Effect of chlorinated aliphatic hydrocarbons on the acetoclastic methanogenic activity of granular sludge

The toxicity of chlorinated aliphatic hydrocarbons on acetoclastic methanogens in anaerobic granular sludge was determined using a standardized anaerobic bioassay method. Most of the chloroaliphatics tested were strong inhibitors of methanogenesis. Tri- and tetrachloride derivatives of methane and ethane were the most highly toxic compounds tested, with concentrations of less than 18 mg/l resulting in 50% inhibition (IC₅₀) of the methanogenic activity. Dichlorinated compounds were less toxic, with IC₅₀ values ranging from 40 mg/l to 100 mg/l. On the other hand, perchlorinated derivatives of ethane and ethene were scarcely inhibitory at concentrations near their maximum water solubility. The toxicity caused by chlorinated aliphatic hydrocarbons was reversible. The comparison of structurally related compounds indicated that unsaturated chloroaliphatics were less toxic than their saturated counterparts. A reverse correlation between the electric dipole moment of these compounds and their methanogenic toxicity is discussed. Received: 9 July 1996 / Received revision: 11 October 1996 / Accepted: 18 October 1996     

Cobalt sorption onto anaerobic granular sludge: isotherm and spatial localization analysis

This study investigated the effect of different feeding regimes on the cobalt sorption capacity of anaerobic granular sludge from a full-scale bioreactor treating paper mill wastewater. Adsorption experiments were done with non-fed granules in monometal (only Co) and competitive conditions (Co and Ni in equimolar concentrations). In order to modify the extracellular polymeric substances and sulfides content of the granules, the sludge was fed for 30 days with glucose (pH 7, 30 degrees C, organic loading rate=1.2 g glucose l(-1) day-1) in the presence (COD/SO4(2-)=1) or absence of sulfate. The partitioning of the sorbed cobalt between the exchangeable, carbonates, organic matter/sulfides and residual fractions was determined using a sequential extraction procedure (modified Tessier). Experimental equilibrium sorption data for cobalt were analysed by the Langmuir, Freundlich and Redlich-Peterson isotherm equations. The total Langmuir maximal sorption capacity of the sludge fed with glucose and sulfate loaded with cobalt alone displayed a significantly higher maximal cobalt sorption (Qmax =18.76 mg g-1 TSS) than the sludge fed with glucose alone (Qmax =13.21 mg g-1 TSS), essentially due to an increased sorption capacity of the exchangeable (30-107%) and organic/sulfides fractions (70-30%). Environmental scanning electron microscopy coupled with an energy dispersive X-ray analysis of granular cross-sections showed that mainly iron minerals (i.e. iron sulfides) were involved in the cobalt accumulation. Moreover, the sorbed cobalt was mainly located at the edge of the granules. The sorption characteristics of the exchangeable and carbonates fractions fitted well to the Redlich-Peterson model (intermediate multi-layer sorption behaviour), whereas the sorption characteristics of the organic matter/sulfides and residual fractions fitted well to the Langmuir model (monolayer sorption behaviour). The organic matter/sulfides fraction displayed the highest affinity for cobalt for the three sludge types investigated.     

Toxicity of inorganic cadmium salts to the microalga Scenedesmus armatus (Chlorophyta) with respect to medium composition, pH and CO2 concentration

