Journal of Environmental Chemical Engineering 7 (2019) 103158

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Journal of Environmental Chemical Engineering

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

Study the performance of continuous bioreactor for the treatment of T

wastewater containing methyl parathion by isolated Alcaligenes species
⁎
Sachin Rameshrao Geed , Kulbhushan Samal, Harsh Srivastava, Bheemiseetty Kartheek
Department of Chemical Engineering, Madhav Institute of Technology & Science, Gwalior 474005, India

A R T I C LE I N FO A B S T R A C T

Keywords: Methyl parathion degradation potential bacterial species was successfully isolated from the agricultural field and
Alcaligenes sp. identified as Alcaligenes sp. SRG. The coconut coir was used as packing media for the bioreactor. The process
Coconut coir parameters were optimized in a batch experiment as pH (7 ± 0.3); temperature (35 ± 3 °C) and methyl
Continuous bioreactor parathion concentration as 150 mg/L. The continuous operation was performed under optimized process
Inhibition kinetics
parameters and at varying flow rate of 10–40 ml/h. The performance of continuous bioreactor was studied and
Methyl parathion
found maximum removal efficiency greater than 90% at a flow rate of 30 mL/h. The residual analysis FT-IR and
GC–MS confirmed the biodegradation of methyl parathion. The phenolsulphonic acid, phenylamine, and benzo-
1,4-quinone are the confirmed metabolites of methyl parathion. The growth and inhibition kinetic parameters
were studied and found μmax; 0.176 per day; Ks; 77.85 mg/L and Ki; 289.01 mg/L.

1. Introduction
The agricultural production generally based upon the type of soil,
nature of water, quality of fertilizers, seeds and effect of pesticides. The
various compounds like herbicides, rodenticides, insecticides, fungi-
cides, molluscicides, nematicides, plant growth regulators and others
are part of the term pesticides [1]. The productions of pesticides were
first started with the manufacturing of BHC (beta-Hexa-
chlorocyclohexane) in a plant at Calcutta in the year1952 [2]. A recent
survey shows that the total production value of $4.9 billion of pesticide
was obtained in the financial year 2017 making India among the top
four largest pesticides producers after the USA, China and Japan [2,3].
Methyl Parathion has chemical formula O,O-diethyl O-4-nitrophenyl
thiophosphate has very wide potentials and still used as a major pes-
ticide worldwide [2]. In generally DDT & other chlorinated hydro-
carbon were used as a major pesticide, but Organophosphates esters
such as parathion have replaced them due to its non-degrading nature
under natural water in the basic medium at a normal temperature [3].
One more characteristic of methyl parathion and its toxic metabolite
Paraoxon is its Hydrolytic half-life which is 108 and 144 days respec-
tively [3]. Methyl Parathion is widely sold to control millets on cotton,
aphids, boll weevils, artichokes, etc. due to its non-systematic contact
and stomach insecticide [4]. Cotton poison is also the name of methyl
parathion and is applied in the production of crops such as corn, apples,
cotton, sugar cane, apricot and corn [2,5]. Along with these, it has high
potential to protect the crop from numerous insect species by inter-
acting with the nervous system of both humans and insects which
contains where essentially methyl parathion attacks its cholinesterase
an essential enzyme of the nervous system [3,6]. Possible ill effects of
Methyl Parathion long time contamination of land and drinking water
leading to high ecological imbalance and environmental problems
which will have an adverse effect on human bodies [7–10].
It is difficult to remove methyl parathion from polluted environ-
ment, in general, a physicochemical process is employed such as ex-
traction, etc. for the ejection of methyl parathion were used to treat
methyl parathion [11–13]. But the problem with these techniques is
they have disadvantages concerning toxic byproduct generation, high
operating cost and energy demand, etc. [13–15]. To overcome the
problems associated with the physicochemical procedure the biological
process was used to treat methyl parathion. Many kinds of literature
have been reported on the removal of methyl parathion by isolated
bacterial species such as Bacillus sp. Pseudomonas sp., Alcaligenes sp.,
etc. [11–17].
The limited literature is available on batch and continuous treat-
ment of methyl parathion. The present work aimed to understand the
biological degradation of methyl parathion with the help of isolated
bacteria isolated in coconut coir employing both batch and continuous
manner. The inhibition kinetics was also studied, and further biode-
gradation of methyl parathion was confirmed by confirmative analysis.

