Biochemical Engineering Journal 130 (2018) 104–112

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Biochemical Engineering Journal

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

Regular article

Effects of pH, dissolved humic acid and Cu2+ on the adsorption of

norfloxacin on montmorillonite-biochar composite derived from
wheat straw
Jinghuan Zhang ∗ , Mingyi Lu, Jun Wan, Yuhuan Sun, Huixia Lan, Xiaoyan Deng
College of Environment and Safety Engineering, Qingdao University of Science and Technology, Qingdao 266042, China

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

Article history: A montmorillonite-biochar (MT-BC) composite was prepared from wheat straw heating at 400 ◦ C. Adsorp-
Received 10 August 2017 tion isotherms and effects of pH, dissolved humic acid (DHA) and Cu2+ on norfloxacin (NOR) sorption
Received in revised form to biochar and MT-BC composite were carried out. Montmorillonite modified biochar showed higher
25 November 2017
adsorption affinity for NOR. The qmax value increased from 10.58 mg g−1 to 25.53 mg g−1 after surface
Accepted 27 November 2017
Available online 2 December 2017
modified by montmorillonite. MT-BC composite had rich pore structure, with specific surface area (SSA)
and pore volume values of 112.6 m2 g−1 and 0.604 cm3 g−1 , respectively. The optimum pH range for NOR
adsorption on MT-BC composite is 5–11. The presence of DHA (20–150 mg L−1 ) greatly suppressed the
Keywords:
Montmorillonite-biochar composite NOR adsorption. The decrease of NOR sorption is attributed to the “competitive adsorption” of DHA with
Adsorption mechanism NOR and “pore blockage” by sorbed DHA. Similarly, reduced NOR sorption with Cu2+ was probably due
Norfloxacin to the competition for sorption sites on biochar surface. Compared to DHA, Cu2+ resulted to a smaller
Dissolved humic acid decrease in NOR sorption. This is because the formation of NOR-Cu2+ complexes can slightly increase
Cu2+ NOR sorption capacity. The dominant adsorption mechanisms of NOR on MT-BC composite are electro-
static interaction, H-bond and pore-filling. This study provided valuable guidance and effective method
for the removal of NOR from aquatic environments.
© 2017 Elsevier B.V. All rights reserved.

1. Introduction A prior study showed that MgO-impregnated biochar had

Norfloxacin (NOR), one of the fluoroquinolone antibiotics, was
widely used in medical and veterinary practice to treat several
infectious diseases in both human and animals [1]. About 30–90%
of the antibiotics, acted as active compounds, are stable and can
not be easily degraded, thus they can stay in the environment for a
long time [2]. High concentrations of antibiotics in dissolved phase
had been detected in surface water [3]. Antibiotics were found to be
toxic to aquatic organisms such as algae, fish, and daphnia, contam-
inate drinking water and threaten human health [4]. Thus, removal
of antibiotics from the water environment is necessary.
Biochars can be used to remove antibiotics from aquatic
environment via adsorption technology. Biochars derived from dif-
ferent sources are reported to be strong adsorbents for organic
pollutants in potable and reuse water [5–7]. Surface modified
biochar composite with high surface area exhibited stronger
adsorption than original biochar for organic pollutants [8,9].
∗ Corresponding author.
E-mail address: vickyhuan@163.com (J. Zhang).
maximum adsorption capabilities of 398 mg g−1 for phosphate,
22 mg g−1 for ammonium, and 247 mg g−1 for humate, respectively.
Struvite crystallization, electrostatic attraction, and ␲-␲ interac-
tions contributed to the adsorption enhancement [10]. It is reported
that coated minerals covered on the biochar surface modified
the pore diameter and surface functional groups, thus improv-
ing the adsorption of propiconazole on biochars [11,12]. Li et al.
also reported that attapulgite-biochar derived from potato stem
had great adsorption affinity for NOR, with the maximum qe of
5.24 mg g−1 , which is about 1.68 times higher than primary biochar
[13]. Previous results showed that biochar coated with montmo-
rillonite was a highly effective adsorbent for methylene blue and
ammonium [14,15]. It is interesting and essential to explore the
adsorption affinity of clay-biochar composite for NOR and the influ-
encing environmental factors. In our study, montmorillonite was
used to modify the surface of wheat straw- derived biochar.
Dissolved organic matter (DOM) and heavy metal are widely
existing compounds in aquatic environment, which may interact
with water, solid matrix and antibiotics. Prior studies showed that
high pH and the presence of DOM (humic acid and sodium algi-
nate) and Cu2+ greatly inhibited the sorption of sulfonamides onto

