Lactic acid production from food waste using a microbial consortium: Focus on key parameters for process upscaling and fermentation residues valorization.

In this study, the production of lactic acid from food waste in industrially relevant conditions was investigated. Laboratory assays were first performed in batch conditions to determine the suitable operational parameters for an efficient lactic acid production. The use of compost as inoculum, the regulation of temperature at 35 °C and pH at 5 enhanced the development of Lactobacillus sp. resulting in the production of 70 g/L of lactic acid with a selectivity of 89% over the other carboxylic acids. Those parameters were then applied at pilot scale in successive fed-batch fermentations. The subsequent high concentration (68 g/L), yield (0.38 g/gTS) and selectivity (77%) in lactic acid demonstrated the applicability of the process. To integrate the process into a complete value chain, fermentation residues were then converted into biogas through anaerobic digestion. Lastly, the experiment was successfully replicated using commercial and municipal waste collected in France.

Robust operation through effluent recycling for hydrogen production from the organic fraction of municipal solid waste.

The stability of fermentative hydrogen production from the organic fraction of municipal solid waste (OFMSW) was evaluated in this work using a strategy of effluent recycling. Three pretreatment conditions were applied on the recycled effluent: a) no heat shock treatment, b) one initial heat shock treatment (90 °C, 30 min) and c) systematic heat shock treatment at the beginning of each fermentation. When a systematic heat shock was applied, a maximal hydrogen yield of 17.2 ± 3.8 mLH2/gVS was attained. The hydrogen productivity was improved by 331% reaching a stable value of 1.51 ± 0.29 mLH2/gVS/h, after 8 cycles of effluent recycling. This strategy caused a sharp decrease of diversity with stable co-dominance of hydrogen- and lactate-producing bacteria, ie. Clostridiales and Lactobacillales, respectively. For the other conditions, a sharp decrease of the hydrogen yields was observed showing the importance of applying a heat shock treatment for optimal hydrogen production with effluent recycling.

Temperature and Inoculum Origin Influence the Performance of Ex-Situ Biological Hydrogen Methanation.

The conversion of H2 into methane can be carried out by microorganisms in a process so-called biomethanation. In ex-situ biomethanation H2 and CO2 gas are exogenous to the system. One of the main limitations of the biomethanation process is the low gas-liquid transfer rate and solubility of H2 which are strongly influenced by the temperature. Hydrogenotrophic methanogens that are responsible for the biomethanation reaction are also very sensitive to temperature variations. The aim of this work was to evaluate the impact of temperature on batch biomethanation process in mixed culture. The performances of mesophilic and thermophilic inocula were assessed at 4 temperatures (24, 35, 55 and 65 °C). A negative impact of the low temperature (24 °C) was observed on microbial kinetics. Although methane production rate was higher at 55 and 65 °C (respectively 290 ± 55 and 309 ± 109 mL CH4/L.day for the mesophilic inoculum) than at 24 and 35 °C (respectively 156 ± 41 and 253 ± 51 mL CH4/L.day), the instability of the system substantially increased, likely because of a strong dominance of only Methanothermobacter species. Considering the maximal methane production rates and their stability all along the experiments, an optimal temperature range of 35 °C or 55 °C is recommended to operate ex-situ biomethanation process.

Biomethanation processes: new insights on the effect of a high H2 partial pressure on microbial communities.