Batch cultures of algae grown at low (0.1 %) and elevated (2.0 %) concentrations of CO₂, as well as in original BBM (Bold Basal Medium) and BBM modified with phosphate, EDTA and a combination of both, were exposed to cadmium (Cd(NO₃)₂·4H₂O, 3CdSO₄·8H₂O and CdCl₂·H₂O) for 24 h. Regardless of the salt applied, the concentration-dependent relationships of Cd toxicity were found to be biphasic, suggesting the different affinity of target sites to cadmium. Nominal values of EC₅₀ obtained for algae grown in original BBM and at low CO₂ were 18.0, 16.44 and 15.37 mg·dm⁻³ for cadmium nitrate, sulphate and chloride, respectively. However, it was estimated that 97 % of the free cadmium in the added salts were bound by components of original BBM such as EDTA, phosphates, chloride and sulphate. The effect of Cd-salts at concentrations corresponding to EC₅₀ values on algae were tested in media with 10-fold reduced phosphates (BBM-P), BBM depleted of EDTA (BBM-EDTA) and of both phosphates and EDTA (BBM-P-EDTA). For algae grown at low CO₂ and BBM-P, cadmium was about 25 % less toxic than those applied in original BBM. Cadmium greatly inhibited (about 85 % of the control) the growth of algae cultured in BBM-EDTA; this effect was only slightly dependent on the CO₂ concentration. Deficits of both EDTA and P led to effects similar to those brought about by the absence of EDTA only. The toxicity of cadmium depends on CO₂ concentration only when algae are grown in original BBM. The growth of algae under high CO₂ conditions was reduced considerably less (about 80% of control) compared with low CO₂ concentrations (about 50 % of control). A relationship was found between the toxicity of cadmium salts and final pH values only in variants of low-CO₂ grown algae; with an increase of medium pH the toxicity decreased. The results suggest that both growth conditions and the binding ability of the medium markedly affect the toxicity of cadmium towards microalgae.     

Effect of cobalt on the anaerobic degradation of methanol

The effect of trace elements on the methanogenesis from methanol and acetate was studied utilizing granular sludge obtained from an anaerobic wastewater treatment plant. The methanogenic activity from methanol was dramatically stimulated by the addition of a cocktail of trace elements in the basal medium. When trace elements were supplied individually, cobalt greatly stimulated methanogenesis which equalled the stimulation observed with the complete trace element mixture. No remarkable influence of any trace element was observed when acetate was used as the substrate. Two Upflow Anaerobic Sludge Blanket (UASB) reactors were operated with and without supplementation of cobalt. Cobalt greatly stimulated both acetogenesis in the initial operational phase and later methanogenesis. The cobalt sufficient column provided almost 3 times the methane productivity compared to the cobalt deprived column. At an organic loading rate of 8 g COD/l·d, 87% of the COD was converted to methane in the cobalt sufficient column. Under low cobalt concentration, methanogens compete better for cobalt than acetogens.     

Cost-effective production of Bacillus thuringiensis biopesticides by solid-state fermentation using wastewater sludge: effects of heavy metals

This study demonstrated the feasibility to produce Bacillus thuringiensis subsp. kurstaki (Btk) based biopesticides using wastewater sludge as raw materials under solid-state fermentation (SSF). More than 10¹⁰ CFU/g viable cells of Btk were obtained using sludge or its mixture with agricultural wastes. This study well considered the effect of heavy metals on Btk growth and their changes of chemical speciation caused by SSF. The IC₅₀ (concentration causing 50% inhibition in total cell biomass) for Pb(II), Cu(II), Cd(II) and Cr(III) on Btk were determined to be 227, 82, 15 and 263 mg/L, respectively. Exposure to 150 mg/L of Cu(II) severely reduced the amount and size of toxin crystals, which decreased the endotoxin synthesis and entomotoxicity potency of Btk cells. Using Tessier’s sequential extraction procedure, the exchangeable heavy metals in sludge were shown to be transformed into residual fractions after SSF, and thus significantly reduced their bioavailability and potential environmental risks.     

Influence of the activated sludge system configuration on heavy metal toxicity reduction

The effect of configuration of activated sludge systems on heavy metal toxicity was investigated. Two bench-scale completely mixed activated sludge systems were operated identically in order to determine the toxic effects of Cr(VI), Zn(II) and industrial wastewater on the activated sludge biomass. One system was operated with an aerobic selector and the other without. Batch experiments based on OECD 209 (Organisation for Economic Cooperation and Development) were performed using a respirometer to find out potential toxicity reduction effect of an aerobic selector. The IC₅₀ (concentration of a chemical that exhibits 50% respiration inhibition) values of Cr(VI), Zn(II) and industrial wastewater in the activated sludge were determined. Results indicated that the heavy metals and industrial wastewater caused less inhibitory effect on the selector activated sludge system in comparison to the conventional activated sludge system. Cr(VI) was found to exert higher inhibition on both systems.     

