Inhibition of Biogas Production by Alkyl Benzene Sulfonates (LAS) in a Screening Test for Anaerobic Biodegradability

The effect of the inoculum source on the digestion of linear alkylbenzene sulfonates (LAS) under anaerobic conditions has been investigated. The potential for primary and ultimate LAS biodegradation of anaerobic sludge samples obtained from wastewater treatment plants (WWTPs) of different geographical locations was studied applying a batch test system. It was found that only 4–22% of the LAS added to the batch anaerobic digesters was primarily transformed suggesting a poor primary degradation of the LAS molecule in anaerobic discontinuous systems. Regarding ultimate biodegradation, the addition of LAS to the batch anaerobic digesters caused a reduction on the extent of biogas production. Significant differences in the inhibition extent of the biogas production were observed (4–26%) depending on the sludge used as inoculum. Effect of the surfactant on the anaerobic microorganisms was correlated with its concentration in the aqueous phase. Sorption of LAS on anaerobic sludge affects its toxicity by depletion of the available fraction of the surfactant. LAS content on sludge was related to the total amount of calcium and magnesium extractable ions. The presence of divalent cations promote the association of LAS with anaerobic sludge reducing its bioavailability and the extent of its inhibitory effect on the biogas production.     

Methanogenic toxicity in anaerobic digesters treating municipal wastewater

In upflow anaerobic sludge blanket (UASB) digesters treating raw sewage at low temperatures, the sludge progressively lost methanogenic activity, indicating the possibility of methanogenic activity inhibition caused by wastewater constituents. To check this fact, batch and semi-continuous methanogenic toxicity assays were carried out with raw and centrifuged sewage. Permanent methanogenic toxicity on anaerobic sludge of approximately 50% was found when the sludge exposure to wastewater was renewed in a semi-continuous way. A stronger methanogenic inhibition of about 70-100% was observed when an active anaerobic sludge was exposed to mixed liquor from the UASB digester treating municipal wastewater. Suspended solids removal from sewage slightly reduced methanogenic toxicity. Effective concentration of municipal wastewater that caused a 50% reduction in methanogenic activity was estimated to be in the range of 150-200 mg CODl(-1). As methanogenic inhibition appeared to be related to remaining COD, higher methanogenic toxicity in digesters operating with low conversion efficiency will be expected.     

Microbial diversity of methanogenic communities in the systems for anaerobic treatment of organic waste

Methane production via anaerobic degradation of organic-contaminated wastewater, semiliquid, or solid municipal waste of complex composition by methanogenic microbial communities is a multistage process involving at least four groups of microorganisms. These are hydrolytic bacteria (polysaccharolytic, proteolytic, and lipolytic), fermentative bacteria, acetogenic bacteria (syntrophic, proton-reducing), and methanogenic archaea; complex trophic interactions exist between these groups. The review provides information concerning the diversity of the major microbial groups identified in the systems for wastewater and concentrated waste treatment, solid-phase anaerobic fermentation, and landfills for disposal of municipal solid waste, and also specifies the sources of isolation of the type strains. The research demonstrates that both new microorganisms and those previously isolated from natural habitats may be found in waste treatment systems. High microbial diversity in the systems for organic waste treatment provides for stable methanogenesis under fluctuating environmental conditions.     

Enhancement of dry anaerobic batch digestion of the organic fraction of municipal solid waste by an aerobic pretreatment step

The start-up of the dry anaerobic batch digestion by the BIOCEL-concept of the organic fraction of municipal solid waste (MSW) is unbalanced when a methanogenic inoculum (digested sewage sludge) is added to a total solids concentration of 35%. The unbalanced conditions are the result of the rapid degradation of easily-degradable compounds which are present in the organic fraction. Enhancement of the first start-up of the dry batch digestion was tried by applying an aerobic partial-composting step. By this aerobic treatment the easily degradable compounds are removed. After the composting step the anaerobic digestion will be limited by the conversion of the ligno-cellulose part of the organic fraction. It appeared that at least 19·5% of the volatile solids (VS) should be converted during the aerobic composting period before acid formation in the digestion was in balance with the methane formation. This amount of aerobically degraded VS means a 40% loss of potential biogas. The loss of a part of the biogas is a major drawback to the partial composting as a method for enhancing the start-up of the dry anaerobic digestion. A shorter composting period which is combined with another start-up method might be a feasible method to decrease the energy input of the dry digestion process.     

