Efficient Fermentation Process Research Articles

Metabolic Versatility of acetogens in syngas Fermentation: Responding to varying CO availability.

Syngas fermentation using acetogenic bacteria offers a promising route for sustainable chemical production. However, gas-liquid mass transfer limitations and efficient co-utilization of CO and H2 pose significant challenges. This study investigated the kinetics of syngas conversion to acetate by Acetobacterium wieringae and Clostridium species in batch conditions under varying initial CO partial pressures (19 - 110 kPa) in batch cultures. A. wieringae strains, exhibited superior growth in all gas compositions, with a maximum growth rate of 0.104 h-1. The distinct CO, H2, and CO2 consumption patterns revealed metabolic flexibility and adaptation to varying syngas compositions. Notably, A. wieringae strains and C. autoethanogenum achieved complete CO and H2 conversion, with C. autoethanogenum also exhibiting net CO2 uptake. These findings provide valuable insights into the distinct metabolic capabilities of these acetogens and contribute to the development of efficient and sustainable syngas fermentation processes.

Coenzyme A biosynthesis in Bacillus subtilis: discovery of a novel precursor metabolite for salvage and its uptake system.

Vitamins are essential components of the diet of animals and humans. Vitamins are thus important targets for biotechnological production. While efficient fermentation processes have been developed for several vitamins, this is not the case for vitamin B5 (pantothenate), the precursor of coenzyme A. We have elucidated the complete pathway for coenzyme A biosynthesis in the biotechnological workhorse Bacillus subtilis. Moreover, a salvage pathway for coenzyme A synthesis was found in this study. Normally, this pathway depends on pantetheine; however, we observed activity of the salvage pathway on complex medium in mutants lacking the pantothenate biosynthesis pathway even in the absence of supplemented pantetheine. This required rewiring of metabolism by expressing a cystine transporter due to acquisition of mutations affecting the regulation of cysteine metabolism. This shows how the hidden "underground metabolism" can give rise to the rapid formation of novel metabolic pathways.

Hydrogen breath test as a diagnostic tool

Introduction and Purpose: Hydrogen breath tests serve the crucial purpose of evaluating the absorption of nutrients in the small intestine, particularly when carbohydrates are not properly absorbed, leading to bacterial overgrowth. These tests are non-invasive, cost-effective, and widely accessible diagnostic tools used to assess various gastrointestinal disorders. The objective of this review is to evaluate the effectiveness and methodology of Hydrogen breath testing (HBT). State of knowledge: The disposal of hydrogen gas is essential for maintaining efficient microbial fermentation processes in the gut. Hydrogenotrophic microbes, including acetogens, methanogenic archaea, and sulfate-reducing bacteria, are responsible for this process. Hydrogen breath tests, conducted with substrates like glucose and lactulose, aid in diagnosing conditions such as small intestinal bacterial overgrowth (SIBO), lactose and fructose intolerance, and other carbohydrate absorption disorders. These tests generate a large volume of data, which can be analyzed using data mining techniques to uncover new hypotheses. Conclusions: Hydrogen breath tests, using various substrates, are effective in diagnosing gastrointestinal disorders such as SIBO and carbohydrate malabsorption. They offer valuable insights into microbial processes in the gut and can inform individualized treatment strategies. However, careful consideration of contraindications and proper test administration protocols is essential for accurate diagnosis and interpretation of results.

Efficient de novo production of bioactive cordycepin by Aspergillus oryzae using a food-grade expression platform

BackgroundCordycepin (3′-deoxyadenosine) is an important bioactive compound in medical and healthcare markets. The drawbacks of commercial cordycepin production using Cordyceps spp. include long cultivation periods and low cordycepin yields. To overcome these limitations and meet the increasing market demand, the efficient production of cordycepin by the GRAS-status Aspergillus oryzae strain using a synthetic biology approach was developed in this study.ResultsAn engineered strain of A. oryzae capable of cordycepin production was successfully constructed by overexpressing two metabolic genes (cns1 and cns2) involved in cordycepin biosynthesis under the control of constitutive promoters. Investigation of the flexibility of carbon utilization for cordycepin production by the engineered A. oryzae strain revealed that it was able to utilize C6-, C5-, and C12-sugars as carbon sources, with glucose being the best carbon source for cordycepin production. High cordycepin productivity (564.64 ± 9.59 mg/L/d) was acquired by optimizing the submerged fermentation conditions.ConclusionsThis study demonstrates a powerful production platform for bioactive cordycepin production by A. oryzae using a synthetic biology approach. An efficient and cost-effective fermentation process for cordycepin production using an engineered strain was established, offering a powerful alternative source for further upscaling.

