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Effect of turbulence on nitrifying biofilms at non-limiting substrate conditions
War. Res. Vol. 26. No. 12. pp. 1629-1638. 1992 Printed in Great Bntain. All rights reserved
0043-135492 $5.00+0.00 Copyright ,~ 1992PergamonPreu Ltd
EFFECT OF TURBULENCE ON NITRIFYING BIOFILMS AT NON-LIMITING SUBSTRATE CONDITIONS S . KUGAPRASATHAM*, H . N A G A O K A ~ " a n d S . OHGAKI ~ Department of Urban Engineering,The Universityof Tokyo, 7-3-I Hongo, Bunkyo-ku, Tokyo 113, Japan
(First received May 1991; accepted in revised form May 1992) Abstract--The effect of turbulence on nitrifying biofilms was studied in five cylindrical PVC (polyvinyl chloride) reactors, each having ten biofilm sampling taps, over a period of 196 days. Bulk water in the reactors was stirred by paddles at 32, 92, 140, 278 and 500 rpm and the turbulent intensities measured at 10ram from the wall were 0.6, 1.5. 2.6, 4.4 and 8.9cm/s. Biofilms appeared as isolated colonies and continued to grow as filament-type biofilms. Higher turbulence resulted in higher NH4-N flux and higher areal biomass density. Turbulent diffusion of substrates and by-products in the vicinity of filament-type biofilms must have resulted in the above phenomena. Photographic observation of the biofilm surfaces on sampling taps showed uniform biofilm filaments at higher turbulent intensities and large variation in the height of filaments at low turbulent intensities. Substrate flux and biofilm structure (areal density, filament height and cross-sectional area of filament) are inter-related parameters and are strongly affected by turbulence near the biofilm. Substrate flux is expressed as a power function of turbulent intensity, volumetric density and substrate concentration for filament-type biofilm when substrates are non-limiting. Key wordviturbulent intensity, non-limitingsubstrates, nitrifying biofilm, biofilm structure, filament-type biofilm, turbulent diffusion, substrate flux
NONIENCLATURE ,4 = cross-sectional area of filament-type biofilm (L 2) C0 = bulk substrate concentration (ML -~) Df = diffusion coefficient in biofilm (L2T - i) Kf = turbulent diffusion coefficient (L~'r - i) k~ = zero.order reaction constant (ML-3T -t) /,.f = height of filament-type biofilm (L) / = average distance between filaments (L) If = characteristic dimension of filament-type biofilm (L) n = filament biofilm density (L-') ro = substrate flux into the biofilm (ML-~T -~) T = sampling time ( T ) u = momentary velocity (LT-') ~--time-averaged velocity (LT -I) = turbulent intensity near biofilm (LT-') X = areal density of biomass (ML -2) z = depth of the biofi]m (L) ct, = constant • 2 = constant Pb = density of biomass (ML -~) X/Lf is referred to as the volumetric density of the biofilm. INTRODUCTION Wastewater and water treatment systems employing biofilm processes are increasing recently due to better process stability of biofilm processes. Many different biofilms are used in these processes at different hyPresent addresses: *Overseas Services Division, Nihon Suido Consultants Co. Ltd, 2-2-60kubo, Shinjuku-ku, Tokyo 169, .Japan and tDepartment of Civil Engineering, Musashi Institute of Technology, 1-28-1 Tamatsuzumi, Setagaya-ku, Tokyo 158. Japan.
draulic conditions. Variation in the physical structure of the biofilm, such as filament-type or colony-type, were observed with hydraulic conditions (Bryers and Characklis, 1981). Picologlou et ai. (1981) observed filament-type heterotrophic biofilms showing viscoelastic movements in the flowing liquid and attributed these movements to energy dissipation. Fluttering of nitrifying biofilms grown under turbulent flow conditions and an increase in the biofilm activity with turbulence were reported previously (Nagaoka and Ohgaki, 1988). Thus, the environment under which a biofilm grows, such as turbulence and substrate affected the biofilm. Growth of the biofilm is a very complex process in which interaction of hydrodynamic and biochemical factors occurs. Turbulence and substrate concentration in the vicinity of the biofilm are external factors affecting the substrate transfer into biofilm. Growth of biomass occurs by utilizing this substrate transferred inside the biofilm. Thus, the amount of biofilm mass and its structure are the result of turbulence and substrate concentration. Investigation into the effect of turbulence on biofilms is important for the understanding of biofilms. The effect of turbulence on nitrifying biofilms at low N H , - N concentrations were reported previously (Kugaprasatham et al., 1991). The objective of this study was to investigate the effect of turbulence on nitrifying biofilms at nonlimiting substrate conditions. Nitrifying biofilms were grown simultaneously in five reactors at different
1629
S. KUGAPRASATHAM el al.
1630
Effluent port / Blofllm samplingport
= 2~
/
r''l -
7-
f-
¢b -i o n-
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$15.4
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t
(b) Position of biofilm sampling port
(a) Reactor and paddies BIoIII~,,,,, 1 ~
Rubber
5
All dimensions In cm
,
t
0 ]
,
v Tap (~2 f
~ ~
_ Reactor will
plln
(d) Biofilm sampling port
(c) Paddle
Fig. I. Rough-walled hard PVC reactors with biofilm sampling ports and paddles. turbulence near the surface by stirring the bulk water in the reactors at different rotational speeds. Biofilm structures (areal density a n d variation o f the height o f filaments) were observed a n d N H , - N fluxes were determined at specific time intervals. T h e results were analyzed to relate the biofilm structure to turbulence a n d the substrate c o n c e n t r a t i o n . EXPERIMENTAL DESIGN Reactors
Figure 1 shows the details of the reactors and paddles used in the experiments. Reactors were made of dark PVC (diameter 4) 15.4 cm and height 24.5 cm). The vertical walls of the five dark PVC reactors were roughened with sand paper No. 60 up to a height of 16.5cm and each of them had l0 ports with closely fitting taps for biofiim sampling as shown in Fig. I. The experimental set-up is shown in Fig. 2
and the paddles were rotated at 32, 92, 140, 278 and 500 rpm.
Measurement of hydraulic conditions Hydraulic measurements were made without biofilm and with only tapwater flowing through the reactor at experimental conditions. Fouling of the hot-film anemometer sensor precluded the velocity measurements when biofilm was grown on the wall as the setting up of the sensor inside the reactor takes time. Temperature of the water was kept at 25"C. A platinum tip hot-film sensor (Mode 1231W, KANOMAX) was used at constant temperature with a Wbeastone bridge/amplifier and the analog data were converted to digital data for storage in diskettes by an A/D converter interface with a personal computer (NEC, PC8801). Sampling frequency was t00Hz and sampling time was 10.3 s. The number of velocity data for each sampling was 1024. A sensor was placed facing the main direction of flow at 10 mm from the wall of the reactor.