BACKGROUND: Biomethanation is a promising solution to upgrade the CH4 content in biogas. This process consists in the injection of H2 into an anaerobic digester, using the capacity of indigenous hydrogenotrophic methanogens for converting the injected H2 and the CO2 generated from the anaerobic digestion process into CH4. However, the injection of H2 could cause process disturbances by impacting the microbial communities of the anaerobic digester. Better understanding on how the indigenous microbial community can adapt to high H2 partial pressures is therefore required. RESULTS: Seven microbial inocula issued from industrial bioprocesses treating different types of waste were exposed to a high H2 partial pressure in semi-continuous reactors. After 12 days of operation, even though both CH4 and volatile fatty acids (VFA) were produced as end products, one of them was the main product. Acetate was the most abundant VFA, representing up to 94% of the total VFA production. VFA accumulation strongly anti-correlated with CH4 production according to the source of inoculum. Three clusters of inocula were distinguished: (1) inocula leading to CH4 production, (2) inocula leading to the production of methane and VFA in a low proportion, and (3) inocula leading to the accumulation of mostly VFA, mainly acetate. Interestingly, VFA accumulation was highly correlated to a low proportion of archaea in the inocula, a higher amount of homoacetogens than hydrogenotrophic methanogens and, the absence or the very low abundance in members from the Methanosarcinales order. The best methanogenic performances were obtained when hydrogenotrophic methanogens and Methanosarcina sp. co-dominated all along the operation. CONCLUSIONS: New insights on the microbial community response to high H2 partial pressure are provided in this work. H2 injection in semi-continuous reactors showed a significant impact on microbial communities and their associated metabolic patterns. Hydrogenotrophic methanogens, Methanobacterium sp. or Methanoculleus sp. were highly selected in the reactors, but the presence of co-dominant Methanosarcinales related species were required to produce higher amounts of CH4 than VFA.

Bioaugmentation enhances dark fermentative hydrogen production in cultures exposed to short-term temperature fluctuations.

Hydrogen-producing mixed cultures were subjected to a 48-h downward or upward temperature fluctuation from 55 to 35 or 75 °C. Hydrogen production was monitored during the fluctuations and for three consecutive batch cultivations at 55 °C to evaluate the impact of temperature fluctuations and bioaugmentation with synthetic mixed culture of known H2 producers either during or after the fluctuation. Without augmentation, H2 production was significantly reduced during the downward temperature fluctuation and no H2 was produced during the upward fluctuation. H2 production improved significantly during temperature fluctuation when bioaugmentation was applied to cultures exposed to downward or upward temperatures. However, when bioaugmentation was applied after the fluctuation, i.e., when the cultures were returned to 55 °C, the H2 yields obtained were between 1.6 and 5% higher than when bioaugmentation was applied during the fluctuation. Thus, the results indicate the usefulness of bioaugmentation in process recovery, especially if bioaugmentation time is optimised.

Assessing practical identifiability during calibration and cross-validation of a structured model for high-solids anaerobic digestion.

High-solids anaerobic digestion (HS-AD) of the organic fraction of municipal solid waste (OFMSW) is operated at a total solid (TS) content ≥ 10% to enhance the waste treatment economy, though it might be associated to free ammonia (NH3) inhibition. This study aimed to calibrate and cross-validate a HS-AD model for homogenized reactors in order to assess the effects of high NH3 levels in HS-AD of OFMSW, but also to evaluate the suitability of the reversible non-competitive inhibition function to reproduce the effect of NH3 on the main acetogenic and methanogenic populations. The practical identifiability of structural/biochemical parameters (i.e. 35) and initial conditions (i.e. 32) was evaluated using batch experiments at different TS and/or inoculum-to-substrate ratios. Variance-based global sensitivity analysis and approximate Bayesian computation were used for parameter optimization. The experimental data in this study permitted to estimate up to 8 biochemical parameters, whereas the rest of parameters and biomass contents were poorly identifiable. The study also showed the relatively high levels of NH3 (i.e. up to 2.3 g N/L) and ionic strength (i.e. up to 0.9 M) when increasing TS in HS-AD of OFMSW. However, the NH3 non-competitive function was unable to capture the acetogenic/methanogenic inhibition. Therefore, the calibration emphasized the need for target-oriented experimental data to enhance the practical identifiability and the predictive capabilities of structured HS-AD models, but also the need for further testing the NH3 inhibition function used in these simulations.

Modelling non-ideal bio-physical-chemical effects on high-solids anaerobic digestion of the organic fraction of municipal solid waste.