The action of antibiotics on the anaerobic digestion process

Antibiotics can disturb the production of biogas during anaerobic digestion. This study shows a systematic approach to understanding how the different bacterial populations involved in the final conversion of organic matter into methane are inhibited by 15 antimicrobial agents with different specificities and modes of action. The results obtained show the following trends: (i) some inhibitors, such as the macrolide erythromycin, lack any inhibitory effect on biogas production; (ii) some antibiotics, with different specificities, have partial inhibitory effects on anaerobic digestion and decrease methane production by interfering with the activity of propionic-acid- and butyric-acid-degrading bacteria,␣(e.g. antibiotics that interfere with cell wall synthesis, RNA polymerase activity and protein synthesis, especially the aminoglycosides); (iii) the protein synthesis inhibitors chlortetracycline (IC₅₀ 40 mg l⁻¹) and chloramphenicol (IC₅₀ 15–20 mg l⁻¹) are very powerful inhibitors of anaerobic digestion. The majority of the antibiotics tested lacked activity against acetoclastic methanogens, being active only on the acetogenic bacteria. However, chloramphenicol and chlortetracycline could cause the complete inhibition of the acetoclastic methanogenic archaea. Received: 6 February 1996 / Received revision: 24 July 1996 / Accepted: 5 August 1996     

Influence of methane enrichment by aeration of recirculated supernatant on microbial activities during anaerobic digestion

A methane enrichment process (MEP) was evaluated that involved air purging of recycled digester contents to strip CO₂ and increase biogas methane content. The objective of this work was to determine if the aeration resulted in oxygen inhibition of microbial activities involved in anaerobic digestion of municipal solid waste. To assess the degree of biological perturbation associated with the MEP, the reactor effluent was sampled twice while the MEP was operating and twice while it was not operating. Analyses were run on composite samples (representing several different sample ports of the non-mixed reactor) and samples of digester effluent entering and leaving the MEP process. The analyses included volatile organic acids (VOA), effluent solids content, dehydrogenase activity, specific methanogenic activities (SMA), and enzyme-linked immunosorbent assay (ELISA) for methanogenic, sulfate-reducing, and cellulolytic bacterial species. Methane enrichment in these experiments occurred with the methane content of the biogas exceeding 90%. There were no effects of the MEP on effluent VOA concentration. The MEP had no effect on volatile solids (VS) levels of composite samples representing the non-mixed digester contents, however, VS was reduced in effluent passing through the MEP. Although MEP had an inhibitory effect on anaerobic populations leaving the MEP process, no inhibitory effects were observed in measurements of microbial activities in digester samples, including specific methane production rate, dehydrogenase, and numbers of specific organisms estimated using ELISA techniques.     

Metabolic interactions between anaerobic bacteria in methanogenic environments

In methanogenic environments organic matter is degraded by associations of fermenting, acetogenic and methanogenic bacteria. Hydrogen and formate consumption, and to some extent also acetate consumption, by methanogens affects the metabolism of the other bacteria. Product formation of fermenting bacteria is shifted to more oxidized products, while acetogenic bacteria are only able to metabolize compounds when methanogens consume hydrogen and formate efficiently. These types of metabolic interaction between anaerobic bacteria is due to the fact that the oxidation of NADH and FADH₂ coupled to proton or bicarbonate reduction is thermodynamically only feasible at low hydrogen and formate concentrations. Syntrophic relationships which depend on interspecies hydrogen or formate transfer were described for the degradation of e.g. fatty acids, amino acids and aromatic compounds.     

