Benzimidazole-based chromogenic chemosensor for the recognition of oxalic acid via counter ion displacement assay in semi-aqueous medium

Article abstract of DOI:10.1016/j.tet.2013.08.043

An azo dye-coupled benzimidazole-based receptor 1 was synthesized and investigated as a receptor for metal ions in semi-aqueous medium. The receptor recognizes Cu²⁺ with high selectivity over other metal ions. The resultant complex 1·Cu²⁺ was found to selectively bind oxalic acid via counter ion displacement.

Full text of DOI:10.1016/j.tet.2013.08.043

Tetrahedron 69 (2013) 9001e9006
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Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Benzimidazole-based chromogenic chemosensor for the recognition
of oxalic acid via counter ion displacement assay in semi-aqueous
medium
,
d,
Preeti Saluja a, Navneet Kaur b, Jongmin Kang c, Narinder Singh a , Doo Ok Jang
*
*
a Department of Chemistry, Indian Institute of Technology Ropar, Rupnagar 140001, Punjab, India
b Centre for Nanoscience & Nanotechnology, Panjab University, Chandigarh 160014, Punjab, India
c Department of Chemistry, Sejong University, Seoul 143-747, Republic of Korea
d Department of Chemistry, Yonsei University, Wonju 220-710, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 11 July 2013
Received in revised form 13 August 2013
Accepted 16 August 2013
Available online 23 August 2013
An azo dye-coupled benzimidazole-based receptor 1 was synthesized and investigated as a receptor
for metal ions in semi-aqueous medium. The receptor recognizes Cu2þ with high selectivity over other
metal ions. The resultant complex 1$Cu2þ was found to selectively bind oxalic acid via counter ion
displacement.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Oxalic acid
Counter ion displacement assay
UVevis
Chromogenic sensor
Cu (II) complex
1. Introduction
acid in food and biological fluids since it is a simple technique and
changes can be detected with the naked eye.
Oxalic acid is essential for the immune system to fight viral,
bacterial, and vascular diseases. With an oxalic acid deficiency, the
immune system cannot eliminate abnormal cells. Radical cells can
also develop, resulting in tumor formation.1 Oxalic acid is an in-
teresting paradox; a small amount is required for body function
while an excess causes calcium oxalate kidney stones.2 Oxalic acid
can also lead to nutrient deficiencies because it binds to vital nu-
trients. Thus, the level of oxalic acid in food is an important crite-
rion for determining food quality.
Oxalic acid determination is performed through a number of
techniques, including electrochemical techniques (voltammetry and
amperometry),3,4 kinetic spectrophotometric methods,5 bio-ther-
mochips,6 electrochemiluminescence,7 electrocatalytic methods,8
liquid chromatography-tandem mass spectrometry,9 flow injection
spectrophotometric methods,10 ion-exclusion chromatography,11
HPLC,12 and fluorescence spectroscopy.13 However, recognition of
oxalic acid with UVevis spectroscopy is rare.14 Chromogenic recog-
nition of oxalic acid would be an interesting tool for analyzing oxalic
The recognition of anionic species in environmentaland biological
samples must be performed in aqueous or semi-aqueous media.
However, this is difficult to carry out because, if the anion receptor is
only composed of hydrogen bond donor sites, the analytes compete
with polar solvents for anion receptor binding sites.15 In recent years,
cation or counter ion displacement has been widely investigated
since it relies on an electrostatic interaction between a metal ion and
an anionic species,16 which is less affected by polar solvents.
To continue our research on the development of (2-aminophenyl)
benzimidazole-based chemosensors,17 we herein report a chromo-
genic sensor that recognizes oxalic acid through counter ion
displacement assay in semi-aqueous medium. Receptor 1 was syn-
thesized by a condensation reaction of 2-(2-aminophenyl)benz-
imidazole and azo compound 218 in dry methanol (Scheme 1).
