Substituent effect on the electron acceptor property of 1,4-benzoquinone towards the formation of molecular complex with sulfamethoxazole

Article abstract of DOI:10.1016/j.saa.2013.01.016

UV-Vis, ¹H NMR, FT-IR, LC-MS and fluorescence spectral techniques were employed to investigate the mechanism of interaction of sulfamethoxazole with varying number of methoxy/chloro substituted 1,4-benzoquinones (MQ_1-4) and to characterize the reaction products. The interactions of MQ_1-4 with sulfamethoxazole (SULF) were found to proceed through the formation of a donor-acceptor complex, containing radical anion and its conversion to the product. Fluorescence quenching studies showed that the interaction between the donor and the acceptors are spontaneous. The results indicated that the progressive replacement of chlorine atom (-I effect) by methoxy group (+M effect) in the quinone decreased the electron acceptor property of the quinone. The results of the correlation of experimentally measured binding constants with electrochemical data and ab initio DFT calculations supported these observations.

Full text of DOI:10.1016/j.saa.2013.01.016

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
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Spectrochimica Acta Part A: Molecular and
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Substituent effect on the electron acceptor property of 1,4-benzoquinone
towards the formation of molecular complex with sulfamethoxazole
⇑
K. Ganesh, A. Satheshkumar, C. Balraj, K.P. Elango
Department of Chemistry, Gandhigram Rural Institute (Deemed University), Gandhigram 624 302, India
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
"
"
"
Novel substituted 1,4-benoquinones
were employed as acceptors in the
CT interaction with a drug.
The mechanism of the interaction
was studied using various spectral
techniques.
Progressive replacement of ACl
(ꢀI effect) by AOMe (+M effect)
makes the acceptor weaker.
0.4
C
0.3
0.2
A
P
0.1
D
0.0
300
400
500
600
700
800
Wavelength (nm)
a r t i c l e i n f o
a b s t r a c t
UV–Vis, 1H NMR, FT-IR, LC–MS and fluorescence spectral techniques were employed to investigate the
mechanism of interaction of sulfamethoxazole with varying number of methoxy/chloro substituted
1,4-benzoquinones (MQ1–4) and to characterize the reaction products. The interactions of MQ1–4 with
sulfamethoxazole (SULF) were found to proceed through the formation of a donor–acceptor complex,
containing radical anion and its conversion to the product. Fluorescence quenching studies showed
that the interaction between the donor and the acceptors are spontaneous. The results indicated that
the progressive replacement of chlorine atom (ꢀI effect) by methoxy group (+M effect) in the quinone
decreased the electron acceptor property of the quinone. The results of the correlation of experimen-
tally measured binding constants with electrochemical data and ab initio DFT calculations supported
these observations.
Article history:
Received 20 June 2012
Received in revised form 28 December 2012
Accepted 10 January 2013
Available online 23 January 2013
Keywords:
Charge transfer
Sulfamethoxazole
Substituent effect
Fluorescence
Ó 2013 Elsevier B.V. All rights reserved.
Theoretical studies
Introduction
sensors [5], organic field effect transistor [6], non-linear optics
[7], magnetic materials [7], organic solar cells [8] and xerogel
Charge transfer (CT) or electron donor acceptor (EDA) com-
plexes constitute a very promising field of study due to its interest-
ing optical and electronic properties [1]. CT is undoubtedly of
fundamental importance in nature and governs a number of
pivotal processes such as photosynthesis and vision [2]. The
fascination of electron donor–acceptor architectures has been
widely recognized in the design of organic optoelectronic materials
for organic light emitting diodes [3], organic photovoltaics [4],
nanoparticles [9]. EDA reactions of certain r and p acceptors have
successfully utilized in pharmaceutical analysis of drugs [10].
Drug-receptor mechanism could be explained by means of CT
phenomenon in addition to weak forces of non-covalent interac-
tions [11]. As a primary step to determine whether CT phenome-
non at any level involved, the ability of the donor drugs and
related compounds to form charge transfer complexes with
acceptors should be studied.
Quinones are important class of organic molecules which nota-
bly plays an essential role in redox reactions such as respiration
and photosynthesis [12]. The key electron acceptors in the
⇑
Corresponding author. Tel.: +91 451 245 2371; fax: +91 451 2454466.
E-mail address: drkpelango@rediffmail.com (K.P. Elango).
1386-1425/$ - see front matter Ó 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.saa.2013.01.016

K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
157
photosynthetic processes are menoquinones, plastoquinones and
O
ubiquinones [13,14]. The biological activity of the quinones is
reportedly due to the redox chemistry of the quinone system
[15]. Quinones are capable of accepting one or two electrons to
form the corresponding radical anion (QÅꢀ) and hydroquinone dian-
ion (Q2ꢀ). Since quinones actively participate in electron transfer
processes, the study of their structural properties is important. It
is well known that any change in the substituent attached to the
quinone ring alters its capability to accept electrons and conse-
quently changes its biological reactions [16–18]. The presence of
methoxy substituent is known to substantially influence the elec-
tron affinity and vibrational spectroscopy of benzoquinones and
has been suggested to be important in determining the function
of ubiquinone as a redox cofactor in bioenergetics [19]. Hence, a
case study of CT interaction between 1,4-benzoquinones possess-
ing varying the number of chloro/methoxy substituents with a
chosen donor drug has been attempted.
