Dry Anaerobic Digestion Research Articles

Cobalt-modified digestate-derived biochar enhances kitchen waste anaerobic dry digestion: Performance, microbial mechanisms, and metabolic pathways

To achieve simultaneous in-situ high-value utilization of digestate and efficient resource recovery of kitchen waste (KW). In this work, Co-modified digestate-derived biochar (DDB-Co) was prepared for the first time, the effects of DDB-Co on KW mesophilic dry anaerobic digestion were investigated, and the underlying mechanisms were elucidated. The results showed that the specific surface area of DDB-Co increased to 98.7 m2/g, which was 37.4 % higher than that of the unmodified group (DDB). DDB-Co exhibited a dose-dependent effect on biogas production from the KW dry digestion, with the optimal dose of 4.0 g/L (calculated on a solid content) DDB-Co yielding 205.3 mL/(g·VS) of biogas, representing an 80.9 % increase compared to the CK group. However, higher doses, such as 8.0 g/L DDB-Co, reduced the cumulative biogas production. Mechanistic analysis indicated that 4.0 g/L DDB-Co facilitated the dissolution of particulate organic matter and accelerates the conversion process of volatile fatty acids (VFA) in KW, thereby preventing excessive acidification. Microbial community structure analysis revealed that DDB-Co reduced microbial richness and diversity but selectively enriched microorganisms such as Lactobacillus and Methanothrix, accelerating the direct interspecific electron transfer (DIET) reaction rate and increasing biogas production. Metagenomic analysis demonstrated that DDB-Co primarily upregulated the relative abundances of key enzymes in the acetate decarboxylation methanogenesis pathway, such as acetate kinase (ackA), phosphate acetyltransferase (pta), and acetyl-CoA synthetase (acs), enhancing biogas yield and maintaining system stability. The findings of this work expand the high-value utilization pathways for digestate and provide an alternative solution for the efficient resource recovery of KW.

Evaluation of the efficiency of dry anaerobic digester in the production of biogas and fertilizer using activated sludge and plant waste.

The process of dry anaerobic digestion (DAD) is an effective economic and environmentally friendly method due to its features such as reducing the volume of waste, destroying pathogens, consuming less energy, and producing biogas rich in energy and fertilizer. This study was conducted on a pilot scale with a reactor volume of 24L in 30 days for three ratios of C/N: 30, 25, and 20 separately using a mixture of activated sludge and plant waste. During the process, the temperature, pH, and ratio of C/N were measured at certain times, and the percentage of gases produced by the Gas chromatography (GC) device was also measured. The results were compared with the National standard of Iran (Compost, Physical, and Chemical Specifications) at the end of the process. The results showed that the average volume of biogas produced in ratios of C/N: 30, 25, 20 is 1.7, 2.5, and 1.5L. d-1, respectively. The results showed that in the C/N ratio of 20, and 25, the amount of, C/N ratio and NO3 are in the range the standard of class 1, and the amount of Phosphorus pentoxide (P2O5) is in the range the standard of class 2. The results showed that the average microbial parameters in ratios of C/N: 30, 25, and 20 are lower than the national standard of Iran. The dry digester is an efficient method in waste treatment, methane gas production, and quality fertilizer close to the national standard of Iran.

Optimization of Dry Anaerobic Digestion of Food Waste in Leachate Bed Reactors

Optimization of Dry Anaerobic Digestion of Food Waste in Leachate Bed Reactors

Fixing collapsed dry anaerobic digestion system of kitchen waste caused by severe VFAs accumulation

Severe inhibition by volatile fatty acids (VFAs) frequently occurs during the dry anaerobic digestion of kitchen waste at high organic load rate (OLR), leading to decreased biogas production and even the collapse of the anaerobic digestion system. In this study, VFAs accumulation was significantly reduced by adding powder-activated carbon (PAC) or alkaline additives (AA), thus restoring the collapsed system to a stable operating state. With the addition of PAC or AA, the maximal OLR reached 11.14 g VS/(L·d), which was 30 % higher than observed without any additives (control group), while the VFAs concentration was maintained below 3000 mg/L. At this OLR, the volumetric biogas production rate stabilized at 5.1 L/(L·d) in the presence of PAC and AA, while that of the control group gradually decreased to zero. The VFAs concentration with PAC addition was 86 % lower than that of the control group, possibly because PAC might stimulate the formation of direct interspecies electron transfer between syntrophic bacteria and methanogens (Methanosarcina), thereby promoting VFAs degradation. The addition of AA, which resulted in a 95 % decrease in ammonia-nitrogen concentration, can provide a good growth environment for the microorganisms involved in acidogenesis and hydrogenotrophic methanogenesis.