Anaerobic waste-activated sludge digestion-a bioconversion mechanism and kinetic model

The anaerobic bioconversion of raw and mechanically lysed waste-activated sludge was kinetically investigated. The hydrolysis of the biopolymers, such as protein, which leaked out from the biological sludge with ultrasonic lysis, was a first-order reaction in anaerobic digestion and the rate constant was much higher that the decay rate constant of the raw waste activated sludge. An anaerobic digestion model that is capable of evaluating the effect of the mechanical sludge lysis on digestive performance was developed. The present model includes four major biological processes-the release of intracellular matter with sludge lysis; hydrolysis of biopolymers to volatile acids; the degradation of various volatile acids to acetate; and the conversion of acetate and hydrogen to methane. Each process was assumed to follow first order kinetics. The model suggested that when the lysed waste-activated sludge was fed, the overall digestive performance remarkably increased in the two-phase system consisting of an acid forming process and a methanogenic process, which ensured the symbiotic growth of acetogenic and methanogenic bacteria. (c) 1993 Wiley & Sons, Inc.     

Exocellular electron transfer in anaerobic microbial communities

Exocellular electron transfer plays an important role in anaerobic microbial communities that degrade organic matter. Interspecies hydrogen transfer between microorganisms is the driving force for complete biodegradation in methanogenic environments. Many organic compounds are degraded by obligatory syntrophic consortia of proton-reducing acetogenic bacteria and hydrogen-consuming methanogenic archaea. Anaerobic microorganisms that use insoluble electron acceptors for growth, such as iron- and manganese-oxide as well as inert graphite electrodes in microbial fuel cells, also transfer electrons exocellularly. Soluble compounds, like humic substances, quinones, phenazines and riboflavin, can function as exocellular electron mediators enhancing this type of anaerobic respiration. However, direct electron transfer by cell-cell contact is important as well. This review addresses the mechanisms of exocellular electron transfer in anaerobic microbial communities. There are fundamental differences but also similarities between electron transfer to another microorganism or to an insoluble electron acceptor. The physical separation of the electron donor and electron acceptor metabolism allows energy conservation in compounds as methane and hydrogen or as electricity. Furthermore, this separation is essential in the donation or acceptance of electrons in some environmental technological processes, e.g. soil remediation, wastewater purification and corrosion.     

Anaerobic treatment of baker's yeast wastewater: I. Start-up and sodium molybdate addition

The anaerobic treatment of baker's yeast wastewater was studied using an anaerobic biological contact reactor (AnRBC) and a fixed-film reactor. The AnRBC had an active biomass developed within the reactor before this study commenced; however, the fixed-film reactor was started without attached biomass in a support structure. The gas production rates obtained for the AnRBC were between 0·55 and 0·61 litre methane per litre reactor per day. However, a gas production rate of only 0·46 litre methane per litre reactor per day was achieved after a four-month operating period for the fixed-film reactor. Higher chemical oxygen demand reduction was also found in the AnRBC. The results indicated that the presence of high sulfate concentration in baker's yeast wastewater affected teh start-up process. The reactor with fully developed active biomass was less susceptible to sulfate inhibition and showed improved anaerobic digestion. Results indicate that the reactor should be innoculated by feeding nutrient-balanced substrate before it was subjected to the digestion of baker's yeast wastewater. The fixed-film reactor was also fed with the substrate contianing sodium molybdate, an inhibitor of sulfate-reducing bacteria. The results indicated that both methanogenic and sulfate-reducing bacteria were inhibited.     

Feasibility of methanogenic digestion applied to a low pH acetic acid solution

This paper discusses the methanogenic digestion of a synthetic acetic acid wastewater inoculated using a mixed culture obtained from an anaerobic digester at a municipal wastewater treatment facility. Experiments were conducted in 500mL batch reactors containing an unbuffered acetic acid solution. Test conditions compared methane production and acetic acid degradation at both acidic (pH 4.5) and neutral (pH 7.0) initial system conditions. Results showed that methane production increased by 30% when the initial pH was decreased from 7.0 to 4.5.     

