Design, synthesis, preliminary pharmacological evaluation, and docking studies of pyrazoline derivatives

Article abstract of DOI:10.2478/s11696-011-0106-2

Ten derivatives of N1 substituted/unsubstituted 5-(4-chlorophenyl)-3-(2- thienyl) pyrazoline were synthesised from chalcone-like intermediate and substituted phenyl hydrazines, hydrazine hydrate, and semi/thiosemicarbazide. The chemical structure of compo

Full text of DOI:10.2478/s11696-011-0106-2

Chemical Papers 66 (1) 67–74 (2012)
DOI: 10.2478/s11696-011-0106-2
ORIGINAL PAPER
Design, synthesis, preliminary pharmacological evaluation,
and docking studies of pyrazoline derivatives
Nirupam Das, Biswajit Dash, Meenakshi Dhanawat, Sushant Kumar Shrivastava*
Department of Pharmaceutics, Institute of Technology, Banaras Hindu University, 221 005 Varanasi, India
Received 30 June 2011; Revised 14 September 2011; Accepted 16 September 2011
Ten derivatives of N1 substituted/unsubstituted 5-(4-chlorophenyl)-3-(2-thienyl) pyrazoline were
synthesised from chalcone-like intermediate and substituted phenyl hydrazines, hydrazine hydrate,
and semi/thiosemicarbazide. The chemical structure of compounds was confirmed by means of
IR, 1H NMR, mass spectroscopy, and elemental analysis. The antidepressant and anticonvulsant
activities were investigated by Porsolt’s behavioural despair test (forced swimming) and maximum
electroshock seizure test, respectively. Rota–Rod test was performed to assess any probable changes
in motor coordination induced by the test compounds. Four compounds (IId, IIg, IIi, and IIj)
exhibited good activity profile against depression and docking studies confirmed their consensual
interaction with monoamine oxidase A. In addition, compounds IIc and IIe showed protection
against MES-induced seizures.
c
ꢀ 2011 Institute of Chemistry, Slovak Academy of Sciences
Keywords: pyrazoline, antidepressant, anticonvulsant, docking
Introduction
the pyrazoline nucleus. The compounds were found
to be highly anti-convulsant while di-substitution of
2-thienyl in position 3,5 confirmed them as potent
anti-depressants. Chimenti et al. (2004, 2008, 2010)
demonstrated that either the N1 acetyl group or N1-
propanoyl substitution in position 1 increased the
potency and selectivity towards the monoamine ox-
idase A (MAO-A) inhibition. The authors recently
reported on several diverse N1-thiocarbamoyl-3,5-
di(hetero)aryl pyrazoline derivatives and observed
that the presence of the chlorine atom in posi-
tion 4 of the 5-phenyl substituent on the pyrazoline
ring was significant for the MAO inhibitory activity.
Both MAO-A and MAO-B are enzymes which con-
tain flavin adenine dinucleotide (FAD) and catalyse
the α-carbon oxidation of a variety of monoamines
such as 5-hydroxytryptamine (serotonin, 5-HT), 4-(2-
aminoethyl)benzene-1,2-diol (dopamine, DA), and 4-
((1R)-2-amino-1-hydroxyethyl)benzene-1,2-diol (nore-
pinephrine, NE) localised in human brain neurons
(Youdim et al., 2006). Since 5-HT is metabolised by
MAO-A in the brain, selective MAO-A inhibitors have
Compounds with pyrazoline ring have received
widespread attention in recent years. They have been
reported as possessing a wide range of biological activ-
ities such as anti-inflammatory, anti-microbial, anti-
androgenic, and anti-thrombotic properties (Fiora-
vanti et al., 2010; Rani et al., 2011; El-Wahab et al.,
2011; Amr et al., 2006; Casimiro-Garcia et al., 2006).
In addition to these effects, in the last decade pyra-
zolines and substituted pyrazolines have emerged as
promising anti-depressant and anti-convulsant agents
(Kaplancikli et al., 2010; Gok et al., 2010; Amnerkar &
Bhusari, 2010; Siddiqui et al., 2009). Of all the synthe-
sised pyrazoline derivatives, the 1,3,5 tri-substituted
derivatives are of particular importance. Prasad et
al. (2005) reported that the compounds possessing
electron-releasing groups on both aromatic rings in
positions 3 and 5 of substituted pyrazolines consider-
ably enhanced the anti-depressant activity. In addi-
tion, Ozdemir et al. (2008) adopted the thienyl func-
tion (a bio-isostere of phenyl group) in position 3 of
*Corresponding author, e-mail: skshrivastava.phe@itbhu.ac.in

68
N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
filtered, dried, and recrystallised from ethanol (Manna
et al., 2002).