A multistage biogas slurry reflux and spray anaerobic digestion reactor for high solid anaerobic digestion: Performance and application evaluation

In this study, a new vertical plug-flow continuous dry anaerobic digestion reactor equipped with a spray percolation system was designed for rice straw and cow manure in dry AD. The findings indicated that the methane yield was 290.63 ± 19.16 mL/g VS when the optimal spray time (9 times/d) was used in batch AD. In addition, the results of continuous experiments under optimal spray time conditions showed that the volumetric methane production rate of the system was 1.55 L/(L·d) when the organic loading rate was 20 g VS/(L·d). In addition, hydrolysis and acidification steps were enhanced. This study demonstrated the potential of the new reactor for stable operation of high OLR dry AD. Digestate is already used as a substitute for synthetic fertilizers precisely because of the environmental benefits associated with its agronomic use.

Comparing the effects and mechanisms of granular activated carbon and magnetite nanoparticles in dry anaerobic digestion: Focusing on microbial electron transfer and metagenomic evidence

Enhancing direct interspecies electron transfer (DIET) via granular activated carbon (GAC) and magnetite nanoparticles (MNPs) to promote methanation performance in dry anaerobic digestion (AD) systems has been extensively studied in recent years. However, the underlying mechanisms, particularly how GAC and MNPs influence DIET and extracellular polymeric substances (EPSs), remain poorly understood. In this study, the different effects and mechanisms of GAC and MNPs on dry AD fed with pig manure (PM) and corn straw were compared, with additions of 10, 20, and 50% of the volatile solid (VS) of PM. The results indicated that GAC improved the cumulative specific methane yields (CSMYs) by 18.8–20.9%, while a high dose of MNPs (50% of the VSPM) decreased the CSMY and hydrolysis rate constant by 27.0 and 75.4%, respectively. GAC provided microorganism attachment sites and functional groups for electron transfer, improving hydrolysis by enriching fermenters such as Fastidiosipila and Rikenellaceae. Nevertheless, MNPs stimulated intense dissimilatory iron reduction by Clostridium sp. in the early stage, competing with methanogens for substrates. Low and medium GAC doses (10 and 20% of the VSPM) increased the utilization of redox-active substances (e.g., humic substances and proteins) in EPS, while the high MNP dose stimulated their secretion due to microbial self-protection, impairing electron transfer. Metagenomic evidence showed that the medium GAC dose improved four methanogenic pathways and DIET mediated by microbial nanowires, while a high MNP dose impeded the final step of methanogenesis. These results provide deeper insights into the mechanisms of different conductive materials on dry AD.

Insight into response mechanism of short-chain fatty acids to refined microbial transformation order of dissolved organic matter ranked by molecular weight during dry anaerobic digestion

Dynamic transformation of dissolved organic matter (DOM) contributes to short-chain fatty acids (SCFAs) production during anaerobic digestion. However, the impact of refined transformation of DOM ranked by molecular weight (MW) on SCFAs has never been investigated. Results indicated that DOM conversion order was 3500–7000 Da>(MW>14000 Da) > 7000–4000 Da during hydrolysis stage, while it was independent of their MW in acidogenesis phase and followed a low to high MW order during methanogenesis stage. Proteins-like DOMs with different MW were closely related to SCFAs. Eight groups of microorganisms (e.g., Bacillus and Caldicoprobacter) responsible for the conversion of proteins-like DOMs to SCFAs. The possible routes linking environmental properties to microorganisms-proteins-like DOMs-SCFAs connections were constructed. Microbial activity modifications by regulating moisture, pH, NO3−-N and NH4+-N can expedite the conversion of proteins-like DOMs to SCFAs. The study emphasizes the importance of MW-classification-based biotransformation of organic waste, offering a potential strategy to enhance anaerobic digestion performance.

