Journal of Water Process Engineering 17 (2017) 135142

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Journal of Water Process Engineering

journal homepage: www.elsevier.com/locate/jwpe

Treatment of wastewater by mixed bacterial consortium in continuous MARK

reactors packed with solid waste

Bhaswati Chakraborty , Srabanti Basu
Department of Biotechnology, Heritage Institute, Anandapur, Kolkata 700107, India

A R T I C L E I N F O A B S T R A C T

Keywords: Phenol is one of the priority pollutants as declared by EPA (USA). Very few comparative studies have been
Packed bed reactor carried out with continuous packed bed reactors (PBR) for bacterial degradation of phenol and applying the
Co-current result in real time situation. The present study compared the performance of a co-current and a counter-current
Counter-current continuous PBR in degrading phenol by a mixed consortium of bacteria isolated from the East Calcutta Wetlands
Removal eciency
based on both rate of phenol degradation and phenol removal eciency. Dierent process parameters like air
Phenol
Industrial euent
ow rate, hydraulic retention time and temperature were varied for both the systems. Performance of the co-
current continuous PBR was found to be more ecient. To validate the performance of this system in actual
application in industry a real time phenolic industrial euent was fed to this system. Phenol degradation rate
was 1.43 mg/l min and the phenol removal eciency of this co-current continuous PBR was 72.85%.

1. Introduction tion of phenol chemically leads to formation of toxic intermediates. On

In developing countries like India urbanization and industrialization
go hand in hand. The necessity for prolic industrialization is undeni-
able. Setting up of industry in a region leads to an overall development
of the region primarily by establishment of townships in and around the
industrial belt. The bulk methods for treatment fail to reduce the levels
of specic organic and inorganic pollutants to meet the environmental
regulations. As a result, the surface water gets contaminated with
organic and inorganic pollutants discharged by industrial euents and
due to seepage the ground water is also contaminated. Industries like
petrochemicals, coal coking, coal gasication, tanneries etc discharge
phenolic euent into the environment [1,2]. Ground water contamina-
tion may occur due to leakage in the gaskets and bearings of the
pipelines carrying phenolic wastewater [3]. Phenolic compounds are
aromatic pollutants which are toxic to human health. Exposure to
phenol above the safety limit leads to central nervous system disorder,
myocardial depression, hypothermia, renal failure, hepatitis damage,
gastrointestinal problem and even blindness. Phenol is carcinogenic too
[4]. Phenol is the representative of all phenolic compounds and the
safety limit of phenol concentration in euent is set as 0.168 mg/l by
EPA (USA) [5].
To reduce the phenol concentration present in industrial euent
within safety limit degradation of phenol by both chemical and
biological treatments are prevalent. The chemical treatments like
adsorption, stripping are though fast are expensive. Moreover, degrada-
the other hand, biological treatments are economic, could process a
varied range of phenol concentration and more importantly lead to
complete mineralization of phenol [2,68].
Aerobic and anaerobic bacteria, fungi had been acclimatized to
degrade hydrocarbons and it had been reported that a mixed bacterial
consortium was more ecient in degrading phenol as compared to its
constituent bacteria degrading individually [911]. Majority of the
work done in phenol degradation by biological method considered
batch process [12,10,11] and concentrated primarily on kinetic para-
meter evaluation. Very few works [13,3] studied the continuous mode
of operation in phenol degradation by mixed culture. In this work both
co-current and counter-current continuous phenol biodegradation
processes were studied and compared.
Micro-organisms for biodegradation of phenol were mostly sourced
from industrial sewage water in previous works [14]. The present study
had used soil microbes from a novel site for this purpose. The site was
known as East Calcutta Wetlands (ECW, 2224N-2236N; 8832E,
12,500 ha in area) and was declared as a Ramsar site on 19th August
2002 by Ramsar Convention Bureau. Since 1868 ECW acted as the most
important urban and industrial waste treatment and recycling site of
Kolkata, a major metropolitan city of India. It was the dumping ground
of domestic waste of the city as well as the industrial sewage euent
from the industries at the outskirts. The soil bacteria of ECW were
expected to have a resistance against organic pollutants as this area was
receiving a mixed waste for a long time and was thus rich in

Corresponding author at: Department of Biotechnology Heritage Institute of Technology Chowbaga Road, Anandapur, Kolkata 700107, India.
E-mail addresses: bhaswati77@gmail.com (B. Chakraborty), srabanti_b@yahoo.co.uk (S. Basu).

http://dx.doi.org/10.1016/j.jwpe.2017.04.002
Received 27 January 2017; Received in revised form 29 March 2017; Accepted 4 April 2017
2214-7144/ 2017 Elsevier Ltd. All rights reserved.
B. Chakraborty, S. Basu Journal of Water Process Engineering 17 (2017) 135142

biodiversity.
The degradation rates and removal eciencies of phenol by these
soil microbes under various mean hydraulic retention times, air ow
rates and temperature conditions were observed and process optimiza-
tions were done for both co-current and counter current processes.
Moreover, the performance characteristics under the optimized condi-
tions were determined against real time industrial euent.