R4
R3
R1
R2
2
3
6
5
1
4
O
Acceptor
R1
R2
R3
R4
MQ1
MQ2
MQ3
MQ4
OCH3
OCH3
OCH3
OCH3
Cl
Cl
OCH3
OCH3
Cl
Cl
Cl
Cl
OCH3
OCH3
OCH3
OCH3
The mechanism of interaction of quinone with drugs, in
general, is a research topic of significant interest and hence the
present study. The objective, therefore, of the present article is
to study the spectral, thermodynamic and kinetic aspects of the
interaction of 1,4-benzoquinones possessing varying number of
methoxy/chloro substituents as electron acceptors (MQ1–4) with
sulfamethoxazole (SULF) drug with an aim to investigate the
mechanism of these interactions and to characterize the structure
of the products formed in these interactions. In general, drugs are
poly-functional organic molecules and the present study aims at
to investigate the actual site of attack during the formation of
charge transfer interaction. Such a study would undeniably shed
some light on the mechanism of the drug action in real
pharmacokinetic study. Sulfamethoxazole is chemically known as
4-amino-N-(5-methylisoxazol-3yl)-benzene sulfonamide which is
used as an antibacterial agent and to treat urinary tract infections
[20]. It is also used in the treatment of sinusitis, toxoplasmosis
and pneumocystis pneumonia which affects primarily patients
with HIV. Though these 1,4-benzoquinones are known to organic
chemists as intermediates, it is the first systematic attempt to
utilize them as acceptors in the study of CT complexes with the
drug. Such a structural variation of the quinones would certainly
helps to tune the redox chemistry of them and hence its
biological activity.
Solutions for the spectroscopic measurements were prepared by
dissolving accurately weighed amounts of donor (D) and acceptor
(A) in the appropriate volume of solvent immediately before run-
ning the spectra. The electronic absorption spectra were recorded
on a JASCO (V 630, Japan) UV–Vis double beam spectrophotometer
using 1 cm matched quartz cells by using corresponding pure sol-
vent as reference. The temperature of the cell holder was controlled
with a water flow ( 0.2 °C). The steady state fluorescence spectra
were obtained on a JASCO (FP 6200, Japan) spectrofluorimeter.
The emission slit width (5 nm) and the scan rate (250 nm) was kept
constant for all of the experiments. FT-IR spectra were recorded in a
JASCO (FT-IR 460 Plus, Japan) spectrometer. 1H NMR spectra were
recorded at Madurai Kamaraj University, Madurai in a Brucker
NMR spectrometer (300 MHz, Switzerland). The LCMS spectra were
obtained from University of Hyderabad in a Shimadzu LCMS 2010
with ionization potential at 70 eV (LCMS 2010, Japan).
Synthesis and characterization of substituted quinones
1,4-Benzoquinones possessing varying number of chloro and
methoxy substituents were synthesized and purified as reported
elsewhere [21]. An excess amount of sodium methoxide was added
into a stirred solution of chloranil in methanol at RT under N2 atm.
The reaction mixture was stirred for 12 h at 70 °C and cooled to
room temperature. Then the reaction mixture was added into
200 ml of water and stirred for 1 h at RT. The crude material
formed was filtered through a filter paper and residue was purified
using column chromatography (Silica gel 60–120 by using 5–10%
ethyl acetate pet ether mixture). The percentage yields of various
quinones obtained by this method are shown in Scheme 1. For
the preparation of MQ4, four equivalents of sodium methoxide
were added into MQ1 under the same preparation condition.
Experimental
Material and methodology
The electron acceptors viz. varying number of methoxy/chloro
substituted 1,4-benzoquinones were synthesized and purified by
the reported method [21]. The electron donor drug sulfamethoxa-
zole was obtained as gift sample from a locally available pharma-
ceutical company and was used as received. The purity of the
drug was checked by its melting point (observed 169 °C; literature
169 °C), 1H NMR and FT-IR spectra. Commercially available spec-
troscopic grade solvents (Merck, India) were used without further
purification. The structures of the drug and the acceptors are
shown below.
2,3,5-Trichloro-6-methoxycyclohexa-2,5-diene-1,4-dione (MQ1)
1H NMR (DMSO-d6, 300 MHz), d(ppm) 4.19 (s, 3H), FT-IR
(cmꢀ1): 1680 (C@O), 1668 (C@O), 1567 (C@C), UV–Vis in ethanol
(kmax): 410 nm (n–pꢁ), log
e 2.50, Anal. Calcd. for C7H3Cl3O3: C,
34.82; H, 1.25; found: C, 36.17; H, 1.74; m.p. 172 °C.
