistry analysis, there were no significant differences in total protein, albumin, glucose, cholesterol, bile acids, plasma alkaline phosphatase, alanine aminotransferase and aspartate aminotransferase activities in all treatments. Cellular infiltration, presence of goblet cells and the width of lamina propria in the distal intestine were partly improved in fish fed FSBM 75 and 100 compared to the fish fed FSBM 50 and basal diet. Glycogen granulation, inflammation and nuclear change condition in the liver of pompano were better as the inclusion of fermented product increased. Results of this study indicate that FSBM can be utilized as an alternative protein source and microbial fermentation process could improve the functional properties of SBM.

Purification of biomevalonate from fermentation broth and conversion of biomevalonate into biomevalonolactone

Mevalonate (MVA) is a key compound of living organisms including bacteria, plants, and humans. MVA and mevalonolactone (MVL), a lactonized form of MVA, are important for pharmaceutical, cosmeceutical, and biotechnological applications. Although (R, S)-MVA with 50% enantiomeric purity is mainly produced by chemical synthesis, recently, microbial fermentation processes for MVA production have been considered as an alternative to the chemical synthesis because of high enantiomeric purity [(R)-MVA] and high titer. In the present study, bio-MVA produced by a fermentative process was decolorized by a charcoal-based method and then chemically transformed into bio-MVL without byproducts by means of phosphoric acid as an acid catalyst. The final bio-MVL was (R)-MVL with over 99% enantiomeric purity according to 1H NMR analysis.

Necrotic and apoptotic cell death induced by Captan on Saccharomyces cerevisiae

Captan is one of the most widely used broad-spectrum fungicide applied to control several early and late diseases of grapes, apples, and other fruits and vegetables, and as other phthalimide fungicides is defined as a multisite compound with thiol-reactivity. Captan can affect non-target organisms as yeasts, modifying microbial populations and fermentation processes. In this study, we asked whether Captan thiol-reactivity and other mechanisms are involved in acute Captan-induced cell death on aerobic growing Saccharomyces cerevisiae. Thus for, we analyze cellular protein and non-protein thiols, cell membrane integrity, reactive oxygen species accumulation, phosphatidylserine externalization, and apoptotic mutants behavior. The results showed that when submitted to acute Captan treatment most cells lost their membrane integrity and died by necrosis due to Captan reaction with thiols. However, part of the cells, even maintaining their membrane integrity, lost their culture ability. These cells showed an apoptotic behavior that may be the result of non-protein thiol depletion and consequent increase of reactive oxygen species (ROS). ROS accumulation triggers a metacaspase-dependent apoptotic cascade, as shown by the higher viability of the yca1-deleted mutant. Together, necrosis and apoptosis are responsible for the high mortality detected after acute Captan treatment of aerobically growing cells of S. cerevisiae.

Johnson Matthey teams with Intrexon

Johnson Matthey and and Intrexon, a Budapest-based biopharmaceutical firm, will jointly develop microbial strains for the production of peptide-based active pharmaceutical ingredients (APIs) via fermentation. Intrexon will contribute its proprietary microbial hosts and fermentation processes to the initial fermentation, isolation, and purification of APIs. Johnson Matthey will be responsible for scale-up, formulation, and commercialization of microbial strains.

Gene modification of the acetate biosynthesis pathway in Escherichia coli and implementation of the cell recycling technology to increase L-tryptophan production.

The implementation of a novel cell recycling technology based on a special disk centrifuge during microbial fermentation process can continuously separate the product and harmful intermediates, while maintaining the cell viability owing to the installed cooling system. Acetate accumulation is an often encountered problem in L-tryptophan fermentation by Escherichia coli. To extend our previous studies, the current study deleted the key genes underlying acetate biosynthesis to improve l-tryptophan production. The deletion of the phosphotransacetylase (pta)–acetate kinase (ackA) pathway in a gltB (encoding glutamate synthase) mutant of E. coli TRTHB, led to the highest production of l-tryptophan (47.18 g/L) and glucose conversion rate (17.83%), with a marked reduction in acetate accumulation (1.22 g/L). This strain, TRTHBPA, was then used to investigate the effects of the cell recycling process on L-tryptophan fermentation. Four different strategies were developed concerning two issues, the volume ratio of the concentrated cell solution and clear solution and the cell recycling period. With strategy I (concentrated cell solution: clear solution, 1: 1; cell recycling within 24–30 h), L-tryptophan production and the glucose conversion rate increased to 55.12 g/L and 19.75%, respectively, 17.55% and 10.77% higher than those without the cell recycling. In addition, the biomass increased by 13.52% and the fermentation period was shortened from 40 h to 32 h. These results indicated that the cell recycling technology significantly improved L-tryptophan production by E. coli.