2. Materials and methods

2.1. Chemicals and reagents

Glucose and all the inorganic salts used in preparation of MS media

were reagent grade. All other chemicals used were of analytical grade.
All these chemicals and other reagents for analytical study were
purchased from Merck, India.

2.2. Collection and maintenance of samples

An indigenous mixed microbial culture expected to have potential

to degrade phenol and phenolic compounds were collected from a
sewage-based agricultural eld of the East Calcutta Wetlands, India.
10gm of collected soil sample was suspended in 0.8% saline water and
incubated at 32 C overnight in a shaker at 100 rpm. Then the soil
suspension was centrifuged at 3000 rpm for 3 min. The supernatant
containing the mixed microbial cells was added to nutrient broth and
cultured overnight for enhanced growth in nutrient rich media at 32 C.
The mixed culture was maintained on nutrient agar slant and stored at
4 C for future use.

2.3. Micro-organisms and culture conditions

Inoculum was prepared in nutrient broth with the microorganisms

preserved in nutrient agar slant. 24-h old culture was inoculated into
MS media (10%, v/v) in an Erlenmeyer ask. The composition of MS
media was (mg/L): KH2PO4:680, K2HPO4: 1730, FeSO4:30,
NH4NO3:100, CaCl2: 20, MnSO4:30 supplemented with glucose.
Glucose in the MS media was then gradually replaced by increasing
concentration of phenol till phenol served as the sole source of carbon. Fig. 1. (a) Schematic diagram of continuous co-current reactor system.(b) Schematic
By 16 s rDNA sequence data analysis it was found that Psychrobacter sp., diagram of continuous counter current reactor system.
Stenotrophomonas maltophilia, Bacillus subtilis and Escherichia coli were
present in the mixed culture [15]. Dp = 3(6 Vp/), where Vp = mean volume of a chip in cubic cen-
timeter. The value of Dp was found to be 1.08 cm. The bed volume was
2.4. Reactor, packing and immobilization procedure 80% of the total reactor volume.

Two 1000 ml bench scale reactors with aspect ratio (H:D) of 10:1
2.5. Scanning electron microscopy (SEM) and energy dispersive X-ray
made of Borosil glass packed with claychips and designed to function in
spectrometry (EDS) study
co-current and counter-current modes respectively were used as packed
bed reactors (PBR). One reactor had an inlet port 110 mm from the base
SEM and EDS were done with clay chips, bacteria immobilized clay
of the reactor and the other had an inlet port 15 mm from the base of
chips to obtain their topographical characterization and mineral
the PBR. There were two outlet ports at 25 mm and 50 mm from the
composition. SEM photographs were taken after coating the samples
base of each PBR respectively. Oxygen was supplied from the lower end
with palladium by JFC-1600 Autone Coater with a scanning electron
of the reactors from a compressed air source at varying ow rates from
microscope (JSM 6360) using 17 kV. EDS was done to ascertain the in
0 to 3 Lpm. The air ow rate was controlled and measured by a
carbon percentage using instrument INCA-MICS, 01736-03-04.
rotameter. The rate of aeration was sucient to maintain saturated DO
(dissolved oxygen) level (4.5 mg O2/L) in the reactor medium so as to
assume growth not to be limited by DO level. Schematic diagrams of the 2.6. Process description for removal of phenol from synthetic media and
reactors are given in Fig. 1a and b. real time industrial euent
The clay chips used here as the packing materials were broken
pieces of earthen tea cups used in Kolkata(India) by tea vendors to serve The phenol acclimatized mixed bacterial culture was suspended in
tea. The cups were non-biodegradable and caused environmental 800 mg/l of phenol in MS media. The phenol was the sole source of
pollution as bulk garbage. The used tea cups were collected from the carbon for the mixed culture. The mixed culture was allowed to grow in
local tea vendors, washed and dried under the sun. Then they were phenol media for 24 h. The optical density of this 24h-old cell
broken into smaller pieces and similar sized clay chips were selected for suspension was measured at 600 nm by UV-VIS spectrophotometer
packing the reactor. The void volume () was 0.81707 and the (Shimadzu). This mixed bacterial culture was circulated through the
equivalent diameter, Dp, was calculated from the following formula: PBR with the help of peristaltic pump at the rate of 6 ml/min for 12 h

