Bioreduction of para-chloronitrobenzene in a hydrogen-based hollow-fiber membrane biofilm reactor: effects of nitrate and sulfate

A continuous-stirred, hydrogen-based, hollow-fiber membrane biofilm reactor (HFMBfR) that was active in nitrate and sulfate reductions was shown to be effective for degradation or detoxification of para-chloronitrobenzene (p-CNB) in water by biotransforming it first to para-chloroaniline (nitro-reduction) and then to aniline (reductive dechlorination) with hydrogen (H₂) as an electron donor. A series of short-term experiments examined the effects of nitrate and sulfate on p-CNB bioreduction. The results obtained showed both higher nitrate and sulfate concentration declined the p-CNB bioreduction in the biofilm, and this suggests the competition for H₂ caused less H₂ available for the p-CNB bioreduction when the H₂ demand for the reductions was larger. Denitrification and sulfate reduction intermediates were thought to be potential factors inhibiting the p-CNB bioreduction. Analysis of electron-equivalent fluxes and reaction orders in the biofilm further demonstrated both denitrification and sulfate reduction competed more strongly for H₂ availability than p-CNB bioreduction. These findings have significant implications for the HFMBfR used for degrading p-CNB under denitrifying and/or sulfate reducing conditions.     

Performance of a membrane biofilm reactor for denitrification with methane

In this study, a membrane biofilm reactor was investigated for aerobic methane oxidation coupled indirectly to denitrification, a process potentially useful for denitrification of nitrate-contaminated waters and wastewaters using methane as external electron donor. Methane and oxygen were supplied from the interior of a silicone tube to a biofilm growing on its surface. We found that the membrane biofilm reactor was to some extent self-regulating in the supply of methane and oxygen. Although the intramembrane partial pressures of methane and oxygen were varied, the oxygen-to-methane ratio penetrating the membrane tended towards 1.68. Both nitrate removal rate and dissolved organic carbon (DOC) production rate appeared to be positively correlated with intramembrane methane pressure. Based on measured nitrate removal rates, DOC production rates, and nitrate removal efficiency, the possibility of using this method for treatment of a hypothetical wastewater was evaluated.     

Biodegradation of acetonitrile by adapted biofilm in a membrane-aerated biofilm reactor

A membrane-aerated biofilm reactor (MABR) was developed to degrade acetonitrile (ACN) in aqueous solutions. The reactor was seeded with an adapted activated sludge consortium as the inoculum and operated under step increases in ACN loading rate through increasing ACN concentrations in the influent. Initially, the MABR started at a moderate selection pressure, with a hydraulic retention time of 16 h, a recirculation rate of 8 cm/s and a starting ACN concentration of 250 mg/l to boost the growth of the biofilm mass on the membrane and to avoid its loss by hydraulic washout. The step increase in the influent ACN concentration was implemented once ACN concentration in the effluent showed almost complete removal in each stage. The specific ACN degradation rate achieved the highest at the loading rate of 101.1 mg ACN/g-VSS h (VSS, volatile suspended solids) and then declined with the further increases in the influent ACN concentration, attributed to the substrate inhibition effect. The adapted membrane-aerated biofilm was capable of completely removing ACN at the removal capacity of up to 21.1 g ACN/m² day, and generated negligible amount of suspended sludge in the effluent. Batch incubation experiments also demonstrated that the ACN-degrading biofilm can degrade other organonitriles, such as acrylonitrile and benzonitrile as well. Denaturing gradient gel electrophoresis studies showed that the ACN-degrading biofilms contained a stable microbial population with a low diversity of sequence of community 16S rRNA gene fragments. Specific oxygen utilization rates were found to increase with the increases in the biofilm thickness, suggesting that the biofilm formation process can enhance the metabolic degradation efficiency towards ACN in the MABR. The study contributes to a better understanding in microbial adaptation in a MABR for biodegradation of ACN. It also highlights the potential benefits in using MABRs for biodegradation of organonitrile contaminants in industrial wastewater.     

