Design, synthesis and potential 6 Hz psychomotor seizure test activity of some novel 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one

Article abstract of DOI:10.1016/j.ejmech.2011.01.009

Thirty new 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one were designed and synthesized keeping in view the structural requirement of pharmacophore and evaluated for anticonvulsant activity and neurotoxicity. The anticonvulsant activity of the titled compounds was assessed using the 6 Hz psychomotor seizure test. The most active compound of the series was 3-({(E)-[3-(4-chloro-3-methylphenoxy)phenyl]methylidene}amino) -2-phenylquinazolin-4(3H)-one PhQZ 7, which showed 100% protection (4/4, 0.5 h) and 75% protection (3/4, 0.25 h) at a dose of 100 mg/kg in mice. A computational study was carried out for calculation of pharmacophore pattern and prediction of pharmacokinetic properties. Titled compounds have also exhibited good binding properties with epilepsy molecular targets such as glutamate, GABA (A) delta and GABA (A) alpha-1 receptors, in Lamarckian genetic algorithm based flexible docking studies.

Full text of DOI:10.1016/j.ejmech.2011.01.009

European Journal of Medicinal Chemistry 46 (2011) 1006e1018
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Design, synthesis and potential 6 Hz psychomotor seizure test activity of some
novel 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one
,
c
a
b
Praveen Kumar c , Birendra Shrivastava , Surendra N. Pandeya , James P. Stables
*
a Saroj Institute of Technology and Management, Lucknow 226 002, Uttar Pradesh, India
b Preclinical Pharmacology Section, Epilepsy Branch, National Institute of Health, Bethesda, MD 20892-9020, USA
c School of Pharmaceutical Sciences, Jaipur National University, Jaipur-302025, Rajasthan, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Thirty new 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one were designed and synthesized
keeping in view the structural requirement of pharmacophore and evaluated for anticonvulsant activity
and neurotoxicity. The anticonvulsant activity of the titled compounds was assessed using the 6 Hz
psychomotor seizure test. The most active compound of the series was 3-({(E)-[3-(4-chloro-3-methyl-
phenoxy)phenyl]methylidene}amino)-2-phenylquinazolin-4(3H)-one PhQZ 7, which showed 100%
protection (4/4, 0.5 h) and 75% protection (3/4, 0.25 h) at a dose of 100 mg/kg in mice. A computational
study was carried out for calculation of pharmacophore pattern and prediction of pharmacokinetic
properties. Titled compounds have also exhibited good binding properties with epilepsy molecular
targets such as glutamate, GABA (A) delta and GABA (A) alpha-1 receptors, in Lamarckian genetic
algorithm based flexible docking studies.
Received 17 November 2010
Received in revised form
25 December 2010
Accepted 10 January 2011
Available online 15 January 2011
Keywords:
Quinazolin-4(3H)-one
6 Hz psychomotor seizure test activity
Neurotoxicity
Ó 2011 Elsevier Masson SAS. All rights reserved.
Computational study
1. Introduction
methyl group at position 2 of synthesized compounds by phenyl
and n-propyl group and to report their effect on anticonvulsant
Quinazolin-4(3H)-one 1 and their derivatives constitute an
important class of heterocyclic compounds and are shown to have
potent CNS activities such as anticonvulsant [1e5] and CNS
depressant [6,7]. A literature survey revealed that the presence of
a substituted aromatic ring at position 3 and a methyl group at
activity. The second objective was based upon the hypothesis that
methyl group at the second position of quinazolin-4(3H)-one is not
always necessary for the CNS activity and other groups when
placed at this position can also lead to potent CNS active agents
[9,10]. The 2-substituted quinazolin-4(3H)-one was also coupled
with isatin, a well known anticonvulsant.
position
2 on quinazolin-4(3H)-one nucleus is a necessary
requirement for CNS depression and anticonvulsant activities.
Modification of the methyl group by some other chemical moiety
yielded structural analogs with potent CNS activity. Methaqualone
Based on the literature review, we are the first to report the
synthesis and anticonvulsant activities of 2-(substituted)-3-
{[substituted]amino}quinazolin-4(3H)-one. Their chemical struc-
tures were characterized using IR, 1H NMR, MS and elemental
analysis techniques. All the synthesized titled compounds
comprised of the essential pharmacophoric elements (Fig. 3) that
are necessary for good anticonvulsant activity as suggested by
Unverferth et al. [11]. In addition, their anticonvulsant activity was
evaluated by using 6 Hz psychomotor seizure test in mice. The
rotarod assay was performed in mice to evaluate the neurotoxicity
of the compounds. Computational study was also carried out to
highlight the pharmacophore distance mapping, log P calculation
and pharmacokinetic parameters. In this study, we have used
Autodoc 4.0 along with its LGA algorithm for automated flexible
ligand docking of compounds with six established epilepsy
molecular targets and evaluated their affinity and hydrogen
bonding.
2
(2-methyl-3-o-tolyl-4(3H)-quinazolinone) is a well known
sedative-hypnotic containing quinazolin-4(3H)-one nucleus. In
fact, these evidences suggest that the quinazolin-4(3H)-one
nucleus possess a pharmacophoric character for CNS activity. In
addition, the 4-(aryloxy) phenyl semicarbazones 3 (Fig. 1) were
reported as potential anticonvulsant [8]. The design and synthesis
of the titled compounds were carried out with two objectives: the
first was the molecular hybridization of Methaqualone, having
Quinazolin-4(3H)-one nucleus and 4-(aryloxy) phenyl semi-
carbazones (Fig. 2). The second objective was the replacement of
* Corresponding author. Tel.: þ91 9897112288.
E-mail address: praveensha1977@rediffmail.com (P. Kumar).
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.01.009

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1007
H3C
N
The structural assignments to new compounds were based on
their elemental analysis and spectral (FT-IR, 1H NMR and mass)
data. The formation of 4-substituted benzaldehydes 3a-i from 4-
substituted phenols 1a-i was confirmed by its IR and 1H NMR
spectral studies. The IR spectrum of 4-(4-fluorophenoxy) benzal-
dehydes 3f showed bands at 1665, 3035 and 1240 cmꢀ1 indicating
the presence of C]O stretching, CeH stretching and diaryl ether
linkage (eOe). The absence of broad band of phenolic eOH at
2900e3000 cmꢀ1 confirms the conversion of 4-fluoro phenol 1f to
O
O
N
O
N
C
H
O Ar
NH
H2N
N
H
N
CH3
(1)
(2)
(3)
Fig. 1. Chemical structure of quinazolin-4(3H)-one (1), Methaqualone (2) and 4-(ary-
loxy)phenyl semicarbazone (3).
3f. In its 1H NMR spectrum a singlet at
presence of eCHO group, where as absence of a singlet around
d 9.87 ppm indicating the
2. Chemistry
d
9.288 ppm for phenolic eOH confirms the conversion of 1f to 3f.
The IR spectrum of the titled compound PhQZ 7 over the
The reaction sequence leading to the synthesis of titled
compounds, viz. 2-(substituted)-3-{[substituted]amino}quinazo-
lin-4(3H)-one (PhQZ 1e10, MtQZ 1e10 and PrQZ 1e10) is shown in
Scheme 1. The 4-substituted benzaldehydes 3aei were prepared
by refluxing various substituted phenol 1aei with 4-fluoro benz-
aldehyde 2 in N,N-DMF in presence of potassium carbonate. The
2-phenyl-4H-3,1-benzoxazin-4-one 5 was obtained by reacting
anthranilic acid 4 with benzoyl chloride in pyridine. The 2-methyl-
4H-3,1-benzoxazin-4-one 6 was obtained by refluxing anthranilic
acid 4 in acetic anhydride. Anthranilic acid 4 was refluxed in butyric
anhydride to obtain N-propyl anthranilic acid 7, which was further
refluxed with acetic anhydride to obtained 2-propyl-4H-3,1-ben-
zoxazin-4-one 8. The 2-substituted-4H-3,1-benzoxazin-4-ones (5,
6, 8) were converted to their corresponding amines i.e., 3-amino-2-
(substituted)-quinazolin-4(3H)-one (9, 10, 11) by reacting with
hydrazine hydrate. The 3-amino-2-(substituted)-quinazolin-4(3H)-
ones (9, 10, 11) were condensed with various 4-substituted benz-
aldehyde 3aei/isatin 3j to yield the titled compounds PhQZ 1e10,
MtQZ 1e10 and PrQZ 1e10. Thin layer chromatography (TLC) was
run throughout the reactions to optimize the reactions for purity
and completion. The physical and elemental analyses data of the
titled compounds are summarized in Table 1.
3067 cmꢀ1 showed multiple weak absorption peak corresponding
to Qu-H and AreH stretching vibration. The strong absorption at
1685 cmꢀ1 is due to the C]O stretching vibration and the moderate
intensity absorption at 1596 cmꢀ1 corresponds to a CH]N
stretching vibration. The 1447e1526 cmꢀ1 absorptions are due to
the skeleton vibration of the aryl and heterocyclic rings. The strong
absorption at 1240 cmꢀ1 corresponds to diaryl ether linkage (O).
The absorption peaks at 755 and 699 cmꢀ1 arise due to the phenyl
substitution at 2-position in the quinazolinone. Its 1H NMR spec-
trum showed a singlet at
the imine carbon. A set of signals appear between
8.39 ppm corresponds to aromatic and quinazolinone protons. The
methyl protons appear at 2.36 ppm. The presence of CH]N
stretching vibration at 1596 cmꢀ1 in IR spectrum and a singlet for
proton attached to the imine carbon at 8.45 ppm confirm the
d
8.45 ppm due to the proton attached to
d
6.82 and
d
d
formation of PhQZ 7. Further mass spectrum confirmed their purity
and molecular weight.
3. Pharmacology
The newly synthesized 2-(substituted)-3-{[substituted]amino}
quinazolin-4(3H)-one derivatives (PhQZ 1e10, MtQZ 1e10, PrQZ
1e10) were subjected to anticonvulsant screening according to
anticonvulsant drug development (ADD) program protocol. The
profile of anticonvulsant activity of six titled compounds was
established after i.p. injections into mice and compounds were
evaluated in the maximal electroshock (MES) and subcutaneous
metrazole (scMET) model using doses of 30, 100 and 300 mg/kg at
two different time intervals. The twenty four titled compounds
were screened in 6 Hz psychomotor seizure test to identify their
anticonvulsant activity at five different time points, i.e., 0.25 h,
0.5 h, 1.0 h, 2.0 h and 4.0 h after i.p. administration in mice. The
results are shown in Table 2. Compound showing potential activity
was also subjected to quantification studies in 6 Hz test and
neurotoxicity. The corresponding an ED50 and a TD50 were reported
in Table 3. The most active compound was screened in in-vitro
hippocampal slice culture neuroprotection assay (NP) and results
are shown in Table 4. Neurotoxicity was observed by minimal
motor impairment which was measured by the rotarod test.
CH3
O
O
N
H2N
C
N
H
N
C
H
O Ar
N
CH3
(3)
(2)
Molecular Hybridization
O
N
N
C
O
Ar
H
N
CH3
(MtQZ 1-9)
4. Computational study
Replacement of 2-methyl by
2-phenyl and 2-propyl group
The pharmacophore pattern studies in which distance between
the various groups postulated as essential for anticonvulsant
activity were done on the 3D optimized structures using ACD/3D
viewer version 12.0 and Argus Lab 4.0 Mark A. Thompson Planaria
Software LLC. A computational study of all titled compounds was
performed for prediction of ADME properties such as absorption (%
ABS), polar surface area (TPSA), milog P, number of rotatable bonds,
and violations of Lipinski’s rule of five by using molinspiration
online property calculation toolkit. Calculated milog P for synthe-
sized compounds were then compared with the experimental log P
O
O
N
N
N
C
O Ar
N
N
C
O Ar
H
H
N
C3H7
C6H5
(PhQZ 1-9)
(PrQZ 1-9)
Fig. 2. Rational concept to new 2-(substituted)-3-{[substituted]amino}quinazolin-4
(3H)-ones PhQZ 1e10, MtQZ 1e10 and PrQZ 1e10.