The effect of calcium on the treatment of fresh leachate in an expanded granular sludge bed bioreactor

This research investigated the calcium effect on the anaerobic treatment of fresh leachate in an expanded granular sludge bed (EGSB) bioreactor under mesophilic conditions. The observations show that the bioreactor, inoculated with anaerobic granular sludge, can be started up only in about 40 days for the treatment of calcium-containing fresh leachate with chemical oxygen demand (COD) removal efficiency above 90% and organic loading rate up to 72.84 kg COD/m³ day. The calcium accumulation onto the granules was monotonically related to the calcium concentration, accounting for 17-18 wt.% of Ca in the suspended solid in the form of calcium carbonate, phosphates/phosphonates and carboxylates. The mineral formation significantly increased the granule settling velocity (by ∼50%) and the suspended solid concentration (by ∼100%). However, the effect of calcium precipitation on the specific methanogenic activity and the CH₄ production rate was complex, first positive during the start-up but later on negative.     

Inhibition of Methanogenesis from Acetate in Granular Sludge by Long-Chain Fatty Acids

The effect of four saturated long-chain fatty acids (caprylic, capric, lauric, and myristic) and one unsaturated long-chain fatty acid (oleic) on the microbial formation of methane from acetate was investigated in batch anaerobic toxicity assays. The tests were carried out with granular sludge from an upflow anaerobic sludge bed reactor. In this sludge, Methanothrix spp. are the predominant acetoclastic methanogens. Lauric acid appeared to be the most versatile inhibitor: inhibition started at 1.6 mM, and at 4.3 mM the maximum specific acetoclastic methanogenic activity had been reduced to 50%. Caprylic acid appeared to be only slightly inhibitory. Oleic acid was almost as inhibitory as lauric acid. Although adsorption of the inhibitor on the cell wall might play an important role in the mechanism of inhibition, the inhibition was found to be correlated with concentration rather than with the amount per unit of biomass. In practical situations, as in anaerobic waste treatment processes, synergism can be expected to enhance the inhibition of methanogenesis. In the present research a background concentration of lauric acid below its MIC strongly enhanced the toxicity of capric acid and (to an even greater extent) myristic acid.     

Aerobic granulation and nitrogen removal with the effluent of internal circulation reactor in start-up of a pilot-scale sequencing batch reactor

Aerobic granular sludge was successfully cultivated with the effluent of internal circulation (IC) reactor in a pilot-scale sequencing batch reactor (SBR) using activated sludge as seeding sludge. N removal was investigated in the start-up of aerobic granulation process. Initially, the phenomenon of partial nitrification was observed and nitrite accumulation rates (NO₂ ^?-N/NO _x ^? -N) were between 84.6 and 99.1?%. It was potentially caused by ammonium oxidizing bacteria (AOB) in the seeding activated sludge, high external environmental temperature (~32?°C) and free ammonia (FA) concentration. After 50?days’ running, the aerobic granules-based bioreactor demonstrated perfect performance in simultaneous removal of organic matter and ammonia nitrogen, and average removal efficiencies were maintained above 93 and 96?%, respectively. The maximum nitrogen removal efficiency of 83.1?% was achieved after the formation of aerobic granules. The average diameter of mature aerobic granular sludge mostly ranged from 0.5 to 1.0?mm. Furthermore, one typical cyclic test indicated that pH and DO profiles could be used as effective parameters for biological reactions occurring in the aerobic/anoxic process. The obtained results could provide further information on the cultivation of aerobic granular sludge with practical wastewater, especially with regard to nitrogen-rich industrial wastewater.     