⁎
Corresponding author.
E-mail address: gsachinr.rs.che15@itbhu.ac.in (S.R. Geed).

https://doi.org/10.1016/j.jece.2019.103158
Received 10 April 2019; Received in revised form 30 April 2019; Accepted 11 May 2019
Available online 14 May 2019
2213-3437/ © 2019 Elsevier Ltd. All rights reserved.
S.R. Geed, et al.
2. Material method
2.1. Chemicals and media
Analytical standard of methyl parathion (> 99.9% purity) was ob-
tained from Sigma Aldrich, India where as chemicals (analytical grade)
were procured from Merck (Germany). Membrane filtration technique
with a pore size of 0.2 μm was used to filter the stock of methyl para-
thion solution(Pall scientific, India). The Basal salt medium (BSM) and
methyl parathion were used for the degradation studies with compo-
sitions 2.5.g of (NH4)2SO4, 0.3 g of MgSO4·7H2O, 0.02 g of CaCl2 · 2H2O,
0.002 g of FeSO4·7H2O, 1.7 g of Na2HPO4·12H2O, and 1.7 g of KH2PO4
per liter. The pH value was adjusted to 7.2 ± 0.2 and decontaminating
at 121 °C for 25 min.
2.2. Isolation of methyl parathion degrading isolates
The potential bacterial strain was isolated from soil contaminated
with methyl parathion by an enrichment culture technique and was
used in the degradation experiments. The isolated bacterial species
were grown in BSM agar and methyl parathion to check the potential
species for degradation. The biochemical test as Catalase, Motility,
Gram’s staining, Oxidase, etc. were carried out for the isolated bacterial
species using the biochemical kit as per guidelines of Bergey’s Manual
[16,17].
The molecular characterization was carried out to identify bacterial
species and performed the DNA pattern of the 16S rRNA of microbes.
The isolate was preserved on Potato Dextrose Agar (PDA, Merck) at
normal temperature and co-cultured periodically. Genomic DNA was
extracted from fresh cultured bacterial species using DNA removal Kit
(Vivantis Technologies, USA). Further, PCR amplification was con-
ducted using universal primer pairs of ITS 1 (50-TCCGTAGGTGAACC
TGCGG-30) and ITS 4 (50-TCCTCCGCTTATTGATATGC-30).
The succession of the bacterial isolates was carried out from Triyat
Scientific Hyderabad India. The obtained sequences were oriented with
sequences in the NCBI database by using CLUSTALW. The evolutionary
history was contingent using the Neighbor-Joining method. The evo-
lutionary distances were computed using the Maximum Composite
Likelihood method [18]. Evolutionary analyses were performed by
MEGA6 [19].
2.3. Bioreactor: startup, inoculation
The bioreactor was built using borosilicate glass containing a
working volume 1000 ml and total volume 141.3 ml with having a
height of 50 cm and an internal radius of 3.0 cm as shown in Fig. 1. The
openings for incoming and outgoing liquids were provided at 2.5 cm
and 45 cm respectively from the bottom whereas a 0.2 μm membrane is
Fig. 1. Schematic of continuous bioreactor used for the treatment of waste-
water containing methyl parathion.
2
Journal of Environmental Chemical Engineering 7 (2019) 103158
provided at the outlet to hold back the bacteria. To reduce the leakages,
the openings of the port were sealed by pinch cork using silicon tubing.
The inlet 2.5 cm from bottom and outlet 45 cm from bottom ports were
provided at appropriate positions to facilitate inlet and outlet feed of
reactants or products. The outlet was fixed with 0.2 μm membrane to
hold back the drop of bacteria from the bioreactor. All the opening of
the ports was closed by pinch cork with silicon tubing. Air was supplied
to the bioreactor using a compressor (Khosla, India) to preserve the
aerobic condition. Air sparger was used to maintain the uniform dis-
tribution and placed 1.5 mm above the bottom of the reactor.
The coconut coir was used as a packing media in the bioreactor; 15 g
of sterile coconut coir was filled into the reactor. For the immobilization
of bacterial species on coconut coir 10 g/L of glucose were added into
the reactor over 20 days. To check the biofilm formation, the SEM (SEM
QUANTA 200 F) analysis was performed, and after confirming the
biofilm, the biodegradation experiment was performed.
Before going for a continuous process, the batch reactor studies
were done to analyze the adsorption capacity of methyl parathion.
Batch experiments were performed by placing 100 mg/L, and 150 mg/L
of methyl parathion concentration along with coconut coir in two screw
cap bottles closed with a cork. Mechanical shaking at 160 rpm was
provided by placing it in a shaker for 60 h. The results of GC which were
performed to measure the methyl Parathion concentration in the head
