Background: Elevated CO₂ partial pressure (pCO₂) has been proposed as a potential steering parameter for selective carboxylate production in mixed culture fermentation. It is anticipated that intermediate product spectrum and production rates, as well as changes in the microbial community, are (in)directly influenced by elevated pCO₂. However, it remains unclear how pCO₂ interacts with other operational conditions, namely substrate specificity, substrate-to-biomass (S/X) ratio and the presence of an additional electron donor, and what effect pCO₂ has on the exact composition of fermentation products. Here, we investigated possible steering effects of elevated pCO₂ combined with (1) mixed substrate (glycerol/glucose) provision; (2) subsequent increments in substrate concentration to increase the S/X ratio; and (3) formate as an additional electron donor. Results: Metabolite predominance, e.g., propionate vs. butyrate/acetate, and cell density, depended on interaction effects between pCO₂–S/X ratio and pCO₂–formate. Individual substrate consumption rates were negatively impacted by the interaction effect between pCO₂–S/X ratio and were not re-established after lowering the S/X ratio and adding formate. The product spectrum was influenced by the microbial community composition, which in turn, was modified by substrate type and the interaction effect between pCO₂–formate. High propionate and butyrate levels strongly correlated with Negativicutes and Clostridia predominance, respectively. After subsequent pressurized fermentation phases, the interaction effect between pCO₂–formate enabled a shift from propionate towards succinate production when mixed substrate was provided. Conclusions: Overall, interaction effects between elevated pCO₂, substrate specificity, high S/X ratio and availability of reducing equivalents from formate, rather than an isolated pCO₂ effect, modified the proportionality of propionate, butyrate and acetate in pressurized mixed substrate fermentations at the expense of reduced consumption rates and increased lag-phases. The interaction effect between elevated pCO₂ and formate was beneficial for succinate production and biomass growth with a glycerol/glucose mixture as the substrate. The positive effect may be attributed to the availability of extra reducing equivalents, likely enhanced carbon fixating activity and hindered propionate conversion due to increased concentration of undissociated carboxylic acids.

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A reliable drinking water supply and appropriate sanitation in the western world are the product of numerous innovations which addressed pressing historical challenges. This paper discusses that to tackle emerging water supply and sanitation challenges in the Global South, the innovation system cannot be a simple translation of the western “sustaining innovation” system. In this chapter, the authors claim that “frugal innovation” is not only an alternative for the Global South but also for societies concerned about resource overexploitation. The chapter provides various examples of incorporating “frugal thinking” into different water treatment and sanitation aspects in resource affluent and resource-constrained societies. The authors present life-support system engineering concepts as a source of inspiration to design frugal circular solutions. Through these solutions, independency (in time and space) from natural resources and waste valorization in all types of societies can be achieved.@en

Anaerobic processes such as Anaerobic Digestion (AD) and mixed culture fermentation (MCF) are important technologies in the bioeconomy context since they can be used to convert (waste) biomass feedstock into gaseous energy carriers and chemical commodities, theoretically without the use of any additional energy source. AD is a multi-step bioconversion process pursuing organic matter stabilization whose final product, i.e., biogas, can be used as an energy source. On the other hand, MCF employs open mixed cultures under non-sterilized conditions to produce carboxylates, i.e., short and medium-chain organic acids, which will serve as chemical building blocks after downstream processing. Limitations of biogas production are associated with the low CH4 content (≈50-60%), presence of impurities (like H2S) and unsuitable final pressure for direct connection to national grids. Thus, in recent years, the topic of biogas upgrading to biomethane (i.e., CH4>90%) has gained momentum and in-situ and ex-situ alternatives have been proposed with differences in financial and technical viability as well as achieved final CH4 content. While for the carboxylate production, major limitations are associated with process selectivity, presence of trace pollutants and too low broth concentrations for direct application inducing a need for “wet” and energy-intensive downstream processing. High-Pressure Anaerobic digestion (HPAD) is an innovative technology designed for simultaneous digestion and biogas upgrading. HPAD takes advantage of the large differences in solubility between biogas constituents, i.e., CH4 and CO2. Consequently, CH4 will predominantly remain in the gas phase after a pressure increase, whereas ionisable gases like CO2 and H2S will increasingly dissolve in the liquid. Thus, from a biogas production perspective, the proposed technology accomplishes higher CH4 content in the gas phase at the cost of increased dissolved CO2 levels. The process's overall performance under elevated pCO2 has not been adequately addressed. Mechanistic explanations for the role of increased dissolved CO2 in the fermentation process remain speculative, partially due to the limited amount of published experimental work on high-pressure fermentation with open cultures. Since CO2 exerts multiple roles in biological systems, increased dissolved CO2 could impact the kinetic and energetic feasibility of the reaction chain in AD and MCF, as well as the microbial community dynamics. These effects constitute a notorious knowledge gap that requires urgent attention… @en

