A possible mechanism of metabolic regulation in Gibberella fujikuroi using a mixed carbon source of glucose and corn oil inferred from analysis of the kinetics data obtained in a stirrer tank bioreactor

A nonstructured model was used to study the dynamics of gibberellic acid production in a stirred tank bioreactor. Experimental data were obtained from submerged batch cultures of Gibberella fujikuroi (CDBB H‐984) grown in varying ratios of glucose‐corn oil as the carbon source. The nitrogen depletion effect was included in mathematical model by considering the specific kinetic constants as a linear function of the normalized nitrogen consumption rate. The kinetics of biomass growth and consumption of phosphate and nitrogen were based on the logistic model. The traditional first‐order kinetic model was used to describe the specific consumption of glucose and corn oil. The nitrogen effect was solely included in the phosphate and corn oil consumption and biomass growth. The model fit was satisfactory, revealing the dependence of the kinetics with respect to the nitrogen assimilation rate. Through simulations, it was possible to make diagrams of specific growth rate and specific rate of substrate consumptions, which was a powerful tool for understanding the metabolic interactions that occurred during the various stages of fermentation process. This kinetic analysis provided the proposal of a possible mechanism of regulation on growth, substrate consumptions, and production of gibberellic acid (GA₃) in G. fujikuroi. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1169–1180, 2013     

Model-based optimization of biosurfactant production in fed-batch culture Azotobacter vinelandii

Fed-batch cultivation of Azotobacter vinelandii 21was optimized for biosurfactant production. Optimization of feed-rate time profile and concentration of nutrient medium components in feeding solution is based on a hybrid mathematical model consisting of mass-balance equations for biomass, biosurfactant, volume of cultural liquid and substrate components: glucose, ammonia nitrogen and phosphate phosphorus. The rate of cultural liquid emulsification activity growth as well as the rates of ammonia nitrogen and phosphate phosphorus consumption is modelled by means of artificial neural network, while the rates of the other biochemical transformations are modelled by adequate kinetic relationships.     

Change in cereal production caused by climate change in Malaysia

This study attempts to estimate the effects of climate change variables, such as average temperature, CO₂ emissions and average rainfall, on cereal production in Malaysia from 1969 to 2018. After preliminary tests on time series data, we employed a novel autoregressive distributed lag (ARDL) method known as the dynamic ARDL simulations technique. The results showed that a long-run co-integration relationship exists between cereal production and climatic and non-climatic factors. All climate variables have a negative impact on cereal yield, while energy consumption and cultivated land have a positive effect on cereal yield in the short- and long-run. Granger causality analyses also showed that a unidirectional causality link exists between rainfall and temperature with cereal production and between CO₂ emissions and cereal production. Energy consumption, as a proxy for technology, has a one-way Granger cause with cereal production. The results of the dynamic ARDL simulations suggest that cereal yield was most sensitive to CO₂ emissions, rainfall and temperature. In the long run, a 1% increase in temperature is associated with a 2.87% and 3.52% decrease in general and predicted estimates of cereal production, respectively. The dynamic ARDL simulations methodology provides a better understanding of the variability of cereal production in Malaysia as a result of climate change.     

Conditions for the production of Agrocin 84 byAgrobacterium radiobacter K84

Summary A chemically defined medium was developed that supported the growth ofAgrobacterium radiobacter K84 and the production of agrocin 84. Various supplements were investigated for their effect on growth rate and production of agrocin 84 using a well-diffusion assay method. Mannitol was found to be a better substrate for growth ofA. radiobacter K84 compared to the other sugar alcohols and sugars tested. By contrast,d-fructose was the better substrate for the production of agrocin 84. Biotin supplementation stimulated production of agrocin 84 but did not eliminate the diauxic lag seen with the basal medium. The opine, octopine, inhibited growth ofA. radiobacter K84 and production of agrocin 84 as did coenzyme B₁₂. By contrast, the cytokinin, isopentenyl adenosine, was marginally stimulatory to production, as was vitamin B₁₂. Acetosyringone supplementation had a negligible effect on growth rate and production of agrocin 84.     

