Combination of hydrophobic effect and electrostatic interaction in imprinting for achieving efficient recognition in aqueous media

Article abstract of DOI:10.1016/j.carbpol.2009.09.010

An acifluorfen-imprinted polymer (P₁) was prepared by the combined use of bismethacryloyl-β-cyclodextrin (BMA-β-CD) and 4-vinylpyridine (4-VP) as functional monomers. Compared with the molecularly imprinted polymers (MIPs) using only BMA-β-CD or 4-VP as a functional monomer, P₂ and P₃, respectively, P₁ showed higher binding capacity (C_p) and imprinting factor (IF) for acifluorfen in aqueous media. Scatchard plot analysis revealed that two classes of binding sites were formed in the imprinted polymer P₁ with dissociation constants of 0.435 and 0.868 μmol/ml, respectively. The results of competitive binding experiments showed that P₁ can separate acifluorfen from its structural analogs. The study indicated that the combination of hydrophobic effect and electrostatic interaction in imprinting process is a feasible approach for achieving molecular recognition in aqueous media.

Full text of DOI:10.1016/j.carbpol.2009.09.010

Carbohydrate Polymers 79 (2010) 642–647
Contents lists available at ScienceDirect
Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol
Combination of hydrophobic effect and electrostatic interaction in imprinting
for achieving efficient recognition in aqueous media
*
Zhifeng Xu , Daizhi Kuang, Yonglan Feng, Fuxing Zhang
Department of Chemistry and Material Science, Hengyang Normal University, Key Laboratory of Functional Organometallic Materials of Hengyang Normal University,
College of Hunan Province, Hengyang 421008, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 30 June 2009
Received in revised form 29 July 2009
Accepted 14 September 2009
Available online 18 September 2009
An acifluorfen-imprinted polymer (P1) was prepared by the combined use of bismethacryloyl-b-cyclodex-
trin (BMA-b-CD) and 4-vinylpyridine (4-VP) as functional monomers. Compared with the molecularly
imprinted polymers (MIPs) using only BMA-b-CD or 4-VP as a functional monomer, P2 and P3, respec-
tively, P1 showed higher binding capacity (Cp) and imprinting factor (IF) for acifluorfen in aqueous media.
Scatchard plot analysis revealed that two classes of binding sites were formed in the imprinted polymer
P1 with dissociation constants of 0.435 and 0.868 lmol/ml, respectively. The results of competitive bind-
Keywords:
Molecular imprinting
Acifluorfen
b-Cyclodextrin
Molecular recognition
ing experiments showed that P1 can separate acifluorfen from its structural analogs. The study indicated
that the combination of hydrophobic effect and electrostatic interaction in imprinting process is a feasi-
ble approach for achieving molecular recognition in aqueous media.
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
Whitcombe, & Vulfson, 2000; Urraca, Moreno-Bondi, Hall, &
Sellergren, 2007), have been investigated during the past few years.
Molecular imprinting technique is one of the most promising
methodologies for synthesizing artificial receptors and can be
extensively used in chromatographic separation (Baggiani, Bara-
valle, Giraudi, & Tozzi, 2007; Haginaka, 2002), solid-phase extrac-
tion (Puoci, Garreffa, Iemma, & Muzzalupo, 2005; Zhu, Yang, Su,
Cai, & Gao, 2005), catalysis (Matsui, Nicholls, Karube, & Mosbach,
1996; Meng, Yamazaki, & Sode, 2004), binding assays and sensor
(Hsu & Yang, 2008; Kröger, Turner, Mosbach, & Haupt, 1999). So
far, most of the reported molecularly imprinted polymers (MIPs)
only function in organic solvent. Reports on imprinted polymers,
which can function in aqueous systems, are still limited. The main
barrier preventing imprinting in aqueous media, is due to the nat-
ure of hydrogen bonding. Hydrogen bonding is the most commonly
exploited interaction for the non-covalent molecular imprinting.