2,5-dichloro-3,6-dimethoxycyclohexa-2,5-diene-1,4-dione (MQ2)
1H NMR (DMSO-d6, 300 MHz), d(ppm) 4.07 (s, 3H), 4.12 (s, 3H)
FT-IR (cmꢀ1): 1668 (C@O), 1568 (C@C), UV–Vis in ethanol (kmax):
e 2.50, Anal. Calcd. for C8H6Cl2O4: C, 40.54; H,
2.55; found: C, 41.24; H, 2.66; m.p. 141 °C.
O
H
N
S
412 nm (n–pꢁ), log
O
N
O
2-chloro-3,5,6-trimethoxycyclohexa-2,5-diene-1,4-dione (MQ3)
1H NMR (DMSO-d6, 300 MHz), d(ppm) 3.92 (s, 3H), 4.07 (s, 3H),
4.12 (s, 3H); FT-IR (cmꢀ1): 1666 (C@O), 1568 (C@C), UV–Vis in
NH2
Chemical structure of sulfamethoxazole

158
K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
O
O
O
O
Excess eqv. NaOCH3,
methanol
Cl
Cl
Cl
Cl
Cl
Cl
O
+
Cl
Cl
O
Cl
O
O
+
70°C, 12 h
O
Cl
O
O
O
CHL
O
O
MQ3
10%
MQ1
MQ2
15%
63%
O
O
O
O
O
Cl
Cl
O
4 eqv. NaOCH3,
methanol
70°C, 12 h
O
O
Cl
O
MQ4
72%
Scheme 1. Preparation method for methoxy/chloro substituted quinones (MQ1–4).
ethanol (kmax): 413 nm (n–pꢁ), log
e
2.51, Anal. Calcd. for C9H9ClO5:
C, 46.47; H, 3.90; found: C, 46.17; H, 3.78; m.p. 83 °C.
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
2,3,5,6-Tetramethoxycyclohexa-2,5-diene-1,4-dione (MQ4)
1H NMR (DMSO-d6, 300 MHz), d(ppm) 3.92 (s, 12H), FT-IR
(cmꢀ1): 1668 (C@O), 1568 (C@C), UV–Vis in ethanol (kmax):
416 nm (n–pꢁ), log
e 2.50, Anal. Calcd. for C10H12O6: C, 52.63; H,
5.30; found: C, 53.12; H, 5.58; m.p. 134 °C.
Kinetic procedure
The kinetics of the interaction of SULF with the acceptors (MQ1–4
)
different quinones was followed at three different temperatures in
ethanol solvent under pseudo-first-order conditions, keeping
[D] ꢂ [A]. The increase in absorbance of the new peak at 340 nm
for (SULF–MQ1) and 350 nm for (SULF–MQ2, SULF–MQ3, SULF–
MQ4) with elapse of time were recorded. The pseudo-first-order rate
constants (k1) were calculated from the gradients of log (A1–At)
SULF-MQ1
SULF-MQ2
SULF-MQ3
SULF-MQ4
against time plots, where A and At represent the absorbance at
1
infinity and time t, respectively. The second order rate constants
were calculated by dividing k1 by [D] [22].
0.2
0.3
0.4
0.5
0.6
0.7
0.8
mole fraction of the donor
Results and discussion
Fig. 1. Job’s continuous variation plots for SULF with MQ1, MQ2, MQ3 and MQ4 in
ethanol at 298 K.
Stoichiometry of the interaction
modifications in both donor and acceptor units in the formed prod-
uct relative to the free molecules.
The stoichiometry of the CT complex formed, in all the cases,
was determined by applying Job’s continuous variation method
[23]. In all the cases (SULF–MQ1–4) the symmetrical curve with a
maximum at 0.5 mole fraction indicated the formation of a 1:1
(D:A) CT complex (Fig. 1). The photometric titration measurements
were also performed for the determination of the stoichiometry in
these interactions. The results of the photometric titration studies
(Fig. 2) confirmed the observed stoichiometry of the interaction
[24]. The stoichiometry of the SULF–MQ1–4 systems is further con-
firmed by Jobs continuous variation method using emission studies
also [25] [(Fig. 1Sa) Supplemental information].
In SULF–MQ1 system, some of the significant shifts are: the
peaks due to asymmetric and symmetric vibrations of the aromatic
ring ANH2 group of the free SULF occurred at 3469 cmꢀ1 and
3376 cmꢀ1, respectively and in SULF–MQ1 product it appeared at
3249 cmꢀ1 indicating the participation of NAH moiety in the
product formation. The m(C@O), m(OACH3) and m(CACl) stretching
vibrations in the free MQ1 species appeared at 1684, 1270 and
909 cmꢀ1, respectively. In the product these stretching vibrations
occurred at 1671, 1265 and 896 cmꢀ1
, respectively. Such a
bathochromic shift could be indicative of a higher charge density
on the carbonyl and chloro groups of the MQ1 molecule [26]. These
observations suggested that the NH2 moiety of SULF participated in
the product formation with MQ1.