Citric acid production from sugar cane molasses by 2-deoxyglucose-resistant mutant strain ofAspergillus niger

Citric acid production from sugar cane molasses byAspergillus niger NIAB 280 was studied in a batch cultivation process. A maximum of 90 g/L total sugar was utilized in citric acid production medium. From the parental strainA. niger, mutant strains showing resistance to 2-deoxyglucose in Vogal's medium containing molasses as a carbon source were induced by γ-irradiation. Among the new series of mutant strains, strain RP7 produced 120 g/L while the parental strain produced 80 g/L citric acid (1.5-fold improvement) from 150 g/L of molasses sugars. The period of citric acid production was shortened from 10 d for the wild-type strain to 6–7 d for the mutant strain. The efficiency of substrate uptake rate with respect to total volume substrate consumption rate,Q _s (g per L per h) and specific substrate consumption rate,q _s (g substrate per g cells per h) revealed that the mutant grew faster than its parent. This indicated that the selected mutant is insensitive to catabolite repression by higher concentrations of sugars for citric acid production. With respect to the product yield coefficient (Y _p/x), volume productivity (Q _p) and specific product yields (q _p), the mutant strain is significantly (p≤0.05) improved over the parental strain.     

Effect of ammonium feeding on production of thiostrepton by fed-batch culture

Summary The effect of ammonium ions in the medium on production of thiostrepton byStreptomyces laurentii was investigated. In batch cultures, the excessive initial concentration of ammonium ions inhibited thiostrepton production. Moderate feeding of ammonia accelerated, however, not only microbial growth but also thiostrepton production. Fed-batch cultures with various feed rates of ammonia and a kinetic study clarified the effect of ammonium ion consumption rate on thiostrepton production. A modified kinetic model is proposed that takes product inhibition and the influence of maximum thiostrepton content into account.     

Modelling of cyclic fed-batch plus batch polygalacturonase production by Aureobasidium pullulans on raw orange peel

Summary Cyclic fed-batch plus batch polygalacturonase production by Aureobasidium pullulans in slurry fermentation systems using raw orange peel as substrate was studied in a 3-dm³ stirred fermentor by setting the main operating variables (T=297°K; pH₀=3.2; OP₀=3% w/v; n=700 rpm) to optimal values determined previously. In this way, it was possible to stabilize enzyme excretion at 130–140 VU cm^–3. The time course of this fermentation process in terms of cell growth, substrate consumption and enzyme synthesis was reconstructed with a mean standard error less than 10%, by applying an unstructured model set up in a batch run and further refined in a series of cyclic fed-batch plus batch operations. In particular, the enzyme formation rate was related to the effect of reducing sugars as inhibitors at higher concentrations and as activators at lower levels by using an exponential equation. Moreover, the consumption rate of reducing sugars was found to be linearly related to the cell growth rate, its specific date being of pseudo-first order with respect to the reducing sugar concentration.Offprint requests to: M. Moresi     

Mathematical description of yessotoxin production by Protoceratium reticulatum in culture

The production dynamics of yessotoxin (YTX) by Protoceratium reticulatum and phosphate uptake in culture were investigated in relation to cell growth. The equations used were: the reparametrized logistic for cell production, Luedeking–Piret model for yessotoxin production and maintenance energy model for phosphate consumption. Thus, the YTX formation rate was proportional to producer biomass at the end of the asymptotic phase of culture as a secondary metabolite. Moreover, the equations proposed showed a high accuracy to predict these bioproductions in different experimental conditions and culture scales.     

Simultaneous NH3 oxidation and N2 production at reduced O2 tensions by sewage sludge subcultured with chemolithotrophic medium

The ammonia oxidation rate by sewage sludge was determined as a function of the dissolved oxygen tension. Samples of sludge were taken from a domestic waste water treatment pilot plant in which sludge was completely retained by membrane filtration. The samples were subcultured chemolithotrophically in recycling reactors. The gas supplied was a mixture of pure argon and oxygen. The K _O2 for ammonia oxidation was estimated to be 0.97 (±0.16) kPa dissolved oxygen. Together with ammonia oxidation and oxygen consumption, dinitrogen gas was produced. So, aerobic denitrification occurred. At dissolved oxygen tensions of 1.25 kPa and higher, the dinitrogen production rate (per N-mole) equalled 20% of the ammonia oxidation rate. This proportion was even 58% at 0.3 kPa dissolved oxygen. At 0.15 kPa dissolved oxygen, however, nitrification hardly proceeded, while dinitrogen production soon stopped. Most likely, a nitrifier concomitantly oxidized ammonia and reduced nitrite to dinitrogen.     