General procedure for the synthesis of com-
pounds IIa–IIh
Equimolar concentrations (0.1 mol) of I and 2-, 4-
substituted phenylhydrazine, semi/thiosemicarbazide,
and hydrazine hydrate in ethanol were refluxed for 4
h. The reaction mixture was then poured into ice-cold
water and the precipitate so obtained was washed with
water, dried, and recrystallised from ethanol to afford
the target compounds IIa–IIh.
Fig. 1. Scaffold of the designed pyrazoline derivatives.
been used in the treatment of depression. The in-
crease in the levels of the mood-elevating neurotrans-
mitters 5-HT in the brain alleviates the symptoms of
mood disorders (Fowler et al., 2010). On the basis of
the above findings, we aimed to obtain some N1 sub-
stituted/unsubstituted pyrazoline derivatives with 5-
(4-chlorophenyl)-3-(2-thienyl) pyrazoline as the com-
mon scaffold (Fig. 1), with anticipated anti-depressant
and/or anti-convulsant activities.
General procedure for the Synthesis of com-
pounds IIi–IIj
A mixture of compound I (1.0 mol) and semi/thio-
semicarbazide (1.0 mol) in ethanolic NaOH (0.02 mol,
50 mL) was refluxed for about 2 h. The reaction mix-
ture was then poured into ice-cold water and the
precipitate so obtained was separated by filtration,
washed with water, dried, and recrystallised from
ethanol to afford the target compounds IIi and IIj.
Experimental
All reagents and solvents used in the study were
of analytical grade purity and procured from Sigma–
Aldrich (India). The progress of the reaction was
monitored by thin layer chromatography with hex-
ane/ethyl acetate (ϕr = 3 : 2) as the mobile phase
and performed on Merck silica gel 60 F254 aluminium
sheets (Merck, Germany); the products were purified
by recrystallisation. Melting points were determined
in open capillaries using Stuart SMP10 (Barloworld
Scientific Ltd., UK), electrothermal melting point ap-
paratus and were not corrected. IR spectra were
recorded on a Shimadzu 8400S FTIR (Shimadzu Cor-
poration, Japan) spectrophotometer using KBr pellets
Biological assays
The present biological study was approved by the
Banaras Hindu University Animal Ethical Committee
(No. Dean/10-11/163). All the newly synthesised com-
pounds (IIa–IIj) were tested for their anti-depressant
and anti-convulsant activities. The Rota–Rod test was
performed to assess any probable changes in motor
coordination induced by the test compounds. Seizure
assay was carried out in accordance with the phase-
1 test of the anti-epileptic drug development (ADD)
programme which was developed by the National In-
stitute of Neurological and Communicative Disorders
and Stroke (Krall et al., 1978).
and ν˜ were recorded in cm−1
.
1H NMR (300 MHz)
spectra were acquired on a JEOL AL300 FT-NMR
(Jeol Ltd., Japan) in CDCl3 using TMS as the in-
ternal standard and the chemical shifts were reported
in δ. The mass spectrum was obtained on a Hewlett
Packard model GCD-1800A (Hewlett Packard, USA)
electron impact mass spectrometer at 70 eV ionising
beam and using a direct insertion probe. Elemental
analyses for C, H, and N were performed on Exeter
CE-440 (Hewlett–Packard, USA) elemental analyser.
Anti-depressant activity (Porsolt’s behavioural
despair test)
The synthesised compounds were screened for their
anti-depressant activity using Porsolt’s behavioural
despair (forced swimming) test (Porsolt et al., 1977).