Characterization of reaction products
In all the cases, the reaction products were obtained by allowing
the reactants (4 mmol of SULF and 4 mmol of MQ1–4 in ethanol) to
react for 24 h under stoichiometric conditions and subjected to
MPLC separation. The FT-IR spectra of the pure SULF, MQ1–4 and
their products were recorded and the peak assignments for
important peaks for SULF–MQ1–4 systems are given in Tables 1–4.
The results indicated that the shifts in positions of some of the peaks
could be attributed to the symmetry and electronic structure
As a representative case, the 1H NMR spectra of the donor SULF
and its reaction product with MQ1 were recorded in DMSO d6 and
are given in [(Figs. 3Sa and 3Sb), supplemental information] respec-
tively. Using the proton NMR technique, we can identify the nature
of interaction between the donor and acceptor in the resulted prod-
uct. Pure SULF molecule exhibited the characteristic peak due to
aromatic ANH2 group of the aromatic ring at 6.087 ppm and in
the product they appeared at 9.620 ppm as ANH group. This

K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
159
observation indicated that the product between SULF and MQ1 is
formed with the elimination of HCl molecule. The aromatic protons
of pure SULF molecule lies in the range of 6.562–7.475 ppm and in
the product/complex lie at 7.144–7.754 ppm. The ANH proton of
the pure SULF occurs at 10.927 ppm and in the product/complex oc-
curs at 11.339 ppm. The down field shifts of most of the signals are
corresponding due to the interaction occurred between the donor
and the acceptor [27,28].
In SULF–MQ1 case, the liberation of HCl was qualitatively ana-
lyzed using litmus paper test (the reaction mixture turns blue lit-
mus to red) and silver nitrate test (water extract of the reaction
mixture gives white precipitate with silver nitrate solution). The
LCMS spectrum, of SULF–MQ1 product was recorded and it exhib-
ited the molecular ion peak at m/z 458 [(Fig. 3Sc) supplemental
information] which confirms the chemical reaction between the
donor and acceptor leading to the product. In the SULF–MQ2–4
cases, the molecular ion peaks at m/z 492, 488, 484 confirm the ad-
duct formation between SULF and the corresponding MQ2–4 [(Figs.
3Sd, 3Se, 3Sf) supplemental information].
colorless SULF and pale yellow colored MQ1, yields a pink colored
solution whose electronic spectrum showed absorption band 460–
600 nm range. This is the characteristic absorption bands of qui-
none radical ion [28,29]. It is observed that with elapse of time
the intensity of these bands decreased (Fig. 3 inset) with a concur-
rent increase in intensity of the band at 415 nm. A clear isosbestic
point is observed at 456 nm. The position of the absorption maxi-
mum of the peak at 415 nm has also been blue shifted with elapse
of time. These observations indicated that the initial reactants
were converted into final product via radical ion formation. For
comparison the electronic spectrum of the product is also shown
in Fig. 3. However, in the case of SULF–MQ2–4 systems, on mixing
the ethanolic solutions of the reactants, the intensity of the kmax
of the acceptor increased with the elapse of time rather than the
appearance of new peaks. A representative electronic spectrum is
shown in Fig. 4. This indicated that the reactants associate through
CT complexation to form an adduct rather than a permanent chem-
ical reaction as occurred in the case of SULF–MQ1 system.
The kinetics of interaction of SULF with MQ1–4 has been followed
by monitoring the increase in absorbance of new peak in ethanol as
a
function of time under pseudo-first-order conditions, i.e.
Interaction of sulfamethoxazole with MQ1–4
[D] ꢂ [A]. The pseudo-first-order rate constant (k1) values for the
formation of the product as a function of [D] and [A] are collected
in Table 5. It is evident from the results in all the cases the rate is
independent of initial concentration of A indicating first order
The electronic spectra of MQ1 in the presence of large excess of
donor, i.e. [D]/[A] > 100 were recorded as a function of time in eth-
anol (Fig. 3). Immediately after mixing ethanolic solutions of
O
N
O
N
O
Cl
Cl
Cl
O
NH
O
NH
O
O
S
S
+
O
O
O
Cl
Cl
Cl
O
O
MQ1
H2N
H2N
CT- Complex
O
O
N
N
O
O
O
O
Cl
Cl
O
NH
NH
O
O
Cl
Cl
S
S
O
+
O
Cl
Cl
O
H
H2N
HN
radical ion pair
-HCl
H
N
O
O
O
S
N
Cl
Cl
O
O
O
N
H
Mechanism of interaction of SULF with MQ1

160
K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
dependence on [A]. The plot of log k1 versus log [D] is linear with a
slope of unity (r > 0.995; slope range 0.007–0.0018) [(Fig. 4S) sup-
plemental information] indicating unit order dependence on [D].
This was further supported by the constancy in k2 values [28].
The pseudo first-order rate constants, for all the systems, were
measured at three different temperatures and the thermodynamic
parameters computed are collected in Table 6. A large negative va-
lue of entropy of activation indicates the involvement of polar tran-
sition state. This may be due to the fact that there is some charge
separation in the transformation of reactants to product. Also the
negative entropy of activation indicated a greater degree of order-
ing in the transition state than in the initial state, due to an in-
crease in solvation during the activation process.