1008
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
D
N
D
CH3
A
D
S
H
H
HD
OH
CH3
A
N
O HA
HD
N
N
H
A
S HA
HA
O
HD H2N
Albutoin
Carbamazepine
Gabapentin
D
HA
O
D
N
H
H
N
O
A
H
NH
HD
N
Cl
HD
C2H5
O
Cl
N
H
N
N
O
N
N
O
HA
D
H
HD
HA
CH3
H
Lamotrignine
Mephobarbital
Phenytoin
HA
O
N
O
A
OH
D
H
HD
H3C
O
S
N
H
O
HA
D
HD
H
N
N
N
A
F
HA
O
H
H
N
H
H
Zonisamide
A
HD
Remacemide
Progabide
Cl
O
O
HA
C6H5
H3C
D
CH3
O
N
N
A
N
N
C
Cl
H
S
HA
N
H
HD
CH3
O
Cl
H
A
D
HD
PhQZ 7
HA
Ralitoline
D
O
O
HA
CH3
N
N
N
CH3
A
N
N
C
HD
NH
H
HD
N
C3H
7 O
O
A
D
MtQZ 3
PrQZ 10
Fig. 3. Pharmacophoric pattern of well-known anticonvulsants and 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one PhQZ 7, MtQZ 3 and PrQZ 10.
data of these compounds. Docking study of titled compounds was
performed with six established epilepsy molecular targets namely
GABA (A) alpha-1 receptor, GABA (A) delta receptor, glutamate
receptor, Na/H exchanger, Na channel receptor, T-type calcium
channel receptor by using Autodoc 4.0 along with its LGA algorithm
for automated flexible ligand docking and affinity (kcal/mol) and
hydrogen bonding were evaluated.
6 Hz psychomotor seizure or minimal clonic seizure test was used
to identify the anticonvulsant activity of compounds at five
different time points, i.e., 0.25 h, 0.5 h, 1.0 h, 2.0 h and 4.0 h after i.p.
administration in mice at a dose of 100 mg/kg. The 6 Hz psycho-
motor seizure test is used to assess a compound’s efficacy against
electrically induced seizures but uses a lower frequency (6 Hz) and
longer duration of stimulation (3 s). It is used to identify clinically
useful AEDs that are ineffective in the standard MES and scMET
tests but still have anticonvulsant activities in-vivo. Neurotoxicity
was observed by minimal motor impairment which was measured
by the rotarod (neurotoxicity) test. The results of screening at five
different points are summarized in Table 2. As observed from the
results of various tested 2-(substituted)-3-{[substituted]amino}
quinazolin-4(3H)-one derivatives PhQZ 7, MtQZ 3 and PrQZ 10
showed good activity with significant protection. Compound PhQZ
7 showed 100% protection (4/4, 0.5 h), 75% protection (3/4, 0.25 h)
and 25% protection (1/4, 1.0 and 2.0 h) at a dose of 100 mg/kg.
Compound PhQZ 6 showed 50% protection (2/4, 0.25 and 0.5 h) and
25% protection (1/4, 1.0 h) at a dose of 100 mg/kg. Compound MtQZ
5. Result and discussion
5.1. Anticonvulsant and neurotoxicity evaluation
The six newly synthesized 2-(substituted)-3-{[substituted]
amino}quinazolin-4(3H)-one derivative (PhQZ 1e4, PhQZ 8 and
10) were screened in the maximal electroshock (MES), subcuta-
neous metrazole (scMET) model to identify their anticonvulsant
activity. None of them showed activity in MES and scMET screen.
The rest twenty six titled compounds were screened in 6 Hz
psychomotor seizure test and the results were found promising.

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1009
O
F
K2CO3, DMF
Ar
Ar OH
CHO
CHO
O
3a-i
1a-i
2
O
NH2NH2.H2O
Pyridine
C6H5COCl
Pyridine
NH2
O
N
N
C6H5
N
C6H5
5
9
O
O
NH2NH2H2O
Methanol
NH2
CH3
COOH
NH2
Ac2O
N
O
N
N
CH3
4
10
6
O
O
N
NH2NH2H2O
Methanol
NH2
(C3H7CO)2O
COOH
Ac2O
N
O
N
C3H7
C3H7
N
C
O
C3H7
H
7
8
11
O
Ar
O
Ar
CHO
N
R
C
O
3a-i
N
H
Ethanol, glacial acetic acid
O
N
NH2
O
N
PhQZ 1-9, MtQZ 1-9, PrQZ 1-9
N
R
O
N
O
O
H
9, 10, 11
NH
N
3j
N
PhQZ, R = -C6H5
MtQZ, R= -CH3
PrQZ, R= -C3H7
Ethanol, glacial acetic acid
N
R
PhQZ 10, MtQZ 10, PrQZ 10
Scheme 1. Synthesis of 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one (PhQZ 1e10, MtQZ 1e10 and PrQZ 1e10).
3 showed 75% protection (3/4, 0.5 h), 50% protection (2/4, 0.25 and
5.2. Computational study
5.2.1. Distance mapping
1.0 h) and 25% protection (1/4, 2.0 h) at a dose of 100 mg/kg.
Compound MtQZ 7 showed 50% protection (2/4, 0.25 and 0.5 h) and
25% protection (1/4, 1.0 h) at a dose of 100 mg/kg. Compounds
MtQZ 5 and 8 showed 50% protection (2/4, 0.5 h) at a dose of
100 mg/kg. Compound MtQZ 6 showed 50% protection (2/4, 1.0 h)
at a dose of 100 mg/kg. Compound PrQZ 10 showed 75% protection
(3/4, 0.25 h), 50% protection (2/4, 0.5 and 1.0 h) and 25% protection
(1/4, 2.0 h) at a dose of 100 mg/kg. Compound PrQZ 5 showed 50%
protection (2/4, 0.25 and 1.0 h) and 25% protection (1/4, 0.5 h) at
a dose of 100 mg/kg. Compound PrQZ 8 showed 50% protection (2/
4, 0.25 and 0.5 h) and 25% protection (1/4,1.0 h) at a dose of 100 mg/
kg. Compounds PrQZ 3, 4 and 7 showed 50% protection (2/4, 0.5 h),
(2/4, 0.25 h) and (2/4, 0.25 h) respectively at a dose of 100 mg/kg.
Other compounds showed mild to moderate activity at single or
two or more time points. None of the compounds showed neuro-
toxicity in the highest administered dose.
Compound PhQZ 7 was most active compound of the series and
subjected to quantitative 6 Hz evaluation and biological response
evaluated at 50, 100, 200 and 350 mg/kg. It showed an ED50 and
a TD50 of >350 mg/kg at a time of peak effect (TPE) of 0.5 h in
quantitative 6 Hz and toxicity test (Table 3). Compound PhQZ 7 was
evaluated in the primary screen experiment of in-vitro hippo-
campal slice culture neuroprotection assay (NP) but no significant
protection was observed against either KA or NMDA induced
cytotoxicity (Table 4).
The present work involves the correlation of the structural
requirement of well known and structurally different anticonvul-
sant compounds with the titled compounds. The two-dimensional
(2D) modeling on anticonvulsants has identified that at least one
aryl unit, one or two electron donor atoms, and/or an NH group in
a special spatial arrangement is recommended for anticonvulsant
activity. In the present study, the 10 well known and structurally
different compounds with anticonvulsant activity-albutoin, carba-
mazepine, gabapentin, lamotrigine, mephobarbital, phenytoin,
progabide, ralitoline, remacemide and zonisamide (Fig. 3) with
different mechanisms of action, were selected so as to propose
a generalized pharmacophore model. The pharmacophore group’s
distance estimation was done by molecular mechanics calculation
with the force fields based on both CHARMm force fields and MM3
parametrization. In the present work, energy minimization was
performed on above mentioned ten well known anticonvulsants
and 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one
derivatives using Argus Lab 4.0. Distance between the various
structural components essential for activity was determined by
ACD/3D viewer. The crucial structural components that were
included in the four-point pharmacophore model (Fig. 4) were the
aryl ring center or the lipophilic group (A), an electron donor atom
(D), a hydrogen bond acceptor (HA), and a hydrogen bond donor

