5436
Ind. Eng. Chem. Res. 2001, 40, 5436-5439
Aerobic Phenol Biodegradation in an Inverse Fluidized-Bed Biofilm Reactor Katerina Kryst and Dimitre G. Karamanev* Department of Chemical and Biochemical Engineering, University of Western Ontario, London, Ontario, N6A 5B9 Canada
A new type of bioreactor, an inverse fluidized-bed biofilm reactor, was used for the continuous aerobic biodegradation of phenol in contaminated water. The process proved to be very stable in time, with a constant and high rate of biodegradation obtained during the three-month operation of the bioreactor. This was because of the efficient and simple biofilm thickness control. The effect of the dilution rate on the rate of phenol biodegradation was studied at different inlet phenol concentrations. The biodegradation rate was found to pass through a maximum. Using a kinetic model, this effect was explained by the presence of a large amount of suspended biomass, produced during the biofilm thickness control in the bioreactor. Introduction Phenol has widespread use, including applications as an antioxidant, a herbicide, and many others. Due to its extensive past and present use, phenol is a pollutant commonly found in the environment.1 Because phenol is a model pollutant for the study of the biodegradation of recalcitrant compounds, it has been widely studied, resulting in a large amount of data being available. Most of the data available on phenol biodegradation is based on free suspended cultivation of microbial cells. However, the use of immobilized cells has proved to be much more efficient because of the higher cell concentration per unit volume of bioreactor,2 the higher cell resistance to the toxic effect of phenol and to variations in temperature and pH, and the higher degree of process robustness in general. Especially promising is the use of naturally immobilized cells, or biofilm. Several different types of biofilm reactors have been studied, including rotating-disk bioreactors, fixed-bed reactors and three-phase fluidized-bed bioreactors. The latter are considered superior because of the large biofilm support surface, appropriate hydrodynamic conditions, high mass-transfer rates of both oxygen and substrate, and excellent contact between the liquid and solid phases.2 However, there is one significant problem in practically all types of biofilm reactors, including fluidizedbed reactors: uncontrolled biofilm growth. Uncontrolled growth leads to substrate mass-transfer limitations, lysis, and detachment of biofilm from the support. In addition, in the case of fluidized-bed bioreactors, the uncontrolled biofilm growth results in overexpansion of the fluidized bed with subsequent elutriation of the particles.3 Some researchers have attempted to “control” this biofilm by removing some of the solid phase and replacing it with biomass-free particles.4,5 The inverse fluidized-bed bioreactor was developed to achieve simple and efficient control of biofilm growth.6 So far, it has been used for the treatment of domestic wastewater,7 as well as for the biologically assisted production of uranium. The bioreactor is a combination of an inverse fluidized-bed and draft tube reactor, as * Corresponding author. Fax: +1 (519) 661 3498. E-mail:
[email protected].
illustrated in Figure 1. Air is introduced at the bottom of the draft tube, creating an upflow through the draft tube and a downflow in the annulus. Inert particles for biofilm support, which are less dense than water, are introduced into the reactor and are fluidized by the downward liquid flow in the annulus. To control the biofilm thickness, a bed of heavy inert particles is supported on a grid placed in the draft tube above the air sparger. Initially, the lower level of inverse fluidized bed is kept well above the lower opening of the draft tube by varying the liquid circulation rate through the gas flow rate. When the reactor is inoculated, the microorganisms attach themselves to the immobilized particles, forming a biofilm. As the biofilm grows, the overall bioparticle (support particle covered with biofilm) density increases. This leads to expansion of the bed downward. Bed expansion continues until a critical biofilm thickness is reached, at which point the lower part of the inverse fluidized bed reaches the lower draft tube opening. At that moment, the heaviest particles (those with the thickest biofilm) enter the draft tube. They proceed through the bed of heavy inert particles, where some of the biofilm is removed by attrition. The particles are then carried to the top of the fluidized bed. Thus, the biofilm thickness in the reactor from this point on remains constant. By changing the gas flow rate or hydraulic resistance the biofilm thickness can, therefore, be easily controlled within a very narrow range.6 Most biofilm reactor models assume that the suspended biomass concentration is negligible compared with that of the biofilm. However, in an inverse fluidized-bed bioreactor, there is a continuous production of suspended cells because of the biofilm thickness control. The amount of these cells could be comparable to the amount of cells in the biofilm. The effect of free suspended cells released by biofilm on the performance of a biofilm reactor can be determined by a kinetic model proposed earlier.8 The model was based on the assumption that the amount of immobilized microorganisms (biofilm) remains constant, but each cell of the biofilm produces two daughter cells in a regular time interval. The excess of cells is released into liquid phase as free suspended biomass, with these cells also producing new cells. The total number of cells
10.1021/ie010036f CCC: $20.00 © 2001 American Chemical Society Published on Web 09/08/2001
Ind. Eng. Chem. Res., Vol. 40, No. 23, 2001 5437
Figure 2. Substrate biodegradation rate as a function of dilution rate of immobilized and suspended cells.
