Study of microbial community structures in UASB sludge treating municipal wastewater by denaturing gradient gel electrophoresis of 16S rDNA

The structures of microbial communities in lab-scale upflow anaerobic sludge blanket (UASB) reactors for treating municipal wastewater with different ratios of COD soluble/COD total were studied using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes.The microbial structure of the inoculum sludge obtained from a full-scale UASB reactor of treating potato processing wastewater was compared with the structures of sludges collected from three lab-scale UASB reactors after eight months feeding with raw municipal wastewater, with CEPS (chemically enhanced primary sedimentation) pretreated municipal wastewater, and with a synthetic municipal sewage, respectively. Computer-aided numerical analysis of the DGGE fingerprints showed that the bacterial community underwent major changes. The sludges for treating raw and CEPS pretreated wastewater had very similar bacterial and archaeal communities (82%and 96% similarity) but were different from that for treating the synthetic sewage. Hence, despite similar % COD in the particulate form in the synthetic and the real wastewater, the two wastewaters were selected for different microbial communities. Prominent DGGE bands of Bacteria and Archaea were purified and sequenced. The 16S rRNA gene sequences of the dominant archaeal bands found in the inoculum, and UASB sludge fed with raw sewage, CEPS pretreated wastewater, and synthetic sewage were closely associated with Methanosaeta concilii. In the UASB sludge fed with synthetic sewage, another dominant band associated with an uncultured archaeon 39-2 was found together with M. concilii.     

Study of microbial community structures in UASB sludge treating municipal wastewater by denaturing gradient gel electrophoresis of 16S rDNA

The structures of microbial communities in lab-scale upflow anaerobic sludge blanket (UASB) reactors for treating municipal wastewater with different ratios of COD soluble/ COD total were studied using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes. The microbial structure of the inoculum sludge obtained from a full-scale UASB reactor of treating potato processing wastewater was compared with the structures of sludges collected from three lab-scale UASB reactors after eight months feeding with raw municipal wastewater, with CEPS (chemically enhanced primary sedimentation) pretreated municipal wastewater, and with a synthetic municipal sewage, respectively. Computer-aided numerical analysis of the DGGE fingerprints showed that the bacterial community underwent major changes. The sludges for treating raw and CEPS pretreated wastewater had very similar bacterial and archaeal communities (82% and 96% similarity) but were different from that for treating the synthetic sewage. Hence, despite     

Study of microbial community structures in UASB sludge treating municipal wastewater by denaturing gradient gel electrophoresis of 16S rDNA

The structures of microbial communities in lab-scale upflow anaerobic sludge blanket (UASB) reactors for treating municipal wastewater with different ratios of COD_soluble/ COD_total were studied using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes. The microbial structure of the inoculum sludge obtained from a full-scale UASB reactor of treating potato processing wastewater was compared with the structures of sludges collected from three lab-scale UASB reactors after eight months feeding with raw municipal wastewater, with CEPS (chemically enhanced primary sedimentation) pretreated municipal wastewater, and with a synthetic municipal sewage, respectively. Computer-aided numerical analysis of the DGGE fingerprints showed that the bacterial community underwent major changes. The sludges for treating raw and CEPS pretreated wastewater had very similar bacterial and archaeal communities (82% and 96% similarity) but were different from that for treating the synthetic sewage. Hence, despite similar % COD in the particulate form in the synthetic and the real wastewater, the two wastewaters were selected for different microbial communities. Prominent DGGE bands of Bacteria and Archaea were purified and sequenced. The 16S rRNA gene sequences of the dominant archaeal bands found in the inoculum, and UASB sludge fed with raw sewage, CEPS pretreated wastewater, and synthetic sewage were closely associated withMethanosaeta concilii. In the UASB sludge fed with synthetic sewage, another dominant band associated with an uncultured archaeon 39-2 was found together withM. concilii.     