https://doi.org/10.1016/j.bej.2017.11.018
1369-703X/© 2017 Elsevier B.V. All rights reserved.
 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112
Table 1
Selected physiochemical properties of NOR.
Chemical MW (g mol−1 ) Sw (mg L−1 )
NOR 319.3 161000 (pH = 5)
400 (pH = 7)
910 (pH = 9)
Notes: MW: molecular weight; Tm : melting point; Kow : octanol–water partition coefficient; Sw : aqueous solubility.
pine wood char and graphene oxides [16,17]. This can be explained
by the competition with antibiotic molecules for surface sorption
sites. Lian et al. reported that high levels of humic acid suppressed
the adsorption of sulfonamides by crop straw-derived biochars, but
low levels of humic acid promoted the adsorption [18]. Moreover,
introduction of humic acid was able to decrease the adsorption
of NOR probably due to blockage of sorption sites by DOM [19].
Therefore, this is strongly necessary to investigate the impacts of
pH, coexisted dissolved humic acid and divalent heavy metal cation
Cu2+ on the adsorption of NOR by clay-biochar composite in aquatic
environment.
The aims of this study are to (1) prepare montmorillonite-
biochar (MT-BC) composite from wheat straw heating at 400 ◦ C,
(2) determine the adsorption isotherms and the effects of pH, dis-
solved humic acid (DHA) and Cu2+ on the adsorption of NOR on BC
and MT-BC composite, and (3) explore the adsorption mechanisms
of NOR on MT-BC composite.
2. Materials and methods
2.1. Chemicals
NOR (>98%; Aldrich Chemical Co., USA) was selected as flu-
oroquinolone antibiotics probe to determine the equilibrium
adsorption isotherms and impacts of pH, DHA and Cu2+ on sorption.
DHA (>98%) was purchased from Aldrich Chemical Company in USA
and directly used in sorption experiments. The molecular weight of
DHA was 1500–3000 g mol−1 . The C, H, N, O and ash contents of DHA
were 49.7%, 6.48%, 6.03%, 28.5% and 9.10%, respectively. Selected
physiochemical properties of NOR are given in Table 1 [20]. Stock
solutions of NOR were prepared in HPLC-grade methanol. Methanol
concentrations in the aqueous solutions were always less than 0.2%,
a level at which methanol has no measurable effect on sorption [21].
2.2. Preparation and characterization of MT-BC composite from
wheat straw
Montmorillonite-biochar composites were prepared from
wheat straw as described in a prior study [22]. First, a stable
montmorillonite suspension was prepared by adding 2.5 g of mont-
morillonite powder to 100 mL deionized water and mixed in the
ultrasonicator for 30 min. Then, 10 g of the wheat straw were
dipped into the montmorillonite suspensions and stirred for 2 h.
Finally, the wheat straw were separated from the mixture and oven
dried at 80 ◦ C. The mixture of montmorillonite and wheat straw
were heated in a muffle furnace at 400 ◦ C under a flow of N2 and
held there for 6 h. Then the biochars were washed with deionized
water, freeze-dried, passed though a 100-mesh sieve, and used for
sorption experiments. Wheat straw without montmorillonite was
observed through the same pyrolysis process. The biochars with-
out and with montmorillonite modification were denoted as BC and
MT-BC composite, respectively.
The elemental compositions (C, H, N, O) were analyzed with
a high-temperature combustion method (elementar Vario EL,
Germany). SSA was determined by N2 adsorption at −196 ◦ C
105
logKow Tm (◦ C) Chemical structure
−1.7 (pH = 5) 220
−1.0 (pH = 7)
−1.63(pH = 9)
using an ASAP 2010 system and interpreted with the Brunauer-
Emmet-Teller (BET) model. The FTIR spectra were recorded in the
transmission mode by a NEXUS 670X spectrophotometer using
KBr pellet. The 13 C NMR spectrum was acquired on a Bruker AU-
300 spectrometer using cross-polarization magic angle spinning
(CP/MAS) techniques.
2.3. Sorption experiments
Adsorption isotherms of NOR on BC, MT-BC composite and
montmorillonite were conducted in replicates in 50 mL glass tubes
with Teflon-lined caps. 0.01 mol L−1 CaCl2 and 200 mg L−1 NaN3
were added to stock solutions to minimize biological activity [23].
The ratios of water to solids (40 mL: 50 mg) were adjusted to
achieve 30–80% sorption of NOR. Our preliminary adsorption kinet-
ics experiments showed that apparent sorption equilibrium was
reached in less than 48 h (Fig. S1). The vials with adsorbents and
initial NOR solutions (0.4–15 mg L−1 ) were shaken at 150 rpm at
25 ± 0.5 ◦ C for 48 d and then centrifuged at 4500 rpm for 30 min
to separate solid and aqueous phases. NOR in the supernatant was
filtered through 0.45 ␮m nylon membrane and analyzed by high
performance liquid chromatography (HPLC). The sorbed amounts
were computed from the difference of the initial and final solute
concentrations. Control reactors prepared similarly but with no sor-
bent were run simultaneously for assessing loss of solute to reactor
during sorption. Results showed that average system losses were
consistently less than 4% of the initial concentration, indicating that
microbial degradation and volatilization during sorption and the
uptake by the glass walls were negligible.
The effect of pH on the adsorption of NOR was determined by
varying the pH from 3.0 to 11.0 while containing 50 mg of adsorbent
and the initial NOR concentration at 10 mg L−1 under 25 ◦ C. The ini-
tial pH values of NOR solutions were adjusted using 0.1 mol L−1 HCl
or NaOH solutions. The impacts of DHA and Cu2+ on the adsorption
of NOR on BC and MT-BC composite were carried out at pH = 7.0
with adding concentrations of 20–150 mg L−1 . Sorption isotherms
of DHA on BC and MT-BC composite were also analyzed at 25 ◦ C.
Two replicates for each point were used in all sorption experiments.
2.4. Analytical methods
Norfloxacin concentrations were analyzed by reversed-phase
HPLC, using a Waters 484 HPLC (Waters, America) with a UV detec-
tor [19]. A Diamon-sil 5U C18 column (4.6 mm × 250 mm, Dikma
technologies) was used. Isocratic elution was performed at a flow
rate of 1.0 mL min−1 using a mixed solution of 0.025 mol L−1 phos-
phoric acid-acetonitrile (80:20, V/V) as the mobile phase. The UV
detection wavelength was set at 278 nm. The retention time for
NOR was 3.8 min. The concentration of humic acid was quantified
by UV absorption at 254 nm as described in the reference of Lin
et al. [24].
2.5. Data analysis
Both Langmuir and Freundlich models were used to quantify
the adsorption of NOR on BC and MT-BC composite. The Lang-
muir model indicates a monolayer sorption of a target compound
106 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112