Fermentation at elevated hydrostatic pressure is a novel strategy targeting product selectivity. However, the role of inoculum history and cross-resistance, that is, acquired tolerance from incubation under distinctive environmental stress, remains unclear in high-pressure operation. In our here presented work, we studied fermentation and microbial community responses of halotolerant marine sediment inoculum (MSI) and anaerobic digester inoculum (ADI), pre-incubated in serum bottles at different temperatures and subsequently exposed to mild hydrostatic pressure (MHP; < 10 MPa) in stainless steel reactors. Results showed that MHP effects on microbial growth, activity, and community structure were strongly temperature-dependent. At moderate temperature (20°C), biomass yield and fermentation were not limited by MHP; suggesting a cross-resistance effect from incubation temperature and halotolerance. Low temperatures (10°C) and MHP imposed kinetic and bioenergetic limitations, constraining growth and product formation. Fermentation remained favorable in MSI at 28°C and ADI at 37°C, despite reduced biomass yield resulting from maintenance and decay proportionally increasing with temperature. Microbial community structure was modified by temperature during the enrichment, and slight differences observed after MHP-exposure did not compromise functionality. Results showed that the relation incubation temperature—halotolerance proved to be a modifier of microbial responses to MHP and could be potentially exploited in fermentations to modulate product/biomass ratio.

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Background: Pressurised anaerobic digestion allows the production of biogas with a high content of methane and, at the same time, avoid the energy costs for the biogas upgrading and injection into the distribution grid. The technology carries potential, but the research faces practical constraints by a.o. the capital investment needed in high-pressure reactors and sensors and associated sampling limitations. In this work, the kinetic model of an autogenerative high-pressure anaerobic digestion of acetate, as the representative compound of the aceticlastic methanogenesis route, in batch configuration, is proposed to predict the dynamic performance of pressurised digesters and support future experimental work. The modelling of autogenerative high-pressure anaerobic digestion in batch configuration, which is not extensively studied and simulated in the present literature, was developed, calibrated, and validated by using experimental results available from the literature. Results: Under high-pressure conditions, the assessment of the Monod maximum specific uptake rate, the half-saturation constant and the first-order decay rate was carried out, and the values of 5.9 kg COD kg COD⁻¹ d⁻¹, 0.05 kg COD m⁻³ and 0.02 d⁻¹ were determined, respectively. By using the predicted values, excellent fittings of the final pressure, the CH₄ molar fraction and the specific methanogenic yield calculation were obtained. Likewise, the variation in the gas–liquid mass transfer coefficient by several orders of magnitude showed negligible effects on the model predictive values in terms of methane molar fraction of the produced biogas, while the final pressure seemed to be slightly influenced. Conclusions: The proposed model allowed to estimate the Monod maximum specific uptake rate for acetate, the half-saturation rate for acetate and the first-order decay rate constant, which were comparable with literature values reported for well-studied methanogens under anaerobic digestion at atmospheric pressure. The methane molar fraction and the final pressure predicted by the model showed different responses towards the variation of the gas–liquid mass transfer coefficient since the former seemed not to be affected by the variation of the gas–liquid mass transfer coefficient; in contrast, the final pressure seemed to be slightly influenced. The proposed approach may also allow to potentially identify the methanogens species able to be predominant at high pressure.