The absorption spectra of receptor 1 in HEPES-buffered CH3OH/
H2O (1:1, v/v, 20 mM, pH¼7.0) had absorption maxima at 365 nm
and 490 nm (Fig. 1A). The band at 365 nm is characteristic for imine
linkage due to an internal charge transfer (ICT),19a while the band at
490 nm arises due to incorporation of an azo group.19b,c UVevis
absorption spectra of receptor 1 shows formation of new band at
365 nm, which is different from the spectra of both of its reactants,
confirming development of a new compound (Fig. S1). Furthermore,
* Corresponding authors. E-mail addresses: nsingh@iitrpr.ac.in (N. Singh),
dojang@yonsei.ac.kr (D.O. Jang).
0040-4020/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2013.08.043

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P. Saluja et al. / Tetrahedron 69 (2013) 9001e9006
binds with receptor 1 causing a bathochromic shift in the absorp-
tion maxima from 365 nm to 415 nm. The recognition profile
reveals that the cavity of receptor 1 has compatibility to bind with
Cu2þ. Receptor 1 was titrated with Cu2þ to investigate its photo-
R
MeOH
reflux
O
N
NH2
HN
HN
+
N
N
HO
physical properties. Upon addition of Cu2þ (0.5
mM) to a solution of
HO
R
receptor 1 (10
mM) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM,
2
pH¼7.0), the absorbance peak at 365 nm decreased while the
absorbance peak at 415 nm increased with a clear isosbestic point
at 390 nm. The observed changes continued gradually with gradual
increases in Cu2þ until reaching a saturation point that was attained
1
R =
N
N
NO2
Scheme 1.
1.66
1.88
2.63
3.44
4.23
4.75
5.43
6.27
6.75
7
1
0.8
2.28
2.98
3.88
4.52
5.12
5.89
6.54
6.84
7.59
7.89
9.1
A
B
Absorbance at 365 nm
Absorbance at 490 nm
0.8
0.6
0.4
0.2
0
0.6
7.7
8.57
9.28
10.12
11.18
12.11
0.4
0.2
0
9.36
10.58
11.44
300
400
500
600
700
0
5
10
15
Wavelength (nm)
pH
Fig. 1. (A) Changes in the UVevis spectra of receptor 1 (10 mM) in CH3OH/H2O (1:1, v/v) with variable pH. (B) Changes in absorbance at 365 nm and 490 nm with varied pH.
literature supports that arising of band at 365 nm is due to imine
linkage. However, in the case of receptor 1, the band at 365 nm is due
to superimposition of both imine linkage and azophenol group. The
effect of pH on the UVevis absorption spectra of receptor 1 was
by adding 1.0 equiv (10
in the UVevis spectra of receptor 1 with the addition of 1 equiv of
Cu2þ indicates the formation of a 1:1 complex between 1 and Cu2þ
m
M) of Cu2þ, as shown in Fig. 2B. Saturation
.