Dynamics and stability analysis of the growth and astaxanthin production system of Haematococcus pluvialis

This paper investigated high cell density cultivation of Haematococcus pluvialis for astaxanthin production in 3.7-L bioreactors. A biomass concentration of 2.74 g L⁻¹and an astaxanthin yield of 64.4 mg L⁻¹ were obtained. Based on the experimental results, a new and simple dynamic model is proposed, differing from Monod kinetics, to describe cell growth, product formation and substrate consumption. Good agreement was found between the model predictions and experimental data. The model revealed that there was cell growth inhibition on product formation and product feedback compensation for substrate consumption, but no substrate inhibition or product inhibition of cell growth. Stability analysis demonstrated that no multiplicity of steady states was observed; the unique positive steady state was locally asymptotically stable; and the effect of dilution rate on steady states was greater than that of the initial substrate concentration. Received 23 February 1999/ Accepted in revised form 08 June 1999     

A kinetic model incorporating energy spilling for substrate removal in substrate-sufficient batch culture of activated sludge

Batch assays are currently used to study the kinetic behavior of microbial growth. However, it has been shown that the outcome of batch experiments is greatly influenced by the initial ratio of substrate concentration (S _o) to biomass concentration (X _o). Substrate-sufficient batch culture is known to have mechanisms of spilling energy that lead to significant nongrowth-associated substrate consumption, and the Monod equation is no longer appropriate. By incorporating substrate consumption associated with energy spilling into the balance of the substrate oxidation reaction, a kinetic model for the observed specific substrate consumption rate was developed for substrate-sufficient batch culture of activated sludge, and was further verified by experimental data. It was demonstrated that the specific substrate consumption rate increased with the increase of the S _o/X _o ratio, and the majority of substrate was consumed through energy spilling at high S _o/X _o ratios. It appears that the S _o/X _o ratio is a key parameter in regulating metabolic pathways of microorganisms. Received: 18 January 1999 / Received revision: 7 May 1999 / Accepted: 28 May 1999     

Enzyme production in a cell recycle fermentation system evaluated by computer simulations

