N-{[(6-Substituted-1,3-benzothiazole-2-yl)amino]carbonothioyl}-2/4-substituted benzamides: Synthesis and pharmacological evaluation

Article abstract of DOI:10.1016/j.ejmech.2007.07.008

A series of 1,3-benzothiazol-2-yl benzamides (11-30) were prepared in satisfactory yield and evaluated for their anticonvulsant, neurotoxicity, CNS depressant study and other toxicity studies. All the synthesized compounds were in good agreement with elemental and spectral data. Majority of the compounds were active in MES and scPTZ screen and showed the decrease in the immobility time. None of the compounds had shown neurotoxicity or liver toxicity.

Full text of DOI:10.1016/j.ejmech.2007.07.008

Available online at www.sciencedirect.com
European Journal of Medicinal Chemistry 43 (2008) 1114e1122
http://www.elsevier.com/locate/ejmech
Preliminary communication
N-{[(6-Substituted-1,3-benzothiazole-2-yl)amino]carbonothioyl}-2/4-
substituted benzamides: Synthesis and pharmacological evaluation
a
b
Arpana Rana a, Nadeem Siddiqui a, , Suroor A. Khan , Syed Ehtaishamul Haque ,
*
Mashooq A. Bhat a
a Hamdard University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry, New Delhi 110 062, India
b Hamdard University, Faculty of Pharmacy, Department of Pharmacology, New Delhi 110 062, India
Received 4 February 2007; received in revised form 4 July 2007; accepted 9 July 2007
Available online 29 July 2007
Abstract
A series of 1,3-benzothiazol-2-yl benzamides (11e30) were prepared in satisfactory yield and evaluated for their anticonvulsant, neurotox-
icity, CNS depressant study and other toxicity studies. All the synthesized compounds were in good agreement with elemental and spectral data.
Majority of the compounds were active in MES and scPTZ screen and showed the decrease in the immobility time. None of the compounds had
shown neurotoxicity or liver toxicity.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Benzamides; Benzothiazole; Anticonvulsant; Neurotoxicity; CNS depressant activity
1. Introduction
of epilepsy. Another way is to modify already existing drugs
and formulations. The long established anticonvulsants
control seizures in 50% of patients developing partial
seizures and 60e70% of those developing generalized seizures
[5e7]. Hence there is an urgent need to develop new anticon-
vulsants [8].
The benzamides are a class of compounds presenting
a wide range of biological applications and anticonvulsant
activity [9e12]. A variety of benzamides reported to possess
anticonvulsant activity in a number of animal models used
to predict potential anticonvulsant activity in man. Recently
pharmacophore model was proposed based on the semicarba-
zones [13] according to which essential features for the
anticonvulsant activity are (a) a hydrophobic aryl ring, (b) a hy-
drogen bonding domain, (c) an electron donor acceptor system
and (d) an another hydrophobic aryl ring responsible for
metabolism, the size of ring may differ Fig. 1.
The term epilepsy refers to a disorder of the brain function
characterized by the periodic and unpredictable occurrence of
seizures. Epilepsies are common and frequently devastating
and affect around 1e2% of the world population. The convul-
sions of approximately 25% of epileptics are inadequately
controlled by the standard drug therapy [1,2]. The number
of drugs useful for the treatment of epilepsy is remarkably
small. Fewer than 20 drugs are currently marketed in the
United States and of these only five or six are widely used [2].
The search for antiepileptic compounds with a more selec-
tive activity and lower toxicity continues to be an area of in-
vestigation in medicinal chemistry. A rational drug design
process of a new anticonvulsant could be achieved in several
ways [3,4]. The first strategy is the identification of new tar-
gets through better understanding of molecular mechanisms
In our study N-{[(6-substituted-1,3-benzothiazole-2-yl)-
amino]carbonothioyl}-2/4-substituted benzamides were syn-
thesized and evaluated for pharmacological activity to test
* Corresponding author. Tel.: þ91 011 2605 9688x5639; fax: þ91 011 2605
9688 5307.
E-mail address: nadeems_03@yahoo.co.in (N. Siddiqui).
this hypothesis. Benzothiazole, a versatile heterocyclic
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.07.008