458.4; Anal. Calcd. for C17H13Cl2N3O6S: C, 44.55; H, 2.86;: N,
9.17; found: C, 45.75; H, 2.70;: N, 9.06.
Characteristics of the CT complexes
In all the SULF–MQ1–4 systems, an attempt was made to charac-
terize the CT complexes formed in these reactions. For that the
absorbance of the new bands were measured using constant accep-
tor concentration in ethanol and varying concentrations of the do-
nor but always [D] ꢂ [A]. A representative spectra is shown in
Fig. 6S, [supplemental information]. The nature of the spectra indi-
cated that the interactions between the donor and the acceptor are
of CT type. The formation constants (K) and molar extinction coef-
ficients (e) of the CT complexes were determined spectrophoto-
Based on the foregoing results and discussions the following
plausible mechanism for the interaction SULF with MQ1–4 has been
proposed.
metrically using the Scott equation [30].
½Dꢃ½Aꢃ=d ¼ ½Dꢃ= þ ð1=K
e
eÞ
ð1Þ
where [D] and [A] are the initial molar concentrations of the donor
and acceptor, respectively and d the absorbance. The values of K and
Mechanism of interaction of SULF with MQ1.
Mechanism of interaction of SULF with MQ2–4
.
e
are determined from the gradient and intercept of the linear plot
of [D][A]/d against [D]. Representative Scott plots are shown in
[(Fig. 5S) Supplemental information] and the values of K and thus
Characterization of the interaction product
e
determined are given in Table 7. The observed high values of K sug-
gested that the formed CT complexes are of a strong type [28] and
the linearity of the Scott plots further supported this result.
SULF–MQ1 [3,6-dichloro-2-methoxy-5-(4-amino-N-(5-meth-
ylisoxazol-3-yl)-benzene
dione].
sulfonamide)cyclohexa-2,5-diene-1,4-
1H NMR (DMSO-d6, 300 MHz), d(ppm) 2.29 (s, 3H), 4.17 (s, 3H),
6.14 (s, 1H), 7.14 (d, J = 8.7 Hz, 2H), 7.72 (d, J = 8.7 Hz, 2H), 9.62 (s,
1H), 11.33 (s, 3H). FT-IR (cmꢀ1, KBr): 3249 (NH), 1671 (C@O), 1265,
1037 (AOCH3); LCMS: Calcd. For C17H13Cl2N3O6S: 458.2; found:
Fluorescence studies
The nature and magnitude of the interaction of drugs with
receptors play an important role in the pharmacokinetics of the
O
N
O
N
O
R4
R3
R1
R2
NH
O
NH
O
O
S
S
+
O
O
O
R4
R3
R1
R2
O
MQ2-4
H2N
H2N
CT- Complex
O
N
O
R4
R3
R1
R2
NH
O
S
O
O
H2N
radical ion pair/adduct
Mechanism of interaction of SULF with MQ2-4

K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
161
SULF-MQ1
1.2
0.9
0.6
0.3
0.0
SULF-MQ2
SULF-MQ3
SULF-MQ4
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
Volume of acceptor (ml)
Fig. 2. Photometric titration plots for SULF with MQ1–4 in ethanol at 298 K.
Table 1
Table 2
FT-IR wave numbers (cmꢀ1), intensitya and tentative band assignments for the SULF
and its product with MQ1.
FT-IR wave numbers (cmꢀ1), intensitya and tentative band assignments for the SULF
and its product with MQ2.
SULF
MQ1
SULF–MQ1 product
Assignments
SULF
MQ2
SULF–MQ2 product
Assignments
3469s
3376s
3299s
3012m
2927m
2854m
3432b
3249s
3469s
3378s
3300s
3012m
2927m
2854m
3465m
3378s
3296m
3006w
2925m
2857
1660m
1619
1597
1504w
1471
1365m
1313
1267m
1062m
887w
734
m
(NAH);
m
(NAH);
3006w
2923m
2854
1745m
1671
m
m
m
m
(CAH); aromatic,
(CACH3)
(CAH)
(C@O)
m
m
m
m
m
m
(CAH); aromatic,
(CACH3)
(CAH)
(C@O)
(C@N); isoxazole
(C@C)
1684s
1575s
1663s
1568s
1621
1596s
1502
1471
1365
1310
1621
1596s
1502
1471
1365
1310
1617
1581
m
m
(C@N); isoxazole
(C@C)
d(NAH)
1486w
1363m
1308
1265m
1091m
896w
769
d(NAH)
m
m
m
(CAN)
(SO2)
(OACH3)
m
m
m
(CAN)
(SO2)
(OACH3)
1270m
1060s
908m
734m
1270m
1099s
909m
778m
722m
m(CACl)
m(CACl)
a
s, Strong; m, medium; w, weak; m, stretching; d, bending.
a
s, Strong; m, medium; w, weak; m, stretching; d, bending.
h ¼ ðF0 ꢀ FÞ=F0
ð2Þ
where F and F0 denote the fluorescence intensities of donor in the
presence of acceptor and in the absence of acceptor, respectively.