Selective Inhibition by 2-Bromoethanesulfonate of Methanogenesis from Acetate in a Thermophilic Anaerobic Digestor

The effects of 2-bromoethanesulfonate, an inhibitor of methanogenesis, on metabolism in sludge from a thermophilic (58°C) anaerobic digestor were studied. It was found from short-term experiments that 1 μmol of 2-bromoethanesulfonate per ml completely inhibited methanogenesis from ¹⁴CH₃COO⁻, whereas 50 μmol/ml was required for complete inhibition of ¹⁴CO₂ reduction. When 1 μmol of 2-bromoethanesulfonate per ml was added to actively metabolizing sludge which was then incubated for 24 h. it caused a 60% reduction in methanogenesis and a corresponding increase in acetate accumulation; at 50 μmol/ml it caused complete inhibition of methanogenesis and accumulation of acetate. H₂, and ethanol.     

Extending the diffuse layer model of surface acidity constant behaviour: II. Estimation of intrinsic acidity and electrolyte ion site binding constants

Abstract

The main objective of this work was to study the capacity of bioaccumulation of copper in an upflow anaerobic sludge blanket (UASB) reactor, for the treatment of acid mine drainages (AMD). UASB reactor of 0.8 L was fed with sodium acetate, the organic loading rate (OLR) was set at 2.0gCOD/Ld, and experiments were performed in the reactor during 115 days. After 30 days, the COD removal efficiency was 90%. In the stationary stage, the higher production methane was 0.253 L/Ld. The reactor was added with 20 mgCu(II)/L during 30 days. The copper bioaccumulation was 98% and it was accumulated gradually in sludge until reaching a 61.6 mgCu(II)/gTS. The toxicity study demonstrated that the copper concentration at which the level of specific methanogenic activity (SMA) of the biogranules was reduced to 50% of the control (IC₅₀), was 280 mgCu(N)/L     

Anaerobic digestion of glycerol derived from biodiesel manufacturing

The anaerobic digestion of glycerol derived from biodiesel manufacturing, in which COD was found to be 1010 g/kg, was studied in batch laboratory-scale reactors at mesophilic temperature using granular and non-granular sludge. Due to the high KOH concentration of this by-product, H₃PO₄ was added to recover this alkaline catalyst as agricultural fertilizer (potassium phosphates). Although it would not be economically viable, a volume of glycerol was distilled and utilised as reference substrate. The anaerobic revalorisation of glycerol using granular sludge achieved a biodegradability of around 100%, while the methane yield coefficient was 0.306 m³ CH₄/kg acidified glycerol. Anaerobic digestion could be a good option for revalorising this available, impure and low priced by-product derived from the surplus of biodiesel companies. The organic loading rate studied was 0.21–0.38 g COD/g VSS d, although an inhibition phenomenon was observed at the highest load.     

Addressing speciation in the effect factor for characterisation of freshwater ecotoxicity—the case of copper

Purpose  

Determination of the ecotoxicity effect factor (EF) in life cycle impact assessment (LCIA) is based on test data reporting the total dissolved concentration of a substance. In spite of the recognised influence of chemical speciation and physico-chemical characteristics of the aquatic systems on toxicity of dissolved metals, these properties are not considered when calculating characterization factors (CFs) for metals. It is hypothesised that the main cause of the variation in reported EC50 values of Cu among published test results lies in different speciation patterns for Cu in the test media, and that the toxicity of Cu is predominantly caused by the free Cu²⁺ ion. Hence, the free Cu²⁺ ion concentration should substitute the total dissolved metal concentration when determining the EF.     

The effect of electrodialytic treatment and Na2H2EDTA addition on methanogenic activity of copper-amended anaerobic granular sludge: treatment costs and energy consumption

The effect of electrodialytic treatment in terms of a current density, pH and Na₂H₂EDTA addition on the methanogenic activity of copper-amended anaerobic granular sludge taken from the UASB reactor from paper mill was evaluated. Moreover, the specific energy consumption and simplified operational and treatment costs were calculated. Addition of Na₂H₂EDTA (at pH 7.7) to copper-amended sludge resulted in the highest microbial activity (62 mg CH₄-COD g VSS⁻¹ day⁻¹) suggesting that Na₂H₂EDTA decreased the toxic effects of copper on the methanogenic activity of the anaerobic granular sludge. The highest methane production (159 %) was also observed upon Na₂H₂EDTA addition and simultaneous electricity application (pH 7.7). The energy consumption during the treatment was 560, 840, 1400 and 1680 kW h m⁻³ at current densities of 0.23, 0.34, 0.57 and 0.69 mA cm⁻², respectively. This corresponded to a treatment costs in terms of electricity expenditure from 39.2 to 117.6 € per cubic meter of sludge.     