Pure oxygen (",- 0.5 1/ min at 20 cm Hg)
Dechlorinated tapwater Substrate (additional)
Substrate
Effluent L'/",
Fig. 2. Experimental set-up.
J/
Constant temperature bath 25 °C
1631
Effect of turbulence on nitrifying biofilms Table I. Average velocity and turbulent intensity measured at different depths and at 10 mm from the wall in roughened cylindrical hard PVC reactors Paddle Rotational Velocity Turbulent width (cm) speed (rpm) (cm/s) intensity (cm;'s) 5 500 59. I 8.9 5 278 34.0 4.4 5
140
17,7
2.6
5 5
92 32
8.8 2.4
1.5 0.6
Time-series velocity data were analyzed to obtain average
velocity and turbulent intensity. Turbulent intensity
Winkler methods as in Water Examination Methods (Japan Water Works Association, 1988).
Estimation of biomass Nitrifying biofilms found at the initial stages of growth in the small reactors were too small to be measured by gravimetric methods. As an alternative to gravimetric methods the turbidity of the homogenized suspension of biomass in a known volume of distilled water was measured. The measured turbidity was related to the suspended solids concentration by a calibration curve using known concentrations of stock biomass and their turbidity values. Turbidity was measured after homogenizing the biomass by a sonicator at 70 W for I min.
Photograph was calculated as follows, /
£r
=/(i/T)jo
\o5
(U --t~)2dr)
(I)
where u is momentary velocity ( L T - I ) , 6 is time-average velocity ( L T -I) and T is the sampling time (T). Average velocity and turbulent intensity are shown in Table I. These
velocities and turbulent intensities are in the range of velocity and turbulent intensities measured for submerged honeycomb units made of smooth PVC. which are used in actual wastewater treatment systems (Kugaprasatham. 1990).
The biofilm sampling taps were removed carefully from the reactors. The biofilm sampling taps were kept in water and were photographed with a camera (with a bellows frame and close-up lens). Since the biofilm was made from individual filaments, biofilm on the tap was removed with a spatula leaving a thin biofilm strip of about 2-3 mm strip width on the tap. The front and side elevation of this biofilm-strip was also photographed. By enlarging the photograph a magnification of about 15 could be obtained. Variations in biofilm filament height were traced with a digitizer from the side elevation of the thin biofilm strip and the average thickness was obtained.
Reactor operation The experiments were conducted such that the hydraulic condition or the rotational speed ofeach reactor throughout the experimental period was kept constant for each reactor. Substrates NH4-N and DO (dissolved oxygen) concentration were kept high in each reactor so that substrates were non-limiting, especially when the biofilms were thin and fully-penetrated. Reactors were operated at a fill and draw condition for 59 days followed by continuous feeding for 137 days. During the fill and draw condition, suspended nitrifying biomass, from a stock culture initially added as a seed, and suspended biomass concentrations in the reactors were between 2 and 5 mg/I. Reactors were stopped for I h once in 2 days and 3 I. of supernatants from each reactor were removed. Substrate was added to each reactor and mixing was continued. Whenever the suspended biomass concentration in any reactor increased above 5 rag/I, the mixed suspension was removed from that reactor just before settling. Constituents of the substrate for stock culture and for the reactors are as shown in Table 2 and diluted with tap water to the desired NH4-N concentration. Reactors were kept in a water bath at 25~C. Biofilm samples from the sampling taps showed considerable attachment of biomass on all reactors at the end of the fill and draw period. Thus, continuous feeding of substrate was commenced thereafter at an HRT of about 3 h, as shown in Fig. 2. lnfluent NH4-N concentration was 15-40 mg/I. Feeding of additional substrate and saturation of the influent with pure oxygen were commenced for the 500 rpm reactor when the concentrations of NH4-N and DO were found to be lower at 24 and 51 days after the commencement of the continuous feeding. During continuous operation, biofilm could be seen by the naked eye. Thus, biofilm samples were photographed and their masses were also measured. NH4-N flux was calculated from the measurement of influent and effluent concentration, the removal rate of NH4-N by suspended biomass and the flow rate. The removal rate of NH,-N by suspended biomass for each reactor was determined by incubating .500 ml of reactor water collected during sampling over 6-8 h. DO concentrations were also monitored. NH4-N and DO concentrations were measured according to the indophenol and
THEORETICAL DERIVATION T h e main objective o f this derivation is to describe the interaction between turbulence, substrate transfer, substrate c o n c e n t r a t i o n a n d biofiim structure for filament-type biofilms. Biofilms having filamentshaped mass which are observed to flutter with a t u r b u l e n t flow past them are defined as filament-type biofilms. For filament-type biofilms, the biofilm structure is described mathematically. T h e relationship between biofilm structure a n d t u r b u l e n t diffusion between the filaments is assumed. T h e resulting e q u a t i o n s o n biofilm structure and t u r b u l e n t diffusion are c o m b i n e d with a mass conservation e q u a t i o n o f substrate for biofilms under non-limiting substrate conditions. C o n s i d e r a filament-type biofilm as shown in microscopic scale in Fig. 3. F i l a m e n t dimensions arc height, L n , cross-sectional area, A ~, a n d the distances between adjacent filaments is /~.2 etc. Assume a n average height o f filament, Lf, a n d its cross-sectional area, A. Areal density of biomass can be related to filament density as follows,
X = pbLfAn
(2)
where X is the areal density o f the biomass (ML-2), Pb is the density o f the biomass ( M L -3) a n d n is the Table 2. Constituents of substrate for nitrifying biomass each in m8/l
Substrate I NH 4CI CaCI 2 MgSO 4
17,200 1560 1280
Substrate 2 K zHPO4 KH zPO4 Na 2HPO 4
3190 4980
NaHCO~
56200
960
Note: Substrate I and substrate 2 were pumped at equal rates to the reactors.
1632
S. KUGAPItASATHAMet al.
(a) Isometric view
Filaments
,2
/
where ,,: is a constant. Substituting in equation (5) for
tf, -r2
D r = x/u'Z~,: !.
(7)
For a partly-penetrated biofilm under zero-order kinetics of biofilm, substrate flux into the biofilm was shown to be (Harremoes, 1978),
,. = ~
.
is)
Thus, for uniform turbulent diffusion in a filamenttype biofilm, Dr of equation (7) is substituted for D in equation (8), r. = ~/ 2 ~ / u ~ , . l k o f C , .
Biofilsurface mattachment
Substituting equation (4) for ! in equation (9) the following expression relating substrate flux with turbulent intensity, biofilm structure (Lr, X and A), substrate concentration and turbulent intensity is obtained:
(b)Plan Filaments A, ~ , ~ ~ L A =
r,/(( Lt/ X)°z'C ° s) m (Phil A )0'25(2":ka) °'s (~.')o.5.