A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
1115
pentylenetetrazole (scPTZ) [19] and neurotoxicity screen [20],
using doses of 30, 100, 300 mg/kg and observations carried
out at two different time intervals. These data are presented
in Table 2. These compounds were then evaluated for their
CNS behavioral activity in mice using porsolt’s swim pool
test [21] in rats, results are presented in Table 2. The selected
compounds were evaluated for liver toxicity by enzyme esti-
mation [22e24] and histopathological studies [25]. Data are
presented in Table 3 and Fig. 2 and 3.
Fig. 1. Suggested pharmacophore model for anticonvulsant activity. A: hydro-
phobic domain, B: hydrogen bonding site, C: distal hydrophobic domain, D:
electron donor moiety.
4. Results and discussions
The preliminary evaluation of the synthesized N-{[(6-
substituted-1,3-benzothiazole-2-yl)amino]carbonothioyl}-2/4-
substituted benzamides 11e30 were evaluated for
anticonvulsant activity by the use of predictable animal
models and the neurotoxicity was checked by rotarod test
method. Of these, the MES and scPTZ seizure models repre-
sent the two animal seizure models, most widely used in the
search for new anticonvulsants. Data is presented in Table 2
after the 0.5 and 4 h time intervals at the dose level of 30,
100 and 300 mg/kg. Phenytoin and carbamazepine were used
as the standard drugs for the comparison.
Maximal electroshock seizure (MES) test is a proven
method to check the generalized toniceclonic seizure and
identifies clinical candidates that prevent seizure spread. The
synthesized compounds 12, 18, 19, 20, 22, 24 and 26 had
shown activity at the dose level of 30 mg/kg against the sei-
zure spread at both the time intervals except compounds 18,
20 and 22 which had shown protection at the dose level of
100 mg/kg after 4 h indicating rapid onset but shorter duration
of action. Whereas compounds 15, 21 and 23 had shown pro-
tection at both the time intervals 0.5 and 4 h after the dose
level of 100 mg/kg, compound 23 had shown activity at
300 mg/kg after the 4 h interval. Compounds 11, 13, 14, 16,
17, 25, 29 and 30 had shown protection at the dose level of
300 mg/kg at both the time intervals. In the scPTZ screen
compounds 12, 19 and 26 had shown activity at 30 mg/kg
dose level after both the time intervals except 12, 26 which
had shown activity at the dose of 100 mg/kg at both the
time intervals but compounds 11, 18, 22 had shown activity
at the dose level of 300 mg/kg. Rest of the compounds had ac-
tivity at the dose level of 300 mg/kg at both the time intervals
except compounds 23, 27e29 which had inactivity. Selected
compounds which had shown activity at 30 mg/kg dose level
were further tested for their neurotoxicity. None of the com-
pounds showed any sign of neurotoxicity.
hydrophobic molecule possessing preliminary anticonvulsant
properties [14,15] has been selected for the hydrophobic bind-
ing site. Benzothiazole were condensed with the benzamides
with the hope to potentiate the biological activities with lesser
or limited amount of toxicities.
2. Chemistry
The synthesis of N-{[(6-substituted-1,3-benzothiazole-2-yl)-
amino]carbonothioyl}-2/4-substituted benzamides 11e30 was
accomplished as presented in Scheme 1. The 6-substituted-
1,3-benzothiazole-2-amines 1e6 were synthesized by reacting
aryl amines with potassium thiocyanates in a satisfactory yield
by a known preparation method [16]. In FTIR spectrum bands
at 3490e3210 cmꢀ1 and 1580e1520 cmꢀ1 confirms presence
of NH and C]N stretching vibrations, respectively. The
1H NMR spectrum showed a singlet at d 5.33e7.21 ppm due
to NH2 protons (D2O exchangeable). 2/4-Substituted benzoyli-
sothiocyanates 7e10 were obtained by reacting substituted ben-
zoic acid and thionyl chloride, substituted benzoyl chloride
was obtained which was further treated with ammonium thiocy-
anate to get 1e6. The FTIR bands showed bands for C]O and
NeCeS at 1670e1628 cmꢀ1 and 2180e2100 cmꢀ1, respec-
1
tively. The H NMR spectrum showed multiplet at d 7.20,
7.79e7.48, 8.01 for aromatic protons.
Final compounds N-{[(6-substituted-1,3-benzothiazole-
2-yl)amino]carbonothioyl}-2/4-substituted benzamides 11e30
were synthesized by refluxing 6-substituted-1,3-benzothiazol-
2-amines 1e6 and 2/4-substituted benzoylisothiocyanates
7e10. The synthesized compounds were characterized by ele-
1
mental analysis, FTIR, H NMR and mass spectrum. FTIR
spectrum revealed three bands for NH, C]O and C]S at
3448e3110 cmꢀ1, 1790e1628 cmꢀ1 and 1150e1045 cmꢀ1
,
1
respectively. The H NMR of 11e30 confirmed presence of
NHC]O and NHC]S by showing singlet at d 7.78e9.81
and broad singlet at d 9.88e13.27, respectively, exchangeable
with D2O. The physical and chemical data for newly synthe-
sized compounds are presented in Table 1.
Selected compounds which were showing protection
against seizures at the dose level of 30 mg/kg in MES and
scPTZ screen were further tested for their neurotoxicity.
None of the compounds showed any sign of neurotoxicity.
Basic structure of the compounds fulfilled all the pharma-
cophoric structural requirements like presence of benzothia-
zole as hydrophobic aryl ring, N as electron donor acceptor
system and another hydrophobic aryl ring responsible for me-
tabolism proposed by Pandeya et al. [26] except the presence
of hydrophobic domain Fig. 4.
3. Pharmacology
The synthesized compounds (11e30) obtained from the re-
actional sequence, were injected intraperitoneally into mice
and in the maximal electroshock (MES) [17,18], subcutaneous