Solubility and mobility of copper,zinc and lead in acidic environments

Understanding the chemical speciation of metals in solution is necessary for evaluating their toxicity and mobility in soils. Soil samples from the Powder River Basin, Wyoming were extracted with distilled deionised H₂O. Soil water extracts were subjected to chemical speciation to determine the relative distribution and chemical forms of copper (Cu), zinc (Zn) and lead (Pb) in acidic environments. As pyrite oxidised, the pH decreased from 6.6 to 2.4, concentration of dissolved sulfate (S_T) increased from 259 to 4,388 mg L^-1 and concentration of dissolved organic carbon (DOC) decreased from 56.9 to 14.4 mg L^-1. Dissolved Cu concentrations ranged from 0.06 to 0.42 mg L^-1 and dissolved Zn concentrations ranged from 0.084 to 4.60 mg L^-1. Dissolved concentrations of Pb were found to be 0.003 to 0.046 mg L^-1. Chemical speciation indicated that at near neutral pH, dissolved metal concentration in soil water extracts was dominated by DOC- metal complexes. At low pH, dissolved metal concentration in soil water extracts was dominated by free ionic forms (e.g. Cu²⁺, Zn²⁺, Pb²⁺) followed by ion pairs (e.g. CuSO₄ ⁰, ZnSO _inf4 ^sup0 , PbSO _in4 ^sup0 ). Results obtained in this study suggest that as soil pH decreased, the availability and mobility of metal ions increased due to the chemical form in which these metal ions are present in soil solutions.     

Effect of Iron Hydroxide on Phosphate Removal during Anaerobic Digestion of Activated Sludge

The addition of iron (III) hydroxide during methanogenic digestion of activated sludge by anaerobic sludge displaying an iron-reducing activity resulted in a microbial reduction of iron (III) with the formation of iron (II), capable of precipitating phosphates. The feasibility of eliminating 66.6 to 99.6% of the dissolved phosphate at initial concentrations of 1000 to 3500 mg PO^3- ₄/l by adding 6420 mg/l iron (III) hydroxide into a reactor for anaerobic fermentation of activated sludge was analyzed. The optimal ratio of iron (III) added to dissolved phosphate removed (mg) providing a 95% removal amounted to 2 : 1. These results may be used in new technology for anaerobic wastewater treatment with phosphate removal.     

芦岭煤田微生物群落结构和生物成因气的产甲烷类型研究

【目的】揭示芦岭煤田微生物群落组成,并分析其潜在的产甲烷类型及产甲烷途径。【方法】采集芦岭煤田的煤层气样品和产出水样品,分别分析样品的地球化学性质特征;利用Illumina HiSeq高通量测序技术分析产出水中的微生物群落结构;采用添加不同底物的厌氧培养实验进一步证实芦岭煤田生物成因气的产甲烷类型。【结果】该地区煤层气为生物成因和热成因的混合成因气;古菌16S rRNA基因分析表明在产出水中含有乙酸营养型、氢营养型和甲基营养型的产甲烷菌。丰度较高的细菌具有降解煤中芳香族和纤维素衍生化合物的潜力。厌氧富集培养结果表明,添加乙酸盐、甲酸盐、H2+CO2为底物的矿井水样均有明显的甲烷产生。【结论】芦岭煤田具有丰富的生物多样性,该地区同时存在三种产甲烷类型。本研究为利用微生物技术提高煤层气的采收率,实现煤层气的可持续开采提供科学依据。     

Divalent metal addition restores sulfide-inhibited N2O reduction in Pseudomonas aeruginosa

