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Degradation of ciprofloxacin using Fenton's oxidation: Effect of operating

parameters, identification of oxidized by-products and toxicity assessment

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DOI: 10.1016/j.chemosphere.2017.11.046

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 Chemosphere 193 (2018) 1181e1188

Contents lists available at ScienceDirect

Chemosphere
journal homepage: www.elsevier.com/locate/chemosphere

Degradation of ciprofloxacin using Fenton's oxidation: Effect of

operating parameters, identification of oxidized by-products and
toxicity assessment
Anirudh Gupta, Anurag Garg*
Centre for Environmental Science and Engineering, Indian Institute of Technology Bombay, Powai, Mumbai, India

h i g h l i g h t s g r a p h i c a l a b s t r a c t

Effect of various reaction parameters

on Fenton's oxidation of ciprofloxacin
(CPX).

Kinetic evolution and/or decay pro-
files of CPX and its degradation
products.

Determination of dissolved inorganic
ions (Fe2þ, Fe3þ, F, NO þ
3 and NH4 )
and H2O2.

Addition of t-butanol reduced CPX
degradation due to its radical scav-
enging action.

Reduction in toxicity of wastewater
after Fenton treatment using
respirometer.

a r t i c l e i n f o a b s t r a c t

Article history: Ciprofloxacin (CPX), a fluoroquinolone antibiotic, is found in the bulk effluents emerged from pharma-
Received 5 July 2017 ceutical industries. Its presence in the effluent may lead to toxicity to the aquatic life and antimicrobial
Received in revised form resistance. Hence, the oxidative degradation of CPX by classical Fenton's process was investigated for the
28 October 2017
present study. The maximum CPX and total organic carbon (TOC) removal from the synthetic wastewater
Accepted 10 November 2017
Available online 13 November 2017
(initial CPX concentration ¼ 100 mg L1) were 70% and 55%, respectively, under the best reaction con-
ditions ([H2O2]:[Fe2þ] ¼ 10, stoichiometric H2O2 concentration ¼ 14.2 mM and initial wastewater
Handling Editor: Hyunook Kim pH ¼ 3.0). Five major degradation products could be identified by high resolution liquid chromatography
emass spectrometry (HR LC-MS) which could have resulted by the occurrence of decarboxylation,
Keywords: defluorination, hydroxylation and cleavage of the piperazine ring. The time based detection of ferrous
Ciprofloxacin and ferric ions in aqueous phase confirmed the CPX degradation by hydroxyl radicals. A pathway has
Degradation by-products been proposed for CPX degradation by Fenton's process based on the kinetic decay and/or evolution
Fenton's oxidation profiles of CPX and intermediates as well as the release of inorganic ions into the aqueous solution. It was
Reaction kinetics
indicated from the rapid respirometric test that the wastewater toxicity was lowered after Fenton's
Respirometric test
treatment.
© 2017 Elsevier Ltd. All rights reserved.

1. Introduction

The discharge of persistent pharmaceutically active compounds

* Corresponding author.
E-mail address: a.garg@iitb.ac.in (A. Garg). in aquatic water bodies is a major environmental issue. Such