Toxic effects of thiol-reactive compounds on anaerobic biomass

The objective of this study was to characterize the toxic effects of three well known thiol-reactive electrophilic compounds, N-ethylmaleimide (NEM), pentachlorophenol (PCP) and 1-chloro-2,4-dinitrobenzene (CDNB) on anaerobic biotransformation process. The work was part of a larger investigation on potassium efflux as a possible response mechanism of anaerobic microorganisms to the presence of thiol-reactive organic compounds and the interference of such compounds on the reductive dehalogenation process. Using anaerobic toxicity assay (ATA) and granular anaerobic biomass from a full-scale upflow anaerobic sludge blanket (UASB) reactor, inhibitory concentrations of these compounds that reduced the microbial activity of granular biomass to 50% of a control (IC50) were determined to be 592, 0.97, and 450 mg/l for NEM, PCP, and CDNB, respectively. Toxicity of NEM was also tested on anaerobic biomass from a municipal wastewater treatment plant digester and slightly lower IC50 of 532 mg/l was obtained. The results presented here indicate that anaerobic biomass can acclimate to the three thiol-reactive compounds studied and recover from inhibition as long as the toxicant concentration is below a threshold level. That threshold concentration was found to be 500 mg/l for NEM on biomass from the municipal digester, 1 mg/l for PCP, and 500 mg/l for CDNB, both on granular biomass. Granular anaerobic biomass showed recovery even at NEM concentrations of 1000 mg/l.     

The effect of aromatic structure on the inhibition of acetoclastic methanogenesis in granular sludge

Summary Benzene derivatives are important constituents of certain effluents discharged by pulp and paper, petrochemical and chemical industries. The anaerobic treatment of these waste-waters can be limited due to methanogenic inhibition exerted by aromatic compounds. The objective of this study was to evaluate the effect of aromatic structure on acetoclastic methanogenic inhibition. The toxicity to acetoclastic methanogens was assayed in serum flasks utilizing granular sludge as inoculum. Among the monosubstituted benzenes, chlorobenzene, methoxybenzene and benzaldehyde were the most toxic with 50% inhibition occurring at concentrations of 3.4, 4.2 and 5.2 mm, respectively. In contrast, benzoate was the least inhibitory: concentrations up to 57.3 mm were non-toxic. In general, the toxicity of aromatic compounds increased with increasing length of aliphatic side-chains, increasing the number of alkyl or chlorine groups. The logarithm of the partition coefficient octanol/water (log P), an indicator of hydrophobicity, was observed to be positively correlated with the methanogenic inhibition. The results indicate that hydrophobicity is an important factor contributing to the high toxicity of the most inhibitory aromatic compounds.     

The effect of calcium on anaerobic digestion

Summary The ability of methanogenic bacteria to adapt to high concentrations of calcium was investigated using a 9 l Upflow Anaerobic Sludge Blanket (UASB) reactor fed semi-continuously with a synthetic waste. It was found that a calcium concentration of up to 7000 mg/l had no inhibitory effect on anaerobic digestion.     

Removal of molecular weight fractions of COD and phenolic compounds in an integrated treatment of olive oil mill effluents

Previous works (Beccari et al. 1999b; Beccari et al. 2001a; Beccari et al. 2001b)on the anaerobic treatment of olive oil mill effluents (OME) have shown: (a) apre-treatment based on the addition of Ca(OH)₂ and bentonite was able toremove lipids (i.e. the most inhibiting substances present in OME) almostquantitatively; (b) the mixture OME – Ca(OH)₂ – bentonite, fed to amethanogenic reactor without providing an intermediate phase separation,gave way to high biogas production even at very low dilution ratios; (c) theeffluent from the methanogenic reactor still contained significant concentrationsof residual phenolic compounds (i.e. the most biorecalcitrant substances present inOME). Consequently, this paper was aimed at evaluating the fate of the phenolicfractions with different molecular weights during the sequence of operations(adsorption on bentonite, methanogenic digestion, activated sludge post-treatment).The results show that a very high percentage (above 80%) of the phenolic fractionbelow 500 D is removed by the methanogenic process whereas the phenolic fractionsabove 1,000 D are significantly adsorbed on bentonite; the 8-day activated sludgepost-treatment allows an additional removal of about 40% of total filtered phenoliccompounds. The complete sequence of treatments was able to remove more than the96% of the phenolic fraction below 500 D (i.e. the most toxic fraction towards plantgermination). Preliminary respirometric tests show low level of inhibition exerted bythe effluent from the methanogenic reactor on aerobic activated sludges taken fromfull-scale municipal wastewater plants.     

