Point source detoxification of an industrially produced 3,4-dichloroaniline-manufacture wastewater using a membrane bioreactor

A membrane bioreactor has been used to treat an industrially produced waste-water containing aniline, 4-chloroaniline, 2,3-dichloroaniline and 3,4-dichloroaniline. Conventional direct biological treatment of such effluents cannot be implemented without some form of pretreatment or dilution because of the hostile inorganic composition of the waste-water. In order to overcome this problem a membrane separation step selectively removes the organics from the waste-water and subsequent biodegradation takes place in the biological growth compartment of the reactor system. At a waste-water flow rate of 69 ml h^–1 (corresponding to a contact time of approximately 1.5 h) over 99% of the organic compounds quoted above were removed and biodegraded. Correspondence to: A. G. Livingston     

Mineralization of diuron [3-(3,4-dichlorophenyl)-1, 1-dimethylurea] by co-immobilized <Emphasis Type="Italic">Arthrobacter</Emphasis> sp. and <Emphasis Type="Italic">Delftia acidovorans</Emphasis>

Mineralization of diuron has not been previously demonstrated despite the availability of some bacteria to degrade diuron into 3,4-dichloroaniline (3,4-DCA) and others that can mineralize 3,4-DCA. A bacterial co-culture of Arthrobacter sp. N4 and Delftia acidovorans W34, which respectively degraded diuron (20 mg l⁻¹) to 3,4-DCA and mineralized 3,4-DCA, were able to mineralize diuron. Total diuron mineralization (20 mg l⁻¹) was achieved with free cells in co-culture. When the bacteria were immobilized (either one bacteria or both), the degradation rate was higher. Best results were obtained with free Arthrobacter sp. N4 cells co-cultivated with immobilized cells of D. acidovorans W34 (mineralization of diuron in 96 h, i.e., 0.21 mg l⁻¹ h⁻¹ vs. 0.06 mg l⁻¹ h⁻¹ with free cells in co-culture).     

A novel bioreactor system for the destruction of volatile organic compounds

The biological treatment of waste-waters containing 1,2-dichloroethane (DCE) in conventional bioreactors results in air-stripping of DCE. In the present work, a novel bioreactor system intended to overcome this problem has been developed for the treatment of a synthetically concocted DCE-containing waste-water (1000 mg DCE l^–1). The operation of a conventional air-lift bioreactor at a waste-water flow rate of 0.24 l h^–1 led to 33% of the DCE supplied to the reactor being lost to the exit gas stream. The use of the novel enclosed system, operated with a recycling O₂ sparge instead of air, resulted in negligible air-stripping at the same waste-water flow rate. A control system was implemented to add O₂ as required to maintain the pressure of the recycle gas stream, and a scrubber removed the CO₂ produced. Over 99% of DCE supplied was biodegraded during operation of this system, and virtually all carbon entering the system was evolved as CO₂. Correspondence to: A. G. Livingston Correspondence to: A. G. Livingston     

Biodegradation of pyridine by freely suspended and immobilized Pimelobacter sp.

Freely suspended and Ca-alginate-immobilized cells of Pimelobacter sp. were used for degradation of pyridine. When the pyridine concentration was up to 2 g l^–1, freely suspended cells completely degraded pyridine regardless of the initial cell concentrations used. However, when the pyridine concentration increased to 4 g l^–1, the initial cell concentration in freely suspended cell culture should be higher than 1.5 g dry cell weight l^–1 for complete degradation of pyridine. In addition, a freely suspended cell culture with a high initial cell concentration resulted in a high volumetric pyridine-degradation rate, suggesting the potential use of immobilized cells for pyridine-degradation. When the immobilized cells were used for pyridine-degradation, neither specific pyridine-degradation rate nor tolerance against pyridine was improved. However, a high volumetric pyridine-degradation rate in the range 0.082–0.129 g l^–1 hr^–1 could be achieved by the immobilized cells because of the high cell concentration. Furthermore, when the immobilized cells were reused in degrading pyridine at a concentration of 2–4 g l^–1 they did not lose their pyridine-degrading activity for 2 weeks. Taken together, the data obtained here showed the feasibility of using immobilized cells for pyridine-degradation.     