Fig. 1. SEM images of BC (a) and MT-BC (b).

Table 2
Elemental composition and SSA of BC and MT-BC composite.

Sample elemental composition (%) SSA (m2 g−1 ) Pore volume Dp Clay content(%)
a a a a 3 −1 b
C H N O H/C O/C C/N (N + O)/C (cm g ) (nm)

BC 52.4 10.3 2.03 25.2 2.36 0.36 30.1 0.39 20.1 0.138 9.85 –
MT-BC 58.8 1.16 1.85 27.0 0.24 0.34 37.1 0.37 112.6 0.604 21.7 8.91
MT – – – – – – – – 148.3 0.202 48.2 –
a
Atomic ratio.
b
Dp is average pore diameter calculated from N2 adsorption isotherm by the Berret-Joyner-Halenda method.

on a homogenous surface, but the Freundlich model illustrates a
multi-layer adsorption process [25]. The fitting curves and statisti-
cal analysis were processed using SigmaPlot 12.5.
Langmuirmodel :qe = (qmax ·K L ·C e )/(1 + K L ·C e )
where Ce is the liquid phase concentration in mg L−1 , qe is the solid
phase concentration in mg kg−1 ,
KL is the Langmuir equilibrium constant (L mol−1 ), qmax is the
Langmuir maximum adsorption capacity in mg kg−1 .
Freundlichmodel :qe = K f ·C e n
where Kf is the Freundlich sorption capacity-related parameter
[(mg kg−1 )/(mg L−1 ) n )], n is the isotherm nonlinearity index.
However, a precise comparison cannot be made according to the
Kf values because of their different units as a result of nonlinear-
ity. The concentration dependent sorption capacity coefficient Kd
(L g−1 ) at three selected concentration (Ce = 0.5, 1.5, and 5.0 mg L−1 )
were employed to compare the sorption capacity. The Kd value is
calculated using the formula:
K d = K f ·C e n−1
In general, the Kd value decreased as a function of Ce because of
isotherm nonlinearity.
3. Results and discussion
ratio indicates a higher aromatic structure of MT-BC composite. The
BC and MT-BC composite have polarity index (N + O)/C of 0.40 and
0.37. The SSA and pore volume values of BC were 20.1 m2 g−1 and
0.138 cm3 g−1 , respectively, which was lower than that of bamboo
(1)
char [27], but slightly higher than that of zein-derived biochar [28].
Surface treated by montmorillonite greatly increased the SSA and
pore volume values to 112.6 m2 g−1 and 0.604 cm3 g−1 , respectively.
The FTIR spectra of BC and MT-BC composite are presented
in Fig. 2. As shown in Fig. 2, the broad band observed at about
3400 cm−1 is assigned to the stretching vibration of hydroxyl
(2) groups ( OH). Peaks at 2925 cm−1 and 2850 cm−1 are ascribed to
aliphatic −CH2 asymmetric and symmetric stretching and C H
band, respectively [29]. These peaks in MT-BC composite became
a decrease than those in the FTIR spectrum of BC. The sharp and
strong band at 1640 cm−1 can be assigned to aromatic C C stretch-
ing [30]. The sharp peak at 660 cm−1 is associated with mineral
compounds such as carbonates and phosphates. The absorbance
at 1190 cm−1 for MT-BC composite is the C H stretching and
OH deformation of COOH or C O stretching of aryl esters, which
was considered larger than that of BC. Introduction of montmoril-
(3) lonite increased the surface oxygen-containing functional groups
of biochar.
3.2. Effect of pH on the adsorption of NOR on BC and MT-BC
composite

3.1. Characteristics of BC and MT-BC composite
The SEM images in Fig. 1 shows the different surface morphol-
ogy of BC (a) and MT-BC composite (b). The BC sample had porous
structure (Fig. 1a). Fig. 1b indicates that montmorillonite particles
were deposited on the biochar surface.
In Table 2, the C and O content of BC is 52.4% and 25.2%, which
is close to the C content of rice straw-derived biochar, but lower
than that of pine wood-derived biochar [26]. The MT-BC composite
had higher C and O content of 58.8% and 27.0% than original BC.
The H/C and O/C atomic ratios are 2.36 and 0.36 for BC, and 0.24