This is further confirmed by Job’s plot (Fig. S4).21 Mass spectra also
support a 1:1 complex because the m/z peak at 524.1 corresponds
to the Mþ peak, where M is [1ꢀHþþCu2þ] (Fig. S5). As receptor 1
and Cu2þ form a 1:1 complex, the BenesieHildebrand plot was used
to calculate the binding constant, giving 1.10 (ꢁ0.1)ꢂ106 Mꢀ1
(Fig. S6).22 Using titration data, the detection limit was calculated as
investigated by recording receptor 1 spectra (10 mM) in CH3OH/H2O
(1:1, v/v) with varying pH. In acidic pH, the spectra showed marginal
changes. However, as pH increased above 7, the absorbance band at
365 nm began decreasing and the absorbance band at 490 nm began
increasing, as shown in Fig. 1B. The results are due to deprotonation
by a base, which leads to a bathochromic shift with increasing pH.20
To avoid the effect of variable pH, further recognition studies
were carried out in HEPES buffer solution. Cation recognition
0.56 m
M (Fig. S7).23
A comparison has been made between the diffraction pattern of
receptor 1 and its metal complex 1$Cu2þ by executing the PXRD
technique. Fig. S8 shows the XRD pattern of receptor 1, its complex
studies were performed by recording receptor 1 (10
m
M) spectra in
the presence of metal ions (10
m
M), including Naþ, Kþ, Ba2þ, Sr2þ
,
1$Cu2þ, and copper nitrate measured over a range of 2
q values lying
Ca2þ, Mg2þ, Mn2þ, Cr3þ, Fe3þ, Co2þ, Ni2þ, Cu2þ, Zn2þ, Pb2þ, Agþ,
Hg2þ, and Cd2þ in their nitrate forms in a HEPES-buffered CH3OH/
H2O (1:1, v/v, 20 mM, pH¼7.0) solution. Fig. 2A illustrates that Cu2þ
between 0 and 100ꢃ. The scattering angles found from the dif-
fraction pattern for 1$Cu2þ (6.68, 9.45, 9.69, 10.06, 10.39, 12.06,
13.22, 17.06, 18.09, 18.99, 19.66, 19.99, 21.66, 23.05, 23.59, 24.83,
0.4
0.5
0.4
0.3
0.2
0.1
0
B
0.3
0.2
Absorbance at 365nm
Absorbance at 415nm
0.1
0
5
Cu2+
]
10
[
,
Cd2+
300
350
400
450
500
550
600
Wavelength (nm)
Fig. 2. (A) Changes in UVevis absorption spectra of receptor 1 (10
changes in UVevis absorption spectra of receptor 1 (10
M) with increasing concentrations of Cu2þ (from 0 to 10
shows the absorbance of receptor 1 at 365 nm and 415 nm with respect to Cu2þ concentration (0e10
M).
m
M) in the presence of metal nitrate salts (10
m
M) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0); (B)
m
m
M) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0). Inset
m

P. Saluja et al. / Tetrahedron 69 (2013) 9001e9006
9003
25.53, 25.99, 26.74, 27.91, 30.67, 33.96, 36.79, 38.60, 41.63, 48.80)
are different from the scattering angles of receptor 1 (5.25, 7.40,
10.53, 12.44, 13.04, 14.92, 15.51, 16.78, 18.39, 19.53, 21.35, 22.56,
23.74, 26.57, 27.20, 28.68, 31.14, 32.11) and copper nitrate (14.86,
15.02, 18.65, 22.00, 26.32, 28.47, 30.09, 37.11, 38.73, 42.43, 44.14,
48.51, 54.07, 77.17, 85.98, 87.33, 88.13, 89.42). This indicates the
formation of a new compound with different composition.
with tetrabutylammonium salt of sulfide (Fig. S11). S2ꢀ is not ef-
fective for the sensor application of 1$Cu2þ because the modulation
in its UVevis spectrum with S2ꢀ is at different wavelength.
Among all tested dicarboxylic acids, complex 1$Cu2þ selectively
bound oxalic acid. Selectivity was verified by carrying out com-
petitive experiments in the presence of all tested dicarboxylic acids.
The absorption spectra of complex 1$Cu2þ (10
mM) with oxalic acid
The most important feature of a receptor is its selectivity in the
presence of other cations. Competitive experiments were per-
formed in the presence of background cations, such as Naþ, Kþ,
were unchanged in the presence of all tested dicarboxylic acids
(Fig. 5). Rationale for selective binding of oxalic acid by 1$Cu2þ
complex is smallest size of oxalic acid among all dicarboxylic acid. It
can easily fit into the coordination sphere of Cu2þ binding to re-
ceptor 1 replacing small counter anion, i.e., nitrate, supporting the
mechanism as counter ion displacement mechanism. Furthermore,
oxalic acid has strong chelating effect as it forms a stable five
member ring on binding to a metal ion.24
Ba2þ, Sr2þ, Ca2þ, Mg2þ, Mn2þ, Cr3þ, Fe3þ, Co2þ, Ni2þ, Zn2þ, Pb2þ
,
Agþ, Hg2þ, and Cd2þ. Fig. 3 shows that the absorption spectra of
receptor 1 (10 M) in HEPES-buffered CH3OH/
m
M) with Cu2þ (10
m
H2O (1:1, v/v, 20 mM, pH¼7.0) remained almost unaltered in the
presence of the same equivalent of another cation.