Enzyme production in a cell recycle fermentation system was studied by computer simulations, using a mathematical model of -amylase production by Bacillus amyloliquefaciens. The model was modified so as to enable simulation of enzyme production by hypothetical organisms having different production kinetics at different fermentation conditions important for growth and production. The simulations were designed as a two-level factorial assay, the factor studied being fermentation with or without cell recycling, repression of product synthesis by glucose, kinetic production constants, product degradation by a protease, mode of fermentation, and starch versus glucose as the substrate carbon source.The main factor of importance for ensuring high enzyme production was cell recycling. Product formation kinetics related to the stationary growth phase combined with continuous fermentation with cell recycling also had a positive impact. The effect was greatest when two or more of these three factors were present in combinations, none of them alone guaranteeing a good result. Product degradation by a protease decreased the amount of product obtained; however, when combined with cell recycling, the protease effect was overshadowed by the increased production. Simulation of this type should prove a useful tool for analyzing troublesome fermentations and for identifying production organisms for further study in integrated fermentation systems.List of Symbols a proportionality constant relating the specific growth rate to the logarithm of G (h) - a ₁ reaction order with respect to starch concentration - a ₂ reaction order with respect to glucose concentration - c starch concentration (g/l) - c ₀ starch concentration in the feed (g/l) - D dilution rate (h^–1) - e intrinsic intracellular amylase concentration (g product/g cell mass) - E extracellular amylase concentration (g/l) - F volumetric flow rate (l/h) - G average number of genome equivalents of DNA/cell - K ₁ intracellular repression constant - K ₂ intracellular repression constant - K _s Monod saturation constant (g/l) - k ₃ product excretion rate constant (h^–1) - k _I translation constant (g product/g mRNA/h) - k _d first order decay constant (h^–1) - k _dw first order decay constant (h^–1) - k _gl rate constant for glucose production (g/l/h) - k _{m, dgr} saturation constant for product degradation (g/l) - k _st rate constant for starch hydrolysis (g/l/h) - k _t1 proportionality constant for amylase production (g mRNA/g substrate) - k _t2 proportionality constant for amylase production (g mRNA *h/g substrate) - k _w protease excretion rate constant (h^–1) - k _wt1 proportionality constant for protease production (g mRNA/g substrate) - k _wt2 proportionality constant for protease production (g mRNA *h/g substrate) - k _wI translation constant (g protease/g mRNA/h) - m maintenance coefficient (g substrate/g cell mass/h) - n number of binding sites for the co-repressor on the cytoplasmic repressor - Q repression function, K₁/K₂ less than or equal to 1.0 - Q _w repression function, K₁/K₂ less than or equal to 1.0 - r intrinsic amylase mRNA concentration (g mRNA/g cell mass) - r _m intrinsic protease mRNA concentration (g mRNA/g cell mass) - R _ex retention by the filter of the compounds x=: C starch, E amylase, or S glucose - R _t amylase transport rate (g product/g cell mass/h) - R _wt protease transport rate (g protease/g cell mass/h) - R _s rate of glucose production (g/l/h) - R _c rate of starch hydrolysis (g/l/h) - S ₀ feed concentration of free reducing sugar (g/l) - s extracellular concentration of reducing sugar (g/l) - t time (h) - V volume (1) - w intracellular protease concentration (g/l) - W extracellular protease concentration (g/l) - X cell mass concentration (dry weight) (g/l) - Y yield coefficient (g cell mass/g substrate) - substrate uptake (g substrate/g cell mass/h) - specific growth rate of cell mass (h^–1) - _d specific death rate of cells (h^–1) - _m maximum specific growth rate of cell mass (h^–1) - _m,dgr maximum specific rate of amylase degradation (h^–1) This study was supported by the Nordic Industrial Foundation Bioprocess Engineering Programme and the Center for Process Biotechnology, The Technical University of Denmark.     

A study of photorespiratory ammonia production in the C4 plant Amaranthus edulis, using mutants with altered photosynthetic capacities

Previous studies have indicated that the rate of photorespiration in C₄ plants is low or negligible. In this study, wild-type and mutant leaves of the C₄ plant Amaranthus edulis were treated with the glutamine synthetase inhibitor, phosphinothricin and the glycine decarboxylase inhibitor, aminoacetonitrile, at different concentrations of CO₂. The time course of ammonia accumulation in leaves of the wild type was compared with a mutant lacking phosphoenolpyruvate carboxylase activity (EC 4.1.1.31), and with three different mutants that accumulated glycine. The increase in the concentration of ammonia in the leaves, stimulated by the treatments was used as a measurement of the rate of photorespiration in C₄ plants. The application of glutamine and glycine maintained the rate of photorespiratory ammonia production for a longer period in the wild type, and increased the rate in a mutant lacking phosphoenolpyruvate carboxylase suggesting that there was a lack of amino donors in these plants. The calculated rate of photorespiration in Amaranthus edulis wild-type leaves when the supply of amino donors was enough to maintain the photorespiratory nitrogen flow, accounted for approximately 6% of the total net photosynthetic CO₂ assimilation rate. In a mutant lacking phosphoenolpyruvate carboxylase, however, this rate increased to 48%, when glutamine was fed to the leaf, a value higher than that found in some C₃ plants. In mutants of Amaranthus edulis that accumulated glycine, the rate of photorespiration was reduced to 3% of the total net CO₂ assimilation rate. The rate of ammonia produced during photorespiration was 60% of the total produced by all metabolic reactions in the leaves. The data suggests that photorespiration is an active process in C₄ plants, which can play an important role in photosynthetic metabolism in these plants.     