as well as liquid phase showed extremely low adsorption capacity of
methyl parathion. These experiments were conducted in every 10 h.
The detailed procedure of such experiments were mentioned and
broadly described in our previous literatures [16,17]
2.4. Batch bioreactor: parametric optimization
The batch experiment was carried out to optimize the process
parameters. The process parameters like pH, temperature and methyl
parathion concentration were optimized. The pH (4.0–10.0), tempera-
ture (25–40 °C) and concentration 25–200 mg/L was used. For initial
experiments of the methyl parathion concentration and pH were kept
constant and the temperature was optimized. Then at optimized tem-
perature concentration kept constant and pH was optimized. Again for
concentration set of experiments were carried out at optimized pH and
temperature.
2.5. Continuous bioreactor operation
Data analyzed from the batch reactor results were exfoliated to
operate the continuous bioreactor under optimum parameters as shown
in Fig. 1. After identifying appropriate microbial species growth on
Coconut coir, it was acclimatized with Alcaligenes sp. SRG for 20 days
before biodegradation experiments. The feed solution was prepared by
placing 150 mg/L of methyl parathion in wastewater and placed in a
200 L container. This wastewater container was connected to a con-
tinuous bioreactor, and the feed is pumped with a varying flow rate of
10–40 ml/L using a peristaltic pump (McLins PP10). The continuous
bioreactor was functioned for 86 days and results were investigated in
terms of % Removal efficiency (Eq. (1)), Hydraulic Retention Time
(HRT) (Eq. (2)) and inlet methyl parathion concentration
Cin − Cout
% RE = × 100
Cin (1)
V
HRT =
Q (2)
where, Cin is inlet methyl parathion concentration in a bioreactor, Cout is
outlet methyl parathion concentration from the bioreactor, V is the
volume of the bioreactor and Q is volumetric flow rate (mL/h).
S.R. Geed, et al.
2.6. Residual analysis
Characterization of the biodegraded sample was done using FT-IR in
which functional group determination and bands formation were ana-
lyzed. The FT-IR spectra of the biodegraded sample after the end of the
experiments and control methyl parathion sample were analyzed by FT-
IR (NICOLET 5700 FT-IR, Japan) in wave number range of 400–4000
per cm.
Metabolites/intermediates formed during methyl parathion biode-
gradation were determined using GC–MS-QP2010 Ultra (Shimadzu,
Japan). In case of preparation of sample two equal volumes of biode-
graded solution and chloroform were added, chloroform will help in
extracting remaining methyl parathion. This mixture was stirred vig-
orously and then separate into two immiscible organic layers extract
residual methyl parathion. The program details are reported elsewhere.
Geed et al. [16].
2.7. Growth inhibition kinetics of parathion
For non-inhibitory condition, the Monod kinetic model (Eq. (3)) was
used
1 dx μ
μ= = maxC
X dt Ks + C (3)
where μ is a specific growth rate (1/day), μmax is maximum specific
growth rate (1/day), Ks is half-saturation constant (mg/L), X, C, and t
are microba ial cell, initial substrate concentrations (mg/L), and time,
respectively.
The kinetic parameters including growth inhibition constant Ki
(mg/L) were calculated using Andrews–Haldane model (Eq. (4))
[20,21].
C
μ= μmax
C2
Ks + C + Ki (4)
3. Results and discussion
3.1. Isolation and identification of methyl parathion degradating species
To identify the unknown species, two different characterization
process was used on bacterial isolates. To gather the information re-
garding metabolism capability in case of nutrients, media, and chemi-
cals biochemical test was conducted. This test helped in understanding
identifiable variations in the species. The bacterial isolate showed the
results for catalase, oxidase and motile test whereas gram staining
showed the negative result. The molecular characterization of the given
nucleotide sequence of 16S-rRNA was determined for bacterium species
and deposited in the NCBI GenBank database under the accession
number MG591722. The sequence-based on 16S-rRNA and the phylo-
genetic tree construction was done by BLAST software identified and as
Alcaligenes sp. SRG. The phylogenetic tree constructed with the help of
MEGA 6 as shown in Fig. 2. The kinds of literature are found on the
application of Alcaligenes sp. species for the biodegradation of a variety
of organic pollutants [17,22–24].
3.2. SEM analysis of Coconut coir before and after inoculation
SEM analysis of 0thday before immunization and end of bioreactor
operation in the presence of Parathion as a carbon source were per-