The mice (22–25 g) were housed in groups of six under
standard conditions with access to food and water ad
−1),
libitum. The synthesised compounds (10 mg kg
Synthesis of (E)-3-(4-chlorophenyl)-1-(thio-
phen-2-yl)prop-2-en-1-one (I)
and 3-(10,11-dihydro-5H-dibenzo(b,f )azepin-5-yl)-N,
N-dimethylpropan-1-amine (imipramine) sulphate, as
a reference anti-depressant drug (10 mg kg−1), were
suspended in a 1 % aqueous solution of polyoxyethy-
lene (20) sorbitan monooleate (Tween 80). On the
test-day, the drugs were injected intraperitoneally into
mice in a standard volume of 0.5 mL per 20 g body
mass, 30 min prior to the test. The reference ani-
mals received 1 % aqueous solution of Tween 80. One
Equimolar concentrations (0.04 mol) of 1-thiophen-
2-yl-ethanone and 4-chlorobenzaldehyde were added
to the 10 % aqueous solution of NaOH and ethanol (30
mL) in order to carry out Claisen–Schmidt condensa-
tion. The reaction mixture was then stirred at room
temperature for 3 h and the product thus obtained was

N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
Table 1. Characteristics of the prepared compounds
69
wi(calc.)/%
Compound
Formula
Mr
wi(found)/%
Yield
M.p.
C
H
N
–
%
◦C
I
C13H9ClOS
248.81
338.12
372.67
417.38
368.74
383.09
372.12
352.21
362.13
321.04
305.06
62.78
63.01
67.35
67.53
61.13
60.92
54.63
54.81
65.12
64.89
59.45
59.53
61.13
61.24
68.07
67.42
59.42
59.27
52.25
52.06
54.99
54.85
3.65
3.62
4.46
4.61
3.78
3.65
3.38
3.37
4.65
4.63
3.68
3.69
3.78
3.79
4.86
4.84
4.22
4.29
3.76
3.75
3.96
3.97
82
104–106
IIa
IIb
IIc
IId
IIe
IIf
IIg
IIh
IIi
IIj
C19H15ClN2S
C19H14Cl2N2S
C19H14BrClN2S
C20H17ClN2OS
C19H14ClN3O2S
C19H14Cl2N2S
C20H17ClN2S
C13H11ClN2S
C14H12ClN3S2
C14H12ClN3OS
8.27
8.25
7.50
7.48
6.71
6.69
7.59
7.56
10.95
10.91
7.50
7.48
7.94
71
69
70
67
63
66
68
70
75
87
135–137
130–132
121–123
140–142
137–139
125–127
134–136
133–135
180–182
195–198
7.96
10.66
10.68
13.06
13.01
13.74
13.77
Fig. 2. Synthesis of compounds IIa–IIj.
mouse at a time was transferred into a plexiglass cylin-
der (height of 25 cm, diameter of 20 cm), contain-
ing water of (25
and the mice were observed for 6 min. At the end
of the first 2 min, the animals showing initial vigorous
struggling became immobile. Then the duration of the
2)◦C up to a height of 15 cm

70
N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
phase of immobility in each mouse was measured in
the next 4 min period. The period of immobility was
accounted as passive floating without struggling and
making only those movements which were necessary to
keep its head above the surface of the water. Changes
in the duration of immobilisation were evaluated using
one-way analysis of variance (ANOVA) Dunnet’s post
hoc test (GraphPad Instat Version 3.01) expressed as
binding mode of the ligand with MAO-A, the com-
pounds IId and IIj were computationally docked into
MAO-A (Enzyme Commission number: EC 1.4.3.4)
using Glide. The crystal structure of MAO-A (PDB
ID: 2BXR) retrieved from the protein data bank was
used for molecular docking (De Colibus et al., 2005).
2BXR was subsequently optimised with the “protein
preparation wizard” workflow, as implemented in the
Schro¨dinger 2011 package. A cycle of constrained min-
imisations allowing a maximum root-mean-square de-
means
standard error of the mean (SEM) (Motul-
sky, 1984). A p-value of less than 0.05 was considered
statistically significant.
˚
viation (RMSD) of 0.30 A from the original structure
was carried out. The ligands were built using Maestro
9.2 build panel and prepared by LigPrep 2.5 version
v25111 (Schr¨odinger, LLC, USA) application that uses
OPLS 2005 force field. OPLS stands for optimised po-
tential liquid simulations and it gave the correspond-
ing energy minima 3D conformers of the ligands. A
grid of 17.2612, –25.248, and 13.1624 points in x, y,
and z directions was built, centred on the co-factor
FAD N5 atom. The default settings were used for all
other parameters. The extra precision (GLIDE-XP)
protocol implemented in GLIDE was used for dock-
ing (Glide 5.7, Schro¨dinger Inc., USA) (Halgren et al.,
2004).
