Acetic acid, growth rate, and mass transfer govern shifts in CO metabolism of Clostridium autoethanogenum.

Syngas fermentation is a leading microbial process for the conversion of carbon monoxide, carbon dioxide, and hydrogen to valuable biochemicals. Clostridium autoethanogenum stands as a model organism for this process, showcasing its ability to convert syngas into ethanol industrially with simultaneous fixation of carbon and reduction of greenhouse gas emissions. A deep understanding on the metabolism of this microorganism and the influence of operational conditions on fermentation performance is key to advance the technology and enhancement of production yields. In this work, we studied the individual impact of acetic acid concentration, growth rate, and mass transfer rate on metabolic shifts, product titres, and rates in CO fermentation by C. autoethanogenum. Through continuous fermentations performed at a low mass transfer rate, we measured the production of formate in addition to acetate and ethanol. We hypothesise that low mass transfer results in low CO concentrations, leading to reduced activity of the Wood-Ljungdahl pathway and a bottleneck in formate conversion, thereby resulting in the accumulation of formate. The supplementation of the medium with exogenous acetate revealed that undissociated acetic acid concentration increases and governs ethanol yield and production rates, assumedly to counteract the inhibition by undissociated acetic acid. Since acetic acid concentration is determined by growth rate (via dilution rate), mass transfer rate, and working pH, these variables jointly determine ethanol production rates. These findings have significant implications for process optimisation as targeting an optimal undissociated acetic acid concentration can shift metabolism towards ethanol production. KEY POINTS: • Very low CO mass transfer rate leads to leaking of intermediate metabolite formate. • Undissociated acetic acid concentration governs ethanol yield on CO and productivity. • Impact of growth rate, mass transfer rate, and pH were considered jointly.

Volatile Fatty Acids (VFA) Production and Recovery from Chicken Manure Using a High-Solid Anaerobic Membrane Bioreactor (AnMBR).

Acidogenic fermentation of chicken manure (CM) for production and recovery of volatile fatty acids (VFA) is an interesting biological waste-to-value approach compared to benchmark organic waste management strategies. Considering the wide range of high value applications of VFA, a semi-continuous immersed anaerobic membrane bioreactor (AnMBR) was applied to boost VFA productivity and yield, while reducing downstream processing stages assisting the recovery of VFA. In this regard, the effect of parameters such as pH and organic loading rates (OLR) on the overall bioconversion and filtration performance was investigated. Thermal-shocked CM was applied both as inoculum and substrate. A very high VFA yield (0.90 g-VFA/g-VS) was obtained in the treatment with no pH control (~8.2) at an OLR of 2 g-VS/(L·d), presenting 24% higher yield compared to that of the controlled pH. Batch assays further demonstrated the enhanced hydrolysis and acidogenesis activities at weak alkaline conditions. A long-term (78 days) fermentation and filtration was successfully performed, where stable membrane filtration performance was experienced for about 50 days under high-solid (suspended solid of 37-45 g/L) and high flux (20 L/(m2·h)) conditions. Results suggest that AnMBR of CM is a feasible and promising process for VFA production and recovery.

Membrane bioreactors for syngas permeation and fermentation.

Syngas fermentation to biofuels and chemicals is an emerging technology in the biobased economy. Mass transfer is usually limiting the syngas fermentation rate, due to the low aqueous solubilities of the gaseous substrates. Membrane bioreactors, as efficient gas-liquid contactors, are a promising configuration for overcoming this gas-to-liquid mass transfer limitation, so that sufficient productivity can be achieved. We summarize the published performances of these reactors. Moreover, we highlight numerous parameters settings that need to be used for the enhancement of membrane bioreactor performance. To facilitate this enhancement, we relate mass transfer and other performance indicators to the type of membrane material, module, and flow configuration. Hollow fiber modules with dense or asymmetric membranes on which biofilm might form seem suitable. A model-based approach is advocated to optimize their performance.

Effects of moderately elevated pressure on gas fermentation processes.

Industrial biotechnology has a potential to tackle harmful CO2 emissions and turn CO2 into a valuable commodity. However, a major technical obstacle in gas fermentations is the limited gas mass transfer rate. Increasing system pressure is a way to increase the driving force for mass transfer. This review presents critical aspects of gas fermentation at elevated pressure, with a specific focus on results obtained at 5-10 bar. While a solid foundation for high pressure fermentations has already been laid in the past, mainly to enhance oxygen transfer rates, it can be concluded that fermentations at moderately elevated pressures using gases such as CO2, CH4, CO, H2, O2 are still underexplored. Microbial growth rates and product formation can be improved at higher pressures, but in general, titers and productivities need to be increased to allow a further industrialization. Hence, more systematic investigations and techno-economic assessments are required.

Effect of the electric supply interruption on a microbial electrosynthesis system converting inorganic carbon into acetate.