and 0.34 for MT-BC composite, respectively. The lower H/C atomic
NOR has two proton-binding sites (carboxyl and piperazinyl
group) with its acid dissociation constants pKa values of 6.22 and
8.51, respectively. Fig. 3 shows the molecular structure of NOR and
its ionic forms as a function of pH and pKa values [31]. NOR can
exist in cationic form, zwitterionic/neutral form, or anionic form
depending on the solution pH. The effects of pH on the adsorption
of NOR to BC, MT-BC composite and montmorillonite are given in
Fig. 4. Montmorillonite modified biochar exhibited greater adsorp-
tion affinity than original biochar for NOR at different pH (3–11).
This can be explained by the increase of SSA and pore volume shown
in Table 2. Modification by montmorillonite may lead to the forma-
tion of new pores and plenty of reactive sites on biochar surface.
 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112
Fig. 2. FTIR spectra analysis of BC (a) and MT-BC (b).
Fig. 3. Molecular structure of NOR and its ionic forms as a function of pH and pKa values.
Fig. 4. Effects of pH on the adsorption of NOR on BC, MT-BC composite and mont-
morillonite.
The qe for MT-BC composite enhanced 2.29–2.60 times higher than
BC with the initial NOR concentration of 10 mg L−1 . Compared to
BC and MT-BC composite, montmorillonite had lower adsorption
capacity for NOR with qe of 0.244–2.553 mg g−1 .
As shown in Fig. 4, the maximum NOR sorption capacity on BC
was observed at pH = 7.0. The NOR adsorption on MT-BC composite
enhanced with pH ranging from 3.0 to 5.0. When the pH increased
to 5.0, the adsorption stayed high and steady with the maximum
amount at pH = 6.5. This is closely associated with the pH-regulated
distribution of the deprotonated species of NOR. The point of zero
charge (pzc) describes the condition when the electrical charge
107
density on a surface is zero. The pH of zero point charges (pHpzc) of
BC and MT-BC composite was determined by titration. The pHpzc of
BC and MT-BC composite is 8.86 and 6.92, respectively. The surface
of biochar samples was positively charged at pH lower than pHpzc
and negatively charged at pH higher than pHpzc. At pH<5.0, the
adsorption was depressed, because the NOR molecules and MT-BC
had the same charge and can repel each other. At pH>5.0, the NOR
molecules and MT-BC had opposite charges, which results in the
enhancement of adsorption. Thus, electrostatic attraction between
charged NOR and biochar surface was expected to be essential in
the adsorption process.
At pH = 6.22–8.51, NOR existed in zwitterioinic/neutral form.
The zwitterioinic/neutral form was much more hydrophobic than
the anionic or cationic form according to the Sw and logKow of NOR
presented in Table 1. The rich aromatic structure of biochar sam-
ples provided more hydrophobic sites, which can interact with NOR
molecules by hydrophobic effect. This suggested that hydrophobic
effect was an important factor determining the increase of NOR
adsorption. Yang et al. also reported that the adsorption of NOR to
porous resins and carbon nanotubes enhanced with the increase of
zwitterionic form, with the highest sorption amount at pH = 7 [20].
Our results showed that the optimum pH range for NOR adsorption
on MT-BC composite is 5–11. However, Wang et al. reported that
the effect of pH on the adsorption of NOR on activated magnetic
biochar derived from corn stalks, reed stalks and willow branches
was not obvious [1]. Good adsorption affinity of NOR was observed
in a wide pH range of 2.0–10.0. It was also reported that more than
80.1% of NOR was removed by a clay-biochar composite from aque-
ous solution at 2.0–11.0, with the maximum amount at pH = 3.0
[13].
108 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112