Distribution of microplastics and phthalic acid esters during dry anaerobic digestion of food waste and potential microbial degradation analysis

Food waste (FW) and its biogas residue were considered as sources of terrestrial microplastics (MPs) and phthalic acid esters (PAEs) contamination. However, there was a lack of research and understanding of the MPs and PAEs pollution problem in FW dry anaerobic digestion process (DADP). The MPs and PAEs in three stages of the DADP with the largest monomer disposal scale in China were identified. At the biogas residue extrusion stage, MPs abundance and PAEs concentration reached the highest values, which were 3.63 ± 0.45 × 103 N·kg-1 and 3.62 ± 0.72 mg·kg-1, respectively. Furthermore, there was a significant positive correlation between MPs and PAEs throughout the process (p < 0.05). Although bacteria and fungi with plastic degradation potential were present in all stages, the contamination problem of MPs and PAEs cannot be completely solved through DADP. This study provides a scientific basis for preventing and controlling the pollution of MPs and PAEs.

Evolution of quorum sensing process and their regulatory role on biochemical metabolism during the organic loading rate increase in dry anaerobic digestion

The organic loading rate (OLR) is a critical parameter affecting the stability of dry anaerobic digestion (AD) of kitchen waste (KW), and significantly impacting the variations in physicochemical parameters and microbial communities. However, the evolution of quorum sensing (QS) and their role on anaerobic biochemical metabolism during the increase in OLR in dry AD remain unknown. Therefore, this study systematically elucidated the matter through multi-omics analysis based on a pilot-scale dry AD of KW. The results demonstrated that fluctuations in the OLR significantly influenced the microbial QS in dry AD. When the OLR ≤4.0 g·VS/L·d, the system operated stably, and methane production increased. The enrichment of Proteobacteria was crucial for sustaining high levels of functional genes associated with various types of QS, including acyl-homoserine lactones (AI-1), autoinducer-2 (AI-2), autoinducer-3 (AI-3), and gamma-aminobutyric acid (GABA). This enabled cooperative communication among microbes under low OLR. Furthermore, most genes associated with these QS processes positively affected hydrolysis, acidogenesis, and methanogenesis. When the OLR increased to 6.0 g·VS/L·d, the fatty acids and hydrogen partial pressure increased significantly. The autoinducing peptides (AIP)-type became the predominant QS and was positively correlated with fatty acids abundance. Syntrophaceticus and Syntrophomonas may promote syntrophic oxidation of acetate at high OLR through AIP-type QS. These findings provided new insights into the QS processes of microbes during dry AD of KW and a theoretical foundation for optimizing biochemical metabolic processes in dry AD through QS.

Comparative evaluation of high solids anaerobic digestion of rice husk and rice straw: Impact of substrate characteristics and solids concentration on biogas yield and digestate quality

Open burning of crop residues in agricultural fields causes soil infertility and contributes to global warming. Solid-state or dry anaerobic digestion (AD) is an emerging process for converting high solids (90 ̶ 95%) biomass such as rice husk (RH) and rice straw (RS) into energy and resources without the need for large volmes of water. This paper describes laboratory experiments investigating the dry AD of RH and RS across a range of total solids (TS) concentrations from 20 to 60%. Conducted in 500 mL bioreactors under batch mode at mesophilic temperatures, the experiments had a batch retention time of 45 days. An increase in the F/I ratio from 8.37 to 25.01 correlated with a rise in biogas generation from 2.9 to 4.6 L in RH reactors. The highest biogas yield from RS (0.49 L/g VS) and RH (0.17 L/g VS) was obtained at 20 and 50% TS, respectively. From the mass balance, 68.7 g of biomanure and 4.6 g of biogas were produced from the 50H reactor, while the 20S reactor yielded 20.9 g of biomanure and 8.4 g of biogas. From the modified Gompertz model a shorter lag phase of 1.9 days was observed in reactor 50H compared to the 4.6 days observed in reactor 20S. HIGHLIGHTS Dry AD of rice straw and husk at variable total solids (20–60% TS) Maximum biogas yield of 0.49 L/g VSreduced was observed at 20% TS from rice straw Maximum biogas yield of 0.17 L/g VSreduced was observed at 50% TS from rice husk Rice husk yielded concentrated digestate than rice straw with good nutrient quality Shorter lag phase was observed for 20S and 50H reactors

Unveiling the synergy of volatile fatty acids and ammonia nitrogen in optimizing methane production during dry anaerobic digestion of pig manure and corn straw