From the resulting values of h, the association constant Kf for
SULF–MQ1–4 systems was computed using the method described
by Ward [31]. It has been shown that for equivalent and indepen-
dent binding sites (Eq. (3)):
drugs. CT interaction is one of the non covalent binding forces in
the drug–receptor mechanism. In the present study, an attempt
was made to study the CT interaction of SULF with MQ1–4 by means
of fluorescence study. Fluorescence spectra were recorded at room
temperature in ethanol in the range of 300–700 nm using an exci-
tation at 270 nm for SULF. It was observed that the fluorescence of
SULF was quenched by MQ1–4 as a result of formation of CT com-
plex. The experimental results indicated that the quenching effi-
ciency increased with increasing concentration of the electron
acceptor (Figs. 5 and 6) [(Figs. 7Sa, 7Sb), supplemental informa-
tion] and with increasing time. The fraction of acceptors bound
to the donors was determined by using the following Eq. (2).
1=ð1 ꢀ hÞKf ¼ ½ATꢃ=h ꢀ n½DTꢃ
ð3Þ
where n is the number of binding sites [AT], is the total acceptor
concentration and [DT] is the total donor concentration. The plot
1/(1 ꢀ h) versus [AT]/h is linear (r > 0.97, for SULF–MQ1–4 systems)
indicating that under the experimental conditions all the binding

162
K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
Table 3
0.4
FT-IR wave numbers (cmꢀ1), intensitya and tentative band assignments for the SULF
and its complex with MQ3.
C
SULF
MQ3
SULF–MQ3 complex
Assignments
3469s
3376s
3299s
3012m
2927m
2854m
3465m
3378s
3296
3006w
2927m
2854
1679m
1660
1623
1617
1581
1504w 1467
1367m
1308
1280m
1091m
1029m
927w 730
0.3
0.2
0.1
0.0
m
(NAH);
m
m
m
m
(CAH); aromatic,
(CACH3)
(CAH)
(C@O)
400
500
600
700
800
Wavelength (nm)
1674s
1662
1634
A
1621
m
m
(C@N); isoxazole
(C@C)
1596s
1502 1471
1365
1595s
d(NAH)
m
m
m
(CAN)
(SO2)
(OACH3)
P
1310
1277m
1052s
D
940m 747m
v(CACl)
a
s, Strong; m, medium; w, weak; m, stretching; d, bending.
300
400
500
600
700
800
Wavelength (nm)
Fig. 3. Electronic absorption spectra of SULF with MQ1 in ethanol at 298 K (D:
Donor; A: Acceptor; C: complex; P: product) inset: clear isosbestic point at 456 nm.
Table 4
FT-IR wave numbers (cmꢀ1), intensitya and tentative band assignments for the SULF
and its complex with MQ4.
SULF
MQ4
SULF–MQ complex
Assignments
4
3469s
3376s
3299s
3012m
2927m
2854m
3465s
3365s
3301
3006w
2927m
2854
1621m
1598
1598
0 .5
0.35
m
(NAH);
0 .4
m
m
m
m
m
m
(CAH); aromatic,
(CACH3)
(CAH)
(C@O)
(C@N); isoxazole
(C@C)
0.30
P
0 .3
1666s
1606s
0 .2
0.25
1621
1596s
1502
1471
1365
1310
0 .1
1506
d(NAH)
1471w
1367m
1311
1267m
1025m
C
0.20
0.15
0.10
0.05
0.00
0 .0
3 0 0
3 5 0
4 0 0
4 5 0
5 0 0
5 5 0
6 0 0
6 5 0
m
m
m
(CAN)
(SO2)
(OACH3)
A
1276m
1059s
a
s, Strong; m, medium; w, weak; m, stretching; d, bending.
D
sites are equivalent and independent. The value of Kf obtained, from
the plots, for SULF–MQ1, SULF–MQ2, SULF–MQ3 and SULF–MQ4 sys-
tems are found to be 3.6 ꢄ 103, 2.4 ꢄ 103, 2.2 ꢄ 103, and 0.9 ꢄ 103
mol Lꢀ1, respectively. The standard Gibbs energy change
D
G° was
350
400
450
500
550
600
calculated from Kf values using the relation
D
G° = ꢀ2.303 RT log
Kf. The
DG° values for SULF–MQ1–4 systems were found to be
Wavelength (nm)
ꢀ19.6, ꢀ18.7, ꢀ18.4 and ꢀ16.4 kJ molꢀ1 respectively, indicating
that the interaction between the drug and the acceptors is exother-
mic and spontaneous in nature.
Fig. 4. Electronic absorption spectra of SULF with MQ3 in ethanol at 298 K (D:
Donor; A: Acceptor; C: complex; P: product).
