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Livestock waste is stored and used as soil fertilizer or directly as substrate for biogas production. Methane emissions from manure storages and ammonia inhibition of anaerobic digesters fed with manure, are well-known problems related to manure management. This study examines the effect of adding tannic acid with fluoride (TA-NaF) and lignosulfonic acid (LS) on methanogenic activity in batch reactors with ammonia inhibited maize silage digestate and in batch reactors with manure. Lignosulfonic acid counteracted urea induced ammonia inhibition of methanogenesis, whereas TA-NaF inhibited methanogenesis itself. Stable carbon isotope ratio analysis and methanogen community analysis suggested that TA-NaF affected acetoclastic methanogens the most. The combined findings suggest that TA-NaF could be used to reduce methane emissions from stored manure. Conversely, LS could be used as supplement in anaerobic digesters prone to urea induced ammonia inhibition.

Biomethanation is a promising solution to convertH2 (produced fromsurplus electricity) and CO2 to CH4 by using hydrogenotrophic methanogens. In ex situ biomethanation with mixed cultures, homoacetogens and methanogens compete for H2/CO2. We enriched a hydrogenotrophic microbiota on CO2 and H2 as sole carbon and energy sources, respectively, to investigate these competing reactions. The microbial community structure and dynamics of bacteria and methanogenic archaea were evaluated through 16S rRNA and mcrA gene amplicon sequencing, respectively. Hydrogenotrophic methanogens and homoacetogens were enriched, as acetate was concomitantly produced alongside CH4. By controlling the media composition, especially changing the reducing agent, the formation of acetate was lowered and grid quality CH4 (≥97%) was obtained. Formate was identified as an intermediate that was produced and consumed during the bioprocess. Stirring intensities ≥ 1000 rpm were detrimental, probably due to shear force stress. The predominating methanogens belonged to the genera Methanobacterium and Methanoculleus. The bacterial community was dominated byLutispora. The methanogenic community was stable, whereas the bacterial community was more dynamic. Our results suggest that hydrogenotrophic communities can be steered towards the selective production of CH4 from H2/CO2 by adapting the media composition, the reducing agent and the stirring intensity.

Sugarcane is the most produced agricultural commodity in tropical and subtropical regions, where it is primarily used for the production of sugar and ethanol. The latter is mostly used to produce alcoholic beverages as well as low carbon biofuel. Despite well-established production chains, their respective residues and by-products present unexploited potentials for further product portfolio diversification. These fully or partially untapped product streams are (a) sugarcane trash or straw that usually remain on the fields after mechanized harvest, (b) ashes derived from bagasse combustion in cogeneration plants, (c) filter cake from clarification of the sugarcane juice, (d) vinasse which is the liquid residue after distillation of ethanol, and (e) biogenic CO2 emitted during bagasse combustion and ethanol fermentation. The development of innovative cascading processes using these residual biomass fractions could significantly reduce final disposal costs, improve the energy output, reduce greenhouse gas emissions, and extend the product portfolio of sugarcane mills. This study reviews not only the state-of-the-art sugarcane biorefinery concepts, but also proposes innovative ways for further valorizing residual biomass. This study is therefore structured in four main areas, namely: (i) Cascading use of organic residues for carboxylates, bioplastic, and bio-fertilizer production, (ii) recovery of unexploited organic residues via anaerobic digestion (AD) to produce biogas, (iii) valorization of biogenic CO2 sources, and (iv) recovery of silicon from bagasse ashes.

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Cover cropping is important for nutrient management of agricultural systems and has largely unexploited potential for biogas production. Co-harvest of cover crops and straw and storage as silage blends prior to biogas is promising as it could enhance the long-term storability and lead to synergies for anaerobic digestion. However, it is necessary to evaluate the feasibility of using co-ensiled blends and process optimization based on continuous test. In this study, semi-continuous thermophilic anaerobic digestion experiments were carried out with feeding of cover crop silage, co-ensiled cover crop and straw (with mixing ratios in relation to various harvest strategies) with or without addition of cattle manure. The main objective is to determine the feasibility of aforementioned mixtures for biogas production and the influence on microbial community structures in response to various feeding compositions. Results demonstrated that cover crop (silage) is feasible for digestion alone or with addition of barley straw, cattle manure or both, while manure addition led to higher CH4 yield/buffer capacity, and enhanced the volatile solids reduction. Microbial community compositions were found to have been affected by the feeding, while high straw addition led to a distinct community structure.

