Biofilm development in laboratory methanogenic fluidized bed reactors

Biofilm development on sand with different heterogeneous inocula was studied in laboratory-scale methanogenic fluidized bed reactors. Both the course of biofilm formation during reactor start-up and the bacterial composition of newly developed biofilms at steady-state were found to be similar irrespective of the type of inoculum applied. Biofilm formation proceeded according to a fixed pattern that could be subdivided in three consecutive phases, designated as the lag phase, biofilm production phase, and steady-state phase. Methanogenic activity and biomass content of the fluidized bed granules were found to be accurate parameters of the course of biofilm formation. More indirect parameters monitored did not give unambiguous results in all instances. The composition of the newly developed biomass as assessed on the basis of potential methanogenic activities on different substrates and of the concentration of specific methanogenic cofactors was consistent with electron microscopic observations.     

Inhibition of propionate degradation by acetate in methanogenic fluidized bed reactors

Summary The degradation of acetate, propionate and butyrate was monitored during start-up of five lab-scale methanogenic fluidized bed reactors on an artificially prepared waste water. The acetate concentration in the reactor content was found to influence the degradation of propionate but not of butyrate. In general, at acetate levels over 200 mg/l the degradation of propionate was below 60%, whereas the degradation was complete at acetate levels under 100 mg/l. The rationale of the inhibition of propionate degradation by acetate is discussed.     

Early stages in biofilm development in methanogenic fluidized-bed reactors

Summary Biofilm development in methanogenic fluidized-bed reactors with sand as the carrier was studied on a laboratory scale. The microorganisms present in consecutive layers of the biofilm of mature sludge granules were preliminarily characterized on the basis of their morphology, element composition and adhesion capacity and were compared to bacteria which take part in the initial colonization of sand. The early phase of biofilm development was monitored with reactors receiving waste-waters containing different mixtures of volatile fatty acids and inoculated with fluidized-bed reactor effluent for different lengths of time. The results obtained indicate that facultative anaerobic bacteria abundantly present in the outermost biofilm layers of mature sludge granules are probably the main primary colonizers of the sand. Methanothrix spp. or other methanogens were rarely observed among the primary colonizers. The course of biofilm formation was comparable under the various start-up conditions employed including variations in waste-water composition, inoculation and anaerobicity. However, omission of waste-water and thus of substrate resulted in rapid wash-out of the attached biomass. Offprint requests to: W. Heinen     

Variation of parameters affecting the start-up of methanogenic fluidized bed reactors

Summary The influence of the medium composition, the inoculum and the inoculation procedure on initial biofilm development in methanogenic fluidized bed reactors was studied on laboratory scale. Trace minerals but not vitamins were found to be essential for biofilm development. Inoculation with heterogeneous bacterial cultures of potentially sand-colonizing microorganisms and/or with pure cultures ofMethanothrix soehngenii did not accelerate biofilm development significantly as compared to inoculation with effluent from a fully operative fluidized bed reactor.     

Startup of anaerobic biofilm fluidized bed reactors on molasses and phenol under various conditions

Fluidized sand bed anaerobic biofilm reactors were operated in parallel to study the effects of inoculum, loading, residence time and carrier type on the startup dynamics for the degradation of molasses and phenol. Degradation rates generally depended most directly on concentrations rather than on other operating variables. Residence times did not appear to directly influence startup. Short residence times and high loadings gave the highest specific activities for both substrates. The type of inoculum was found to be most important for the molasses system, and inoculation on fresh carrier was found to be better than reinoculation. The two times higher specific biomass retention on Siran porous glass gave essentially the same degradation rates on a volume basis.List of Symbols L kg/h loading of reactor - M kg/kg biomass per carrier mass - Red. % reduction of feed concentration due to degradation - R kg/(m³ · h) reaction rate - S kg/m³ substrate concentration in reactor and effluent - S ₀ kg/m³ substrate concentration in feed - t h time     

Influence of hydrodynamic conditions on biofilm behavior in a methanogenic inverse turbulent bed reactor

This paper presents a study about the influence of gas velocity on a methanogenic biofilm in an inverse turbulent bed reactor. Experimental results indicate a dynamic response of the growing attached biomass to the changes of hydrodynamic conditions, mainly attrition constraints. Short but intensive increases of gas velocity (U(g)) are shown to induce more detachment than a high but constant gas flow rate. Hydrodynamic conditions control the composition of the growing biofilm in terms of cells and exocellular polymeric substances (EPS). The cell fraction within the biofilm (R(cell)) was found to be inversely proportional to the gas velocity. The specific activity expressed in methane production rate or COD removal rate is higher in biofilms formed under high hydrodynamic constraints. The control of the hydrodynamic conditions in a biofilm reactor should make it possible to obtain a resistant and active biofilm.     