L-Sorbose production in a continuous fermenter with and without cell recycle

The kinetics of continuous l-sorbose fermentation using Acetobacter suboxydans with and without cell recycle (100%) were investigated at dilution rates (D) of 0.05, 0.10, 0.15 and 0.3 h^–1. The biomass and sorbose concentrations for continuous fermentation without recycle increased as the dilution rate was increased from 0.05 to 0.10 h^–1. A maximum biomass concentration of 8.44 g l^–1 and sorbose concentration of 176.90 g l^–1 were obtained at D=0.10 h^–1. The specific rate of sorbose production and volumetric sorbose productivity at this dilution rate were 2.09 g g^–1 h^–1 and 17.69 g l^–1 h^–1. However, on further increasing the dilution rate to 0.3 h^–1, both biomass and sorbose concentrations decreased to 2.93 and 73.20 g l^–1 respectively, mainly due to washout of the reactor contents. However, the specific rate of sorbose formation and volumetric sorbose productivity at this dilution rate increased to 7.49 g g^–1 h^–1 and 21.96 g l^–1 h^–1 respectively. Continuous fermentation with 100% cell recycle served to further enhance the concentration of biomass and sorbose to 28.27 and 184.32 g l^–1 respectively (in the reactor at a dilution rate of 0.05 h^–1). Even though, there was a decline in the biomass and sorbose concentrations to 6.8 and 83.40 g l^–1 at a dilution rate of 0.3 h^–1, the specific rates of sorbose formation and volumetric sorbose productivity increased to 3.67 g g^–1h^–1 and 25.02 g l^–1 h^–1.     

Scale-up performance of a partitioning bioreactor for the degradation of polyaromatic hydrocarbons by Sphingomonas aromaticivorans

A partitioning bioreactor, consisting of an aqueous phase containing Sphingomonas aromaticivorans and an immiscible organic phase (dodecane), loaded with naphthalene and phenanthrene, was operated at two scales, 5 l and 150 l. Complete degradation of 15 g and 300 g, respectively, of these polyaromatic hydrocarbon (PAH) mixtures was achieved in 21 h in both cases resulting in a volumetric PAH degradation rate of 238 mg l^–1 h^–1 based on reactor aqueous volumes.     

Degradation of sodium anthraquinone sulphonate by free and immobilized bacterial cultures

Aerobic biodegradation of a xenobiotic recalcitrant compound sodium anthraquinone-2-sulphonate (SAS), was investigated using as an inoculum a mixed microbial culture, which was activated sludge from industrial and domestic waste-water treatment plants. The difference in SAS degradation was examined using two main systems: (1) suspended cells and (2) immobilized cells, both in batch and in continuous culture. In the suspended cell system, under continuous culture conditions using SAS as a unique source of carbon and energy, it was possible to degrade about 95% of this substrate after 6 days. Maximal SAS removal rates in the suspended-cell system were 593 mg SAS l^–1 h^–1 and 88.7 mg SAS l^–1 h^–1 for dilution rates (D) of 0.05 h^–1 and 0.075 h^–1, respectively. In the immobilized-cell system, almost all SAS was degraded in 6 days and the maximal removal rate reached 88.7 mg SAS l^–1 h^–1 at D=0.05 h^–1. Application of a continuous-flow enrichment procedure resulted in selection of several kinds of micro-organisms and led to a progressive elimination of some species of Aeromonas. A stable microbial community of 11 strains has been established and characterized at D=0.075 h^–1. Most of them were Gram-negative and belonged to the genus Pseudomonas.     

Degradation kinetics of phenol by immobilized cells of Candida tropicalis in a fluidized bed reactor

Degradation kinetics of phenol by free and agar-entrapped cells of Candida tropicalis was studied in batch cultures. The initial phenol degradation rate achieved with free cells was higher than that obtained with immobilized cells, when phenol concentrations up to 1000 mg l^–1 were used. However, at higher phenol concentrations, the behaviour was quite different. The initial degradation rate of the immobilized yeast cells was about 10 times higher than that of the free cells, at a phenol concentration of 3500 mg l^–1. The semicontinuous and continuous degradation of phenol by immobilized yeast cells was also investigated in a multi-stage fluidized bed reactor. The highest phenol removal efficiencies and degradation rates as well as the lowest values of residual phenol and chemical oxygen demand were obtained in the semicontinuous culture when phenol concentrations up to 1560 mg l^–1 were used.     