However, hydrogen bonding interactions between template and
functional monomers are easily destroyed in aqueous media be-
cause aqueous solvents can compete with the template for the
functional monomers. Previous studies have shown that an
imprinted polymer prepared based on hydrogen bonding interac-
tions lost its molecular recognition ability in aqueous systems or
organic solvents with high polarity (Xu, Liu, & Deng, 2006; Yu &
Mosbach, 2000). In order to achieve effective imprinting in aqueous
solution, different methods, such as metal ion mediated imprinting
Recently, b-cyclodextrin(b-CD) and its derivatives have been chosen
as functional monomers for making MIPs, and the synthesized MIPs
were successfullyusedto separate some biological compounds, such
as steroids (Hishiya, Shibata, Kakazu, Asanuma, & Komiyama, 1999),
antibiotics (Asanuma, Akiyama, Kajiya, Hishiya, & Komiyama,
2001; Hishiya, Akiyama, Asanuma, & Komiyama, 2002), peptides
(Akiyama, Hishiya, Asanuma, & Komiyama, 2001), cholesterols
(Asanuma, Kakazu, Shibata, Hishiya, & Komiyama, 1998) and other
compounds (Xu, Xu, Kuang, Zhang, & Wang, 2008) in aqueous
solution. The main strategies to exploit b-CD and its derivatives in
molecular imprinting are as follows: for those comparatively big
templates, several b-CD molecules are assembled around the tem-
plate and each b-CD molecule accommodates a part of the template,
so that the assembled b-CD molecules can work as a whole to recog-
nize the template precisely (Asanuma et al., 1998; Hishiya et al.,
1999; Xu et al., 2008). When making MIPs for those templates whose
main body could be accommodated in the cavity of b-CD, b-CD and
another compound are used as combinatorial functional monomers
(Piletsky, Andersson, & Nicholls, 1999).
Acifluorfen is an effective and selective herbicide for control
most broad-leaved weeds. The widespread use of acifluorfen has
lead to increased food, soil, and water pollution by its residues. It
is important to develop analytic methods for determining its pres-
ent in environment at low levels. In this study, the utilization of
b-CD as functional monomer in molecular imprinting is further
investigated. Combining the features of b-CD and molecularly
imprinting method, a MIP was fabricated using acifluorfen as the
(Wu
& Li, 2003), stoichiometric imprinting (Lübke, Lübke,
* Corresponding author. Tel.: +86 734 8486630; fax: +86 734 8486629.
E-mail address: xuzhifenghw@163.com (Z. Xu).
0144-8617/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carbpol.2009.09.010

Z. Xu et al. / Carbohydrate Polymers 79 (2010) 642–647
643
template, 4-vinylpyridine (4-VP) and bismethacryloyl-cyclodextrin
(BMA-b-CD) as functional monomers. To the best of our knowledge
this is the first time a MIP designed for recognition of acifluorfen in
aqueous media. In order to gain more insight into the origin of the
recognition ability of the imprinted polymer, two referenced MIPs
using only BMA-b-CD or 4-VP as a functional monomer were also
prepared. The recognition abilities and binding characteristics of
the synthesized polymers were evaluated by equilibrium binding
experiments and competitive binding experiments.
substitutional isomers with 2.0 methacryloyl substitutions per b-
cyclodextrin determined from the elemental analysis (Anal. Calcd.
for BMA-b-CD: C 46.09, H 5.99%; found: C 46.31, H 5.67%). FT-IR
(KBr) m
(cmꢁ1): 3600–3000, 2928, 1726, 1635, 1489, 1406, 1154,
1080, 1031, 756, 683, 579.
2.3. 1H NMR experiments
1H NMR experiments were used to study the interactions be-
tween b-CD and acifluorfen. The 1H NMR spectra were recorded
in DMSO-d6 using TMS as the internal standard. The concentration
of the acifluorfen or b-CD unitary solution in DMSO-d6 was kept at
0.02 mmol mlꢁ1. The initial b-CD and acifluorfen concentrations of
the mixed solution were 0.02 mmol mlꢁ1 also.