Fig. 3. Conceptual diagram of a filament-type biofilm. filament density (L-Z). Filament density and the distance between adjacent filaments could be correlated by the following expression, n = =,/! 2
(3)
where I is the average distance between filaments (L) and =, is a constant. Substituting equation (3) for n in equation (2), X = Pb LrAa,/12. (4) Fluttering of filaments due to fluid flow increases the mass transfer. The turbulent diffusion co-efficient for fluttering biofilms with the assumption of uniform diffusivity was defined by Nagaoka et al. 0988) as follows.
where
is the turbulent intensity near filaments and If is the characteristic length of the biofilm. For biofilms grown under uniform turbulence conditions lr is the average characteristic length of the biofilm. Turbulent mass transfer in between filaments depends on the distance between adjacent filaments. Assume a linear relationship between the average distance between filaments, I and If, as follows, ~=~:1
(9)
(6)
(10)
The parameter X / L r, which is a measure of the porousness of a biofilm, affects the substrate flux r, and is affected by the turbulence. Substrate concentration also affects the flux. Due to this inter-relationship between the above parameters the variation in X / L f and C,, the flux should be standardized in order to evaluate the effect of turbulence on substrate flux as shown in the right hand side of equation (10). The values of,Q, "2, ka and A are assumed to be constant. RESULTS AND DISCUSSION
N H , - N flux
Figure 4 shows the variation of flux during continuous feeding for five reactors. The NH4-N flux increased in all reactors with time until about 70 days. After 70 days fluxes could be considered as steady but showed some fluctuations, which were especially high in the reactors stirred at turbulence intensities of 2.6 and 1.5 cm/s. The flux of the reactor mixed at higher turbulence was higher than that of the reactor mixed at lower turbulence. Between 80 and 100 days the reactor mixed at 2.6 cm/s showed a decreased flux and was less than that mixed at 1.5cm/s. This decrease also corresponded with a decrease in the areal biomass density measured at 95 days as shown in Fig. 5. Biomass and biofilm structure Biomass. Figure 5 shows the variation of biomass density during both the fill and draw mode operation and continuous feeding mode operation. During the fill and draw period, attached biomass density was not much different for the reactors stirred at 8.9 and 4.4 cm/s. All the other reactors also showed similar values but were smaller than that of the reactor mixed at 8.9cm/s. During the continuous feeding mode rapid growth of biomass was observed after 22 days.
Effect of turbulence on nitrifying biofilms Turbulent intensity (cm / s)
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I 20
I 40
I 60
I 80
I 100
I 120
I 140
Time (d) Fig. 4. Variation of NH+-N flux during continuous feeding for nitrifying biofilms grown on roughened cylindrical PVC walls. Specific growth rate calculated using the biomass density values between 13 and 95 days did not show any difference between each reactor and was 0.03 d - ' . The growth rate of 0.03 d -I is very low when compared to the values reported in the literature for
nitrifying bacteria (0.3-3d-'). The reason for this may be the loss of biomass due to biofilm shearing. Biomass density in the reactors stirred at higher turbulence was higher than that of the reactors stirred at lower turbulence with only a few exceptions, e.g. 95 days for a reactor stirred at 2.6 cm/s, 134 and 138 days for a reactor stirred at 1.5 cm/s. Even though the Ntt4-N fluxes reached a steady state around 70 days biomass continued to increase and may be closer to the steady state at the very end of the operation.
Biofilm structure. Biofilms initially appeared as small colonies in all reactors and were located densely in the reactors stirred at higher turbulence compared to those mixed at lower turbulence. The number of colonies and their height increased with time. Observation 32 days after continuous feeding showed that biofilm did not cover the entire surface area of the biofilm sampling taps. At higher turbulences (8.9, 4.4 and 2.6 cm/s) large filaments were observed while at lower turbulences (I.5 and 0.6 cm/s) biofilm colonies were still observed. Biofilm surfaces were not smooth and this caused turbulcnt diffusion to occur in the spaces between colonies or filaments. When biofilms grew further, the surface and the structure of the biofilm changed. Individual filaments and spaces in
12
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Continuous feeding
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--
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:'
7_-.3"
I 0
I 20
I 40
0 I 59
Time after seeding (days)
50
1O0
150
Time after continuous feeding (days)
Fig. 5. Variation of the areal density of the biofilm mass for nitrifying biofilms grown on roughened cylindrical PVC walls.
S. K U G ^ P ~ T H A M
1634
A E ::L u) (n C:
et
oi.
4000
2000
E 0
<
3o
-~
t~
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®
NH4-N (103 d) -
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E
Z
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o 0
400
200
c c o
"~ c o 0
-
!
z
0
1
I
I
I
2
4
6
8
10
Turbulent intensity (cm / s) Fig. 7. Variation of the average flux at a steady state, typical concentration of substrates in the reactor and biofilm thickness with turbulent intensity for nitrifying biofilms grown on roughened cylindrical PVC walls.
between the filaments could be seen. The surface showed many ridges and valleys because of the filaments and the spaces between them. Due to high velocity and turbulence filaments seemed to be aligned in a perpendicular direction. The thickness and width of the filaments are about 100-200 ;am and the space between them seems to be less than 100 pm. Figure 6(a) and (b) shows the side elevation of a thin strip of biofilms from the reactors stirred at the turbulent intensities of 8.9 and 1.5 cm/s 95 days after continuous feeding. At high turbulence the variations in the height of the filaments were small and the side elevation appeared as a smooth line. With decreasing turbulence the variation in the height of the filaments was high. The side elevation at 1.5 cm/s shows a filament-type structure on top and a continuous biofilm mass near the attachment surface. However, the photos of the front elevations showed that most of the filaments were independent of each other. Some filaments appeared to be wide due to many filaments being very close to each other. These bigger filaments most probably attached at the very beginning of the operation compared to the smaller ones. Visual examination of similar photographs also showed that the wide filaments are distributed randomly. Since the nitrifying organisms are slow growing a difference could be seen between the ones that attached earlier than the others. This difference could only be seen clearly at the beginning of the operation, in the reactors stirred at high turbulence. because of the increasing density of the filaments. Thus turbulence increased the initial attachment from
random coverage of the surface to a more uniform coverage. An increased number of filaments also increased the surface area available for reaction. However increased numbers of filaments decreases the distance between the filaments and thus decreases the turbulent diffusion coefficient Df [equation (5)]. Figure 7 shows the effect of turbulent intensity on the biofilm thickness (at the latter part of the operation) and the NH4-N flux (averaged value between 70 and 137 days). NH4-N and DO concentrations in the reactors were also plotted in the figure. Average NH4-N flux and biofilm thickness increased with increasing turbulent intensity. Volumetric density of the biofilm (ML -J). This was calculated by dividing the areal density of hiofilm (X) with the average height of the filaments (Lf). Figure 8 shows the variation of the volumetric density of the biofilm with turbulent intensity at different times of operation. Volumetric density showed a slightly increasing trend with increasing turbulence. The exception was with reactor mixed at 0.6cm/s at 111 and 134 days. It is possible that with increasing turbulence a more compact structure could have resulted. The range of turbulence in this experiment may be too narrow to show such a difference. Biofilm experimental methods have yet to be developed to accurately determine the volumetric density of biofilms.