Biodegradation of dichloromethane in waste water using a fluidized bed bioreactor

Summary Biological treatment of a synthetic waste water containing 120 mM dichloromethane (10.2g/l) was carried out under aerobic conditions using dichloromethane-degrading bacteria as an inoculum. The bacteria were adsorbed to support particles and grown in a fluidized bed bioreactor. Charcoal and sand particles were compared as support materials with regard to abrasion, the maximum degradation rate for dichloromethane and the stability of the biological activity in the system.The use of charcoal led to the incorporation of coal dust into the biomass and to an uncontrollable thickness of the biofilm. Therefore the system became unstable and the biological activity decreased. In contrast sand as support material was indifferent to abrasion and allowed biofilm thickness to be controlled. The dichloromethane degrading capacity of the bioreactor increased during the first 30 days. It reached a steady state level of 1.6 g CH₂Cl₂/lxh. Dichloromethane concentration in the effluent was <0.01 mM (<0.85 mg/l) and consequently the degradation efficiency better than 99.99%.     

Influence of environmental conditions on methanogenic compositions in anaerobic biogas reactors

The influence of environmental parameters on the diversity of methanogenic communities in 15 full-scale biogas plants operating under different conditions with either manure or sludge as feedstock was studied. Fluorescence in situ hybridization was used to identify dominant methanogenic members of the Archaea in the reactor samples; enriched and pure cultures were used to support the in situ identification. Dominance could be identified by a positive response by more than 90% of the total members of the Archaea to a specific group- or order-level probe. There was a clear dichotomy between the manure digesters and the sludge digesters. The manure digesters contained high levels of ammonia and of volatile fatty acids (VFA) and were dominated by members of the Methanosarcinaceae, while the sludge digesters contained low levels of ammonia and of VFA and were dominated by members of the Methanosaetaceae. The methanogenic diversity was greater in reactors operating under mesophilic temperatures. The impact of the original inoculum used for the reactor start-up was also investigated by assessment of the present population in the reactor. The inoculum population appeared to have no influence on the eventual population.     

Detachment of multi species biofilm in circulating fluidized bed bioreactor

In this study, the detachment rates of various microbial species from the aerobic and anoxic biofilms in a circulating fluidized bed bioreactor (CFBB) with two entirely separate aerobic and anoxic beds were investigated. Overall detachment rate coefficients for biomass, determined on the basis of volatile suspended solids (VSS), glucose and protein as well as for specific microbial groups, i.e., for nitrifiers, denitrifiers, and phosphorous accumulating organisms (PAOs), were established. Biomass detachment rates were found to increase with biomass attachment on carrier media in both beds. The detachment rate coefficients based on VSS were significantly affected by shear stress, whereas for protein, glucose and specific microbial groups, no significant effect of shear stress was observed. High detachment rates were observed for the more porous biofilm structure. The presence of nitrifiers in the anoxic biofilm and denitrifiers in the aerobic biofilm was established by the specific activity measurements. Detachment rates of PAOs in aerobic and anoxic biofilms were evaluated.     

Phenol degradation in a three-phase biofilm fluidized sand bed reactor

A previous three phase fluidized sand bed reactor design was improved by adding a draft tube to improve fluidization and submerged effluent tubes for sand separation. The changes had little influence on the oxygen transfer coefficients(K _L a), but greatly reduced the aeration rate required for sand suspension. The resulting 12.5 dm³ reactor was operated with 1 h liquid residence time, 10.2dm³/min aeration rate, and 1.7–2.3 kg sand (0.25–0.35 mm diameter) for the degradation of phenol as sole carbon source. The K _La of 0.015 s^–1 gave more than adequate oxygen transfer to support rates of 180g phenol/h · m³ and 216 g oxygen/h · m³. The biomass-sand ratios of 20–35 mg volatiles/g gave estimated biomass concentrations of 3–6 g volatiles/dm³. Offline kinetic measurements showed weak inhibition kinetics with constants ofK _s=0.2 mg phenol/dm³, K _o2=0.5 mg oxygen/dm³ and KinI= 122.5 mg phenol/dm³. Very small biofilm diffusion effects were observed. Dynamic experiments demonstrated rapid response of dissolved oxygen to phenol changes below the inhibition level. Experimentally simulated continuous stagewise operation required three stages, each with 1 h residence time, for complete degradation of 300 mg phenol/dm³ · h.     