2. Experimental
2.1. Materials and instruments
b-CD and 2,2-azobisisobutyronitrile (AIBN) were purchased
from Shanghai Chemical Plant (Shanghai, China). AIBN was recrys-
tallized from methanol and b-CD was recrystallized from water
and dried under vacuum at 110.0 °C for 24 h. Acifluorfen, fomesa-
fen were obtained from Shandong Shenxing Pharmaceutical Indus-
try Limited Company (Qingzhou, China). Ibuprofen was obtained
from South Hospital (Guangzhou, China). Ethylene dimethacrylate
(EDMA) was obtained from Jiangsu Anli Chemical Plant (Suzhou,
China), it was distilled under vacuum after being extracted with
10% sodium hydroxide and dried over anhydrous magnesium sul-
fate. 4-VP was purchased from Acros (NJ, USA) and was distilled
under vacuum to remove the inhibitor. DMSO and pyridine were
purchased from Guangzhou Chemical Plant (Guangzhou). Before
use, they were dried by molecular sieve 3A and distilled under vac-
uum. DMSO-d6 was purchased from Cambridge Isotope Laborato-
ries Inc. (USA). HPLC grade methanol was purchased from
Tianjing Damao Chemical Plant (Tianjing, China). Water was dou-
2.4. Polymer synthesis
The components of the reaction mixtures for making imprinted
(P1) and non-imprinted (NP1) polymers are listed in Table 1. P1 was
prepared by the ‘‘one-step method” (Hishiya et al., 1999). The syn-
thesis procedure of P1 is as follows: BMA-b-CD and acifluorfen
were dissolved in DMSO in a 50 ml glass ampoule. After being
magnetically stirred at room temperature for 2.0 h, 4-VP, EDMA
and AIBN were added. After degassing and nitrogen purging, the
ampoule was sealed under vacuum and the mixture was kept in
a water bath at 60 °C for 36 h. The resultant rigid polymer was
ground and passed through a 90 lm sieve. Fine particles were re-
moved by repeated sedimentation in acetone. The obtained parti-
cles were Soxhlet extracted with a mixture of methanol–acetic
acid (9:1, v/v) until acifluorfen in the elution could no longer be de-
tected by spectrophotometer. Then the particles were washed with
methanol to remove residual acetic acid and dried under vacuum
at 60 °C. The corresponding non-imprinted polymer (NP1) was pre-
pared similarly. Two reference imprinted polymers (P2, P3) using
only BMA-b-CD or 4-VP as a functional monomer and their corre-
sponding non-imprinted polymers (NP2, NP3) were also prepared
(Table 1).
ble deionised and filtered through a 0.45 lm filter. All the other
solvents were of analytical grade and used as received.
Methacryloyl chloride was synthesized by the reaction of meth-
acrylic acid and dichlorosulfoxide, 74% yield: bp 97–98.5 °C; 1H
NMR (CDCl3) d (ppm): 2.02 (s, 3H, CH3); 5.03 (s, 1H, ACHa@CA),
6.50 (s, 1H, ACHb@CA).
1H NMR spectrum was recorded on a Varian INOVA 500NB (Var-
ian, USA). FT-IR spectrum was recorded on an EQUINOX55 FT-IR
spectrophotometer (Bruker, Germany). The thermal gravimetric
analysis (TGA) was performed using a TGS-2 (Perkin–Elmer, USA)
at heating rate 10 °C/min under air atmosphere. UV–vis was per-
formed on a UV2100 spectrophotometer (Unico, China). The ele-
mental analysis was measured on a Vario EL CHNS Elemental
Analyzer (Elementar, Germany).
A Shimadzu CTO-10AVP HPLC system (Shimadzu, Kyoto, Japan)
equipped with a LC-10AD pump, a SPD-10AVP UV–vis detector and
a Shim-pack VP-ODS C18 analytic column (150 ꢀ 6.0 mm) was
used for the chromatographic experiments. The mobile phase
was prepared with methanol and water (50:50, v/v). The UV detec-
tion wavelength was 254 nm and the flow rate of mobile phase
was 0.8 ml/min. The column temperature was set at 33 °C and
2.5. Equilibrium binding experiments
Equilibrium binding experiments were conducted to evaluate
the binding properties of the polymers. The polymer particles
(20.0 mg) were put in a 10 ml conical flask and mixed with
3.0 ml of a known concentration of selected substrate solution.
The conical flask was oscillated for 6 h at 30 °C. Then, the mixture
was filtrated through a 0.45
trate was determined by spectrophotometer. The amount of sub-
strate bound to the polymer Q ( mol) was calculated according to
lm filter. The concentration of the fil-
l
Q ð molÞ ¼ VðCi ꢁ ClÞ
l
where V, Ci and Cl represent the volume of the solution (ml), initial
solution concentration and the concentration after adsorption (con-
centration at equilibrium) (lmol/ml), respectively. The average data
of triplicate independent results were used for the following
sample injection volume was 5 ll.
discussion.