1010
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
Table 1
Physical and elemental analyses data of (i) 2-phenyl-3-{[substituted]amino}quinazolin-4(3H)-ones PhQZ 1-9, (ii) 2-methyl-3-{[substituted]amino}quinazolin-4(3H)-ones
MtQZ 1-9, (iii) 2-propyl-3-{[substituted]amino}quinazolin-4(3H)-ones PrQZ 1-9, and (iv) 3-[(2-oxo-1,2-dihydro-3H-indol-3-ylidene)amino]-2-(substituted)quinazolin-4(3H)-
one PhQZ 10, MtQZ 10, PrQZ 10.
O
O
N
C
H
O
Ar
N
C
H
O
Ar
N
N
N
C6H5
N
CH3
Z
QZ 1-9
O
O
NH
O
N
R
N
N
C
H
O
Ar
N
N
N
C3H7
Z
QZ 10, MtQZ 10, PrQZ 10
Compd.
Ar
R
Molecular Formula (M. W.)
Mp ꢀ(C)
Yielda (%)
Elemental analysis (%): Found (cal.)
C
H
N
PhQZ 1
PhQZ 2
PhQZ 3
PhQZ 4
PhQZ 5
PhQZ 6
PhQZ 7
PhQZ 8
PhQZ 9
MtQZ 1
MtQZ 2
MtQZ 3
MtQZ 4
MtQZ 5
MtQZ 6
MtQZ 7
MtQZ 8
MtQZ 9
PrQZ 1
PrQZ 2
PrQZ 3
PrQZ 4
PrQZ 5
PrQZ 6
PrQZ 7
PrQZ 8
PrQZ 9
PhQZ 10
MtQZ 10
PrQZ 10
Phenyl
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
C27H19N3O2 (417.46)
C27H18N4O4 (462.46)
C28H21N3O2 (431.49)
120-122
203-204
149-151
228-230
110-112
182-184
183-185
207-208
101-102
118-120
175-176
196-198
178-180
177-178
196-197
131-133
165-167
106-107
80-82
137-138
118-119
103-105
102-103
78-80
86-88
101-102
92-94
70
68
74
64
68
72
70
73
61
68
71
72
58
61
66
68
72
55
67
65
62
56
59
62
58
68
54
77
79
75
77.63 (77.68)
70.07 (70.12)
77.91 (77.94)
71.72 (71.76)
65.30 (65.33)
74.48 (74.47)
72.17 (72.18)
79.60 (79.64)
72.83 (72.88)
74.31 (74.35)
65.95 (66.00)
74.72 (74.78)
67.74 (67.78)
60.82 (60.84)
70.75 (70.77)
68.37 (68.40)
76.51 (76.52)
69.16 (69.17)
75.17 (75.18)
67.25 (67.28)
75.51 (75.54)
68.93 (68.98)
62.31 (62.35)
71.80 (71.81)
69.50 (69.52)
77.55 (77.58)
70.24 (70.25)
72.09 (72.12)
67.07 (67.10)
68.62 (68.66)
4.54 (4.59)
3.90 (3.92)
4.87 (4.91)
4.02 (4.01)
3.65 (3.66)
4.13 (4.17)
4.29 (4.33)
4.54 (4.53)
4.11 (4.15)
4.80 (4.82)
4.01 (4.03)
5.14 (5.18)
4.11 (4.14)
3.68 (3.71)
4.30 (4.32)
4.46 (4.49)
4.65 (4.62)
4.26 (4.29)
5.51 (5.52)
4.69 (4.71)
5.80 (5.83)
4.80 (4.82)
4.37 (4.36)
5.01 (5.02)
5.13 (5.13)
5.31 (5.35)
4.91 (4.95)
3.81 (3.85)
3.94 (3.97)
4.83 (4.85)
10.06 (10.07)
12.08 (12.12)
9.71 (9.74)
9.24 (9.30)
8.41 (8.47)
9.61 (9.65)
9.01 (9.02)
8.93 (8.99)
9.10 (9.11)
11.83 (11.82)
13.97 (13.99)
11.35 (11.37)
10.76 (10.78)
9.69 (9.68)
11.24 (11.25)
10.39 (10.40)
10.67 (10.71)
10.50(10.52)
10.97 (10.96)
13.04 (13.08)
10.55 (10.57)
10.07 (10.06)
9.07 (9.09)
4-NO2 phenyl
4-CH3 phenyl
4-Cl phenyl
4-Br phenyl
4- F phenyl
3-CH3, 4-Cl phenyl
Naphthalene-2-yl
1,3-benzodioxol-5-yl
Phenyl
4-NO2 phenyl
4-CH3 phenyl
4-Cl phenyl
4-Br phenyl
4- F phenyl
3-CH3, 4-Cl phenyl
Naphthalene-2-yl
1,3-benzodioxol-5-yl
Phenyl
4-NO2 phenyl
4-CH3 phenyl
4-Cl phenyl
4-Br phenyl
4- F phenyl
3-CH3, 4-Cl phenyl
Naphthalene-2-yl
1,3-benzodioxol-5-yl
-
C27H18ClN3O2 (451.90)
C27H18BrN3O2 (496.35)
C27H18FN3O2 (435.45)
C28H20ClN3O2 (465.93)
C31H21N3O2 (467.52)
C28H19N3O4 (461.47)
C22H17N3O2 (355.39)
C22H16N4O4 (400.39)
C23H19N3O2 (369.42)
C22H16ClN3O2 (389.83)
C22H16BrN3O2 (434.29)
C22H16FN3O2 (373.38)
C23H18ClN3O2 (403.86)
C25H18N3O2 (392.43)
C23H17N3O4 (399.40)
C24H21N3O2 (383.44)
C24H20N4O4 (428.44)
C25H23N3O2 (397.47)
C24H20ClN3O2 (417.89)
C24H20BrN3O2 (462.34)
C24H20FN3O2 (401.43)
C25H22ClN3O2 (431.91)
C28H23N3O2 (433.50)
10.44 (10.47)
9.70 (9.73)
9.68 (9.69)
C25H21N3O4 (427.45)
9.84 (9.83)
C6H5
CH3
n-C3H7
C22H14N4O2 (366.37)
C17H12N4O2 (304.30)
C19H16N4O2 (332.36)
234-236
116-118
173-174
15.28 (15.29)
18.38 (18.41)
16.83 (16.86)
-
-
a
Solvent used for recrystallization e ethanol : water (9:1).
(HD). An average distance range for every point was obtained and
compared to the 2-(substituted)-3-{[substituted]amino}quinazo-
lin-4(3H)-one derivatives. Now it may be interesting to examine
whether the 2-(substituted)-3-{[substituted]amino}quinazolin-4
(3H)-one derivatives reflect the conditions of the derived pharma-
cophore model. Our analyses of the distance relationship showed
that 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-one
derivatives did fulfill the essential demands of the pharmacophore
when compared to the average distance requirement (Table 5).
5.2.2. Prediction of ADME properties
A computational study for prediction of ADME properties of
titled compounds was performed. Topological polar surface area
(TPSA), i.e., surface belonging to polar atoms, is a descriptor that
was shown to correlate well with passive molecular transport
through membranes and, therefore, allows prediction of transport
properties of drugs in the intestines and bloodebrain barrier
crossing [12]. The percentage of absorption (%ABS) was calculated
using TPSA. From all these parameters, it can be observed that all

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1011
Table 2
Table 4
6 Hz anticonvulsant activity and neurotoxicity of 2-(substituted)-3-{[substituted]
Results of in-vitro hippocampal slice culture neuroprotection assay (Test 76).
amino}quinazolin-4(3H)-one PhQZ 1e10, MtQZ 1e10, PrQZ 1e10.
Compound
Excitotoxin
Insult
duration (h)
Primary screen
results
Compound
Time (h) to peak effect (N/F)a
Toxicitya
PhQZ 7
N-methyl-
(NMDA)
D
-aspartate
4
No neuroprotection
observed
0.25
0.5
1.0
2.0
4.0
PhQZ 6
PhQZ 7
PhQZ 9
MtQZ 1
MtQZ 2
MtQZ 3
MtQZ 5
MtQZ 6
MtQZ 7
MtQZ 8
MtQZ 9
MtQZ 10
PrQZ 1
PrQZ 2
PrQZ 3
PrQZ 4
PrQZ 5
PrQZ 6
PrQZ 7
PrQZ 8
PrQZ 10
2/4
3/4
1/4
0/4
1/4
2/4
0/4
0/4
2/4
0/4
0/4
0/4
0/4
1/4
0/4
2/4
2/4
1/4
2/4
2/4
3/4
2/4
4/4
0/4
1/4
1/4
3/4
2/4
1/4
2/4
2/4
1/4
1/4
1/4
0/4
2/4
0/4
1/4
0/4
1/4
2/4
2/4
1/4
1/4
0/4
0/4
1/4
2/4
1/4
2/4
1/4
0/4
0/4
0/4
1/4
1/4
0/4
0/4
2/4
0/4
1/4
1/4
2/4
0/4
1/4
0/4
0/4
1/4
1/4
1/4
1/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
1/4
0/4
1/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
0/4
Kainic acid (KA)
4
No neuroprotection
observed
value ꢀ6.8 kcal/mol and 1H-bond) and GABA (A) delta receptor
(Affinity value ꢀ7.1 kcal/mol and 1H-bond). It also showed affinity
with Na/H exchanger (Affinity value ꢀ5.5 kcal/mol and 0H-bond).
Compound MtQZ 3 has exhibited good binding properties with
glutamate receptor (Affinity value ꢀ7.8 kcal/mol and 4H-bonds)
and GABA (A) alpha-1 receptor (Affinity value ꢀ7.2 kcal/mol and
1H-bond). It also showed affinity with GABA (A) delta receptor
(Affinity value ꢀ7.2 kcal/mol and 0H-bond) and Na/H exchanger
(Affinity value ꢀ6.3 kcal/mol and 0H-bond). Compound PrQZ 10
has exhibited good binding properties with glutamate receptor
(Affinity value ꢀ6.9 kcal/mol and 2H-bond). It also showed affinity
with GABA (A) alpha-1 receptor (Affinity value ꢀ5.9 kcal/mol and
0H-bond), GABA (A) delta receptor (Affinity value ꢀ5.8 kcal/mol
and 0H-bond), and Na/H exchanger (Affinity value ꢀ4.9 kcal/mol
and 0H-bond). Compounds PhQZ 7, MtQZ 3 and PrQZ 10 did not
show affinity and binding with Na channel and T-type calcium
channel receptor. The docking study results of PhQZ 7, MtQZ 3 and
PrQZ 10 with these six receptors are shown in Table 8. The docking
images are shown in Fig. 5. Docking study results show that the
titled compounds exhibited good affinity and binding properties
with glutamate, GABA (A) alpha-1 and GABA (A) delta receptors.
Affinity was also observed with Na/H exchanger without H-bond.
a
N/F ¼ number of animals active or toxic over the number tested at a dose of
100 mg/kg.
titled compounds exhibited a great %ABS ranging from 73.7 to
89.5% (Table 6). Furthermore, compounds PhQZ 1e9; MtQZ 4, 5 and
8 and PrQZ 1e9 violated only one Lipinski’s parameter. None of the
other compounds violated Lipinski’s parameters, making them
potentially promising agents for epilepsy therapy.
5.2.3. log P determination
5.3. Structureeactivity relationship (SAR)
Titled compounds showed dependence of biological activity on
lipophilic character in a congeneric series. In particular, for drugs
acting on central nervous system to be potent, they have to cross
bloodebrain barrier (BBB), thus potency has been correlated with
optimum lipophilicity (log P) near 2. In this study, we attempted to
correlate the anticonvulsant activity of congeners with their
calculated log P value. The experimental log P values were deter-
mined using the octanolephosphate buffer method. The data is
presented in Table 7. As observed some of the experimental values
were in good agreement with the theoretical values. All the titled
compounds showed lipophilic character.
In the present study, various 2-(substituted)-3-{[substituted]
amino}quinazolin-4(3H)-ones were synthesized by using different
substituent, in order to investigate the pharmacophoric substit-
uent, responsible for better activity. In the synthetic design, the
various substituents were introduced at positions 6 and 20 of
quinazolin-4(3H)-one backbone structure. At position 6, we intro-
duced methyl, phenyl and n-propyl group, chosen on the basis of
literature and hypothesis as discussed in Introduction section. At
A- Hydrophobic Unit
D- Electron Donar Group
HA- Hydrogen Bond Acceptor Unit
5.2.4. Docking studies
In this study, we have used Autodoc 4.0 along with its LGA
algorithm for automated flexible ligand docking of compounds
PhQZ 7, MtQZ 3 and PrQZ 10 with six established epilepsy
molecular targets namely GABA (A) alpha-1 receptor, GABA (A)
delta receptor, glutamate receptor, Na/H exchanger, Na channel
receptor, T-type calcium channel receptor and evaluated their
affinity (kcal/mol) and hydrogen bonding. PhQZ 7 has exhibited
good binding properties with glutamate receptor (Affinity value
ꢀ6.9 kcal/mol and 2H-bonds), GABA (A) alpha-1 receptor (Affinity
HD- Hydrogen Bond Donar Unit
D
4.30
(3.01-5.38)
4.86
(2.43-6.79)
C
C
C
C
4.37
(2.52-6.72)
A
C
C
6.05
(3.78-9.51)
Table 3
Quantitative 6 Hz evaluation: ED50 value and biological response data.
5.86
HA
(2.72-9.23)
Compd.
Test
Time (h)
ED50(mg/kg)
ED50 biological response
Dose (mg/kg)
2.54
(1.78-3.87)
HD
50
100
200
350
PhQZ 7
6 Hz
TOX
0.5
0.5
>350
>350
1/8
e
3/8
e
1/8
e
2/8
0/8
Fig. 4. Four-point 3D pharmacophore model for anticonvulsants derived by using
MM3 and CHARMm parametrization (Argus Lab 4.0 and ACD/3D viewer).

