Synthesis, X-ray crystal structure and optical properties of novel 1,3,5-triarylpyrazoline derivatives and the fluorescent sensor for Cu 2+

Article abstract of DOI:10.1016/j.saa.2012.12.062

A series of novel 1,3,5-triarylpyrazoline derivatives was synthesized by the reaction of chalcone and 5-aryl-2-hydrazinyl-1,3,4-thiadiazole in 43.3-84.7% yields. The structures of compounds were characterized using IR, ¹H NMR and HRMS spectroscopy and X-ray diffraction analysis. The absorption and fluorescence characteristics of the compounds were investigated in dichloromethane, toluene, acetonitrile, N,N-dimethylformamide and tetrahydrofuran. The results showed that the absorption maxima of the compounds vary from 366 to 370 nm depending on the group bound to benzene rings. The maximum emission spectra of the compounds in dichloromethane were dependent on nature of groups in benzene ring. Furthermore, the compound 3b can be used to determine Cu²⁺ ion with high selectivity and a low detection limit in the DMF:H₂O = 1:1 (v/v) solution.

Full text of DOI:10.1016/j.saa.2012.12.062

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
Contents lists available at SciVerse ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
Synthesis, X-ray crystal structure and optical properties of novel
1,3,5-triarylpyrazoline derivatives and the fluorescent sensor for Cu2+
Sheng-Qing Wang, Ying Gao, Hao-Yan Wang, Xiao-Xin Zheng, Shi-Li Shen, Yan-Ru Zhang,
⇑
Bao-Xiang Zhao
Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, PR China
g r a p h i c a l a b s t r a c t
h i g h l i g h t s
A series of 1,3,5-triarylpyrazoline derivatives were synthesized. These derivatives exhibited large Stokes
shifts. The compound 3b can be used to determine Cu2+ ion. The sensor is very sensitive with fluorometric
detection limit of 2.46 ꢀ 10ꢁ8 M.
"
"
"
"
A series of 1,3,5-triarylpyrazoline
derivatives were synthesized.
These derivatives exhibited large
Stokes shifts.
The compound 3b can be used to
determine Cu2+ ion.
The sensor is very sensitive with
fluorometric detection limit of
2.46 ꢀ 10ꢁ8 M.
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 12 October 2012
Received in revised form 29 November 2012
Accepted 13 December 2012
Available online 10 January 2013
A series of novel 1,3,5-triarylpyrazoline derivatives was synthesized by the reaction of chalcone and 5-
aryl-2-hydrazinyl-1,3,4-thiadiazole in 43.3–84.7% yields. The structures of compounds were character-
ized using IR, 1H NMR and HRMS spectroscopy and X-ray diffraction analysis. The absorption and fluores-
cence characteristics of the compounds were investigated in dichloromethane, toluene, acetonitrile, N,N-
dimethylformamide and tetrahydrofuran. The results showed that the absorption maxima of the com-
pounds vary from 366 to 370 nm depending on the group bound to benzene rings. The maximum emis-
sion spectra of the compounds in dichloromethane were dependent on nature of groups in benzene ring.
Furthermore, the compound 3b can be used to determine Cu2+ ion with high selectivity and a low detec-
tion limit in the DMF:H2O = 1:1 (v/v) solution.
Keywords:
Pyrazoline
Synthesis
X-ray
Ó 2012 Elsevier B.V. All rights reserved.
UV absorption
Fluorescent sensor
Copper ion
Introduction
depressant [4], antihepatotoxic [5], antitubercular [6], antimalarial
[7] and anti-inflammatory [8]. Furthermore, pyrazoline derivatives
Pyrazolines are important nitrogen-containing 5-membered
heterocyclic compounds with a wide range of interesting proper-
ties, such as antimicrobial [1], antiamoebic [2], anticancer [3], anti-
show stronger fluorescence, and have higher hole-transport effi-
ciency and excellent emitting blueness property. Therefore, pyraz-
oline derivatives have widely been used as whitening or
brightening reagents for synthetic fibers, fluorescent chemosensors
for recognition of transition metal ions, hole-transport materials in
the electrophotography and electroluminescence fields [9–17].
⇑
Corresponding author. Tel.: +86 531 88366425; fax: +86 531 88564464.
E-mail address: bxzhao@sdu.edu.cn (B.-X. Zhao).
1386-1425/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.saa.2012.12.062

S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
111
Meanwhile, pyrazolone-5 derivatives which are similar to pyrazo-
Materials and methods
line form an important class of organic compounds due to their
coordination behaviour with metals [18,19] and biological activity
[20]. Considerable attempts have been made to synthesize and elu-
cidate the effects of substituent on the absorption and fluorescence
properties of pyrazolines with structure diversity [21–26]. More-
over, pyrazoline with membrane permeability, low toxicity, and
high quantum yield render the fluorophore attractive for biological
applications [27,28]. Recently, a pyrazoline as effective ‘‘turn on’’
fluorescent sensors in acetonitrile for Cd2+ and Zn2+ was also re-
ported [29].
The production of fluorescent devices for the sensing and
reporting of chemical events is currently of significant importance
for both chemistry and biology [30]. Among them, sensing of Cu2+
has received ever-increasing attention, since Cu2+ plays a crucial
role as a catalytic co-factor for a variety of metalloenzymes, includ-
ing superoxide dismutase, cytochrome c oxidase, tyrosinase and so
on [31]. However, under overloading conditions, Cu2+ can cause
oxidative stress and disorders associated with neurodegenerative
diseases, such as Alzheimer’s disease, Wilson’s disease, and Men-
ke’s disease, probably by its involvement in the production of reac-
tive oxygen species [32]. Therefore, selective and rapid detection of
Cu2+ in physiological samples is of toxicological and environmental
concern [33–35].
Analytical procedure
A 1.0 ꢀ 10ꢁ3 M of stock solution of compound 3b was prepared
in DMF. The cationic stocks were all in H2O with a concentration of
10ꢁ1 M for UV–vis absorption and fluorescence spectra analysis.