This study evaluates the main effects of including 'non-ideal' bio-physical-chemical corrections in high-solids anaerobic digestion (HS-AD) of the organic fraction of municipal solid waste (OFMSW), at total solid (TS) between 10 and 40%. As a novel approach, a simple 'non-ideal' module, accounting for the effects of ionic strength (I) on the main acid-base equilibriums, was coupled to a HS-AD model, to jointly evaluate the effects of 'non-ideality' and the TS content dynamics on the HS-AD bio-physical-chemistry. 'Non-ideality' influenced the pH, concentration of inhibitors (i.e. NH3), and liquid-gas transfer (i.e. CO2), particularly at higher TS (i.e. ≥ 20%). Meanwhile, fitting the experimental data for batch assays at 15% TS showed that HS-AD of OFMSW might be operated at I ≥ 0.5 M. Therefore, all HS-AD simulations should account for 'non-ideal' corrections, when assessing the main inhibitory mechanisms (i.e. NH3 buildup and acidification) potentially occurring in HS-AD of OFMSW.

Solid-state anaerobic digestion of wheat straw: Impact of S/I ratio and pilot-scale fungal pretreatment.

Solid State Anaerobic Digestion (SSAD) of fungal pretreated wheat straw was evaluated in a leach bed reactor. During a first experiment, the effect of Substrate/Inoculum (S/I) ratios on the start-up phase was investigated. High S/I increased methane productivity but also raised the risk of reactor failure due to Volatile Fatty Acid (VFA) accumulation. With S/I ratios between 1.2 and 3.6 (Volatile Solid (VS) basis), the SSAD start-up using wheat straw was successful. Moreover, reactors were able to recover from acidification when the Total VFA/alkalinity ratio was lower than 2 gHAc_eq/gCaCO3, with VFA concentrations lower than 10 g/L and a pH close to 5.5. The conventional threshold of 0.6 gHAc_eq/gCaCO3 for stable wet AD is therefore not adapted to SSAD. During a second experiment, after the wheat straw was submitted to a fungal pretreatment in a non-sterile pilot-scale reactor, it was digested with an S/I ratio of 2.8-2.9. Under batch SSAD conditions, the biodegradability of pretreated wheat straw was slightly improved in comparison to the control (254 versus 215 NmL/g VS, respectively). Considering mass losses occurring during the pretreatment step, suboptimal pretreatment conditions caused a slightly lower methane production (161 versus 171 NmL/gTSinitial after 60-days anaerobic digestion). Nevertheless, pretreatment improved the start-up phase with lower acidification relative to controls. It would be particularly beneficial to improve the methane production in reactors with short reaction times.

Reversibility of hydrolysis inhibition at high hydrogen partial pressure in dry anaerobic digestion processes fed with wheat straw and inoculated with anaerobic granular sludge.

In dry anaerobic digestion (AD), methanogenic performances are lowered by high solid contents. Low performances are often caused by a decrease of the gas-liquid transfer kinetics leading to local accumulation of inhibitory by-products. Hydrogen was previously identified as an inhibitor of hydrolytic and acetogenic microbial activities in dry AD. CO2 is also generated but its impact on the microbial activity remains unknown. In this study, the reversibility of dry AD inhibition at high H2 partial pressure (PH2 of 1 bar) was investigated by adding CO2 (400 mbars) after 11 and 18 days of methanogenesis inhibition, in an AD process operated at 25% TS, using wheat straw as substrate and inoculated with anaerobic granular sludge. As soon as CO2 was added, the methanogenic activity rapidly recovered within 3 days, from 0.41 ±â€¯0.1 to 3.77 ±â€¯0.8 and then 2.25 ±â€¯0.3, likely through the hydrogenotrophic pathway followed by the acetoclastic pathway, respectively. This result was confirmed by the high abundance of Methanomicrobiales (83%) and the emergence of Methanosarcinales sp (up to 17%) within the methanogenic community. Furthermore, the recovery kinetics were impacted by the duration of the inhibition period suggesting a different impact of the high PH2 on hydrogenotrophic and acetoclastic methanogens.

Effect of ammonium, electron donor and sulphate transient feeding conditions on sulphidogenesis in sequencing batch bioreactors.