1012
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
Table 5
Distance range between the essential structural elements A, D and HAeHD.a
Compounds
AeHA
AeHD
AeD
HAeHD
HDeD
HAeD
Albutoin
5.37
2.72
4.51
2.72
4.03
5.40
Carbamazepine
Gabapentin
Lamotrigine
Mephobarbital
Phenytoin
Progabide
Ralitoline
Remacemide
Zonisamide
Av distance
(range)
4.28
4.26
5.30
3.78
6.20
9.51
8.30
7.51
6.02
6.05
(3.78e9.51)
5.26
4.28
4.93
7.42
5.50
4.01
9.23
5.55
8.75
6.22
5.86
(2.72e9.23)
6.06
4.25
3.83
4.54
4.81
4.35
3.79
4.56
5.38
3.01
4.30
(3.01e5.38)
3.81
2.33
2.23
2.42
2.34
2.63
2.41
2.75
3.87
1.78
2.54
(1.78e3.87)
1.37
5.75
3.57
4.94
4.63
3.88
6.72
2.52
3.96
3.71
4.37
(2.52e6.72)
4.28
5.67
4.50
4.25
5.23
5.17
6.79
4.85
2.43
4.31
4.86
(2.43e6.79)
2.93
PhQZ 7
MtQZ 3
5.28
6.13
3.96
1.37
4.26
2.90
PrQZ 10
5.27
7.90
3.93
3.57
5.71
2.90
a
Distances calculated for 3D optimized structures using MM3 and CHARMm parameterization (Argus Lab 4.0 and ACD/3D viewer).
position 20, the substituent viz. phenyl, 4-nitro phenyl, 4-methyl
phenyl, 4-chloro phenyl, 4-bromo phenyl, 4-flouro phenyl, 4-
chloro-3-methyl phenyl, naphthalene-2-yl, 1,3-benzodioxol-5-yl
were introduced. Keeping in view, the pharmacophoric character of
isatin for anticonvulsant activity, the isatinyl moiety was also
attached to position N-11. These substituents were chosen based on
the literature review and computational study.
The most active compound PhQZ 7 has phenyl at position 6 and
4-chloro-3-methyl phenyl at position 20. It has been observed that
at position 6, the presence of phenyl was more significant than
methyl and n-propyl, and yields more active compounds. This
observation support the hypothesis that methyl group at second
position of quinazolin-4(3H)-one is not always necessary for the
CNS activity and other groups when placed at this position can also
lead to potent CNS active agents. In addition, at position 20, the
presence of halogenated phenyl was more significant than phenyl,
non-halogenated phenyl, naphthalene-2-yl, 1,3-benzodioxol-5-yl
substituents. This observation clearly indicates the effect of elec-
tronic factors on activity. The 4-chloro-3-methyl phenyl substituent
is most active one and also increases the duration of action. This
increase in duration of action might be because of blockade of
metabolism at position 25 by methyl group. The presence of isatinyl
at position N-11 and n-propyl at position 6, also yields the active
compound PrQZ 10 with good duration of action. The presence of
naphthalene-2-yl and 1,3-benzodioxol-5-yl substituent at position
20 does not yield any active compound.
Table 6
Pharmacokinetic parameters important for good oral bioavailability of compounds PhQZ 1e10, MtQZ 1e10, PrQZ 1e10.a
Compound
%ABS
TPSA (A2)
n-ROTB
MW
MV
n-OHNH
donors
n-ON
acceptors
Lipinski’s
violations
Rule
e
e
e
5
6
5
5
5
5
5
5
5
2
4
5
4
4
4
4
4
4
4
1
6
7
6
6
6
6
6
6
6
3
<500
e
<5
0
0
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
0
0
1
<10
5
8
5
5
5
5
5
5
7
6
5
8
5
5
5
5
5
5
7
6
5
8
5
5
5
5
5
5
7
6
ꢁ1
1
1
1
1
1
1
1
1
1
0
0
0
0
1
1
0
0
1
0
0
1
1
1
1
1
1
1
1
1
0
PhQZ 1
PhQZ 2
PhQZ 3
PhQZ 4
PhQZ 5
PhQZ 6
PhQZ 7
PhQZ 8
PhQZ 9
PhQZ 10
MtQZ 1
MtQZ 2
MtQZ 3
MtQZ 4
MtQZ 5
MtQZ 6
MtQZ 7
MtQZ 8
MtQZ 9
MtQZ 10
PrQZ 1
PrQZ 2
PrQZ 3
PrQZ 4
PrQZ 5
PrQZ 6
PrQZ 7
PrQZ 8
PrQZ 9
PrQZ 10
89.5
73.7
89.5
89.5
89.5
89.5
89.5
89.5
83.1
81.4
89.5
73.7
89.5
89.5
89.5
89.5
89.5
89.5
83.1
81.4
89.5
73.7
89.5
89.5
89.5
89.5
89.5
89.5
83.1
81.4
56.49
102.3
56.50
56.50
56.50
56.50
56.50
56.50
74.97
80.12
56.50
102.3
56.50
56.50
56.50
56.50
56.50
56.50
74.96
80.12
56.49
102.3
56.50
56.50
56.50
56.50
56.50
56.50
74.96
80.12
417.47
462.47
431.50
451.91
496.37
435.46
465.94
467.53
461.48
366.38
355.39
400.39
369.42
389.84
434.29
373.39
405.88
405.46
399.41
304.31
383.45
428.45
397.48
417.90
462.35
401.44
431.92
433.51
427.46
332.36
374.7
398.04
391.26
388.24
392.59
379.63
404.80
418.70
398.63
314.85
319.85
343.19
336.41
333.39
337.74
324.78
356.16
363.85
343.78
260.00
353.46
376.79
370.02
366.99
371.34
358.39
383.55
397.45
377.39
293.61
a
%ABS, percentage of absorption; TPSA, topological polar surface area; n-ROTB, number of rotatable bonds; MW, molecular weight; MV, molecular volume; n-OHNH,
number of hydrogen bond donors; n-ON, number of hydrogen bond acceptors.

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1013
Table 7
anticonvulsant activity was evaluated. The titled compounds dis-
played the significant 6 Hz psychomotor seizure test activity. The
neurotoxicity evaluation was done by rotarod test. A computational
study was also carried out, including calculation of pharmacophore
pattern, prediction of pharmacokinetic properties and docking
studies. The compound PhQZ 7 displayed 100% protection and
emerged as a lead in this series. Further, compounds MtQZ 3 and
PrQZ 10 came out as a potential candidate for further investigation.
Compounds PhQZ 6, MtQZ 5e8, PrQZ 3e5 and PrQZ 7 and 8
exhibited moderate protection. None of the compound showed
neurotoxicity in rotarod test. Furthermore, PhQZ 10; MtQZ 1e3, 6,
7, 9, 10; and PrQZ 10 also did not show Lipinski’s violation, making
them potentially promising agent for epilepsy therapy. Docking
study results show that the compounds exhibited good binding
properties with glutamate, GABA (A) delta and GABA (A) alpha-1
receptor. The docking study data strongly support the assumption
that these receptors may be involved in observed anticonvulsant
activity of 2-(substituted)-3-{[substituted]amino}quinazolin-4(3H)-
one derivatives. However, further studies need to be carried out
to ascertain the precise mechanism of anticonvulsant activity of
these molecules.
log P value for titled compounds PhQZ 1e10, MtQZ 1e10, PrQZ 1e10.
Compounds
Experimental log P
Theoretical log P
(milog Pa)
Rule
e
ꢁ5
PhQZ 1
PhQZ 2
PhQZ 3
PhQZ 4
PhQZ 5
PhQZ 6
PhQZ 7
PhQZ 8
PhQZ 9
PhQZ 10
MtQZ 1
MtQZ 2
MtQZ 3
MtQZ 4
MtQZ 5
MtQZ 6
MtQZ 7
MtQZ 8
MtQZ 9
MtQZ 10
PrQZ 1
PrQZ 2
PrQZ 3
PrQZ 4
PrQZ 5
PrQZ 6
PrQZ 7
PrQZ 8
PrQZ 9
PrQZ 10
6.134
6.123
6.291
6.381
6.394
6.212
6.517
6.526
6.012
4.032
4.012
4.013
4.321
4.598
4.643
4.212
4.412
5.221
4.011
2.232
5.202
5.178
5.451
5.782
5.878
5.221
6.019
6.123
4.823
3.234
6.357
6.316
6.805
7.035
7.166
6.521
7.411
7.54
6.247
4.371
4.329
4.288
4.777
5.007
5.138
4.493
4.812
5.512
4.219
2.343
5.404
5.363
5.853
6.082
6.213
5.568
6.459
6.587
5.294
3.418
7. Experimental protocol
7.1. Chemistry
All the chemicals and solvents, purchased from Merck (India),
Spectrochem (India), Himedia (India) and S. d. Fine were used
without further purification. The progress of reaction was moni-
tored by thin layer chromatography, performed on a silica gel 60
F254 coated aluminum sheet. The melting points were determined
by using ThomaseHoover melting point apparatus and are uncor-
rected. The FT-IR spectra were recorded on PerkineElmer Spectrum
BX-II Spectrophotometer. The 1H NMR spectra were recorded on
Bruker 300 MHz High Resolution NMR spectrometer using TMS as
a
milog P, logarithm of compound partition coefficient between n-octanol and
water calculated as per molinspiration online property toolkit.
6. Conclusion
A series of 2-(substituted)-3-{[substituted]amino}quinazolin-4
an internal standard. Chemical shifts were reported in ppm (
d) and
(3H)-one derivatives were designed, synthesized, and their
signals were described as singlet (s), doublet (d), triplet (t) and
Table 8
Docking results of compounds PhQZ 7, MtQZ 3 and PrQZ 10.a
Ligand
Receptor
Affinity (kcal/mol)
H-bonds
H-binding ligand
H-binding receptor
Element
Atom No.
Type
Residue
Element
Atom No.
Type
PhQZ 7
GABA (A) alpha-1
GABA (A) delta
Glutamate
ꢀ6.8
ꢀ7.1
ꢀ6.9
01
01
02
N
N
O
N
e
e
e
O
e
N
N
O
N
e
e
e
e
e
O
N
e
e
e
19
19
08
16
e
Donor
Donor
Acceptor
Acceptor
Phe323
Lue325
Try 456
Ser 554
e
O
O
O
N
e
e
e
N
e
O
O
O
O
e
e
e
e
e
O
O
e
e
e
407
458
146
1067
e
Acceptor
Acceptor
Both
Donor
e
Na/H exchanger
Na channel
T-type calcium
GABA (A) alpha-1
GABA (A) delta
Glutamate
ꢀ5.5
ꢀ0.0
ꢀ0.0
ꢀ7.2
ꢀ7.2
ꢀ7.8
00
00
00
01
00
04
e
e
e
e
e
e
e
e
e
e
e
MtQZ 3
00
e
Acceptor
e
Lys339
e
567
e
Donor
e
08
08
00
11
e
Donor
Donor
Acceptor
Donor
Pro531
Thr533
Try459
Try459
e
846
865
146
146
e
Acceptor
Both
Both
Both
e
Na/H exchanger
Na channel
T-type calcium
GABA (A) alpha-1
GABA (A) delta
Glutamate
ꢀ6.3
ꢀ0.0
ꢀ0.0
ꢀ5.9
ꢀ5.8
ꢀ6.9
00
00
00
00
00
02
e
e
e
e
e
e
e
e
e
e
e
PrQZ 10
e
e
e
e
e
e
e
e
e
e
25
16
e
Acceptor
Acceptor
e
e
e
Try459
Thr533
e
146
865
e
Both
Both
e
Na/H exchanger
Na channel
T-type calcium
ꢀ4.9
ꢀ0.0
ꢀ0.0
00
00
00
e
e
e
e
e
e
e
e
a
Affinity and H-bonds calculations were determined by docking studies using Autodoc 4.0 software.