Fig. 5. Effects of DHA on the adsorption of NOR on BC (a) and MT-BC composite (b).

3.3. Effects of DHA on the adsorption of NOR on BC and MT-BC

composites

Sorption isotherms and fitting parameters of NOR on BC and MT-

BC composite with and without DHA are given in Fig. 5 and Table 3.
Data points and fitting curves in Fig. 5 shows that sorption of NOR
was fitted better by the Langmuir model than Freundlich model.
The Langmuir model fitting results revealed that the monolayer
sorption was the main adsorption mechanism of NOR on BC and
MT-BC.
Montmorillonite modified biochar greatly increased the sorp-
tion of NOR with and without DHA. The Kd values of
NOR (0.5 mg L−1 < Ce < 5.0 mg L−1 ) on MT-BC composite were of
0.578–11.42 L g−1 , about 3.4–6.4 times higher than BC. More-
over, the qmax value for MT-BC composite with DHA enhanced
2.3–4.5 times higher than BC. As shown in Fig. 5 and Table 3,
the presence of DHA reduced the sorption of NOR on BC and
MT-BC composite. The qmax value reduced from 10.58 mg g−1 to Fig. 6. Sorption isotherms of DHA on BC and MT-BC composite.
1.586 mg g−1 for BC, and from 25.53 mg g−1 to 6.968 mg g−1 for MT-
BC composite, with DHA concentration increasing from 0 mg L−1
on BC and MT-BC composite depends on the extent and competi-
to 150 mg L−1 , respectively. Furthermore, when the concentration
tion of each interaction. To testify the hypothesis, the adsorption of
of DHA was 150 mg L−1 , the Kd value (at Ce = 0.5 mg L−1 ) decreased
DHA on BC and MT-BC composite was carried out. Notably, BC and
from 2.833 L g−1 to 0.213 L g−1 for BC, and from 11.42 L g−1 to
MT-BC composite showed high adsorption for DHA in Fig. 6. The
1.497 L g−1 for MT-BC composite, respectively. Moreover, negative
sorbed amounts of DHA are 5.822 mg g−1 and 9.445 mg g−1 on BC
correlations were found between qmax and Kd values of NOR and
and MT-BC composite, respectively, when the initial concentration
DHA concentration (R2 ≥ 0.936) (Fig. 8). Reduced NOR sorption with
of DHA is 200 mg L−1 . Therefore, the reduced NOR adsorption was
DHA was probably due to the fact that coexisted DHA may block the
mainly attributed to the competitive adsorption of DHA with NOR
adsorption sites on modified biochar surface. Peng et al. found that
and the pore blockage by DHA.
humic acid (50 mg L−1 ) showed a depressed effect on the adsorp-
tion of NOR onto titanium oxide [19]. Prior studies also showed
that the presence of low weight DOM was found to significantly 3.4. Effects of Cu2+ on the adsorption of NOR on BC and MT-BC
suppress the sorption of sulfonamides on wood-based biochar and composites
graphene oxides [17,32]. However, suspended carbon nanotubes
exhibited enhanced sorption of sulfamethoxazole in the presence Fig. 7 shows the impacts of Cu2+ with adding concentrations
of dissolved and coated humic acid [33]. Citric acid and malic acid of 20–150 mg L−1 on the adsorption of NOR on BC and MT-BC
(0–100 mmol L−1 ) also increased the adsorption of sulfamethoxa- composite. All sorption isotherms of NOR were nonlinear. The n
zole on crop straw biochar more than 5 times, which was attributed values of NOR with and without Cu2+ were of 0.446–0.648 for
to the elevated microporosity of biochar after treated by low weight BC, and of 0.500–0.611 for MT-BC composite, respectively. Com-
DOM [34]. pared to BC, MT-BC composite exhibited higher sorption for NOR
Hypothetically, the impact of DHA on NOR sorption may be with and without Cu2+ . As shown in Fig. 7, the adsorption of NOR
explained by the following mechanisms [35,36]: (1) competitive on BC and MT-BC composite greatly reduced in the presence of
sorption between NOR and DHA for sorptive sites on biochar sur- Cu2+ . With Cu2+ concentration of 150 mg L−1 , the qmax and Kd value
face, resulting in adsorption reduction; (2) coexisted DHA may (at Ce = 1.5 mg L−1 ) decreased from 10.58 mg g−1 and 1.703 L g−1 to
block the pores on biochar surface to decrease NOR adsorption; 1.779 mg g−1 and 0.350 L g−1 for BC, and from 25.53 mg g−1 and
and (3) partition of NOR to sorbed DHA on biochar surface, which 6.594 L g−1 to 8.076 mg g−1 and 1.878 L g−1 for MT-BC composite,
leads to adsorption enhancement. The apparent sorption of NOR respectively. Also, there were negative correlations between Cu2+
concentration with qmax and Kd values of NOR (R2 ≥ 0.968) (Fig. 8).
 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112 109