Dry anaerobic digestion (AD) is vulnerable to volatile fatty acids (VFA) and ammonia nitrogen. Understanding their synergy is crucial for optimizing strategies. In this study, the digestive performance, microbial dynamics, metabolic pathways, and synergistic relationships were investigated in the dry AD process by adding different concentrations of acetic acid (2, 5, and 8 mg/g) and urea (1, 5 mg/g N). Results showed that acid inhibition was caused by excessive pH reduction due to VFA accumulation. Propionic acid was the primary form of VFA accumulation. The inhibition of VFA accumulation could be alleviated using a low concentration of ammonia nitrogen (1 mg/g N urea) or be strengthened using a high concentration of ammonia nitrogen (5 mg/g N urea). Compared with the control group (82.2 mL/g VS), VFA and 1 mg/g N urea had a synergistic facilitation effect on the methane production (increasing by 1.7 %-18.1 %). This synergistic effect shortened the lag phases, enhanced the activity of Caldicoprobacter and Methanosarcina, increased the expression of genes encoding functional enzymes involved in the acetoclastic-methanogenic pathway. By contrast, VFA and 5 mg/g N urea had a synergistic inhibition effect on methane production (decreasing by 65.7 %–69.8 %). This synergistic effect prolonged lag phases, increased the relative abundance of ammonia-tolerant and acid-tolerant microbial (such as Methanoculleus) and reduced the expression of genes encoding functional enzymes involved in the hydrogenotrophic-methanogenic pathway. These results improve the understanding of methane regulation during dry AD.

Food waste impact on dry anaerobic digestion of straw in a novel reactor: Biogas yield, stability, and hydrolysis-methanogenesis processes

Gradient anaerobic digestion reactor (GADR) can improve substrate utilization efficiency by solving the problem of the “short circuit” of materials. However, the substrate’s composition significantly affects the reactor’s performance. This study investigated the impact of food waste (FW) levels on corn straw’s dry anaerobic digestion (AD) in a novel GADR. The results show that biomethane production can be improved by coupling urban and agricultural solid waste recycling. The mechanism is to increase the hydrolysis and acid production efficiency, and the abundance of enzymes related to methanogenesis. The maximum methane yield (494.2 mL CH4/g VS) and the highest anaerobic biodegradability (85.7 %) were obtained when the FW was added at 60 %. The co-digestion of FW and straw can improve the hydrolysis and acid production efficiency and methane yield, which improves the buffering capacity and stability of the system compared with the single digestion of FW.

Enhancement of Biodegradability of Chicken Manure via the Addition of Zeolite in a Two-Stage Dry Anaerobic Digestion Configuration.

Leach bed reactors (LBRs) are dry anaerobic systems that can handle feedstocks with high solid content, like chicken manure, with minimal water addition. In this study, the chicken manure was mixed with zeolite, a novel addition, and packed in the LBR to improve biogas production. The resulting leachate was then processed in a continuous stirred tank reactor (CSTR), where most of the methane was produced. The supernatant of the CSTR was returned to the LBR. The batch mode operation of the LBR led to a varying methane production rate (MPR) with a peak in the beginning of each batch cycle when the leachate was rich in organic matter. Comparing the MPR in both systems, the peaks in the zeolite system were higher and more acute than in the control system, which was under stress, as indicated by the acetate accumulation at 2328 mg L-1. Moreover, the presence of zeolite in the LBR played a crucial role, increasing the overall methane yield from 0.142 (control experiment) to 0.171 NL CH4 per g of volatile solids of chicken manure entering the system at a solid retention time of 14 d. Zeolite also improved the stability of the system. The ammonia concentration increased gradually due to the little water entering the system and reached 3220 mg L-1 (control system) and 2730 mg L-1 (zeolite system) at the end of the experiment. It seems that zeolite favored the accumulation of the ammonia at a lower rate (14.0 mg L-1 d-1) compared to the control experiment (17.3 mg L-1 d-1). The microbial analysis of the CSTR fed on the leachate from the LBR amended with zeolite showed a higher relative abundance of Methanosaeta (83.6%) compared to the control experiment (69.1%). Both CSTRs established significantly different bacterial profiles from the inoculum after 120 days of operation (p < 0.05). Regarding the archaeal communities, there were no significant statistical differences between the CSTRs and the inoculum (p > 0.05).