The aim of this review is to give a summary of natural lignocellulose-degrading systems focusing mainly on animal digestive tracts of wood-feeding insects and ruminants in order to find effective strategies that can be applied to improve anaerobic digestion processes in engineered systems. Wood-feeding animals co-evolved with symbiotic microorganisms to digest lignocellulose-rich biomass in a very successful way. Considering the similarities between these animal gut systems and the lignocellulose-based biotechnological processes, the gut with its microbial consortium can be a perfect model for an advanced lignocellulose-degrading biorefinery. The physicochemical properties and structure of the gut may provide a scheme for the process design, and the microbial consortium may be applied as genetic resource for the up-scaled bioreactor communities. Manipulation of the gut microbiota is also discussed in relation to the management of the reactor communities.

Anaerobic digestion (AD) of sugarcane filter cake (SFC) was investigated by comparing the performance of pre-treatment methods in biochemical methane potential (BMP) tests and semi-continuous experiments. For that, SFC was pre-treated by autoclaving the substrate alone or with sodium hydroxide (NaOH). Experimental data from BMP tests were fitted to a kinetics model and further used for simulating the AD process in a continuous stirred-tank reactor (CSTR). BMP tests showed differences (p < 0.05) in total methane potential (SBMP), which have affected methane yields during simulation in a CSTR. Untreated produced 185 mL CH4 gVS−1, autoclaved pre-treatment 174 mL CH4 gVS−1 and autoclaved with NaOH pre-treatment 222 mL CH4 gVS−1. Interestingly, such higher performance of autoclaved with NaOH pre-treatment was only observed at earlier stages during semi-continuous feeding experiment. At steady-state no significant differences (p > 0.05) in terms of methane yield were observed among the reactors (average of 224 mL CH4 gVS−1). These results demonstrate that the benefits of pre-treatment could only be observed in BMP tests, which is likely explained by a better adaptation of the microbial community to the substrate during long term semi-continuous experiment, making SFC pre-treatment ineffective in a single-stage CSTR and under this feeding regime.

Anaerobic digestion of nitrogen-rich substrate often causes process inhibition due to the susceptibility of the microbial community facing ammonia accumulation. However, the precise response of the microbial community has remained largely unknown. To explore the reasons, bacterial communities in ammonia-stressed reactors and control reactors were studied by amplicon pyrosequencing of 16S rRNA genes and the active methanogens were followed by terminal restriction fragment length polymorphism (T-RFLP) analyses of mcrA/mrtA gene transcripts. The results showed that the diversity of bacterial communities decreased in two parallel ammonia-inhibited reactors compared with two control reactors, but different levels of inhibitions coinciding with different community shifts were observed. In one reactor, the process was completely inhibited, which was preceded by a decreasing relative abundance of the phylum Firmicutes. Despite the same operating conditions, the process was stabilized in the parallel, partially inhibited reactor, in which the relative abundance of Firmicutes greatly increased. In particular, both ammonia-inhibited reactors lacked taxa assumed to be syntrophic bacteria (Thermoanaerobacteraceae, Syntrophomonadaceae, and Synergistaceae). Besides the predominance of the hydrogenotrophic methanogens Methanoculleus and Methanobacterium, activity of Methanosarcina and even of the strictly aceticlastic genus Methanosaeta were found to contribute at very high ammonia levels (> 9 g NH4-N L−1) in the stabilized reactor (partial inhibition). In contrast, the lack of aceticlastic activity in the parallel reactor might have led to acetate accumulation and thus process failure (complete inhibition). Collectively, ammonia was found to be a general inhibitor while accumulating acetate and thus acidification might be the key factor of complete process failure.