https://doi.org/10.1016/j.chemosphere.2017.11.046
0045-6535/© 2017 Elsevier Ltd. All rights reserved.
1182 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188
pollutants are not amenable to the conventional wastewater
treatment processes (Heberer, 2002). Ciprofloxacin (CPX) is one of
the most widely used antibiotics (Haddad et al., 2015). It belongs to
class of fluoroquinolones (chemical formula ¼ C17H18FO3N3; mo-
lecular weight ¼ 331) and is bactericidal in nature. The industrial
wastewater may contain CPX concentration of 0.5 mM (i.e.
165.5 mg L1) (Rakshit et al., 2013). Proper measures need to be
taken for CPX removal from wastewater to prevent its ecotoxico-
logical effects and antimicrobial resistance on the receiving water
bodies. In aqueous environment, CPX exists in the cationic and
anionic form in acidic and alkaline solution, respectively (pKa1 ¼ 6.1
and pKa2 ¼ 8.7) due to the presence of carboxylic and amine groups
in the piperazine moiety (Gu and Karthikeyan, 2005).
Advanced oxidation processes (AOPs) are considered promising
option for the removal of pharmaceutically active compounds
(especially fluoroquinolones) from aqueous systems (Rivera-Utrilla
et al., 2013; Giri and Golder, 2015; Feng et al., 2017). A recently
published review article provides deep insight into the various
aspects associated with metal mediated oxidation of fluo-
roquinolone antibiotics. These aspects include kinetics, trans-
formation products and toxicity assessment. It has been reported
that the application of AOPs can reduce aquatic toxicity of the
fluoroquinolones.
Among AOPs, Fenton's oxidation is very popular and its per-
formance largely depends upon the generation of highly reactive
hydroxyl (
OH) radicals (oxidation potential ¼ 2.8 V). Extreme
acidic conditions favour the decomposition of hydrogen peroxide
into hydroxyl radicals in the presence of ferrous (Fe2þ) catalyst
(Feng et al., 2015; Biglarijoo et al., 2016). The recent studies pub-
lished on the degradation of other fluoroquinolones (such as lev-
ofloxacin and norfloxacin) by Fenton's oxidation showed promising
results (Epold et al., 2015; Santos et al., 2015). Few research groups
have performed Fenton's oxidation for CPX degradation also in the
recent past (Sun et al., 2009; Yang and Chen, 2013; Giri and Golder,
2014). Sun et al. (2009) reported around 91% CPX degradation from
synthetic wastewater (CPX concentration ¼ 15 mg L1) after Fen-
ton's oxidation performed for 120 min duration. Among other
conditions, the initial reaction pH of the wastewater was set at 4.0
while H2O2 to Fe2þ molar ratio was used as 100. Yang and Chen
(2013) reported CPX removal efficiency of more than 90% within
30 min from synthetic wastewater (initial CPX
concentration ¼ 50 mg L1) under best operating conditions
(pH ¼ 2, [H2O2]:[Fe2þ] ¼ 2.7). In another study, CPX degradation
from a synthetic wastewater (initial CPX
concentration ¼ 15 mg L1) was studied by Fenton's process under
the following conditions: [H2O2]:[Fe2þ] ¼ 5.6e20; pH ¼ 2e4.5,
temperature ¼ ambient (~30  C) and reaction time ¼ 45 min (Giri
and Golder, 2014). The CPX and total organic carbon (TOC) re-
movals of 74% and 38%, respectively were reported within 45 min at
the optimum reaction conditions (i.e., [H2O2]:[Fe2þ] ¼ 8; pH ¼ 3.5).
In this study, CPX degradation pathway was also suggested based
on the intermediates identified in the wastewater sample with-
drawn after 10 min of reaction.
Apart from the above studies, some researchers employed
photo-catalytic oxidation and ozonation for CPX removal from the
wastewater (CPX concentration ¼ 5e10 mg L1) (Liu et al., 2012;
Zhang et al., 2015). In both the studies, the complete removal of
CPX was reported and upto 20 degradation products were detected.
It is evident from the literature that there is limited information
available on the detailed degradation pathway of CPX during Fen-
ton's oxidation process. In particular, the time based profiles of
various intermediates are not available up to the best of our