Degradation of 2,4-Dinitrophenol by Free and Immobilized Cells of Rhodococcus erythropolis HL PM-1

The degradation of 2,4-dinitrophenol (2,4-DNP) by Rhodococcus erythropolis HL PM-1 was studied. The enzymes involved in 2,4-DNP degradation were inducible, and their resynthesis took place during the process. Cell immobilization by embedding into agar gels decreased the degrader activity. The maximum rates of 2,4-DNP degradation by free and immobilized cells were 10.0 and 5.4 nmol/min per mg cells, respectively. The concentration dependence of 2,4-DNP degradation was typical of substrate inhibition kinetics. The immobilized cells were used in a model reactor designed for 2,4-DNP biodegradation. Its maximum capacity was 0.45 nmol/min per mg cells at a volumetric flow rate of 20 h^–1. The reactor operated for 14 days without losing capacity; its half-life equaled 16 days.     

Production of citric acid with immobilized Yarrowia lipolytica

Summary Citric acid was produced with immobilized Yarrowia lipolytica yeast in repeated batch-shake-flask and air-lift fermentations. In active and passive immobilization methods calcium alginate, -carrageenan, polyurethane gel, nylon web and polyurethane foams were tested as carriers in repeated-batch fermentations. The highest citric acid productivity of 155 mg l^–1 h^–1 was reached with alginate-bead-immobilized cells in the first batch. A decrease in bead diameter from 5–6 mm to 2–3 mm increased the volumetric citric acid productivity threefold. In an air-lift bioreactor the highest citric acid productivity of 120 mg l^–1 h^–1 with a product concentration of 16.4 g l^–1 was obtained with cells immobilized in -carrageenan beads. Offprint requests to: H. Kautola     

2-Propanol degradation by Sulfolobus solfataricus

Sulfolobus solfataricus used 2-propanol and 2-propanone (acetone) when grown in static cultures at 78 °C with or without glucose at 10 g l^–1. The presence of 3.92 g 2-propanol l^–1 in both cases inhibited growth. However, acetone accumulation following 2-propanol depletion suggested that 2-propanol was co-metabolized via the acetone metabolic pathway. Glucose at 10 g l^–1 increased 2-propanol and acetone utilization from 0.93 g l^–1 to 1.77 g l^–1 and from 0.11 g l^–1 to 1.62 g l^–1, respectively. Without glucose, immobilized S. solfataricus cells increased the 2-propanol removal rate to 0.035 g l^–1 h^–1, compared to 0.0012 g l^–1 h^–1 by its suspended counterpart. The results suggest the establishment of an immobilized reactor configuration is preferential for the treatment of high temperature solvent waste streams by this acidothermophilic Crenarchaeon.     

Direct alcoholic fermentation of starchy biomass using amylolytic yeast strains in batch and immobilized cell systems

Summary Direct alcoholic fermentation of dextrin or soluble starch with selected amylolytic yeasts was studied in both batch and immobilized cell systems. In batch fermentations, Saccharomyces diastaticus was capable of fermenting high dextrin concentrations much more efficiently than Schwanniomyces castellii. From 200 g·l^–1 of dextrin S. diastaticus produced 77 g·l^–1 of ethanol (75% conversion efficiency). The conversion efficiency decreased to 59% but a higher final ethanol concentration of 120 g·l^–1 was obtained with a medium containing 400 g·l^–1 of dextrin. With a mixed culture of S. diastaticus and Schw. castellii 136 g·l^–1 of ethanol was produced from 400 g·l^–1 of dextrin (67% conversion efficiency). S. diastaticus cells attached well to polyurethane foam cubes and a S. diastaticus immobilized cell reactor produced 69 g·l^–1 of ethanol from 200 g·l^–1 of dextrin, corresponding to an ethanol productivity of 7.6g·l^–1·h^–1. The effluent from a two-stage immobilized cell reactor with S. diastaticus and Endomycopsis fibuligera contained 70 g·l^–1 and 80 g·l^–1 of ethanol using initial dextrin concentrations of 200 and 250 g·l^–1 respectively. The corresponding values for ethanol productivity were 12.7 and 9.6 g·l^–1·h^–1. The productivity of the immobilized cell systems was higher than for the batch systems, but much lower than for glucose fermentation.     