1116
A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
N
Glacial acetic acid
Br2
R
+
KSCN
NH2
R
S
NH2
6-Substitued-1,3-benzothiazol
-2-amines
(1-6)
Reflux/Benzene
1 h
COOH + SOCl2
COCl
R1
R1
NH4SCN
Reflux, 30 min
CONCS
R
1
2/4-Substitutedbenzoylisothiocyanates
(7-10)
N
+
CONCS
S
NH2
R
R
1
(7-10)
(1-6)
Reflux/Benzene
5 h
N
R
S
N
H
N
H
O
S
R
1
R = Br, Cl, F, NO2, CH3, OCH3
= H, 2-Cl, 4-Cl, 4-OCH3
R
1
Scheme 1. N-{[(6-Substituted-1,3-benzothiazole-2-yl)amino]carbonothioyl}-2/4-substituted benzamides (11e30).
In general compounds bearing the groups like F, CH3, OCH3
and total protein suggested that none of the compounds had
shown any significant increase or decrease. Thus considering
the biochemical parameters estimated to establish liver func-
tion test with selected compounds, it was clearly indicated
that none of the compounds showed any malfunctioning or
toxicity of the liver as compared to control. Further histopath-
ological study of liver confirmed that there is no liver toxicity
by showing normal hepatic parenchyma with portal triad,
central vein and heptocytes in comparison to control (Figs. 2
and 3).
The forced swim test is a well validated and extensively used
screen for compounds with antidepressant activity. Compounds
11e30 were also evaluated for their antidepressant activity with
the standard fluoxetine at the dose level of 30 mg/kg. Antide-
pressant activity was assessed as mean average immobility
time in seconds and data is presented as mean ꢁ SEM (Table 3).
Data was analyzed by using students‘t’ test against the individ-
ual untreated controls with the treated compounds. Compounds
13, 15e17, 20, 22, 24 and 30 showed decrease in the immobility
time and were highly potent ( p < 0.001) equivalent to the
standard fluoxetine ( p < 0.001). Compounds 11, 21, 23, 25,
29 had shown highly significant activity with ( p < 0.01).
Compounds 18 and 19 had shown statistically significant
at the 6-position of benzothiazole ring with H, 2-Cl, 4-Cl
substituted distant phenyl ring showed highly potent activity
in MES and scPTZ tests. Whereas replacement with lesser elec-
tronegative Br group at the 6-position of benzothiazole ring with
4-OCH3 substituent at the distant phenyl ring resulted in com-
pounds with decrease in activity to a lesser extent. Presence of
substituent like NO2 at the 6-position of benzothiazole ring,
with the unsubstituted distant phenyl ring in compound, showed
significant activity whereas substitution with 4-OCH3 at the dis-
tant phenyl ring renders the compound inactive.
Hepatotoxic reactions have contributed to the decline of
many therapies. Table 3 shows the liver function tests with ref-
erence to the control. Studies reported earlier have shown that
phenytoin is a possible cause of acetaminophen hepatotoxicity
[27] and anticonvlsants such as carbamazepine [28] and
valporic acid [29] were also found to enhance or show hepto-
toxic side effects. In the present study some selective
compounds showing protection at 30 mg/kg in both the tests
were administered chronically to animals for 15 days and
the biochemical parameters were estimated. The values of
alkaline phosphatase, serum glutamate oxaloacetate transami-
nase (SGOT), serum glutamate pyruvate transaminase (SGPT)

