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limnospira indica pcc8005 growth in photobioreactor2020-11-25; International audience; the arthrospira-b experiment is the first experiment in space ever allowing the online measurements of both oxygen production rate and growth rate of limnospira indica pcc8005 in batch photobioreactors running on-board iss. four bioreactors were integrated in the iss biolab facility. each reactor was composed of two chambers (gas and liquid) separated by a ptfe membrane and was run in batch conditions. oxygen production was monitored by online measurement of the total pressure increase in the gas chamber. the experiments are composed of several successive batch cultures for each reactor, performed in parallel on iss and on ground. in this work, a model for the growth of the cyanobacterium limnospira indica pcc8005 (also known as arthrospira or spirulina) in these space membrane photobioreactors was proposed and the simulation results obtained are compared to the experimental results gathered in space and on ground. the photobioreactor model was based on a light transfer limitation model, already used to describe and predict the growth and oxygen production in small to large scale ground photobioreactors. it was completed by a model for ph prediction in the liquid phase allowing assessment of the ph increase associated to the bicarbonate consumption for the biomass growth. a membrane gas-liquid transfer model is used to predict the gas pressure increase in the gas chamber. substrate limitation is considered in the biological model. a quite satisfactory fit was achieved between experimental and simulation results when a suitable mixing of the liquid phase was maintained. the data showed that microgravity has no first order effect on the oxygen production rate of limnospira indica pcc8005 in a photobioreactor operating in space in zero gravity conditions.

2. International Conference on Alternative Fuels and Energy (ICAFE) ; Daegu (Corée (Populaire)) - (2017-10-23 - 2017-10-25) / Conférence; Studies of the effects of electromagnetic waves on Saccharomyces cerevisiae emphasize the need to develop instrumented experimental systems ensuring a characterization of the exposition level to enable unambiguous assessment of their potential effects on living organisms. A bioreactor constituted with two separate compartments has been designed. The main element (75% of total volume) supporting all measurement and control systems (temperature, pH, agitation, and aeration) is placed outside the exposure room whereas the secondary element is exposed to irradiation. Measurements of the medium dielectric properties allow the determination of the electromagnetic field at any point inside the irradiated part of the reactor and are consistent with numerical simulations. In these conditions, the growth rate of Saccharomyces cerevisiae and the ethanol yield in aerobic conditions are not significantly modified when submitted to an electromagnetic field of 900 and 2400 MHz with an average exposition of 6.11 V.m−1 and 3.44 V.m−1 respectively.

The aim of this study was to investigate ammonia load removable by a fixed-bed bioreactor. In this study, high ammonia loads, up to 4.5 kgN/m3 d were applied on a fixed bed reactor colonised by two pures nitrifying bacteria: Nitrosomonas europaea ATCC® 19718 and Nitrobacter winogradskyi ATCC® 25391. The loads were similar to the highest values reported in literature especially with a two bacteria population. The complete ammonia removal efficiency in our installation has reached 2.5 kgN/m3.d for a full nitrification. An N-tank in series model was used to represent the fixed bed reactor and to predict the nitrification in the installation. The developed model was accurate and representative of the nitrification assumed by Nitrosomonas europaea ATCC® 19718 and Nitrobacter winogradskyi ATCC® 25391. This model was also used to predict the biomass and oxygen concentrations during the experiment under various operational conditions. The biofilm fixed on the solid support (polysterene beads 2 mm diameter) was mainly present in the bottom of the reactor where the ammonia load was applied. It is demonstrated that in this region of the column the oxygen concentrations reached the lowest values.