This work aimed to study the effect of transient feeding conditions on sulphidogenesis in 8 sequencing batch bioreactors (SBR). SBR L1 and H1, operated under steady-state conditions were used as the control reactors, while four SBR were tested under transient feeding conditions using moderate (L2 and L3, feast and famine: 2.5 and 0 g SO42-·L-1) and high (H2 and H3, feast and famine: 15 and 0 g SO42-·L-1) loads. The sulphate removal efficiency (RE) was ≥90% in SBR L2, L3 and H1. The NH4+ famine conditions resulted in a higher sulphate RE (≥40% H3) compared to feast conditions (≤20% H2). Besides, the sulphidogenic first-order kinetic constant was 4% larger and the use of electron donor was 16.6% more efficient under NH4+ famine conditions. Sulphidogenesis is robust to transient feeding conditions, but not when applying high loading rates (SBR H2 and H3).

Enhancement of corn stover conversion to carboxylates by extrusion and biotic triggers in solid-state fermentation.

Solid-state fermentation is a potential technology for developing lignocellulosic biomass-based biorefineries. This work dealt with solid-state fermentation for carboxylates production from corn stover, as building blocks for a lignocellulosic feedstock-based biorefinery. The effect of extrusion pretreatment, together with the action of a microbial consortia and hydrolytic enzymes as biotic triggers, was investigated on corn stover conversion, microbial metabolic pathways, and populations. The extrusion caused changes in the physical and morphological characteristics, without altering the biochemical composition of the corn stover. Extrusion also led to remarkable differences in the composition of the indigenous microbial population of the substrate. Consequently, it affected the structure of community developed after fermentation and the substrate conversion yield, which increased by 118% (from 23 ± 4 gCOD/kgVSi obtained with raw substrate to 51 ± 1 gCOD/kgVSi with extruded corn stover) with regard to self-fermentation experiments. The use of activated sludge as inoculum further increased the total substrate conversion into carboxylates, up to 60 ± 2 gCOD/kgVSi, and shaped the microbial communities (mainly composed of bacteria from the Clostridia and Bacteroidia classes) with subsequent homogenization of the fermentation pathways. The addition of hydrolytic enzymes into the reactors further increased the corn stover conversion, leading to a maximum yield of 142 ± 1 gCOD/kgVSi. Thus, extrusion pretreatment combined with the use of an inoculum and enzyme addition increased by 506% corn stover conversion into carboxylates. Beside biomass pretreatment, the results of this study indicated that biotic factor greatly impacted solid-state fermentation by shaping the microbial communities and related metabolic pathways.

Semi-continuous mono-digestion of OFMSW and Co-digestion of OFMSW with beech sawdust: Assessment of the maximum operational total solid content.

In this study, mono-digestion of the organic fraction of municipal solid waste (OFMSW) and co-digestion of OFMSW with beech sawdust, simulating green waste, were used to investigate the maximum operational total solid (TS) content in semi-continuous high-solids anaerobic digestion (HS-AD). To alleviate substrate overloading in HS-AD, the effluent mass was relatively reduced compared to the influent mass, extending the mass retention time. To this aim, the reactor mass was daily evaluated, permitting to assess the reactor content removal by biogas production. During mono-digestion of OFMSW, the NH3 inhibition and the rapid TS removal prevented to maintain HS-AD conditions (i.e. TS ≥ 10%), without exacerbating the risk of reactor acidification. In contrast, the inclusion of sawdust in OFMSW permitted to operate HS-AD up to 30% TS, before acidification occurred. Therefore, including a lignocellulosic substrate in OFMSW can prevent acidification and stabilize HS-AD at very high TS contents (i.e. 20-30%).

Lime Pretreatment of Miscanthus: Impact on BMP and Batch Dry Co-Digestion with Cattle Manure.