Laccases as a Potential Tool for the Efficient Conversion of Lignocellulosic Biomass: A Review

The continuous increase in the world energy and chemicals demand requires the development of sustainable alternatives to non-renewable sources of energy. Biomass facilities and biorefineries represent interesting options to gradually replace the present industry based on fossil fuels. Lignocellulose is the most promising feedstock to be used in biorefineries. From a sugar platform perspective, a wide range of fuels and chemicals can be obtained via microbial fermentation processes, being ethanol the most significant lignocellulose-derived fuel. Before fermentation, lignocellulose must be pretreated to overcome its inherent recalcitrant structure and obtain the fermentable sugars. Usually, harsh conditions are required for pretreatment of lignocellulose, producing biomass degradation and releasing different compounds that are inhibitors of the hydrolytic enzymes and fermenting microorganisms. Moreover, the lignin polymer that remains in pretreated materials also affects biomass conversion by limiting the enzymatic hydrolysis. The use of laccases has been considered as a very powerful tool for delignification and detoxification of pretreated lignocellulosic materials, boosting subsequent saccharification and fermentation processes. This review compiles the latest studies about the application of laccases as useful and environmentally friendly delignification and detoxification technology, highlighting the main challenges and possible ways to make possible the integration of these enzymes in future lignocellulose-based industries.

A Sequential Steam Explosion and Reactive Extrusion Pretreatment for Lignocellulosic Biomass Conversion within a Fermentation-Based Biorefinery Perspective

The present work evaluates a two-step pretreatment process based on steam explosion and extrusion technologies for the optimal fractionation of lignocellulosic biomass. Two-step pretreatment of barley straw resulted in overall glucan, hemicellulose and lignin recovery yields of 84%, 91% and 87%, respectively. Precipitation of the collected lignin-rich liquid fraction yielded a solid residue with high lignin content, offering possibilities for subsequent applications. Moreover, hydrolysability tests showed almost complete saccharification of the pretreated solid residue, which when combined with the low concentration of the generated inhibitory compounds, is representative of a good pretreatment approach. Scheffersomyces stipitis was capable of fermenting all of the glucose and xylose from the non-diluted hemicellulose fraction, resulting in an ethanol concentration of 17.5 g/L with 0.34 g/g yields. Similarly, Saccharomyces cerevisiae produced about 4% (v/v) ethanol concentration with 0.40 g/g yields, during simultaneous saccharification and fermentation (SSF) of the two-step pretreated solid residue at 10% (w/w) consistency. These results increased the overall conversion yields from a one-step steam explosion pretreatment by 1.4-fold, showing the effectiveness of including an extrusion step to enhance overall biomass fractionation and carbohydrates conversion via microbial fermentation processes.

Washout and non-washout solutions of a system describing microbial fermentation process under the influence of growth inhibitions and maximal concentration of yeast cells

An unstructured model for the growth of yeast cell on glucose due to growth inhibitions by substrate, products, and cell density is discussed. The proposed model describes the dynamical behavior of fermentation system that shows multiple steady states for a certain regime of operating parameters such as inlet glucose and dilution rate. Two types of steady state solutions are found, namely washout and non-washout solutions. Furthermore, different numerical impositions to the two parameters put in evidence three results regarding non-washout solution: a unique locally stable non-washout solution, a unique locally stable non-washout solution towards which other nearby solutions exhibit damped oscillations, and multiple non-washout solutions where one is locally stable while the other is unstable. It is also found an optimal inlet glucose which produces the highest cell and ethanol concentration.