Microbial electrosynthesis (MES) technology relies on the direct use of electrons to convert CO2 into long chain organic chemicals. Therefore, MES has been proposed to be coupled to the renewable electricity supply, mainly from solar and wind sources. However, those energies suffer fluctuations and interruptions of variable duration, which can have an adverse effect on MES performance. Such effects on MES has been evaluated for the first time under different interruptions time. H-cell-MES reactors were disconnected from power supply during 4, 6, 8, 16, 32 and 64 h. Interruptions affected the acetate production rate, causing a decrease of until 77% after 64 h off. However, after all the interruptions, the acetate production was restored, taking between 7 and 16 h for the reduction current to turn steady. Therefore, microbial community on MES proved to be resilient and able to recover the electro-autotrophic activity despite the duration of current supply interruptions.

Targeted poly(3-hydroxybutyrate-co-3-hydroxyvalerate) bioplastic production from carbon dioxide.

A microbial production process was developed to convert CO2 and valeric acid into tailored poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) bioplastics. The aim was to understand microbial PHBV production in mixotrophic conditions and to control the monomer distribution in the polymer. Continuous sparging of CO2 with pulse and pH-stat feeding of valeric acid were evaluated to produce PHBV copolyesters with predefined properties. The desired random monomer distribution was obtained by limiting the valeric acid concentration (below 1 gL-1). 1H-NMR, 13C-NMR and chromatographic analysis of the PHBV copolymer confirmed both the monomer distribution and the 3-hydroxyvalerate (3HV) fraction in the produced PHBV. A physical-based model was developed for mixotrophic PHBV production, which was calibrated and validated with independent experimental datasets. To produce PHBV with a predefined 3HV fraction, an operating diagram was constructed. This tool was able to predict the 3HV fraction with a very good accuracy (2% deviation).

Valorisation of CO2-rich off-gases to biopolymers through biotechnological process.

As one of the key enabling technologies, industrial biotechnology has a high potential to tackle harmful CO2 emissions and to turn CO2 into a valuable commodity. So far, experimental work mainly focused on the bioconversion of pure CO2 to chemicals and plastics and little is known about the tolerance of the bioprocesses to the presence of impurities. This work is the first to investigate the impact of real CO2-rich off-gases on autotrophic production of polyhydroxybutyrate. To this end, two-phase heterotrophic-autotrophic fermentation experiments were set up, consisting of heterothrophic cell mass growth using glucose as substrate followed by autotrophic biopolymer production using either pure synthetic CO2 or industrial off-gases sampled at two point sources. The use of real off-gases did not affect the bacterial performance. High biopolymer content (up to 73%) and productivities (up to 0.227 g/lh) were obtained. Characterisation of the polymers showed that all biopolymers had similar properties, independent of the CO2 source. Moreover, the CO2-derived biopolymers' properties were comparable to commercial ones and biopolymers reported in literature, which are all produced from organic carbon sources.

Continuous bioethanol fermentation from wheat straw hydrolysate with high suspended solid content using an immersed flat sheet membrane bioreactor.

Finding a technological approach that eases the production of lignocellulosic bioethanol has long been considered as a great industrial challenge. In the current study a membrane bioreactor (MBR) set-up using integrated permeate channel (IPC) membrane panels was used to simultaneously ferment pentose and hexose sugars to ethanol in continuous fermentation of high suspended solid wheat straw hydrolysate. The MBR was optimized to flawlessly operated at high SS concentrations of up to 20% without any significant changes in the permeate flux and transmembrane pressure. By the help of the retained high cell concentration, the yeast cells were capable of tolerating and detoxifying the inhibitory medium and succeeded to co-consume all glucose and up to 83% of xylose in a continuous fermentation mode leading to up to 83% of the theoretical ethanol yield.

Electro-Fermentation - Merging Electrochemistry with Fermentation in Industrial Applications.

Electro-fermentation (EF) merges traditional industrial fermentation with electrochemistry. An imposed electrical field influences the fermentation environment and microbial metabolism in either a reductive or oxidative manner. The benefit of this approach is to produce target biochemicals with improved selectivity, increase carbon efficiency, limit the use of additives for redox balance or pH control, enhance microbial growth, or in some cases enhance product recovery. We discuss the principles of electrically driven fermentations and how EF can be used to steer both pure culture and microbiota-based fermentations. An overview is given on which advantages EF may bring to both existing and innovative industrial fermentation processes, and which doors might be opened in waste biomass utilization towards added-value biorefineries.

Reverse membrane bioreactor: Introduction to a new technology for biofuel production.