For all measurements of fluorescence spectra, excitation was at
377 nm with 10.5 nm of excitation slit width and scan speed was
set at 600 nm minꢁ1. UV–vis and fluorescence titration experi-
ments were performed using 5 ꢀ 10ꢁ5 M and 1 ꢀ 10ꢁ5 M of com-
pound 3b in the DMF:H2O = 1:1, respectively. For Cu2+ ion
absorption and fluorescence titration experiments, 3 mL solution
of compound 3b (5 ꢀ 10ꢁ5 M) were filled in the quartz cell of
1 cm optical path length, and each time 1.5 l
L solution of Cu2+
(10ꢁ2 M) and 1.0
the quartz cell gradually by using a micro-syringe, respectively.
After each addition of Cu2+ ion, the solution was stirred for
3 min. The volume of cationic stock solution added was less than
l
L solution of Cu2+ (3 ꢀ 10ꢁ3 M) were added into
100 lL with the purpose of keeping the total volume of testing
solution without obvious change.
Oscillator strength and quantum yields
It is known that the aryl substituents in the 1- and 3-position of
the pyrazoline ring influence the photophysical properties of the
fluorophores in distinctly different ways [28,36,37]. Thus, one
might expect to modify the pyrazoline structure by changes of
the substituent in the 1- and 3-position. As an extension of our
work on the development of fluorescent probe for monitoring me-
tal ions [9,10,38–40], in this paper, we synthesized a series of novel
pyrazoline compounds containing 1,3,4-thiadiazole groups in 1-
position using 5-aryl-2-hydrazinyl-1,3,4-thiadiazole and chalcones
(Scheme 1), which had good blue light fluorescence. These com-
pounds have not been reported previously. The compounds were
characterized by IR, 1H NMR, HRMS, X-ray diffraction analysis
and fluorescence techniques. It is found that ICT (intramolecular
charge transfer) process exists between the nitrogen atom at the
1-position and the carbon atom at the 3-position in pyrazoline
moiety, according to the fluorescence analysis. The fluorescence
spectra of compound 3b for detecting Cu2+ were studied. It was
found that the compound 3b exhibited Cu2+-selective and sensitive
chromogenic signaling behavior over other common physiologi-
cally important metal ions. The association constant Ka measured
for coordination of sensor 3b with Cu2+ was 4.56 ꢀ 104 Mꢁ1, and
the detection limit of the sensor 3b toward Cu2+ was 2.46 ꢀ
10ꢁ8 M.
The Stokes shift (Dv) and oscillator strength (f) are important
characteristics for the fluorescent compounds [41]. The Stokes shift
is a parameter that indicates differences in the properties and
structure of the fluorophores between the ground state S0 and
the first excited state S1. The Stokes shift (cmꢁ1) was calculated
by the following equation:
ðVA ꢁ VFÞ ¼ ð1=VA ꢁ 1=VFÞ ꢀ 107
ð1Þ
The oscillator strength (f) shows the effective number of elec-
trons whose transition from ground to excited state gives the
absorption area in the electron spectrum. The oscillator strength
was calculated by Eq. (2). The emax is molar extinction coefficient
and
D
v
1/2 is the width of the absorption band (cmꢁ1) at 1/2 (
e
max).
f ¼ 4:32 ꢀ 10ꢁ9
D
m1=2emax
ð2Þ
The ability for the molecules to emit the absorbed light energy is
characterized quantitatively by the fluorescence quantum yield
UF). Quantum yield was determined by the relative comparison
procedure, using quinine sulfate dehydrate (P99.0%) in 0.1 N
H2SO4 as the main standard. The corrected emission spectra were
measured for the quinine sulfate dehydrate standard (kex = 380 nm;
A (Absorption) <0.01; UF = 0.510) [42]. For all the measurements of
(
Scheme 1. Synthesis of 1,3,5-triaryl pyrazoline derivatives.

112
S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
fluorescence spectra, scan speed was 600 nm minꢁ1 using a quartz
cell of 1 cm optical path length. And the UV–vis absorption spectra
were recorded in a standard 1 cm path length quartz cell in range
250–600 nm with spectral resolution 1 nm. The general equation
used in the determination of relative quantum yields from earlier re-
search was given in Eq. (3) [21].
4-Methyl-2-(5-phenyl-1-(5-phenyl-1,3,4-thiadiazol-2-yl)-4,5-
dihydro-1H-pyrazol-3-yl)phenol (3d). Yellow solid, yield 63.9%; mp
236–238 °C; IR (KBr, cmꢁ1): 3166.4, 3063.1, 3027.2, 2920.1,
1622.3, 1594.3, 1525.2, 1493.6; 1H NMR (300 MHz, CDCl3): d 2.28
(s, 3H, CH3), 3.52 (dd, 1H, J = 6.0, 17.7 Hz, 4-Htrans), 4.12 (dd, 1H,
J = 11.1, 17.7 Hz, 4-Hcis), 5.84 (dd, 1H, J = 6.0, 11.1 Hz, 5-H of pyraz-
oline), 6.99 (d, 2H, J = 8.4 Hz, ArAH), 7.17 (dd, 1H, J = 1.5, 8.4 Hz,
ArAH), 7.30–7.42 (m, 8H, ArAH), 8.05 (dd, 2H, J = 3.0, 6.6 Hz,
ArAH), 9.67 (s, 1H, OH); HRMS: calcd for [M+H]+ C24H20N4OS:
413.1436; found: 413.1461.
UF ¼ ðUFSÞðFAuÞðAsÞðg2uÞ=ðFAsÞðAuÞðgs2
Þ
ð3Þ
where UF and FA are fluorescence quantum yield and integrated
area under the corrected emission spectrum, respectively; A is
2-(1-(5-(4-Methoxyphenyl)-1,3,4-thiadiazol-2-yl)-5-phenyl-4,5-
dihydro-1H-pyrazol-3-yl)-4-methylphenol (3e). Yellow solid, yield
84.7%; mp 236–237 °C; IR (KBr, cmꢁ1): 3206.6, 3068.8, 3031.0,
2931.7, 2910.8, 1606.2, 1572.9, 1493.7, 1483.5; 1H NMR
(300 MHz, CDCl3): 2.29 (s, 3H, CH3), 3.50 (dd, 1H, J = 6.0, 16.8 Hz,
4-Htrans), 3.84 (s, 3H, CH3), 4.10 (dd, 1H, J = 13.2, 16.8 Hz, 4-Hcis),
5.80 (dd, 1H, J = 6.0, 13.2 Hz, 5-H of pyrazoline), 6.93 (d, 2H,
J = 9.0 Hz, ArAH), 7.00 (d, 2H, J = 8.4 Hz, ArAH), 7.17 (d, 1H,
J = 9.0 Hz, ArAH), 7.29–7.42 (m, 5H, ArAH), 7.74 (d, 2H, J = 8.7 Hz,
ArAH), 9.68 (s, 1H, OH); HRMS: calcd for [M+H]+ C25H22N4O2S:
443.1542; found: 443.1522.