136
B. Chakraborty, S. Basu
through the inlet port at 110 mm from the base of the reactor for
counter current and 15 mm from the base of the reactor for co-current
reactor. The cell-OD was again measured after immobilization process.
The cell loading factor was the dierence between the initial and nal
cell optical densities with respect to the initial cell-OD. The cell
loadings obtained were 70% of its initial cell mass upon immobilization
of the mixed culture on clay chips in both the reactors. The cells were
grown on the clay chips for 5 days in the reactor by passing 800 mg/l of
phenol through the PBR semi-continuously. The supernatant was
drained out each morning for 5 days till sucient cell growth occurred
on the clay chips.
The MS media supplemented with 800 mg/l of phenol as the sole
source of carbon was introduced into the reactor once from the upper
inlet port for counter-current continuous process and through the lower
inlet port for co-current continuous process. The air was sparged into
the reactors at the bottom ends of both the reactors and the air ow
rates were controlled by a rotameter. Both the counter current and co-
current continuous systems were studied at steady state at dierent
substrate ow rates, air ow rates and temperatures. The phenol
degradation rate and removal eciency at these varying conditions
were studied and optimized for each system.
Real time industrial euent was collected from a coke oven renery
in West Bengal. The initial phenol concentration was determined by
amino-antipyrene assay. Removal of phenol from real time industrial
euent was studied using the continuous system with better perfor-
mance.
3. Results and discussion
3.1. Scanning electron microscopy (SEM) and energy dispersive X-ray
spectrometry (EDS) study
SEM photographs of clay chips and clay chips containing immobi-
lized cells are given in Fig. 2a and b respectively. A reduction in pores
on the surface of clay chips was observed after immobilization. EDS
proles of clay chips and clay chips containing immobilized cells are
Fig. 2. (a) SEM image of clay chips, (b) SEM image of immobilized cells on clay chips (c) EDS spectrum of clay chips (d) EDS spectrum immobilized cells on clay chips.
137
Journal of Water Process Engineering 17 (2017) 135142
given in Fig. 2c and d respectively. It was observed that carbon content
increased with immobilization. Percentage of carbon content was
5.78% and 34.17% before and after immobilization respectively which
was indicative of successful immobilization of cells.
3.2. Phenol biodegradation by continuous PBR in co-current mode
In the co-current mode both the air supply and the substrate input
points were at the bottom of the PBR. Thus both the streams were
owing in the upward direction. The initial phenol concentration in the
synthetic feed was 800 mg/l and cell loading was determined to be
70%. The substrate ow rates were varied from 1 ml/min to 6 ml/min
at 30 C. The intermediate substrate ow rates were 2 ml/min, 2.5 ml/
min, 3 ml/min, 3.5 ml/min and 4 ml/min. For each phenol ow rate,
the air ow rates were varied as 0 Lpm, 1 Lpm, 1.5 Lpm, 2 Lpm,
2.5 Lpm and 3 Lpm. The parameters that were analyzed during the
degradation process were hydraulic retention time (HRT), removal
eciency (R.E), degradation rate (qs), maximum degradation rate
(qmax), saturation constant (Ks).
The present phenol degradation process followed Haldane model for
substrate inhibition. But it had been already established that at high
substrate inhibition coecient (Ki) values Haldane equation simplied
to Monod model [16]. The mass balance equation for substrate
utilization was as followed:
VR (dS/dt) = FS0 FS (VR g X)/YX/S (1)
Or, dS/dt = (F/VR) S0 (F/VR)S (g X)/YX/S,
At steady state, dS/dt = 0,
Or, D(S0 S) = (g X)/YX/S, = qS, (2)
where F is substrate(phenol) ow rate in mL/min, VR is reaction volume
in ml and F/VR is dilution rate, D. Degradation rate can be expressed as
the rate of substrate ow per unit length of the reactor as,
NS/L = dS/dt = qS = qmaxS/(KS + S)
B. Chakraborty, S. Basu Journal of Water Process Engineering 17 (2017) 135142