formed. No bacterial growth was observed on the 0th day shown in
Fig. 3(a), whereas, uniform size and population density of bacterial
species were observed on coconut coir on the 20th day of operation
Fig. 3(b). This result was in agreement with the results of successful
acclimatization of bacterial species on coconut coir which depend on
the various factors such as the adaptive capability of microorganism in
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Journal of Environmental Chemical Engineering 7 (2019) 103158
a bioreactor, nutrient availability, Methyl Parathion concentration, etc
[15]. Fig. 3(c) shows the stable biofilm on coconut coir at the end of the
experiment. Similar results for the morphology of immobilized micro-
organisms were also observed by several researchers for different pes-
ticides [16–19].
3.3. Parameters optimization
In order to adjust the process parameters, i.e. pH, temperature and
methyl parathion concentration, the temperature was varied between
24–42 °C with an increase of 3 °C. The percentage of removal increased
from 39% to 71% with increasing temperature from 24 °C to 34 °C.
Further increase in temperature up to 42 °C, the percentage removal
decreased to 49% (Fig. 5a).
At optimized pH (7 ± 0.3) and temperature (35 ± 3 °C), methyl
parathion concentration was varied from 25 to 200 mg/L. The max-
imum percentage removal was observed at an initial concentration of
150 mg/L. The pH was varied between 4–10. The percentage removal
increased with increasing pH up to pH seven then it started decreasing
up to pH 10. The maximum percentage removal (72%) was observed at
pH 7. This trend was obvious as the metabolism of bacterial species was
maximum at neutral pH (Fig. 5b).
3.4. Biodegradation in a continuous bioreactor
In the initial stage, the immobilization of bacterial species was
performed on coconut coir in the bioreactor. The bioreactor was sup-
plied with 15 g/L and 5 mL of glucose and bacterial culture containing
isolated bacterial species respectively to form the bio-film over the
coconut coir. The SEM result showed the growth of bacterial population
on coconut coir and formation of stable biofilm of 20th day SEM images
shown in Fig. 3(a) and (b). Hence, the 20th day was well thought-out as
the most suitable time for the initiation of the bioreactor. Methyl
parathion degradation was carried out in continuous bioreactor with
the changing feed rates from 10 to 40 mL/h under the optimized pro-
cess parametric condition obtained by batch experimental studied. The
coconut coir helped to maintain the high concentration of attached
biofilm on the coconut coir surface and hence the high rate of de-
gradation was obtained.
Methyl parathion concentration of 150 mg/L was fed to bioreactor
from 20th day onwards to the end of operation 86th day at varying feed
rates to estimate the performance of the bioreactor in terms of Removal
efficiency (RE) and Hydraulic retention time (HRT) shown in Fig. 6. The
bioreactor was managed at a 10 mL/h feed flow rate corresponding
HRT is 100 h to achieved the maximum removal efficiency 75% on the
26th day. As degradation further proceeds the RE was decreasing to
38%. Further increased the flow rate to 20 mL/h on the 37th day, the
corresponding HRT was 50 h and performance of bioreactor was im-
proved. The maximum RE was obtained at 20 mL/h was 84% on the
43rd day as the operation proceeds the RE was decreasing from 84 to
41% on 46 days. As obtained the constant RE 41% again increased the
flow rate 30 mL/h on the 47th day (Fig. 6). After a sharp dip further
increased the RE from 47 to 92% was observed. The HRT was 33.33 h,
and maximum RE was obtained 92 on the 64th-day operation. The
performance of bioreactor was observed better at this flow rate and
HRT. As the operation was proceeded after 64th further again bior-
eactor performance down from 92% to 51% constant RE(Fig. 6). After
getting the constant RE further increased the Flow rate 40 mL/h on the
75th day. The slight improved RE value from 51% to 72% on the 80th-
day operation. The reactor performed getting down further as proceeds
at an HRT 25 h. The performance of bioreactor was decreased may be
toxic byproduct forms and methyl parathion toxicity in the bioreactor.
As HRT decreased the performance of bioreactor improved and further
decreased the maximum RE was obtained at 33.33 h HRT (Fig. 6). The
similar patterns of trends of RE were obtained at the entire flow rate as
initially increased the RE achieved maximum and further decreased.
S.R. Geed, et al. Journal of Environmental Chemical Engineering 7 (2019) 103158