2-(1-(5-(4-Chlorophenyl)-1,3,4-thiadiazol-2-yl)-5-phenyl-4,5-
dihydro-1H-pyrazol-3-yl)-4-methylphenol (3f). Yellow solid, yield
55.4%; mp 246–247 °C; IR (KBr, cmꢁ1): 3251.0, 3026.8, 2921.5,
1619.9, 1587.5, 1520.8, 1491.1; 1H NMR (300 MHz, CDCl3): 2.28
(s, 3H, CH3), 3.51 (dd, 1H, J = 5.4, 18.0 Hz, 4-Htrans), 4.12 (dd, 1H,
J = 12.9, 18.0 Hz, 4-Hcis), 5.80 (dd, 1H, J = 5.4, 12.9 Hz, 5-H of pyraz-
oline), 6.98 (d, 2H, J = 8.4 Hz, ArAH), 7.17 (d, 1H, J = 9.3 Hz, ArAH),
7.30–7.41 (m, 7H, ArAH), 7.72 (d, 2H, J = 8.4 Hz, ArAH), 9.63 (s, 1H,
OH); HRMS: calcd for [M+H]+ C24H19ClN4OS: 447.1046; found:
447.1071.
absorbance at the excitation wavelength;
g represent the refractive
index of the solution; and the subscripts u and s refer to the un-
known and the standard, respectively. Furthermore, the energy
yield off fluorescence (EF), which is calculated by Eq. (4), also can
be used instead of the fluorescence quantum yield (UF).
EF ¼ UFðkA=kFÞ
ð4Þ
General procedure for the synthesis of compounds 3a–f
To a stirred solution of substituted chalcone (1) (1.0 mmol) in
ethanol (15 mL) was added 5-aryl-2-hydrazinyl-1,3,4-thiadiazole
(2) (1.2 mmol) and NaOH (3.0 mmol) and the reaction mixture
was refluxed for 3–6 h as shown in Scheme 1. The progress of
the reaction was monitored by TLC. The reaction mixture was al-
lowed to cool to room temperature, and the solvent was evapo-
rated under reduced pressure. The residue was dissolved in
dichloromethane (30 mL), and then washed with water (50 mL)
and brine (20 mL). The organic layer was collected and dried over
MgSO4. After filtration, the solvent was removed under reduced
pressure. The crude product was purified by silica gel column chro-
matography with petroleum dichloromethane/ethyl acetate (15:1;
v/v) as the eluting solvent to give the desired products 3a–3f.
X-ray crystallography
Suitable single crystals of 3b for X-ray structural analysis were
obtained by slow evaporation of a solution of the solid in dichloro-
methane at room temperature for 14 days. The diffraction data
were collected with a Bruker SMART CCD diffractometer using a
The spectroscopic data of compounds 3a–3f
4-Chloro-2-(5-phenyl-1-(5-phenyl-1,3,4-thiadiazol-2-yl)-4,5-
dihydro-1H-pyrazol-3-yl)phenol (3a). Yellow solid, yield 67.4%; mp
228–230 °C; IR (KBr, cmꢁ1): 3192.6, 3063.1, 3026.4, 2921.7,
1614.3, 1587.9, 1522.5, 1483.0; 1H NMR (400 MHz, CDCl3): d 3.48
(dd, 1H, J = 6.6, 17.6 Hz, 4-Htrans), 4.05 (dd, 1H, J = 12.0, 17.6 Hz,
4-Hcis), 5.72 (dd, 1H, J = 6.6, 12.0 Hz, 5-H of pyrazoline), 7.04 (d,
1H, J = 8.9 Hz, ArAH), 7.18 (d, 1H, J = 2.7 Hz, ArAH), 7.31–7.48 (m,
9H, ArAH), 7.80–7.82 (m, 2H, ArAH), 9.89 (s, 1H, OH); HRMS: calcd
for [M+H]+ C23H17ClN4OS: 433.0890; found: 433.0913.
4-Chloro-2-(1-(5-(4-methoxyphenyl)-1,3,4-thiadiazol-2-yl)-5-
phenyl-4,5-dihydro-1H-pyrazol-3-yl)phenol (3b). Yellow solid, yield
80.3%; mp 241–243 °C; IR (KBr, cmꢁ1): 3242.2, 3058.2, 3028.7,
2931.3, 1607.7, 1574.9, 1524.7, 1458.9; 1H NMR (300 MHz, CDCl3):
d 3.46 (dd, 1H, J = 6.3, 17.7 Hz, 4-Htrans), 3.84 (s, 3H, CH3), 4.07 (dd,
1H, J = 12.0, 17.7 Hz, 4-Hcis), 5.76 (dd, 1H, J = 6.3, 12.0 Hz, 5-H of
pyrazoline), 6.92 (d, 2H, J = 9.6 Hz, ArAH), 7.02 (d, 1H, J = 9.0 Hz,
ArAH), 7.16 (d, 1H, J = 2.4 Hz, ArAH), 7.27–7.40 (m, 6H, ArAH),
7.73 (d, 2H, J = 8.7 Hz, ArAH), 9.87 (s, 1H, OH); HRMS: calcd for
[M+H]+ C24H19ClN4O2S: 463.0995; found: 463.0993.