Effect of sulfur source on the performance and metal retention of methanol-fed UASB reactors

The effect of a sulfur source on the performance and metal retention of methanol-fed upflow anaerobic sludge bed (UASB) reactors was investigated. For this purpose, two UASB reactors were operated with cobalt preloaded granular sludge (1 mM CoCl2; 30 degrees C; 24 h) at an organic loading rate (OLR) of 5 g COD.L reactor(-1).d(-1). One UASB reactor (R1) was operated without a sulfur source in the influent during the first 37 days. In this period the methanol conversion to methane remained very poor, apparently due to the absence of a sulfur source, because once cysteine, a sulfur-containing amino acid, was added to the influent of R1 (day 37) a full conversion of methanol to methane occurred within 6 days. The second reactor (R2) was operated with sulfate (0.41 mM) in the influent during the first 86 days of operation, during which no limitation in the methanol conversion to methane manifested. Cobalt washed out from the sludge at similar rates in both reactors. The leaching of cobalt occurred at two distinct rates, first at a high rate of 22 microg.g TSS(-1).d(-1), which proceeded mainly from the exchangeable and carbonate fraction and later at a relatively slow rate of 9 mug.g TSS(-1).d(-1) from the organic/sulfide fraction. This study showed that the supply of the sulfur source L-cysteine has a pronounced positive effect on the methanogenic activity and the retention of metals such as iron, zinc and molybdenum.     

The inhibitory effects and removal of dieldrin in continuous upflow anaerobic sludge blanket reactors

The inhibitory effects and removal efficiency of dieldrin (DLD) in anaerobic reactors were investigated. Anaerobic toxicity assay (ATA) experiments conducted in batch reactors revealed that 30 mg/l DLD had inhibitory effects on the unacclimated mixed anaerobic cultures. Continuous reactor experiments performed in a lab-scale two-stage upflow anaerobic sludge blanket (UASB) reactor system which was fed with ethanol as the sole carbon source, indicated that anaerobic granular cultures could be successfully acclimated to DLD. Chemical oxygen demand (COD) removal efficiencies were 88-92% for the two-stage system. The influent DLD concentration of 10 mg/l was removed by 44-86% and 86-94% in the second stage and overall UASB system, respectively. Biosorption of DLD on granular anaerobic biomass was found to be a significant mechanism for DLD removal in the UASB system. The maximum DLD loading rate and minimum HRT achievable for the first stage UASB reactor were 0.5 mg/lday (76 microg DLD/g VSS.day) and 10 h, respectively, which resulted in the overall COD removal efficiency of 85%.     

Controlled shear affinity filtration (CSAF): a new technology for integration of cell separation and protein isolation from mammalian cell cultures

Up-flow anaerobic sludge blanket (UASB) reactors are being used with increasing regularity all over the world, especially in India, for a variety of wastewater treatment operations. Consequently, there is a need to develop methodologies enabling one to determine UASB reactor performance, not only for designing more efficient UASB reactors but also for predicting the performance of existing reactors under various conditions of influent wastewater flows and characteristics. This work explores the feasibility of application of an artificial neural network-based model for simulating the performance of an existing UASB reactor. Accordingly, a neural network model was designed and trained to predict the steady-state performance of a UASB reactor treating high-strength (unrefined sugar based) wastewater. The model inputs were organic loading rate, hydraulic retention time, and influent bicarbonate alkalinity. The output variables were one or more of the following, effluent substrate concentration (Se), reactor bicarbonate alkalinity, reactor pH, reactor volatile fatty acid concentration, average gas production rate, and percent methane content of the gas. Training of the neural network model was achieved using a large amount of experimentally obtained reactor performance data from the reactor mentioned above as the training set. Training was followed by validation using independent sets of performance data obtained from the same UASB reactor. Subsequently, simulations were performed using the validated neural network model to determine the impact of changes in parameters like influent chemical oxygen demand (COD) concentration and hydraulic retention time on the reactor performance. Simulation results thus obtained were carefully analyzed based on qualitative understanding of UASB process and were found to provide important insights into key variables that were responsible for influencing the working of the UASB reactor under varying input conditions.     

Comparison of long-term performances and final microbial compositions of anaerobic reactors treating landfill leachate

Laboratory scale anaerobic upflow filter, sludge blanket and hybrid bed reactors were operated for 860 days in the treatment of high ammonia landfill leachate. Organic loading was gradually increased from 1.3 to 23.5 kg COD/m3 day in the start-up period and then fluctuated according to the COD concentration of raw leachate. To prevent free ammonia inhibition, influent pH was reduced to 4.5 after Day 181 and consequently COD removal efficiencies above 80% were achieved in all reactors. However, the anaerobic filter and hybrid bed reactor were generally found slightly more efficient and stable than the UASB reactor. In addition to conventional anaerobic reactor control parameters, the complementary techniques of denaturing gradient gel electrophoresis (DGGE), cloning and fluorescent in situ hybridization (FISH) were used to identify and compare the microbial profiles in the reactors at Day 830. Molecular analyses revealed that acetoclastic Methanosaeta species were prevalent in all reactors and configuration did not have an impact on microbial diversity in the long-term.     