International audience; An epoxy-activated monolithic Convective Interaction Media (CIM®) disk was used for the immobilization of endodextranase D8144 from Penicillium sp. (EC 3.2.1.11) in order to produce on-line isomalto-oligosaccharides (IMOs) from Dextran T40. Enzymatic parameters, molecular weight of IMOs and performance of the IMmobilized Enzymes Reactor (IMER) were investigated. The immobilization yield of enzymes was about 45.3% (w/w), and the real specific activity close to 3.26 U mg−1. The Km values did not significantly change between free (12.8 g L−1) and immobilized enzymes (14.2 g L−1), due to the absence of diffusional limitation. The IMER system presented more than 80% of its residual activity after 5000 column volumes, highlighting the high stability of the immobilized endodextranases. Response surface methodology was used to enhance the performance of the IMER. Depending on dextran concentrations and flow rates, specific patterns of IMOs distributions were observed during the enzymatic hydrolysis. Finally, prebiotic activity was also investigated on IMOs produced by medium conditions (flow rate 0.3 mL min−1 and dextran concentrations 4% w/w) against Lactobacillus rhamnosus GG (ATCC 53103). Their scores were at least as good as two commercialized fructo-oligosaccharides (FOS), Fibrulose® F97 and Orafti® P95.

The aim of the present work was to study the growth of two nitrifying bacteria. For modelling the nitrifying subsystem of the MELiSSA loop, Nitrosomonas europaea ATCC® 19718 and Nitrobacter winogradskyi ATCC® 25931 were grown separately and in cocultures. The kinetic parameters of a stoichiometric mass balanced Pirt model were identified: μ max = 0.054 h −1 , decay rate b = 0.003 h −1 and maintenance rate m = 0.135 gN-NH 4 + ·gX −1 ·h −1 for Nitrosomonas europaea ; μ max = 0.024 h −1 , b = 0.001 h −1 and m = 0.467 gN-NO 2 − ·gX −1 ·h −1 for Nitrobacter winogradskyi . A predictive structured model of nitrification in co-culture was developed. The online evolution of the addition of KOH is correlated to the nitritation; the dissolved oxygen concentration is correlated to both nitritation and nitratation. The model suitably represents these two variables so that transient partial nitrification is assessed. This is a clue for avoiding partial nitrification by predictive functional control.

Continuous cultures of Fibrobacter succinogenes S85 were performed on a standardized fully synthetic culture medium with glucose as carbon source at a dilution rate (D = 0.02 h(-1)) in a 5-L bioreactor. The culture was stabilized during 20 days and demonstrated the ability of Fibrobacter succinogenes to grow in this synthetic medium. CO(2) partial pressure and redox potential probes were used to check the anaerobic state of the culture. The biomass yield was calculated 0.206 g (g glucose)(-1) and the production yield of succinate, the major end-product, was 0.63 mol (mol glucose)(-1). The consistency of the experimental data was checked by proton and mass (C, N) balances. The results were satisfactory (90-110% recovery) leading to derive a stoichiometric equation representative of the growth on glucose. The stoichiometric coefficients were calculated using data reconciliation and linear algebra methods enabling to obtain a complete modeling of all conversion yields possible.

Fibrobacter succinogenes S85, a strictly anaerobic Gram-negative bacterium, was grown in continuous culture in a bioreactor at different dilution rates (0.02 to 0.092 h(-1)) on a fully synthetic culture medium with glucose as carbon source. Glucose and ammonium sulfate consumption, as well as biomass, succinate, acetate, formate, and carbohydrate production were regularly measured. The relevant biomass elemental compositions were established for each dilution rate. Robustness of the experimental information was checked by C and N mass balances estimation, which were satisfactory. A detailed overall stoichiometry analysis of the process, including all substrates and products of the culture, was proposed. Online and off-line parameters measured during the culture brought a large number of data which were weighted by their respective variance associated to the measured value. The material balance resulted in an overdetermined linear system of equations made of weighted relationships including experimental data, elemental balances (C, H, O, N, S, Na), and an additional constraint. The mass balances involved in stoichiometric equations were solved using data reconciliation and linear algebra methods to take into account error measurements. This methodology allowed to establish the overall stoichiometric equation for each dilution rate studied.