Optimization of Rare Ginsenosides and Antioxidant Activity Quality of Ginseng Jiaosu Based on Probiotic Strains and Fermentation Technology

The primary active components of plant ginseng are saponins, and rare saponins play a specific antitumor action. In this study, Lactobacillus that can efficiently transform rare saponins were screened as fermentation strains of ginseng enzymes. The fermentation process was optimized to investigate the changes in various biochemical indicators and antioxidant activities of the enzymes before and after fermentation. The results showed that Lactiplantibacillus plantarum had the highest conversion efficiency for diol saponins, among which the content of rare ginsenoside-CK increased by 256%. The optimum optimizing method was a 1 : 1 : 10 ratio of ginseng extract/concentrated apple juice/water, fermentation time of 16 days, initial pH of 6.0, fermentation temperature of 37°C, and sterilization amount of 1.0%. The contents of rare ginsenoside-Rh1, ginsenoside-F2, ginsenoside-Rg3, and ginsenoside-CK increased by 18.48%, 135.73%, 343.03%, and 441.80%, respectively, and the process was stable and controllable. After fermentation, the scavenging rates for hydroxyl radical, DPPH radical, and superoxide anion radical were increased by more than 9%, and new organic acids were produced. In general, this study established an efficient, stable, and controllable ginseng fermentation process to improve the rare saponin content and biological activity of ginseng enzymes, which can provide new ideas for traditional Chinese medicine and medicinal health food in China.

Statistical Medium Optimization for the Production of Anti-Methicillin-Resistant Staphylococcus aureus Metabolites from a Coal-Mining-Soil-Derived Streptomyces rochei CMB47

The development of novel antibacterial drugs needs urgent action due to the global emergence of antibiotic resistance. In this challenge, actinobacterial strains from arid ecosystems are proving to be promising sources of new bioactive metabolites. The identified Streptomyces rochei strain CMB47, isolated from coal mine Saharan soil, provided an ethyl acetate extract which tested against a series of pathogens. It displayed a minimum inhibitory concentration of <0.439 µg/mL against MRSA. A statistical experimental design using a response surface methodology (RSM) based on the second-order rotatable central composite design (RCCD) was planned to develop an efficient fermentation process able to improve the bioactive metabolite production. The optimal conditions were determined for starch and NaNO3 concentrations, incubation time and the initial pH value, reaching the inhibition zone diameter of 20 mm, close to the experimental value, after validation of the model. A bioassay-guided fractionation of the crude extract provided the most active fractions, which were analyzed by HPLC equipped with a photodiode array detector and coupled online with an electrospray mass spectrometer (HPLC-DAD/ESI-MS), obtaining preliminary indications on the molecular structures of the metabolites. These results support the potential interest in further investigations into the purification and full characterization of the metabolites responsible for the biological activity observed so far.

Development of an oil-sealed anaerobic fermentation process for high production of γ-aminobutyric acid with Lactobacillus brevis isolated by directional colorimetric screening

γ-aminobutyric acid (GABA) is an important inhibitory neurotransmitter in the brain with many physiological functions. Therefore, this work establishes an efficient biosynthesis process to achieve the high production of food-grade GABA. Firstly, we adopted a directional colorimetric screening method and then obtained the strain Lactobacillus brevis (Lb. brevis) CGMCC 24975 with 22.61 g/L GABA in shake flasks, much higher than Lb. brevis ATCC 367 (3.07 g/L) due to its more efficient and stable expression of its glutamate decarboxylase (GAD) system. After exploring the effects of optimal fermentation conditions and oxygen on GABA production, we found that anaerobic fermentation could increase GABA production by up-regulating gad operon expression and activating GADase. Finally, we developed an oil-sealed anaerobic fermentation process with 157.90 g/L GABA in a 5 L bioreactor, which was 6.98-fold of that in shake flasks. This work includes a complete process from screening GABA-producing strains to developing efficient fermentation processes, laying the foundation for producing food-safe GABA.

