Reduction of biomass in a bioscrubber for waste gas treatment by limited supply of phosphate and potassium ions

  Elimination of n-butanol from the gas phase was examined with a mixed culture in a compact bioscrubber. The extent of the cell concentration was limited by the supply of n-butanol, phosphate or potassium, and the growth rate was determined by the dilution rate. With n-butanol as the limiting substrate the cellular yield was 0.53 g dry cell weight/g n-butanol. Phosphate limitation decreased this yield to 0.34 g and potassium limitation to 0.31 g dry cell weight/g n-butanol at a dilution rate of 0.1/h. Under these conditions n-butanol was eliminated from the gas phase by 84%–100%. In the same order of limitations the specific degradation rate ranged from 0.19 g to 0.32 g n-butanol g dry cell weight⁻¹ h⁻¹. The fraction of n-butanol required to satisfy the needs for maintenance energy increased significantly depending on the limiting nutrient. Limitation by n-butanol, phosphate or potassium caused a maintenance requirement of 0.07, 0.16 and 0.34 g n-butanol g dry cell weight⁻¹ h⁻¹, thus showing a fivefold increase. This high demand for the carbon source demonstrated the feasibility of operating a bioscrubber under mineral limitation to reduce biomass formation significantly, and to maintain a high degree of substrate elimination from the gas phase. Received: 22 May 1996 / Received revision: 23 July 1996 / Accepted: 5 August 1996     

Effects of nitrogen limitation on biofilm formation in a hydrocarbon-degrading trickle-bed filter

The effect of nitrogen limitation on young and mature steady-state biofilm in a trickle-bed filter was studied. Toluene and n-heptane were the sole carbon source. Biomass concentration, respiration, substrate-induced respiration, metabolic quotient, and total hydrocarbon degradation efficiency were measured. The aim of the experiment was to control excess biomass production in the trickle-bed filter by limiting the mineral nutrients and to achieve increased mineralization of the carbon source. Biofilm growth responded strongly to the amount of available nitrogen, whereas hydrocarbon degradation efficiency reached a maximum of 60% and could not be increased even by further addition of nitrogen. The experiments showed that 95% of the adsorbed carbon was mineralized completely and only 5% was used for biofilm formation. This complete mineralization can also be concluded from the metabolic quotient. The value of the latter was about 6–10 mg CO₂-C g⁻¹C_mic h⁻¹, indicating an expanded energy demand due to stress effects in the presence of nutrient deficiency. It was postulated that determination of the metabolic quotient could be an simple instrument to measure the rate of mineralization of carbon sources and also the rate of biomass formation in trickle-bed filters or biofilters. Received: 11 November 1998 / Accepted: 5 December 1998     

Removal of organic air pollutants from exhaust gases in the trickle-bed bioreactor. Effect of oxygen

 The paper describes some reaction engineering fundamentals of the separation of organic air pollutants (volatile organic compounds) from waste gases using fixed-bacteria monocultures (biocatalysts) in a trickle-bed reactor. In particular the influence of pollutant concentration and oxygen concentration are investigated. The separation efficiency of certain substances such as acetone and isopropanol depends strongly on the oxygen concentration. The results obtained can be described by a mathematical model based on the diffusion of oxygen into the biofilm (diffusion regime of the catalyst). The non-stationary operation of the reactor – interruption of the oxygen stream and strong fluctuation in the exhaust gas stream – showed that other components such as propionaldehyde and n-propanol could be eliminated for a certain time without oxygen. Propionic acid is formed. Received: 9 June 1995/Received revision: 28 September 1995/Accepted: 4 October 1995     

Controlled biomass formation and kinetics of toluene degradation in a bioscrubber and in a reactor with a periodically moved trickle-bed