2.2. Synthesis of bismethacryloyl-b-cyclodextrin (BMA-b-CD)
2.6. Competitive binding experiments
The synthesis procedure of BMA-b-CD was as below: b-CD
(18.50 g, 16.30 mmol) was dissolved in 120 ml of pyridine, the
solution was cooled to 0 °C. Then, an ice-chilled solution of meth-
acryloyl chloride (3.91 g, 37.50 mmol) in ether (20 ml) was added
dropwise to this solution with stirring magnetically under nitro-
gen. The reactive mixture was slowly warmed to room tempera-
ture and stirred for 24 h. After that, the solvent was evaporated
in rotary evaporator. The residue was washed with ethanol for sev-
eral times and gave a white product in 71% yield. The product was
Acifluorfen was mixed with its analog compounds, fomesafen
and ibuprofen to form a mixture solution. The initial concentration
of each compound was the same (2.0 mmol/l). The polymer parti-
cles (P1 or NP1) (30.0 mg) were placed in a conical flask and mixed
with 3.0 ml of the mixture solution. After being shaken for 6.0 h at
30 °C, the mixture was filtrated through a 0.45 lm filter. The fil-
trate (1.0 ml) was diluted to 5.0 ml with solvent. Then, the analytes
in the solution were analyzed by HPLC.

644
Z. Xu et al. / Carbohydrate Polymers 79 (2010) 642–647
Table 1
regularly placed. The positions and mutual conformations of
BMA-b-CD molecule and 4-VP molecule are fixed by cross-linking.
This imprinting process is schematic illustrated in Fig. 2.
To investigate the contribution of the used functional mono-
mers, the other two reference imprinted polymers (P2, P3) and their
corresponding non-imprinted polymers were also prepared. To en-
sure that the concentrations of the b-CD residues in P2 and NP2 and
the 4-VP residues in P3 and NP3 are equal to that in the P1 and NP1,
different amounts of cross-linkers were used (Table 1).
Composition of the polymerization mixtures for synthesizing polymers.
Polymer BMA-b-CD
4-VP
(mmol)
Acifluorfen
(mmol)
EDMA
(mmol)
DMSO
(ml)
(mmol)
P1
NP1
P2
NP2
P3
NP3
0.50
0.50
0.50
0.50
–
0.50
0.50
–
–
0.50
0.50
0.50
–
0.50
–
0.50
–
5.0
5.0
5.25
5.25
8.26
8.26
8
8
7
7
6
6
–
3.2. Properties of the polymers P1 and NP1
3. Results and discussion
3.1. Method for preparation of MIPs
The results of the elemental analysis of P1 and NP1 were as fol-
lows: P1 found: C 55.18, H 6.35, N 0.396%; NP1 found: C 55.34, H
6.42, N 0.385%. Anal. Calcd. for P1 and NP1 (calculated according
to the chemical composition for making these polymers): C
55.60, H 6.64, N 0.41%. These indicated that most of the compo-
nents for making the polymers had participated in the polymeric
reactions. The stability of the polymers was investigated by the
TGA analysis. The results indicated that both P1 and NP1 begin to
In non-covalent approaches to molecular imprinting, non-cova-
lent interactions such as hydrogen bonding, bonding, electro-
p–p
static interaction, hydrophobic effect and metal ion-coordination,
can be exploited to organize the functional monomers around
the template. While hydrogen bonding interactions between tem-
plate and functional monomers are destroyed in aqueous media,
hydrophobic effect can be exploited. However, hydrophobic effect
is less specific because it is applicable to broad groups of com-
pounds. A solution to this problem is to combine hydrophobic
effect with other types of interactions (for instance, electrostatic
interaction, metal ion-coordination, etc.) in the imprinting process
(Piletsky et al., 1999).
decompose at around 270 °C. FT-IR (KBr)
m
(cmꢁ1) of P1: 3600–
3000, 2927, 1730, 1637, 1457, 1391, 1260, 1156, 925, 735, 621.
The location and shape of the major bands in FT-IR spectrum of
NP1 are very similar to those of P1.