A dicyclic-type electrode-based biofilm reactor for simultaneous nitrate and Cr(VI) reduction

Bioprocess and Biosystems Engineering - A dicyclic-type electrode-based biofilm-electrode reactor (BER) was investigated for simultaneous removal of nitrate and Cr(VI). In the absence of Cr(VI),...     

Enhancement on the simultaneous removal of nitrate and organic pollutants from groundwater by a three-dimensional bio-electrochemical reactor

To improve denitrification performance and effective degradation of organic pollutants from micro-polluted groundwater simultaneously, a novel three-dimensional (3D) bio-electrochemical reactor was developed, which introduced activated carbon into a traditional two-dimensional (2D) reactor as the third electrode. The static and dynamic characteristics of the reactor were investigated with special attentions paid to the performance comparison of these two reactors. In the 3D reactor both TOC and nitrate removal efficiency were greatly improved, and the formation of nitrite byproduct is considerably reduced, comparing with that of the 2D reactor. The role of activated carbon biofilm was explored and possible remediation mechanisms for the 2D and 3D reactors were suggested. In such a 3D reactor, the denitrification rate improved greatly to 0.288 mg NO₃–N/cm²/d and the current efficiency could reach as high as 285%. Further, it demonstrated good performance stably against variable conditions, indicating very promising in application for groundwater remediation.     

Denitrifying and methanogenic bacteria in the biofilm of a fixed-film reactor operated with methanol/nitrate demonstrated by immunofluorescence and microscopy

 A denitrifying bacterial biofilm population established on a polypropylene substratum of a fixed-film reactor was characterized by microscopy, scanning electron microscopy and immunofluorescence after 120 days of operation. The reactor, operated at pH 7.0, 22°C, and −180 mV with synthetic wastewater containing methanol/nitrate, achieved a denitrification rate of 0.24 mol NO^- ₃ l^-1 day^-1 with a removal efficiency for nitrate of 95%–99% at an organic loading rate of 0.325 mol methanol l^-1 day^-1. The gas produced contained 2%–3% (v/v) methane and 3%–4% (v/v) carbon dioxide in addition to nitrogen. The biofilm contained mainly cells of Methanobrevibacter arboriphilus antigenically related to strain DC, short, flagellated, gram-negatively staining rods of Pseudomonas sp. antigenically related to Pseudomonas stutzeri strain AN11, non-identified pink-pigmented rods and small lemon-shaped cells with mono- and bipolar appendages resembling prosthecate Hyphomicrobium sp. The biofilm analysis provided evidence for a syntrophy between the denitrifying, methylotrophic, bacterial consortium and hydrogenotrophic methanogens, which were identified by antigenic fingerprinting with 17 antibody probes. Received: 11 July 1994/Received revision: 23 September 1994/Accepted: 28 September 1994     

Denitrification of high strength nitrate waste

The aim of the present work was to study the treatment of high strength nitrate waste (40000 ppm NO(3) i.e., 9032 ppm NO(3)-N) by acclimatizing sludge initially capable of degrading dilute streams (100-200 ppm NO(3)-N). Sludge from an effluent treatment plant of a fertilizer industry was acclimatized for 15 d each at 1694, 3388, 6774 and 9032 ppm NO(3)-N in a 4 L sequencing batch reactor. Complete denitrification of extremely concentrated nitrate waste (9032 ppm NO(3)-N) using acclimatized sludge was achieved in just 6 h. During the acclimatization period, increase in nitrite peak value from zero to 5907 ppm NO(2)-N was observed, as the concentration was increased from 1694 to 9032 ppm NO(3)-N. Kinetic analysis of the nitrate and nitrite profile could reasonably support microbiological explanations for nitrite build up and changes in sludge composition.     