Anaerobic digestion is a biochemical process where complex organic matter such as carbohydrates, proteins, and lipids degrade in the absence of oxygen and are converted into methane and carbon dioxide by the action of different groups of microorganisms. It is a sustainable waste management technology, which reduces and stabilizes organic wastes, recycles its nutrient and water content, while producing energy. Biogas reduces the demand for fossil fuels, since it can be used for the production of electric power and heat, or converted into vehicle fuel. Currently, methane production via anaerobic digestion is a steadily increasing industry in Europe and all over the world. This chapter focuses on the anaerobic digestion process and the parameters affecting its performance. It also describes briefly the current technologies for anaerobic digestion. Finally, since the degradation of organic material requires a synchronized action of different groups of microorganisms with different metabolic capabilities, this chapter also presents recent developments in molecular biology techniques as valuable tools to obtain in-depth understanding about this complex microbiological system.

The concept of year-round biogas production to increase the capacity factor of anaerobic digestion (AD) plants in sugarcane biorefineries was investigated for the first time in semi-continuous feeding mode. To simulate the use of sugarcane vinasse during the sugarcane season and sugarcane filter cake (SFC) during the off-season period, a two-stage reactor system based on an acidogenic continuous stirred-tank reactor (1st stage) followed by solid–liquid separation and an upflow anaerobic sludge blanket (UASB) reactor (2nd stage) to convert the COD-rich liquid fraction into biogas was operated. Additionally, to optimize the biogas production from SFC, the effects of its thermo-chemical pre-treatment on AD were investigated in a parallel reactor set-up. The saponification effect provided by autoclaving the substrate with sodium hydroxide improved the hydrolysis/fermentation of SFC in the acidogenic reactor, which in turn resulted in a 28% higher volumetric methane production in the methanogenic reactor (p < 0.05). However, the methane yields observed during operation of the two-stage reactor system were markedly lower than previously found in biochemical methane potential tests using SFC. In this case, the feed-in with low suspended solids required by UASB reactors prevented the utilization of the non-hydrolyzed/fermented solid fraction of SFC (> 60% of the substrate’s methane potential). Nevertheless, the capacity factor of the AD plants in sugarcane biorefineries could be increased from 0.55 up to 0.69 when considering a 200 d a−1 sugarcane season (0.66–0.83 for a longer season of 240 d a−1), representing an increase of 25.7%. The average capacity factor for biogas combined heat and power and upgrading units of around 0.91 (8000 h a−1) would be reached if further developments could improve the solubilization of non-hydrolyzed/fermented solids or alternatively allow their direct use in the methanogenic reactor.

Three different bioaugmentation cultures enriched from natural and engineered cellulolytic environments (cow and goat rumen, a biogas reactor digesting sorghum biomass) were compared for their enhancement potential on the anaerobic digestion of wheat straw. Methane yields were determined in batch tests using the Automatic Methane Potential Test System operated for 30 days under mesophilic conditions. All cultures had positive effects on substrate degradation, and higher methane yields were observed in the bioaugmented reactors compared to control reactors set up with standard inoculum. However, the level of enhancement differed according to the type of the enrichment culture. Methane yield in batch reactors augmented with 2% cow rumen derived enrichment culture was increased by only 6%. In contrast, reactors amended with 2% goat rumen derived enrichment culture or with the bioaugmentation culture obtained from the biogas reactor digesting sorghum biomass produced 27 and 20% more methane, respectively. The highest methane yield was recorded in reactors amended with 6% goat rumen derived enrichment culture, which represented an increase by 36%. The microbial communities were quite similar at the end of the batch tests independently of the bioaugmentation sources, indicating that the introduced microbial communities of the enrichment cultures did not dominate the reactors.

Anaerobic digestion of poultry manure poses many challenges due to ammonia inhibition by nitrogen‐containing compounds. Here, the effect of NH4Cl or urea‐induced inhibition was investigated in bioreactors fed with maize silage or cellulose and compared to previous experiments with chicken manure as ammonia source. The hydrogen and carbon stable isotope compositions of biogas samples were evaluated to assess the shifts in predominant methanogenic pathways during increasing ammonia inhibition. Due to the higher free ammonia concentrations, stronger inhibition was observed in the urea added reactors compared to the NH4Cl treated ones. Collectively, the results suggested that ammonium salts are not really appropriate to investigate ammonia inhibition during anaerobic digestion of manure.