Heterogeneous biofilm activity in a segregated anaerobic fluidized bed bioreactor

Bed segregation in a fluidized bed bioreactor profoundly influenced biofilm thickness and microbial activities of the biofilm along the bed height. Bioparticles coated with a thin biofilm, observed at the bottom of the reactor, had a higher specific activity in propylene glycol and n-propanol degradation than in thick biofilms developed at the top of the reactor. Although no significant difference was observed in specific activity for propionate and acetate along the reactor flow axis, more total propionate and acetate conversion occurred in regions of thicker biofilm accumulation.     

Evaluation of methanogenic kinetics in an anaerobic fluidized bed reactor (AFBR)

A kinetic study of the methanogenic phase was carried out on a pilot lab scale anaerobic fluidized bed reactor (AFBR) in batch mode. An examination of the effect of initial acetate concentration, bed expansion and bed segregation is presented.Experimental data observed for the acetate removal against time were adjusted to a zero-order kinetic equation, over the chemical oxygen demand (COD) range studied (1430–5340 mg litre⁻¹), independently of the bed expansion (11–37%). The kinetic constant was calculated using robust regression analysis. The zero-order kinetic constant, K₀ was between 1180–1380 mg COD litre⁻¹ h⁻¹ on the fixed bed volume basis, and the maximum specific substrate utilization rate, k, was between 145–198 mg COD g VS⁻¹ h⁻¹.The kinetic behaviour was found to be different throughout the reactor, on the fixed bed volume basis and the activity at the bottom of the bed was lower than the activity in the upper region. However, on an attached volatile solids basis, the activity at the bottom level was the greatest.     

Modeling of experiments on biofilm penetration effects in a fluidized bed nitrification reactor

A four-component, diffusion-reaction model with double Michaelis-Menten kinetics was used to describe the experimental data obtained from a laboratory biofilm, fluidized-bed nitrification reactor. Theory and experiment demonstrated that the stoichiometric ratio (3.5 mg O(2)/mg NH(4) (+)-N) can be employed as a criterion to determine whether the limiting substrate is oxygen or ammonia. For the present work, in the range of concentrations where limitation occurred, 4 mg/L NH(4) (+)-N and 14 mg/L O(2), the ratio of oxygen to ammonia in the bulk liquid determined which substrate was penetration-limiting-O(2) if <3.5 and NH(4) (+) if > 3.5. Halforder kinetics with respect to the limiting substrate described the apparent overall rates. Simulations provided biofilm concentration profiles which demonstrated the role of the oxygen-ammonia ratio. Experiments indicated that, generally, high NO(2) (-) concentrations can be expected. These depend on the residence time, biofilm area, and oxygen concentration. This dependency was investigated with the model, as was the parametric sensitivity with respect to the saturation constants. Particularly important for the NO(2) (-) levels were the ratios of the saturation constants for oxygen.     

Growth and productivity of Beta vulgaris cell culture in fluidized bed reactors

Beta vulgaris cells were cultured in specially constructed fluidized bed reactors on Murashige and Skoog medium with 3% sucrose as a carbon source. Biomass density levels of approximately 17–18 g dry cell weight liter^–1 for 5 liter fermenters and 9–10 g dry cell weight liter^–1 for 50 liter reactor were observed. Depending on particular cell line, cell content of betalains reached levels ranging from 12 to 25 mg g dry cell weight^–1, representing the productivities of 14 to 17 mg of pigments per liter and day.The authors would like to thank the Chemap Tetra Laval group and the Kommission zur Förderung der wissenschaftlichen Forschung for the financial support of this project (KWF No. 2057.1).     