Relationship between NH4-N flux, biofllm structure, NH4-N Concentration and turbulent intensity. Experimental results showed that the NH4-N flux increased with
a) Turbulent intensity 8.9 cm/s
0
5
10ram
b) Turbulent intensity 1.5 cmls
Fig. 6. Photographs of the side-view of nitrifying biofilms grown on rough cylindrical PVC walls 95 days after continuous feeding.
1635
Effect of turbulence on nitrifying biofilms 100
E .~ .o ~-" .OE 0.~..
>,_
Time (days) o 45 o 66 & 95 • 111 t 134
80
~ o
60 •
0"
"%
.
,0
-.
~
._o ~ 40
>0 I
I
I
'1
I
2
4
6
8
10
Turbulent intensity (cm / s) Fig. 8. Variation of volumetric density with turbulent intensity during continuous feeding.
time and reached the steady state around 70 days (Fig. 4). The areal density of the biomass was increased in all the reactors (Fig. 5) and might have reached steady state at the end of the experiment. NH4-N concentrations in the reactors were 6.7-24.6 mg/I. For Monod kinetics for nitrification the half-saturation constant is about I mg/I. If NH4N has fully penetrated the biofilm the ratio of flux/areal density of the biofilm should bc constant and equal to the zero-order reaction constant. Here the part of the biofilm closer to the wall must be at concentrations lower than I mg/I and be at first-order kinetics but this is neglected. Here the penetration of substrate means the penetration in between filaments and into the filaments. Figure 9 shows the variation of the ratio of flux/areal density for each reactor. Except at a turbulence of 1.5 cm/s the ratio showed a peak at 22 days but decreased with time after that. At the initial stages the filaments were far apart thus the ratio depends on the penetration of substrate into the filaments. During these periods coverage of the surface with new filaments occurs. Substrate penetration into new filaments (thin) must be higher
~ 0.02
n o • A •
~
~ E 0.01
Z "" E
A &
0.0
o x _
o
_
"-- E
o~ •o .9 ''6
8.9 4.4 2•6 1•5
~-..; ~..~.
zT
than that of mature filaments• The increase in the ratio up to 22 days must be the result of many new filaments• However, after 22 days the areal density increased more and the distance between the filaments also decreased. Since the distance between filaments decreased the penetration or the diffusivity decreased with time resulting in the decrease of the ratio. Thus the biofilms during 45-134 days can be assumed to be partly penetrated. Further, neglecting the effect of first-order kinetics at the bottom part of the biofilm when the NH4-N concentration decreases, zero-order kinetics can be assumed. Though the DO concentrations were in the range of 3.7-5.7mg/!, which was lower than the N H , - N concentrations, there was a marked difference in the NH4-N concentrations in the reactors as shown in Fig. 7. Thus, NH4-N concentrations were substituted in equation (10) for C, instead of DO concentrations in the following analysis. The interaction of turbulence, substrate flux, binfilm structure and substrate concentration is very complex, A simpler expression [equation (10)] derived under the assumption of zero-order kinetics and a partly-penetrated biofilm was applied to the experimental data. The left-hand side of equation (I0) was calculated from experimental data and Fig. 10 shows the relationship between turbulent intensity and the value of left-hand side of equation (10) in a log-log scale. Even with many assumptions a good correlation with turbulent intensity can be seen. The slope is 0.69, which is the power of turbulent intensity. The =~, =2 and zero-order reaction constants of all the reactors need to be same for the power to be 0.5 as in equation (10). However, experimental values of zero-order reaction constants could be different between reactors. The constants ~,j, =2 may also vary with turbulent intensity. Further, the turbulent diffusion model assumes a constant diffusivity inside the filaments. However, turbulence should die down as the wall is approached. Thus, diffusivity between filaments decreases and a representative value needs to be used. The relationship between the turbulence measured without the biofilm and that inside biofilm was assumed to be direct.
Turbulent Intensity(cm I s)
,~.'0 x
1637
0
I
I
50
1 O0
A
,,]
J t50
Time (d)
Fig. 9. Variation of the ratio NH4-N flux/areal density of the biomass during continuous operation for nitrifying biofilms grown on rough cylindrical PVC walls.
0.1
1
lO
~/u '2 (cm / s)
Fig. 10. Relationship between r,/((Lr/X)°25C°5) and turbulent intensity.
1638
S. KUG^F'IUL~TI4AMet al.
From the actual observations, lower turbulent intensities showed a large width of filaments (possibly many filaments being held together), compared to those of a higher turbulence. The effects of difference in the biofilm configuration (the width of the filaments or the distance between the filaments) on the constants are not known. The above analysis showed that substrate flux and biofilm structure (areal density, filament height and cross-sectional area of filament) are interrelated parameters and are strongly affected by turbulence near the biofilm. Substrate flux is expressed as a power function of turbulent intensity, volumetric density and substrate concentration for a filamenttype biofiim when substrates are non-limiting. SUMMARY AND CONCLUSIONS Hydraulic conditions measured without biofilm were used to study the effect of hydraulic conditions on biofilms. The effects on substrate flux, biomass density and biofilm structure using nitrifying biofilms at non-limiting substrate conditions were studied. Based on the experimental results and analysis the following conclusions are made. Increase in turbulence increased biomass and substrate flux within the range of experimental conditions used when substrates were not limiting. During the initial period of attachment small colonies could be observed randomly at any turbulence. However, biofiim structure was different according to turbulence with the growth of biofilm. Biofilm structure at higher turbulence consisted of uniform filaments covering the entire surface. At lower
turbulent intensities the filaments showed large variation in their height. The volumetric density of the biofilm showed a slight increase with increasing turbulence. An expression was [equation (10)] derived for substrate flux, using turbulent diffusion in between filaments for partly-penetrated filament-type biofilms under zero-order reaction kinetics, relating substrate concentration, turbulent intensity and biofilm structure (volumetric density of biofilm, filament crosssectional area). Experimental results for nitrifying biofilms grown on roughened PVC reactors correlated well with this expression. REFERENCES
Bryers J. and Characklis W. G. (1981) Early fouling biofilm formation in a turbulent flow system; overall kinetics. ;Vat. Res. 15, 483-491. Harremos P. (1978) Biofilm kinetics. In Water Pollution Microbiology: Vol. 2 (Edited by Mitchell R.). Wiley, New York. Japan Water Works Association (1985) Water Examination Methods (in Japanese). Kugaprasatham S. (1990) Effect of hydraulic conditions on biofilms: a Dissertation, Department of Urban Engineering, The University of Tokyo, Japan. Kugaprasatham S., Nagaoka H. and Ohgaki S. (1991) Effect of short-term and long-term changes in hydraulic conditions on nitrifying biofilms. War. Sci. Technol. 23, 1487-1494. Nagaoka H. and Ohgaki S. (1988) Effect of turbulence on biofilm activity. In Water Pollution Control in Asia (Edited by Pansawad T. et al.), pp. 155-161. Pergamon Press, Oxford. Picologlou B. F., Zelver N. And Characklis W. O. (1980) Biofilm growth and hydraulic performance. J. Hydraul. Div., Am. Soc. civ. Engrs 106, 733-746.