The kinetics of degradation of toluene from a model waste gas and of biomass formation were examined in a bioscrubber operated under different nutrient limitations with a mixed culture. The applicability of the kinetics of continuous cultivation of the mixed culture was examined for a special trickle-bed reactor with a periodically moved filter bed. The efficiency of toluene elimination of the bioscrubber was 50 to 57% and depended on the toluene mass transfer as evident from a constant productivity of 0.026 g dry cell weight/L . h over the dilution rate. Under potassium limitation the biomass productivity was reduced by 60% to 0.011 g dry cell weight/L . h at a dilution rate of 0.013/h. Conversely, at low dilution rates the specific toluene degradation rates increased. Excess biomass in a trickle-bed reactor causes reduction of interfacial area and mass transfer, and increase in pressure drop. To avoid these disadvantages, the trickle-bed was moved periodically and biomass was removed with outflowing medium. The concentration of steady state biomass fixed on polyamide beads decreased hyperbolically with the dilution rate. Also, the efficiency of toluene degradation decreased from 72 to 56% with increasing dilution rate while the productivity increased. Potassium limitation generally caused a reduction in biomass, productivity, and yield while the specific degradation increased with dilution rate. This allowed the application of the principles of the chemostat to the trickle-bed reactor described here, for toluene degradation from waste gases. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 686-692, 1997.     

Continuous biological waste gas treatment in stirred trickle-bed reactor with discontinuous removal of biomass

A new reactor for biological waste gas treatment was developed to eliminate continuous solvents from waste gases. A trickle-bed reactor was chosen with discontinuous movement of the packed bed and intermittent percolation. The reactor was operated with toluene as the solvent and an optimum average biomass concentration of between 5 and 30 kg dry cell weight per cubic meter packed bed (m3pb). This biomass concentration resulted in a high volumetric degradation rate. Reduction of surplus biomass by stirring and trickling caused a prolonged service life and prevented clogging of the trickle bed and a pressure drop increase. The pressure drop after biomass reduction was almost identical to the theoretical pressure drop as calculated for the irregular packed bed without biomass. The reduction in biomass and intermittent percolation of mineral medium resulted in high volumetric degradation rates of about 100 g of toluene m-3pb h-1 at a load of 150 g of toluene m-3pb h-1. Such a removal rate with a trickle-bed reactor was not reported before.     

On-line enzyme activity determination using the stopped-flow technique: application to laccase activity in pulp mill waste-water treatment

An automated system for on-line measurement of enzyme activity is proposed. The system uses a flow injection manifold in the stopped-flow mode to measure initial reaction rates. The time during which the flow is halted is selected in such a way as to optimise the enzyme/substrate ratio for the correct determination of activity values. The proposed system was used to determine the activity of laccase produced by the fungus Trametes versicolor immobilised on nylon in a fixed-bed reactor used for treating pulp mill waste water. Received: 17 February 1997 / Received revision: 23 April 1997 / Accepted: 27 April 1997     

Physiological aspects of continuous lactic acid fermentations at high dilution rates

The effects of the substrate conditions on the volumetric productivity of Lactobacillus helveticus at different cell densities up to 60 g l⁻¹ in a continuous stirred-tank reactor with microfiltration to retain the biomass were investigated. At low dilution rates, D, the steady-state volumetric productivity, r _p, gradually increased to a maximum at D = 1.2–1.5 h⁻¹, because of reduced product inhibition. At higher D values, r _p unexpectedly decreased, although the substrate conditions further improved. The maxima of r _p at different cell densities coincided with a critical specific substrate utilization rate beyond which the cell metabolism seems to be controlled through a catabolic modulator factor, and r _p decreases. Received: 8 September 1997 / Received last revision: 31 December 1997 / Accepted: 2 January 1998     

Cloning, expression in Streptomyces lividans and biochemical characterization of a thermostable endo-β-1,4-xylanase of Thermomonospora alba UL JB1 with cellulose-binding ability

Several thermophilic actinomycetes were isolated from urban solid waste. One of them, Thermomonospora alba ULJB1, showed a broad degradative activity on xylan, cellulose, starch and other polymers. Xylanase and cellulase activities were quantified and compared with those of Thermomonospora fusca. Genes encoding two different endo-β-1,4-xylanases were cloned from T.␣alba ULJB1. One of them, xylA, was sequenced, subcloned and overexpressed in Streptomyces lividans. It encodes a protein of 482 amino acids with a deduced molecular mass of 48 456 Da. The protein contains a 38-amino-acid leader peptide with six Arg⁺ residues in its amino-terminal end, a catalytic domain and a cellulose-binding domain connected by a linker region rich in proline and glycine. The XylA protein was purified to near homogeneity from S. lividans/xylA cultures. Two forms of the extracellular xylanase, of 48 kDa and 38 kDa, were produced that differed in their cellulose-binding ability. The 48-kDa protein showed a strong binding to cellulose whereas the 38-kDa form did not bind to this polymer, apparently because of the removal during processing of the cellulose-binding domain. Both forms were able to degrade xylans form different origins but not lichenam or carboxymethylcellulose. The major degradation product was xylobiose with traces of xylose. The xylanase activity was thermostable, showing a good activity up to 95 °C, and had broad pH stability in the range from pH 4.0 to pH 10.0. Received: 9 January 1997 / Received revision: 27 March 1997 / Accepted: 13 April 1997     