Table 1 layer at very low phenol ow rate. The rate of mass transfer of phenol
Evaluation of kinetic parameters for the co-current and counter current PBR for dierent through this layer by diusion was inversely proportional to the
air ow rates.
thickness of the layer. Thickness of this diusion layer was inversely
Air Flow Rate qmax (mg/l min) KS (mg/l) proportional to the phenol ow rate in the PBR [17]. In general, for
(Lpm) almost all substrate ow rates it was found that at higher ow rate of air
Co-current Counter-current Co-current Counter-current the removal eciency decreased due to foaming in the reactor. The
PBR PBR PBR PBR maximum removal eciency of around 82% was observed at a phenol
0 3.759 2 372.28 74.562 ow rate of 3 ml/min when the air ow rate was maintained at
1 4 5.128 616.04 437.644 22.5 Lpm.
1.5 7.3475 5.37 1523.71 394.22
2 8.33 3.477 976.67 90.89
2.5 8.33 1.738 661.95 19.57 3.2.2. Eect of airow rate on phenol degradation rate at various substrate
3 8.33 1.567 175.4904 12.567 ow rate
The phenol degradation rate, qS in mg/l min, was highest at phenol
ow rate of 6 ml/min at all the dierent air ow rates used in the
Or, 1/qS = KS/qmaxS + 1/qmax. (3) present study (Fig. 3b). But the system was continuously at the verge of
ooding at the given ow rate of 6.8 ml/min. Thus the optimum
For surface immobilization of cells on clay chips the diusion
limitation of substrate to cells was found to be negligible for limited degradation rate of 1.972 mg/l-h was at a substrate ow rate of
3.33 ml/min and air ow rates of 22.5 Lpm.
biolm thickness. Thus, assuming = 1, the kinetic parameters KS and
qmax were calculated for dierent physicochemical conditions observed.
The calculated values of the kinetic parameters for phenol degradation 3.2.3. Eect of hydraulic retention time on phenol degradation rate
by immobilized mixed culture in co-current continuous reactor were The phenol degradation rate was maximum at a hydraulic retention
given in Table 1. time of 155 min for all the dierent air ow rates (Table 2). At steady
The hydraulic retention time was calculated as VR/F for dierent state operations the rate of phenol degradation reduces with increase in
substrate ow rate. The phenol removal eciency of the co-current hydraulic retention time. At steady state the maximum phenol degrada-
PBR was calculated as R.E = [1 (S/S0)] 100. tion rate of 3 mg/l min was observed for air ow rate of 2 Lpm at
hydraulic retention time of155 min. The reason of this trend can be
explained in the similar manner as the previous observation since
3.2.1. Eect of airow rate on phenol removal eciency at various hydraulic retention time is inversely proportional to substrate ow rate.
substrate ow rate
Phenol removal eciency increased with decreasing substrate ow
3.2.4. Eect of temperature on phenol degradation rate and removal
rates. The reason being that hydraulic retention time of the substrate, in
eciency at various substrate ow rate
this case phenol, in the reactor increased with decreasing substrate ow
Phenol degradation rate and removal eciency of phenol by the
rates. However, at very low phenol ow rates (1 ml/min and 2 ml/min)
mixed consortium of bacteria was studied at dierent temperatures.
the removal eciency initially was less than the next higher ow rate
tested (3.33 ml/min) (Fig. 3a). The initial decrease may be due to
Table 2
saturation of cell metabolism and formation of an external diusion Eect of hydraulic retention time on phenol degradation rate at various air ow rates for
co-current continuous PBR.

Air ow HRT, min Phenol degradation rate, Phenol removal

rate, Lpm qs, mg/l min eciency, RE,%

0 147.0588 2.48 51.739

208.3333 1.961 49.047
300.3003 1.76 53.809
480.7692 1.7213 73.478
1000 1.106 69.130
1 147.0588 2.274 47.391
208.3333 1.580952 39.523
300.3003 1.763478 58.571
480.7692 1.874286 73.478
1000 1.175652 73.478
1.5 147.0588 2.692 56.087
208.3333 1.771 44.285
300.3003 1.659 72.857
480.7692 2.331 69.140
1000 1.2452 77.826
2 147.0588 2.90087 60.434
208.3333 1.771429 44.285
300.3003 1.972174 68.095
480.7692 2.179 82.173
1000 1.314783 82.173
2.5 147.0588 2.2747 47.391
208.3333 2.1523 53.809
300.3003 1.972174 58.571
480.7692 1.8742 82.173
1000 1.3843 86.521
3 147.0588 2.692 56.087
208.3333 1.961 39.523
300.3003 1.4171 44.285
Fig. 3. (a) Eect of airow rate on phenol removal eciency in co-current packed bed 480.7692 1.7634 73.478
reactor at 30 C.(b) Eect of airow rate on phenol degradation rate at various substrate 1000 1.2452 77.826
ow rates in co-current packed bed reactor at 30 C.

138
B. Chakraborty, S. Basu Journal of Water Process Engineering 17 (2017) 135142

Table 3
Comparative study of phenol removal eciency by continuous co-current reactor at
dierent temperatures at dierent substrate and air ow rates.