Performance of staged and non-staged up-flow anaerobic sludge bed (USSB and UASB) reactors treating low strength complex wastewater

The use of anaerobic processes to treat low-strength wastewater has been increasing in recent years due to their favourable performance-costs balance. For optimal results, it is necessary to identify reactor configurations that are best suited for this kind of application. This paper reports on the comparative study carried out with two high-rate anaerobic reactor systems with the objective of evaluating their performances when used for the treatment of low-strength, complex wastewater. One of the systems is the commonly used up-flow anaerobic sludge blanket (UASB) reactor. The other is the up-flow staged sludge bed (USSB) system in which the reactor was divided longitudinally into 3, 5 and 7 compartments by the use of baffles. The reactors (9 l) were fed with a synthetic, soluble and colloidal waste (chemical oxygen demand (COD) < 1000 mg/l) and operated at 28°C and 24 h hydraulic retention time. Intermediate flow hydraulics, between plug-flow and completely-mixed, in the UASB and 7 stages USSB reactors allowed efficient degradation of substrates with minimum effluent concentrations. Low number of compartments in the USSB reactors increased the levels of short-circuiting thus reducing substrate removal efficiencies. All reactors showed high COD removal efficiencies (93–98%) and thus can be regarded as suitable for the treatment of low strength, complex wastewater. Staged anaerobic reactors can be a good alternative for this kind of application provided they are fitted with a large enough (≥7) number of compartments to fully take advantage of their strengths. Scale factors seem to have influenced importantly on the comparison between one and multi staged sludge-bed reactors and, therefore, observations made here could change at larger reactor volumes.     

Treatment of low strength soluble wastewaters in UASB reactors

Low strength wastewaters can be those with chemical oxygen demand (COD) below 2,000 mg/l. The anaerobic treatment of such wastewaters has not been fully explored so far. The suboptimal reaction rates with low substrate concentrations, and the presence of dissolved oxygen in the influent are regarded as possible constraints. In this study, the treatment of low strength soluble wastewaters containing ethanol or whey was studied in lab-scale upflow anaerobic sludged bed (UASB) reactors at 30°C. The high treatment performance obtained demonstrates that UASB reactors are viable for treating both types of wastewaters at low COD concentrations. The treatment of the ethanol containing wastewater resulted in COD removal efficiencies exceeding 95% at organic loading rates (OLR) between 0.3 to 6.8 g COD/l-d with influent concentrations in the range of 422 to 943 mg COD/l. In the case of the more complex whey containing wastewater, COD removal efficiencies exceeded 86% at OLRs up to 3.9 g COD/l·, as long as the COD influent was above 630 mg/l. Lowering the COD influent resulted in decreased efficiency with sharper decrease at values below 200 mg/l. Acidification instead of methanogenesis was found to be the rate limiting step in the COD removal at low concentrations, which was not the case when treating ethanol. The effect of dissolved oxygen in the influent as a potential danger in anaerobic treatment was investigated in reactors fed with and without dissolved oxygen. Compared with the control reactor, the reactor receiving oxygen showed no detrimental effects in the treatment performance. Thus, the presence of dissolved oxygen in dilute wastewaters is expected to be of minor importance in practice.     

Statistical modeling and optimization of biomass granulation and COD removal in UASB reactors treating low strength wastewaters

The aim of this work was to study the influence of influent chemical oxygen demand (COD), upflow velocity of wastewater, and cationic polymer additives in inoculum, on biomass granulation and COD removal efficiency in upflow anaerobic sludge blanket (UASB) reactor for treating low strength wastewater. Statistical models were formulated based on these three variables to optimize the biomass granulation and COD removal efficiency in UASB reactors using a two-level, full factorial design. For the thick inoculum used in this study, having suspended solids (SS) >80 g/l and volatile suspended solids (VSS) to SS ratio <0.3, cationic polymer additives in the inoculum showed adverse effect on biomass granulation and COD removal efficiency. It is concluded that for such thick inoculum, granulation can be obtained while treating low strength wastewaters in UASB reactor by selecting proper combination of influent COD and liquid upflow velocity so as to represent the organic loading rate (OLR) greater than 1.0 kg COD/m(3) d. Validation of model predictions for treatment of synthetic wastewater and actual sewage reveals the efficacy of these models for enhancing granulation and COD removal efficiency.     