knowledge. In addition, the release of inorganic ions (such as F,
NO þ
3 and NH4 ) during the reaction is not discussed. In addition, the
above studies used upto 50 mg L1 CPX concentration in
wastewater though the majority of these works investigated the
performance of AOP with much lower concentration of CPX. Only
limited information is available on the treatment of high concen-
tration levels of pharmaceuticals typically found in industrial dis-
charges (Larsson, 2014).
Therefore, a systematic study was performed to enhance un-
derstanding on CPX degradation from synthetic wastewater by
Fenton's process. The major objectives of the study were (i) to
determine the effect of various reaction parameters (such as pH,
[H2O2]:[Fe2þ] ratio, substrate concentration and temperature) on
CPX removal, (ii) to examine the time based degradation and/or
evolution profiles for parent and intermediate compounds, (iii) to
determine the consumption of H2O2 during the oxidation reaction
and release of inorganic ionic species (F, NO þ
3 and NH4 ) from CPX
degradation, (v) to estimate reaction rate constants and (vi) to
predict change in toxicity of Fenton treated wastewater.
2. Materials and methods
2.1. Materials
High performance Liquid chromatography (HPLC) grade CPX
was procured from Sigma Aldrich Chemical Ltd., Mumbai, INDIA.
The other chemicals such as H2O2 (30% w/v), FeSO4$7H2O, H2SO4
(98% by wt.) and acetonitrile were purchased from Merck Chem-
icals, Mumbai, INDIA. For respirometric analysis, activated sludge
sample was collected from a local wastewater treatment plant.
The synthetic wastewater was prepared by dissolving a pre-
determined amount of CPX in 1 L of distilled water. The main
characteristics of the wastewater were as follows: CPX
concentration ¼ 50e200 mg L1, pH ¼ 7.0 and
TOC ¼ 30.75e123 mg L1.
2.2. Procedure for Fenton's oxidation runs
Batch experiments were conducted with 100 mL of the synthetic
wastewater aliquots in a 250 mL capacity Pyrex reactor (in dupli-
cates or triplicates). Predetermined concentration of ferrous ions
(as FeSO4.7H2O salt) and the stoichiometric concentration of
hydrogen peroxide were added to the wastewater in the same
sequence while mixing the reactants with a magnetic stirrer
(agitation speed ¼ 300 rpm). The stoichiometric amount of H2O2
was calculated using the following equation (Sun et al., 2009):
C17H18N3O3F þ 47H2O2 / 17CO2 þ 54H2O þ 3HNO3 þ HF (1)
To find the effect of various operating parameters on the Fen-
ton's oxidation process, H2O2 to Fe2þ molar ratio was varied 5 to 50,
and the initial reaction pH was adjusted to 2e4 using H2SO4. The
reaction temperature was varied from ambient (¼ 30  C) to 60  C.
The reaction was supposed to commence at the instant when H2O2
was added to the reactor. The samples were withdrawn periodically
and 1 mL of 1 N NaOH solution was added to 5 mL sample for
quenching the reaction. The determination of optimum NaOH so-
lution concentration required for quenching the oxidation process
is discussed in supporting information (Section S.1). Subsequently,
the samples were filtered using 0.2 mm nylon membrane filter
before subjecting to the analysis for CPX and TOC concentrations.
The measured concentrations of CPX and TOC were corrected by
multiplying a correction factor of 1.2 taking the dilution of 5 mL
sample with 1 mL of 1 N NaOH solution into account. The dissolved
ferrous and ferric ion concentrations in the solution were obtained
using 1, 10-Phenanthroline method (APHA, 2005). The residual
H2O2 concentration in a wastewater sample was determined by
titanium oxalate colorimetric method with a UV/vis
 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188
spectrophotometer (UV 210 A, Schimadzu, Kyoto, JAPAN)
(Brandhuber and Korshin, 2009). To confirm whether the degra-
dation of CPX is free radicals mediated or not, radical scavenger t-
Butanol was added before the commencement of reaction (Bae
et al., 2013). The release of nitrogen species (i.e., NHþ4 and NO3