Anaerobic digestion of sugarcane straw co-digested with sugarcane filter cake was investigated with a special focus on macronutrients supplementation for an optimized conversion process. Experimental data from batch tests and a semi-continuous experiment operated in different supplementation phases were used for modeling the conversion kinetics based on continuous stirred-tank reactors. The semi-continuous experiment showed an overall decrease in the performance along the inoculum washout from the reactors. By supplementing nitrogen alone or in combination to phosphorus and sulfur the specific methane production significantly increased (P < 0.05) by 17% and 44%, respectively. Although the two-pool one-step model has fitted well to the batch experimental data (R2 > 0.99), the use of the depicted kinetics did not provide a good estimation for process simulation of the semi-continuous process (in any supplementation phase), possibly due to the different feeding modes and inoculum source, activity and adaptation.

The aim of this study was to develop an effective bioaugmentation concept for anaerobic digesters treating lignocellulosic biomass such as straw. For that purpose, lignocellulose-degrading methanogenic communities were enriched on wheat straw from cow and goat rumen fluid as well as from a biogas reactor acclimated to lignocellulosic biomass (sorghum as mono-substrate). The bacterial communities of the enriched cultures and the different inocula were examined by 454 amplicon sequencing of 16S rRNA genes while the methanogenic archaeal communities were analyzed by terminal restriction fragment length polymorphism (T-RFLP) fingerprinting of the mcrA gene. Bacteroidetes was the most abundant phylum in all samples. Within the Bacteroidetes phylum, Bacteroidaceae was the most abundant family in the rumen-derived enrichment cultures, whereas Porphyromonadaceae was the predominant one in the reactor-derived culture. Additionally, the enrichment procedure increased the relative abundance of Ruminococcaceae (phylum: Firmicutes) in all cultures. T-RFLP profiles of the mcrA gene amplicons highlighted that the ruminal methanogenic communities were composed of hydrogenotrophic methanogens dominated by the order Methanobacteriales regardless of the host species. The methanogenic communities changed significantly during the enrichment procedure, but still the strict hydrogenotrophic Methanobacteriales and Methanomicrobiales were the predominant orders in the enrichment cultures. The bioaugmentation potential of the enriched methanogenic cultures will be evaluated in further studies.

The aim of this study was to determine the potential of bioaugmentation with cellulolytic rumen microbiota to enhance the anaerobic digestion of lignocellulosic feedstock. An anaerobic cellulolytic culture was enriched from sheep rumen fluid using wheat straw as substrate under mesophilic conditions. To investigate the effects of bioaugmentation on methane production from straw, the enrichment culture was added to batch reactors in proportions of 2% (Set-1) and 4% (Set-2) of the microbial cell number of the standard inoculum slurry. The methane production in the bioaugmented reactors was higher than in the control reactors. After 30 days of batch incubation, the average methane yield was 154 mLN CH4 gVS−1 in the control reactors. Addition of 2% enrichment culture did not enhance methane production, whereas in Set-2 the methane yield was increased by 27%. The bacterial communities were examined by 454 amplicon sequencing of 16S rRNA genes, while terminal restriction fragment length polymorphism (T-RFLP) fingerprinting of mcrA genes was applied to analyze the methanogenic communities. The results highlighted that relative abundances of Ruminococcaceae and Lachnospiraceae increased during the enrichment. However, Cloacamonaceae, which were abundant in the standard inoculum, dominated the bacterial communities of all batch reactors. T-RFLP profiles revealed that Methanobacteriales were predominant in the rumen fluid, whereas the enrichment culture was dominated by Methanosarcinales. In the batch rectors, the most abundant methanogens were affiliated to Methanobacteriales and Methanomicrobiales. Our results suggest that bioaugmentation with sheep rumen enrichment cultures can enhance the performance of digesters treating lignocellulosic feedstock.