Hollow-fibre bioreactors compared to batch and chemostat culture for the production of a recombinant toxoid by a marine Vibrio

Bioreactor selection is important for maximising the productivity of recombinant organisms. In this paper a comparison is made between growth and recombinant protein synthesis in three types of bioreactor containing a marine Vibrio capable of heterologous expression and secretion of the non-toxic B-subunit pentamer of Escherichia coli heat-labile enterotoxin, EtxB. The heterologous gene was located on the plasmid pMMB68. Resistance to carbenicillin was used to select for plasmid-containing cells. In batch and continuous culture, volumetric productivities were highest when cells were grown in the presence of carbenicillin. Without antibiotic selection, the highest volumetric productivity (9.4 mg EtxB⁻¹ h⁻¹) was observed in hollow-fibre bioreactors, and the production phase could be maintained for over 50 h. The highest specific productivity under these conditions was found in batch culture, but the maximal production phase was only of 5 h duration. In hollow-fibre reactors the type of fibre used significantly affected productivity, both with regards to the maintenance of reactor integrity and by allowing passage of the recombinant toxoid through the selectively permeable membrane. Where contamination of the product with carbenicillin is to be avoided, these bioreactors are superior to batch or continuous culture. Received: 29 January 1997 / Received revision: 9 April 1997 / Accepted: 13 April 1997     

Mesophilic and thermophilic anaerobic digestion of source-sorted organic wastes: effect of ammonia on glucose degradation and methane production

The wet organic fraction of household wastes was digested anaerobically at 37 °C and 55 °C. At both temperatures the volatile solids loading was increased from 1 g l⁻¹ day⁻¹ to 9.65 g l⁻¹ day⁻¹, by reducing the nominal hydraulic retention time from 93 days to 19 days. The volatile solids removal in the reactors at both temperatures for the same loading rates was in a similar range and was still 65% at 19 days hydraulic retention time. Although more biogas was produced in the thermophilic reactor, the energy conservation in methane was slightly lower, because of a lower methane content, compared to the biogas of the mesophilic reactor. The slightly lower amount of energy conserved in the methane of the thermophilic digester was presumably balanced by the hydrogen that escaped into the gas phase and thus was no longer available for methanogenesis. In the thermophilic process, 1.4 g/l ammonia was released, whereas in the mesophilic process only 1 g/l ammonia was generated, presumably from protein degradation. Inhibition studies of methane production and glucose fermentation revealed a K _i (50%) of 3 g/l and 3.7 g/l ammonia (equivalent to 0.22 g/l and 0.28 g/l free NH₃) at 37 °C and a K _i (50%) of 3.5 g/l and 3.4 g/l ammonia (equivalent to 0.69 g/l and 0.68 g/l free NH₃) at 55 °C. This indicated that the thermophilic flora tolerated at least twice as much of free NH₃ than the mesophilic flora and, furthermore, that the thermophilic flora was able to degrade more protein. The apparent ammonia concentrations in the mesophilic and in the thermophilic biowaste reactor were low enough not to inhibit glucose fermentation and methane production of either process significantly, but may have been high enough to inhibit protein degradation. The data indicated either that the mesophilic and thermophilic protein degraders revealed a different sensitivity towards free ammonia or that the mesophilic population contained less versatile protein degraders, leaving more protein undegraded. Received: 26 March 1997 / Received revision: 13 May 1997 / Accepted: 19 May 1997     

Effects of support material on the pattern of volatile fatty acid accumulation at overload in anaerobic digestion of semi-solid waste