Long-term effects of multi-walled carbon nanotubes on the performance and microbial community structures of an anaerobic granular sludge system

Multi-walled carbon nanotubes (MWCNTs) released into the sewage may cause negative and/or positive effects on the treatment system. The objective of this study was to explore over 110 days’ effect of MWCNTs on the performance of anaerobic granular sludge and microbial community structures in an upflow anaerobic sludge blanket (UASB) reactor. The results showed that MWCNTs had no significant effect on the removal of chemical oxidation demand (COD) and ammonia in UASB reactor, but the total phosphorus (TP) removal efficiency increased by 29.34%. The biogas production of the reactor did not change. The anaerobic granular sludge tended to excrete more EPS to resist the effects of MWCNTs during the long-term impact. Illumina MiSeq sequencing of 16S rRNA gene revealed that MWCNTs did not affect the microbial diversity, but altered the composition and structure of microbial community in the reactor. In this process, Saccharibacteria replaced Proteobacteria as the highest abundant bacterial phylum. MWCNTs promoted the differentiation of methanogen structure, resulting in increase of Methanomassiliicoccus, Methanoculleus, and the uncultured WCHA1–57. These results indicated that MWCNTs impacted the performance of UASB reactor and the structures of the microbial community in anaerobic granular sludge.

     

The effect of pretreatment on anaerobic activity of olive mill wastewater using batch and continuous systems

This study was focused on several physico-chemical and biological treatment methods that may affect the reduction of the organic load in olive mill wastewater (OMW). In this study, removal of 95% of the phenolic compounds present in OMW was achieved using sand filtration and subsequent treatment with powdered activated carbon in a batch system. This pretreatment for OMW was found to enhance the anaerobic activity of the sludge in the batch system significantly. The efficiency of organic load removal achieved by the anaerobic treatment of untreated OMW in batch reactors with tap water dilution factors below 1:10, reached approximately 65% chemical oxygen demand (COD) removal. However, in the up-flow sludge anaerobic blanket (UASB) reactor, COD removal efficiency of 80–85% was reached at a hydraulic retention time (HRT) of 5 days with an influent COD concentration of 40 g l⁻¹ and organic loading rate (OLR)=8 g⁻¹ COD l⁻¹ per day.     

Thermophilic (55-65 degrees C) and extreme thermophilic (70-80 degrees C) sulfate reduction in methanol and formate-fed UASB reactors

The feasibility of thermophilic (55-65 degrees C) and extreme thermophilic (70-80 degrees C) sulfate-reducing processes was investigated in three lab-scale upflow anaerobic sludge bed (UASB) reactors fed with either methanol or formate as the sole substrates and inoculated with mesophilic granular sludge previously not exposed to high temperatures. Full methanol and formate degradation at temperatures up to, respectively, 70 and 75 degrees C, were achieved when operating UASB reactors fed with sulfate rich (COD/SO4(2-)=0.5) synthetic wastewater. Methane-producing archaea (MPA) outcompeted sulfate-reducing bacteria (SRB) in the formate-fed UASB reactor at all temperatures tested (65-75 degrees C). In contrast, SRB outcompeted MPA in methanol-fed UASB reactors at temperatures equal to or exceeding 65 degrees C, whereas strong competition between SRB and MPA was observed in these reactors at 55 degrees C. A short-term (5 days) temperature increase from 55 to 65 degrees C was an effective strategy to suppress methanogenesis in methanol-fed sulfidogenic UASB reactors operated at 55 degrees C. Methanol was found to be a suitable electron donor for sulfate-reducing processes at a maximal temperature of 70 degrees C, with sulfide as the sole mineralization product of methanol degradation at that temperature.     