0043-135492 $5.00+0.00 Copyright ,~ 1992PergamonPreu Ltd
EFFECT OF TURBULENCE ON NITRIFYING BIOFILMS AT NON-LIMITING SUBSTRATE CONDITIONS S . KUGAPRASATHAM*, H . N A G A O K A ~ " a n d S . OHGAKI ~ Department of Urban Engineering,The Universityof Tokyo, 7-3-I Hongo, Bunkyo-ku, Tokyo 113, Japan
(First received May 1991; accepted in revised form May 1992) Abstract--The effect of turbulence on nitrifying biofilms was studied in five cylindrical PVC (polyvinyl chloride) reactors, each having ten biofilm sampling taps, over a period of 196 days. Bulk water in the reactors was stirred by paddles at 32, 92, 140, 278 and 500 rpm and the turbulent intensities measured at 10ram from the wall were 0.6, 1.5. 2.6, 4.4 and 8.9cm/s. Biofilms appeared as isolated colonies and continued to grow as filament-type biofilms. Higher turbulence resulted in higher NH4-N flux and higher areal biomass density. Turbulent diffusion of substrates and by-products in the vicinity of filament-type biofilms must have resulted in the above phenomena. Photographic observation of the biofilm surfaces on sampling taps showed uniform biofilm filaments at higher turbulent intensities and large variation in the height of filaments at low turbulent intensities. Substrate flux and biofilm structure (areal density, filament height and cross-sectional area of filament) are inter-related parameters and are strongly affected by turbulence near the biofilm. Substrate flux is expressed as a power function of turbulent intensity, volumetric density and substrate concentration for filament-type biofilm when substrates are non-limiting. Key wordviturbulent intensity, non-limitingsubstrates, nitrifying biofilm, biofilm structure, filament-type biofilm, turbulent diffusion, substrate flux
NONIENCLATURE ,4 = cross-sectional area of filament-type biofilm (L 2) C0 = bulk substrate concentration (ML -~) Df = diffusion coefficient in biofilm (L2T - i) Kf = turbulent diffusion coefficient (L~'r - i) k~ = zero.order reaction constant (ML-3T -t) /,.f = height of filament-type biofilm (L) / = average distance between filaments (L) If = characteristic dimension of filament-type biofilm (L) n = filament biofilm density (L-') ro = substrate flux into the biofilm (ML-~T -~) T = sampling time ( T ) u = momentary velocity (LT-') ~--time-averaged velocity (LT -I) = turbulent intensity near biofilm (LT-') X = areal density of biomass (ML -2) z = depth of the biofi]m (L) ct, = constant • 2 = constant Pb = density of biomass (ML -~) X/Lf is referred to as the volumetric density of the biofilm. INTRODUCTION Wastewater and water treatment systems employing biofilm processes are increasing recently due to better process stability of biofilm processes. Many different biofilms are used in these processes at different hyPresent addresses: *Overseas Services Division, Nihon Suido Consultants Co. Ltd, 2-2-60kubo, Shinjuku-ku, Tokyo 169, .Japan and tDepartment of Civil Engineering, Musashi Institute of Technology, 1-28-1 Tamatsuzumi, Setagaya-ku, Tokyo 158. Japan.
draulic conditions. Variation in the physical structure of the biofilm, such as filament-type or colony-type, were observed with hydraulic conditions (Bryers and Characklis, 1981). Picologlou et ai. (1981) observed filament-type heterotrophic biofilms showing viscoelastic movements in the flowing liquid and attributed these movements to energy dissipation. Fluttering of nitrifying biofilms grown under turbulent flow conditions and an increase in the biofilm activity with turbulence were reported previously (Nagaoka and Ohgaki, 1988). Thus, the environment under which a biofilm grows, such as turbulence and substrate affected the biofilm. Growth of the biofilm is a very complex process in which interaction of hydrodynamic and biochemical factors occurs. Turbulence and substrate concentration in the vicinity of the biofilm are external factors affecting the substrate transfer into biofilm. Growth of biomass occurs by utilizing this substrate transferred inside the biofilm. Thus, the amount of biofilm mass and its structure are the result of turbulence and substrate concentration. Investigation into the effect of turbulence on biofilms is important for the understanding of biofilms. The effect of turbulence on nitrifying biofilms at low N H , - N concentrations were reported previously (Kugaprasatham et al., 1991). The objective of this study was to investigate the effect of turbulence on nitrifying biofilms at nonlimiting substrate conditions. Nitrifying biofilms were grown simultaneously in five reactors at different
1629
S. KUGAPRASATHAM el al.
1630
Effluent port / Blofllm samplingport
= 2~
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,
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,
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(d) Biofilm sampling port
(c) Paddle
Fig. I. Rough-walled hard PVC reactors with biofilm sampling ports and paddles. turbulence near the surface by stirring the bulk water in the reactors at different rotational speeds. Biofilm structures (areal density a n d variation o f the height o f filaments) were observed a n d N H , - N fluxes were determined at specific time intervals. T h e results were analyzed to relate the biofilm structure to turbulence a n d the substrate c o n c e n t r a t i o n . EXPERIMENTAL DESIGN Reactors
Figure 1 shows the details of the reactors and paddles used in the experiments. Reactors were made of dark PVC (diameter 4) 15.4 cm and height 24.5 cm). The vertical walls of the five dark PVC reactors were roughened with sand paper No. 60 up to a height of 16.5cm and each of them had l0 ports with closely fitting taps for biofiim sampling as shown in Fig. I. The experimental set-up is shown in Fig. 2
and the paddles were rotated at 32, 92, 140, 278 and 500 rpm.
Measurement of hydraulic conditions Hydraulic measurements were made without biofilm and with only tapwater flowing through the reactor at experimental conditions. Fouling of the hot-film anemometer sensor precluded the velocity measurements when biofilm was grown on the wall as the setting up of the sensor inside the reactor takes time. Temperature of the water was kept at 25"C. A platinum tip hot-film sensor (Mode 1231W, KANOMAX) was used at constant temperature with a Wbeastone bridge/amplifier and the analog data were converted to digital data for storage in diskettes by an A/D converter interface with a personal computer (NEC, PC8801). Sampling frequency was t00Hz and sampling time was 10.3 s. The number of velocity data for each sampling was 1024. A sensor was placed facing the main direction of flow at 10 mm from the wall of the reactor.