Anti-convulsant activity (maximal electroshock
seizure test)
Albino mice of either sex, weighing 20–25 g were
used. Food was withdrawn 12–15 h before commenc-
ing the experiment while water was withdrawn imme-
diately before the experiment. The synthesised com-
pounds were suspended in 30 % of aqueous solu-
tion of poly(ethylene glycol) (PEG 400) and admin-
istered to the mice intraperitoneally in a standard
volume of 0.5 mL per 20 g body mass at a dose of
30 mg kg−1, 100 mg kg−1, 300 mg kg−1. Reference
animals received 30 % aqueous PEG 400 and 5,5-
diphenylimidazolidine-2,4-dione (phenytoin) was used
as a reference drug (10 mg kg−1). Maximal seizure
was induced by application of an electrical stimulus
(50 mA at 60 Hz) of 0.2 s in duration transmitted via
corneal electrodes across the brain after 30 min and
4 h following the drug administration. After applying
the shock, the animals were observed for the type of
convulsion produced and the hind limb extensor re-
sponse was taken as the end point.
Results and discussion
Spectral characterisation
The proposed derivatives were synthesised as il-
lustrated in Fig. 2. Physical properties and elemen-
tal analysis (Table 1) as well as all the spectral data
(Table 2) are in accordance with the structures of the
synthesised compounds. The spectra of I displa−ye1d the
—
characteristic —C O stretching at 1708 cm
and
—
—
—
Rota–Rod performance test
a sharp peak at 1658 cm−1 due to α,β —CH CH.
The derivatives showed diagnostic infrared absorp-
The Rota–Rod test was carried out in accordance
with the method described by Dunham and Miya (Vo-
gel, 2002). The cardinal feature of the test is to ascer-
tain the impairment of motor performance, ataxia, loss
of skeletal muscular strength, and acute neurotoxicity
produced by drugs in pre-clinical studies. Albino mice
weighing between 20–24 g (n = 4–8), where n repre-
sents the number of mice in a group, were trained to
balance on the knurled wooden rotating rod (3.2 cm
diameter) that rotated at 6 min−1. Trained animals
were treated with the test compounds at different dose
tions at 1578–1598 cm−1 for —C N stretching of
—
—
the pyrazoline nucleus. In addition, all the compounds
displayed C4—H deformation (1354–1442 cm−1) and
C5—N1 stretching (1068–1142 cm−1). Compounds IIi
and IIj showed additional thiocarbamoyl NH stretch-
ing vibration (3481 cm−1), carbamoyl NH stretch-
ing (3464–3481 cm−1), —C S stretching at a lower
—
—
frequency of 1345 cm−1, and —C O stretching at
—
—
1684 cm−1 1H NMR spectra of IIa–IIj exhibited
.
three spectral ranges in which each appears as a dou-
ble doublet due to the presence of non-magnetically
equivalent pyrazoline Ha, Hb, and Hx protons. The
geminal pyrazoline proton in position 4 represented
by the Ha and Hb methylene protons displayed sig-
nals at δ 3.04–3.13 (upfield) and δ 3.13–3.93 (down-
field), respectively. The methine proton Hx appeared
further downfield at δ 5.16–5.98 and the aromatic
and thienyl protons at chemical shift in the range
of δ 6.44–7.82. All the other protons belonging to
the methyl and methoxy groups were seen accord-
ing to the expected chemical shift. The mass spec-
levels (30 mg kg−1, 100 mg kg−1, and 300 mg kg−1
)
administered intraperitoneally. After 30 min and 4 h,
respectively, the mice were placed onto the rotating
rod for one minute. Neurological impairment was de-
termined as the inability of the animal to remain on
the rod for 1 min.