0.8
Cu(II) only
Na(I)+Cu(II)
K(I)+Cu(II)
0.8
A
B
Mn(II)+Cu(II)
Ba(II)+Cu(II)
Sr(II)+Cu(II)
Ca(II)+Cu(II)
Mg(II)+Cu(II)
Cr(III)+Cu(II)
Fe(III)+Cu(II)
Co(II)+Cu(II)
Ni(II)+Cu(II)
Cu(II)+Zn(II)
Pb(II)+Cu(II)
Ag(I)+Cu(II)
Hg(II)+Cu(II)
Cd(II)+Cu(II)
0.6
0.4
0.2
0
0.6
0.4
0.2
0
300
400
500
600
700
Wavelength (nm)
Fig. 3. (A) UVevis absorption spectra of receptor 1 (10
background cations; (B) bar diagram showing absorbance of receptor 1 (10
20 mM, pH¼7.0) at 415 nm.
m
M) with Cu2þ (10
m
M) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0) in the presence of all other tested
m
M) bound to Cu2þ (10
m
M) in the presence of other tested cations (10 M) in CH3OH/H2O (1:1, v/v,
m
To evaluate the anion/dicarboxylic acid recognition ability of
complex 1$Cu2þ, UVevis absorption spectra of 1$Cu2þ in HEPES-
buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0) were recorded in
the presence of different anions (Fꢀ, Clꢀ, Brꢀ, Iꢀ, NO3ꢀ, CNꢀ, ClO4ꢀ,
AcOꢀ, HSOꢀ4 , and H2POꢀ4 ) and various dicarboxylic acids (succinic
acid, malonic acid, terephthalic acid, isophthalic acid, oxalic acid,
glutaric acid, adipic acid, suberic acid, and pimelic acid). As shown
in Fig. 4, there was no change in UVevis absorption spectra upon
addition of any anion or dicarboxylic acid except for oxalic acid.
Oxalic acid selectively shifts the UVevis absorption spectra of
1$Cu2þ with a hypsochromic shift from 415 nm to 375 nm. The
interaction of oxalic acid with 1$Cu2þ was verified by titrating
The CV profile of receptor 1 indicates that receptor 1 is inactive
in the selected potential window (Fig. 6). However, upon addition
of Cu2þ to receptor 1, there was one oxidation peak at ꢀ0.7 V. With
further addition of oxalic acid to the 1$Cu2þ complex, this peak
shifted to ꢀ0.5 V. A comparison of the CV profiles of receptor 1,
1$Cu2þ, and 1$Cu2þþoxalic acid in HEPES-buffered CH3OH/H2O
(1:1, v/v, 20 mM, pH¼7.0) implies that binding with Cu2þ changes
the electrochemical structure of the sensor. In addition, the CV
profile shows the interaction of 1$Cu2þ complex with oxalic acid.
The CV profile supports a counter ion displacement mechanism
that senses oxalic acid because the CV profile of 1$Cu2þþoxalic acid
was different from the profiles of receptor 1 and 1$Cu2þ
.