Substrate ow Air ow Removal Removal Removal

rate, ml/min rate, eciency at eciency at eciency at
Lpm 15 C, % 30 C, % 50 C, %

6.8 0 28.69 51.739 38.69565

1 33.04 47.3913 43.04348
1.5 33.04 56.087 43.04348
2 37.39 60.43478 47.3913
2.5 41.739 47.3913 41.73913
3 37.3913 56.087 37.23
4.8 0 33.0434 49.04762 43.04348
1 37.3913 39.52381 47.3913
1.5 37.3913 44.28571 47.3913
2 37.3913 44.28571 47.3913
2.5 41.739 53.80952 44.23
3 41.739 39.52381 41.67
4.2 0 41.739 53.80952 51.739
1 41.739 58.57143 51.739
1.5 46.08696 72.85714 56.08696
2 46.08696 68.09524 56.08696
2.5 41.739 58.57143 51.739
3 47.3913 44.28571 47.3913
3.33 0 29.09091 73.47826 39.09091
1 33.636 73.47826 43.63636
1.5 38.18182 69.14043 48.18182
2 33.636 82.17391 43.63636
2.5 33.636 82.17391 43.63636
3 38.18182 73.47826 48.18182
Fig. 4. (a) Eect of air ow rate on rate of phenol degradation at dierent substrate ow
2.08 0 33.636 69.13043 43.63636
rates at 15 C in co-current packed bed reactor.(b) Eect of air ow rate on rate of phenol
1 29.09091 73.47826 39.09091
degradation at dierent substrate ow rates at 50 C in co-current packed bed reactor.
1.5 29.09091 77.82609 39.09091
2 38.18182 82.17391 48.18182
The cells being mesophiles the temperature selected for this study were 2.5 33.636 86.52174 49.67
15 centigrade and 50 centigrade. The rate of phenol degradation (qs) 3 33.636 77.82609 50.23
increased with the increase in the ow rate of substrate as also with air
ow rate. But once foaming set in the packed bed reactor at higher air
Table 4
ow rates the rate of phenol degradation decreased at all temperature Temperature Optimization based on phenol degradation rate for co-current PBR system.
ranges. The rate of degradation of phenol was highest at 30 C as the
mixed consortium consisted mainly of mesophiles. The degradation rate Temperature,C Air Flow Rate, Lpm qs, mg/l min
was higher at 50 C as compared to 15 C (as shown in Fig. 4a and b).
15 0 1.47
The rate of phenol degradation at very low substrate ow rates were 1 1.7
comparable and very less at all temperatures. 1.5 1.7
The phenol removing eciency was maximum at 30 C for all 2 1.92
substrate ow rates (Table 3). At a substrate ow rate of 6.8 ml/min for 2.5 2.14
3 1.92
higher air ow rates the phenol removal eciency decreased due to
30 0 2.48
foaming at higher temperatures [18]. The rate of foaming was less at 1 2.27
15 C and thus the removal eciency more or less increased with 1.5 2.69
increasing air ow rates. Higher rate of air ow in absence of foaming 2 2.90
increases the rate of phenol removal by increasing the metabolic 2.5 2.27
3 2.69
activities. The removal eciency increased markedly at lower substrate 50 0 1.99
ow rates of 3.33 ml/min and 2.08 ml/min at 30 C due to optimal 1 2.21
residence time of the phenol in the reactor, rate of aeration and 1.5 2.21
temperature (Table 3). The phenol removal eciencies were 82% and 2 2.43
2.5 2.13
86.5% respectively where as at 15 C and 50 C the phenol removal
3 1.91
eciencies were 33% and 50% respectively under similar conditions.

All further analyses were done assuming the process optimum tem-
3.2.5. Optimization of removal eciency of phenol in co-current PBR
perature to be 30 C. Analyzing the experimental data of variation of
The experimental data obtained from experiments performed on the
rate of phenol degradation with substrate and air ow rates simulta-
co-current PBR system in duplicate varying the principal process
neously it was observed that the maximum phenol degradation rate of
parameters like substrate ow rate, air ow rate and temperature were
2.901 mg/l min occurred at a substrate ow rate of 6 ml/min and air
analyzed for optimization of process conditions based on phenol
ow rate of 2 Lpm (Table 5). It was found that the maximum rate of
degradation rate, (qs), and percentage phenol removal eciency, RE.
phenol degradation occurred at a substrate ow rate of 6 ml/min
Phenol degradation was a kinetic parameter whereas removal eciency
irrespective of the air ow rate into the reactor. However, the rate of
was a performance parameter of the reactor.
phenol degradation is not independent of aeration rate. When the rate
Initially, process temperature was optimized based on phenol
of supply of oxygen into the reactor was below the critical level the rate
degradation rate and the optimum temperature was found to be 30 C
of phenol degradation was comparatively less than the maximum value.
as shown in Table 4. This nding was in accordance to the known fact
When the air ow rate was in excess to 2 Lpm foam formation occurred
that the constituent microorganisms were predominantly mesophilic.

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B. Chakraborty, S. Basu Journal of Water Process Engineering 17 (2017) 135142

Table 5
Optimization based on phenol degradation rate and phenol removal eciency for a co-
current PBR system at 30 C.