Immobilization of invertase on a new type of macroporous glycidyl methacrylate

Invertase was immobilized via its carbohydrate moiety. The immobilized enzyme has a specific activity of 5500 IU g^–1, with 45% activity yield on immobilization. In a packed bed reactor, 90% 2.5 M sucrose was converted at a flow rate of 4 bed volumes h^–1. The obtained specific productivity at 40 °C of 3 kg l^–1 h^–1 is the best one so far. Long-term stability was 290 days in 2.5 M sucrose at 40 °C and at a flow rate of 3 bed volumes h^–1.     

Continuous fermentation of sweet whey permeate for propionic acid production in a CSTR with UF recycle

Summary Propionic acid was produced byPropionibacterium acidi-propionici from sweet-whey permeate in a stirred tank reactor (CSTR) with cell recycle by ultrafiltration. The highest volumetric productivity achieved was 14.3 g.l^–1. h^–1, with a biomass of 100 g.l^–1 (dry weight). More concentrated product can be obtained by electrodialysis of the cell free fermentation medium.     

Enrichment and kinetics of biodegradation of 1-naphthylamine in activated sludge

Sequencing-batch reactors were used to develop an activated sludge enrichment culture capable of degrading 1-naphthylamine (1NA). Approximately 5 months acclimation with salicylic acid (1600 mg l^–1) as the primary source of carbon were required to obtain an enrichment culture able to degrade even small quantities of 1NA. After an additional 4 months acclimation, during which the concentration of salicyclic acid was decreased to 50 mg l^–1, a culture developed that degraded 1NA concentrations as high as 300 mg l^–1. Kinetic determinations showed that 1NA degradation (in the presence of salicylate) followed Michaelis-Menten kinetics with K _m and V _m values of 32.5±2.2 mg l^–1 and 375±18 ng 1NA mg^–1 cells h^–1, respectively. The same enrichement was able to degrade 1NA when present as the sole source of carbon and energy and to convert approximately 87% to CO₂.     

Propionic acid and biomass production using continuous ultrafiltration fermentation of whey

Summary This paper presents a study of propionic acid and propionibacteria production from whey by usingPropionibacterium acidi-propionici in continuous fermentation with cell recycle. The highest propionic acid volumetric productivity achieved was 5 g.l^–1.h^–1 with no biomass bleeding. A maximal biomass concentration of 130 g.l^–1 was achieved before initiating biomass bleeding to give a biomass volumetric productivity of 3.2 g.l^–1.h^–1 with a biomass of 75 g.l^–1 and a propionic acid productivity of 3.6 g.l^–1.h^–1 (for about 100 hours i.e. more than 50 residence times).     

Degradation of caffeine by immobilized cells of Pseudomonas putida strain C 3024

Summary A caffeine-resistant strain of Pseudomonas putida was isolated from soil and was grown with caffeine as the sole source of carbon, energy and nitrogen. Cells were immobilized in agar gel particles which were continuously supplied with a caffeine solution (0.52 g · l^–1, D=1.0 h^–1) in a homogeneously mixed aerated reaction vessel. In the presence of the ATPase inhibitor arsenate the caffeine was removed by the immobilized cells at an average rate of 0.25 mg caffeine · h^–1 · (mg cell carbon)^–1 during 6 days. Thereafter a rapid decline of activity was observed. From a similar system without arsenate supplied with a growth medium containing a limiting amount of caffeine (0.13 g · l^–1) the caffeine was almost completely oxidized by the immobilized cells. The concentration of the remaining caffeine was 1.4 mg · l^–1, which is much lower than the substrate constant for caffeine (9.7 mg · l^–1) observed with freshly harvested suspended resting cells.     

Improvement of the culture stability of non-anchorage-dependent animal cells grown in serum-free media through immobilization