Comparative analysis of methanogenic archaea compositions and dynamics in 11 laboratory-scale continuous stirred tank reactors fed with different agricultural materials (chicken manure, cattle manure, maize straw, maize silage, distillers grains, and Jatropha press cake) was carried out by analysis of the methyl coenzyme-M reductase -subunit (mcrA) gene. Various taxa within Methanomicrobiales, Methanobacteriaceae, Methanosarcinaceae, Methanosaetaceae, and Methanomassiliicoccales were detected in the biogas reactors but in different proportions depending on the substrate type utilized as well as various process parameters. Improved coverage and higher taxonomic resolution of methanogens were obtained compared to a previous 16S rRNA gene based study of the same reactors. Some members of the genus Methanoculleus positively correlated with the relative methane content, whereas opposite correlations were found for Methanobacterium. Specific biogas production was found to be significantly correlating with Methanosarcinaceae. Statistical analysis also disclosed that some members of the genus Methanoculleus positively correlated with the ammonia level, whereas the prevalence of Methanocorpusculum, Methanobacterium, and Methanosaeta was negatively correlated with this parameter. These results suggest that the application of methanogenic archaea adapted to specific feedstock might enhance the anaerobic digestion of such waste materials in full-scale biogas reactors.

BackgroundDuring strategic planning of a biogas plant, the local availability of resources for start-up and operation should be taken into consideration for a cost-efficient process. Because most bioethanol/sugar industries in Brazil are located in remote areas, the use of fresh cattle manure from local farms could be a solution for the inoculation of the biogas process. This study investigated the diversity and dynamics of bacterial and archaeal communities and the performance of biogas reactors inoculated with manure and a mixed inoculum from different biogas reactors as for a controlled start-up until steady state.ResultsLaboratory-scale biogas reactors were fed semi-continuously with sugarcane filter cake alone (mono-digestion) or together with bagasse (co-digestion). At the initial start-up, the reactors inoculated with the mixed inoculum displayed a less diverse taxonomic composition, but with higher presence of significant abundances compared to reactors inoculated with manure. However, in the final steady state, the communities of the differently inoculated reactors were very similarly characterized by predominance of the methanogenic genera Methanosarcina and Methanobacterium, the bacterial families Bacteroidaceae, Prevotellaceae and Porphyromonadaceae (phylum Bacteroidetes) and Synergistaceae (phylum Synergistetes). In the mono-digestion reactors, the methanogenic communities varied greater than in the co-digestion reactors independently of the inoculation strategy.ConclusionThe microbial communities involved in the biogas production from waste products of the Brazilian bioethanol/sugar industry were relatively similar and stable at the reactor’s steady phase independently of the inoculum source (manure or mixed inoculum). Therefore, the locally available manure can be used as inoculum for start-up of the biogas process, since it also contains the microbial resources needed. The strong fluctuation of methanogenic communities in mono-digestion reactors indi

The effects of hydraulic retention time (HRT) reduction at constant high organic loading rate on the activity of hydrogen-producing bacteria and methanogens were investigated in reactors digesting thin stillage. Stable isotope fingerprinting was additionally applied to assess methanogenic pathways. Based on hydA gene transcripts, Clostridiales was the most active hydrogen-producing order in continuous stirred tank reactor (CSTR), fixed-bed reactor (FBR) and anaerobic sequencing batch reactor (ASBR), but shorter HRT stimulated the activity of Spirochaetales. Further decreasing HRT diminished Spirochaetales activity in systems with biomass retention. Based on mcrA gene transcripts, Methanoculleus and Methanosarcina were the predominantly active in CSTR and ASBR, whereas Methanosaeta and Methanospirillum activity was more significant in stably performing FBR. Isotope values indicated the predominance of aceticlastic pathway in FBR. Interestingly, an increased activity of Methanosaeta was observed during shortening HRT in CSTR and ASBR despite high organic acids concentrations, what was supported by stable isotope data.