4-Chloro-2-(1-(5-(4-chlorophenyl)-1,3,4-thiadiazol-2-yl)-5-phe-
nyl-4,5-dihydro-1H-pyrazol-3-yl)phenol (3c). Yellow solid, yield
43.3%; mp 267–268 °C; IR (KBr, cmꢁ1): 3435.2, 3199.3, 3057.6,
3023.2, 2922.3, 1615.1, 1588.1, 1519.9, 1483.4; 1H NMR
(300 MHz, CDCl3): d 3.48 (dd, 1H, J = 6.0, 18.0 Hz, 4-Htrans), 4.11
(dd, 1H, J = 12.0, 18.0 Hz, 4-Hcis), 5.82 (dd, 1H, J = 6.0, 12.0 Hz, 5-
H of pyrazoline), 7.01 (d, 1H, J = 9.0 Hz, ArAH), 7.16 (d, 1H,
J = 2.4 Hz, ArAH), 7.27–7.39 (m, 8H, ArAH), 7.70 (d, 2H, J = 8.1 Hz,
ArAH), 9.78 (s, 1H, OH); HRMS: calcd for [M+H]+ C23H16Cl2N4OS:
467.0500; found: 467.0512.
graphite monochromated Mo
Ka radiation (k = 0.71073 Å) at
298 K. The structures were solved by direct methods with SHEL-
XS-97 program and refinements on F2 were performed with SHEL-
XL-97 program by full-matrix least-squares techniques with
anisotropic thermal parameters for the non-hydrogen atoms. All
H atoms were initially located in a difference Fourier map. The
methyl H atoms were then constrained to an ideal geometry, with
CAH = 0.96 Å and Uiso(H) = 1.5 Ueq(C). All other H atoms were
placed in geometrically idealized positions and constrained to ride
on their parent atoms, with CAH = 0.93 Å and Uiso(H) = 1.2 Ueq(C).
Results and discussion
Synthesis of compounds 3
The synthetic routes of the proposed compounds 3 are outlined
in Scheme 1. The chalcone derivatives (1) and 5-aryl-2-hydrazinyl-
1,3,4-thiadiazole (2) were prepared according to the literature
[43,44]. The 3,5-diarylpyrazoline derivatives 3a–f were obtained
by the reaction of chalcone 1 with 5-aryl-2-hydrazinyl-1,3,4-thia-
diazole 2 under reflux condition in 43.3–84.7% yields. The struc-
tures of compounds (3a–f) were confirmed by IR, 1H NMR and
HRMS spectral data. The IR spectra of all the compounds 3a–f
showed v
(C@N) stretch at 1587–1607 cmꢁ1 consisting with pyraz-
oline and thiadiazole moiety. In the 1H NMR spectra of compounds
that is shown in Fig. 1, the CH2 protons of the pyrazoline ring res-
onated as a pair of doublets at d 3.46–3.52 ppm (HA), 4.05–

S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
113
4.12 ppm (HB). The CH proton at C5 also appeared as a doublet of
doublets at d 5.72–5.84 ppm due to vicinal coupling with the
two magnetically non-equivalent protons of the methylene. The
hydroxy proton signal appeared at the range of d = 9.63–
9.89 ppm as a single peak. HRMS showed that found [M+H]-ion
peak accorded with calculated value. Moreover, typically, the
structure of compound 3b was confirmed by X-ray diffraction
analysis.
X-ray crystallography
The spatial structure of compounds 3b was determined by using
X-ray diffraction analysis as shown in Fig. 2. A summary of crystal-
lographic data collection parameters and refinement parameters
for 3b is compiled in Table S1. The structure of compound 3b is
crystallized in monoclinic space group C2/c. Two benzene moieties
and a thiadiazole moiety are bonded to the pyrazoline ring at the
atoms of C7, C9 and N2, respectively. In the crystal of 3b, torsion
angle C(10)AC(9)AN(2)AC(16) of ꢁ73.5(4), shows C10 in the thia-
diazole moiety adopts an antiperiplanar conformation with respect
to the C16 atom of the benzene ring. In the asymmetry unit, the
thiadiazole ring, benzene rings and pyrazoline are almost coplanar.
And the pyrazoline ring makes dihedral angles with benzene A and
thiadiazole ring of 5.0(19)° and 9.4(18)°, respectively, while the
dihedral angle between pyrazoline and benzene A moiety is
82.5(2)°. The dihedral angle of benzene and thiadiazole is 6.8(18).
Regarding the crystal structure of 3b, there is one intramolecu-
lar C(23)AH(23)ꢂ ꢂ ꢂS hydrogen bond forming a pseudo five-mem-
Fig. 2. Molecular structure of compound 3b with displacement ellipsoids drawn at
the 30% probability level.
p–
pꢃ transition of the conjugated backbone localized on the pyraz-
oline ring system [45]. It can be seen that the spectral shapes are very
similar due to these compounds possess a similar structure. But a
small definite substituent effect was observed such that electron-
withdrawing groups (Cl) result in a red shift with respect to an elec-
tron-donating group (MeO). The data indicated that, when a chlorine
group is located on the phenyl ring (C ring), the absorption peaks of
3c and 3f are at longer wavelengths than those of 3b and 3e in which
the methoxyl group is bonded.
Our group has previously reported the synthesis of several pyraz-
oline or bis-pyrazoline derivatives containing pyridazine, thiazole or
ferrocene moiety, which have good blue fluorescence [36,37,46,47].
But the maximum absorptions were fixed at 327, 342, and 347 nm,
respectively, and except the bis-pyrazoline derivatives which the
maximum absorption was fixed at 370 nm. Otherwise, the single-
pyrazoline derivatives we reported in this paper have the minimum
absorption at 366 nm and the maximum at 370 nm. The phenome-
non are attributed to the incorporation of the 1,3,4-thiadiazole en-
hanced the conjugated structure of pyrazoline ring.
bered ring. The molecules are connected by weak
p–p
interactions and further assigned into layers via intramolecular
0
hydrogen0 bond of C(8)A H(8)ꢂ ꢂ ꢂCl, H(8)ꢂ ꢂ ꢂCl, 2.79 ÅA, C(8)ꢂ ꢂ ꢂCl
3.549(3) ÅA, C(8)AH(8)ꢂ ꢂ ꢂCl 140° with the symmetry code of 1/
2 ꢁ x, ꢁ1/2 + y, 3/2 ꢁ z along the b-axis (Fig. S1). Besides the afore-
mentioned hydrogen bonds, there is also a weak CAHꢂ ꢂ ꢂ
p interac-
tion, which is, however, important for the packing modes
C(15)ꢂ ꢂ ꢂCg(2) 2.974(4)°.