Continuous pullulan fermentation in a biofilm reactor

Biofilm is a natural form of cell immobilization in which microorganisms attach onto solid support. In this study, a pigment-reduced pullulan-producing strain, Aureobasidium pullulans (ATCC 201253), was used for continuous pullulan fermentation in a plastic composite support (PCS) biofilm reactor. Optimal conditions for the continuous pullulan production were determined by evaluating the effects of the feeding medium with various concentrations of ammonium sulfate and sucrose and dilution rate. Pullulan concentration and production rate reached maximum (8.3 g/l and 1.33 g/l/h) when 15 g/l of sucrose, 0.9 g/l of ammonium sulfate, and 0.4 g/l of yeast extract were applied in the medium, and the dilution rate was at 0.16 h⁻¹. The purity of produced pullulan was 93.0%. The ratio of hyphal cells of A. pullulans increased when it was grown on the PCS shaft. Overall, the increased pullulan productivity can be achieved through biomass retention by using PCS biofilm reactor.     

Denitrification in lucerne nodules and bacteroids supplied with nitrate

Nodulated lucerne plants ( Medicago sativa L. cv. Aragón) were supplied with 20 m M nitrate. Anaerobically isolated bacteroids of Rhizobium meliloti from these plants were able to denitrify after 48 h treatment. R. meliloti bacteroids behave as total denitrifiers, reducing nitrate to dinitrogen: when acetylene was omitted from the assay medium very little nitrous oxide was recovered. The onset of denitrification activity was coincident with the induction of nitrite reductase activity (EC 1.7.99.3) whereas nitrate reductase activity (EC 1.7.99.4) was constitutive. Whole nodules from plants receiving several doses of nitrate were assayed, in a nitrate-free medium, to monitor denitrification activity dependent on nitrate within the nodules. Denitrification activity was detected after 2 days of 20 m M nitrate supply or after 3 days in the presence of 10 or 5 m M nitrate. These results are discussed in relation to current controversy about nitrate entry into the infection region of nodules. It is concluded that this process occurs more rapidly than suggested in recent research.     

Denitrification of high nitrate wastewater in a cloth strip bioreactor with immobilized sludge

Denitrification of synthetic high nitrate wastewater containing 40,000?ppm NO₃ (9,032?ppm NO₃-N) was achieved using immobilized activated sludge in a column reactor. Active anoxic sludge adsorbed onto Terry cloth was used in the denitrification of high nitrate wastewater. The operational stability of the immobilized sludge system was studied both in a batch reactor and in a continuous reactor. The immobilized sludge showed complete degradation of different concentrations of NO₃-N (1,129, 1,693, 3,387, 6,774, and 9,032?ppm) in a batch process. The reactors were successfully run for 90?days without any loss in activity. The immobilized cell process has yielded promising results in attaining high denitrifying efficiency.     

Denitrification capacity in response to increasing nitrate loads and decreasing organic carbon contents in injected leachate of a simulated landfill reactor

Ex situ nitrification followed by denitrification inside the landfill has been recommended to remove ammonia from leachate. The effects of increasing nitrate load and decreasing organic carbon content in the injected leachate on the denitrifying capacity of municipal solid waste (MSW) were investigated. Results showed that MSW possesses a high denitrification capacity. Nitrate reduction could be initiated within 48 h after the first addition of nitrate. Nitrate reduction rate increased with the increasing nitrate loading concentration. When the nitrate loading concentration was increased to 850 mg L^?1, nitrate reduction rate reached up to 35 mg L^?1 h^?1. Nitrite accumulation could be found after the addition of nitrate in each test. However, the maximum nitrite accumulation efficiency declined with increased nitrate load. Organic carbon played an important role in the reduction of nitrate, and both endogenous and exogenous organic materials could act as electron donors.     