ions) into the wastewater was determined using HQ40d multi-
parameter probe supplied by HACH, Loveland, USA while F ions
were measured by standard SPADNS colorimetric method (APHA,
2005).
2.3. High performance liquid chromatography (HPLC) analysis
To determine CPX concentration in wastewater, HPLC analysis
was performed on the liquid chromatograph equipped with
UVevisible detector (Pump: Model PU-2089i, Intelligent HPLC
Pump, JASCO, PDA Detector: MD-2010/2015, Intelligent UV/VIS
Multi wavelength Detector, JASCO, Easton, USA). A 20 mL sample
was injected directly in a Kinetex C18 [(150 mm * 4.6 mm) with
(5 mm particle size)] reverse phase analytical column. A mixture of
high purity water and acetonitrile (ratio ¼ 20:80 v/v) was used as
the mobile phase (flow rate ¼ 1 mL/min). pH of the mobile phase
was adjusted to 3.0 using 5% (v/v) o-phosphoric acid. The sus-
pended particles were removed by filtering the samples through
0.2 mm nylon membrane filter. For the determination of CPX, the
absorbance of samples was measured at 280 nm wavelength.
2.4. High resolution liquid chromatography-mass spectrometry (HR
LC-MS) analysis
For HR LC-MS analysis, the wastewater samples
(volume ¼ 100 mL) were passed through HPLC (1290 Infinity UHPLC
system, Agilent Technologies, Santa Clara, USA) followed by PDA
detector-mass spectrometer. The chromatographic separation was
performed on the same Kinetex C18 reverse phase analytical col-
umn. Flow rate of the mobile phase was 0.8 mL min1. A linear
gradient elution was taken from 93% A (water) and 7% B (acetoni-
trile) to 50% A and 50% B in 20 min duration which was returned to
the original composition after 5 min. An electrospray interface (ESI)
was used in positive ionisation mode and full scan acquisition for
the MS and MS/MS measurements. The capillary voltage was set at
3.2 kV and argon was used as collision gas. The source block and
desolvation temperatures were maintained at 130  C and 400  C,
respectively. N2 was purged at a flow rate of 400 L h1 and 150 L h1
for drying and sheathing, respectively. A cone voltage of 25 and
75 V was used to determine the molecular mass to charge ratio (m/
z) of parent ion and for the fragmentation of daughter ions,
respectively. The change in concentration of a compound was
determined using peak area measurements.
2.5. Toxicity assessment
The rapid toxicity assessment of untreated and treated waste-
water was performed according to the procedure described by
Gutierrez et al. (2002) and Dalzell et al. (2002) using respirometric
instrument (PF-8000 Respirometer Systems and Application,
Arkansas, USA). The instrument was equipped with eight bio-
reactors (500 mL volume each), control module, oxygen supply
port, magnetic stirrer and a computer to capture the oxygen con-
sumption patterns. The rapid assessment was based on the respi-
ratory inhibition test for activated sludge. For this test, a nutrient
medium containing minerals, micro-nutrients and buffer solutions,
was prepared (Young and Cowan, 2004). The information on the
composition and preparation method for nutrient medium is dis-
cussed in the supplementary information (Section S.2).
3 mL each of mineral base (i.e. I and II), 5 mL of nutrient base and
1183
5 mL of buffer base were added to each reactor bottle. Sodium
acetate was used as the reference substrate. Oxygen was supplied
by means of oxygen cylinder and the mixture was agitated at
700 rpm. The above respirometric test was performed on control,
CPX laden wastewater and treated wastewater. The compositions of
various samples were taken as: The control sample contained
16 mL nutrient medium, 234 mL activated sludge, 10 mL sodium
acetate and 240 mL distilled water. The untreated wastewater
sample contained 16 mL nutrient medium, 234 mL activated
sludge, 10 mL sodium acetate and 240 mL untreated wastewater
whereas the treated wastewater sample contained 16 mL nutrient
medium, 234 mL activated sludge, 10 mL sodium acetate and
240 mL treated wastewater.
3. Results and discussion
3.1. Effect of various operating parameters on Fenton's oxidation
process
3.1.1. Effect of [H2O2]:[Fe2þ] molar ratio
The performance of Fenton's oxidation on CPX degradation
(initial CPX concentration ¼ 100 mg L1) was studied with H2O2 to
Fe2þ molar ratio ranging from 5 to 50 while using the stoichio-
metric H2O2 concentration (¼ 14.2 mM). Correspondingly, Fe2þ
concentration was changed from 0.284 to 2.84 mM. The experi-
ments were conducted with an initial wastewater pH of 3.0 at
ambient temperature (ca. 30  C). The maximum CPX and TOC re-
movals were 70% and 55%, respectively, after 60 min reaction at
[H2O2]:[Fe2þ] molar ratio of 10 (Fig. 1). The CPX and TOC degrada-
tion patterns could be divided into two parts: (i) fast degradation
from 0 to 15 min, and (ii) very slow or insignificant degradation
between 15 and 60 min. The wastewater was turned brownish-
yellow in colour just after adding Fenton's reagent which was
transformed into yellowish colour after 15 min of reaction. The
yellowish colour may be attributed to presence of the persistent
degradation intermediates/by-products which were detected in the
wastewater. The nature of these compounds is discussed later.
Doubling Fe2þ concentration (i.e., [H2O2]:[Fe2þ] ¼ 5) caused slight
drop in CPX degradation (Fig. 1). Excess Fe2þ ions may cause
destruction of
OH radicals according to equation (2) (Joseph et al.,
2000):
Fe2þ þ
OH/Fe3þ þ OH (2)
Furthermore, scavenging action of resulting OH ions may also
destroy the hydroxyl radicals as per equation (3).