Effect of influent COD/SO4(2-) ratios on mesophilic anaerobic reactor biomass populations: physico-chemical and microbiological properties

The competitive and syntrophic interactions between different anaerobic bacterial trophic groups in sulphate limited expanded granular sludge bed (EGSB) reactors was investigated. The outcome of competition between the sulphate-reducing, methanogenic and syntrophic populations after development in reactors at varying influent COD/SO4 (2-) ratios was examined in batch activity tests with the inclusion of specific sulphate reducing bacteria (SRB) and methane producing archaea (MPA) inhibitors. SRB species could not out-compete MPA species for acetate at influent COD/SO4 (2-) ratios as low as 2. The SRB were seen to play a more significant role in the conversion of hydrogen but did not become completely dominant. HMPA were responsible for hydrogen utilization at an influent COD/SO4 (2-) ratio of 16, and were still dominant when the ratio was reduced to 4. It was only when the COD/SO4 (2-) ratio was reduced to 2 that the HSRB assumed a more influential role. SRB species were significant in the degradation of propionate at all COD/SO4 (2-) ratios applied. Sludge samples were analysed by scanning electron microscopy (SEM), granule size distribution and fluorescent in situ hybridization (FISH), combined with confocal laser scanning microscopy (CLSM), to monitor any changes in granule morphology under the various COD/SO4 (2-) ratios imposed during the reactor trial. In situ hybridization with domain- and species-specific oligonucleotide probes demonstrated a layered architecture with an outer layer harboring mainly Eubacterial cells and an inner layer dominated by Archaeal species.     

Long-term competition between sulfate reducing and methanogenic bacteria in UASB reactors treating volatile fatty acids

The competition between acetate utilizing methane-producing bacteria (MB) and sulfate-reducing bacteria (SRB) was studied in mesophilic (30 degrees C) upflow anaerobic sludge bed (UASB) reactors (upward velocity 1 m h-1; pH 8) treating volatile fatty acids and sulfate. The UASB reactors treated a VFA mixture (with an acetate:propionate:butyrate ratio of 5:3:2 on COD basis) or acetate as the sole substrate at different COD:sulfate ratios. The outcome of the competition was evaluated in terms of conversion rates and specific methanogenic and sulfidogenic activities. The COD:sulfate ratio was a key factor in the partitioning of acetate utilization between MB and SRB. In excess of sulfate (COD:sulfate ratio lower than 0.67), SRB became predominant over MB after prolonged reactor operation: 250 and 400 days were required to increase the amount of acetate used by SRB from 50 to 90% in the reactor treating, respectively, the VFA mixture or acetate as the sole substrate. The competition for acetate was further studied by dynamic simulations using a mathematical model based on the Monod kinetic parameters of acetate utilizing SRB and MB. The simulations confirmed the long term nature of the competition between these acetotrophs. A high reactor pH (+/-8), a short solid retention time (<150 days), and the presence of a substantial SRB population in the inoculum may considerably reduce the time required for acetate-utilising SRB to outcompete MB.     

Effect of starch addition on the biological conversion and microbial community in a methanol-fed UASB reactor during long-term continuous operation

The effect of starch addition on the microbial composition and the biological conversion was investigated using two upflow anaerobic sludge bracket (UASB) reactors treating methanolic wastewater: one reactor was operated with starch addition, and another reactor was operated without starch addition. Approximately 300 days of operation were performed at 30 kg COD/m³/d, and then, the organic load of the reactors was gradually increased to 120 kg COD/m³/d. Successful operation was achieved at 30 kg COD/m³/d in both reactors; however, the methanol-fed reactor did not perform well at 120 kg COD/m³/d while the methanol-starch-fed reactor did. The granule analysis revealed the granule developed further only in the methanol-starch-fed reactor. The results of the microbial community analysis revealed more Methanosaeta cells were present in the methanol-starch-fed reactor, suggesting the degradation of starch produced acetate as an intermediate, which stimulated the growth of Methanosaeta cells responsible for the extension of granules.     

Degradation of 4-chlorophenol in UASB reactor under methanogenic conditions

Treatment of simulated wastewater containing 40 mg/l of 4-chlorophenol (4-CP) was carried out in an upflow anaerobic sludge blanket (UASB) reactor under methanogenic condition. The performance of this test UASB reactor was evaluated in terms of 4-CP removal. Hydraulic retention time (HRT) and substrate:co-substrate ratio for the 4-CP removal was optimized by varying the influent flow rate (13-34.7 ml/min) and sodium acetate concentration (2-5 g/l), respectively. A control UASB reactor, which was not exposed to 4-CP was also operated under similar conditions. Organic loading rate (OLR) was varied in the range of 2-5.3 kg/m(3)/d and 1.7-4.2 kg/m(3)/d, respectively, for HRT and substrate:co-substrate ratio studies, respectively. The optimum HRT and substrate:co-substrate ratio for the removal of 4-CP was 12h and 1:75, respectively. Removal of 4-CP achieved at optimum HRT and substrate:co-substrate ratio was 88.3+/-0.7%. Removal of 4-CP occurred through dehalogenation and caused increase in chloride ion concentration in the effluent by 0.23-0.27 mg/mg 4-CP removed. The ring cleavage test showed the ortho mode of ring cleavage of 4-CP. Change in the elemental composition of the anaerobic biomass of UASB reactors was observed during the study period. Concentration of Ca(2+) increased in the biomass and this could be attributed to the biosoftening. Specific methanogenic activity of the sludge of control and test UASB reactor was 0.832 g CH(4) COD/g VSS d and 0.694 g CH(4) COD/g VSS d, respectively.     