Figure 1. Schematic of an inverse fluidized-bed biofilm reactor.
produced, which at steady state is equal to the number of cells leaving the bioreactor, is due to the cell production from both immobilized and free suspended cells. Although the kinetic model was originally proposed for the case of a biofilm structure different from that observed in this study, the model8 is applicable to the immobilized system used in this study. From the model, the substrate utilization rate, or in this study, the phenol degradation rate (r), is given by
[
r ) D S0 -
]
[-B - xB2 - 4DS0KS(µm - D)] 2(µm - D)
(1)
where B is given by
(
B ) D(S0 - KS) - µm
)
Xfix + S0 Y
(2)
The model represented by eq 1 was developed on the basis of the following assumptions: (1) The concentrations of cells in the biofilm and the cells suspended within the liquid medium are constant in time. (2) The kinetics of substrate utilization, product formation, and biomass growth are the same for free suspended cells and immobilized cells. (3) The liquid in the bioreactor is perfectly mixed. (4) The process is in a continuous steady-state regime. From this model, when a reactor contains a significant concentration of suspended and fixed cells, the reaction rate passes through a maximum at a specific dilution rate. Also, the integrated system has a higher degradation rate than purely immobilized systems or purely suspended systems, as shown in Figure 2. The main goals of this study were to determine the potential of the inverse fluidized-bed bioreactor for longterm operation and efficiency of aerobic phenol biodegradation, to determine the long-term stability of the system, and to use the kinetic model of biofilm plus suspended biomass for a description of the results obtained. Experimental Section The bioreactor was a vertical cylinder constructed of transparent polymethylpentene and had a working volume of 0.9 L (Figure 1). Its diameter was 6.5 cm. A
converging-diverging conical insert with a shape similar to an hourglass was installed just above the draft tube to eliminate dead zones near the reactor wall and to avoid particle accumulation above the liquid level. The support particles were made of expanded polystyrene (Styrofoam) and had a spherical shape and an average density of 800 kg/m3. The bulk volume of particles used was 300 mL, with diameters ranging between 0.8 and 1.6 mm. To aid microbial attachment, the external surface of the Styrofoam particles was coated with carbon powder. The microbial consortium used in this study was developed in an immobilized soil bioreactor9 from a soil sample obtained from a contaminated site. The bioreactor was operated initially in batch mode with increasing initial phenol concentrations, and after a constant rate of biodegradation was obtained, the reactor was switched to continuous operation. The concocted wastewater contained phenol with concentrations ranging between 30 and 350 mg/L (a typical range for wastewaters) and nutrient salts with the following composition: 840 mg of KH2PO4, 750 mg of K2HPO4, and 1200 mg of (NH4)2SO4 per liter. This solution was fed from the top of the reactor through silicone tubing by a Masterflex variable-speed peristaltic pump with flow rates between 0.05 and 6.5 L/h. The reactor was aerated with 260 L/h of air fed through a perforated (0.5-mm hole diameter) plastic tube. The air flow rate was measured with a calibrated rotameter and the dissolved oxygen was measured using an Orion 850 DO meter. An Orion 420a pH meter was used to measure the pH of the liquid. The phenol concentration was analyzed using the standard aminoantipyrine method,10 with absorbance measurements at 500 nm utilizing a Cary-50 (Varian Canada, Mississauga) UV-visible spectrophotometer. The suspended biomass concentration was determined by dry weight.10 Results and Discussion The reactor was inoculated by feeding the outlet from the immobilized soil reactor for approximately 5 days. It was operated in the continuous regime from the beginning of inoculation. After inoculation with the microbial consortium, a visible biofilm developed within 3 days, and the concocted wastewater was added directly into the reactor. About 1 week after inoculation, the reactor began foaming extensively. The foam was very thick and dense and did not collapse. A similar type of foaming has been observed by other researchers
5438
Ind. Eng. Chem. Res., Vol. 40, No. 23, 2001
Figure 3. Effect of the dilution rate on the rate of phenol biodegradation and biomass production in an inverse fluidizedbed biofilm reactor. Input phenol concentration ) 59 mg/L.