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Volatile fatty acid accumulation is a sign of digester perturbation. Previous work showed the thermodynamic limitations of hydrogen and CO₂ in syntrophic propionate oxidation under elevated partial pressure of CO₂ (pCO₂). Here we study the effect of directional selection under increasing substrate load as a strategy to restructure the microbial community and induce cross-protection mechanisms to improve glucose and glycerol conversion performance under elevated pCO₂. After an adaptive laboratory evolution (ALE) process, viable cell density increased and predominant microbial groups were modified: an increase in Methanosaeta and syntrophic propionate oxidizing bacteria (SPOB) associated with the Smithella genus was found with glycerol as the substrate. A modest increase in SPOB along with a shift in the predominance of Methanobacterium toward Methanosaeta was observed with glucose as the substrate. The evolved inoculum showed affected diversity within archaeal spp. under 5 bar initial pCO₂; however, higher CH₄ yield resulted from enhanced propionate conversion linked to the community shifts and biomass adaptation during the ALE process. Moreover, the evolved inoculum attained increased cell viability with glucose and a marginal decrease with glycerol as the substrate. Results showed differences in terms of carbon flux distribution using the evolved inoculum under elevated pCO₂: glucose conversion resulted in a higher cell density and viability, whereas glycerol conversion led to higher propionate production whose enabled conversion reflected in increased CH₄ yield. Our results highlight that limited propionate conversion at elevated pCO₂ resulted from decreased cell viability and low abundance of syntrophic partners. This limitation can be mitigated by promoting alternative and more resilient SPOB and building up biomass adaptation to environmental conditions via directional selection of microbial community.

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Small-scale electrical power generation (<100 kW) from biogas plants to provide off-grid electricity is of growing interest. Currently, gas engines are used to meet this demand. Alternatively, more efficient small-scale solid oxide fuel cells (SOFCs) can be used to enhance electricity generation from small-scale biogas plants. Most electricity generators require a constant gas supply and high gas quality in terms of absence of impurities like H ₂S. Therefore, to efficiently use the biogas from existing decentralized anaerobic digesters for electricity production, higher quality and stable biogas flow must be guaranteed. The installation of a biogas upgrading and buffer system could be considered; however, the cost implication could be high at a small scale as compared to locally available alterna-tives such as co-digestion and improved digester operation. Therefore, this study initially describes relevant literature related to feedstock pre-treatment, co-digestion and user operational practices of small-scale digesters, which theoretically could lead to major improvements of anaerobic digestion process efficiency. The theoretical preamble is then coupled to the results of a field study, which demonstrated that many locally available resources and user practices constitute frugal innovations with potential to improve biogas quality and digester performance in off-grid settings.

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Simultaneous digestion and in situ biogas upgrading in high-pressure bioreactors will result in elevated CO2 partial pressure (pCO2). With the concomitant increase in dissolved CO2, microbial conversion processes may be affected beyond the impact of increased acidity. Elevated pCO2 was reported to affect the kinetics and thermodynamics of biochemical conversions because CO2 is an intermediate and end-product of the digestion process and modifies the carbonate equilibrium. Our results showed that increasing pCO2 from 0.3 to 8 bar in lab-scale batch reactors decreased the maximum substrate utilization rate (rsmax) for both syntrophic propionate and butyrate oxidation. These kinetic limitations are linked to an increased overall Gibbs free energy change (ΔGOverall) and a potential biochemical energy redistribution among syntrophic partners, which showed interdependence with hydrogen partial pressure (pH2). The bioenergetics analysis identified a moderate, direct impact of elevated pCO2 on propionate oxidation and a pH-mediated effect on butyrate oxidation. These constraints, combined with physiological limitations on growth exerted by increased acidity and inhibition due to higher concentrations of undissociated volatile fatty acids, help to explain the observed phenomena. Overall, this investigation sheds light on the role of elevated pCO2 in delicate biochemical syntrophic conversions by connecting kinetic, bioenergetic, and physiological effects.

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