1$Cu2þ (10
(0
mM) with increasing concentrations of oxalic acid
In summary, the azo dye-coupled benzimidazole-based receptor
1 was synthesized. Receptor 1 showed high selectivity for Cu2þ over
other metal ions. The resultant 1$Cu2þ complex was used as
a highly selective chromogenic receptor for oxalic acid in semi-
aqueous medium, shifting the back absorption peak from 415 nm
m
Me1 mM). On stepwise addition of oxalic acid, the absorption
spectra of 1$Cu2þ gradually shifted from 415 nm to 375 nm with
decreased absorbance at 415 nm and increased absorbance at
375 nm. The binding constant was calculated to be 6.25 (ꢁ0.3)ꢂ
107 Mꢀ1 using a BenesieHildebrand plot (Fig. S9).22 The detection
limit was also calculated from titration data. Complex 1$Cu2þ can
to 375 nm with a detection limit of 70.8 mM. The counter ion dis-
placement mechanism was confirmed by recording CV profiles of 1,
detect oxalic acid to a limit of 70.8
m
M (Fig. S10).23 Changes in
1$Cu2þ, and 1$Cu2þ with oxalic acid.
UVevis absorption spectra of 1$Cu2þ in HEPES-buffered CH3OH/
H2O (1:1, v/v, 20 mM, pH¼7.0) with dianions of dicarboxylic acids
(succinic acid, malonic acid, terephthalic acid, isophthalic acid,
oxalic acid, glutaric acid, adipic acid, suberic acid, and pimelic acid)
were also similar to changes with dicarboxylic acids (Fig. S11). As
Cu2þ has strong affinity with S2ꢀ, the effect of sulfide on complex
1$Cu2þ was also examined by recording UVevis absorption spectra
of 1$Cu2þ in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0)
2. Experimental
2.1. General
All chemicals were purchased from commercial suppliers and
used without further purification. 1H NMR and 13C NMR spectra
were recorded on an Avance-II (Bruker) instrument operating at

9004
P. Saluja et al. / Tetrahedron 69 (2013) 9001e9006
0.8
0.6
0.4
0.2
0
0.8
0.6
0.4
0.2
0
1
1
A
B
1.Cu(II)
1.Cu(II)
Succinic acid
Malonic acid
Terephthalic acid
Isophthalic acid
Oxalic acid
Glutaric acid
Adipic acid
Suberic acid
Pimelic acid
Fluoride
Chloride
Bromide
Iodide
Cyanide
Nitrate
Dihydrogen phosphate
Hydrogen sulphate
Perchlorate
Acetate
300
400
500
600
700
300
400
500
600
700
Wavelength (nm)
Wavelength (nm)
0.45
0.4
0.35
0.3
0.25
0.2
0.5
0.4
0.3
0.2
0.1
0
C
Absorbance at 375nm
Absorbance at 415nm
0
200 400 600 8001000
[Oxalic acid]
300
350
400
450
Wavelength (nm)
500
550
600
Fig. 4. Changes in UVevis absorption spectra of complex 1$Cu2þ (10
mM) in the presence of (A) tetrabutylammonium salts of different anions (1 mM) and (B) dicarboxylic acids
(1 mM) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0); (C) changes in UVevis absorption spectra of complex 1$Cu2þ (10
mM) with increasing concentrations of oxalic acid
(0e1 mM) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0). Inset shows absorbance of 1$Cu2 at 375 nm and 415 nm with respect to concentration of oxalic acid (0e1 mM).
0.5
0.5
0.4
0.3
0.2
0.1
0
A
B
0.4
0.3
0.2
0.1
0
a
b
c
d
e
f
g
h
i
300
350
400
450
500
a
b
c
d
e
f
g
h
i
Wavelength (nm)
Fig. 5. (A) UVevis absorption spectra of complex 1$Cu2þ (10
tested background dicarboxylic acids; (B) bar diagram showing absorbance of complex 1$Cu2þ (10
m
M) with oxalic acid (1 mM) in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0) in the presence of all other
m
M) binding with oxalic acid (1 mM) in the presence of other tested dicarboxylic
acids (1 mM) in CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0) at 375 nm with (a) oxalic acid only, (b) oxalic acidþsuccinic acid, (c) oxalic acidþmalonic acid, (d) oxalic acidþterephthalic
acid, (e) oxalic acidþisophthalic acid, (f) oxalic acidþglutaric acid, (g) oxalic acidþadipic acid, (h) oxalic acidþsuberic acid, and (i) oxalic acidþpimelic acid.