In life support systems, such as the MELiSSA (Micro-Ecological Life Support Alternative) project, developed by the European Space Agency, the aim is to understand and assemble artificial ecosystems for ensuring human subsistence in space. Fibrobacter succinogenes, an anaerobic bacterium, was used for the degradation of vegetable wastes produced in higher plants chambers, but the process does not allow the monitoring of biomass concentration and degradation rates. This study proposes a growth and a degradation monitoring technique using pressure measurements. First, volatile fatty acids (VFA) production was compared with biomass growth and with CO(2) production. The experiments were carried out in batch and fed-batch processes on glucose and on vegetables. The results have shown that a link could be established between VFA production, degradation rate and gas pressure measurements. Thus, the pressure could be used both as a relevant variable for online evaluation of biomass growth and of degradation of complex vegetable wastes.

An operational reactor has been designed for the fermentation of a pure culture of Fibrobacter succinogenes with the constraints of strict anaerobic condition. The process is controlled by measurements of pH, redox, temperature and CO2 pressure; it allows an efficient degradation (67%) of lignocellulosic wastes such as a mixture of wheat straw, soya bean cake and green cabbage.

National audience

2,5-Dimethylpyrazine (2,5-DMP) and tetramethylpyrazine (TTMP) were produced using Bacillus subtilis IFO 3013 grown on soybeans. Solid-state cultivations were carried out either in 100-ml bottles or in a fixed-bed column reactor, both systems being at 27 °C. Optimization studies showed that the best way to produce the two above aroma compounds involved two separate processes. 2,5-DMP was obtained using soybeans enriched with 75 g threonine/kg initial dry weight (i.d.w.), giving 0.85 g metabolite/kg i.d.w. after 6 days. TTMP production involved addition of 90 g/kg i.d.w. acetoin to soybeans, and 2.5 g/kg i.d.w. was recovered after 14 days. These results demonstrated the suitability of solid-state cultivation for production of high-added-value compounds.

The artificial ecosystem MELiSSA, supported by the European Space Agency is a closed loop system consisting of 5 compartments in which food, water and oxygen are produced out of organic waste. The first compartment is conceived as a thermophilic anaerobic membrane bioreactor liquefying organic waste into VFAs, ammonium and CO 2 without methane. A 20 L reactor was assembled to demonstrate the selected design and process at prototype scale. We characterized system performance from start-up to steady state and evaluated process efficiencies with special attention drawn to the mass balances. An overall efficiency for organic matter biodegradation of 50% was achieved. The dry matter content was stabilized around 40–50 g L −1 and VFA production around 5–6 g L −1 . The results were consistent for the considered substrate mixture and can also be considered relevant in a broader context, as a first processing step to produce building blocks for synthesis of primary energy vectors.

As a part of a natural biological N-cycle, nitrification is one of the steps included in the conception of artificial ecosystems designed for extraterrestrial life support systems (LSS) such as Micro-Ecological Life Support System Alternative (MELiSSA) project, which is the LSS project of the European Space Agency. Nitrification in aerobic environments is carried out by two groups of bacteria in a two-step process. The ammonia-oxidizing bacteria (Nitrosomonas europaea) realize the oxidation of ammonia to nitrite, and the nitrite-oxidizing bacteria (Nitrobacter winogradskyi), the oxidation of nitrite to nitrate. In both cases, the bacteria achieve these oxidations to obtain an energy and reductant source for their growth and maintenance. Furthermore, both groups also use CO2 predominantly as their carbon source. They are typically found together in ecosystems, and consequently, nitrite accumulation is rare. Due to the necessity of modeling accurately conversion yields and transformation rates to achieve a complete modeling of MELiSSA, the present study focuses on the experimental determination of nitrogen to biomass conversion yields. Kinetic and mass balance studies for axenic cultures of Nitrosomonas europaea and Nitrobacter winogradskyi in autotrophic conditions are performed. The follow-up of these cultures is done using flow cytometry for assessing biomass concentrations and ionic chromatography for ammonium, nitrite, and nitrate concentrations. A linear correlation is observed between cell count and optical density (OD) measurement (within a 10 % accuracy) validating OD measurements for an on-line estimation of biomass quantity even at very low biomass concentrations. The conversion between cell count and biomass concentration has been determined: 7.1 × 1012 cells g dry matter (DM)−1 for Nitrobacter and 6.3 × 1012 cells g DM−1 for Nitrosomonas. Nitrogen substrates and products are assessed redundantly showing excellent agreement for mass balance purposes and conversion yields determination. Although the dominant phenomena are the oxidation of NH 4 + into nitrite (0.95 mol mol N−1 for Nitrosomonas europaea within an accuracy of 3 %) and nitrite into nitrate (0.975 mol mol N−1 for Nitrobacter winogradskyi within an accuracy of 2 %), the Nitrosomonas europaea conversion yield is estimated to be 0.42 g DM mol N−1, and Nitrobacter winogradskyi conversion yield is estimated to be 0.27 g DM mol N−1. The growth rates of both strains appear to be dominated by the oxygen transfer into the experimental setups.