Autotrophic denitrification via a novel membrane-attached biofilm reactor

AIMS: A laboratory-scale autotrophic membrane-attached biofilm reactor was developed to remove nitrate from drinking water. METHODS AND RESULTS: Hydrogen and carbon dioxide flowed together into the lumem side of a gas-permeable silicone tube. The gases diffused through the membrane wall to feed Alcaligenes eutrophus that formed a biofilm on the surface of the silicone tube for autotrophic denitrification. Hydrogen provided the energy source, and carbon dioxide, besides serving as the carbon source, was employed to neutralize the alkalinity from denitrification. The optimal carbon dioxide concentration in the silicone tube was between 20% and 50%. CONCLUSION: This study has demonstrated that a gas-permeable silicone tube is a convenient and efficient method to feed A. eutrophus for autotrophic denitrification. Supplying a suitable amount of carbon dioxide together with hydrogen into the silicone tube solved the problem that alkalinity formation caused during denitrification. The pH of the bioreactor was maintained at about 7 to avoid nitrite accumulation, and then the nitrogen removal rate was increased. A high specific nitrogen removal rate (1.6-5.4 g Nm(2)d(-1-1) of surface area of silicone tube) was achieved. SIGNIFICANCE AND IMPACT OF THE STUDY: In addition to combining the advantages of the hydrogenotrophic denitrification process and a membrane feeding substrate bioreactor (MFSB), this bioreactor achieved a high nitrogen removal rate and is simple to operate. It therefore is highly promising in drinking-water treatment.     

Characteristics of a methanotrophic culture in a membrane-aerated biofilm reactor

The membrane-aerated biofilm reactor (MABR) shows considerable potential as a bioprocess that can exploit methanotrophic biodegradation and offers several advantages over both conventional biofilm reactors and suspended-cell processes. This work seeks primarily to investigate the oxidation efficiency in a methanotrophic MABR. A mixed methanotrophic biofilm was immobilized on an oxygen-permeable silicone membrane in a single tube hollow fiber configuration. Under the conditions used the maximum oxygen uptake rate reached values of 16 g/m2.d, and the rate of biofilm growth achieved was 300 microm/d. Both indicators reflect a very high metabolic rate. It was shown that the biofilm was predominantly in a dual-substrate limitation regime but below about 250 microm was fully penetrated by both substrates. Oxygen limitation was not observed. Analysis indicated that microbial activity stratification was evident and the location of stratified layers of oxygen-consuming components of the consortium could be manipulated via the intramembrane oxygen pressure. The results confirm that an MABR can be employed to minimize substrate diffusion limitations in thick biofilms.     

Denitrification by thermophilic soil bacteria with ethanol as substrate in a USB reactor

Summary Thermophilic biological denitrification was studied in a laboratory-upflow sludge blanket reactor fed with ethanol as carbon and energy source. High denitrification efficiency (>98%) was obtained at an ethanol: nitrate ratio >2 and at a hydraulic retention time (HRT) of 5 hours. The performance of the system with respect to nitrate removal was very satisfactory (>95%), even at high nitrate (235 mg NO₃-N/L) and hydraulic (3 hours HRT) loading rates applied.A stable sludge was formed by spherical granules 1 to 3 mm in diameter with a content of 25,8 g VS/L and were almost exclusively composed of bacteria belonging to the genus Bacillus.     

Continuous biocatalytic synthesis of epoxypropane using a biofilm reactor

Mixed culture methanotrophic attached biofilms immobilized on diatomite particles in a three-phase fluidized bed reaction system were developed. Methane monooxygenase (MMO) activity on diatomite particles increased as soon as the lag phase ended. More than 90% of the MMO activity in the fluidized bed was attached. A biofilm concentration of 3.3c3.7mg dry weight cell (dwc) per g dry solid (DS) was observed. Batch experiments were performed to explore the possibility of producing epoxypropane by a propene–methane co-oxidation process. The effect of methane on the epoxidation of propene and the effect of propene on the growth of methanotroph was also studied. In continuous experiments, optimum mixed gas containing 35 methane, 20 propene and 45% oxygen were continuously circulated through the fluidized bed reactor to deliver substrates and extract product. Initial epoxypropane productivity was 110–150 μmol/day. The bioreactor operated continuously for 53 days without obvious loss of epoxypropane productivity.     