Optical properties of compounds 3a–f
UV–vis absorption spectra
The UV–vis absorption spectra of compounds 3a–f shown in
Fig. S2 and Table 1 have been recorded in dichloromethane solution
with the concentration of 2.5 ꢀ 10ꢁ5 M. As we all know, the pyrazo-
Effect of solvent on the absorption spectra
Typical absorption spectra of compound 3b in different solvents
were shown in Fig. S3 in Supplementary data, and the correspond-
ing absorption wavelength maxima and molar extinction coeffi-
line derivative is chromophoric p-system composed of two aryl sub-
stituents in the 1- and 3-position and three out of the five pyrazoline
ring atoms (N1AN2AC3). The remaining two carbon atoms (C4 and
C5) of the ring are sp3 hybridized and are not part of the conjugated
p-system. Absorption spectra of each compound exhibit a strong,
featureless absorption band, which can be assigned to an allowed
cient (e) values were presented in Table 2. As shown in Table 2,
absorption wavelength maxima and molar extinction coefficient
values of compound 3b in different solvents varied from 371 to
366 nm and from 2.46 to 2.90 (104 Mꢁ1 cmꢁ1), respectively. It
was observed that the absorption spectra of these compounds
change very little with increasing solvent polarity indicating that
there is no charge transfer in the ground state [48].
Fluorescence spectra
Following the absorption experiments, steady-state fluores-
cence spectra of compounds 3a–f were obtained in dichlorometh-
ane solution with concentration of 2.5 ꢀ 10ꢁ6 M. As shown in
Table 1 and Fig. 3 the maximal emission bands were all in the
range of 431–457 nm. It can be observed that an electron-with-
drawing group such as chlorine group in benzene ring (A ring)
made the emission wavelength of 3a, 3b and 3c red shifted than
that of compounds 3d, 3e and 3f with methyl group in the benzene
ring. The relative shifts in the emission spectra were attributed to
the nature of the electronic transition (
p–
pꢃ) involving an intramo-
Fig. 1. Structure of compounds 3.
lecular charge transfer from the ArAN1 donor to the ArAAC3@N2

114
S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
Table 1
The optical characteristics of the compounds 3a–f in dichloromethane solution.
Compounds
kabs (nm)
e
(104 Mꢁ1 cmꢁ1
)
kex (nm)
kem (nm)
D
v (cmꢁ1
)
f
UF
EF
3a
3b
3c
3d
3e
3f
368
368
370
368
366
370
2.75
2.90
2.92
2.69
2.74
2.94
373
376
376
369
374
376
443
457
439
431
442
432
4236
4714
3817
3898
4114
3448
0.69
0.76
0.73
0.66
0.63
0.73
0.10
0.20
0.13
0.02
0.07
0.04
0.083
0.161
0.109
0.017
0.058
0.034
Table 2
The optical characteristics of the compound 3b in different solvents.
Compound
kabs (nm)
e
(104 Mꢁ1 cmꢁ1
)
kex (nm)
kem (nm)
v
(cmꢁ1
)
f
UF
EF
DCM
THF
DMF
Acetonitrile
Toluene
368
369
371
367
366
2.90
2.77
2.76
2.75
2.46
376
377
376
363
377
457
457
455
459
455
4714
4643
4618
5762
4547
0.76
0.70
0.72
0.71
0.69
0.20
0.24
0.27
0.19
0.25
0.161
0.194
0.220
0.152
0.201
acceptor. Along with the charge transfer, increased charge densi-
ties were found for the atoms N2 and C3 of pyrazoline ring in
the first excited state S1 [49]. With increasing electron-withdraw-
ing capacity of the ArA, the lone pair electrons on N1 were more
delocalized toward the 3-substituted phenyl ring, and hence the
charge transfer character of the emission state increased. Further-
more, for given substituent ArA, electronic character of substituent
ArC also affected the emission wavelength and intensity, as effec-
tive as the substituent ArA. For example, compounds 3b and 3e
which possess substituent methoxy group in the phenyl ring (C
ring) have red-shift phenomenon compared with 3c and 3f which
possess substituent chlorine. Besides above-mentioned phenome-
non, we also found that the substituent ArC and ArA affected the
fluorescence quantum yields of compounds 3a–f. As listed in Ta-
ble 1, when ArC with p-methoxyl group, the fluorescence quantum
yield was higher than that when ArC with p-chlorophenyl group.
For example, the quantum yields of compound 3b and 3c were
0.20 and 0.13, respectively. In addition, when ArA is with m-chloro
group, the fluorescence quantum yield was higher than that of ArA
with m-CH3 group.
Fig. 3. Fluorescence excitation and emission spectra of compounds 3a–g in
dichloromethane solution with the concentration of 2.5 ꢀ 10ꢁ6 M. (The excitation
and emission slit widths were 10 and 2.5 nm under normal light source,
respectively.)
Effect of solvent on the emission spectra
Fluorescence spectra of compound 3b in various solvents with
different polarity were recorded in Fig. 4 and Table 2. As depicted
in Fig. 4, the emission maxima had a red-shift from 455 to 459 nm
and the Stokes shifts varied from 4547 to 5762 cmꢁ1 with increas-
ing the polarity of the solvent from toluene to acetonitrile. The re-
sults might be resulted from the difference in the dipole moment
of compound 3b in the excited state and the ground state (i.e. ue > -
ug). The relaxed excited state would be energetically stabilized rel-
ative to the ground state with increasing the polarity of solvents
and a significant red-shift of the fluorescence band was observed.
The stronger the interaction of solute and solvent, the lower the
energy of the excited state, and the larger the red-shift of the emis-
sion band and the corresponding Stokes shift [50].
Effect of the concentration on the emission spectra
The effect of the concentration of compound 3b on the fluores-
cence emission intensity was also studied. As shown in Fig. 5 the
fluorescence intensity of the compound 3b in dichloromethane in-
creased initially and then decreased with increasing the concentra-
tion. This phenomenon was attributed to that the increase of
molecule number of the compound 3b with the increase of the
concentration initially resulted in the increase of fluorescence
intensity. However, when the concentration increased over to
Fig. 4. Fluorescence excitation and emission spectra of compound 3b in different
solvents with the concentration of 2.5 ꢀ 10ꢁ6 M. (The excitation and emission slit
widths were 10 and 2.5 nm under normal light source, respectively.)