Fig. 2. Phylogenetic evaluation of bacterial isolate Alcaligenes sp. SRG used for methyl parathion biodegrdadation.

Fig. 3. SEM/morphological images of (a) coconut coir, (b) biofilm over coconut coir, (c) biofilm at the end of experiment, (d) FTIR spectra of control methyl
parathion and biodegraded sample.

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S.R. Geed, et al. Journal of Environmental Chemical Engineering 7 (2019) 103158

Fig. 4. Fragmented GC–MS result of (a) Control methyl parathion, (b) metabolite phenolsulphonic acid, (c) metabolite phenylamine, (d) metabolite benzo-1,4-
quinone.

Fig. 5. (a) Effect of temperature on removal of methyl parathion, (b) effect of pH on removal of methyl parathion, (c) growth inhibition kinetics.

Similar studies have been reported by various literature [16,17,25–27].
3.5. Analysis of residual methyl parathion: FT-IR and GC–MS
FTIR transmittance spectra of methyl parathion and other metabo-
lites formed during biodegradation were recorded from 600 to 4000 per
cm (Fig. 3d). The change in intensity of peaks and shifting of peaks
were identified. The peaks for methyl parathion were observed at 3372
per cm for C–H stretching, 1508 per cm for N]O stretching, 2328 per
cm for CeH bending, 1064 per cm for CeN stretching, 673 per cm for
P]S stretching and 3610 per cm for OeH stretching. The characteristic
peaks of metabolites were observed at 3581 per cm for eOH stretching,
3410 per cm for eNH stretching, 1640 per cm for C]C stretching, 1221
per cm for CeO stretching, 1008 per cm for CeH bending and 627 per
cm for CeS stretching.
The GC–MS analysis of control methyl parathion sample and treated