Real evidence about zeolite as microorganisms immobilizer in anaerobic fluidized bed reactors

Using the scanning electronic microscopy, it was observed that natural zeolite possesses excellent physical characteristics as a support medium in anaerobic fluidized bed reactors (AFBR). Samples for biomass analysis were taken from two identical laboratory-scale AFBR (R-1 and R-2), which were operated with 25% of fluidization. These reactors treated distillery wastewaters (vinasses) at mesophilic temperature (30 ± 2 °C). The experiments were carried out with 0.25–0.50 and 0.50–0.80 mm zeolite particle diameter in reactors R-1 and R-2, respectively. The biomass concentration attached to zeolite in both reactors was found to be in the range of 40–45 g volatile solids/l. COD removal efficiencies as high as 90% were achieved at organic loading rate (OLRs) of up to 20 g COD/l day. The volatile fatty acid (VFA) levels were always lower that the suggested limits for digester failure. The yield coefficient of methane production was 0.29 l CH₄(at STP)/g COD consumed and was virtually independent of the OLR applied. A hybridization technique (fluorescence in situ hybridization, FISH) helped determine the predominant anaerobic microorganisms that colonized the natural zeolite, which were found to be Methanosaeta and Methanosarcinaceae, observing a reduced number of sulphate reducing bacteria. The results obtained for reactors R-1 and R-2 were very similar, showing that the particle size did not significantly influence the microbial community immobilized on zeolite.     

Microbial community structure and operational performance of a fluidized bed biofilm reactor treating oil sands process-affected water

This study evaluated the treatment of oil sands process-affected water (OSPW) using a fluidized bed biofilm reactor (FBBR) with granular activated carbon (GAC) as support media. The bioreactor was operated for 120 days at different organic and hydraulic loading rates. The combined GAC adsorption and biodegradation process removed 51% of chemical oxygen demand (COD), 56% of acid-extractable fraction (AEF) and 96% of classical naphthenic acids (NAs) under optimized operational conditions. Bioreactor treatment efficiencies were dependent on the organic loading rate (OLR), and to a lower degree, on the hydraulic loading rate (HLR). Further ultra performance liquid chromatography/high resolution mass spectroscopy (UPLC/HRMS) analysis showed that the removal of classical NAs increased as the carbon number increased. Compared with planktonic bacterial community in OSPW, more diverse microbial structures were found in biofilms colonized on the surface of GAC after 120-day treatment, with various carbon degraders namely Polaromonas jejuensis, Algoriphagus sp., Chelatococcus sp. and Methylobacterium fujisawaense in the GAC-biofilm reactor. The results of this study, therefore, showed that the GAC-biofilm seems to be a promising biological treatment method for OSPW remediation.     

Scanning electron microscopy of biofilm development in anaerobic fixed-bed reactors: Influence of the inoculum

Summary Scanning electron microscopy was applied to evaluate the influence of inoculum on efficiency of initial biofilm formation and reactor performance. Five anaerobic fixed-bed reactors were inoculated with anaerobic sludges from different sources and operated in parallel under identical conditions with defined wastewater and acetate, propionate and butyrate as constituents In all sludges Methanothrix sp. was the predominant acetotroph. The reactors inoculated with anaerobic sludge adapted to the wastewater achieved the highest space loading with 21.0 g COD/l·d after 58 days. The inoculation with granular sludge from an upflow anaerobic sludge blanket (UASB) reactor resulted in significantly less reactor efficiency. Time course of biofilm formation and biofilm thickness (ranging from 20–200 m) depended on the type of inoculum.     

Mathematical model for a three-phase fluidized bed biofilm reactor in wastewater treatment

A mathematical model for a three phase fluidized bed bioreactor (TFBBR) was proposed to describe oxygen utilization rate, biomass concentration and the removal efficiency of Chemical Oxygen Demand (COD) in wastewater treatment. The model consisted of the biofilm model to describe the oxygen uptake rate and the hydraulic model to describe flow characteristics to cause the oxygen distribution in the reactor. The biofilm model represented the oxygen uptake rate by individual bioparticle and the hydrodynamics of fluids presented an axial dispersion flow with back mixing in the liquid phase and a plug flow in the gas phase. The difference of settling velocity along the column height due to the distributions of size and number of bioparticle was considered. The proposed model was able to predict the biomass concentration and the dissolved oxygen concentration along the column height. The removal efficiency of COD was calculated based on the oxygen consumption amounts that were obtained from the dissolved oxygen concentration. The predicted oxygen concentration by the proposed model agreed reasonably well with experimental measurement in a TFBBR. The effects of various operating parameters on the oxygen concentration were simulated based on the proposed model. The media size and media density affected the performance of a TFBBR. The dissolved oxygen concentration was significantly affected by the superficial liquid velocity but the removal efficiency of COD was significantly affected by the superficial gas velocity. An erratum to this article can be found online at .