In the Micro-Ecological Life Support System Alternative (MELiSSA), the liquefying compartment is the first step in the loop; the fraction of organic wastes that can be further recycled in the loop is determined by the efficiency of liquefaction. The first compartment is colonized by mixed group of bacteria which digests wastes by anaerobic process (hydrolysis, acidogenesis) knowing that the production of methane is not favoured by low pH and high temperature. The semi-continuous feeding of solid wastes in a perfectly mixed tank leads to consider that the residence times of the different solid particles are distributed with a mean (average age of the particles) and a variance which depend on hydraulic characteristics of the reactor. In order to make the solid particles residence time a controllable variable, a solid particles-recycled reactor has been imagined and operated. However, this calls for the rationale study of the influence of solid particles residence time on global efficiency of the reactor. This is the aim of this study. A reactor (RuSiTec: Rumen Simulating Technique) operating with controlled residence time of solid particles has been designed and built. The influence of residence time and of the degradability of solid particles is tested with pure culture of a rumen bacteria, Fibrobacter succinogenes . The results show that the mean residence time of solid particle can be shortened by using this kind of device in comparison with perfectly mixed conditions. This leads to a significant reduction of the volume of the reactor maintaining the same percentage of waste degradation. Furthermore, the kinetics of waste degradation can be assessed, enabling to define digestible and recalcitrant fractions of wastes.

The production of 2-heptanone from octanoic acid may be performed by free and entrapped spores of Penicillium roquefortii in a water-organic solvent two phase system.An industrial, isoparafflnic solvent, i.e. Hydrosol IP 230 O.S., which may be considered as tetradecane, is well suited for the process. Activities nearly double those achieved with aqueous systems are observed using an initial fatty acid content in the organic layer close to 100 mM and a ratio of the volume of the organic phase to the total volume of the medium of 0.88. The presence of the solvent allows a better recovery of the metabolite by lowering its activity coefficient.Fed-batch experiments performed in an aerated, stirred reactor show that the bioconversion may proceed in the two-phase system for at least 300 h. These conditions allow conversion of 750 mM (108 g · 1-1) fatty acid, and production of 600 mM (68.5 g · 1-1) 2-heptanone.

Manmade ecosystems differ from their prototype biosphere by the principle of control. The Earth Biosphere is sustainable by stochastic control and very large time constants. By contrast, in a closed ecosystem such as the micro-ecological life support system alternative (MELiSSA system) developed by the European Space Agency for space exploration, a deterministic control is a prerequisite of sustainable existence. MELiSSA is an integrated sum of interconnected biological subsystems. On one hand, all unit operations in charge of the elementary functions constitutive of the entire life support system are studied until a thorough understanding and mathematical modelling. On the other hand, the systemic approach of complex, highly branched systems with feedback loops is performed. This leads to study in the same perspective, with the same degree of accuracy and with the same language, waste degradation, water recycling, atmosphere revitalisation and food production systems prior to the integration of knowledge-based control models. This paper presents the mathematical modelling of the MELiSSA system and the interface between the control strategy of the entire system and the control of the bioreactors.