Fluorescence quenching can occur by different mechanisms viz.
static or dynamic or both. Stern–Volmer equation (Eq. (4)) is useful
in understanding the mechanism of fluorescence quenching.
The relationship between the fluorescence quenching intensity
and the concentration of quenchers can be described by the follow-
ing equation.
F0=F ¼ 1 þ KSV½Qꢃ
ð4Þ
logðF0 ꢀ FÞ=F ¼ log KA þ n log½Qꢃ
ð5Þ
where F0 is the initial fluorescence intensity measured in the
absence of quencher and F is that in the presence of quencher
concentration [Q]. The Stern–Volmer constant KSV is obtained by
plotting F0/F against [Q]. In all the cases, linear Stern–Volmer
relationship observed (Fig. 7) indicated that either static or dynamic
quenching is dominant [32,33].
where KA is the binding constant and n is the number of binding
sites per donor molecule [34]. In the present study, in all the cases,
a plot of log (F0–F)/F versus log [Q] is linear [(Fig. 8S Supplemental
information 10, r > 0.98)] and the binding constant values
computed are collected in Table 8. The results indicated that, the

K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
163
Table 5
Effect of concentration of the donor and acceptors on the rate of the interaction at 298 K.
[D] (10ꢀ2 M)
[A] (10ꢀ4 M)
k1 (10ꢀ4), sꢀ1
k2 sꢀ1 molꢀ1 dm3
SULF–MQ1
SULF–MQ2
SULF–MQ3
SULF–MQ4
SULF–MQ1
SULF–MQ2
SULF–MQ3
SULF–MQ4
0.5
0.7
0.9
1.1
1.1
1.1
1.1
5.5
5.5
5.5
5.5
4.4
3.3
2.2
1.65
2.45
3.12
3.76
3.06
3.06
3.06
1.24
1.84
2.42
2.94
2.14
2.14
2.14
0.92
1.35
1.69
2.03
1.53
1.53
1.53
0.66
0.85
1.02
1.32
1.17
1.17
1.17
3.4
3.5
3.4
3.4
2.5
2.6
2.6
2.6
1.8
1.9
1.8
1.8
1.3
1.2
1.2
1.2
Table 6
40
30
20
10
0
Kinetic and thermodynamic parameters for the interaction of SULF with MQ1–4 in
ethanol.
D
a
Systems
k
k1 (10ꢀ4) sꢀ1
D
H#
ꢀ
D
S#
D
G#
(nm)
298
305
313 K
SULF–MQ1
SULF–MQ2
SULF–MQ3
SULF–MQ4
346
355
355
355
3.76
2.98
2.03
1.32
4.70
3.42
2.46
1.92
5.60
3.78
3.03
2.12
17
18
19
22
250
253
255
246
93
93
94
94
b
c
d
e
f
D
H# kJ molꢀ1
;
D
S# J Kꢀ1 molꢀ1
;
D
G# kJ molꢀ1
.
g
Table 7
Formation constants of the CT complexes formed between SULF and MQ1–4 in ethanol
at 298 K.
Property
SULF–
MQ1
SULF–
MQ2
SULF–
MQ3
SULF–
MQ4
K (dm3 molꢀ1) Absorption method 2980
2390
5550
1810
3215
1220
1404
e (dm3 molꢀ1 cmꢀ1) Absorption
method
6410
300
350
400
450
500
550
600
650
700
Kf (mol Lꢀ1) Emission method
Stern–Volmer constant KSV
Emission method
3615
3891
2441
2577
2207
2148
939
1918
Wavelength (nm)
Fig. 6. Variation of fluorescence spectra of SULF–MQ2 system in ethanol at fixed
concentration [D] = {2.8125 ꢄ 10ꢀ3 (curve D)} and variable concentration of
[A] ꢄ 10ꢀ4 = {1.3875 (curve a)}, 2.775 (curve b), 4.1625 (curve c), 5.55 (curve d),
6.9375 (curve e), 8.325 (curve f), 9.7125 (curve g) mol Lꢀ1 at 298 K.
60
D
5
SULF-MQ1 r=0.99
50
SULF-MQ2 r=0.99
SULF-MQ3 r=0.99
SULF-MQ4 r=0.98
a
40
b
c
d
4
3
2
1
30
e
f
20
10
0
320
360
400
440
480
Wavelength (nm)
0.0000
0.0002
0.0004
0.0006
0.0008
0.0010
Fig. 5. Variation of fluorescence spectra of SULF–MQ1 system in ethanol at fixed
concentration [D] = {2.8125 ꢄ 10ꢀ3 (curve D)} and variable concentration of
[A] ꢄ 10ꢀ4 = {1.3875 (curve a)}, 2.775 (curve b), 4.1625 (curve c), 5.55 (curve d),
6.9375 (curve e), 8.325(curve f) mol Lꢀ1 at 298 K.
[Q]
Fig. 7. Stern–Volmer plots for the fluorescence quenching of SULF with the
acceptors MQ1–4 in ethanol at 298 K.