3.3. Binding characteristics of the polymers for acifluorfen
The amount of acifluorfen bound to the polymers was deter-
mined by equilibrium binding experiments. The affinity of the
polymers was estimated by the distribution coefficients of acifluor-
fen between polymer and solution. The distribution coefficient Kd
(ml/g) is defined as (Zhu, Haupt, & Knopp, 2002):
b-CD molecule is a torus-shaped cyclicoligosaccharide consist-
ing of 1,4-linked D-glucopyranose units with an internal hydropho-
bic cavity. This structure enables it to form inclusion compounds
with many compounds in aqueous media or organic solvents with
high polarity through hydrophobic interactions. In the present
study, the lipophilic aromatic rings of acifluorfen can be inserted
into the cavity of BMA-b-CD to form inclusion complex in DMSO.
1H NMR experiments were conducted to study the interactions be-
tween b-CD and acifluorfen in DMSO. The results of the 1H NMR
experiments indicated that the proton chemical shifts of b-CD
and acifluorfen almost did not change whether b-CD and BPP are
mixed or not. But, the NMR signals shapes of the protons on the
C-3, C-5, C-6 (marked as H3, H5 and H6, respectively) in the gluco-
pyranose ring of b-CD have changed, as shown in Fig. 1. This indi-
cates that acifluorfen can be inserted into the cavity of b-CD and
can change the circumstance of the inner core of b-CD (Tong,
2001).
In the pre-polymerization mixture for synthesis P1, when BMA-
b-CD was mixed with acifluorfen in DMSO, the aromatic rings of
acifluorfen were entrapped inside the hydrophobic core of BMA-
b-CD and the more polar carboxyl group (–COOH) of acifluorfen
was left outside the cavity. When 4-VP was added, the carboxyl
group can form strong ionic interactions with this basic functional
monomer. Thus, the BMA-b-CD molecule and 4-VP molecule are
Kdðml=gÞ ¼ Cp=Cl
Cp (
lmol/g) is the binding capacity of the polymer, it was calculated
according to
Qð
l
molÞ
Cpð mol=gÞ ¼
l
ðmass of polymer in gÞ
The specific binding capacity
to
DCp (lmol/g) was calculated according
D
CP ¼ CpðMIPÞ ꢁ CpðNPÞ
The molecular imprinting factor IF was used to evaluate the
imprinting effect. IF was calculated according to
IF ¼ KdðMIPÞ=KdðNPÞ
The obtained Cp,
:
D
Cp, Kd and IF values are listed in Table 2.
The polymer P1, containing both functional monomers, BMA-b-
CD and 4-VP, and imprinted with acifluorfen, demonstrated supe-
rior selectivity to the reference polymer (NP1). Compared with P2
Fig. 1. Part of enlarged 1H NMR spectra of b-CD: before (a) and after (b) mixed with acifluorfen in DMSO-d6. H3, H5 and H6 represent the protons on the C-3, C-5 and C-6 in the
glucopyranose ring of b-CD, respectively.

Z. Xu et al. / Carbohydrate Polymers 79 (2010) 642–647
645
O
C
O
EDMA, AIBN
Polymerization
Cl
N
+
F3C
O
NO2
COOH
C
O
O
O
C
O
O
C
O
O
O
N
Dissociation
Association
HN
Cl
C
Acifluorfen
+
NO2
F3C
O
C
C
O
O
O
O
Fig. 2. Schematic illustration of the molecular imprinting procedure.
Table 2
50
45
40
35
30
25
20
15
10
P1
NP1
Recognition of acifluorfen on polymers prepared with different functional
monomers.a
P1
NP1
P2
NP2
P3
NP3
Cp
47.63
19.18
28.32
1.80
28.45
23.54
1.87
12.77
1.09
21.67
31.24
11.08
17.43
1.61
20.16
D
Kd
IF
Cp
15.72
11.68
10.81
a
Initial concentration: 2.0 mmol/l; solvent:methanol/water (1:1, v/v); volume:
p
C
3 ml.