Pure oxygen (",- 0.5 1/ min at 20 cm Hg)
Dechlorinated tapwater Substrate (additional)
Substrate
Effluent L'/",
Fig. 2. Experimental set-up.
J/
Constant temperature bath 25 °C
1631
Effect of turbulence on nitrifying biofilms Table I. Average velocity and turbulent intensity measured at different depths and at 10 mm from the wall in roughened cylindrical hard PVC reactors Paddle Rotational Velocity Turbulent width (cm) speed (rpm) (cm/s) intensity (cm;'s) 5 500 59. I 8.9 5 278 34.0 4.4 5
140
17,7
2.6
5 5
92 32
8.8 2.4
1.5 0.6
Time-series velocity data were analyzed to obtain average
velocity and turbulent intensity. Turbulent intensity
Winkler methods as in Water Examination Methods (Japan Water Works Association, 1988).
Estimation of biomass Nitrifying biofilms found at the initial stages of growth in the small reactors were too small to be measured by gravimetric methods. As an alternative to gravimetric methods the turbidity of the homogenized suspension of biomass in a known volume of distilled water was measured. The measured turbidity was related to the suspended solids concentration by a calibration curve using known concentrations of stock biomass and their turbidity values. Turbidity was measured after homogenizing the biomass by a sonicator at 70 W for I min.
Photograph was calculated as follows, /
£r
=/(i/T)jo
\o5
(U --t~)2dr)
(I)
where u is momentary velocity ( L T - I ) , 6 is time-average velocity ( L T -I) and T is the sampling time (T). Average velocity and turbulent intensity are shown in Table I. These
velocities and turbulent intensities are in the range of velocity and turbulent intensities measured for submerged honeycomb units made of smooth PVC. which are used in actual wastewater treatment systems (Kugaprasatham. 1990).
The biofilm sampling taps were removed carefully from the reactors. The biofilm sampling taps were kept in water and were photographed with a camera (with a bellows frame and close-up lens). Since the biofilm was made from individual filaments, biofilm on the tap was removed with a spatula leaving a thin biofilm strip of about 2-3 mm strip width on the tap. The front and side elevation of this biofilm-strip was also photographed. By enlarging the photograph a magnification of about 15 could be obtained. Variations in biofilm filament height were traced with a digitizer from the side elevation of the thin biofilm strip and the average thickness was obtained.
Reactor operation The experiments were conducted such that the hydraulic condition or the rotational speed ofeach reactor throughout the experimental period was kept constant for each reactor. Substrates NH4-N and DO (dissolved oxygen) concentration were kept high in each reactor so that substrates were non-limiting, especially when the biofilms were thin and fully-penetrated. Reactors were operated at a fill and draw condition for 59 days followed by continuous feeding for 137 days. During the fill and draw condition, suspended nitrifying biomass, from a stock culture initially added as a seed, and suspended biomass concentrations in the reactors were between 2 and 5 mg/I. Reactors were stopped for I h once in 2 days and 3 I. of supernatants from each reactor were removed. Substrate was added to each reactor and mixing was continued. Whenever the suspended biomass concentration in any reactor increased above 5 rag/I, the mixed suspension was removed from that reactor just before settling. Constituents of the substrate for stock culture and for the reactors are as shown in Table 2 and diluted with tap water to the desired NH4-N concentration. Reactors were kept in a water bath at 25~C. Biofilm samples from the sampling taps showed considerable attachment of biomass on all reactors at the end of the fill and draw period. Thus, continuous feeding of substrate was commenced thereafter at an HRT of about 3 h, as shown in Fig. 2. lnfluent NH4-N concentration was 15-40 mg/I. Feeding of additional substrate and saturation of the influent with pure oxygen were commenced for the 500 rpm reactor when the concentrations of NH4-N and DO were found to be lower at 24 and 51 days after the commencement of the continuous feeding. During continuous operation, biofilm could be seen by the naked eye. Thus, biofilm samples were photographed and their masses were also measured. NH4-N flux was calculated from the measurement of influent and effluent concentration, the removal rate of NH4-N by suspended biomass and the flow rate. The removal rate of NH,-N by suspended biomass for each reactor was determined by incubating .500 ml of reactor water collected during sampling over 6-8 h. DO concentrations were also monitored. NH4-N and DO concentrations were measured according to the indophenol and
THEORETICAL DERIVATION T h e main objective o f this derivation is to describe the interaction between turbulence, substrate transfer, substrate c o n c e n t r a t i o n a n d biofiim structure for filament-type biofilms. Biofilms having filamentshaped mass which are observed to flutter with a t u r b u l e n t flow past them are defined as filament-type biofilms. For filament-type biofilms, the biofilm structure is described mathematically. T h e relationship between biofilm structure a n d t u r b u l e n t diffusion between the filaments is assumed. T h e resulting e q u a t i o n s o n biofilm structure and t u r b u l e n t diffusion are c o m b i n e d with a mass conservation e q u a t i o n o f substrate for biofilms under non-limiting substrate conditions. C o n s i d e r a filament-type biofilm as shown in microscopic scale in Fig. 3. F i l a m e n t dimensions arc height, L n , cross-sectional area, A ~, a n d the distances between adjacent filaments is /~.2 etc. Assume a n average height o f filament, Lf, a n d its cross-sectional area, A. Areal density of biomass can be related to filament density as follows,
X = pbLfAn
(2)
where X is the areal density o f the biomass (ML-2), Pb is the density o f the biomass ( M L -3) a n d n is the Table 2. Constituents of substrate for nitrifying biomass each in m8/l
Substrate I NH 4CI CaCI 2 MgSO 4
17,200 1560 1280
Substrate 2 K zHPO4 KH zPO4 Na 2HPO 4
3190 4980
NaHCO~
56200
960
Note: Substrate I and substrate 2 were pumped at equal rates to the reactors.
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S. KUGAPItASATHAMet al.
(a) Isometric view
Filaments
,2
/
where ,,: is a constant. Substituting in equation (5) for
tf, -r2
D r = x/u'Z~,: !.
(7)
For a partly-penetrated biofilm under zero-order kinetics of biofilm, substrate flux into the biofilm was shown to be (Harremoes, 1978),
,. = ~
.
is)
Thus, for uniform turbulent diffusion in a filamenttype biofilm, Dr of equation (7) is substituted for D in equation (8), r. = ~/ 2 ~ / u ~ , . l k o f C , .
Biofilsurface mattachment
Substituting equation (4) for ! in equation (9) the following expression relating substrate flux with turbulent intensity, biofilm structure (Lr, X and A), substrate concentration and turbulent intensity is obtained:
(b)Plan Filaments A, ~ , ~ ~ L A =
r,/(( Lt/ X)°z'C ° s) m (Phil A )0'25(2":ka) °'s (~.')o.5.