164
K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
Table 8
Table 9
Binding constants (KA) and number of binding sites (n) for SULF–MQ1–4 systems in
Reversible voltammetric parameters for the acceptors.
ethanol.
Acceptor
Epc
Epa
E1/2
ipa/ipc
Acceptors
KA (molꢀ1 L)
n
MQ1
MQ2
MQ3
MQ4
ꢀ0.12
ꢀ0.29
ꢀ0.43
ꢀ0.62
ꢀ0.02
ꢀ0.18
ꢀ0.33
ꢀ0.50
ꢀ69
1.1
0.9
1.1
1.1
SULF–MQ1
SULF–MQ2
SULF–MQ3
SULF–MQ4
3.2 ꢄ 103
2.6 ꢄ 103
1.7 ꢄ 103
0.6 ꢄ 103
1.1
0.9
0.9
0.8
ꢀ232
ꢀ383
ꢀ556
Fig. 8. The optimized structures of the SULF and MQ1–4 along with frontier orbitals.

K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
165
computed using absorption spectral data follows the same order
(Table 6). Also, the rate constant for the interaction, in these cases,
decreased with an increase in methoxy groups in the acceptor
(Tables 6 and 8). The value of n, for the systems, is nearly constant
in ethanol indicating the presence of equivalent binding sites.
-3.0
-3.5
-4.0
-4.5
-5.0
-5.5
-6.0
-6.5
MQ4
LUMO
MQ3
MQ2
Electrochemical studies
MQ1
The redox potentials of the acceptors were measured by cyclic
voltametry at room temperature using a conventional two-com-
partment three electrode cell with a 3 mm Glassy Carbon (GC)
disk as working electrode and 0.1 M tetrabutylammonium per-
chlorate as the electrolyte in acetonitrile. For all the acceptors,
the voltagrams, recorded in the potential range from +1.5 V to
ꢀ0.8 V versus Ag/AgCl are shown in [(Fig. 9S) Supplemental infor-
mation]. To compare the electron accepting properties of these
acceptors the electrochemical data of the first wave alone has
been considered here. The half wave potential values (E1/2) were
evaluated from the voltammograms obtained at a sweep rate of
100 mV sꢀ1, E1/2 = (Epa + Epc)/2, where Epa and Epc correspond to
anodic and cathodic peak potentials, respectively. In all the cases,
the ratio of ipa/ipc, was found to be nearly unity (Table 9) indicat-
ing reversible nature of the systems [15,16]. The results obtained
indicated that the E1/2 values become more negative from MQ1 to
MQ4. That is the electron accepting property (reduction) of MQ4 is
relatively low when compared to MQ1 or in other words MQ4 is
comparatively a weaker acceptor. This may be due to the fact that
progressive replacement of electron withdrawing chlorine atom
(ꢀI effect) by electron releasing methoxy group (+M effect) ren-
dered the quinone increasingly electron rich and consequently
make it as a weak acceptor. This observation corroborates well
with the results obtained in the spectral and kinetic studies enu-
merated above.
HOMO
Fig. 9. Energy of HOMO of the donor and LUMO of the acceptors.
4.0
SULF-MQ1
3.5
3.0
SULF-MQ2
2.5
SULF-MQ3
Theoretical calculations
2.0
To understand the foregoing experimental observations on the
CT complex formed between SULF and the acceptors, we have per-
formed the optimization of SULF, MQ1–4 using Density Functional
Theory with the Backle3LYP hybrid functional, by using a basis
set of 6-31G. Computations have been performed using the Gauss-
ian 03 Revision D.01 program package [35]. The optimized geome-
try of the donor along with HOMO and the acceptors along with
LUMO are depicted in Fig. 8. In the case of SULF, the HOMO is con-
centrated on the ANH2 group and in the case of the acceptors the
LUMO resides on the quinone ring.
1.5
SULF-MQ4
1.0
0.5
2.0
2.2
2.4
2.6
2.8
3.0
3.2
3.4
The energies of the frontier orbitals of the donor and the accep-
tors along with the energy corresponds to the CT transition,
ΔΕ
Fig. 10. Correlation between association constant Kf and
DE.
D
E(=HOMOSULF–LUMOacceptor
)
[36,37], for all the systems are
E depends
shown in Fig. 9. It is evident from the figure that the
D
on the nature of the substituent present in the quinone. Also, a
good linear correlation obtained between the theoretical energy
gap values and the experimentally determined stability constant
values (Fig. 10) indicated that the strength of the acceptor decrease
in the order:
binding constant value decreased from MQ1 to MQ4, i.e. the
magnitude of binding constant is in the order of SULF–MQ1 >
SULF–MQ2 > SULF–MQ3 > SULF–MQ4. These observations are in cor-
roboration with the results of absorption spectral studies as enu-
merated earlier in this paper. That is the formation constant
O
O
O
O
Cl
O
O
O
O
O
Cl
O
Cl
O
O
>
>
>
O
Cl
Cl
Cl
O
O
MQ2
O
MQ4
O
MQ1
O
MQ3

166
K. Ganesh et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 107 (2013) 156–166
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