Table 3
Freundlich and Langmuir sorption parameters of NOR on BC and MT-BC composite in the presence of DHA and Cu2+ .

Coexisting Freundlich model Langmuir model

pollutants (mg/L)

Kd c (L g−1 )

n Kf R2
N a
Ce = 0.5 mg L−1 Ce = 1.5 mg L−1 Ce = 5.0 mg L−1 qmax (mg g−1 ) KL (L mol−1 ) R2 Na

BC
Control 0 0.637 ± 0.004b 2.055 ± 0.005 0.985 22 2.833 1.703 0.975 10.58 ± 0.02 0.194 ± 0.003 0.986 22
DHA 20 0.584 ± 0.005 1.213 ± 0.004 0.983 22 1.581 1.039 0.656 9.326 ± 0.03 0.120 ± 0.002 0.998 22
50 0.605 ± 0.006 0.959 ± 0.004 0.973 22 1.346 0.787 0.437 4.506 ± 0.007 0.219 ± 0.003 0.986 22
100 0.503 ± 0.003 0.744 ± 0.005 0.940 22 1.092 0.594 0.305 2.759 ± 0.004 0.300 ± 0.004 0.975 22
150 0.476 ± 0.004 0.167 ± 0.003 0.943 22 0.213 0.145 0.094 1.586 ± 0.005 0.092 ± 0.002 0.962 22
Cu2+ 20 0.618 ± 0.005 1.538 ± 0.004 0.990 22 2.052 1.299 0.787 9.536 ± 0.008 0.171 ± 0.002 0.992 22
50 0.511 ± 0.003 1.120 ± 0.005 0.990 22 1.472 0.953 0.593 7.586 ± 0.006 0.142 ± 0.001 0.995 22
100 0.446 ± 0.005 0.962 ± 0.004 0.969 22 1.408 0.769 0.397 3.369 ± 0.004 0.347 ± 0.004 0.980 22
150 0.648 ± 0.006 0.433 ± 0.003 0.920 22 0.623 0.350 0.186 1.779 ± 0.005 0.266 ± 0.004 0.980 22
MT-BC composite
Control 0 0.500 ± 0.003 8.076 ± 0.007 0.990 22 11.42 6.594 3.612 25.53 ± 0.11 0.474 ± 0.003 0.991 22
DHA 20 0.448 ± 0.004 4.995 ± 0.006 0.980 22 6.541 4.266 2.671 19.94 ± 0.07 0.252 ± 0.003 0.994 22
50 0.361 ± 0.003 3.674 ± 0.004 0.969 22 4.916 3.099 1.869 15.50 ± 0.04 0.204 ± 0.004 0.985 22
100 0.489 ± 0.005 2.599 ± 0.005 0.980 22 3.620 2.141 1.204 11.45 ± 0.01 0.252 ± 0.004 0.983 22
150 0.443 ± 0.003 1.124 ± 0.004 0.960 22 1.497 0.951 0.578 6.968 ± 0.005 0.158 ± 0.002 0.976 22
Cu2+ 20 0.611 ± 0.005 6.590 ± 0.005 0.970 22 9.662 5.268 2.711 22.64 ± 0.02 0.488 ± 0.005 0.975 22
50 0.580 ± 0.004 6.053 ± 0.003 0.960 22 9.426 4.671 2.164 17.23 ± 0.01 0.772 ± 0.007 0.970 22
100 0.522 ± 0.003 3.625 ± 0.005 0.975 22 5.166 2.947 1.593 13.95 ± 0.009 0.315 ± 0.004 0.987 22
150 0.587 ± 0.005 2.355 ± 0.004 0.955 22 3.465 1.878 0.961 8.076 ± 0.005 0.361 ± 0.005 0.987 22
a
Number of data.
b
Standard deviation.
c
the concentration dependent sorption capacity coefficient Kd = Kf Ce n−1 .