Hydrogen sulfide (H₂S) inhibits the last step of the denitrification process, i.e. the reduction of nitrous oxide (N₂O) to dinitrogen gas (N₂), both in natural environments (marine sediments) and industrial processes (activated sludge, methanogenic sludge, BioDeNOx process). In a previously published study, we showed that the inhibitory effect of sulfide to N₂O reduction in mixed microbial communities is reversible and can be counteracted by dosing trace amounts of copper. It remained, however, unclear if this was due to copper sulfide precipitation or a retrofitting of the copper containing N₂O-reductase (N₂OR). The present study aimed to elucidate the mechanism of the restoration of sulfide-inhibited N₂O reducing activity by metal addition to a pure Pseudomonas aeruginosa culture. This was done by using other metals (zinc, cobalt and iron) in comparison with copper. Zinc and cobalt clearly alleviated the sulfide inhibition of N₂OR to the same extent as copper and the activity restoration was extremely fast (within 15 min, Fig. 3) for zinc, cobalt and copper. This suggests that the alleviation of the inhibitory effect of sulfide is due to metal sulfide precipitation and thus not exclusively limited to Cu. This work also underlines the importance of metal speciation: supply of iron did not restore the N₂OR activity because it was precipitated by the phosphates present in the medium and thus could not precipitate the sulfide.     

Removal of malachite green (MG) from aqueous solutions by native and heat-treated anaerobic granular sludge

The performance of native and heat-treated anaerobic granular sludge in removing of malachite green (MG) from aqueous solution was investigated with different conditions, such as pH, ionic strength, initial concentration and temperature. The maximum biosorption was both observed at pH 5.0 on the native and heat-treated anaerobic granular sludge. The ionic strength had negative effect on MG removal. Kinetic studies showed that the biosorption process followed pseudo-second-order and q_e for native and heat-treated anaerobic granular sludge is 61.73 and 59.17 mg/g at initial concentration 150 mg/L, respectively. Intraparticle diffusion model could well illuminate adsorption process and faster adsorption rate of native anaerobic granular sludge than heat-treated anaerobic granular sludge. The equilibrium data were analyzed using Langmuir and Freundlich model, and well fitted Langmuir model. The negative values of ΔG° and ΔH° suggested that the interaction of MG adsorbed by native and heat-treated anaerobic granular sludge was spontaneous and exothermic. Desorption studies revealed that MG could be well removed from anaerobic granular sludge by 1% (v/v) of HCl–alcohol solution.     

Diversity of Dominant Bacterial Taxa in Activated Sludge Promotes Functional Resistance following Toxic Shock Loading

Examining the relationship between biodiversity and functional stability (resistance and resilience) of activated sludge bacterial communities following disturbance is an important first step towards developing strategies for the design of robust biological wastewater treatment systems. This study investigates the relationship between functional resistance and biodiversity of dominant bacterial taxa by subjecting activated sludge samples, with different levels of biodiversity, to toxic shock loading with cupric sulfate (Cu[II]), 3,5-dichlorophenol (3,5-DCP), or 4-nitrophenol (4-NP). Respirometric batch experiments were performed to determine the functional resistance of activated sludge bacterial community to the three toxicants. Functional resistance was estimated as the 30 min IC₅₀ or the concentration of toxicant that results in a 50% reduction in oxygen utilization rate compared to a referential state represented by a control receiving no toxicant. Biodiversity of dominant bacterial taxa was assessed using polymerase chain reaction-terminal restriction fragment length polymorphism (PCR-T-RFLP) targeting the 16S ribosomal RNA (16S rRNA) gene. Statistical analysis of 30 min IC₅₀ values and PCR-T-RFLP data showed a significant positive correlation (P < 0.05) between functional resistance and microbial diversity for each of the three toxicants tested. To our knowledge, this is the first study showing a positive correlation between biodiversity of dominant bacterial taxa in activated sludge and functional resistance. In this system, activated sludge bacterial communities with higher biodiversity are functionally more resistant to disturbance caused by toxic shock loading.