and P3, P1 showed the highest Cp,
DCp, Kd and IF values. In the rec-
ognition process, while the lipophilic aromatic rings of acifluorfen
insert in the cavity of the modified b-CD through hydrophobic ef-
fects, the carboxyl group of acifluorfen can form strong ionic inter-
actions with the basic functional group of 4-VP residues. The
results of binding experiments demonstrated that the combined
functional monomers imprinting strategy can increase the specific
binding sites in the cavities of the polymer and thus enhances the
molecular recognition ability. The polymer P2, which was im-
printed with acifluorfen only using the BMA-b-CD as the functional
monomer, showed almost no imprinting effect. Table 2 shows that
P3 also has binding affinity for the template. P3 was prepared in the
absence of the BMA-b-CD monomer, there have electrostatic inter-
actions between the templates and the basic monomers, and the
electrostatic interactions also induced the imprinting effect. It is
worthy of note that the binding capacity (Cp) of P3 is much lower
than P1, although the concentrations of 4-VP residues in P3 is equal
to that in the P1.
0.5
1.0
1.5
2.0
2.5
Initial concentration of acifluorfen (mmol/l)
Fig. 3. Binding isotherm of P1 and NP1. solvent:methanol/water (1:1, v/v); volume:
3 ml.
Cp Cpmax ꢁ Cp
¼
Cl
K
where Cpmax
sites and K (
(l
mol/g) is the apparent maximum number of binding
mol/ml) is the dissociation constant. As shown in
l
Fig. 4, the Scatchard plot is not a single straight line, but consists
of two linear parts with different slope. The linear regression equa-
tions for the two linear regions are Cp/Cl = ꢁ1.152Cp + 83.87
(r = 0.994) and Cp/Cl = ꢁ2.298Cp + 136.04 (r = 0.980), respectively.
This suggests that the binding sites in P1 are heterogeneous in re-
spect to the affinity for acifluorfen, and indicates that the binding
sites could be classified into two groups with specific binding prop-
erties. The K and Cpmax of the higher affinity binding sites can be cal-
culated to be 0.435 lmol/ml and 59.20 lmol/g, respectively, from
the slope and the intercept of the Scatchard plot. Similarly, the K
and Cpmax of the lower affinity binding sites were found to be
0.868 lmol/ml and 72.80 lmol/g, respectively.
3.4. Adsorption isotherm
In an attempt to investigate the binding performance of the
polymer, the binding isotherm of acifluorfen to polymers P1 and
NP1 was determined in the 0–2.5 mmol/l range of initial concentra-
tion of acifluorfen. The results are shown in Fig. 3. It can be seen
that the amount of acifluorfen bound to both polymers at equilib-
rium Cp increased along with increasing the initial concentration of
acifluorfen, but the binding amount of acifluorfen on the imprinted
polymer P1 was more than that on the non-imprinted polymer NP1
in the whole concentration range, reflecting the molecular imprint-
ing effect.
3.5. Influence of H2O content in methanol on adsorption
Different contents of H2O in methanol (v/v) were used as sol-
vents to evaluate the influence of H2O on the binding of acifluorfen
on the polymers P1 and NP1 (Fig. 5). It can be seen that the distri-
bution coefficients of acifluorfen on P1 and NP1 increased with
increasing the H2O content in the range of 10–50% (v/v). As we
The obtained binding data of acifluorfen to P1 were further pro-
cessed with Scatchard equation to estimate the binding properties
of MIP (Matsui, Miyoshi, Doblhoff-Dier, & Takeuchi, 1995).

646
Z. Xu et al. / Carbohydrate Polymers 79 (2010) 642–647
65
60
55
50
45
40
35
30
25
20
both in shape and functional group arrangement to the template
molecule, (2) insertion of acifluorfen into the cavities of b-CD res-
idues which were random arrangement, and (3) unspecific interac-
tions (e.g. electrostatic interaction, hydrophobic effect) between
acifluorfen and polymers. The binding of acifluorfen to NP1 was
caused by points (2) and (3) mentioned above. The difference in
binding mechanism between P1 and NP1 gives rise to the observed
difference in the values of Kd between P1 and NP1.
l
C
p
3.6. Specific binding effect of the polymer P1 in mixture solution
C
In this work, the recognition ability of the imprinted polymer P1
toward acifluorfen was directly observed from the competitive
adsorption experiment. To demonstrate the binding specificity of
P1, two analog molecules of acifluorfen, fomesafen and ibuprofen,
were chosen. They were mixed with acifluorfen to form a mixture
solution for adsorption. The adsorption amount of each component
(Cp) by the polymer was determined by measuring the residual
concentration of each component in the bulk solution. The samples
were analyzed with HPLC and external standard method was used
for quantification.