Flux distribution for a wild and a mutant strain of Saccharomyces cerevisiae are compared and investigated in terms of metabolic flux calculation and thermodynamic analysis of central metabolism under anaerobic conditions. Starting from a redundant set of measured rates obtained from batch cultures on glucose or fructose as carbon source, an original data reconciliation technique associated with the calculation of metabolic flux is used. Comparative analysis of carbon split in the metabolic network for the mutant yeast strain lacking the glucose6P-dehydrogenase (CD101-1A) and for the reference wild strain (ATCC 7754) allows to conclude that the pentose phosphate is in priority devoted to its anabolic function rather than to the production of NADPH cofactors. This last function seems to be as well assumed by the specific NADP acetaldehyde dehydrogenase enzyme; this explains the significantly higher production of acetate by the mutant strain.

Fibrobacter succinogenes, an anaerobic bacteria is cultivated in a batch reactor. A detailed stoichiometric model of metabolism is developed. It includes all reactions of catabolic pathways that have been proved to exist and anabolic pathways. The measured conversion yields are correlated in terms of metabolic flux distribution using a mathematical technique of yields calculation associatedwith a methodology of data reconciliation. This approach validates the metabolic network built on Fibrobacter succinogenes.

The feasibility of nearly-complete conversion of lignocellulosic waste (70% food crops, 20% faecal matter and 10% green algae) into biogas was investigated in the context of a life support project. The treatment comprised a series of processes, i.e., a mesophilic laboratory scale CSTR (continuously stirred tank reactor), an upflow biofilm reactor, a fiber liquefaction reactor employing the rumen bacterium Fibrobacter succinogenes and a hydrothermolysis system in near-critical water. By the one-stage CSTR, a biogas yield of 75% with a specific biogas production of 0.37 l biogas g(-1) VSS (volatile suspended solids) added at a RT (hydraulic retention time) of 20-25 d was obtained. Biogas yields could not be increased considerably at higher RT, indicating the depletion of readily available substrate after 25 d. The solids present in the CSTR-effluent were subsequently treated in two ways. Hydrothermal treatment (T approximately 310-350 degrees C, p approximately 240 bar) resulted in effective carbon liquefaction (50-60% without and 83% with carbon dioxide saturation) and complete sanitation of the residue. Application of the cellulolytic Fibrobacter succinogenes converted remaining cellulose contained in the CSTR-effluent into acetate and propionate mainly. Subsequent anaerobic digestion of the hydrothermolysis and the Fibrobacter hydrolysates allowed conversion of 48-60% and 30%, respectively. Thus, the total process yielded biogas corresponding with conversions up to 90% of the original organic matter. It appears that particularly mesophilic digestion in conjunction with hydrothermolysis at near-critical conditions offers interesting features for (nearly) complete and hygienic carbon and energy recovery from human waste in a bioregenerative life support context.

A fed-batch technique for the production of 2-heptanone from octanoic acid by Ca-alginate/Eudragit RL-entrapped spores of Penicillium roquefortii is presented. It involves the use of a pH-stat apparatus, the increase in pH arising due to the reaction being overcome by direct substrate addition. When the reaction is performed at pH 5.5, no true constant rate period can be achieved due to a gradual buffering of the medium connected with a decrease in the catalytic activity of the spores. At pH 6.5 the system exhibits high stability and the reaction rate remains constant for at least 300 h with a value of 0.975 mM/h and a substrate concentration close to 4 mM. The process used may be very useful in the study of long-term behaviour of biocatalysts when the medium is not renewed and when the reaction involves an ionic compound and a neutral one.

International audience