OH þ OH/H2O þ O (3)
The results were compared with two control runs performed
with only one of the Fenton's reagent (either Fe2þ or H2O2). With
Fe2þ ions, only 6% and 5% CPX and TOC removals, respectively, were
obtained slightly lower than those with H2O2 only.
The pseudo first and second order reaction rate constants for the
fast reaction step (i.e., 0e15 min) were determined for the run
performed at the reaction conditions showed best CPX and TOC
removals (i.e., [H2O2]:[Fe2þ] ¼ 10 and initial wastewater pH ¼ 3.0).
The first and second order rate constants obtained for CPX degra-
dation were found to be 0.055 ± 0.005 min1 and
0.6 ± 0.07  103 L mg1min1, respectively while the corre-
sponding rate constants for TOC removal were found to be
0.04 ± 0.006 min1 and 1.2 ± 0.08  103 L mg1min1. The details
are provided in supporting information (Section S.3, Fig. S1 and
Table S1).
1184 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188

3.1.3. Effect of reaction temperature

The influence of temperature on CPX and TOC degradation
during Fenton's oxidation was also observed. The reaction tem-
perature was varied between ambient (30  C) to 60  C. The cipro-
floxacin, [H2O2]:[Fe2þ] and pH were 100 mg L1, 10 and 3,
respectively. Only slight enhancement in the degradation of CPX
was observed with rise in temperature (ranging from 58 to 61%)
(section S.5; Fig. S3).

3.1.4. Effect of initial CPX concentration

The effectiveness of Fenton's oxidation was also investigated by
varying initial CPX concentration (viz., 50, 100 and 200 mg L1) in
synthetic wastewater. The corresponding stoichiometric amount of
H2O2 was added to the wastewater (7.1, 14.2 and 28.4 mM,
respectively). For the runs, [H2O2]:[Fe2þ]molar ratio and waste-
water pH were selected as 10 and 3.0, respectively. It can be
observed from Fig. S4 that an increase in substrate concentration
resulted a slight decrease in percent CPX and TOC removals (CPX
removal ¼ 67.6e72.4% and TOC removal ¼ 53.2e56.8%).
One-way ANOVA statistical analysis was performed at 1% and 5%
significance levels to determine the parameters having significant
impact on CPX and TOC removals. According to the analysis,
[H2O2]:[Fe2þ] and initial reaction pH had substantial effect on CPX
and TOC degradation while CPX concentration and temperature
showed no considerable effect on the oxidation process (Tables S2
and S3, respectively).