S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
115
Fig. 6. Absorption spectra of 3b (10 l
M) upon the addition of Cu2+ (0–3.0 equiv.) in
DMF:H2O (1:1, v/v). Insert: Absorption changes of compound 3b at 368 nm upon
Fig. 5. Fluorescence emission spectra of compound 3b in dichloromethane solution
with different concentrations varied from10ꢁ4 to 10ꢁ7 M. (The excitation and
emission slit widths were 10 and 10 nm under 1% attenuation, respectively.)
the addition of Cu2+ (0–3.0 ꢀ 10ꢁ5 M).
function of Cu2+ concentration (Fig. S5). The sensor can be used as
a ratiometric sensor for detecting Cu2+. According to the above phe-
nomena, we can observe the transformation from free compound 3b
to the Cu2+-coordinated species. The coordination stoichiometry be-
tween 3b and Cu2+ ion was estimated to be 1:1 by a nonlinear curve
fitting of the UV–vis titration results (inset) similar to Job’s plot as
depicted in Fig. S6.
5 ꢀ 10ꢁ5 M, the collision of fluorescent molecules with each other
took place to quench the fluorescence [15]. In addition, maximum
emission spectra of the compound 3b in dichloromethane solution
with varied concentrations were unchanged, which means that the
fluorescent molecule structure was not changed, and kept single
molecule in solution with increasing concentration, instead of di-
aggregates, tri-aggregates or poly-aggregates molecule.
Fluorescent sensor for Cu2+
Fluorescent selectivity for Cu2+
To obtain an excellent chemosensor, high selectivity is essential.
Herein, the fluorescence spectra of 3b with various metal ions were
conducted to examine the selectivity. As indicated in Fig. S7 there
was no response to other metal ions except Cu2+ ions. Similarly,
achieving a high selectivity for the analyte of interest over a com-
plex background of potentially competing species is a challenge in
the development of sensors. Thus, the competition experiments
were also conducted in the presence of 5.0 equiv. Cu2+ ion mixed
with 50.0 equiv. other cations. As shown in Fig. S8 only Cr3+, Fe3+
and Al3+ slightly disturbed the intensity by comparison with that
without the other metal ions besides Cu2+. Therefore, these results
UV–vis selectivity for Cu2+ studies
The optical response of the product 3b to various metal ions
was investigated by the UV–visible and emission spectroscopy. It
is found that the compound 3b can be used to determine Cu2+
ion with high selectivity and a low detection limit in the DMF:H2-
O = 1:1 (v/v) solution. The absorption spectrum of compound 3b
exhibits a broad band at 375 nm at room temperature in the
DMF:H2O (1:1, v/v). Binding affinities of compound 3b toward me-
tal ions, Ag+, Al3+, Ba2+, Ca2+, Cd2+, Co2+, Cr3+, Cu2+, Fe3+, Hg2+, K+,
Mg2+, Na+, Ni2+, Pb2+ and Zn2+ ions were evaluated by UV–vis spec-
troscopy measurements. As shown in Fig. S4 upon interaction with
metal ions, significant changes in absorption spectra were ob-
served particularly with Cu2+, the addition of Cu2+ caused a de-
crease of absorption intensity to some extent at 375 nm, and an
new absorption peak appeared obviously at 390–410 nm. Other
metal ions caused relatively minor changes in the absorption spec-
tra of 3b.
UV–vis absorption spectra titration studies
To investigate the binding property of compound 3b toward Cu2+
,
we measured the UV–vis absorption spectra of compound 3b
(1 ꢀ 10ꢁ5 M) in the presence of various concentrations of Cu2+ ion
(0–3 ꢀ 10ꢁ5 M), as shown in Fig. 6. The absorbance of compound
3b at 368 nm gradually decreases with an increasing concentration
of Cu2+ ion. Moreover, two isobestic points at 330 nm and 390 nm
were observed and a new absorption peak appears at the range of
390–410 nm, and its absorption intensity gradually increases with
the addition of Cu2+ ion. This absorption peak is likely due to the
coordination of compound 3b with Cu2+ ion. Under comparable
experimental conditions, the UV–vis spectrum of Cu2+ displayed
no appreciable absorption between 220 and 600 nm within the
appropriate concentration range. There is a linear dependence of
the ratio of absorbance at 400 nm and absorbance at 368 nm as a
Fig. 7. Fluorescence emission spectra of 3b (10 lM) in DMF:H2O (1:1, v/v) upon the
addition of Cu2+ (0–3.0 equiv.). Excitation wavelength was 377 nm. Inset: variations
of fluorescence intensity of compound 3b (10ꢁ5 M) at 454 nm vs. equivalents of
[Cu2+].

116
S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
suggest that 3b has a high fluorescent selectivity for Cu2+ in the
presence of these tested foreign metal ions.
The fluorescence spectra of 3b in different ratios of DMF/H2O
solution were studied. As shown in Fig. S9, we can arrive the con-
clusion that when the percent of DMF from 40 to 80, the compound
3 can well response to Cu2+. Considering the practicability, the
DMF/H2O (v/v = 1:1) is selected for further experiments.
tion of the compound 3b increased over to 5 ꢀ 10ꢁ5 M, the fluores-
cence quenching could be attributed to the collision of fluorescent
molecules with each other. The compound 3b was used for the
determination of Cu2+ ion with high selectivity and a low detection
limit in DMF:H2O = 1:1 (v/v). This sensor formed a 1:1 complex with
Cu2+ and showed a fluorescent quench with good tolerance of other
metal ions. Moreover, this sensor is very sensitive with fluorometric
detection limit of 2.46 ꢀ 10ꢁ8 M.
Fluorescence titration studies
To illustrate the sensitivity of 3b for Cu2+ ion fluorescence titra-
tion experiment was carried out by increasing the concentration of
Cu2+ ion to the solution of 3b in DMF/H2O (v/v = 1:1) at room tem-
perature. As shown in Fig. 7, as Cu2+ ion was gradually titrated, the
fluorescence intensity of compound 3b gradually reduced, and
Acknowledgment
This study was supported by 973 Program (2010CB933504).