In Europe, the agricultural biogas sector is currently undergoing fast developments, and cattle manure constitutes an important feedstock. Batch dry digester processes with leachate recirculation prove to be particularly interesting for small-scale plants. However, their startup being relatively slow, the process could be facilitated by co-digestion with energy crops. In this study, Miscanthus xgiganteus was chosen for its high biomass yields and low input requirements. The carbohydrate accessibility of this lignocellulosic biomass is limited but may be improved with alkali pretreatment. The efficiency of lime (CaO) pretreatment with low water addition on the biochemical methane potential (BMP) of miscanthus was investigated through two experimental designs (CaO concentrations ranged between 2.5 and 17.5% and pretreatment lasted 1, 3, or 5 days). The pretreated miscanthus was then co-digested with cattle manure in dry leach bed reactors. CaO pretreatments led to a 14⁻37% improvement of miscanthus BMP, and a 67⁻227% increase in the first-order kinetics constant; a high contact time was shown to favor methane production. According to these results and to industrial requirements, miscanthus was pretreated with 5 and 10% CaO for 5 days, then co-digested with manure in dry leach bed reactors. Nevertheless, the promising results of the BMP tests were not validated. This could be related to the high water absorption capacity of miscanthus.

High-solids anaerobic digestion model for homogenized reactors.

During high-solids anaerobic digestion (HS-AD) of the organic fraction of municipal solid waste (OFMSW), an important total solid (TS) removal occurs, leading to the modification of the reactor content mass/volume, in contrast to 'wet' anaerobic digestion (AD). Therefore, HS-AD mathematical simulations need to be approached differently than 'wet' AD simulations. This study aimed to develop a modelling tool based on the anaerobic digestion model 1 (ADM1) capable of simulating the TS and the reactor mass/volume dynamics in HS-AD of OFMSW. Four hypotheses were used, including the effects of apparent concentrations at high TS. The model simulated adequately HS-AD of OFMSW in batch and continuous mode, particularly the evolution of TS, reactor mass, ammonia and volatile fatty acids. By adequately simulating the reactor content mass/volume and the TS, the HS-AD model might bring further insight about potentially inhibitory mechanisms (i.e. NH3 buildup and/or acidification) occurring in HS-AD of OFMSW.

Addition of granular activated carbon and trace elements to favor volatile fatty acid consumption during anaerobic digestion of food waste.

The effect of supplementing granular activated carbon and trace elements on the anaerobic digestion performance of consecutive batch reactors treating food waste was investigated. The results from the first batch suggest that addition of activated carbon favored biomass acclimation, improving acetic acid consumption and enhancing methane production. Adding trace elements allowed a faster consumption of propionic acid. A second batch proved that a synergy existed when activated carbon and trace elements were supplemented simultaneously. The degradation kinetics of propionate oxidation were particularly improved, reducing significantly the batch duration and improving the average methane productivities. Addition of activated carbon favored the growth of archaea and syntrophic bacteria, suggesting that interactions between these microorganisms were enhanced. Interestingly, microbial analyses showed that hydrogenotrophic methanogens were predominant. This study shows for the first time that addition of granular activated carbon and trace elements may be a feasible solution to stabilize food waste anaerobic digestion.

Methanosarcina plays a main role during methanogenesis of high-solids food waste and cardboard.

Anaerobic digestion of food waste is a complex process often hindered by high concentrations of volatile fatty acids and ammonia. Methanogenic archaea are more sensitive to these inhibitors than bacteria and thus the structure of their community is critical to avoid reactor acidification. In this study, the performances of three different inocula were compared using batch digestion tests of food waste and cardboard mixtures. Particular attention was paid to the archaeal communities in the inocula and after digestion. While the tests started with inocula rich in Methanosarcina led to efficient methane production, VFAs accumulated in the reactors where inocula initially were poor in this archaea and no methane was produced. In addition, higher substrate loads were tolerated when greater proportions of Methanosarcina were initially present in the inoculum. Independently of the inoculum origin, Methanosarcina were the dominant methanogens in the digestates from the experiments that efficiently produced methane. These results suggest that the initial archaeal composition of the inoculum is crucial during reactor start-up to achieve stable anaerobic digestion at high concentrations of ammonia and organic acids.

Effect of total solids content on biohydrogen production and lactic acid accumulation during dark fermentation of organic waste biomass.