during biodegradation of phenol.11 This entrapped some support particles, which were expelled out of the reactor with the foam. Eventually, it was determined that the foaming greatly decreased if the biomass at the bottom of the reactor was not allowed to settle. Therefore, a magnetic stirrer at the bottom of the bioreactor rotating at a very slow speed kept the suspended biomass afloat, ensuring that it would not accumulate, but would be expelled with the outlet flow. Once steady-state conditions were achieved with continuous liquid feed, the effect of the dilution rate on the phenol biodegradation was studied. The effect of dilution rate on the effluent phenol concentration at a constant influent phenol concentration of 59 mg/L is shown in Figure 3. From the graph, it can be seen that the degradation rate passed through a maximum at a specific dilution rate. In this case, the maximum rate of 180 mg/(L h) occurred at a dilution rate of about 3.6 h-1. At lower dilution rates, the biofilm production rate gradually increased, but at dilution rates beyond approximately 3.5 h-1, the apparent biomass production rate greatly increased, as can be seen in Figure 3. One reason for this increase might be that the diameter of the tube for the outlet liquid flow was relatively small and the flow resistance at higher liquid flow rates became so large that the height of the liquid level in the reactor increased. As a result, the biofilm thickness on the support particles was decreased, and more biomass was produced during these unsteady-state conditions, temporarily increasing the apparent biomass production rate. The experimental phenol biodegradation rate was compared to the value calculated by eq 1. The resulting theoretical curve is also shown in Figure 3. A very good fit between the experimental and theoretical results can be noted. The curve was produced by least-squares fitting using kinetic constants of the process as independent variables. The values of the kinetic parameters representing the best-fit curve are Ks ) 0.10 mg/L, µm ) 2.18 h-1, and Xfix/Y ) 33.6 mg/L. The second study was conducted at an inlet phenol concentration of 80 mg/L. The resulting data are shown in Figure 5. Again, the degradation rate passed through a maximum at a specific dilution rate. In this case, the peak was not as strongly pronounced as in Figure 3; however, the maximum degradation rate of above 170 mg/(L h) could be observed at a dilution rate of around 2.4 h-1. One thing that did differ from Figure 3 was the
Figure 4. Photograph of bioparticles in inverse fluidized bed.
Figure 5. Effect of dilution rate on the rate of phenol biodegradation and biomass production in an inverse fluidized-bed biofilm reactor. Input phenol concentration ) 80 mg/L.
constant biomass production rate of around 25 mg/(L h) in the studied dilution range of 1.4-4 h-1. According to our explanation above, no increase was observed because of the lower dilution rates, or liquid flow rates, than those in Figure 3. The maximum degradation rate of 180 mg/(L h) was observed for the inverse fluidized-bed bioreactor. The highest value found published in the literature was a degradation rate of 638 mg/(L h) in a tapered fluidizedbed bioreactor using a mutant strain of Pseudomonas bacterium.12 However, this rate occurred when only 30% of the inlet phenol was degraded. To ensure that the reactor was not operating under oxygen-limiting conditions, which would make the model used invalid, the oxygen concentration profile along the height of the inverse fluidized bed was measured. Because almost all support particles were in the annulus, it can be assumed that the majority of the reaction occurred there. Because only the riser was aerated, the highest concentration of oxygen in the annulus was expected to be at the top of the inverse fluidized bed, as liquid flows downward. This was confirmed by the data, which are shown in Figure 6. The minimum oxygen concentration at the bottom of the bed was found to be 3.6 mg/L. The previous mathematical model of an inverse fluidized-bed biofilm reactor13 showed that, in a biofilm with similar thickness, sub-
Ind. Eng. Chem. Res., Vol. 40, No. 23, 2001 5439 KS ) saturation constant, mg/L r ) phenol degradation rate, mg/(L h) S ) outlet phenol concentration, mg/L S0 ) input phenol concentration, mg/L Xfix ) fixed biofilm concentration, mg/L Y ) yield coefficient, mg/mg µm ) maximum specific growth rate, 1/h kbf ) biofilm density (wet), kg/m3 kliq ) liquid density, kg/m3 kpart ) support particle density, kg/m3
Acknowledgment This work was supported by the Natural Sciences and Engineering Research Council of Canada. Figure 6. Profile of oxygen concentration in liquid as a function of fluidized-bed height.