High Production of γ-Aminobutyric Acid by Activating the xyl Operon of Lactobacillus brevis.

γ-Aminobutyric acid (GABA) is an inhibitory neurotransmitter with important physiological functions such as sleep assistance and anti-depression. In this study, we developed a fermentation process for the high-efficiency production of GABA by Lactobacillus brevis (Lb. brevis) CE701. First, xylose was found as the optimal carbon source that could improve the GABA production and OD600 in shake flasks to 40.35 g/L and 8.64, respectively, which were 1.78-fold and 1.67-fold of the glucose. Subsequently, the analysis of the carbon source metabolic pathway indicated that xylose activated the expression of the xyl operon, and xylose metabolism produced more ATP and organic acids than glucose, which significantly promoted the growth and GABA production of Lb. brevis CE701. Then, an efficient GABA fermentation process was developed by optimizing the medium components using response surface methodology. Finally, the production of GABA reached 176.04 g/L in a 5 L fermenter, which was 336% higher than that in a shake flask. This work enables the efficient synthesis of GABA using xylose, which will provide guidance for the industrial production of GABA.

Enhanced biosurfactant production by Bacillus subtilis SPB1 using developed fed-batch fermentation: effects of glucose levels and feeding systems.

Biosurfactants stand for highly useful and promising compounds. They basically serve for a variety of applications in multiple industries and aspects of human life. Therefore, it is highly required to improve their production yield especially through the development of new and more efficient fermentation processes. In this aim, batch and fed-batch were studied and compared in terms of their effective biosurfactant production by Bacillus subtilis SPB1. Experiments of fed-batch fermentations were carried out through three different glucose feeding strategies, namely the pulsed, the constant Donespeed and the exponential feeding. The comparison between different fermentation processes revealed that fed-batch process proved to be a more efficient cultivation strategy than the batch process in terms of cell biomass, biosurfactant production and productivity. Among the three different feeding strategies, the exponential feeding process achieved the highest fermentation results of final biosurfactant concentration. The latter increased more than twofolds compared to batch fermentation.

Enhanced production of poly-γ-glutamic acid by Bacillus licheniformis ATCC 9945a using simultaneous pulse-feedings of citrate and glutamate

Poly-γ-glutamic acid (γ-PGA) is a versatile biopolymer with widespread applications in the food, pharmaceutical, and medical industries. One of the main challenges in expanding γ-PGA industrial applications is the high cost of production. Developing an efficient and low-cost fermentation process such as bacterial cultivation with pulsed feeding can significantly reduce production costs. Thus, initially, a new pulsed-feeding strategy of citrate and glutamate was developed for γ-PGA production enhancement in the fed-batch culture of Bacillus licheniformis ATCC 9945a. Then, the effects of pulse number, feeding amount, feeding times, the addition time of calcium and manganese solutions, the pH of the added citrate solution, and the concentration of feed stock solutions of pulse-feeds on γ-PGA production were investigated. Under optimal conditions: feeding two pulses at 8 and 24 hours of culture, 20 g citrate and glutamate per liter of culture medium per pulse (about 52 mL of each of citrate and glutamate feeding solutions prepared with a concentration of 384 g/L by adding distilled water) about 88 ± 4 g/L of γ-PGA was obtained. It is one of the highest values ever reported for γ-PGA production with Bacillus licheniformis ATCC 9945a, of course with a much simpler process than the other fed-batch processes.

Enhanced ATP and antioxidant levels for cAMP biosynthesis by Arthrobacter sp. CCTCC 2013431 with polyphosphate addition.