Appendix A. Supplementary material
when the amount of Cu2+ ion added was about 1 equiv. (10
lM),
the fluorescence intensity almost reached minimum. When more
Cu2+ was titrated, the fluorescence intensity showed negligible
changes. The nonlinear curve fitting of the fluorescence titration
(inset) also gives a 1:1 stoichiometric ratio between compound
3b and Cu2+ which consist with one by UV–vis spectra as shown
in Fig. 6. The association constant (Ka) between compound 3b
and Cu2+ ion in the DMF:H2O (1:1, v/v) was calculated to be
4.56 ꢀ 104 Mꢁ1 according to the literature [51] (Fig. S10).
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.saa.2012.12.062.
References
[1] Z.N. Siddiqui, T.N.M. Musthafa, A. Ahmad, A.U. Khan, Bioorg. Med. Chem. Lett.
21 (2011) 2860–2865.
[2] F. Hayat, A. Salahuddin, S. Umar, A. Azam, Eur. J. Med. Chem. 45 (2010) 4669–
4675.
[3] P.C. Lv, D.D. Li, Q.S. Li, X. Lu, Z.P. Xiao, H.L. Zhu, Bioorg. Med. Chem. Lett. 21
(2011) 5374–5377.
[4] Z.A. Kaplancıklı, A. Özdemir, G. Turan-Zitouni, M.D. Altıntop, Ö.D. Can, Eur. J.
Med. Chem. 45 (2010) 4383–4387.
[5] H. Khalilullah, S. Khan, M.J. Ahsan, B. Ahmed, Bioorg. Med. Chem. Lett. 21
(2011) 7251–7254.
[6] K. Manna, Y.K. Agrawal, Eur. J. Med. Chem. 45 (2010) 3831–3839.
[7] N.D. Amnerkar, K.P. Bhusari, Eur. J. Med. Chem. 45 (2010) 149–159.
[8] S. Bano, K. Javed, S. Ahmad, I.G. Rathish, S. Singh, M.S. Alam, Eur. J. Med. Chem.
46 (2011) 5763–5768.
Effect of pH on the fluorescence
The pH value of the environment around the fluorescent probe
for metal ions usually shows somewhat of an effect on its perfor-
mance because of the protonation or deprotonation reaction for
the fluorophore or the hydrolysis reaction for the metal ions in the
alkaline condition. The effects of pH on the fluorescence response
of the new probe 3b to Cu2+ were therefore investigated as shown
in Fig. S11. We found that in a solution of DMF/water (1:1) the suit-
able pH span for Cu2+ determination is between pH 4–10. In this re-
gion, the free 3b has no response, while addition of Cu2+ ion can lead
to a remarkable response. The facts suggest efficient complexation
between probe and Cu2+ ion. As a result, our Cu2+-selective receptors
would be an ideal colorimetric chemosensor for monitoring Cu2+ in
aqueous solution in the pH range of 4–10.
[9] Z.L. Gong, B.X. Zhao, W.Y. Liu, H.S. Lv, J. Photochem. Photobiol., A 218 (2011) 6–
10.
[10] Z.L. Gong, F. Ge, B.X. Zhao, Sens. Actuators, B 159 (2011) 148–153.
[11] H.B. Shi, S.J. Ji, B. Bian, Dyes Pigments 73 (2007) 394–396.
[12] J.L. Bricks, A. Kovalchuk, C. Trieflinger, M. Nofz, M. Büschel, A.I. Tolmachev, J.
Daub, K. Rurack, J. Am. Chem. Soc. 127 (2005) 13522–13529.
[13] X.H. Zhang, W.Y. Lai, T.C. Wong, Z.Q. Gao, Y.C. Jiang, S.K. Wu, H.L. Kwong, S.T.
Lee, Synth. Met. 114 (2000) 115–117.
[14] X.C. Gao, H. Cao, L.Q. Zhang, B.W. Zhang, Y. Cao, C.H. Huang, J. Mater. Chem. 9
(1999) 1077–1080.
[15] G. Bai, J.F. Li, D.X. Li, C. Dong, X.Y. Han, P.H. Lin, Dyes Pigments 75 (2007) 93–
98.
[16] G. Chen, H.Y. Wang, Y. Liu, X.P. Xu, S.J. Ji, Dyes Pigments 85 (2010) 194–200.
[17] Y.F. Sun, S.H. Xu, R.T. Wu, Z.Y. Wang, Z.B. Zheng, J.K. Li, Y.P. Cui, Dyes Pigments
87 (2010) 109–118.
[18] K.M. Vyas, R.N. Jadeja, V.K. Gupta, K.R. Surati, J. Mol. Struct. 990 (2011) 110–
120.
[19] K.R. Surati, Spectrochim. Acta Part A Mol. Biomol. Spectrosc. 79 (2011) 272–
277.
[20] H. Huang, Y. Yu, Z.T. Gao, Y. Zhang, C.J. Li, X. Xu, H. Jin, W.Z. Yan, R.Q. Ma, J. Zhu,
X. Shen, H.L. Jiang, L.L. Chen, J. Li, J. Med. Chem. 55 (2012) 7037–7053.
[21] S.M. Song, D. Ju, J.F. Li, D.X. Li, Y.L. Wei, C. Dong, P.H. Lin, S.M. Shuang, Talanta
77 (2009) 1707–1714.
[22] Y.F. Sun, Y.P. Cui, Dyes Pigments 81 (2009) 27–34.
[23] Q. Liu, L. Gao, L. Wang, Z. Xie, D. Li, Spectrosc. Spect. Anal. 29 (2009) 2810–
2814.
Quantitative studies
The quantitative response of compound 3b toward Cu2+ ion was
studied by the fluorescence titration and the linear calibration
plots as shown in Fig. S12. The dynamic range for the determina-
tion of Cu2+ was determined to be linear up to 0–10
l
M with cor-
relation coefficient (R2) of 0.995 [52]. The limit of detection (LOD)
is evaluated using 3 bi/m [53], where bi is the standard deviation
r
r
of the blank signals and m is the slope of the linear calibration plot.