Anaerobic degradation of a semi-solid waste with a total solids content of 4% particulate matter, much of it insoluble, was investigated in four laboratory-scale reactors. Two of the reactors were equipped with different textile materials for immobilisation of microorganisms, while the other two were used as continuously-stirred-tank reactor references. A constant organic loading rate and hydraulic retention time were used in the start-up period; the hydraulic retention time was then decreased and the effects of this change were monitored. Volatile fatty acid (VFA) concentration and pH were chosen as indicators of the microbial status in the reactors. The reactors with support material showed a greater resistance to overload than did the continuously-stirred-tank reactors. This is in agreement with many studies undertaken on the anaerobic treatment of wastewater. However, no problems with clogging occurred, showing that a support material is also applicable in systems treating waste containing large amounts of insoluble, particulate matter. The pH was comparable to VFA for indicating an approaching process failure. However, the pattern of VFA accumulation was qualitatively different between the reactors with and without support material. Obviously the metabolic pattern of mixed cultures changes when the microorganisms are immobilised. Received: 3 December 1996 / Received revision: 7 February 1997 / Accepted: 14 February 1997     

Hydrogen production with high yield and high evolution rate by self-flocculated cells of Enterobacter aerogenes in a packed-bed reactor

Continuous hydrogen gas evolution by self-flocculated cells of Enterobacter aerogenes, a natural isolate HU-101 and its mutant AY-2, was performed in a packed-bed reactor under glucose-limiting conditions in a minimal medium. The flocs that formed during the continuous culture were retained even when the dilution rate was increased to 0.9 h⁻¹. The H₂ production rate increased linearly with increases in the dilution rate up to 0.67 h⁻¹, giving maximum H₂ production rates of 31 and 58 mmol l⁻¹ h⁻¹ in HU-101 and AY-2 respectively, at a dilution rate of more than 0.67 h⁻¹. The molar H₂ yield from glucose in AY-2 was maintained at about 1.1 at dilution rates between 0.08 h⁻¹ and 0.67 h⁻¹, but it decreased rapidly at dilution rates more than 0.8 h⁻¹. Received: 27 August 1997 / Received revision: 11 November 1997 / Accepted: 14 December 1997     

Non-toluene-associated respiration in a Pseudomonas putida 54G biofilm grown on toluene in a flat-plate vapor-phase bioreactor

Non-toluene-associated respiration (NTAR) within a Pseudomonas putida 54G biofilm growing on toluene as sole external carbon source was evaluated using oxygen microelectrodes in a flat-plate vapor-phase biological reactor. Two fluorescent probes, 2,4-diamidino-2-phenylindole and 5-cyano-2,3-ditolyltetrazolium chloride, were used to evaluate the number of total and respiring cells respectively within the biofilm. Biofilm samples were also analyzed for viable and toluene-culturable cells by spread-plating on non-selective and selective media respectively. Fractions of viable stressed, respiring and non-respiring cells within the biofilm were evaluated. The NTAR rate was positively correlated with the fraction of viable stressed and non-respiring cells within the biofilm, which suggested the capability of some cells to grow at the expense of leakage and lysis products coming from injured and dead cells. This effect was more pronounced at higher toluene concentration. Results suggest that NTAR should be incorporated into mathematical models of biofilm reactors degrading volatile organic carbon compounds. Received: 4 January 1997 / Received revision: 20 March 1997 / Accepted: 27 March 1997     

Microbial cleaning of waste gas containing volatile organic compounds in a bioreactor system with a closed gas circuit