3.2. Relation between H2O2 consumption and CPX/TOC removal

(XCPX/XH2O2 and XTOC/XH2O2)
2þ
Fig. 1. Effect of Fe :H2O2 ratio on (a) CPX removal and (b) TOC removal (Initial CPX
concentration ¼ 100 mg L1, H2O2 concentration ¼ 14.2 mM, Initial pH ¼ 3.0,
Temperature ¼ ambient). To define the efficiency of H2O2 consumption, X1 (XTOC/XH2O2)
and X2 (XCPX/XH2O2) were determined. X1 and X2 are defined as the
ratio of amount of TOC (or CPX) removal to the amount of H2O2
consumed within a specified duration (Pinho et al., 2015; Messele
et al., 2015). The change in these ratios with time is presented for
3.1.2. Effect of initial pH
the run performed at following conditions: CPX
To determine the effect of pH, Fenton's oxidation was conducted
concentration ¼ 100 mg L1, [H2O2]:[Fe2þ] ¼ 10 and pH ¼ 3.0. The
at different initial pH (ranging from 2 to 4) under ambient condi-
change in X ratios and H2O2 consumption with time followed the
tions. CPX concentration in the synthetic wastewater was
same patterns (Fig. 2). Approximately 90% of H2O2 was consumed
100 mg L1 while H2O2 to Fe2þ molar ratio was selected as 10.
within 30 min. No significant removal of CPX or TOC was obtained
The maximum CPX and TOC removals were found at an initial
beyond this period possibly due to the lack of H2O2.
reaction pH of 3.0 (i.e., 70% and 55%, respectively) (presented in
supporting information, section S.4; Fig. S2). Higher or lower initial
wastewater pH exhibited lower CPX degradation. At initial waste-
water pH values of 2.0 and 4.0, CPX degradation was 55% and 60%,
respectively while the corresponding TOC removal was 42.5% and
47.5%. After the treatment, final pH of the wastewater treated at
different initial pH was varied between 1.5 and 3.75. The drop in pH
may be attributed to the formation of inorganics (like HF, NH4OH
and HNO3) and low molecular weight organic species (such as
carboxylic acids). Short chain carboxylic acids may be formed
before total mineralization (Yahya et al., 2014).
At reaction pH higher than 3.0, the formation of insoluble iron
species [Fe(OH)þ 2 ] and dissociation of H2O2 into H2O and oxygen
might be responsible for the reduced TOC and CPX removals
(Tamimi et al., 2008). The maximum concentration of soluble Fe2þ
ions has been reported at a reaction pH of ~3.0 (Brillas et al., 2009).
The drop in efficacy of Fenton's reaction at lower pH (i.e., <3.0) can
be attributed to the stable nature of H2O2 and formation of oxonium
ions (H3Oþ

2 ) as well as the scavenging of OH radicals by H ions
þ Fig. 2. Change in X1 and X2 ratio and H2O2 concentration during Fenton's oxidation of
CPX (Initial CPX concentration ¼ 100 mg L1, H2O2 concentration ¼ 14.2 mM, Initial
(Kwon et al., 1999). pH ¼ 3.0, Temperature ¼ ambient).
 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188
3.3. Time based profile of inorganic species and residual ferrous/
ferric ions in wastewater during the reaction
The time based release of inorganic ions (i.e. NHþ  
4 , NO3 and F )
from CPX was determined during the degradation of CPX at the best
reaction conditions (i.e., [H2O2]:[Fe2þ] ¼ 10 and initial wastewater
pH ¼ 3.0) (Fig. 3). The heteroatoms present in CPX were released as
inorganic ions during the reaction. The representative reactions are
shown as follows (Yahya et al., 2014):
CPX þ
OH///Intermediates
Intermediates þ
OH///CO2 þ H2O þ NHþ 
4 /NO3 þ F