The LOD for determination of Cu2+ was thus calculated to be
2.46 ꢀ 10ꢁ8 M.
[24] K.R. Suratia, B.T. Thaker, Spectrochim. Acta Part A Mol. Biomol. Spectrosc. 75
(2010) 235–242.
Conclusions
[25] W. Kuznik, J. Ebothe, I.V. Kitykc, K.J. Plucinskic, E. Gondekd, P. Szlachcice, T.
Uchaczf, P. Armatysg, Spectrochim. Acta Part A Mol. Biomol. Spectrosc. 77
(2010) 130–134.
In summary, a series of novel pyrazoline derivatives were de-
signed and synthesized in 43.3–84.7% yields. The structures of the
compounds were determined by IR, 1H NMR, HRMS spectra. Repre-
sentatively, the spatial structure of the compound 3b was deter-
mined using X-ray diffraction analysis. The absorption maxima of
the compounds vary from 366 to 370 nm depending on the group
bound to benzene rings. The maximum emission spectra and the
fluorescence quantum yield of the compounds in dichloromethane
are dependent on group in benzene ring. The intensity of absorption
and fluorescence was also correlated with substituents on two aryl
rings. The absorption spectra of these compounds change very little
with increasing solvent polarity. In addition, when the concentra-
[26] Q. Peng, X.H. Tang, Chin. Chem. Lett. 20 (2009) 13–16.
[27] J.F. Li, D.X. Li, Y.Y. Han, S.M. Shuang, C. Dong, Spectrochim. Acta Part A Mol.
Biomol. Spectrosc. 73 (2009) 221–225.
[28] C.J. Fahrni, L.C. Yang, D.G. VanDerveer, J. Am. Chem. Soc. 125 (2003) 3799–
3812.
[29] A. Ciupa, M.F. Mahon, P.A.D. Bank, L. Caggiano, Org. Biomol. Chem. 10 (2012)
8753–8757.
[30] A.P. Silva, H.Q.N. Gunaratne, T. Gunnlaugsson, A.J.M. Huxley, C.P. McCoy, J.T.
Rademacher, T.E. Rice, Chem. Rev. 197 (1997) 1515–1566.
[31] H. Tapiero, D.M. Townsend, K.D. Tew, Copper Biomed. Pharmac. 57 (2003)
386–398.
[32] R.A. Lovstad, Biometals 17 (2004) 111–113.
[33] M.Y. Pamukoglu, F. Kargi, J. Hazard. Mater. 148 (2007) 274–280.
[34] P.G. Welsha, J. Liptona, C.A. Mebaneb, J.C.A. Marra, Ecotoxicol. Environ. Saf. 69
(2008) 199–208.

S.-Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 106 (2013) 110–117
117
[35] P. Kumar, R.K. Tewari, P.N. Sharma, Plant Cell Rep. 27 (2008) 399–409.
[36] Z.L. Gong, L.W. Zheng, B.X. Zhao, D.Z. Yang, H.S. Lv, W.Y. Liu, S. Lian, J.
Photochem. Photobiol., A 209 (2010) 49–55.
[37] W.Y. Liu, Y.S. Xie, B.X. Zhao, B.S. Wang, H.S. Lv, Z.L. Gong, S. Lian, L.W. Zheng, J.
Photochem. Photobiol., A 214 (2010) 135–144.
[44] S.A. Carvalho, E.F. Silva, R.M. Santa-Rita, S.L. Castrod, C.A.M Fraga, Bioorg. Med.
Chem. Lett. 14 (2004) 5967–5970.
[45] C.J. Fahrni, L. Yang, D.G. VanDerveer, J. Am. Chem. Soc. 125 (2003) 3799–3812.
[46] Z.L. Gong, Y.S. Xie, B.X. Zhao, H.S. Lv, W.Y. Liu, L.W. Zheng, S. Lian, J. Fluoresc. 21
(2011) 355–364.
[38] W.Y. Liu, H.Y. Li, H.S. Lv, B.X. Zhao, J.Y. Miao, Spectrochim. Acta Part A Mol.
Biomol. Spectrosc. 95 (2012) 658–663.
[39] W.Y. Liu, H.Y. Li, B.X. Zhao, J.Y. Miao, Org. Biomol. Chem. 9 (2011) 4802–4805.
[40] W.Y. Liu, H.Y. Li, B.X. Zhao, J.Y. Miao, Analyst 137 (2012) 3466–3469.
[41] H. Shakia, K. Gharanjiga, S. Rouhania, A. Khosravi, J. Photochem. Photobiol., A
216 (2010) 44–50.
[42] R.A. Velapoldi, K.D. Mielenz. A Fluorescence Standard Reference Material:
Quinine Sulfate Dihydrate. National Bureau of Standards (Now, The National
Institute of Standards and Technology, NIST) Special Publication, Washington
DC, US, 1980, pp. 260–264.
[47] Z.L. Gong, L.W. Zheng, B.X. Zhao, J. Lumin. 132 (2012) 318–324.
[48] J.F. Li, B. Guan, D.X. Li, C. Dong, Spectrochim. Acta Part
Spectrosc. 68 (2007) 404–408.
A Mol. Biomol.
[49] K. Rurack, J.L. Bricks, B. Schulz, M. Maus, G. Reck, U. Resch-Genger, J. Phys.
Chem. A 104 (2000) 6171–6188.
[50] C. Reichardt, Solvents and solvent effects in organic chemistry, third ed.,
Wiley-VCH, Marburg, 2004. pp. 352–353.
[51] M.J. Yuan, W.D. Zhou, X.F. Liu, M. Zhu, J.B. Li, X.D. Yin, H.Y. Zheng, Z.C. Zuo, C.B.
Ouyang, H.B. Liu, Y.L. Li, D.B. Zhu, J. Org. Chem. 73 (2008) 5008–5014.
[52] N. Li, Y. Xiang, X.T. Chen, A.J. Tong, Talanta 79 (2009) 327–332.
[53] B.P. Joshi, J. Park, W.I. Lee, K.H. Lee, Talanta 78 (2009) 903–909.
[43] A. Budakoti, A.R. Bhat, A. Azam, Eur. J. Med. Chem. 44 (2009) 1317–1325.