The cleaning of the exhaust gases of a bioreactor containing volatile hydrocarbons in a bioreactor system with a closed gas circuit is described. The bioreactor system consisted of three different reactor types: a stirred tank which was filled with hydrocarbon-containing waste water to simulate the exhaust gases of a remediation process; a trickle-bed reactor for aerobic treatment of the exhaust gas from the stirred tank; and a photoreactor containing an algae culture which assimilated CO₂ from the trickle-bed reactor and also produced O₂. With this bioreactor system, it was possible to efficiently remove volatile organic compounds (VOC) from the waste gases. Depending on the type of waste water investigated, elimination rates of 41% to 93% of BTEX (benzene, ethylbenzene, toluene, xylene) and 29% to 53% of VCH (volatile chlorinated hydrocarbons) were obtained. Due to the photosynthesis of the algae in the system's photoreactor, oxygen concentrations between 12% and 18% [v/v], equivalent to about 57% to 83% DOT, were obtained. This concentration permitted the aerobic degradation to be carried out without having to add fresh air. The trickle-bed reactor and the photoreactor worked continuously, whereas the waste water in the stirred bioreactor was replaced in different batches. The accumulation of toxic compounds in the nutrient solutions of the trickle-bed (EC-50 > 30 g/l) and of the photoreactor (EC-50 > 35 g/l) was low. Carbon dioxide concentrations in the gas flow were higher than in fresh air (1% to 3% [vol/vol]), but no long-term accumulation of CO₂ occurred. This means that the algae in the photoreactor were active enough to assimilate the CO₂ which had been produced. They were also able to produce sufficient oxygen for aerobic hydrocarbon degradation. The system described is a first step towards treating waste gases which results from the bioremediation of hydrocarbon-contaminated media in a closed gas circuit without any emission (e.g. VOC, CO₂, germs).     

Effects of directly soluble and fibrous rapidly acidifying chemical oxygen demand and reactor liquid surface tension on granulation and sludge-bed stability in upflow anaerobic sludge-blanket reactors

 In recent years, it has become clear that the rapidly acidifying chemical oxygen demand (RACOD) content of the waste water and the surface tension of the reactor liquid contribute to the phenomenon of granular growth in upflow anaerobic sludge-blanket reactors (UASB). By adding 20% of directly soluble RACOD, in the form of a sucrose/starch mixture, on top of the original COD load and by adjusting the reactor liquid surface tension below 50 mN m⁻¹ with linear alkylbenzenesulphonate, granular growth and sludge-bed stability could be enhanced significantly within 40 days. Carrot pulp, a waste product having a high short-chain fatty acid precursor potential, was applied as an alternative fibrous RACOD source. Best results were obtained when adding the carrot pulp freshly to the laboratory-scale UASB reactor in an in-recycle liquefying chamber. This concept of adding carrot pulp waste product as a granular growth supplement by means of an in-recycle liquefying chamber therefore merits testing in practice. Received: 30 October 1996 / Received version: 3 February 1997 / Accepted: 10 February 1997     

Aerobic 4-nitrophenol degradation by microorganisms fixed in a continuously working aerated solid-bed reactor

Studies of microbial purification of a model waste water containing 4-nitrophenol were carried out in a continuously working aerobic solid-bed reactor. The main emphasis was on the dynamic behaviour of the system after a sudden change in cultivation conditions and on the steady-state performance of the reactor as a function of the pollution load. A change from ammonium-free to ammonium-containing medium hardly influenced the nitrophenol degradation. The reactor responded differently to an increase in pollutant load, which was brought about by increasing either the 4-nitrophenol content or the flow of the waste water. Up to a load of 270 mg l⁻¹h⁻¹ the pollutant was stably and almost completely degraded. At a higher load, only a partial 4-nitrophenol degradation took place. A mathematical model was derived to describe the processes that occurred in the reactor. By segregation into two compartments – the aqueous phase and the biofilm – account was taken of the fact that the pollutant is carried into the biofilm by diffusion and is degraded there. The observed relations between the pollutant load, the pollutant concentration in the outlet of the reactor and the reactor performance agreed with the simulated process behaviour. As the model simulation showed, the incomplete pollutant degradation at a higher reactor load was caused by oxygen limitation. Received: 5 August 1998 / Received revision: 22 October 1998 / Accepted: 24 October 1998     

Benzene/toluene/p-xylene degradation. Part I. Solvent selection and toluene degradation in a two-phase partitioning bioreactor