1014
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
Fig. 5. Docking Images. (a) PhQZ 7 with GABA (A) alpha-1 receptor, (b) PhQZ 7 with GABA (A) delta receptor, (c) PhQZ 7 with glutamate receptor, (d) PhQZ 7 with Na/H exchanger;
(e) MtQZ 3 with GABA (A) alpha-1 receptor, (f) MtQZ 3 with GABA (A) delta receptor, (g) MtQZ 3 with glutamate receptor, (h) MtQZ 3 with Na/H exchanger; (i) PrQZ 10 with GABA
(A) alpha-1 receptor, (j) PrQZ 10 with GABA (A) delta receptor, (k) PrQZ 10 with glutamate receptor, (l) PrQZ 10 with Na/H exchanger.

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1015
multiplet (m). All exchangeable protons were confirmed by addi-
tion of D2O. The mass spectra were recorded on a Waters Micro-
mass ZQ 2000 mass spectrometer. Elemental analysis (C, H, N) was
undertaken with PerkineElmer Model 240C analyzer.
3-Amino-2-methyl-quꢀin1azolin-4(3H)-one (10) Yield: 82%; mp:
139e141 ꢂC; IR (KBr, cm
) y: 3310-3262 (NH2), 1678 (C]O), 1617
(C]N), 1595 (C]C); 1H NMR (CDCl3, 300 MHz)
d: 6.71e8.18 (m, 4H,
Qu-H), 4.63 (s, 2H, NH2), 2.38 (s, 3H, CH3); MS (m/z, %): 176.08 (M þ
þ 1, 38.13).
3-Amino-2-propyl-quinazolin-4(3H)-one (11) Yield: 77%; mp:
7.1.1. Synthesis of 4-substituted benzaldehyde (3aei)
84e86 ꢂC; IR (KBr, cmꢀ1
)
y
: 3304e3260 (NH2), 1672 (C]O), 1617
A mixture of substituted phenol 1aei (37.4 mmol), 4-fluoro
benzaldehyde 2 (37.4 mmol) and potassium carbonate (38.8 mmol)
in N,N-dimethylformamide (30 ml) was refluxed for 16e18 h under
nitrogen. After cooling, the product was extracted from the reaction
mixture and purified by chromatography.
(C]N),1602 (C]C); 1H NMR (CDCl3, 300 MHz)
d
: 6.78e8.27 (m, 4H,
Qu-H), 4.68 (s, 2H, NH2), 2.73 (t, 2H, CH2Qu ), 1.82 (sextet, 2H,
CH2Me ), 1.07 (t, 3H, CH3); MS (m/z, %): 204.08 (M þ þ 1, 43.18).
4-(4-Fluoro-phenoxy)-benzaldehyde (3f): Yield: 70%; m.p.: 41 ꢂC;
7.1.7. Synthesis of 2-(substituted)-3-{[substituted]amino}
IR (KBr, cmꢀ1 : 1665 (C]O), 1240 (O); 1H NMR (CDCl3, 300 MHz)
) y
quinazolin-4(3H)-one (PhQZ 1ePhQZ , MtQZ 1e10, PrQZ 1e10)
Equimolar quantities (0.01 mol) of 4-substituted benzaldehydes
3aei/isatin 3j and 3-amino-2-(substituted)-quinazolin-4(3H)-one
(9, 10, 11) were dissolved in warm ethanol containing 0.5 ml of
glacial acetic acid. The reaction mixture was refluxed for 4e6 h and
set aside. The resultant solid was washed with ethanol and
recrystallized from 90% ethanol. The physical data and elemental
analysis data of the titled compounds PhQZ 1e10, MtQZ 1e10,
PrQZ 1e10 are presented in Table 1. The spectral data of titled
compounds PhQZ 1e10, MtQZ 1e10, PrQZ 1e10 are given below.
d
: 9.87 (s, 1H, CHO), 6.83e7.87 (a set of signals, 8H, Ar-H); MS (m/z,
%): 216.17 (Mþ þ 1, 67.2)
7.1.2. Synthesis of 2-phenyl-4H-3,1-benzoxazin-4-one (5)
To a stirred solution of anthranilic acid 4 (0.05 mol) in pyridine
(50 mL), benzoyl chloride (0.05 mol) was added drop wise, main-
taining the temperature near 0e6 ꢂC for 1 h. The reaction mixture
was stirred for another 3 h at room temperature until a solid
product was separated. The reaction mixture was neutralized with
saturated sodium bicarbonate solution and the pale yellow solid
which separated was filtered, washed with water and recꢀr1ystallised
7.1.7.1. 2-Phenyl-3-{[(E)-(3-phenoxyphenyl)methylidene]amino}qui-
nazolin-4(3H)-one (PhQZ 1). IR (KBr, cmꢀ1
) y: 3046 (Qu-H, Ar-H),
from ethanol. Yield: 86%; mp. 112e114 ꢂC; IR (KBr, cm
) y: 3033
1687 (C]O), 1599 (N]CH), 1446e1524 (C]C, benzene and Qu-
(Qu-H, Ar-H), 1680 (C]O), 1602 (C]N), 1178 (CeO); 1H NMR
ring), 1243 (O), 754, 696 (mono substituted benzene); 1H NMR
(CDCl3, 300 MHz) d: 6.88e8.18 (m, 9H, Qu-H and Ar-H), MS (m/z, %):
(CDCl3, 300 MHz) d: 8.43 (s, 1H, CH]N), 6.83e8.17 (a set of signals,
224.06 (Mþ þ 1, 54.27).
18H, Qu-H and Ar-H); MS (m/z, %): 418.16 (Mþ þ 1, 57.31).
7.1.3. Synthesis of 2-methyl-4H-3,1-benzoxazin-4-one (6)
7.1.7.2. 3-({(E)-[3-(4-Nitrophenoxy)phenyl]methylidene}amino)-2-
phenylquinazolin-4(3H)-one (PhQZ 2). IR (KBr, cmꢀ1
) y: 3058 (Qu-
A mixture of anthranilic acid 4 (0.01 mol) and acetic anhydride
(0.1 mol) was refluxed on gentle flame for 2 h. The excess acetic
anhydride was distilled off under reduced pressure and the residue
was dissolved in petroleum ether and kepꢀt 1aside for 1 h, to yield 6.
H, Ar-H), 1682 (C]O), 1594 (N]CH), 1525 (N]O), 1449e1528 (C]
C, benzene and Qu-ring), 1247 (O), 759, 696 (mono substituted
benzene); 1H NMR (CDCl3, 300 MHz)
d in ppm: 8.47 (s, 1H, CH]N),
Yield: 78%; mp: 180e182 ꢂC; IR (KBr, cm
)
y
: 1665 (C]O), 1605
6.78e8.11 (a set of signals,17H, Qu-H and Ar-H); MS (m/z, %): 463.16
(C]N), 1169 (CeO); 1H NMR (CDCl3, 300 MHz)
d: 6.91e8.09 (m, 4H,
(Mþ þ 1, 37.11).
Qu-H), 2.39 (s, 3H, CH3); MS (m/z, %): 162.11 (Mþ þ 1, 34.12).
7.1.7.3. 3-({(E)-[3-(4-Methylphenoxy)phenyl]methylidene}amino)-2-
phenylquinazolin-4(3H)-one (PhQZ 3). IR (KBr, cmꢀ1
) y: 3030 (Qu-
7.1.4. Synthesis of 2-propyl-4H-3,1-benzoxazin-4-one (8)
Anthranilic acid 4 (0.1 mol) was refluxed with butyric anhydride
for 3 h. Excess of anhydride was distilled off by vacuum distillation
to obtain the precipitate of N-propyl anthranilic acid 7, which was
further refluxed with acetic anhydride for 2 h to yield 8. Yield: 72%;
H, Ar-H), 1662 (C]O), 1599 (N]CH), 1450e1540 (C]C, benzene
and Qu-ring), 1243 (O), 751, 702 (mono substituted benzene); 1H
NMR (CDCl3, 300 MHz) d in ppm: 8.31 (s, 1H, CH]N), 6.92e8.04 (a
set of signals, 17H, Qu-H and Ar-H), 2.35 (s, 3H, CH3); MS (m/z, %):
mp: 58e60 ꢂC; IR (KBr, cmꢀ1
(CeO); 1H NMR (CDCl3, 300 MHz)
)
y
: 1732 (C]O), 1608 (C]N), 1160
: 6.84e8.28 (m, 4H, Qu-H), 2.71
432.12 (Mþ þ 1, 21.22).
d
(t, 2H, CH2Qu ), 1.83 (sextet, 2H, CH2Me ), 1.03 (t, 3H, CH3); MS (m/z,
7.1.7.4. 3-({(E)-[3-(4-Chlorophenoxy)phenyl]methylidene}amino)-2-
%): 190.05 (Mþ þ 1).
phenylquinazolin-4(3H)-one (PhQZ 4). IR (KBr, cmꢀ1
) y: 3041
(Qu-H, Ar-H), 1690 (C]O), 1596 (N]CH), 1449e1528 (C]C,
7.1.5. Synthesis of 3-amino-2-phenylquinazolin-4(3H)-one (9)
To a stirred solution of 5 (0.05 mol) in pyridine (20 mL), 80%
hydrazine hydrate (0.15 mol) was added. The reaction mixture was
stirred and refluxed for 30 min. After cooling, the crude product
was obtained by filtration and recrystallized from 80% ethanolꢀt1o
benzene and Qu-ring), 1246 (O), 756, 697 (mono substituted
benzene); 1H NMR (CDCl3, 300 MHz)
d in ppm: 8.44 (s, 1H, CH]N),
6.82e8.18 (a set of signals,17H, Qu-H and Ar-H); MS (m/z, %): 452.14
(Mþ þ 1 for 35Cl, 61.20), 454.10 (Mþ þ 1 for 37Cl, 19.3).
afford 9 as a white solid. yield: 86%; m.p. 178e180 ꢂC; IR (KBr, cm
)
7.1.7.5. 3-({(E)-[3-(4-Bromophenoxy)phenyl]methylidene}amino)-2-
: 3423 (NH2), 3033 (CeH, Ar-H),1687 (C]O),1591 (C]N); 1H NMR
phenylquinazolin-4(3H)-one (PhQZ 5). IR (KBr, cmꢀ1
) y: 3039 (Qu-
y
(CDCl3, 300 MHz) d: 6.81e8.20 (m, 9H, Qu-H and Ar-H), 4.67 (s, 2H,
H, Ar-H), 1681 (C]O), 1594 (N]CH), 1451e1528 (C]C, benzene
Qu-NH2); MS (m/z, %): 238.12 (Mþ þ 1, 38.16).
and Qu-ring), 1246 (O), 757, 693 (mono substituted benzene); 1H
NMR (CDCl3, 300 MHz) d in ppm: 8.49 (s, 1H, CH]N), 6.82e8.27 (a
set of signals, 17H, Qu-H and Ar-H); MS (m/z, %): 498.08 (Mþ þ 1 for
7.1.6. Synthesis of 3-amino-2-(methyl/propyl) quinazolin-4(3H)-
one (10, 11)
A mixture of 2-methyl/propyl-4H-3,1-benzoxazin-4-one (6/8)
(0.01 mol) and 80% hydrazine hydrate (0.03 mol) in methanol was
refluxed for 2e3 h and cooled. The separated solid was recrystal-
lized from ethanol.
81Br, 43.12), 496.03 (Mþ þ 1 for 79Br, 40.43).
7.1.7.6. 3-({(E)-[3-(4-Fluorophenoxy)phenyl]methylidene}amino)-2-
phenylquinazolin-4(3H)-one (PhQZ 6). IR (KBr, cmꢀ1
)
y
: 3058 (Qu-
H, Ar-H), 1686 (C]O), 1597 (N]CH), 1449e1528 (C]C, benzene