Fig. 7. Effects of Cu2+ on the adsorption of NOR on BC (a) and MT-BC composite (b).

The apparent decrease of NOR sorption with Cu2+ may be due to the
“competitive adsorption” associated with sorbed Cu2+ on modified
biochar surface.
A prior study also showed that Cu2+ and DHA inhibited the
adsorption of sulfonamides to pine wood biochars attributed to
the competition for adsorption sites on biochar surface [16]. Kong
et al. reported that the presence of divalent cations (Cu2+ , Zn2+ ,
and Mg2+ , 0–20 mmol L−1 ) prominently suppressed the adsorption
of NOR by paddy and red soils [37]. Moreover, incorporation of
Sn4+ weakened the adsorption ability of NOR on Mg-Al layered
double hydroxides due to the decrease of the SSA [32]. However,
Huang et al. (2017) reported that Cu2+ could significantly enhance
the adsorption of tetracycline on Fe3 O4 @SiO2 -Chitosan/graphene
oxide (MSCG) nanocomposite [38]. The highest adsorption capac-
ity of tetracycline increased about 2.72 times with Cu2+ . The
major mechanism is that the Cu2+ acts as a bridge between
tetracycline and MSCG, and tetracycline-Cu2+ complexes were
formed, which significantly improve the adsorption of tetracy-
cline.
In our study, complexes may also be formed in the BC-Cu2+ -NOR
ternary system. Compared to DHA, the presence of Cu2+ resulted in
a smaller decrease in NOR sorption. With the adding concentra-
tion of 100 mg L−1 , the qmax and Kd values (at Ce = 5.0 mg L−1 ) of
NOR on MT-BC composite were 13.95 mg g−1 and 1.593 L g−1 for
Cu2+ , and 11.45 mg g−1 and 1.204 L g−1 for DHA, respectively. This
can be explained by the formation of NOR-Cu2+ complexes, which
can slightly increase NOR sorption capacity [39].
3.5. Adsorption mechanisms
According to the prior reported results, the adsorption mech-
anisms of antibiotics onto carbon materials mainly involved elec-
trostatic interaction, hydrophobic effect, H-bond, ␲-␲ interaction,
surface precipitation, pore-filling and partition to non-carbonized
fractions [40,41]. In our study, the main mechanisms of NOR on
MT-BC composite may be explained in Fig. 9. FTIR spectra analy-
sis indicated that the −OH, C O, C H, −COOH bonds were covered
on the surface of MT-BC composite. After modified by montmoril-
110 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112

Fig. 8. Correlations of qmax and Kd values (Ce = 1.5 mg L−1 ) of NOR on BC and MT-BC composite with the concentration of Cu2+ and DHA.