Denitrification of nitrate and nitric acid with methanol as carbon source

Summary A methanol/nitrate-medium and anaerobic conditions yielded an enrichment culture which consisted ofHyphomicrobium andParacoccus. This mixed culture proved to be very effective in denitrification of solutions containing high concentrations of nitrate and free nitric acid when grown in a chemostat (D=0.04 h^-1). With 0.1 mol/l nitric acid solution as feed medium the pH in the culture vessel adjusted itself to 5.8. For the reduction of 1 g NO₃–N 2.6 g methanol were consumed and 0.56 g cells were produced.     

Bifurcational and dynamical analysis of a continuous biofilm reactor

A dynamical model of a continuous biofilm reactor is presented. The reactor consists of a three-phase internal loop airlift operated continuously with respect to the liquid and gaseous phases, and batchwise with respect to the immobilized cells. The model has been applied to the conversion of phenol by means of immobilized cells of Pseudomonas sp. OX1 whose metabolic activity was previously characterized (Viggiani, A., Olivieri, G., Siani, L., Di Donato, A., Marzocchella, A., Salatino, P., Barbieri, P., Galli, E., 2006. An airlift biofilm reactor for the biodegradation of phenol by Pseudomonas stutzeri OX1. Journal of Biotechnology 123, 464-477). The model embodies the key processes relevant to the reactor performance, with a particular emphasis on the role of biofilm detachment promoted by the fluidized state. Results indicate that a finite loading of free cells establishes even under operating conditions that would promote wash out of the suspended biophase. The co-operative/competitive effects of free cells and immobilized biofilm result in rich bifurcational patterns of the steady state solutions of the governing equations, which have been investigated in the phase plane of the process parameters. Direct simulation under selected operating conditions confirms the importance of the dynamical equilibrium establishing between the immobilized and the suspended biophase and highlights the effect of the initial value of the biofilm loading on the dynamical pattern.     

Denitrification in a seashore sandy deposit influenced by groundwater discharge

The chemical compositions of ground water and organic matter in sediments were investigated at a sandy shore of Tokyo Bay, Japan to determine the fate of ground water NO₃ ^–. On the basis of Cl^– distribution in ground water, the beach was classified into freshwater (FR)-, transition (TR)-, and seawater (SW)-zones from the land toward the shoreline. The NO₃ ^– and N₂O did not behave conservatively with respect to Cl^– during subsurface mixing of freshwater and seawater, suggesting NO₃ ^– consumption and N₂O production in the TR-zone. Absence of beach vegetation indicated that NO₃ ^– assimilation by higher plants was not as important as NO₃ ^– sink. Low NH₄ ⁺ concentrations in ground water revealed little reduction of NO₃ ^– to NH₄ ⁺. These facts implied that microbial denitrification and assimilation were the likely sinks for ground water NO₃ ^–. The potential activity and number of denitrifiers in water-saturated sediment were highest in the low-chlorinity part of the TR-zone. The location of the highest potential denitrification activity (DN-zone) overlapped with that of the highest NO₃ ^– concentration. The C/N ratio and carbon isotope ratio (¹³C) of organic matter in sediment (< 100 -m) varied from 12.0 to 22.5 and from –22.5 to –25.5, respectively. The ¹³C value was inversely related to the C/N ratio (r ² = 0.968, n = 11), which was explained by the mixing of organic matters of terrestrial and marine origins. In the DN-zone, the fine sediments were rich in organic matters with high C/N ratios and low ¹³C values, implying that dissolved organic matters of terrestrial origin might have been immobilized under slightly saline conditions. A concurrent supply of NO₃ ^– and organic matter to the TR-zone by ground water discharge probably generates favorable conditions for denitrifiers. Ground water NO₃ ^– discharged to the beach is thus partially denitrified and fixed as microbial biomass before it enters the sea. Further studies are necessary to determine the relative contribution of these processes for NO₃ ^– removal.