Air ow Substrate ow Phenol degradation Phenol removal

rate, Lpm rate,ml/min rate, qs, mg/l min eciency, RE, %

0 6.8 2.48 51.73

4.8 1.96 49.04
4.2 1.76 53.80
3.33 1.72 73.47
2.08 1.11 69.13
1 6.8 2.27 47.39
4.8 1.58 39.52
4.2 1.76 58.57
3.33 1.87 73.47
2.08 1.17 73.47
1.5 6.8 2.69 56.08
4.8 1.77 44.28
4.2 1.65 72.85
3.33 2.33 69.14
2.08 1.24 77.82
2 6.8 2.90 60.43
4.8 1.77 44.28
4.2 1.97 68.09
3.33 2.18 82.17
2.08 1.31 82.17
2.5 6.8 2.27 47.39
4.8 2.15 53.80
4.2 1.97 58.57
3.33 1.87 82.17
2.08 1.38 86.52
3 6.8 2.69 56.08
4.8 1.96 39.52
4.2 1.41 44.28
Fig. 5. (a) Removal Eciency of phenol at dierent air ow rates in counter current
3.33 1.76 73.47
continuous PBR.(b) Phenol degradation rate at dierent air ow rates in counter current
2.08 1.24 77.82
continuous PBR.

in the reactor thus leading to oxygen depletion in the system resulting
in reduced metabolic rate. Since phenol was the sole carbon source for
the microorganisms in the consortium reduced metabolic rate meant
reduced rate of degradation of phenol. Thus based on rate of phenol
degradation the optimum substrate ow rate and air ow rate were
6 ml/min and 2 Lpm respectively in tandem.
Another optimization criterion was percentage phenol removal
eciency (Table 5). Removal eciency was a measure of the amount
of phenol that was degraded by the microorganisms with respect to the
initial phenol concentration in the synthetic media. So it was more
extrinsic in nature which depended on the initial concentration of the
phenol in the media. The maximum phenol removal eciency was 86%
at a substrate ow rate of 2.08 ml/min and air ow rate of 2.5 Lpm. But
when the overall experimental result was analyzed as in Table 5 it was
noted that more consistent removal eciency was obtained at an air
ow rate of 2 Lpm. The substrate ow rates of both 3.33 ml/min and
2.08 ml/min at air ow rate of 2 Lpm yielded a phenol removal
eciency of 82%.
3.3. Phenol biodegradation by continuous PBR in counter-current mode
Another more often employed continuous ow system is the
counter-current continuous process. The substrate is introduced from
the top section of the reactor and compressed air is sparged into the
reactor from the bottom. The phenol substrate ows in the downward
direction and the air travels in the opposite direction of the substrate
and hence the nomenclature- counter-current process. The study was
done at 30 C. The kinetic constants qmax (maximum specic degrada-
tion rate) and Ks (saturation constants) were calculated by assuming
negligible diusion limitation. The eciency factor was thus assumed
to be equal to 1. By using Eq. (3) the kinetic constants were calculated
and are given in Table 1.
3.3.1. Eect of airow rate on phenol removal eciency at various
substrate ow rates
In contrary to the co-current continuous packed bed reactor where
removal eciency was better at lower substrate ow rates, in counter-
current continuous packed bed reactor it was observed that higher
substrate ow rates gave better phenol removal eciency. Removal
eciency increased with aeration till foaming began in the reactor at
higher air ow rates. The maximum phenol removal eciency of 78.5%
was at a phenolic substrate ow rate of 4.8 ml/min with an air ow rate
of 2 Lpm (Fig. 5a). When the air ow rate was increased beyond 2 Lpm
foams were formed which depleted the oxygen concentration in the
reactor thus inhibiting the metabolic activities of the immobilized
bacterial cells. As a result phenol removal eciency decreased in those
cases. But for lower phenol ow rates even at higher air ow rates
beyond 2 Lpm foaming did not occur in the counter-current ow
reactor and availability of sucient oxygen enhanced phenol removal
eciency of the bacterial cells.
3.3.2. Eect of airow rate on phenol degradation rate at various substrate
ow rate
In the counter current PBR system the rate of phenol degradation
increased as the air ow rate was increased for all substrate ow rates
as the microbes were all aerobes. The degradation rate of phenol at
lower substrate ow rates did not increase appreciably with increasing
air ow rates as at very low substrate ow rates in countercurrent
reactor there were signicant substrate limitations in the system. But at
substrate ow rate of 6 ml/min it was observed that at air ow rates
higher than 2 Lpm phenol degradation rate deceased due to foaming in
the reactor space. The phenol degradation rate was maximum at an air
ow rate of 2 Lpm at a substrate ow rate of 6 ml/min (Fig. 5b).
3.3.3. Eect of hydraulic retention time on phenol degradation rate
The eect of hydraulic retention time on phenol degradation rate in
counter current packed bed reactor system is similar to that in co-

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Table 6 Table 7
Eect of hydraulic retention time on phenol degradation rate at various air ow rates for Optimization based on phenol degradation rate and phenol removal eciency for a
counter- current continuous PBR. counter current PBR system at 30 C.