In this study, the effects of nitrogen, phosphate and trace elements supplementation were investigated in a semi-continuously operated upflow anaerobic sludge blanket system to enhance process stability and biogas production from sugarcane vinasse. Phosphate in form of KH2PO4 induced volatile fatty acids accumulation possibly due to potassium inhibition of the methanogenesis. Although nitrogen in form of urea increased the reactor’s alkalinity, the process was overloaded with an organic loading rate of 6.1 g COD L−1 d−1 and a hydraulic retention time of 3.6 days. However, by supplementing urea and trace elements a stable operation even at an organic loading rate of 9.6 g COD L−1 d−1 and a hydraulic retention time of 2.5 days was possible, resulting in 79% higher methane production rate with a stable specific methane production of 239 mL g COD−1.

The effect of pig manure mixed with rice straw on methane yield and the microbial community involved in a thermophilic (55°C) anaerobic digestion process was investigated. Three substrates composed of mixed pig manure and rice straw at different ratios (95:5; 78:22 and 65:35 w/w, which resulted in C/N ratios of 10:1, 20:1 and 30:1) were used for the experiment. The substrate type had a major influence on the total bacterial community, while the methanogens were less affected. The members of the class Clostridia (phylum Firmicutes) were predominant regardless of mixture ratio (C/N ratio), but at species level there was a major difference between the low and high C/N ratio samples. The hydrogenotrophic methanogenic genus of Methanothermobacter was predominant in all samples but higher C/N ratio sequences affiliated to the genus Methanosarcina were also detected. The appearance of Methanosarcina sp. is most likely due to the less inhibition of ammonia during the anaerobic digestion.

The anaerobic digestion of sugarcane filter cake and the option of co-digestion with bagasse were investigated in a semi-continuous feeding regime to assess the main parameters used for large-scale process designing. Moreover, fresh cattle manure was considered as alternative inoculum for the start-up of biogas reactors in cases where digestate from a biogas plant would not be available in remote rural areas. Experiments were carried out in 6 lab-scale semi-continuous stirred-tank reactors at mesophilic conditions (38 ± 1 °C) while the main anaerobic digestion process parameters monitored. Fresh cattle manure demonstrated to be appropriate for the start-up process. However, an acclimation period was required due to the high initial volatile fatty acids concentration (8.5 g L−1). Regardless the mono-digestion of filter cake presented 50% higher biogas yield (480 mL gVS−1) than co-digestion with bagasse (320 mL gVS−1) during steady state conditions. A large-scale co-digestion system would produce 58% more biogas (1008 m3 h−1) than mono-digestion of filter cake (634 m3 h−1) due to its higher biomass availability for biogas conversion. Considering that the biogas production rate was the technical parameter that displayed the most relevant differences between the analyzed substrate options (0.99–1.45 m3 biogas m3 d−1). The decision of which substrate option should be implemented in practice would be mainly driven by the available construction techniques, since economically efficient tanks could compensate the lower biogas production rate of co-digestion option.

Biogas production from sugarcane waste has large potential for energy generation, however, to enable the optimization of the anaerobic digestion (AD) process each substrate characteristic should be carefully evaluated. In this study, the kinetic challenges for biogas production from different types of sugarcane waste were assessed. Samples of vinasse, filter cake, bagasse, and straw were analyzed in terms of total and volatile solids, chemical oxygen demand, macronutrients, trace elements, and nutritional value. Biochemical methane potential assays were performed to evaluate the energy potential of the substrates according to different types of sugarcane plants. Methane yields varied considerably (5–181 Nm3·tonFM−1), mainly due to the different substrate characteristics and sugar and/or ethanol production processes. Therefore, for the optimization of AD on a large-scale, continuous stirred-tank reactor with long hydraulic retention times (>35 days) should be used for biogas production from bagasse and straw, coupled with pre-treatment process to enhance the degradation of the fibrous carbohydrates. Biomass immobilization systems are recommended in case vinasse is used as substrate, due to its low solid content, while filter cake could complement the biogas production from vinasse during the sugarcane offseason, providing a higher utilization of the biogas system during the entire year.