Mathematical description of bikaverin production in a fluidized bed bioreactor

Summary  Growth of Gibberella fujikuroi in submerged cultures occurs as micelles or filamentous hyphae dispersed in fluid and pellets or stable, spherical agglomerations. Gibberella fujikuroi growth, substrate consumption and bikaverin production kinetics obtained from submerged batch fermentation were fitted to three different sigmoid models: two and three-parameter Gompertz models and one Logistic model. Growth fitting was used to compare between models and select the best one by means of an F test. The best model for describing growth was the two-parameter Gompertz model and was used for glucose consumption and bikaverin production fitting. Data from eight different schemes of fermentations were analysed and parameter estimation was carried out by means of minimization of residual sum of squares. Some characteristic values obtained with the two-parameter Gompertz model fit are: μ=0.028 h⁻¹, Y_x/s=0.1089 g substrate/g biomass, α =0.1384 g product/g biomass.     

Fed-batch propionic acid production by Propionibacterium acidipropionici

The batch fermentations were conducted using lactose as the substrate at pH 6.5 and temperature 30°C. Average batch kinetic data was eventually used to develop an unstructured mathematical model. The kinetic parameters of the model were determined by non-linear regression technique using the batch experimental results. Parametric sensitivity analysis showed the maximum specific substrate consumption rate (r_S^max) and the maintenance energy constant (m_S) to be the most sensitive parameters. The experimental observations in batch fermentation were close to the model predictions. The batch model was extrapolated to identify nutrient feeding strategies, which were tested successfully for two different fed-batch fermentations. It demonstrated enhanced propionic acid productivity. The developed model was found suitable for the design of feeding strategies to increase propionic acid production in fed-batch mode of reactor operation.     

A robust fed-batch feeding strategy for optimal parameter estimation for baker's yeast production

Parameter identification of structured models is often a problem in biotechnology, because the poor data situation and the number of unknown parameters only allow for inaccurate estimates. But often only a subset of all kinetic parameters of the model are of interest for production purposes, e.g. for fed-batch cultivation. These parameters should be estimated with a given accuracy. In addition, the experiments for information acquisition with respect to these parameters should be as simple as possible and should consider some practical restrictions. In this contribution a fed-batch feeding strategy is proposed to allow for an accurate estimation of yield and of critical growth rate of baker's yeast. The feeding also allows for economic and stereotyped use of staff and equipment and is therefore suitable for routine use in screening of strains and media. The overall pattern is similar to that one, usually used in production scale to minimize errors by limited model validity. After an initial phase for achieving a reproducible state three different growth rates are adjusted to cover the range of possible critical growth rates. From biomass and ethanol measurements yield and critical growth rate can be estimated with an accuracy of about 2.1%. The fermentation pattern ends up with a constant feeding rate to simulate a limited oxygen transfer rate and to allow for an uptake of residual sugar and ethanol before a dough test can be carried out. Beside experimental results simulations and sensitivity analyses are shown.List of Symbols P ethanol concentration - S substrate concentration - S _f substrate concentration in feed - T fermentation time - V fermenter volume - X biomass concentration - C measurement error covariance matrix - F Fisher information matrix - X state variables - Y output variables - X _p state sensitivity functions with respect to parameters - Y _p output sensitivity functions - e eigenvectors - k vector of limitation and inhibition parameters - n number of observations - q _in feeding stream - q _b stream for samples and ammonia feed - r vector of specific turnover rates - y vector of yields - specific weight - eigenvalues - specific growth rate - _set exponent in exponential feeding - standard deviation Dedicated to the 65th birthday of Professor Fritz Wagner.A. O. Ejiofor and B. O. Solomon are grateful to the Alexander von Humboldt Stiftung for granting them fellowships and to GBF for providing all the materials necessary for their successful research stay in Germany.     

A model of xylitol production by the yeast Candida mogii

Production of xylitol from xylose in batch fermentations of Candida mogii ATCC 18364 is discussed in the presence of glucose as the cosubstrate. Various initial ratios of glucose and xylose concentrations are assessed for their impact on yield and rate of production of xylitol. Supplementation with glucose at the beginning of the fermentation increased the specific growth rate, biomass yield and volumetric productivity of xylitol compared with fermentation that used xylose as the sole carbon source. A mathematical model is developed for eventual use in predicting the product formation rate and yield. The model parameters were estimated from experimental observations, using a genetic algorithm. Batch fermentations, which were carried out with xylose alone and a mixture of xylose and glucose, were used to validate the model. The model fitted well with the experimental data of cell growth, substrate consumption and xylitol production.     