400 MHz for 1H NMR and 100 MHz for 13C NMR. IR spectra were
measured on a Bruker Tensor 27 spectrophotometer. An Analy-
tikjena Specord 250 Plus spectrometer was used to collect UVevis
absorption spectra. Redox potential was measured on a Potentios-
tateGalvanostat BASI EPSILON. Electrochemical measurements
were made using a three-electrode Pt disk working electrode,
platinum wire counter-electrode, and Ag/AgNO3 reference elec-
trode in a single-compartment cell under nitrogen at 25 ꢃC.
Tetrabutylammonium perchlorate was used as the supporting
electrolyte.

P. Saluja et al. / Tetrahedron 69 (2013) 9001e9006
9005
CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0). To determine the stoichi-
ometry of the complex formed from receptor 1 and Cu2þ, solutions
of receptor 1 and Cu2þ were prepared in ratios of 1:9, 2:8, 3:7, 4:6,
1:1, 6:4, 7:3, 8:2, and 9:1. These solutions were kept for 3 h and
shaken occasionally. UVevis absorbance spectra were then recor-
ded for all solutions. To evaluate any possible interference due to
other metal ions when estimating Cu2þ, solutions were prepared
0.0000060
0.0000045
0.0000030
0.0000015
0.0000000
-0.0000015
1
1.Cu2+
1.Cu2++Oxalic acid
containing receptor 1 (10 mM) along with a fixed concentration of
Cu2þ both with and without other background cations in HEPES-
buffered CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0). The UVevis ab-
sorbance spectra of each solution were recorded.
2.1.4. Anion recognition properties of 1$Cu2þ. Studies similar to
those conducted to determine cation recognition properties were
performed, except tetrabutylammonium salts of the anions and
dicarboxylic acids were used. Detailed concentrations are given in
the manuscript text.
Acknowledgements
-0.8 -0.6 -0.4 -0.2 0.0 0.2
Voltage
This work was supported by the Korean and Indian Government
for the India-Korea Joint Program of Cooperation in Science &
Technology (NRF-2011-0027710) and the CSIR Project (project no:
01(2417)/10/EMR-II). P.S. would like to thank UGC India for her
research fellowship.
Fig. 6. CV profiles of receptor 1, 1$Cu2þ, and 1$Cu2þþoxalic acid in HEPES-buffered
CH3OH/H2O (1:1, v/v, 20 mM, pH¼7.0).
2.1.1. Synthesis of 1. A solution of 2-(2-aminophenyl)-1H-benz-
imidazole (209 mg, 1.0 mmol) and compound 2 (406 mg, 1.5 mmol)
was heated in dry MeOH (100 mL) at reflux for 8 h. The orange solid
precipitate was filtered and washed with cold MeOH three times.
An orange solid was obtained in 89% yield (411 mg). Mp>200 ꢃC. IR
Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.tet.2013.08.043.
(KBr): NeH and OeH (3395), C]N (1592), NO2 (1342, 1522) cmꢀ1
;
1H NMR (400 MHz, DMSO-d6)
d
6.82 (t, 1H, J¼6.88 Hz), 6.88 (d, 1H,
References and notes
J¼7.8 Hz), 7.05e7.25 (m, 6H), 7.35 (s, 1H), 7.38 (d, 1H, J¼2.28 Hz),
7.41 (s, 1H), 7.67 (d, 1H, J¼7.8 Hz), 7.80e7.88 (m, 3H), 7.95e8.00 (m,
1. US Patent: 6133318: Oxalic Acid or Oxalate Compositions and Methods for
Bacterial, Viral, and Other Diseases or Conditions.