Molecular docking
In order to gain some structural insights into the

N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
Table 2. Spectral characterisation of newly prepared compounds
71
Compound
Spectral dataa
—
—
—
I
IR, ν˜/cm−1: 855 (C—Cl stretching), 1658 (α,β CH CH), 1708 (C O)
—
1
—
—
—
H-NMR (CDCl3), δ: 6.65 (d, 1H, —CO—CH ), 7.20–7.68 (m, 7H, Ar—H and thiophene), 7.81 (d, 1H, CH—Ar)
—
MS, m/z: 248.81 (M+
)
IIa
IIb
IR, ν˜/cm−1: 1069 (C5—N1 stretching), 1373 (C4—H deformation), 1591 (C N), 3010 (CH thienyl)
—
—
1H-NMR (CDCl3), δ: 3.05 (1H, dd, Ha), 3.89 (1H, dd, Hb), 5.26 (1H, dd, Hx), 6.93–7.71 (12H, m, thiophene and
Ar—H)
MS, m/z: 338.12 (M+
)
IR, ν˜/cm−1: 1076 (C5—N1 stretching), 1089 (C—S—C stretching), 1383 (C4—H deformation), 1597 (C N), 3012
—
—
(CH thienyl)
1H-NMR (CDCl3), δ: 3.07 (1H, dd, Ha), 3.83 (1H, dd, Hb), 5.21 (1H, dd, Hx), 6.94–7.68 (11H, m, thiophene and
Ar—H)
MS, m/z: 372.67 (M+
)
—
IIc
IId
IIe
IIf
IR, ν˜/cm−1: 1081 (C5—N1 stretching), 1085 (C—S—C stretching), 1367 (C4—H deformation), 1593 (C N), 3050
(CH thienyl)
—
1H-NMR (CDCl3), δ: 3.07 (1H, dd, Ha), 3.86 (1H, dd, Hb), 5.24 (1H, dd, Hx), 6.85–7.64 (11H, m, thiophene and
Ar—H)
MS, m/z: 417.38 (M+
)
IR, ν˜/cm−1: 1068 (C5—N1 stretching), 1082 (C—S—C stretching), 1132 (C—O stretching), 1358 (C4—H defor-
—
mation), 1595 (C N), 3041 (CH thienyl)
—
1H-NMR (CDCl3), δ: 2.78 (3H, s, OCH3), 3.04 (1H, dd, Ha), 3.82 (1H, dd, Hb), 5.16 (1H; dd, Hx), 6.73–7.73 (11H,
m, thiophene and Ar—H)
MS, m/z: 368.74 (M+
)
IR, ν˜/cm−1: 1072 (C5—N1 stretching), 1088 (C—S—C stretching), 1350 (N—O stretching), 1359 (C4—H defor-
—
mation), 1586 (C N), 3017 (CH thienyl)
—
1H-NMR (CDCl3), δ: 3.08 (1H, dd, Ha), 3.75 (1H, dd, Hb), 5.53 (1H, dd, Hx), 7.05–7.82 (11H, m, thiophene and
Ar—H)
MS, m/z: 383.09 (M+
)
IR, ν˜/cm−1: 1087 (C—S—C stretching), 1138 (C5—N1 stretching), 1442 (C4—H deformation), 1595 (C N), 3045
—
—
(CH thienyl)
1H-NMR (CDCl3), δ: 3.13 (1H, dd, Ha), 3.71 (1H, dd, Hb), 5.27 (1H, dd, Hx), 6.79–7.60 (11H, m, thiophene and
Ar—H)
MS, m/z: 372.12 (M+
)
—
IIg
IIh
IIi
IR, ν˜/cm−1: 1079 (C—S—C stretching), 1142 (C5—N1 stretching), 1378 (C4—H deformation), 1578 (C N), 3032
(CH thienyl)
—
1H-NMR (CDCl3), δ: 2.12 (1H, s, CH3), 3.10 (1H, dd, Ha), 3.78 (1H, d, Hb), 5.22 (1H, d, Hx), 6.89–7.65 (11H, m,
thiophene and Ar—H)
MS, m/z: 352.21 (M+
)
IR, ν˜/cm−1: 1075 (C5—N1 stretching), 1091 (C—S—C stretching), 1354 (C4—H deformation), 1597 (C N), 3012
—
—
(CH thienyl), 3290 (NH streching)
1H-NMR (CDCl3), δ: 3.12 (1H, dd, Ha), 3.93 (1H, dd, Hb), 5.29 (1H, dd, Hx), 6.44–7.49 (7H, m, thiophene and
Ar—H), 7.51 (1H, s, NH)
MS, m/z: 362.13 (M+
)
IR, ν˜/cm−1: 1071 (C—S—C stretching), 1085 (C5—N1 stretching), 1345 (C S stretching), 1425 (C4—H deforma-
—
—
—
tion), 1592 (C N stretching), 3024 (CH thienyl), 3481 (NH stretching)
—
1H-NMR (CDCl3), δ: 3.12 (1H, dd, Ha), 3.81 (1H, dd, Hb), 5.98 (1H; dd, Hx), 7.07–7.63 (7H, m, thiophene and
Ar—H), 7.75 (1H, s, NH2)
MS, m/z: 321.04 (M+
)
IIj
IR, ν˜/cm−1: 1072 (C—S—C stretching), 1095 (C5—N1 stretching), 1433 (C4—H deformation), 1595 (C N stretch-
—
—
—
ing), 1684 (C O stretching), 3021 (CH thienyl), 3464 (NH stretching)
—
1H-NMR (CDCl3), δ: 3.08 (1H, dd, Ha), 3.84 (1H, dd, Hb), 5.29 (1H, dd, Hx), 7.04–7.52 (7H, m, thiophene and
Ar—H), 7.56 (2H, s, NH2)
MS, m/z: 305.06 (M+
)
a) Ha and Hb denote the methylene protons and Hx denotes the methine proton of pyrazoline nucleus.