Fig. 3. Conceptual diagram of a filament-type biofilm. filament density (L-Z). Filament density and the distance between adjacent filaments could be correlated by the following expression, n = =,/! 2
(3)
where I is the average distance between filaments (L) and =, is a constant. Substituting equation (3) for n in equation (2), X = Pb LrAa,/12. (4) Fluttering of filaments due to fluid flow increases the mass transfer. The turbulent diffusion co-efficient for fluttering biofilms with the assumption of uniform diffusivity was defined by Nagaoka et al. 0988) as follows.
where
is the turbulent intensity near filaments and If is the characteristic length of the biofilm. For biofilms grown under uniform turbulence conditions lr is the average characteristic length of the biofilm. Turbulent mass transfer in between filaments depends on the distance between adjacent filaments. Assume a linear relationship between the average distance between filaments, I and If, as follows, ~=~:1
(9)
(6)
(10)
The parameter X / L r, which is a measure of the porousness of a biofilm, affects the substrate flux r, and is affected by the turbulence. Substrate concentration also affects the flux. Due to this inter-relationship between the above parameters the variation in X / L f and C,, the flux should be standardized in order to evaluate the effect of turbulence on substrate flux as shown in the right hand side of equation (10). The values of,Q, "2, ka and A are assumed to be constant. RESULTS AND DISCUSSION
N H , - N flux
Figure 4 shows the variation of flux during continuous feeding for five reactors. The NH4-N flux increased in all reactors with time until about 70 days. After 70 days fluxes could be considered as steady but showed some fluctuations, which were especially high in the reactors stirred at turbulence intensities of 2.6 and 1.5 cm/s. The flux of the reactor mixed at higher turbulence was higher than that of the reactor mixed at lower turbulence. Between 80 and 100 days the reactor mixed at 2.6 cm/s showed a decreased flux and was less than that mixed at 1.5cm/s. This decrease also corresponded with a decrease in the areal biomass density measured at 95 days as shown in Fig. 5. Biomass and biofilm structure Biomass. Figure 5 shows the variation of biomass density during both the fill and draw mode operation and continuous feeding mode operation. During the fill and draw period, attached biomass density was not much different for the reactors stirred at 8.9 and 4.4 cm/s. All the other reactors also showed similar values but were smaller than that of the reactor mixed at 8.9cm/s. During the continuous feeding mode rapid growth of biomass was observed after 22 days.
Effect of turbulence on nitrifying biofilms Turbulent intensity (cm / s)
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Time (d) Fig. 4. Variation of NH+-N flux during continuous feeding for nitrifying biofilms grown on roughened cylindrical PVC walls. Specific growth rate calculated using the biomass density values between 13 and 95 days did not show any difference between each reactor and was 0.03 d - ' . The growth rate of 0.03 d -I is very low when compared to the values reported in the literature for
nitrifying bacteria (0.3-3d-'). The reason for this may be the loss of biomass due to biofilm shearing. Biomass density in the reactors stirred at higher turbulence was higher than that of the reactors stirred at lower turbulence with only a few exceptions, e.g. 95 days for a reactor stirred at 2.6 cm/s, 134 and 138 days for a reactor stirred at 1.5 cm/s. Even though the Ntt4-N fluxes reached a steady state around 70 days biomass continued to increase and may be closer to the steady state at the very end of the operation.
Biofilm structure. Biofilms initially appeared as small colonies in all reactors and were located densely in the reactors stirred at higher turbulence compared to those mixed at lower turbulence. The number of colonies and their height increased with time. Observation 32 days after continuous feeding showed that biofilm did not cover the entire surface area of the biofilm sampling taps. At higher turbulences (8.9, 4.4 and 2.6 cm/s) large filaments were observed while at lower turbulences (I.5 and 0.6 cm/s) biofilm colonies were still observed. Biofilm surfaces were not smooth and this caused turbulcnt diffusion to occur in the spaces between colonies or filaments. When biofilms grew further, the surface and the structure of the biofilm changed. Individual filaments and spaces in
12
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:'
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I 0
I 20
I 40
0 I 59
Time after seeding (days)
50
1O0
150
Time after continuous feeding (days)
Fig. 5. Variation of the areal density of the biofilm mass for nitrifying biofilms grown on roughened cylindrical PVC walls.
S. K U G ^ P ~ T H A M
1634
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oi.
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z
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Turbulent intensity (cm / s) Fig. 7. Variation of the average flux at a steady state, typical concentration of substrates in the reactor and biofilm thickness with turbulent intensity for nitrifying biofilms grown on roughened cylindrical PVC walls.
between the filaments could be seen. The surface showed many ridges and valleys because of the filaments and the spaces between them. Due to high velocity and turbulence filaments seemed to be aligned in a perpendicular direction. The thickness and width of the filaments are about 100-200 ;am and the space between them seems to be less than 100 pm. Figure 6(a) and (b) shows the side elevation of a thin strip of biofilms from the reactors stirred at the turbulent intensities of 8.9 and 1.5 cm/s 95 days after continuous feeding. At high turbulence the variations in the height of the filaments were small and the side elevation appeared as a smooth line. With decreasing turbulence the variation in the height of the filaments was high. The side elevation at 1.5 cm/s shows a filament-type structure on top and a continuous biofilm mass near the attachment surface. However, the photos of the front elevations showed that most of the filaments were independent of each other. Some filaments appeared to be wide due to many filaments being very close to each other. These bigger filaments most probably attached at the very beginning of the operation compared to the smaller ones. Visual examination of similar photographs also showed that the wide filaments are distributed randomly. Since the nitrifying organisms are slow growing a difference could be seen between the ones that attached earlier than the others. This difference could only be seen clearly at the beginning of the operation, in the reactors stirred at high turbulence. because of the increasing density of the filaments. Thus turbulence increased the initial attachment from
random coverage of the surface to a more uniform coverage. An increased number of filaments also increased the surface area available for reaction. However increased numbers of filaments decreases the distance between the filaments and thus decreases the turbulent diffusion coefficient Df [equation (5)]. Figure 7 shows the effect of turbulent intensity on the biofilm thickness (at the latter part of the operation) and the NH4-N flux (averaged value between 70 and 137 days). NH4-N and DO concentrations in the reactors were also plotted in the figure. Average NH4-N flux and biofilm thickness increased with increasing turbulent intensity. Volumetric density of the biofilm (ML -J). This was calculated by dividing the areal density of hiofilm (X) with the average height of the filaments (Lf). Figure 8 shows the variation of the volumetric density of the biofilm with turbulent intensity at different times of operation. Volumetric density showed a slightly increasing trend with increasing turbulence. The exception was with reactor mixed at 0.6cm/s at 111 and 134 days. It is possible that with increasing turbulence a more compact structure could have resulted. The range of turbulence in this experiment may be too narrow to show such a difference. Biofilm experimental methods have yet to be developed to accurately determine the volumetric density of biofilms.