5
S.R. Geed, et al.
Fig. 6. Performance evaluation of continuous bioreactor for methyl parathion biodegradation using Alcaligenes species with varying flow rate.
effluent was performed. The GC–MS shattered or fragmented pattern of
methyl parathion showed parent ion peak at m/z 29, 65, 81, 97, 109,
125, 137, 142, 155, 172, 186, 218, 235, 246, 263 & 291 as shown in
Fig. 4(a). The GC–MS result confirmed that the methyl parathion de-
graded during the treatment process. The Fig. 4(b) showed a frag-
mented pattern of phenol sulphonic acid parent ion peak at m/z 39, 50,
66, 74 & 94. Similar fragmented pattern for phenyl amine showed
Fig. 4(c) m/z ion peak at 24, 39, 55, 66, 77 & 94. The Fig. 4(d) showed a
fragmented pattern for benzo- 1,4, quinone parent ion peak at m/z 28,
39, 55, 63, 81, 84, 110 & 112. In the present work, methyl parathion
was degraded during the treatment process and converted to three
metabolites (phenolsulphonic acid, phenylamine, and benzo-1,4-qui-
none) as confirmed by GC–MS result.
3.6. Growth inhibition kinetics
The growth inhibition kinetics was determined for methyl parathion
biodegradation. The separate bench scale series of batch (flask) study
were performed. The experiments were conducted with initial methyl
parathion concentration was ranged in between 25–500 mg/L. The es-
timation of biomass concentration was on a dry weight basis (g/L), and
the specific growth rate was calculated. The important growth inhibi-
tion constant (Ki) was determined underneath substrate (methyl para-
thion) inhibition using Andrew–Haldane model as shown in Fig. 5 (c)
The growth kinetic parameters half saturation rate constant (Ks),
maximum specific growth rate (μmax) and inhibition kinetic constant
(Ki) were estimated 77.84 mg/L, 0.176 per day and 289.01 mg/L re-
spectively. The Fig. 5(c) shows the inhibition kinetics by Andre-
w–Haldane model. The Ki (289.01 mg/L) exhibits the strong inhibition
in attended in the system. This value shows the chances of inhibition at
an increased concentration above 289.01 mg/L. Availability of litera-
ture on the inhibition kinetics of methyl parathion biodegradation is
extremely rare. The similar literature are found on growth inhibition
kinetics by (Andrew–Haldane model) and stated by various authors for
different organic compound [16,17].
4. Conclusion
In this work, the methyl parathion biodegradating microbial species
Alcaligenes sp. SRG was successfully isolated. The degradation of methyl
parathion containing synthetic wastewater was studied in a bioreactor
6
Journal of Environmental Chemical Engineering 7 (2019) 103158
with a varying flow rate. The best performance of bioreactor was ob-
served at a flow rate of 30 mL/h at a methyl parathion concentration of
150 mg/L was used and achieved 92% RE on 64th day of operation.
While analyzing the output of biodegraded methyl parathion through
GC–MS some common metabolite like phenolsulphonic acid,
Phenylamine and Benzo-1,4-quinoen existence was seen. The growth
inhibition Andrew–Haldane kinetic model was used to know the growth
inhibition constant (Ki; 289.01; mg/L). This procedure was used to treat
wastewater containing impurity in an aerobic bioreactor at high con-
centration.
Acknowledgment
I (S.R. Geed), thanks to MHRD India, TEQUIP-III (NPIU) and also
thankful to Prof B.N. Rai from, FRO Department of Chemical
Engineering & Technology, IIT (BHU) Varanasi for encouragement,
guidance and help in my research work.
References
[1] W. Aktar, D. Sengupta, A. Chowdhury, Impact of pesticides use in agriculture: their
benefits and hazards, Interdiscip. Toxicol. 2 (2009) 1–12.
[2] OIPI Outlook of Indian Pesticide Industry (2017).
[3] A. Kotronarou, G. Mills, M.R. Hoffmann, Decomposition of parathion in aqueous
solution by ultrasonic irradiation, Environ. Sci. Technol. 26 (1992) 1460–1462.
[4] S. Garcia, A. Abu-Qare, W. Meeker-O’Connell, A. Borton, M. Abou-Donia, Methyl
parathion: a review of health effects, J. Toxicol. Environ. Health B Crit. Rev. 6
(2003) 185–210.
[5] F.L. Edwards, P.B. Tchounwou, Environmental toxicology and health effects asso-
ciated with methyl parathion exposure — a scientific review, Int. J. Environ. Res.
Public Health 2 (2005) 430–441.
[6] O.A. Timofeeva, D. Sanders, K. Seemann, L. Yang, D. Hermanson, S. Regenbogen,
S. Agoos, A. Kallepalli, A. Rastogi, D. Braddy, C. Wells, Persistent behavioral al-
terations in rats neonatally exposed to low doses of the organophosphate pesticide,
parathion, Brain Res. Bull. 77 (2008) 404–411.
[7] M.J. Lydy, T.W. Lohner, S.W. Fisher, Influence of pH, temperature and sediment