A murine hybridoma cell line producing a monoclonal antibody against penicillin-G-amidase and a murine transfectoma cell line secreting a monovalent chimeric human/mouse Fab-antibody fragment were cultivated in three different media (serum-containing, low protein serum-free, and iron-rich protein-free) in flask cultures, stirred reactors and a fixed bed reactor. In static batch cultures in flasks both cell lines showed similar good growth in all three media.In suspension in a stirred reactor, the hybridoma cell line could be cultivated satisfactory only in serum-containing medium. In low protein serum-free medium, Pluronic F68 had to be added to protect the hybridoma cells against shear stress. But even with this supplement only batch, not chemostat mode was possible. In iron-rich protein-free medium the hybridoma cells grew also in continuous chemostat mode, but the stability of the culture was low. The transfectoma cell line did not grow in stirred reactors in any of the three media.Good results with both cell lines were obtained in fixed bed experiments, where the cells were immobilized in macroporous Siran^®-carriers. The media, which were optimized in flask cultures, could be used without any further adaptation in the fixed bed reactor. Immobilization improved the stability and reliability of cultures of non-adherent animal cells in serum-free media tremendously compared to suspension cultures in stirred reactors. The volume-specific glucose uptake rate, an, indicator of the activity of the immobilized cells, was similar in all three media. Deviations in the metabolism of immobilized and suspended cells seem to be mainly due to low oxygen concentrations within the macroporous carriers, where the cells are supplied with oxygen only by diffusion.List of symbols c substrate or product concentration mmol l^–1 - c₀ substrate or product concentration in the feed mmol l^–1 - c_Glc glucose concentration mmol l^–1 - c_Gln glutamine concentration mmol l^–1 - c_Amm ammonia concentration mmol l^–1 - c_Lac lactate concentration mmol l^–1 - c_FAB concentration of Fab# 10 antibody fragment g l^–1 - c_MAb monoclonal antibody concentration mg l^–1 - D dilution rate d^–1 - q cell-specific substrate uptake or metabolite production rate mmol cell^–1 h^–1 - q_Glc cell-specific glucose uptake rate mmol cell^–1 h^–1 - q_Gln cell-specific glutamine uptake rate mmol cell^–1 h^–1 - q_MAb cell-specific MAb production rate mg cell^–1 h^–1 - q^* volume-specific substrate uptake or metabolite production rate mmol l^–1 h^–1 - q^*FB volume-specific substrate uptake or metabolite production rate related to the fixed bed volume mmol l_FB ^–1 h^–1 - q^*FB,Glc volume-specific glucose uptake rate related to the fixed bed volume mmol l_FB ^–1 h^–1 - q^*FB,Gln volume-specific glutamine uptake rate related to the fixed volume mmol l_FB ^–1 h^–1 - q^*FB,MAb volume-specific MAb production rate related to the fixed volume mg l_FB ^–1 h^–1 - q^*FB,02 volume-specific oxygen uptake rate related to the fixed bed volume mmol l_FB ^–1 h^–1 - t time h - U superficial flow velocity mm s^–1 - V medium volume in the conditioning vessel of the fixed bed reactor l - V_FB volume of the fixed bed l - x_v viable cell concentration cells ml^–1 - y_Amm,Gln yield of Ammonia from glutamine - y_Lac,Glc yield of lactate from glucose - specific growth rate h^–1 - _d specific death rate h^–1     

Immobilized cells cultivated in semi-continuous mode in a fluidized bed reactor for xylitol production from sugarcane bagasse

Summary Xylitol production from sugarcane bagasse hemicellulosic hydrolyzate was evaluated in a fluidized bed reactor operated in semi-continuous mode, using cells immobilized on porous glass. The fermentative process was performed during five successive cycles of 72 h each one. The lowest xylitol production occurred in the first cycle, where a high cell concentration (12 g l⁻¹) was observed. In the subsequent cycles the xylitol concentration was ever increasing due to the cells adaptation to the medium. In the last one, 18 g xylitol l⁻¹ was obtained with a yield factor of 0.44 g g⁻¹ and volumetric productivity of 0.32 g l⁻¹ h⁻¹.     

Continuous production ofl-lactic acid from whey permeate by immobilizedLactobacillus casei subsp.casei

Summary The production of l-lactic acid from whey permeate, a waste product of the dairy industry, by fermentation with the lactic acid bacterium Lactobacillus casei subsp. casei was investigated. A fermentation medium consisting of permeate and supplements, which enables exponential growth of the organisms, was developed. A fast method for determination of free and immobilized biomass in solid-rich media, based on measurement of cellular ATP, was evolved. Continuous fermentations in a stirred tank reactor (STR) and in a fluidized bed reactor (FBR) with immobilized biomass were compared. In the STR a volumetric productivity of 5.5 g/l per hour at 100% substrate conversion [dilution rate (D) = 0.22 h^–1] was determined. In the FBR porous sintered glass beads were used for immobilization and a maximum biomass concentration of 105 g/kg support was measured. A productivity of 10 g/l per hour was obtained at D = 0.4 h^–1 (substrate conversion 93%) and of 13.5 g/l per hour at D = 1.0 h^–1 (substrate conversion 50%). Offprint requests to: W. Krischke