Relationship between NH4-N flux, biofllm structure, NH4-N Concentration and turbulent intensity. Experimental results showed that the NH4-N flux increased with
a) Turbulent intensity 8.9 cm/s
0
5
10ram
b) Turbulent intensity 1.5 cmls
Fig. 6. Photographs of the side-view of nitrifying biofilms grown on rough cylindrical PVC walls 95 days after continuous feeding.
1635
Effect of turbulence on nitrifying biofilms 100
E .~ .o ~-" .OE 0.~..
>,_
Time (days) o 45 o 66 & 95 • 111 t 134
80
~ o
60 •
0"
"%
.
,0
-.
~
._o ~ 40
>0 I
I
I
'1
I
2
4
6
8
10
Turbulent intensity (cm / s) Fig. 8. Variation of volumetric density with turbulent intensity during continuous feeding.
time and reached the steady state around 70 days (Fig. 4). The areal density of the biomass was increased in all the reactors (Fig. 5) and might have reached steady state at the end of the experiment. NH4-N concentrations in the reactors were 6.7-24.6 mg/I. For Monod kinetics for nitrification the half-saturation constant is about I mg/I. If NH4N has fully penetrated the biofilm the ratio of flux/areal density of the biofilm should bc constant and equal to the zero-order reaction constant. Here the part of the biofilm closer to the wall must be at concentrations lower than I mg/I and be at first-order kinetics but this is neglected. Here the penetration of substrate means the penetration in between filaments and into the filaments. Figure 9 shows the variation of the ratio of flux/areal density for each reactor. Except at a turbulence of 1.5 cm/s the ratio showed a peak at 22 days but decreased with time after that. At the initial stages the filaments were far apart thus the ratio depends on the penetration of substrate into the filaments. During these periods coverage of the surface with new filaments occurs. Substrate penetration into new filaments (thin) must be higher
~ 0.02
n o • A •
~
~ E 0.01
Z "" E
A &
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o x _
o
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than that of mature filaments• The increase in the ratio up to 22 days must be the result of many new filaments• However, after 22 days the areal density increased more and the distance between the filaments also decreased. Since the distance between filaments decreased the penetration or the diffusivity decreased with time resulting in the decrease of the ratio. Thus the biofilms during 45-134 days can be assumed to be partly penetrated. Further, neglecting the effect of first-order kinetics at the bottom part of the biofilm when the NH4-N concentration decreases, zero-order kinetics can be assumed. Though the DO concentrations were in the range of 3.7-5.7mg/!, which was lower than the N H , - N concentrations, there was a marked difference in the NH4-N concentrations in the reactors as shown in Fig. 7. Thus, NH4-N concentrations were substituted in equation (10) for C, instead of DO concentrations in the following analysis. The interaction of turbulence, substrate flux, binfilm structure and substrate concentration is very complex, A simpler expression [equation (10)] derived under the assumption of zero-order kinetics and a partly-penetrated biofilm was applied to the experimental data. The left-hand side of equation (I0) was calculated from experimental data and Fig. 10 shows the relationship between turbulent intensity and the value of left-hand side of equation (10) in a log-log scale. Even with many assumptions a good correlation with turbulent intensity can be seen. The slope is 0.69, which is the power of turbulent intensity. The =~, =2 and zero-order reaction constants of all the reactors need to be same for the power to be 0.5 as in equation (10). However, experimental values of zero-order reaction constants could be different between reactors. The constants ~,j, =2 may also vary with turbulent intensity. Further, the turbulent diffusion model assumes a constant diffusivity inside the filaments. However, turbulence should die down as the wall is approached. Thus, diffusivity between filaments decreases and a representative value needs to be used. The relationship between the turbulence measured without the biofilm and that inside biofilm was assumed to be direct.
Turbulent Intensity(cm I s)
,~.'0 x
1637
0
I
I
50
1 O0
A
,,]
J t50
Time (d)
Fig. 9. Variation of the ratio NH4-N flux/areal density of the biomass during continuous operation for nitrifying biofilms grown on rough cylindrical PVC walls.
0.1
1
lO
~/u '2 (cm / s)
Fig. 10. Relationship between r,/((Lr/X)°25C°5) and turbulent intensity.
1638
S. KUG^F'IUL~TI4AMet al.
From the actual observations, lower turbulent intensities showed a large width of filaments (possibly many filaments being held together), compared to those of a higher turbulence. The effects of difference in the biofilm configuration (the width of the filaments or the distance between the filaments) on the constants are not known. The above analysis showed that substrate flux and biofilm structure (areal density, filament height and cross-sectional area of filament) are interrelated parameters and are strongly affected by turbulence near the biofilm. Substrate flux is expressed as a power function of turbulent intensity, volumetric density and substrate concentration for a filamenttype biofiim when substrates are non-limiting. SUMMARY AND CONCLUSIONS Hydraulic conditions measured without biofilm were used to study the effect of hydraulic conditions on biofilms. The effects on substrate flux, biomass density and biofilm structure using nitrifying biofilms at non-limiting substrate conditions were studied. Based on the experimental results and analysis the following conclusions are made. Increase in turbulence increased biomass and substrate flux within the range of experimental conditions used when substrates were not limiting. During the initial period of attachment small colonies could be observed randomly at any turbulence. However, biofiim structure was different according to turbulence with the growth of biofilm. Biofilm structure at higher turbulence consisted of uniform filaments covering the entire surface. At lower
turbulent intensities the filaments showed large variation in their height. The volumetric density of the biofilm showed a slight increase with increasing turbulence. An expression was [equation (10)] derived for substrate flux, using turbulent diffusion in between filaments for partly-penetrated filament-type biofilms under zero-order reaction kinetics, relating substrate concentration, turbulent intensity and biofilm structure (volumetric density of biofilm, filament crosssectional area). Experimental results for nitrifying biofilms grown on roughened PVC reactors correlated well with this expression. REFERENCES
Bryers J. and Characklis W. G. (1981) Early fouling biofilm formation in a turbulent flow system; overall kinetics. ;Vat. Res. 15, 483-491. Harremos P. (1978) Biofilm kinetics. In Water Pollution Microbiology: Vol. 2 (Edited by Mitchell R.). Wiley, New York. Japan Water Works Association (1985) Water Examination Methods (in Japanese). Kugaprasatham S. (1990) Effect of hydraulic conditions on biofilms: a Dissertation, Department of Urban Engineering, The University of Tokyo, Japan. Kugaprasatham S., Nagaoka H. and Ohgaki S. (1991) Effect of short-term and long-term changes in hydraulic conditions on nitrifying biofilms. War. Sci. Technol. 23, 1487-1494. Nagaoka H. and Ohgaki S. (1988) Effect of turbulence on biofilm activity. In Water Pollution Control in Asia (Edited by Pansawad T. et al.), pp. 155-161. Pergamon Press, Oxford. Picologlou B. F., Zelver N. And Characklis W. O. (1980) Biofilm growth and hydraulic performance. J. Hydraul. Div., Am. Soc. civ. Engrs 106, 733-746.