1016
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
and Qu-ring), 1241 (O), 759, 696 (mono substituted benzene); 1H
Ar-H), 2.63 (s, 3H, Qu-CH3); MS (m/z, %): 390.11 (Mþ þ 1 for 35Cl,
66.22), 392.07 (Mþ þ 1 for 37Cl, 21.8).
NMR (CDCl3, 300 MHz)
d in ppm: 8.44 (s, 1H, CH]N), 6.82e8.19 (a
set of signals, 17H, Qu-H and Ar-H); MS (m/z, %): 436.15 (Mþ þ 1,
61.23).
7.1.7.15. 3-({(E)-[3-(4-Bromophenoxy)phenyl]methylidene}amino)-
2-methylquinazolin-4(3H)-one (MtQZ 5). IR (KBr, cmꢀ1
)
y
: 3049
(Qu-H, Ar-H), 1676 (C]O), 1594 (N]CH), 1465e1562 (C]C,
benzene and Qu-ring), 1245 (O); 1H NMR (CDCl3, 300 MHz)
in
7.1.7.7. 3-({(E)-[3-(4-Chloro-3-methylphenoxy)phenyl]methylidene}
amino)-2-phenylquinazolin-4(3H)-one (PhQZ 7). IR (KBr, cmꢀ1
)
y:
d
3067 (Qu-H, Ar-H), 1685 (C]O), 1596 (N]CH), 1447e1526 (C]C,
ppm: 8.90 (s, 1H, CH]N), 6.87e8.27 (a set of signals, 12H, Qu-H and
Ar-H), 2.64 (s, 3H, Qu-CH3); MS (m/z, %): 436.02 (Mþ þ 1 for 81Br,
32.07), 434.05 (Mþ þ 1 for 79Br, 32.14).
benzene and Qu-ring), 1240 (O), 755, 699 (mono substituted
benzene); 1H NMR (CDCl3, 300 MHz)
d in ppm: 8.45 (s, 1H, CH]N),
6.82e8.39 (a set of signals,16H, Qu-H and Ar-H), 2.36 (s, 3H, CH3); MS
(m/z, %):466.11(Mþ þ 1for35Cl, 67.70), 468.08(Mþ þ 1for37Cl, 21.56).
7.1.7.16. 3-({(E)-[3-(4-Fluorophenoxy)phenyl]methylidene}amino)-
2-methylquinazolin-4(3H)-one (MtQZ 6). IR (KBr, cmꢀ1
)
y
: 3055
(Qu-H, Ar-H), 1671 (C]O), 1597 (N]CH), 1462e1568 (C]C,
benzene and Qu-ring), 1240 (O); 1H NMR (CDCl3, 300 MHz)
in
7.1.7.8. 3-({(E)-[3-(Naphthalen-2-yloxy)phenyl]methylidene}
amino)-2-phenylquinazolin-4(3H)-one (PhQZ 8). IR (KBr, cmꢀ1
)
y
:
d
3061 (Qu-H, Ar-H), 1682 (C]O), 1591 (N]CH), 1440e1519 (C]C,
benzene and Qu-ring), 1244 (O), 836, 821 ( -naphthyl), 754, 696
(mono substituted benzene); 1H NMR (CDCl3, 300 MHz)
in ppm:
ppm: 8.94 (s, 1H, CH]N), 6.84e8.29 (a set of signals,12H, Qu-H and
b
Ar-H), 2.67 (s, 3H, Qu-CH3); MS (m/z, %): 374.08 (Mþ þ 1, 41.12).
d
8.43 (s, 1H, CH]N), 6.83e8.31 (a set of signals, 20H, Qu-H, naph-
thyl-H and Ar-H); MS (m/z, %): 468.16 (Mþ þ 1, 32.89).
7.1.7.17. 3-({(E)-[3-(4-Chloro-3-methylphenoxy)phenyl]methylidene}
amino)-2-methylquinazolin-4(3H)-one (MtQZ 7). IR (KBr, cmꢀ1
) y:
3064 (Qu-H, Ar-H), 1675 (C]O), 1593 (N]CH), 1467e1568 (C]C,
7.1.7.9. 3-({(E)-[3-(1,3-Benzodioxol-5-yloxy)phenyl]methylidene}
benzene and Qu-ring), 1243 (O); 1H NMR (CDCl3, 300 MHz)
d
in
amino)-2-phenylquinazolin-4(3H)-one (PhQZ 9). IR (KBr, cmꢀ1
)
y
:
ppm: 8.92 (s, 1H, CH]N), 6.82e8.31 (a set of signals, 11H, Qu-H and
Ar-H), 2.65 (s, 3H, Qu-CH3), 2.37 (s, 3H, CH3); MS (m/z, %): 404.09
(Mþ þ 1 for 35Cl, 47.67), 406.10 (Mþ þ 1 for 37Cl, 15.7).
3049 (Qu-H, Ar-H), 1685 (C]O), 1597 (N]CH), 1449e1521 (C]C,
benzene and Qu-ring), 1247 (O), 755, 699 (mono substituted
benzene); 1H NMR (CDCl3, 300 MHz)
d in ppm: 8.41 (s, 1H, CH]N),
6.71e8.27 (a set of signals, 16H, Qu-H, sesamol-H and Ar-H), 5.98 (s,
7.1.7.18. 2-Methyl-3-({(E)-[3-(naphthalen-2-yloxy)phenyl]methyl-
2H, CH2); MS (m/z, %): 462.16 (Mþ þ 1, 23.10).
idene}amino)quinazolin-4(3H)-one (MtQZ 8). IR (KBr, cmꢀ1
)
y
: 3061
(Qu-H, Ar-H), 1672 (C]O), 1591 (N]CH), 1469e1570 (C]C,
benzene and Qu-ring), 1244 (O), 834, 823 (
-naphthyl); 1H NMR
(CDCl3, 300 MHz) in ppm: 8.95 (s, 1H, CH]N), 6.83e8.34 (a set of
7.1.7.10. 3-[(2-Oxo-1,2-dihydro-3H-indol-3-ylidene)amino]-2-phe-
b
nylquinazolin-4(3H)-one (PhQZ10). IR (KBr, cmꢀ1
)
y
: 3261 (NH),
d
3068 (Qu-H, Ar-H), 1687 (C]O), 1589 (N]C), 1446e1528 (C]C,
signals, 15H, Qu-H, naphthyl-H and Ar-H), 2.67 (s, 3H, Qu-CH3); MS
benzene and Qu-ring), 757, 698 (mono substituted benzene); 1H
(m/z, %): 393.11 (Mþ þ 1, 41.22).
NMR (CDCl3, 300 MHz)
d in ppm: 12.81 (s, 1H, NH of Isatin, D2O
exchangeable), 6.92e8.19 (a set of signals, 13H, Qu-H, Isatin-H and
7.1.7.19. 3-({(E)-[3-(1,3-Benzodioxol-5-yloxy)phenyl]methylidene}
Ar-H); MS (m/z, %): 367.12 (Mþ þ 1, 64.65).
amino)-2-methylquinazolin-4(3H)-one (MtQZ 9). IR (KBr, cmꢀ1
) y:
3057 (Qu-H, Ar-H), 1671 (C]O), 1597 (N]CH), 1471e1568 (C]C,
7.1.7.11. 2-Methyl-3-{[(E)-(3-phenoxyphenyl)methylidene]amino}
benzene and Qu-ring), 1243 (O); 1H NMR (CDCl3, 300 MHz)
d
in
quinazolin-4(3H)-one (MtQZ 1). IR (KBr, cmꢀ1
)
y
: 3049 (Qu-H, Ar-
ppm: 8.91 (s, 1H, CH]N), 6.79e8.21 (a set of signals, 11H, Qu-H,
sesamol-H and Ar-H), 5.93 (s, 2H, CH2), 2.65 (s, 3H, Qu-CH3); MS (m/
z, %): 400.13 (Mþ þ 1, 33.21).
H), 1667 (C]O), 1594 (N]CH), 1467e1558 (C]C, benzene and Qu-
ring), 1240 (O), 754, 696 (mono substituted benzene); 1H NMR
(CDCl3, 300 MHz) d: 8.93 (s, 1H, CH]N), 6.93e8.28 (a set of signals,
13H, Qu-H and Ar-H), 2.63 (s, 3H, Qu-CH3); MS (m/z, %): 356.09
7.1.7.20. 2-Methyl-3-[(2-oxo-1,2-dihydro-3H-indol-3-ylidene)
(Mþ þ 1, 47.11).
amino]quinazolin-4(3H)-one (MtQZ 10). IR (KBr, cmꢀ1
)
y
: 3263
(NH), 3066 (Qu-H, Isatin-H), 1684 (C]O), 1587 (N]C), 1465e1568
(C]C, benzene and Qu-ring); 1H NMR (CDCl3, 300 MHz)
in ppm:
7.1.7.12. 2-Methyl-3-({(E)-[3-(4-nitrophenoxy)phenyl]methylidene}
d
amino)quinazolin-4(3H)-one (MtQZ 2). IR (KBr, cmꢀ1
)
y
: 3058 (Qu-
H, Ar-H), 1672 (C]O), 1594 (N]CH), 1521 (N]O), 1466e1561 (C]
C, benzene and Qu-ring), 1244 (O); 1H NMR (CDCl3, 300 MHz)
in
12.83 (s, 1H, NH of isatin, D2O exchangeable), 6.95e8.22 (a set of
signals, 8H, Qu-H and isatin-H), 2.63 (s, 3H, Qu-CH3); MS (m/z, %):
305.06 (Mþ þ 1, 61.40).
d
ppm: 8.95 (s, 1H, CH]N), 6.88e8.25 (a set of signals, 12H, Qu-H and
Ar-H), 2.62 (s, 3H, Qu-CH3); MS (m/z, %): 401.11 (Mþ þ 1, 57.04).
7.1.7.21. 3-{[(E)-(3-Phenoxyphenyl)methylidene]amino}-2-pro-
pylquinazolin-4(3H)-one (PrQZ 1). IR (KBr, cmꢀ1
) y: 3049 (Qu-H,
7.1.7.13. 2-Methyl-3-({(E)-[3-(4-methylphenoxy)phenyl]methyl-
Ar-H), 1685 (C]O), 1602 (N]CH), 1447e1521 (C]C, benzene and
idene}amino)quinazolin-4(3H)-one (MtQZ 3). IR (KBr, cmꢀ1
)
y
: 3067
(Qu-H, Ar-H), 1670 (C]O), 1591 (N]CH), 1471e1566 (C]C,
benzene and Qu-ring), 1243 (O); 1H NMR (CDCl3, 300 MHz)
in
Qu-ring), 1241 (O), 751, 697 (mono substituted benzene); 1H NMR
(CDCl3, 300 MHz) d: 8.41 (s, 1H, CH]N), 6.80e8.27 (a set of signals,
d
13H, Qu-H and Ar-H), 2.46 (t, 2H, CH2Qu), 1.80 (sextet, 2H, CH2Me),
ppm: 8.91 (s, 1H, CH]N), 6.97e8.30 (a set of signals, 12H, Qu-H and
Ar-H), 2.64 (s, 3H, Qu-CH3), 2.37 (s, 3H, Ar-CH3); MS (m/z, %): 370.10
(Mþ þ 1, 33.20).
1.01 (t, 3H, CH3); MS (m/z, %): 384.10 (Mþ þ 1, 27.13).
7.1.7.22. 3-({(E)-[3-(4-Nitrophenoxy)phenyl]methylidene}amino)-2-
propylquinazolin-4(3H)-one (PrQZ 2). IR (KBr, cmꢀ1
)
y
: 3062 (Qu-H,
Ar-H), 1677 (C]O), 1604 (N]CH), 1521 (N]O), 1451e1525 (C]C,
benzene and Qu-ring), 1244 (O); 1H NMR (CDCl3, 300 MHz)
in
7.1.7.14. 3-({(E)-[3-(4-Chlorophenoxy)phenyl]methylidene}amino)-
2-methylquinazolin-4(3H)-one (MtQZ 4). IR (KBr, cmꢀ1
)
y
: 3061
(Qu-H, Ar-H), 1673 (C]O), 1596 (N]CH), 1469e1568 (C]C,
benzene and Qu-ring), 1241 (O); 1H NMR (CDCl3, 300 MHz)
in
ppm: 8.94 (s, 1H, CH]N), 6.92e8.28 (a set of signals, 12H, Qu-H and
d
ppm: 8.43 (s, 1H, CH]N), 6.73e8.23 (a set of signals, 12H, Qu-H and
Ar-H), 2.48 (t, 2H, CH2Qu), 1.85 (sextet, 2H, CH2Me), 1.00 (t, 3H,
CH3); MS (m/z, %): 429.08 (Mþ þ 1, 34.10).
d