Air ow HRT, min Phenol degradation rate, Phenol removal Air ow Substrate ow Phenol degradation Phenol removal
rate, Lpm qs, mg/l min eciency, RE, % rate, Lpm rate, ml/min rate, qs, mg/l min eciency, RE, %

0 147.05 1.704 50.12 0 6.8 1.70 50.12

208.33 1.37 57.23 4.8 1.37 57.23
300.30 0.82 45.43 4.2 0.82 45.43
480.76 0.47 49.62 3.33 0.47 49.62
1000 0.18 36.7 2.08 0.18 36.7
1 147.05 1.82 53.75 1 6.8 1.82 53.75
208.33 1.50 62.62 4.8 1.50 62.62
300.30 0.86 49 4.2 0.86 49
480.77 0.50 52 3.33 0.51 52
1000 0.21 42.22 2.08 0.21 42.22
1.5 147.05 2.08 61.37 1.5 6.8 2.08 61.37
208.33 1.5 62.75 4.8 1.50 62.75
300.30 0.97 56.7 4.2 0.97 56.7
480.77 0.59 58.37 3.33 0.59 58.37
1000 0.25 49.87 2.08 0.25 49.87
2 147.05 2.25 66.37 2 6.8 2.25 66.37
208.33 1.88 78.5 4.8 1.88 78.5
300.30 0.89 58.23 4.2 0.89 58.23
480.77 0.60 63.5 3.33 0.60 63.5
1000 0.26 52.35 2.08 0.26 52.35
2.5 147.05 1.78 52.37 2.5 6.8 1.78 52.37
208.33 1.82 76 4.8 1.82 76
300.30 1.18 68.32 4.2 1.184 68.32
480.77 0.62 71.12 3.33 0.62 71.12
1000 0.29 59 2.08 0.29 59
3 147.05 1.62 50.16 3 6.8 1.62 50.16
208.33 1.73 68.53 4.8 1.73 68.53
300.30 1.34 62.27 4.2 1.34 62.27
480.77 1.54 69.78 3.33 1.54 69.78
1000 0.25 49 2.08 0.25 49

Table 8
Degradation of phenol from real time industrial euent by mixed bacterial consortium in
a co-current continuous reactor (substrate ow rate: 3 ml/min, initial phenol concentra-
tion 590.058 mg/l, air ow rate: 2 Lpm, and hydraulic retention time 300 min).

Run Degradation rate, qs, mg/l min RE,%

1 1.24 63.33
2 1.43 72.85
3 1.43 72.85

3.3.4. Eect of temperature on phenol degradation rate and removal

Fig. 6. (a) Phenol degradation rate at dierent temperatures in counter current
continuous PBR.(b) Phenol Removal Eciency at dierent temperatures in counter
current continuous PBR.
current packed bed reactor system, that is, phenol degradation rate
decreases with increasing hydraulic retention time. The maximum
phenol degradation rate of 2.256 mg/l min was observed at an air ow
rate of 2 Lpm at a hydraulic retention time of 147 min (Table 6).
eciency at various substrate ow rates
In the countercurrent PBR system the variation of phenol degrada-
tion rate and phenol removal eciency were observed at dierent
temperatures. Since the mixed consortium was predominantly meso-
philic the best results were obtained at 30 C. Here observations made
at a single substrate ow rate of 6.8 ml/min were analyzed and shown
(Fig. 6a and b) as the behavior was similar at all other substrate ow
rates.
At lower air ow rates when the supply of oxygen was less than
optimal, the phenol degradation rate was higher at 50 C than at 30 C.
The probable reason would be that the mesophiles in the mixed
consortium were under oxygen limitation as well as in a temperature
condition outside the mesophilic range of 30 C37 C and thus were in
a stressed environment. Moreover, at comparatively higher tempera-
tures the rate of oxygen utilization was higher leading to a temporary
higher rate of degradation of phenol and copious foam formation. The
higher bubble residence time led to oxygen depletion in the bubbles
circulating in the reactor. As the rate of aeration was further increased
foam formation was boosted and thus the metabolic rates of the
microorganisms were severely decreased and thus rate of phenol
degradation decreased signicantly. At temperature higher than the
optimum temperature 30 C foaming occurred at air ow rates as low as