20
25
30
35
40
45
50
CP (µmol/g)
Fig. 4. Scatchard plot analysis of the binding of acifluorfen to P1.
Fig. 6a and b are the chromatographs of the mixture solutions
before and after being binding by P1 (both solutions were diluted
five times with THF before HPLC test). The binding capacity of P1
for acifluorfen, ibuprofen and fomesafen can be calculated to be
30
28
26
24
22
20
18
16
14
12
10
P
NP
1
1
38.45, 11.22 and 9.48
of NP1 for acifluorfen, ibuprofen and fomesafen found to be
16.40, 8.55 and 9.12 mol/g, respectively. The results of competi-
lmol/g, respectively. The binding capacity
l
tive adsorption experiment further confirmed that the imprinted
polymer has molecular recognition ability.
After binding the compounds, P1 was Soxhlet extracted with
6.0 ml of a methanol–acetic acid mixture (9:1, v/v). The eluate
was concentrated to dryness by a nitrogen stream. The dry residue
was dissolved in methanol and the volume of the solution was set
at 3 ml. Then, the solution was analyzed by HPLC. The resulting
chromatogram is presented in Fig. 6c. We can see that the main
component of the eluate is acifluorfen. The concentration of aciflu-
orfen is 0.317 mmol/l. The recovery efficiency of the polymer P1
10
20
30
40
50
can be calculated to be 31.70
lmol/g. Compared with its Cp value
Water content in methanol,%(v/v)
(38.45 mol/g), a recovery of 82.4% was obtained. These results
l
indicated that the imprinted polymer P1 can be used as a solid-
phase extraction sorbent to separate the template from its analogs.
Fig. 5. The influence of H2O content on adsorption. Initial concentration: 2.0 mmol/l;
volume: 3 ml.
4. Conclusions
know, the cavity of b-CD is relatively hydrophobic compared to
water. When the content of H2O increased in the solvent, the
hydrophobic effect increased also, more template molecules have
been driven into the cavities of the polymers. The binding of aciflu-
orfen to P1 was caused by (1) insertion of acifluorfen into cavities
created during the imprinting process, which are complementary
In this work, an acifluorfen-imprinted polymer (P1) was pre-
pared by using 4-VP and BMA-b-CD as bi-functional monomers.
Using BMA-b-CD or 4-VP as the functional monomers, two refer-
ence imprinted polymers (P2, P3) were also synthesized. The im-
printed polymer P1 demonstrated superior selectivity to the
Fig. 6. Chromatographs on high performance liquid chromatography. (a) Mixture solution before adsorption, (b) mixture solution after adsorption by P1, (c) eluate of P1. 1 –
Ibuprofen; 2 – acifluorfen; 3 – fomesafen.

Z. Xu et al. / Carbohydrate Polymers 79 (2010) 642–647
647
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polymers synthesized with a single functional monomer (P2, P3).
The study revealed that the combination of hydrophobic and elec-
trostatic interactions in molecular imprinting can effectively im-
prove the recognition ability of the imprinted polymer. The MIP
synthesis strategy is promising for realizing effective imprinting
in aqueous media. The competitive binding experiments indicated
that the imprinted polymer P1 can be used as a solid-phase extrac-
tion sorbent to separate the template from its structural analogs.
Acknowledgement
Meng, Z. H., Yamazaki, T., & Sode, K. (2004). A molecularly imprinted catalyst
designed by
a computational approach in catalysing a transesterification
This study was supported by the Open Fund Project of Key Lab-
oratory in Hunan Universities and Scientific Research Program of
Hengyang Science and Technology Bureau (No. 2008KS035).
process. Biosensors and Bioelectronics, 20, 1068–1075.
Piletsky, S. A., Andersson, H. S., & Nicholls, I. A. (1999). Combined hydrophobic and
electrostatic interaction-based recognition in molecularly imprinted polymers.
Macromolecules, 32, 633–636.
Puoci, F., Garreffa, C., Iemma, F., & Muzzalupo, R. (2005). Molecularly imprinted
solid phase extraction for detection of Sudan I in food matrices. Food Chemistry,
93, 349–353.
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