A two-phase organic/aqueous reactor configuration was developed for use in the biodegradation of benzene, toluene and p-xylene, and tested with toluene. An immiscible organic phase was systematically selected on the basis of predicted and experimentally determined properties, such as high boiling points, low solubilities in the aqueous phase, good phase stability, biocompatibility, and good predicted partition coefficients for benzene, toluene and p-xylene. An industrial grade of oleyl alcohol was ultimately selected for use in the two-phase partitioning bioreactor. In order to examine the behavior of the system, a single-component fermentation of toluene was conducted with Pseudomonas sp. ATCC 55595. A 0.5-l sample of Adol 85 NF was loaded with 10.4 g toluene, which partitioned into the cell containing 1 l aqueous medium at a concentration of approximately 50 mg/l. In consuming the toluene to completion, the organisms were able to achieve a volumetric degradation rate of 0.115 g l⁻¹ h⁻¹. This system is self-regulating with respect to toluene delivery to the aqueous phase, and requires only feedback control of temperature and pH. Received: 16 November 1998 / Received revision: 28 March 1999 / Accepted: 9 April 1999     

Nitrification, denitrification, and dechlorination in bleached kraft pulp mill wastewater

This study deals with combining the biologi cal removal of organic halogens with the removal of nitrogen from bleached kraft pulp mill wastewater in fluidized-bed reactors under nitrifying and denitrifying conditions. Untreated and biotreated bleached kraft pulp mill wastewaters had no detrimental effect on nitrification or denitrification. The nitrifying biofilm reactor, pregrown on synthetic inorganic feed with ammonia, removed without a lag phase adsorbable organic halogens [7.2 mg Cl (g biomass volatile solids)⁻¹day⁻¹] from bleached kraft pulp mill wastewater and selected chlorophenols from synthetic wastewater. Electron microscopical examination of the biofilm showed that bacteria, morphologically similar to the nitrifying species Nitrosomonas or Nitrobacter, and Nitrosospira were dominant. The denitrifying fluidized-bed reactor, pregrown on nitrate and methanol, denitrified without a lag phase bleached kraft pulp mill wastewater. Under denitrifying conditions, 35% of the total organic carbon content of untreated bleached kraft pulp mill waste water was removed. The reducing power delivered by untreated bleached kraft pulp mill wastewater for denitrification was 2 mmol electrons/mmol carbon mineralized. Dechlorination under denitrifying conditions was negligible. Received: 21 November 1996 / Received revision: 27 January 1997 / Accepted: 1 February 1997     

Validation of the doubly labeled water method in Japanese Quail Coturnix c. japonica chicks: is there an effect of growth rate?

The Doubly Labeled Water (DLW) method was validated against respiration gas analysis in growing Japanese Quail chicks (between 1 week and 3 weeks of age) as well as in birds after having achieved sexual maturity (7 weeks of age). A comparison was made between a strain selected for high growth rates (P-strain, n=18), and a non-selected strain (C-strain, n=18). Relative growth rates of individual chicks during the measurement ranged from −13.8% day⁻¹ to 23.1% day⁻¹. When employing a single-pool model (eq. 34, Lifson and McClintock 1966), it was found that the relative error of the DLW method was sensitive to assumptions concerning fractional evaporative water loss. The best fit was obtained after taking a fractional evaporative water loss value of 0.33. When applying this value for all chicks, it was found that neither strain, relative growth rate of the chick during measurement, nor age significantly contributed to the explained variance. When employing two-pool models, it was found that the DLW method significantly underestimated the true rates of CO₂ production at all assumed levels of fractional evaporative water loss. Based on an evaluation of DLW validation studies in growing shorebirds, terns, and quail we recommend Speakman's Eq. 7.17 (Speakman 1997) for general use in young birds. Accepted: 14 April 2000     

Monitoring nitric oxide from immobilised denitrifying bacteria, Pseudomonas stutzeri, by the use of chemiluminescence

A chemiluminescence detector was used to measure the production of nitric oxide, NO, from the denitrifying bacteria Pseudomonas stutzeri. NO is an intermediate when P. stutzeri converts nitrate into nitrogen gas. The reaction between NO and ozone is selective and sensitive in generating chemiluminescence. Calibrations were made down to 1 nM, with a signal-to-noise ratio of 3. Bacteria were immobilised in alginate beads. Denitrification experiments were made in an anaerobic non-growth medium by adding nitrate to a certain concentration in the reactor. The bacteria were exposed to nitrate in the concentration range 1 pM–5 mM. The lowest concentration to give a measurable NO response was 100 nM. Received: 16 October 1997 / Received revision: 20 January 1998 / Accepted: 24 January 1998