141
B. Chakraborty, S. Basu
1.5 Lpm. Higher temperature is conducive to foam formation in the
reactor [18]. At 15 C the metabolic rate of the mesophilic mixed
consortium was less than 30 C and as foaming at low temperature is a
much-controlled phenomenon rate of phenol degradation gradually
increased with increased air ow rates. The maximum phenol degrada-
tion was achieved at 30 C which was the optimum temperature
condition for mesophiles at an air ow rate of 2 Lpm. When the air
ow rate was increased further foam formation started leading to
decrease in the phenol degradation rate.
For substrate ow rate of 6 ml/min the maximum phenol removal
eciency was at 30 C reported a value of 66.4% as opposed to the
maximum phenol removal eciencies at 15 C and 50 C which were
64% and 63.375% respectively as shown in Fig. 6b. The overall
maximum phenol removal eciency in the counter current PBR system
was 78.5% at a substrate ow rate of 4.8 ml/min and air ow rate of
2 Lpm at 30 C.
3.3.5. Optimization of removal eciency of phenol in counter-current PBR
It had been already established that the mixed consortium was
mesophilic in nature and thus the optimum temperature for the counter
current PBR reactor system was 30 C. Therefore, the process para-
meters considered for optimization were substrate and air ow rates. As
in the case of co-current PBR system the optimization criteria were rate
of phenol degradation and percentage phenol removal eciency.
From analysis of Fig. 6a it was noted that phenol degradation rate
(qs) continually increased with increasing substrate ow rate till
ooding of reactor occurred. The phenol degradation rate depended
on the air ow rate in to the reactor also as the microorganisms were
aerobic in nature. Thus it was observed in Table 7 that phenol
degradation rate gradually increased with air ow rate. But when
higher substrate ow rate combined with higher air ow rate in
counter-current PBR system signicant foaming occurred in the reactor
leading to depleted oxygen concentration in the reaction system which
in turn aected the metabolic rate and henceforth decreased the phenol
degradation rate. The optimum substrate ow rate and air ow rate
that resulted in maximum phenol degradation rate of 2.25 mg/l min
were 6 ml/min and 2 Lpm respectively.
The other criterion for optimization of process parameters was
percentage phenol removal eciency as shown in Table 6. Removal of
phenol from the system eciently also depended on substrate ow rate
as well as air ow rates. The optimum hydraulic retention time with
respect to removal eciency was 208.34 min. The removal of phenol
was more ecient as air ow rate was increased. But beyond 2 Lpm air
ow rate foaming began in the reactor system thus reducing the phenol
removal eciency of the counter current PBR system. The maximum
removal eciency in this system was 78.5% at substrate ow rate of
4.8 ml/min and air ow rate of 2 Lpm.
3.4. Performance study of the co-current continuous packed bed reactor
with real time industrial euent
Comparing the performance of the co-current and counter current
packed bed reactor systems under identical process conditions it was
evident that the co-current PBR system was superior to counter current
PBR system. Thus the co-current PBR system was opted for further
study. Based on phenol degradation rate the optimum conditions were
temperature at 30 C, substrate ow rate at 6 ml/min and air ow rate
at 2 Lpm. But when the phenol removal eciency was considered the
performance of the said reactor system was 60% only which was far less
than the maximum removal eciency of the system, which was 86%.
On the other hand, at 3.33 ml/min media ow rate both degradation
rate and removal eciency of phenol were appreciable (2 mg/l min and
82% respectively). Therefore the process variables selected as optimum
conditions were temperature at 30 C, substrate ow rate at 3.33 ml/
min, air ow rate at 2 Lpm and hydraulic retention time, HRT, at
300.3 min.
142
Journal of Water Process Engineering 17 (2017) 135142
The initial phenol concentration was determined as 590.058 mg/l.
The industrial euent was then treated in the co-current PBR reactor
system at the nal optimized conditions mentioned above. Three
consecutive runs were performed and the result was given in Table 8.
In the rst run, the removal eciency as well as phenol degradation
rate was less compared to the second and third runs. The probable
reason could be introduction of new media instead of the synthetic
media that the mixed consortium was used to. The second and third
runs results were in accord. Phenol degradation rate was 1.43 mg/l min
and the phenol removal eciency of the present system from the real
time industrial euent was 72.85%. Thus by using the present system
in series conguration or by diluting the industrial euent as a
pretreatment to the present system the environmental regulation of
EPA(USA) regarding phenolic discharge could be adhered to by the
industry.
4. Conclusion
The mixed bacterial consortium isolated from East Calcutta
Wetlands (ECW) could degrade phenol at a higher rate in a co-current
than a counter current continuous packed bed reactor. The optimized
process conditions in co-current reactor were temperature at 30 C,
substrate ow rate at 3.33 ml/min, air ow rate at 2 Lpm and hydraulic
retention time, HRT, at 300.3 min. Rate of phenol degradation was
2 mg/l min and phenol removal eciency was 82%. The consortium
could degrade phenol from a real time industrial euent under
optimum conditions in a co-current reactor. Phenol degradation rate
was 1.43 mg/l min and the phenol removal eciency was 72.85%.
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