The novel concept of reverse membrane bioreactors (rMBR) introduced in this review is a new membrane-assisted cell retention technique benefiting from the advantageous properties of both conventional MBRs and cell encapsulation techniques to tackle issues in bioconversion and fermentation of complex feeds. The rMBR applies high local cell density and membrane separation of cell/feed to the conventional immersed membrane bioreactor (iMBR) set up. Moreover, this new membrane configuration functions on basis of concentration-driven diffusion rather than pressure-driven convection previously used in conventional MBRs. These new features bring along the exceptional ability of rMBRs in aiding complex bioconversion and fermentation feeds containing high concentrations of inhibitory compounds, a variety of sugar sources and high suspended solid content. In the current review, the similarities and differences between the rMBR and conventional MBRs and cell encapsulation regarding advantages, disadvantages, principles and applications for biofuel production are presented and compared. Moreover, the potential of rMBRs in bioconversion of specific complex substrates of interest such as lignocellulosic hydrolysate is thoroughly studied.

Laboratory-scale membrane up-concentration and co-anaerobic digestion for energy recovery from sewage and kitchen waste.

This study assessed an alternative concept for co-treatment of sewage and organic kitchen waste in Vietnam. The goal was to apply direct membrane filtration for sewage treatment to generate a permeate that is suitable for discharge. The obtained chemical oxygen demand (COD) concentrations in the permeate of ultrafiltration tests were indeed under the limit value (50 mg/L) of the local municipal discharge standards. The COD of the concentrate was 5.4 times higher than that of the initial feed. These concentrated organics were then co-digested with organic kitchen wastes at an organic loading rate of 2.0 kg VS/m(3).d. The volumetric biogas production of the digester was 1.94 ± 0.34 m(3)/m(3).d. The recovered carbon, in terms of methane gas, accounted for 50% of the total carbon input of the integrated system. Consequently, an electrical production of 64 Wh/capita/d can be obtained when applying the proposed technology with the current wastes generated in Ho Chi Minh City. Thus, it is an approach with great potential in terms of energy recovery and waste treatment.

Biobutanol production from C5/C6 carbohydrates integrated with pervaporation: experimental results and conceptual plant design.

In this study, a simulated lignocellulosic hydrolyzate was used in a continuous two-stage fermentor setup for production of acetone, butanol and ethanol. An organophilic pervaporation unit was coupled to the second fermentor. The dilution rate in the first fermentor was kept constant at 0.109 h(-1), while the dilution rate in the second fermentor was gradually decreased from 0.056 to 0.020 h(-1). Glucose was completely consumed, while 61% of the xylose was consumed at the lowest dilution rate, leading to an overall solvent productivity of 0.65 g L(-1) h(-1) and a high concentration of 185 g kg(-1) solvents in the permeate in the last fermentation zone during 192 h. Based on the experimental results, a process integrated with organophilic pervaporation was conceptually designed and compared with a base-case. Chemcad simulations indicate an energy reduction of ~50% when organophilic pervaporation is used. This study also demonstrates significant reductions in process flows and energy consumption by the use of organophilic pervaporation as in situ product recovery technology.

Improvement of hydrogen production via ethanol-type fermentation in an anaerobic down-flow structured bed reactor.

Although a novel anaerobic down-flow structured bed reactor has shown feasibility and stable performance for a long-term compared to other anaerobic fixed bed systems for continuous hydrogen production, the volumetric rates and yields have so far been too low. In order to improve the performance, an operation strategy was applied by organic loading rate (OLR) variation (12-96 g COD L(-1) d(-1)). Different volumetric hydrogen rates, and yields at the same OLR indicated that the system was mainly driven by the specific organic load (SOL). When SOL was kept between 3.8 and 6.2 g sucrose g(-1) VSS d(-1), the volumetric rates raised from 0.1 to 8.9 L H2 L(-1) d(-1), and the yields were stable around 2.0 mol H2 mol(-1) converted sucrose. Furthermore, hydrogen was produced mainly via ethanol-type fermentation, reaching a total energy conversion rate of 23.40 kJ h(-1) L(-1) based on both hydrogen and ethanol production.

Effect of sodium accumulation on heterotrophic growth and polyhydroxybutyrate (PHB) production by Cupriavidus necator.

This study evaluates the effect of sodium (Na(+)) concentration on the growth and PHB production by Cupriavidus necator. Both biomass growth and PHB production were inhibited by Na(+): biomass growth became zero at 8.9 g/L Na(+) concentration while PHB production was completely stopped at 10.5 g/L Na(+). A mathematical model for pure culture heterotrophic PHB production was set up to describe the Na(+) inhibition effect. The parameters related to Na(+) inhibition were estimated based on shake flask experiments. The accumulated Na(+) showed non-linear inhibition effect on biomass growth but linear inhibition effect on PHB production kinetics. Fed-batch experiments revealed that a high accumulation of Na(+) due to a prolonged growth phase, using NaOH for pH control, decreased the subsequent PHB production. The model was validated based on independent experimental data sets, showing a good agreement between experimental data and simulation results.