1H), 8.27e8.36 (m, 2H); 13C NMR (100 MHz, DMSO-d6)
d 110.0,
2. (a) Zheng, Y.; Yang, C.; Pu, W.; Zhang, J. Food Chem. 2009, 114, 1523e1528; (b)
Sljukic, B.; Baron, R.; Compton, R. G. Electroanalysis 2007, 19, 918e922; (c)
Manea, F.; Radovan, C.; Corb, I.; Pop, A.; Burtica, G.; Malchev, P.; Picken, S.;
Schoonman, J. Sensors 2007, 7, 615e627; (d) Ivandini, T. A.; Rao, T. N.; Fujishima,
A.; Einaga, Y. Anal. Chem. 2006, 78, 3467e3471.
3. Yamazaki, S. I.; Fujiwara, N.; Yasuda, K. Electrochim. Acta 2010, 55, 753e758.
4. Chen, X.; Cai, Z.; Huang, Z.; Oyama, M.; Jiangb, Y.; Chen, X. Nanoscale 2013, 5,
5779e5783.
5. Ensafi, A. A.; Abbasi, S.; Rezaei, B. Spectrochim. Acta Part A Mol. Biomol. Spectrosc.
2001, 57, 1833e1838.
6. Shimohigoshi, M.; Karube, I. Sens. Actuators B Chem. 1996, 30, 17e21.
7. Zhao, C. Z.; Egashira, N.; Kurauchi, Y.; Ohga, K. Electrochim. Acta 1998, 43,
2167e2173.
111.4, 114.7, 116.9, 117.9, 118.6, 122.1, 122.2, 123.0, 123.7, 124.5, 124.6,
124.8, 127.1, 131.5, 132.7, 143.1, 143.8, 144.9, 147.2, 147.8, 155.2, 159.4.
Anal. Calcd for C26H18N6O3: C, 67.53; H, 3.92; N, 18.17. Found: C,
67.15; H, 3.78; N, 18.53.
2.1.2. Synthesis of complex 1$Cu2þ. Compound
1 (462 mg,
1.0 mmol) and Cu(NO3)2$6H2O (295 mg, 1.0 mmol) were placed in
CH3OH (50 mL). The solution was heated to reflux for 4 h. Upon
reaction completion, the product was separated via slow diffusion
of diethyl ether into the reaction mixture. The solid metal complex
1$Cu2þ was washed with cold methanol, and a dark brown product
was obtained in 79% yield (513 mg). Mp>200 ꢃC. IR (KBr): NeH and
OeH (3233), C]N (1612), NO2 (1517, 1338) cmꢀ1; Anal. Calcd for
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C
26H18CuN8O9: C, 48.04; H, 2.79; N, 17.24. Found: C, 48.13; H, 2.88;
N, 17.16.
2.1.3. Metal recognition studies of 1. All recognition studies were
performed at 25ꢁ1 ꢃC. Before recording any spectra, solutions were
sufficiently shaken to ensure uniformity. The effect of pH on the
UVevis absorption spectrum of receptor 1 was investigated with
a solution of receptor 1 prepared in CH3OH/H2O (1:1, v/v). The
cation binding ability of receptor 1 in HEPES-buffered CH3OH/H2O
(1:1, v/v, 20 mM, pH¼7.0) was determined by preparing standard
solutions of receptor 1 along with fixed amounts of a particular
metal nitrate salt in HEPES-buffered CH3OH/H2O (1:1, v/v, 20 mM,
pH¼7.0). The cation recognition behavior of receptor 1 was evalu-
ated from changes in the UVevis absorption spectra of the receptor
after adding that metal salt. Titrations used volumetric flasks con-
taining each standard solution of receptor 1 along with varied
amounts of a particular metal nitrate salt in HEPES-buffered

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