Bulking agent in dry anaerobic digestion as a key factor for the enhancement of biogas production

Dry anaerobic digestion (dry-AD) is an attractive process for solid wastes such as agri-food waste. However, some limitations mainly associated to lack of effective mixing, can hinder the methane production capacity of the systems. Bulking agent (BA) has been proposed as a solution to the compaction issues in systems without mechanical agitation, such as leaching bed reactors. However, effects of BA are still not clear, and, thus, the factors to consider for its dose has not been optimized yet. This work studies the effect of BA in dry-AD. Two substrates with different characteristics were proposed as models, bean peel as a lignocellulosic substrate and a mixture of food waste as a readily biodegradable substrate. Inert plastic rings were used as BA at different BA:S ratios. Assessed BA:S ratio did not affect the performance of methane production for the lignocellulosic waste, but it did significantly affect to the easily biodegradable substrate, showing up to a 28% of methane production increase. This result could be due to the presence of lignocellulosic compounds in the bean peel, behaving like a natural BA. In assays with an increased bed height, the compaction of the system was more severe, resulting in the rapid acidification of the processes. At these conditions, the positive effect of BA addition was more marked, allowing methane production and no acidification of the system. Thus, the addition of BA is a suitable strategy for improving methane production or stability in dry-AD systems without requiring the stirring of the systems.

From grass to gas and beyond: Anaerobic digestion as a key enabling technology for a residual grass biorefinery

Roadside grass clippings hold potential as a sustainable source of bioenergy as they do not compete with crops for land use, and are only partially utilized for low-value applications. In this study, we proposed using roadside grass as a sole feedstock for anaerobic digestion (AD) in three different settings, and assessed the potential of producing biomaterials and fertilizers from grass-based digestate. Wet continuous digestion at pilot scale and dry batch digestion at pilot and large scales resulted in biogas yields up to 700 Nm3.t−1 DOM with a methane content of 49–55 %. Despite promising results, wet AD had operational problems such as clogging and poor mixing; once upscaled, the dry digestion initially also presented an operational problem with acidification, which was overcome by the second trial. Digested grass fibers from the pilot dry AD were processed into biomaterials and performed similarly or better than the undigested fibers, while around 20 % performance reduction was observed when compared to reference wood fibers. A mass balance indicated reduced fiber recovery when higher biogas production was obtained. The liquid fraction from the pilot dry AD was characterized for its nutrient content and used as a biofertilizer in another study. In contrast, the leachate collected from the large-scale dry AD had a low nitrogen content and high chloride content that could hinder its further use. Finally, a regional market analysis was conducted showing that the biocomposites produced with the available grass fibers could substitute at least half of the current European market based on our results.

Switching from wet to dry anaerobic digestion of food waste with different dilution times under no mechanical mixing condition

Previous research on anaerobic digestion of food waste has primarily focused on either wet or dry anaerobic digestion (AD), typically accompanied by continuous mechanical mixing. However, the necessary dilution rates and the extent of mixing required have yet to be addressed. In this study, we investigated switching from wet to dry AD of food waste without mechanical mixing, employing different dilution rates. Lab-scale anaerobic reactors were operated with dilution rates of 10, 5, and 2 times during Phases I (0–56 days), II (57–121 days), and III (122–209 days), respectively. The methane production rates were not significantly different (p > 0.05) across the dilution rates decreased from 10 to 2 times. Remarkably, the methane production in the anaerobic reactors exhibited fluctuations due to variations in feeding, with the methane production rate ranging from 2.0 to 2.7 g CH4-COD/(L d), without mechanical mixing, as the solids content transitioned from wet to near-dry digestion conditions (15 %, food waste). The distribution of sludge volatile solids concentrations remained uniform in the reactor, even at high solids concentrations of up to 15 %. A dynamic microbial community response to changes in dilution rates, with a shift from aceticlastic to hydrogenotrophic methanogenesis pathways. Syntrophic acetate oxidization bacteria (the genus Syner-01 (4.2–8.9 %) and f_Synergistaceae (3.6–4.2 %)) were highly enriched as switching from wet AD to dry AD. The study's findings provide crucial operational insights for anaerobic food waste treatment, potentially resulting in decreased water usage and operational costs, particularly in scenarios with low dilution rates and without mechanical mixing.