When citrate and pyruvate were utilized to strengthen ATP generation for high cAMP production, oxidative stress became more severe in cells resulting in lower cell viability and cAMP formation at the late fermentation phase. To further improve cAMP biosynthesis, the effects of polyphosphate on cAMP fermentation performance together with intracellular ATP and oxidation levels were investigated under high oxidative stress condition and then high efficient cAMP fermentation process based on polyphosphate and salvage synthesis was developed and studied. With 2g/L-broth sodium hexametaphosphate added at 24h was determined as the optimal condition for cAMP production by Arthrobacter sp. CCTCC 2013431 in shake flasks. Under high oxidative stress condition caused by adding 15mg/L-broth menadione, cAMP contents and cell viability were improved greatly due to hexametaphosphate addition and also exceeded those of control (without hexametaphosphate and menadione added) when fermentations were conducted in a 7 L bioreactor. Meanwhile, ATP levels and antioxidant capacity were improved obviously by hexametaphosphate as well. Moreover, a fermentation process with hexametaphosphate and hypoxanthine coupling added was developed by which cAMP concentration reached 7.25g/L with an increment of 87.1% when compared with only hypoxanthine added batch and the high ROS contents generated from salvage synthesis were reduced significantly. Polyphosphate could improve intracellular ATP levels and antioxidant capacity significantly under high oxidative stress condition resulting in enhanced cell viability and cAMP fermentation production no matter by de novo synthesis or salvage synthesis.

Enhanced ethanol production from lignocellulosic hydrolysate using Meyerozyma caribbica biofilm immobilized on modified epoxy foam

In comparison to planktonic cells, microbial biofilm immobilization is known to enhance ethanol production. To maintain this capability during long-term fermentation, the detachment of the biofilm must be prevented. In this study, the enhancement of the biofilm immobilization of the yeast Meyerozyma caribbica YLP01GX on bio-based epoxy foam (EF) was performed through plasma surface modification to improve the production of ethanol from oil palm empty fruit bunch hydrolysate. This newly synthesized EF serves as a suitable carrier due to its superior properties and cost-effectiveness. The modification could induce yeast cell attachment more rapidly with more extracellular polymeric substance (EPS) being produced in the presence of the yeast extract. Therefore, the immobilization period can be shortened. In comparison to untreated EF-immobilized cells (NEF-IC), the modified EF-immobilized cells (MEF-IC) showed an improved binding affinity for proteins in the EPS, which resulted in a higher adhesion force between the carrier and the yeast cell surface. The stable biofilm attachment on the modified EF can prevent biofilm detachment and improve the tolerance of the yeast to inhibitors in the hydrolysate, leading to enhanced ethanol production compared to NEF-IC. Additionally, the MEF-IC was reused several times with similar fermentation activity being observed. This approach offers a more efficient and economical fermentation process.

Self-assembly of CdS@C. Beijerinckii hybrid system for efficient lignocellulosic butanol production

Bioproduction of butanol from lignocellulose has been considered as a sustainable and eco-friendly technology to cope with energy crisis and environmental issues. In the present research, a novel CdS@C. beijerinckii hybrid system was assembled by combining Clostridium beijerinckii NCIMB 8052 and the in situ formed CdS nanoparticles (NPs) to enhance the butanol production in ABE fermentation. 0.1 g/L of Cd(NO3)2·4H2O was identified for the effective in situ formation of CdS NPs. The in situ formed CdS NPs was powerful in generating photoelectrons which resulted in a significant upgradation of NADH/NAD+ in C. beijerinckii NCIMB 8052 under the visible irradiation of 2000 W/m2. With glucose and butyric acid as co-substrates, butanol production in the fermentation system of hybrid CdS@C. beijerinckii reached 10.42 ± 0.15 g/L, much higher than that of 8.08 ± 0.13 g/L in the fermentation system of C. beijerinckii NCIMB 8052. Butanol production from the mixed carbon source of saccharified liquid and acidogenic fermentation broth of rice straw was also improved by 23.6% in the hybrid system of CdS@C. beijerinckii, comparing to that of C. beijerinckii NCIMB 8052. The upgraded NADH/NAD+ was confirmed as the dominant factor for the enhanced butanol production. The research work provided an efficient fermentation process for the production of lignocellulosic butanol.