Treatment of slaughterhouse wastewater in a UASB reactor and an anaerobic filter

A study was performed to assess the feasibility of anaerobic treatment of slaughterhouse wastewaters in a UASB (Upflow Anaerobic Sludge Blanket) reactor and in an AF (Anaerobic Filter). Among the different streams generated, the slaughter line showed the highest organic content with an average COD of 8000 mg/l, of which 70% was proteins. The suspended solids content represented between 15 and 30% of the COD. Both reactors had a working volume of 21. They were operated at 37°C. The UASB reactor was run at OLR (Organic Loading Rates) of 1–6.5 kg COD/m³/day. The COD removal was 90% for OLR up to 5 kg COD/m³/day and 60% for an OLR of 6.5 kg COD/m³/day. For similar organic loading rates, the AF showed lower removal efficiencies and lower percentages of methanization. At higher OLR sludge, flotation occurred and consequently the active biomass was washed out from the filter. The results indicated that anaerobic treatment systems are applicable to slaughterhouse wastewaters and that the UASB reactor shows a better performance, giving higher COD removal efficiencies than the AF.     

Effect of non-feeding period length on the intermittent operation of UASB reactors treating dairy effluents

Recent environmental concerns have prompted a re-evaluation of conventional management strategies and refueled the search of innovative waste management practices. In this sense, the anaerobic digestion of both fat and the remaining complex organic matter present in dairy wastewaters is attractive, although the continuous operation of high rate anaerobic processes treating this type of wastewaters causes the failure of the process. This work accesses the influence of non-feeding period length on the intermittent operation of mesophilic UASB reactors treating dairy wastewater, in order to allow the biological degradation to catch up with adsorption phenomenon. During the experiments, two UASB reactors were subject to three organic loading rates, ranging from 6 to 12 g(COD) x L(-1) x d(-1), with the same daily load applied to both reactors, each one with a different non-feeding period. Both reactors showed good COD removal efficiencies (87-92%). A material balance for COD in the reactors during the feeding and non-feeding periods showed the importance of the feedless period, which allowed the biomass to degrade substrate that was accumulated during the feeding period. The reactor with the longest non-feeding period had a better performance, which resulted in a higher methane production and adsorption capacity for the same organic load applied with a consequent less accumulation of substrate into the biomass. In addition, both reactors had a stable operation for the organic load of 12 g(COD) x L(-1) x d(-1), which is higher than the maximum applicable load reported in literature for continuous systems (3-6 g(COD) x L(-1) x d(-1)).     

Granule development and performance in sucrose fed anaerobic baffled reactors

Two 90 L anaerobic baffled reactors were used to study the granulation of sludge and the effect of the organic loading rate and NaHCO3/COD ratios on reactor performance. Furthermore, it was determined whether an anaerobic baffled reactor would promote phase separation and if additive of bentonite or granular active carbon was capable of enhancing granule formation. In order to minimize feed variations, and have a totally biodegradable substrate, a synthetic sucrose substrate was used. Granulation was achieved in both reactors within 75 days. However, the granules from the granular active carbon amended reactor appeared earlier and were larger and more compact. The reactors were maintained at a hydraulic retention time of 20 h during performance study stage. The results showed that when organic loading rate were changed from 2.15 to 6.29 kg COD m(-3)day(-1), chemical oxygen demand (COD) removal was not decreased (91-93%), but a slight increase in effluent COD was observed. It was found that the COD removals were generally good (87-92%) and had not obviously change with the decreasing NaHCO3/COD ratios. From the bacterial distribution and the concentration of volatile fatty acids in four compartments, it was concluded that a separation of phases occurred within the anaerobic baffled reactors.