The inorganic ions were released at faster rate in first 15 min.
The concentration of NHþ  
4 eN, NO3 -N and F in the treated waste-
water was found to be 3.79, 2.28 and 1.69 mg L1, respectively, after
60 min reaction. These concentrations were ~49% and ~30% of the
initial nitrogen and fluorine in CPX (Initial N ¼ 12.7 mg L1 and
F ¼ 5.74 mg L1).
Fe2þ concentration in the wastewater solution was decreased
from an initial ~80 mg L1 to ~15 mg L1 within first 15 min (Fig. 3)
during which maximum removal of CPX (or TOC) was occurred.
Fe3þ concentration in the wastewater was also increased to
55 mg L1 during the same time interval (Fig. 3). It can be suggested
that the rate of
OH radicals production was rapid in first 15 min of
reaction after which it diminished due to the progressive depletion
of Fe2þ and H2O2. The conversion of Fe2þ into Fe3þ ions takes place
by the following reactions:
Fe2þ þ H2O2/Fe3þ þ OH þ
OH
Fe2þ þ
OH/Fe3þ þ OH
With t-Butanol (
OH scavenger) in the reaction mixture, CPX
removal from the wastewater was ~18% compared to 70% obtained
during the reaction without t-Butanol. Similarly, TOC removal from
the wastewater in the absence and presence of t-Butanol was 55%
and 12%, respectively. It implied that the hydroxyl radicals played
key role in the removal of CPX and other intermediates during
Fenton's oxidation. Addition of t-Butanol could have scavenged
OH
radicals selectively by abstracting hydrogen atom from tertiary
Fig. 3. Monitoring of residual Fe2þ and Fe3þ as well as release of F, NO
3 and NH4 ions
in during the Fenton's oxidation (Initial CPX concentration ¼ 100 mg L1, H2O2
concentration ¼ 14.2 mM, Initial pH ¼ 3.0, Temperature ¼ ambient).
1185
carbon atom as per the following equation (Cole et al., 2007):
(CH3)3COH þ
OH/CH2(CH3)2COH þ H2O (8)
3.4. Sludge quantification
Three oxidation runs for 15, 30 and 60 min duration were per-
(4)
formed in batch mode on the synthetic wastewater (CPX
concentration ¼ 100 mg L1) at the best reaction conditions (i.e.,
(5)
[H2O2]:[Fe2þ] ¼ 10 and initial pH ¼ 3.0). The sludge concentration
after the three runs for different durations (i.e., 15, 30 and 60 min)
was 280, 290 and 300 mg L1, respectively. An effort was made to
estimate the TOC adsorbed on the sludge recovered after 60 min of
reaction. According to the procedure reported by Yadav and Garg
(2014), the oven-dried sludge (¼ 15 mg) was dissolved in 3 mL
aqua-regia and subsequently diluted to 50 mL. The adsorbed TOC
on the sludge was found to be only ca. 4% of the total TOC in syn-
thetic wastewater (i.e., 2.5 mg L1).
3.5. Identification of transformation oxidation products
To predict CPX degradation by-products, the treated wastewater
samples withdrawn periodically during the Fenton's oxidation of
synthetic wastewater (initial CPX concentration ¼ 100 mg L1) at
an initial pH and [H2O2]:[Fe2þ] of 3.0 and 10, respectively, were
(6) subjected to HR LC-MS analysis. From MS/MS analysis, the details of
protonated molecular as well as fragmented ions of CPX and the
(7) oxidation products (i.e. D1eD5) were obtained which are presented
in Table S4. The time-based profiles of CPX degradation products
with time were also recorded (Fig. 4a). All the identified interme-
diate compounds could not be degraded completely even after
60 min of reaction. The maximum concentration of various iden-
tified compounds (except D5) was found after 15e20 min of reac-
tion. The possible reactions causing the degradation of CPX
molecule are shown in Fig. 4b. Decarboxylation and defluorination
are expected due to the presence of two strong electron with-
drawing substituents (i.e. carboxylic group and fluorine) in CPX.
Due to the inductive effect of electronegative fluorine atom,
piperazine ring suffers an angle stress which may lead to partial
piperazine moiety degradation (loss of ethylene group) and for-
mation of desethylene CPX (D1; m/z ¼ 305). The formation of
desethylene CPX during Fenton's oxidation of CPX has been re-
ported earlier (Giri and Golder, 2014). The complete degradation of
the piperazine moiety using various AOPs has been reported earlier
(Liu et al., 2012; Zhang et al., 2015). The complete piperazine moiety
degradation and defluorination can occur simultaneously to pro-
duce D2 and D3, respectively. The compound D2 may partially be
converted into lower molecular weight compound (i.e., D4) after
defluorination. Apart from this, the decarboxylation and hydrox-
ylation of CPX molecule can form another compound D5 (m/
z ¼ 303). It can be observed that the core quinolone structure and
cyclopropyl group remained intact in all the identified products
(i.e., D1 e D5). An et al. (2010) have also reported the formation of
D1 and D2 as end-products after heterogeneous photo-catalytic
CPX oxidation. However, some other possible by-products such as
the analogues of isatin (C8H5NO2) and anthranilic acid (C7H7NO2)
were not found. These compounds have been reported during the
þ ozonation of CPX (Dewitte et al., 2008). In short, it can be stated
that the degradation of CPX could have occurred due to a combi-
nation of series and/or parallel reactions.
1186 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188