tra of the compounds were studied and the molec-
ular ion peaks ((M)+), which were found consistent
for all the compounds. The elemental analysis re-
Biological screening
In vivo anti-depressant activity of all the com-
pounds was assessed in mice by applying the forced
swimming test. The test is effective in predicting the
sults were within
ues.
0.4 % of the theoretical val-

72
N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
activity of a wide variety of anti-depressants including
MAO inhibitors (Bourin et al., 2002; Petit-Demouliere
et al., 2005). The anti-convulsant activity was evalu-
ated by the MES test and the Rota–Rod test was used
to evaluate neurotoxicity. Pharmacological data of the
compounds are shown in Table 3 and Table 4.
Anti-depressant activity
A few of the compounds tested (Table 3, Fig. 3)
were noted as exhibiting considerable anti-depressant
properties, especially IId, IIg, IIi, and IIj which ex-
hibited remarkable activities revealing the lowest du-
ration of immobility comparable with 10 mg kg−1of
imipramine sulphate used as the reference drug. The
structure-activity relationship based on the results ob-
served indicated that the type of substituent attached
to the N1 of the 5-(4-chlorophenyl)-3-(thiophen-2-yl)-
4,5-dihydro-(1H)-pyrazole scaffold modulated the ac-
Fig. 3. Duration of immobility by Porsolt’s behavioural despair
test; ∗ – significant compared with the reference (Dun-
net’s test; p < 0.05) (n = 6, dose = 10 mg kg−1).
tivity. Attachment of the phenyl group alone does not
elicit such favourable activity as compared with the
substituted aryl nucleus. Electron-withdrawing sub-
stituents decrease the activity in descending order i.e.
the higher the electro-negativity the lower the activity,
Table 3. Anti-depressant activities of compounds IIa–IIj
Duration of immobility/s
Change from reference/%
Compound
Substituent
Mean
SEM
IIa
IIb
IIc
IId
IIe
IIf
IIg
IIh
IIi
H
4-Cl
4-Br
4-OCH3
4-NO2
2-Cl
2-CH3
H
130
109
124
71
145
126
92
101
84
86
4.9
8.1
3.9
5.3
7.9
6.5
5.3
3.2
5.2
4.1
–13.33
–27.33
–17.33
–52.60a
–3.33
–16.00
–38.66a
–32.66
–44.00a
–42.66a
S
O
IIj
Imipramine sulphateb
Control (vehicle)
60
150
2.3
3.5
–60.00
–
a) Significant in comparison with the reference (Dunnet’s test; p < 0.05) (n = 6, dose = 10 mg kg−1); b) dose = 10 mg kg−1
,
intraperitoneally.
Fig. 4. Interacting mode and orientations of compound (a) IId and (b) IIj at the active sites of MAO-A. The FAD co-factor and
the interacting key amino acid residues are shown in ball and stick models. The hydrogen bonds are displayed as dashed
yellow.

N. Das et al./Chemical Papers 66 (1) 67–74 (2012)
73
Table 4. Phase-1 anti-convulsant screening of compounds IIa–
ing that they induced rapid onset of the action while
IIf and IIj elicited late onset of the action. Mostly, the
change in motor coordination was observed for IIb, IIc,
IIg, and IIi at the dose level of 300 mg kg−1. Finally,
it may be concluded that IIc and IIe displayed bet-
ter activity profiles compared with other derivatives
as anti-convulsants with the 4-nitro phenyl derivative
(IIe) considered as the most potent at a dose level of
100 mg kg−1 with a sustained action (Table 4).