strate diffusion properties, and oxygen consumption kinetics, there is no internal mass-transfer limitation at a bulk concentration of oxygen of 3.7 mg/L. After the reactor was inoculated, it operated continuously for the whole period of the study, which was over three months. The macrokinetics in the bioreactor were also stable for this period of time. On January 30, at a phenol outlet concentration of 2.4 mg/L, the reaction rate was 108 mg/(L h), and almost two months later, the phenol outlet concentration was 3.3 mg/L and the reaction rate was 105 mg/(L h). Conclusions It has been shown that phenol biodegradation in the inverse fluidized-bed reactor was very stable because of the efficient control of the biofilm thickness. The bioreactor was run continuously for over three months, and the rate of biodegradation remained similar for similar operating conditions. For the range of phenol inlet concentrations studied, it was determined that there was no oxygen limitation. The maximum reported value for the biological degradation of phenol determined in this study was 180 mg/ (l h). This rate can be increased further by optimization of process conditions, such as hydrodynamics and retention time distribution. At higher flow rates, phenol was not completely degraded. Because the required surface water content of phenol is around 0.1 mg/L 2, either the residence time needs to be increased in the reactor or a tank-in-series design needs to be used to meet the guidelines. Nomenclature B ) constant, B ) D(S0 - KS) - µm(Xf/Y + S0), mg/(L h) D ) dilution rate, 1/h
Literature Cited (1) Plumb, R. H. J. Disposal Site Monitoring Data: Observations and Strategy Implications. In Proceedings of the 2nd Canadian/American Conference on Hydrogeology; Hitchon, B., Trudell, M., Eds.; National Water Well Association: Dublin, Ohio, 1985. (2) Tang, W.-T.; Fan, L.-S. Steady-State Phenol Degradation in a Draft-Tube, Gas-Liquid-Solid Fluidized-Bed Bioreactor. AIChE J. 1987, 33, 239. (3) Livingston, A. G.; Chase, H. A. Development of a Phenol Degrading Fluidized Bed Bioreactor for Constant Biomass Holdup. Chem. Eng. J. Biochem. Eng. J. 1991, 45, B35. (4) Donaldson, T. L.; Strandberg, G. W.; Hewitt, J. D. Biooxidation of Coal Gasification Wastewaters Using Fluidized-Bed Bioreactor. Environ. Prog. 1987, 6, 205. (5) Fan, L.-S. Gas-Liquid-Solid Fluidization Engineering; Butterworth: Toronto, Canada, 1989. (6) Nikolov, L.; Karamanev, D. Experimental Study of the Inverse Fluidized Bed Biofilm Reactor. Can. J. Chem. Eng. 1987, 65, 214. (7) Karamanev, D. G.; Nikolov, L. N. Application of Inverse Fluidization to Wastewater Treatment: From Laboratory to FullScale Bioreactors. Environ. Prog. 1996, 15, 194. (8) Karamanev, D. G.; Samson, R. Model of the Biofilm Structure of Thiobacillus ferrooxidans. J. Biotechnol. 1991, 20, 51. (9) Karamanev, D. G.; Chavarie, C.; Samson, R. Soil Immobilization: New Concept for Biotreatment of Soil Contaminants. Biotechnol. Bioeng. 1998, 57, 471. (10) APHA. Standard Methods for the Examination of Water and Wastewater; American Public Health Association: Washington, D.C., 1990. (11) Mueller, J. Disintegration as a Key Sep in Sewage Sludge Treatment. Water Sci. Technol. 2000, 41, 123. (12) Lee, D. D.; Scott, C. D.; Hancher, C. W. Fluidized-Bed Bioreactor for Coal-Conversion Effluents. Water Pollut. Control Fed. J. 1979, 51, 974. (13) Chavarie, C.; Karamanev, D. Use of Inverse Fluidization in Biofilm Reactors. In First International Conference on Bioreactor Fluid Dynamics; BHRA: Cranfield, U.K., 1986.
Received for review January 11, 2001 Revised manuscript received June 19, 2001 Accepted July 7, 2001 IE010036F