Excretory nitrogen metabolism in the Chinese fire-belly newt<Emphasis Type="Italic"> Cynops orientalis</Emphasis> in water,on land,or in high concentrations of environmental ammonia

The Chinese fire-belly newt Cynops orientalis reverts to an aquatic mode of living when sexually mature. Despite living in water, sexually mature C. orientalis maintained high capacity for hepatic urea synthesis. However, it had a lower rate of urea production than other terrestrial amphibians because endogenous ammonia could diffuse out to the external medium as NH₃. This conserves cellular energy because urea synthesis is energetically expensive. Simultaneously, C. orientalis also reduced the rate of urea excretion, and excreted 33% of the total nitrogenous waste as ammonia. Upon exposure to land, C. orientalis increased the rate of urea synthesis from accumulating endogenous ammonia. The increased rate of urea synthesis was within the inherent capacity of the hepatic ornithine–urea cycle; there was no induction of hepatic carbamoyl phosphate synthetase or ornithine transcarbamoylase activities and there was no reduction in ammonia production. When exposed to water containing 75 mmol.l^–1 NH₄Cl, the rates of both urea synthesis and urea excretion increased. Under such experimental conditions, the ornithine–urea cycle may be operating close to its limit; glutamine began to accumulate in the body, and endogenous ammonia production via amino acid catabolism was reduced.Abbreviations CPS carbamoyl phosphate synthetase - FAA free amino acid - OTC ornithine transcarbamoylase - OUC ornithine–urea cycle - TCA trichloroacetic acid Communicated by I.D. Hume     

Microbial N2O consumption in and above marine N2O production hotspots

The ocean is a net source of N₂O, a potent greenhouse gas and ozone-depleting agent. However, the removal of N₂O via microbial N₂O consumption is poorly constrained and rate measurements have been restricted to anoxic waters. Here we expand N₂O consumption measurements from anoxic zones to the sharp oxygen gradient above them, and experimentally determine kinetic parameters in both oxic and anoxic seawater for the first time. We find that the substrate affinity, O₂ tolerance, and community composition of N₂O-consuming microbes in oxic waters differ from those in the underlying anoxic layers. Kinetic parameters determined here are used to model in situ N₂O production and consumption rates. Estimated in situ rates differ from measured rates, confirming the necessity to consider kinetics when predicting N₂O cycling. Microbes from the oxic layer consume N₂O under anoxic conditions at a much faster rate than microbes from anoxic zones. These experimental results are in keeping with model results which indicate that N₂O consumption likely takes place above the oxygen deficient zone (ODZ). Thus, the dynamic layer with steep O₂ and N₂O gradients right above the ODZ is a previously ignored potential gatekeeper of N₂O and should be accounted for in the marine N₂O budget.Subject terms: Water microbiology, Biogeochemistry, Microbial ecology     

A cybernetic model to predict the effect of freely available nitrogen substrate on rifamycin B production in complex media

It is well-known that secondary metabolite production is repressed by excess nitrogen substrate available in the fermentation media. Although the nitrogen catabolite repression has been known, quantitative process models have not been reported to represent this phenomenon in complex medium. In this paper, we present a cybernetic model for rifamycin B production via Amycolatopsis mediterranei S699 in complex medium, which is typically used in industry. Nitrogen substrate is assumed to be present in two forms in the medium; available nitrogen (S _ANS) such as free amino acids and unavailable nitrogen (S _UNS) such as peptides and proteins. The model assumes that an inducible enzyme catalyzes the conversion of S _UNS to S _ANS. Although S _ANS is required for growth and product formation, high concentrations were found to inhibit rifamycin production. To experimentally validate the model, five different organic nitrogen sources were used that differ in the ratio of S _ANS/S _UNS. The model successfully predicts higher rifamycin B productivity for nitrogen sources that contain lower initial S _ANS. The higher productivity is attributed to the sustained availability of S _ANS at low concentration via conversion of S _UNS to S _ANS, thereby minimizing the effects of nitrogen catabolite repression on rifamycin production. The model can have applications in model-based optimization of substrate feeding recipe and in monitoring and control of fed batch processes.