Enhanced surfactin fermentation via advanced repeated fed-batch fermentation with increased cell density stimulated by EDTA–Fe (II)

High-performance production of biosurfactant surfactin is very important for real applications of surfactin but still exhibits significant challenges. In this study, an efficient fermentation process with enhanced cell density is developed to improve the surfactin production by Bacillus subtilis ATCC 21332. The addition of 0.2 mM EDTA–Fe2+ complex resulted in the highest surfactin yield of 5.67 g/L within 3 days, which is 7.8 times higher than that obtained by regular fermentation in the absence of this complex. This significantly improved surfactin production is related to the remarkably increased cell density which was stimulated by the addition of EDTA-Fe2+ complex at molar ratio of 1:1, was significantly better than that obtained by the addition of Fe2+, Fe3+ or EDTA alone. Furthermore, a modified repeated fed-batch fermentation process sustained the high-performance fermentation of surfactin at 1.345 g/L d, and the overall surfactin yield reached 9.414 g/L within a fermentation period of 7 days.

Oriented Fermentation of Food Waste towards High-Value Products: A Review

Food waste has a great potential for resource recovery due to its huge yield and high organic content. Oriented fermentation is a promising method with strong application prospects due to high efficiency, strong robustness, and high-value products. Different fermentation types lead to different products, which can be shifted by adjusting fermentation conditions such as inoculum, pH, oxidation-reduction potential (ORP), organic loading rate (OLR), and nutrients. Compared with other types, lactic acid fermentation has the lowest reliance on artificial intervention. Lactic acid and volatile fatty acids are the common products, and high yield and high purity are the main targets of food waste fermentation. In addition to operational parameters, reactors and processes should be paid more attention to for industrial application. Currently, continuously stirred tank reactors and one-stage processes are used principally for scale-up continuous fermentation of food waste. Electro-fermentation and iron-based or carbon-based additives can improve food waste fermentation, but their mechanisms and application need further investigation. After fermentation, the recovery of target products is a key problem due to the lack of green and economic methods. Precipitation, distillation, extraction, adsorption, and membrane separation can be considered, but the recovery step is still the most expensive in the entire treatment chain. It is expected to develop more efficient fermentation processes and recovery strategies based on food waste composition and market demand.

Assessing the Biological Value of Soluble Organic Fractions from Tomato Pomace Digestates

Anaerobic digestion is proposed for sustainable exploitation of the huge amounts of residues produced yearly by tomato processing. The aim of this study was to analyze the biological effects of water-soluble organic fractions of tomato pomace digestates applied on soil and plant, at a laboratory scale. Digestates are derived from a short-term dark fermentation batch experiment of tomato pomace and buffalo slurry at three decreasing ratios (12:1, 8:1, and 6:1, respectively Set 1, Set 2, and Set 3). Bioassays on soil and plant were carried out after addition of different amounts of dissolved organic carbon (DOC) extracted from the three sets: microbial pool content, respiration activity, root development, and cell division were considered. Digestate from Set 1 showed the highest content of volatile fatty acids (about 80 mg UL−1), suggesting an efficient fermentation process which led to lower water solubility of organic substrates. The highest carbon mineralization rate in soil (4.29%) occurred with DOC additions from Set 1 (p 0.05) was detected in the micronuclei assay. Soluble organic forms of these digestates are a nutrient source for soil microorganism metabolism and at low doses show a hormonal-like action, without toxicological or genotoxicity effects on soil biota and root cell.

Losses, chemical composition and aerobic stability of rye silage cv. Temprano subjected to different pre-flowering cuttings with or without fungicide