Biomass adaptation over anaerobic co-digestion of sewage sludge and trapped grease waste

The feasibility of sewage sludge co-digestion using intermediate waste generated inside a wastewater treatment plant, i.e. trapped grease waste from the dissolved air flotation unit, has been assessed in a continuous stirred lab reactor operating at 35 °C with a hydraulic retention time of 20 days. Three different periods of co-digestion were carried out as the grease waste dose was increased. When the grease waste addition was 23% of the volatile solids fed (organic loading rate 3.0 kg_COD m⁻³ d⁻¹), an increase in methane yield of 138% was reported. Specific activity tests suggested that anaerobic biomass had adapted to the co-substrate. The adapted inoculum showed higher acetoclastic methanogenic and β-oxidation synthrophic acetogenic activities but lower hydrogenotrophic methanogenic activity. The results indicate that a slow increase in the grease waste dose could be a strategy that favours biomass acclimation to fat-rich co-substrate, increases long chain fatty acid degradation and reduces the latter’s inhibitory effect.     

Thermodynamic equilibrium model in anaerobic digestion process

Catabolic reactions provide the chemical energy necessary for the maintenance of living microorganisms. The catabolic reactions in anaerobic digestion process may progress close to the equilibrium state (ΔG = 0) depending strongly on the microorganisms in the digester. The thermodynamic equilibrium of catabolic reactions in the anaerobic digestion process was modelled under isothermal and isobaric conditions. Three thermodynamic models were considered; the ideal, the Debye-Hückel–Praunitz, and the Pitzer–Praunitz. The models in this paper concentrate on the methanogenic equilibrium of the anaerobic digestion process. The thermodynamic equilibrium model shows that the methanogenesis step requires thermal energy and electrons, so that anaerobic digestion may achieve high substrate degradation and high conversion to methane. Some thermodynamic recommendations are suggested for the future development of the methanogenic phase of anaerobic digestion.     

Responses of the biogas process to pulses of oleate in reactors treating mixtures of cattle and pig manure

The effect of oleate on the anaerobic digestion process was investigated. Two thermophilic continuously stirred tank reactors (CSTR) were fed with mixtures of cattle and pig manure with different total solid (TS) and volatile solid (VS) content. The reactors were subjected to increasing pulses of oleate. Following pulses of 0.5 and 1.0 g oleate/L, the most distinct increase in volatile fatty acid (VFA) concentrations were observed in the reactor with the lowest TS/VS content. This suggests a higher adsorption of oleate on the surfaces of biofibers in the reactor with the highest TS/VS and a less pronounced inhibition of the anaerobic digestion process. On the other hand, addition of 2.0 g oleate/L severely inhibited the process in both reactors, and a significant increase in all VFA concentrations combined with an immediate drop in methane production was noticed. However, 20 days after the reactors had been exposed to oleate both reactors showed a lower VFA concentration along with a higher methane production than before the pulses. This indicates that oleate had a stimulating effect on the overall process. The improved acetogenic and methanogenic activity in the reactors was confirmed in batch activity tests. In addition to this, toxicity tests revealed that the oleate pulses induced an increase in the tolerance level of acetotrophic methanogens towards oleate. When evaluating the usability of different process parameters (i.e., VFA and methane production) as indicators of process recovery, following the inhibition by oleate, propionate was found to be most suitable.     

Strain differentiation of epidemic typhus rickettsiae (Rickettsia prowazekii) by DNA restriction endonuclease analysis

We have investigated the number of methanogenic bacteria in different samples of fresh feces and effluents from the anaerobic digestion of cattle and swine waste and municipal sludges. We have further compared conventional anaerobic techniques with the rigorous, oxygen-free, anaerobic, experimental conditions obtained in a controlled glove cabinet atmosphere. It was found that the total number of oxygen-intolerant methanogenic bacteria was about 10–100 times higher than that of anaerobic oxygen-tolerant methanogenic bacteria. We have also developed a simple technique for the growth of methanogenic bacteria that permits the realization of rigorous anaerobic conditions without the use of a glove cabinet.