type on the toxicity, accumulation and degradation of parathion in aquatic systems,
Aquat. Toxicol. 17 (1990) 27–44.
[8] A. Ghauch, J. Rima, C. Amine, M. Martin-Bouyer, Rapid treatment of water con-
tamined with atrazine and parathion with zero-valent iron, Chemosphere 39 (1999)
1309–1315.
[9] L.G. Sultatos, The effects of phenobarbital pretreatment on the metabolism and
acute toxicity of the pesticide parathion in the mouse, Toxicol. Appl. Pharmacol. 86
(1986) 105–111.
[10] A. Muttray, U. Spelmeyer, M. Degirmenci, D. Jung, G. Bäcker, G. Hill, Acute effects
of low doses of methyl parathion on human EEG, Environ. Toxicol. Pharmacol. 19
(2005) 477–483.
S.R. Geed, et al.
[11] M.L. Ortiz-hernández, M. Monterrosas-brisson, G. Yanez-ocampo, E. Sanchez-sal-
inas, Biodegradation of methyl-parathion by bacteria isolated of agricultural soil,
Rev. Int. Contam. Ambie 17 (2001) 147–155.
[12] X. Zhang, Q. Huang, M. Liu, J. Tian, G. Zeng, Z. Li, K. Wang, Q. Zhang, Q. Wan,
F. Deng, Y. Wei, Preparation of amine functionalized carbon nanotubes via a
bioinspired strategy and their application in Cu2+ removal, Appl. Surf. Sci. 343
(2015) 19–27.
[13] X. Zhang, Q. Huang, F. Deng, H. Huang, Q. Wan, M. Liu, Y. Wei, Mussel-inspired
fabrication of functional materials and their environmental applications: progress
and prospects, Appl. Mater. Today 7 (2017) 222–238.
[14] M. Megharaj, D.R. Madhavi, C. Sreenivasulu, A. Umamaheswari, K. Venkateswarlu,
Biodegradation of methyl parathion by soil isolates of microalgae and cyano-
bacteria, Bull. Environ. Contam. Toxicol. 53 (1994) 292–297.
[15] Q. Huang, J. Zhao, M. Liu, J. Chen, X. Zhu, T. Wu, J. Tian, Y. Wen, X. Zhang, Y. Wei,
Preparation of polyethylene polyamine@ tannic acid encapsulated MgAl-layered
double hydroxide for the efficient removal of copper (II) ions from aqueous solu-
tion, J. Taiwan Inst. Chem. Eng. 82 (2018) 92–101.
[16] S.R. Geed, M.K. Kureel, B.S. Giri, R.S. Singh, B.N. Rai, Performance evaluation of
malathion biodegradation in batch and continuous packed bed bioreactor (PBBR),
Bioresour. Technol. Rep. 227 (2017) 56–65.
[17] S.R. Geed, S. Prasad, M.K. Kureel, R.S. Singh, B.N. Rai, Biodegradation of waste-
water in alternating aerobic-anoxic lab scale pilot plant by Alcaligenes sp. S 3 iso-
lated from agricultural field, J. Environ. Manage. 214 (2018) 408–415.
[18] K. Tamura, M. Nei, S. Kumar, Prospects for inferring very large phylogenies by
using the neighbor-joining method, Proc. Natl. Acad. Sci. 101 (2004) 11030–11035.
[19] K. Tamura, D. Peterson, N. Peterson, G. Stecher, M. Nei, S. Kumar, MEGA5:
Journal of Environmental Chemical Engineering 7 (2019) 103158
molecular evolutionary genetics analysis using maximum likelihood, evolutionary
distance, and maximum parsimony methods, Mol. Biol. Evol. 28 (2011) 2731–2739.
[20] J.S.B. Haldane, Enzymes, Longmans, Green, UK, 1930 (Republished by MIT Press,
Cambridge, MA1965).
[21] J.F. Andrews, A mathematical model for the continuous culture of microorganisms
utilizing inhibitory substrates, Biotechnol. Bioeng. 10 (1968) 707–723.
[22] A.E. Abo-Amer, A.E.R.R. El-Shanshoury, O.M. Alzahrani, Isolation and molecular
characterization of heavy metal-resistant Alcaligenes faecalis from sewage waste-
water and synthesis of silver nanoparticles, Geomicrobiol. J. 32 (2015) 836–845.
[23] H. Li, H. Xu, S. Li, X. Feng, H. Xu, P. Ouyang, Effects of dissolved oxygen and shear
stress on the synthesis and molecular weight of welan gum produced from
Alcaligenes p, Process Biochem. 46 (2011) 1172–1178.
[24] I. Yusuf, S.A. Ahmad, L.Y. Phang, M.A. Syed, N.A. Shamaan, K.A. Khalil,
F.A. Dahalan, M.Y. Shukor, Keratinase production and biodegradation of polluted
secondary chicken feather wastes by a newly isolated multi heavy metal tolerant
bacterium-Alcaligenes sp. AQ05-001, J. Environ. Manage. 183 (2016) 182–195.
[25] S.R. Geed, M.K. Kureel, S. Prasad, R.S. Singh, B.N. Rai, Novel study on biode-
gradation of malathion and investigation of mass transfer correlation using alginate
beads immobilized Bacillussp. S4 in bioreactor, J. Environ. Chem. Eng. 6 (2018)
3444–3450.
[26] M.K. Kureel, S.R. Geed, B.S. Giri, B.N. Rai, R.S. Singh, Biodegradation and kinetic
study of benzene in bioreactor packed with PUF and alginate beads and im-
mobilized with Bacillussp. M3, Bioresour. Technol. 242 (2017) 92–100.
[27] M. Yadav, N. Srivastva, R.S. Singh, S.N. Upadhyay, S.K. Dubey, Biodegradation of
chlorpyrifos by Pseudomonas sp. In a continuous packed bed bioreactor, Bioresour.
Technol. 165 (2014) 265–269.