Operational process stability in pilot dry anaerobic digester of source‐sorted organic fraction municipal solid waste

AbstractThe organic fraction of municipal solid waste (OFMSW) is a major portion of solid waste in Malaysia, with 44.5% of the total waste being food waste‐derived sources. This study investigates the performance of dry anaerobic digester (DAD) operation using the pilot dry anaerobic digester (PDAD), a plug flow reactor, in treating source‐sorted organic fraction municipal solid waste (SS‐OFMSW) for biogas production. A commercial Malaysian food waste (CMFW) sample has been used to represent SS‐OFMSW. The anaerobic digestion was performed in a semi‐continuous operation using a 15 m3 PDAD with organic loading rates (OLRs) ranging from .63 to 5.46 kg volatile solid (VS)/m3·day under mesophilic conditions. The maximum methane composition was achieved at 56.0% at OLR 5.17 kg VS/m3·day with specific methane production (SMP) of .57 m3·CH4/kg VSfed and gas production rate (GPR) 5.27 m3·gas/m3·digester·day. As indicated by a pH and alkalinity ratio, the PDAD system was stable ranging from pH 6.7 to 8.3, alkalinity ratio of .3 with an inclination of total ammonia nitrogen (TAN) up to 1056 mg/L. The SMP achieved is between 1.58 and .4 m3·CH4/kg VSfed and potentially to fuelled 475 MW commercial biogas plant fed by CMFW. The DAD deployment strengthened the circular economy and decarbonization initiatives.

A Review on Dry Anaerobic Digestion: Existing Technologies, Performance Factors, Challenges, and Recommendations

With the increase in the growing rate of municipal solid waste throughout the world and due to the high moisture and organic components of the organic fraction of municipal solid waste, dry anaerobic digestion has become the future direction to cope with this waste while reducing the impact on the environment, including climate change. Dry anaerobic digestion has become a promising technology that converts the organic fraction of municipal solid waste into combustible biogases, which can be used as an alternative energy source. However, the technology faces several challenges that must be addressed to enhance its performance and adoption. This paper provides a comprehensive analysis of the current technologies used for dry anaerobic digestion in OFMSW and delves into the various factors that influence the performance of these technologies. This review paper also identifies and discusses the challenges faced in optimizing and scaling up these technologies, such as feedstock pretreatment requirements, characteristics of inoculum, and other crucial parameters.

Analysis of energy return on investment of dry anaerobic digestion for low water alperujo with oxidative, thermal and alkaline pretreatments

AbstractAnaerobic digestion processes for biogas generation using alperujo have been the subject of intensive studies suggesting that phenolic compounds act as microbial inhibitors in anaerobic digestion. Pretreatments are needed to reduce the effects of phenolic compounds and improve biogas production, especially for dry anaerobic digestion. However, industrial‐scale implementation of these pretreatments is challenging, and it is unclear whether the improvement in biogas production justifies the energy expended on pretreatment. This study examines the energy analysis of dry anaerobic digestion and three alternative pretreatments: alkaline, oxidative and thermal. Results indicate that thermal pretreatment at 80°C with added water reduces phenolic compounds in alperujo by 35.4%. Meanwhile, pretreated with hydrogen peroxide in alkaline medium had the highest methane productivity (205 mL CH4/gVS). Even so, thermal pretreatment was only one with an energy return on investment greater than 1, signifying the necessity for energy analysis to ensure the viability of pretreatment processes.

Effect of different organic loads on the performance and microbial community mechanism of dry anaerobic digestion

In this study, the vertical continuous push-flow dry anaerobic digestion (AD) reactor was developed independently. Typical agricultural organic waste straw and cow dung were used as raw materials in this paper to carry out long-term continuous mixed dry AD experiments on a controlled laboratory scale. The experimental results showed that the system was in low-stress operation at an organic load rate (OLR) of 3 g VS L-1 d-1, the VS methane production rate was located at 225 ∼ 272 mL CH4 g VS-1, and the volumetric methane production rate was located at 0.95 ± 0.08 L (L·d)-1, and the combination of the volumetric methane production rate, VS methane production rate, methane content of the gas, and the VS removal rate showed that the system's operational performance in this stage was in the best condition in the whole set of experiments. When the OLR reached 25 g VS L-1 d-1, the content of VFAs all began to climb rapidly above 8000 mg/L, with the content of acetic acid increasing most significantly due to the fact that the activity of acetic acid-type methanogenic bacteria was the first to be inhibited. Eventually, because of the kinetic differences between the hydrolytic acidifying bacteria and the methanogenic archaea, the final acidification destabilisation of the anaerobic digestion reactor occurred and the system was irreversibly destabilised. The system performance at this stage was in the best condition in the whole set of experiments. The study of microbial community succession and metabolic mechanisms showed that Methanoculleus and Methanobacterium play a key role in sustaining methane production, and the increased abundance and dominance of hydrogen methanogens implies that methane is produced via the hydrogen methanogenic pathway under high organic loading conditions.