Comparison of costs for three hypothetical alternative kitchen waste management systems.

Urban water and waste management continues to be a major challenge, with the Earth's population projected to rise to 9 billion by 2050, with 70% of this population expected to live in cities. A combined treatment of wastewater and the organic fraction of municipal solid waste offers opportunities for improved environmental protection and energy recovery, but the collection and transport of organic wastes must be cost effective. This study compares three alternative kitchen waste collection and transportation systems for a virtual modern urban area with 300,000 residents and a population density of 10,000 persons per square kilometre. Door-to-door collection, being the standard practice in modern urban centres, remains the most economically advantageous at a cost of 263 euros per tonne of kitchen waste. Important drawbacks are the difficult logistics, increased city traffic, air and noise pollution. The quieter, cleaner and more hygienic vacuum transport of kitchen waste comes with a higher cost of 367 euros per tonne, mainly resulting from a higher initial investment cost for the system installation. The third option includes the well-known use of under-sink food waste disposers (often called garbage grinders) that are connected to the kitchen's wastewater piping system, with a total yearly cost of 392 euros per tonne. Important advantages with this system are the clean operation and the current availability of a city-wide sewage conveyance pipeline system. Further research is recommended, for instance the application of a life cycle assessment approach, to more fully compare the advantages and disadvantages of each option.

Advances in in-situ product recovery (ISPR) in whole cell biotechnology during the last decade.

The review presents the state-of-the-art in the applications of in-situ product recovery (ISPR) in whole-cell biotechnology over the last 10years. It summarizes various ISPR-integrated fermentation processes for the production of a wide spectrum of bio-based products. A critical assessment of the performance of various ISPR concepts with respect to the degree of product enrichment, improved productivity, reduced process flows and increased yields is provided. Requirements to allow a successful industrial implementation of ISPR are also discussed. Finally, supporting technologies such as online monitoring, mathematical modeling and use of recombinant microorganisms with ISPR are presented.

Modeling pure culture heterotrophic production of polyhydroxybutyrate (PHB).

In this contribution a mechanistic model describing the production of polyhydroxybutyrate (PHB) through pure-culture fermentation was developed, calibrated and validated for two different substrates, namely glucose and waste glycerol. In both cases, non-growth-associated PHB production was triggered by applying nitrogen limitation. The occurrence of some growth-associated PHB production besides non-growth-associated PHB production was demonstrated, although it is inhibited in the presence of nitrogen. Other phenomena observed experimentally and described by the model included biomass growth on PHB and non-linear product inhibition of PHB production. The accumulated impurities from the waste substrate negatively affected the obtained maximum PHB content. Overall, the developed mathematical model provided an accurate prediction of the dynamic behavior of heterotrophic biomass growth and PHB production in a two-phase pure culture system.

Integrated conversion of food waste diluted with sewage into volatile fatty acids through fermentation and electricity through a fuel cell.

In this study, domestic wastewater was given a second life as dilution medium for concentrated organic waste streams, in particular artificial food waste. A two-step continuous process with first volatile fatty acid (VFA)/hydrogen production and second electricity production in microbial fuel cells (MFCs) was employed. For primary treatment, bioreactors were optimized to produce hydrogen and VFAs. Hydrolysis of the solids and formation of fermentation products and hydrogen was monitored. In the second step, MFCs were operated batch-wise using the effluent rich in VFAs specifically acetic acid from the continuous reactor of the first step. The combined system was able to reduce the chemical oxygen demand load by 90%. The concentration of VFAs was also monitored regularly in the MFCs and showed a decreasing trend over time. Further, the anode potential changed from -500 to OmV vs. Ag/AgCl when the VFAs (especially acetate) were depleted in the system. On feeding the system again with the effluent, the anode potential recovered back to -500 mV vs. Ag/AgCl. Thus, the overall aim of converting chemical energy into electrical energy was achieved with a columbic efficiency of 46% generating 65.33 mA/m2 at a specific cell potential of 148 mV.

Valorization of cereal based biorefinery byproducts: reality and expectations.

The growth of the biobased economy will lead to an increase in new biorefinery activities. All biorefineries face the regular challenges of efficiently and economically treating their effluent to be compatible with local discharge requirements and to minimize net water consumption. The amount of wastes resulting from biorefineries industry is exponentially growing. The valorization of such wastes has drawn considerable attention with respect to resources with an observable economic and environmental concern. This has been a promising field which shows great prospective toward byproduct usage and increasing value obtained from the biorefinery. However, full-scale realization of biorefinery wastes valorization is not straightforward because several microbiological, technological and economic challenges need to be resolved. In this review we considered valorization options for cereals based biorefineries wastes while identifying their challenges and exploring the opportunities for future process.

Test of an anaerobic prototype reactor coupled with a filtration unit for production of VFAs.

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 CO2 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.