Production of biohydrogen and related metabolic by-products was investigated in Solid State Dark Fermentation (SSDF) of food waste (FW) and wheat straw (WS). The effect of the total solids (TS) content and H2 partial pressure (ppH2), two of the main operating factors of SSDF, were investigated. Batch tests with FW at 10, 15, 20, 25 and 30% TS showed considerable effects of the TS on metabolites distribution. H2 production was strongly inhibited for TS contents higher than 15% with a concomitant accumulation of lactic acid and a decrease in substrate conversion. Varying the ppH2 had no significant effect on the conversion products and overall degradation of FW and WS, suggesting that ppH2 was not the main limiting factor in SSDF. This study showed that the conversion of complex substrates by SSDF depends on the substrate type and is limited by the TS content.

Cardboard proportions and total solids contents as driving factors in dry co-fermentation of food waste.

This study evaluated the influence of the co-substrate proportions (0-60% of cardboard in dry basis) and the initial total solid contents (20-40%) on the batch fermentation performance. Maximum hydrogen yields were obtained when mono-fermenting food waste at high solids contents (89mlH2·gVS-1). The hydrogen yields were lower when increasing the proportions of cardboard. The lower hydrogen yields at higher proportions of cardboard were translated into higher yields of caproic acid (up to 70.1gCOD·kgCODbio-1), produced by consumption of acetic acid and hydrogen. The highest substrate conversions were achieved at low proportions of cardboard, indicating a stabilization effect due to higher buffering capacities in co-fermentation. Clostridiales were predominant in all operational conditions. This study opens up new possibilities for using the cardboard proportions for controlling the production of high added-value products in dry co-fermentation of food waste.

Co-ensiling as a new technique for long-term storage of agro-industrial waste with low sugar content prior to anaerobic digestion.

Biodegradable wastes produced seasonally need an upstream storage, because of the requirement for a constant feeding of anaerobic digesters. In the present article, the potential of co-ensiling biodegradable agro-industrial waste (sugar beet leaves) and lignocellulosic agricultural residue (wheat straw) to obtain a mixture with low soluble sugar content was evaluated for long-term storage prior to anaerobic digestion. The aim is to store agro-industrial waste while pretreating lignocellulosic biomass. The dynamics of co-ensiling was evaluated in vacuum-packed bags at lab-scale during 180 days. Characterization of the reaction by-products and microbial communities showed a succession of metabolic pathways. Even though the low initial sugars content was not sufficient to lower the pH under 4.5 and avoid undesirable fermentations, the methane potential was not substantially impacted all along the experiment. No lignocellulosic damages were observed during the silage process. Overall, it was shown that co-ensiling was effective to store highly fermentable fresh waste evenly with low sugar content and offers new promising possibilities for constant long-term supply of industrial anaerobic digesters.

Kinetic study of dry anaerobic co-digestion of food waste and cardboard for methane production.

Dry anaerobic digestion is a promising option for food waste treatment and valorization. However, accumulation of ammonia and volatile fatty acids often occurs, leading to inefficient processes and digestion failure. Co-digestion with cardboard may be a solution to overcome this problem. The effect of the initial substrate to inoculum ratio (0.25 to 1gVS·gVS-1) and the initial total solids contents (20-30%) on the kinetics and performance of dry food waste mono-digestion and co-digestion with cardboard was investigated in batch tests. All the conditions produced methane efficiently (71-93% of the biochemical methane potential). However, due to lack of methanogenic activity, volatile fatty acids accumulated at the beginning of the digestion and lag phases in the methane production were observed. At increasing substrate to inoculum ratios, the initial acid accumulation was more pronounced and lower cumulative methane yields were obtained. Higher amounts of soluble organic matter remained undegraded at higher substrate loads. Although causing slightly longer lag phases, high initial total solids contents did not jeopardize the methane yields. Cardboard addition reduced acid accumulation and the decline in the yields at increasing substrate loads. However, cardboard addition also caused higher concentrations of propionic acid, which appeared as the most last acid to be degraded. Nevertheless, dry co-digestion of food waste and cardboard in urban areas is demonstrated asan interesting feasible valorization option.