Fig. 4. a) Relative time based profiles of CPX and various degradation products; b). Possible CPX degradation products identified by HR LC-MS analysis.
 A. Gupta, A. Garg / Chemosphere 193 (2018) 1181e1188
Fig. 5. Toxicity assessment of CPX laden wastewater after Fenton treatment (WW:
original wastewater; FTWW: Fenton treated wastewater).
3.6. Respirometric toxicity assessment
The results from the rapid toxicity tests for untreated and
treated wastewater after Fenton's oxidation are illustrated in Fig. 5.
The results indicate fast microbial utilization of the reference sub-
strate (i.e., sodium acetate) present in control (prepared in distilled
water) caused the highest oxygen uptake rate (29.67 mg L1 h1)
after 6 h. The microbial substrate utilization exhibited 67% inhibi-
tion due to toxicity of CPX. After Fenton's treatment, the waste-
water toxicity was reduced and the better microbial utilization of
the substrate was found with 39% inhibition. The oxygen uptake
rate for untreated and treated wastewater was measured to be 9.8
and 18.1 mg L1 h1 after 6 h, respectively.
4. Conclusions
The results obtained from the Fenton's treatment of CPX laden
synthetic wastewater are reported in the present study. The
maximum CPX and TOC removals of 70% and 55% were obtained at
[H2O2]:[Fe2þ] and initial pH of 10 and 3, respectively. Several series/
parallel reactions like piperazine moiety degradation, defluorina-
tion, hydroxylation and decarboxylation caused the formation of
the five degradation by-products. The drop in reaction pH during
the reaction was resulted due to the formation of short chain car-
boxylic acids, HF and HNO3 during the reaction. The fall in X ratios
and disappearance of ferrous ion from the wastewater evidenced
the degradation of CPX molecule from the beginning of reaction. On
the basis of results, it can be suggested that the majority of the
removed organic carbon was mineralized at the optimised condi-
tions. A rapid toxicity measurement test showed considerable
reduction in toxicity of the treated wastewater.
Acknowledgements
The first author is thankful to Ministry of Human Resources and
Development (MHRD), New Delhi, India for fellowship to pursue
his PhD research work. The authors would like to thank Sophisti-
cated Analytical Instrument Facility (SAIF) of Indian Institute of
Technology (IIT) Bombay, Mumbai, India for their help in per-
forming the HR LC-MS analysis of drug solutions.
Appendix A. Supplementary data
Supplementary data related to this article can be found at
1187
https://doi.org/10.1016/j.chemosphere.2017.11.046.
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