P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
1017
7.1.7.23. 3-{(E)-[3-(4-Methylphenoxy)phenyl]methylidene}amino-2-
propylquinazolin-4(3H)-one (PrQZ 3). IR (KBr, cmꢀ1
: 3042 (Qu-H,
Ar-H), 1668 (C]O), 1607 (N]CH), 1445e1542 (C]C, benzene and
Qu-ring), 1241 (O); 1H NMR (CDCl3, 300 MHz)
in ppm: 8.35 (s, 1H,
CH]N), 6.81e8.24 (a set of signals, 12H, Qu-H and Ar-H), 2.47 (t,
2H, CH2Qu), 2.37 (s, 3H, Ar-CH3), 1.86 (sextet, 2H, CH2Me), 1.06 (t,
3H, CH2CH3); MS (m/z, %): 398.11 (Mþ þ 1, 38.81).
7.2. Pharmacology
)
y
The evaluation of anticonvulsant activity and neurotoxicity was
carried out by the Epilepsy Branch, National Institute of Neuro-
logical Disorder and Stroke, National Institute of Health, Bethesda,
USA following the reported procedures.
d
Male albino mice (CF-1 strain, 18e25 g) and male albino rats
(SpragueeDawley, 100e150 g) were used as experimental animals.
The synthesized derivatives were suspended in 0.5% methyl cellu-
lose and the test compound is usually manipulated with a motar
pestle to help preparation of suspension. The titled compounds
(PhQZ 1e4, PhQZ 8 and 10) were subjected to anticonvulsant
screening by MES and scMET model. The titled compounds (PhQZ
5e7 and 9, MtQZ 1e10 and PrQZ 1e10) were subjected to anti-
convulsant screening by 6 Hz psychomotor seizure test. In the
anticonvulsant screening by MES and scMET tests, each compound
was administered as an i.p. injection at three dose levels (30, 100
and 300 mg/kg) and anticonvulsant and neurotoxic effects were
assessed at 30 min and 4h intervals after administration. Similarly
in 6 Hz psychomotor seizure test screening was performed at five
different time points i.e., 0.25 h, 0.5 h, 1.0 h, 2.0 h and 4.0 h at a dose
of 100 mg/kg administered i.p. Neurotoxicity was observed by
minimal motor impairment which was measured by the rotarod
(neurotoxicity) test. Compounds showing significant protection
were evaluated for quantification studies in 6 Hz test and ED50 and
protection index (PI) value reported. Selected derivatives were also
evaluated in in-vitro hippocampal slice culture neuroprotection
assay.
7.1.7.24. 3-({(E)-[3-(4-Chlorophenoxy)phenyl]methylidene}amino)-
2-propylquinazolin-4(3H)-one (PrQZ 4). IR (KBr, cmꢀ1
)
y
: 3061 (Qu-
H, Ar-H), 1683 (C]O), 1608 (N]CH), 1444e1527 (C]C, benzene
and Qu-ring), 1242 (O); 1H NMR (CDCl3, 300 MHz)
in ppm: 8.47 (s,
d
1H, CH]N), 6.87e8.28 (a set of signals, 12H, Qu-H and Ar-H), 2.48
(t, 2H, CH2Qu), 1.87 (sextet, 2H, CH2Me), 1.05 (t, 3H, CH3); MS (m/z,
%): 418.14 (Mþ þ 1 for 35Cl, 43.12), 420.08 (Mþ þ 1 for 37Cl, 14.02).
7.1.7.25. 3-({(E)-[3-(4-Bromophenoxy)phenyl]methylidene}amino)-
2-propylquinazolin-4(3H)-one (PrQZ 5). IR (KBr, cmꢀ1
)
y
: 3049 (Qu-
H, Ar-H), 1680 (C]O), 1599 (N]CH), 1450e1526 (C]C, benzene
and Qu-ring), 1240 (O); 1H NMR (CDCl3, 300 MHz)
in ppm: 8.47 (s,
d
1H, CH]N), 6.78e8.26 (a set of signals, 12H, Qu-H and Ar-H), 2.47
(t, 2H, CH2Qu), 1.81 (sextet, 2H, CH2Me), 1.01 (t, 3H, CH3); MS (m/z,
%): 462.05 (Mþ þ 1 for 79Br, 28.31), 464.07 (Mþ þ 1 for 81Br, 27.11).
7.1.7.26. 3-({(E)-[3-(4-Fluorophenoxy)phenyl]methylidene}amino)-
2-propylquinazolin-4(3H)-one (PrQZ 6). IR (KBr, cmꢀ1
)
y
: 3052 (Qu-
H, Ar-H), 1684 (C]O), 1596 (N]CH), 1446e1530 (C]C, benzene
and Qu-ring), 1242 (O); 1H NMR (CDCl3, 300 MHz)
in ppm: 8.43 (s,
d
1H, CH]N), 6.80e8.25 (a set of signals, 12H, Qu-H and Ar-H), 2.47
(t, 2H, CH2Qu), 1.82 (sextet, 2H, CH2Me), 1.00 (t, 3H, CH3); MS (m/z,
%): 402.11 (Mþ þ 1, 34.55).
7.2.1. Maximal electroshock (MES) test
For MES test, 60 Hz of alternating current (50 mA in mice,
150 mA in rats) is delivered for 0.2 s by corneal electrodes which
have been primed with an electrolyte solution containing anes-
thetic agent (0.5% tetracaine HCl). An animal is considered pro-
tected from MES-induced seizures upon abolition of hind limb tonic
extensor component of the seizure.
7.1.7.27. 3-({(E)-[3-(4-Chloro-3-methylphenoxy)phenyl]methyl-
idene}amino)-2-propylquinazolin-4(3H)-one (PrQZ 7). IR (KBr,
cmꢀ1
)
y
: 3062 (Qu-H, Ar-H), 1678 (C]O), 1603 (N]CH), 1444e1526
(C]C, benzene and Qu-ring), 1244 (O); 1H NMR (CDCl3, 300 MHz)
in ppm: 8.42 (s, 1H, CH]N), 6.80e8.31 (a set of signals, 11H, Qu-H
d
and Ar-H), 2.44 (t, 2H, CH2Qu), 2.38 (s, 3H, Ar-CH3), 1.82 (sextet, 2H,
CH2Me), 1.06 (t, 3H, CH2CH3); MS (m/z, %): 432.11 (Mþ þ 1 for 35Cl,
47.17), 434.10 (Mþ þ 1 for 37Cl, 15.25).
7.2.2. Subcutaneous metrazol (scMET) seizure threshold test
For scMET test animals are pretreated with various doses of the
test compound. At a previously determined TPE of the test
compound the dose of metrazol which will induce convulsion in
97% of animals is injected into a loose fold of skin in the midline of
the neck. The animals are placed in an isolation cage to minimize
stress and observed for the next 30 min to see the absence of
seizure. An episode of clonic spasms, approximately 3e5 s of the
fore and/or hind limbs, jaws or vibrissae was taken as the end point.
Animals which do not meet this criterion were considered
protected.
7.1.7.28. 3-({(E)-[3-(Naphthalen-2-yloxy)phenyl]methylidene}
amino)-2-propylquinazolin-4(3H)-one (PrQZ 8). IR (KBr, cmꢀ1
) y:
3063 (Qu-H, Ar-H), 1676 (C]O), 1595 (N]CH), 1442e1517 (C]C,
benzene and Qu-ring), 1241 (O), 835, 823 (
-naphthyl); 1H NMR
(CDCl3, 300 MHz) in ppm: 8.48 (s, 1H, CH]N), 6.81e8.26 (a set of
b
d
signals, 15H, Qu-H, naphthyl-H and Ar-H), 2.44 (t, 2H, CH2Qu), 1.81
(sextet, 2H, CH2Me), 1.05 (t, 3H, CH3); MS (m/z, %): 434.16 (Mþ þ 1,
22.48).
7.2.3. Neurotoxicity-minimal motor impairment (MMI)
7.1.7.29. 3-({(E)-[3-(1,3-Benzodioxol-5-yloxy)phenyl]methylidene}
Minimal motor impairment was measured by the rotarod
(neurotoxicity) test. When a mouse is placed on a rod that rotates at
a speed of 6 rpm, the animal can maintain its equilibrium for a long
period of time. The compound was considered toxic if the treated
animal falls off this rotating rod 3 times during 1 min period.
amino)-2-propylquinazolin-4(3H)-one (PrQZ 9). IR (KBr, cmꢀ1
) y:
3057 (Qu-H, Ar-H), 1681 (C]O), 1603 (N]CH), 1455e1520 (C]C,
benzene and Qu-ring),1244 (O); 1H NMR (CDCl3, 300 MHz)
d
inppm:
8.42 (s,1H, CH]N), 6.78e8.26 (a setof signals,11H, Qu-H, sesamol-H
and Ar-H), 5.96 (s, 2H, CH2), 2.46 (t, 2H, CH2Qu), 1.84 (sextet, 2H,
CH2Me), 1.06 (t, 3H, CH3); MS (m/z, %): 428.07 (Mþ þ 1, 34.23).
7.2.4. 6 Hz psychomotor seizure test
6 Hz psychomotor seizure test or minimal clonic seizure (6 Hz)
test was used to assess compound’s efficacy against electrically
induced seizures but used a lower frequency (6 Hz) and longer
duration of stimulation (3 s). Test compounds were pre-adminis-
tered to mice via i.p. injection. At varying times, individual mice
(four mice per time point) were challenged with sufficient current
delivered through corneal electrodes to elicit a psychomotor
seizure in 97% animals (32 mA for 3 s). The untreated mice would
7.1.7.30. 3-[(2-Oxo-1,2-dihydro-3H-indol-3-ylidene)amino]-2-pro-
pylquinazolin-4(3H)-one (PrQZ 10). IR (KBr, cmꢀ1
)
y
: 3212 (NH),
3062 (Qu-H, Isatin-H), 1673 (C]O), 1611 (N]C), 1466e1529 (C]C,
benzene and Qu-ring); 1H NMR (CDCl3, 300 MHz)
in ppm: 10.87
d
(s, 1H, NH of Isatin, D2O exchangeable), 6.77e8.24 (a set of signals,
8H, Qu-H and Isatin-H), 2.42 (t, 2H, CH2Qu), 1.82 (sextet, 2H,
CH2Me), 1.03 (t, 3H, CH3); MS (m/z, %): 333.10 (Mþ þ 1, 21.14).