Flexible biogas production that adapts biogas output to energy demand can be regulated by changing feeding regimes. In this study, the effect of change in feeding intervals on process performance, microbial community structure and methanogenesis pathway was investigated. Three different feeding regimes (once daily, every second day and every 2 hours) at the same organic loading rate were studied in continuous stirred tank reactors treating distillers dried grains with solubles. Higher amount of biogas was produced after feeding in the less frequently fed reactors (once per day and every second day) whereas it remained constant in the more frequently fed reactor (every 2 hours), indicating the suitability of the former for production of biogas flexibly. Compared to the conventional more frequent feeding regimes, higher methane yield of up to 14% and improved process stability against organic overloading were achieved by employing less frequent feeding regimes. Bacteria and methanogenic archaea community structures were monitored by terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA and mcrA genes, respectively. The results showed that the composition of the bacterial community varied under the different feeding regimes and the observed T-RFLP patterns were best explained by the difference in total ammonia nitrogen concentration, H2 level and pH value. However, the methanogenic community remained stable under all feeding regimes, with the dominance of Methanosarcina followed by Methanobacterium genus. Stable isotope analysis showed that the average methane produced during each feeding event by acetoclastic and hydrogenotrophic methanogenesis was not influenced under the three feeding regimes.

Different methods for optimization the anaerobic digestion (AD) of sugarcane filter cake (FC) with a special focus on volatile fatty acids (VFA) production were studied. Sodium hydroxide (NaOH) pretreatment at different concentrations was investigated in batch experiments and the cumulative methane yields fitted to a dual-pool two-step model to provide an initial assessment on AD. The effects of nitrogen supplementation in form of urea and NaOH pretreatment for improved VFA production were evaluated in a semi-continuously operated reactor as well. The results indicated that higher NaOH concentrations during pretreatment accelerated the AD process and increased methane production in batch experiments. Nitrogen supplementation resulted in a VFA loss due to methane formation by buffering the pH value at nearly neutral conditions (∼6.7). However, the alkaline pretreatment with 6 g NaOH/100 g FCFM improved both the COD solubilization and the VFA yield by 37%, mainly consisted by n-butyric and acetic acids.

The anaerobic digestion of filter cake and its co-digestion with bagasse, and the effect of gradual increase of the organic loading rate (OLR) from start-up to overload were investigated. Understanding the influence of environmental and technical parameters on the development of particular methanogenic pathway in the biogas process was an important aim for the prediction and prevention of process failure. The rapid accumulation of volatile organic acids at high OLR of 3.0 to 4.0 gvs·L−1·day−1 indicated strong process inhibition. Methanogenic community dynamics of the reactors was monitored by stable isotope composition of biogas and molecular biological analysis. A potential shift toward the aceticlastic methanogenesis was observed along with the OLR increase under stable reactor operating conditions. Reactor overloading and process failure were indicated by the tendency to return to a predominance of hydrogenotrophic methanogenesis with rising abundances of the orders Methanobacteriales and Methanomicrobiales and drop of the genus Methanosarcina abundance.

Alkaline pretreatment has the potential to enhance the anaerobic digestion of lignocellulosic biomass to biogas. However, the elevated pH of the substrate may require alkalitolerant microbial communities for an effective digestion. Three mixed anaerobic lignocellulolytic cultures were enriched from sediments from two soda lakes with wheat straw as substrate under alkaline (pH 9) mesophilic (37°C) and thermophilic (55°C) conditions. The gas production of the three cultures ceased after 4 to 5 weeks, and the produced gas was composed of carbon dioxide and methane. The main liquid intermediates were acetate and propionate. The physiological behavior of the cultures was stable even after several transfers. The enrichment process was also followed by molecular fingerprinting (terminal restriction fragment length polymorphism) of the bacterial 16S rRNA gene and of the mcrA/mrtA functional gene for methanogens. The main shift in the microbial community composition occurred between the sediment samples and the first enrichment, whereas the structure was stable in the following transfers. The bacterial communities mainly consisted of Sphingobacteriales, Clostridiales and Spirochaeta, but differed at genus level. Methanothermobacter and Methanosarcina genera and the order Methanomicrobiales were predominant methanogenes in the obtained cultures. Additionally, single cellulolytic microorganisms were isolated from enrichment cultures and identified as members of the alkaliphilic or alkalitolerant genera. The results show that anaerobic alkaline habitats harbor diverse microbial communities, which can degrade lignocellulose effectively and are therefore a potential resource for improving anaerobic digestion.