The present study evaluated the fermentation, chemical characteristics and ruminal disappearance of dry matter of silages of rye cv. Temprano harvested at the stage of floury grain, managed with or without cutting at the vegetative stage and with or without application of Fluxapyroxad + Pyraclostrobin-based fungicide at the pre-flowering phenological stage. This was a 2x2 factorial randomized block experimental design, with 2 cutting regimes (0 and 1 cut) and 2 application management (with and without fungicide), with six replications, where each repetition is represented by a plot of 9.45m², with the treatments SC-SF: without cut at the vegetative stage and without fungicide application; SC-CF: without cut at the vegetative stage with fungicide application; UC-SF: a cut at the vegetative stage with pre-flowering fungicide application; UC-CF: a cut at the vegetative stage and without fungicide application. UC-CF silages were better in the general context, due to the better fiber composition with an average 6.75% reduction in neutral detergent fiber and acid detergent fiber and a 17.10% reduction in lignin content, but, due to its higher DM content, it presented a less efficient fermentation process in terms of organic acid concentration and aerobic stability. However, it did not change the DM losses or ruminal degradability, and there is also the use of DM harvested at the vegetative stage that certainly has a superior quality. The use of fungicide provided only better dry matter index and better fiber composition and improved the fermentation process resulting in lower DM losses without altering the aerobic stability of silages.

Valorization of Waste Biomass in Fermentative Production of Cellulases: A Review

The most promising way to achieve the smooth, flexible and sustainable bioeconomy is the utilization of renewable lignocellulosic biomass as a feedstock for the production of fuels, chemicals, enzymes and high-valued products. Cellulolytic enzymes are indispensable for the maintenance of global carbon cycle, since they catalyze the degradation of cellulose. Therefore for solving the forthcoming waste management and energy issues of mankind, cellulase production technology plays significant and vital role. Cellulases are industrial enzymes and have extensive application in various process industries. Its relatively high cost of production has hindered the wider industrial application. Significant cost reduction is required to enhance the commercial viability of cellulase production technology. Utilization of novel and cheap lignocellulosic renewable resources as substrate for enzyme fermentation process is a promising way of efficient and low cost cellulases production. The present paper reveals, a review on cellulase production through various microorganisms employing economical, abundantly available renewable lignocellulosic biomass as carbon source. It also deals with the recent approaches used at microbial as well as feedstock level, making more efficient, flexible and cost effective fermentation process.

Efficient keratinase expression via promoter engineering strategies for degradation of feather wastes

Keratinases are promising alternatives over ordinary proteases in several industrial applications due to their unique properties compared with their counterparts in the protease categories. However, their large-scale industrial application is limited by the low expression and poor fermentation efficiency of keratinase. Here, we demonstrate that the expression level of keratinase can be improved by constructing a more efficient enzyme expression system hereby enables the highest production titer as regarding recombinant keratinase production to date. Specially, ten promoters were evaluated and the aprE promoter exhibits a significant promotion of keratinase (kerBv) titer from 165 U/mL to 2605 U/mL in Bacillus subtilis. The batch fermentation mode resulted in a maximum keratinase activity of 7176 U/mL at 36 h in a 5-L fermenter. Furthermore, the extracellular keratinase activity attained up to 16,860 U/mL via fed-batch fermentation within 30 h. The combination of keratinase with l-cysteine brings about 66.4 % degree of degradation of feather. Our work provides a new insight into the development of efficient keratinase fermentation processes with B. subtilis cell factory.

Application of inorganic phosphate limitation to efficient isoprene production in Pantoea ananatis.

Establishment of an efficient isoprene fermentation process by adopting inorganic phosphate limitation as the trigger to direct metabolic flux to the isoprene synthetic pathway. We constructed isoprene-producing strains of Pantoea ananatis (a member of the Enterobacteriaceae family) by integrating a heterologous mevalonate pathway and a metabolic switch that senses external inorganic phosphate (Pi) levels. This metabolic switch enabled dual-phase isoprene production, where the initial cell growth phase under Pi-saturating conditions was uncoupled from the subsequent isoprene production phase under Pi-limiting conditions. In fed-batch fermentation using our best strain (SWITCH-PphoC/pIspSM) in a 1-l bioreactor, isoprene concentration in the off-gas was maintained between 300 and 460ppm during the production phase and at 20ppm during the cell growth phase, respectively. The strain SWITCH-PphoC/pIspSM produced totally 2·5gl-1 of isoprene from glucose with a 1·8% volumetric yield in 48h. This proof-of-concept study demonstrated that our Pi-dependent dual-phase production system using a P. ananatis strain as a producer has potential for industrial-scale isoprene fermentation. This Pi-dependent dual-phase fermentation process could be an attractive and economically viable option for the production of various commercially valuable isoprenoids.