IIj
Dose
MESa
30 min
Toxicityb
30 min 4 h
Compound
mg kg−1
4 h
IIa
30
100
300
0/1
0/3
0/1
0/1
1/3
1/1
0/4
0/8
0/4
0/2
0/4
0/2
IIb
IIc
30
100
300
0/1
2/3
1/1
0/1
1/3
0/1
0/4
0/8
1/4
0/2
0/4
1/2
Molecular docking
Compounds IId and IIj representing N1 aryl and
carbamoyl substitutions, respectively, were success-
fully docked at the active site of MAO-A. The bind-
ing configuration of IId (Fig. 4) shows that the 4-
methoxy phenyl is well lodged in the pocket which
is formed due to a cavity-shaping loop, characterised
by ILE 207, SER 209, ARG 206, GLU 216, and TRP
441, whereas the thienyl ring is accommodated in a
pocket surrounded by LEU 337 and PHE 208. The 4-
chloro phenyl ring fits into the aromatic cage formed
by FAD, TYR 407, and TYR 444. These binding
modes were in agreement with most of the suggested
ligand–enzyme contact points reported in the litera-
ture (Karuppasamy et al., 2010; Jia & Zhu, 2010) and
suggested that the compounds might probably act via
the MAO-A inhibition. The compound IIj (Fig. 4) dis-
played a similar binding pattern to IId except that
the N1 carbamoyl group showed hydrogen bonding
with SER 209 and ILE 207, the oxygen serving as
the hydrogen bond acceptor and the amino hydrogen
as the donor. It is also worth noting here that the S-
configuration of IIj binds with both the amino acids
while the R-configuration binds only with SER 209.
30
100
300
1/1
1/3
1/1
0/1
1/3
1/1
0/4
2/8
3/4
0/2
1/4
2/2
IId
30
100
300
0/1
0/3
0/1
0/1
0/3
0/1
0/4
1/8
2/4
0/2
1/4
0/2
IIe
30
100
300
0/1
3/3
1/1
1/1
1/3
1/1
0/4
1/8
1/4
0/2
0/4
0/2
IIf
30
100
300
0/1
0/3
0/1
0/1
1/3
1/1
0/4
0/8
1/4
0/2
0/4
0/2
IIg
30
100
300
0/1
0/3
0/1
0/1
0/3
0/1
0/4
0/8
1/4
0/2
0/4
2/2
IIh
30
100
300
0/1
0/3
0/1
0/1
0/3
0/1
0/4
0/8
0/4
0/2
0/4
0/2
IIi
30
100
300
0/1
0/3
0/1
0/1
0/3
1/1
0/4
0/8
1/4
0/2
2/4
0/2
Conclusion
IIj
30
100
300
0/1
0/3
0/1
0/1
1/3
1/1
0/4
0/8
1/4
0/2
0/4
0/2
A new series of pyrazoline derivatives with a com-
mon skeleton was synthesised and evaluated for their
anti-depressant and anti-convulsant properties. A few
of them are considered to be promising compounds
due to their respective activities. Four compounds
(IId, IIg, IIi, and IIj) exhibited a good activity profile
against depression and docking studies confirmed their
consensual interaction with the MAO-A enzyme. Two
compounds (IIc and IIe) showed protection against
MES-induced seizures. Therefore, the study deserves
further investigation, particularly with respect to that
of the in vitro MAO-A inhibitory activity and si-
multaneous derivatisation of the building block 5-(4-
chlorophenyl)-3-(2-thienyl) pyrazoline.
Phenytoin
30
5/6
5/6
–
–
MES – maximum electroshock seizure. a) Number of animals
protected/number of animals tested; b) Rota–Rod test (number
of animals exhibiting toxicity/number of animals tested).
whereas the electron-releasing substituents appear to
be more favourable. The bio-isosterically-related com-
pounds IIi and IIj showed similar and potent anti-
depressant activity.
Anti-convulsant activity
Out of all the compounds evaluated, IIb, IIc, and
IIe exhibited anti-MES activity at either 100 mg kg−1
or 300 mg kg−1 in 30 min; in addition, IIc was also
active at 30 mg kg−1. The compounds IIb, IIc, and IIe
were more active within 30 min than in 4 h, indicat-
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