Fig. 9. Adsorption mechanisms of NOR onto MT-BC composite with DHA and Cu2+.
 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112
Fig. 10. The relationship between the adsorption capacity of MT-BC composite for
NOR and the number of regeneration cycles.
lonite, the amount of surface oxygen-containing functional groups
greatly increased. It was reported that NOR molecules consisted of
several moieties, which can be served as acceptors or donors for
H bond. These moieties may interact with the functional groups
on the surface of carbon materials to form H-bond, and greatly
improved the adsorption affinity for NOR [20]. In our study, the
enhancement of the NOR adsorption was mainly due to the H-band
formation between NOR and oxygen-containing groups in MT-BC
composite. Thus, H-bond is one of the main mechanisms of NOR on
MT-BC composite as well as electrostatic attraction.
In addition to H-bond, the pore-filling effect also contributed to
the adsorption of NOR on MT-BC composite. Prior studies showed
that pore-filling was the major factor for the great adsorption of
antibiotics such as tetracycline and NOR onto biochars derived from
solid digestate and corn stalks [1,8]. Other carbon materials, such
as porous resins, carbon nanotubes and animal hairs-based acti-
vated carbon were also reported to have strong adsorption capacity
for NOR mainly due to micropores [20,42]. Granular activated car-
bon with high surface area and pore volume of 852 m2 g−1 and
0.671 cm3 g−1 , respectively, can improve the removal efficiency of
NOR and ciprofloxacin from water. It is believed that high SSA and
favorable pore characters were the main reasons for the removal
of antibiotics by adsorption technology [43]. The MT-BC composite
had a rich microporous structure with a large SSA and pore vol-
ume, and showed a strong adsorption capacity for NOR. Thus, like
electrostatic interaction, H-bond and pore-filling effect were the
main mechanisms for NOR adsorption on MT-BC composite. Each
interaction provides different contributions at different pH value.
At pH < 5, electrostatic attraction between negatively charged NOR
and biochar surface was the main adsorption mechanism. At pH > 5,
H-bond between NOR molecules and oxygen-containing groups,
and pore-filling were the major reason for the high NOR adsorption.
3.6. Regeneration of MT-BC composite
Methanol solution was selected as the washing agent for the
removal of NOR from the adsorbent with a three-stage washing
procedure reported in a prior study [25]. H-bond between NOR and
methanol was stronger, which facilitated the removal of NOR. The
consecutive sorption processes were conducted for five times to
determine the regeneration and reuse value of MT-BC composite.
The adsorption capacity of MT-BC composite for NOR according to
the number of regeneration cycles is given in Fig. 10. Fig. 10 shows
that the stability and reusability of MT-BC composite was good.
The adsorption capacity of MT-BC composite for NOR was steady
along with the time for reuse. Futhermore, most of NOR could be
111
Fig. 11. The relationship between removal percentage of NOR from wastewater by
MT-BC composite and the number of regeneration cycles.
removed from MT-BC composite by methanol. The removal per-
centage of NOR from MT-BC composite were 86.8%, 88.1%, 87.2%,
87.9%, and 86.5% with the number of regeneration cycle from one
to five, respectively. As a result, MT-BC composite prepared in this
study had the potential to be recycled and reused for the removal
of NOR from wastewater.
3.7. Application of MT-BC composite in the removal of NOR from
wastewater
The application of MT-BC composite was evaluated for the
removal of NOR from two wastewater samples obtained from Bohai
Bay in China. It is reported that the concentrations of NOR in
polluted water of Bohai Bay were 0.003–6.8 ␮g L−1 [44]. In our
study, NOR concentrations in the two wastewater samples were
115 ng L−1 for NO.1 and 367 ng L−1 for NO.2. The relationship
between removal percentage of NOR and the number of regen-
eration cycles is listed in Fig. 11. Fig. 11 reveals that the removal
percentage of NOR from wastewater by MT-BC composite was
85.5–87.2% for NO.1 and 88.3–90.0% for NO.2. Therefore, the MT-
BC composite had good stability and reusability for the removal of
NOR from polluted water in Bohai Bay, China.
4. Conclusion
This study reveals that montmorillonite modified biochar had
higher SSA and pore volume than original wheat straw biochar.
Compared to biochar, MT-BC composite exhibited greater adsorp-
tion capacity for NOR. The qmax and Kd values for MT-BC composite
increased 2.41 times and 3.70–3.96 times, respectively. MT-BC
composite had a high adsorption capacity for NOR in a wide pH
range of 5–11. The presence of DHA and Cu2+ (20–150 mg L−1 )
reduced the adsorption of NOR on BC and MT-BC composite. Com-
petitive adsorption and pore blockage are the major reasons for
the reduction of NOR sorption. At the same concentration, DHA
leads to a higher decrease in NOR sorption than Cu2+ . Electrostatic
interaction, H-bond and pore-filling are the dominant adsorption
mechanisms of NOR on MT-BC composite surface. Our results
showed that both DHA and Cu2+ can inhibit the adsorption affinity
for NOR on BC and MT-BC composite, therefore, effectively making
NOR more mobile in the environment especially for that with high
input of chars.
Acknowledgements
The study was jointly supported by the Natural Science Foun-
dation of Shandong Province in China (No. ZR2016BB38; No.
112 J. Zhang et al. / Biochemical Engineering Journal 130 (2018) 104–112
ZR2013DQ002) and the Project of QingDao Minsheng Science and
Technology (No. 14-2-3-70-nsh).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at https://doi.org/10.1016/j.bej.2017.11.018.
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