1018
P. Kumar et al. / European Journal of Medicinal Chemistry 46 (2011) 1006e1018
display seizure characterized by a minimal clonic phase followed by
stereotyped, automatistic behaviors, described originally as being
similar to the aura of human patients with partial seizure. Animals
not displaying this behavior are considered to be protected. Most
potent derivatives were tested quantitatively in the 6 Hz study and
ED50 reported.
24 h at room temperature and then transferred to a separating
funnel and allowed to dynamic equilibrate for 6 h. The aqueous and
octanol phases were separated and filtered through membrane
filter and drug content in aqueous phase was analyzed by UV
spectroscopy. Theoretical milog P for synthesized compounds was
then compared with the experimental log P data.
7.2.5. In-vitro hippocampal slice culture neuroprotection assay
(NP): primary screen experiment
The "Primary Screen Experiment" is a qualitative assessment of
the ability of a compound to prevent excitotoxic cell death. Orga-
7.3.4. Docking study
Compounds PhQZ 7, MtQZ 3 and PrQZ 10 were selected as
ligands for docking studies with six established epilepsy receptors
namely GABA(A) alpha-1, GABA(A) delta, glutamate, Na/H
exchanger, Na channel and T-type calcium channel receptor. These
receptors are the most important molecular targets in the design
and discovery of successful antiepileptic drugs [18]. In the present
study, AutoDock 4.0 with its Lamarckian genetic algorithm (LGA)
was used for automated flexible ligand docking of PhQZ 7, MtQZ 3
and PrQZ 10 with above mentioned receptors and affinity (kcal/
mol) and H-bond properties were evaluated.
notypic hippocampal slice cultures are treated with N-methyl-
aspartate (NMDA) or kainic acid (KA) to induce neuronal cell death.
Propidium iodide, membrane-impermeant compound, is
D-
a
included in all wells of the culture plate. Dying cells have
compromised cell membranes, thus propidium iodide may diffuse
into the cell, intercalate with DNA and fluoresce. Thus, the intensity
of the propidium iodide fluorescence is proportional to the amount
of cell death in the individual slices. Hippocampal slice cultures are
treated with the excitotoxin alone, or where indicated above, with
the excitotoxin and either one or two investigational compounds at
the concentrations indicated. If neuroprotection occurs as a conse-
quence of the added compound, slice cultures will have a visibly
reduced fluorescent intensity when compared to the slice cultures
that have been treated with the excitotoxin alone.
Acknowledgments
The authors would like to express their gratitude to IIT, Delhi
and CDRI, Lucknow for providing the spectral and elemental data.
One of the authors (Praveen Kumar) is thankful to S. D. College of
Pharmacy and Vocational Studies, Muzaffarnagar-251001, India for
providing research facilities. The authors also appreciate the help
and support by Mr. Om Prakash during docking study.
7.3. Computational study
7.3.1. Distance mapping
References
The pharmacophore pattern studies in which distance between
the various groups postulated as essential for anticonvulsant
activity were done on the 3D optimized structures using ACD/3D
viewer version 12.01 and Argus Lab 4.0 Mark A. Thompson Planaria
Software LLC. In conformational analysis of the ten clinically
effective, well known and structurally different anticonvulsant
drugs such as albutoin, carbamazepine, gabapentin, lamotrigine,
mephobarbital, phenytoin, progabide, ralitoline, remacemide, and
zonisamide; a molecular model was suggested on the basis of
molecular dynamics distance estimations [13].
[1] A.V.K. Srivastava, A. Kumar, Eur. J. Med. Chem. 37 (2002) 873e882.
[2] C.B. Chapleo, M. Mayer, P.L. Myer, J.F. Saville, A.C.B. Smith, M.R. Stilling,
I.F. Tulloch, D.S. Walter, A.P. Welbourn, J. Med. Chem. 29 (1986) 2273e2280.
[3] A.V.K. Srivastava, A. Kumar, Bioorg. Med. Chem. 12 (2004) 1257e1264.
[4] M.A. Aziza, M.K. Ibrahim, A.G. El-Helpy, Al-Azhar J. Pharm. Sci. 14 (1994)
193e201.
[5] A.A. Bekhit, M.A. Khalil, Pharmazie 53 (1998) 539.
[6] N. Ergenc, S. Buyuktimkin, G. Capan, G. Baktir, S. Rollas, Pharmazie 46 (1991)
290e291.
[7] S.K. Kashaw, V. Kashaw, P. Mishra, N.K. Jain, ARKIVOC xiv (2008) 17e26.
[8] Y. Wang, S. Cai, N. Lan, J. Keana, V. Ilyin, E. Weber, PCT/US98/08004 (1998)
1e81.
[9] K.H. Boltze, H.D. Dell, H. Lehwald, D. Loranz, M. Ruberg-Schweer, Arzneim-
Forsch./Drug Res. 13 (1963) 688.
[10] J.F. Wolfe, T.L. Rathman, M.C. Sleevi, J.A. Campbell, T.D. Greenwood, J. Med.
Chem. 33 (1990) 161.
[11] K. Unverferth, J. Engel, N. Hofgen, A. Rostock, R. Gunther, H.J. Lankau,
M. Menzer, A. Rolfs, J. Liebscher, B. Muller, H.J. Hofmann, J. Med. Chem. 41
(1998) 63e73.
[12] P. Ertl, B. Rohde, P. Selzer, J. Med. Chem. 43 (2000) 3714e3717.
[13] P. Yogeeswari, D. Sriram, R. Thirumurugan, J.V. Raghavendran, K. Sudhan,
R.K. Pavana, J.P. Stables, J. Med. Chem. 48 (2005) 6202e6211.
[14] C.A. Lipinski, L. Lombardo, B.W. Dominy, P.J. Feeney, Adv. Drug Deliv. Rev. 46
(2001) 3e26.
7.3.2. Calculation of physicochemical parameters
A computational study of titled compounds was performed for
prediction of ADME properties. Polar surface area (TPSA) [12],
milog P, number of rotatable bonds, molecular volume, number of
hydrogen donor and acceptor atoms and violations of Lipinski’s rule
of five [14] were calculated using Molinspiration online property
calculation toolkit [15]. Absorption (%ABS) was calculated by:
%ABS ¼ 109-(0.345 ꢃ TPSA) [16].
[15] Molinspiration Cheminformatics, Bratislava, Slovak Republic, Available from:
http://www.molinspiration.com/services/properties.html, (accessed 16.09.10).
[16] Y. Zhao, M.H. Abraham, J. Lee, A. Hersey, N.C. Luscombe, G. Beck, B. Sherborne,
I. Cooper, Pharm. Res. 19 (2002) 1446e1457.
[17] V.A. Farrar, M.R. Ciechanowicz, J. Grochowski, P. Serda, T. Pilati, G. Filippini,
C.N. Hinko, A. El-Assadi, J.A. Moore, I.O. Edafiogho, C.W. Andrews, M. Cory,
J.M. Nicholson, J.R. Scott, J. Med. Chem. 36 (1993) 3517e3525.
[18] C.J. Landmark, Med. Sci. Monit. 13 (2007) RA1e7.
7.3.3. log P determination
The partition coefficient between octanol and phosphate buffer
was determined at room temperature [17]. 10 mL of octanol and
10 mL phosphate buffer were taken in a glass stoppered graduated
tube and 5 mg of accurately weighed compound was added. The
mixture was then shaken with the help of mechanical shaker for