A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
1117
Table 1
Physico-chemical properties of compounds 11e30
d
Compd. no.
R
R1
Mol. formula (MW)a
Yield (%)
bMp. (ꢂC)
Rfc (Rm)
Found (calcd.)%
C
H
N
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
a
Cl
F
H
C
15H10ClN3OS2 (347.84)
80
82
69
68
75
67
88
75
79
80
65
85
83
80
78
75
73
61
70
71
252
217
295
198
212
180
217
170
215
175
187
195
173
213
178
203
157
257
214
162
0.71 (ꢀ0.38)
0.88 (ꢀ0.86)
0.69 (ꢀ0.34)
0.81 (ꢀ0.62)
0.82 (ꢀ0.65)
0.94 (ꢀ1.19)
0.73 (ꢀ0.43)
0.87 (ꢀ0.82)
0.86 (ꢀ0.78)
0.81 (ꢀ0.62)
0.92 (ꢀ1.06)
0.77 (ꢀ0.52)
0.89 (ꢀ0.90)
0.88 (ꢀ0.86)
0.85 (ꢀ0.75)
0.69 (ꢀ0.34)
0.91 (ꢀ1.00)
0.90 (ꢀ0.95)
0.83 (ꢀ0.68)
0.93 (ꢀ1.12)
51.87 (51.79)
54.32 (54.37)
50.47 (50.27)
58.88 (58.69)
56.00 (55.96)
42.42 (42.22)
47.34 (47.13)
49.46 (49.25)
53.32 (53.11)
51.01 (50.86)
44.20 (42.22)
47.34 (47.13)
50.14 (49.25)
53.17 (53.11)
51.08 (50.86)
51.07 (50.80)
53.27 (53.17)
49.69 (49.47)
57.32 (57.12)
54.88 (54.67)
2.96 (2.90)
3.06 (3.04)
2.84 (2.81)
3.97 (4.00)
3.79 (3.82)
2.16 (2.10)
2.30 (2.37)
2.58 (2.48)
3.30 (3.34)
2.98 (3.20)
2.16 (2.13)
2.34 (2.37)
2.45 (2.48)
3.37 (3.34)
3.15 (3.20)
3.19 (3.20)
3.85 (3.35)
3.15 (3.11)
4.13 (4.23)
4.06 (4.05)
10.21 (10.19)
12.86 (12.68)
15.60 (15.63)
12.72 (12.83)
12.20 (12.24)
9.81 (9.85)
H
C15H10FN3OS2 (331.38)
C15H10N4O3S2 (358.39)
C16H13N3OS2 (327.42)
C16H13N3O2S2 (343.42)
C15H9ClBrN3OS2 (426.73)
C15H9Cl2N3OS2 (382.28)
C15H9ClFN3OS2 (365.83)
C16H12ClN3OS2 (361.86)
C16H12ClN3O2S2 (377.86)
C15H9ClBrN3OS2 (426.73)
C15H9Cl2N3OS2 (382.28)
C15H9ClFN3OS2 (365.83)
C16H12ClN3OS2 (361.86)
C16H12Cl N3O2S2 (377.86)
C16H12ClN3O2S2 (377.86)
C16H12FN3O2S2 (361.41)
C16H12N4O4S2 (388.42)
C17H15N3O2S2 (357.44)
C17H15N3O3S2 (373.44)
NO2
CH3
CH3O
Br
H
H
H
2-Cl
2-Cl
2-Cl
2-Cl
2-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
CH3O
CH3O
CH3O
CH3O
CH3O
F
10.95 (10.99)
11.44 (11.49)
11.56 (11.61)
11.07 (11.12)
10.81 (9.85)
11.03 (10.99)
11.79 (11.49)
11.66 (11. 61)
11.17 (11.12)
11.08 (11.12)
11.02 (11.63)
14.38 (14.42)
11.70 (11.76)
11.20 (11.25)
NO2
CH3
CH3O
Br
F
NO2
CH3
CH3O
Br
F
NO2
CH3
CH3O
Solvent of crystallization: benzene.
Melting point of the compounds at their decomposition.
Solvent system T:E:F (5:4:1).
b
c
d
A logarithmic function of Rf value was also calculated; Rm ¼ log (1 ꢀ 1/Rf).
results ( p < 0.05), whereas remaining compounds were devoid
of activity. Compounds with substitution with Br, Cl, F, NO2,
CH3 and OCH3 at the 6-position of benzothiazoleanddistantphe-
nyl ring substituted with H, 2-Cl, 4-Cl, 4-OCH3 were highly
potent.
were performed on the CHNS Elimentar (Analysen systime,
GmbH) Germany Vario EL III. All the Fourier transform infra
red (FTIR) spectra were recorded in KBr pellets on a Jasco
1
FT/IR 410 spectrometer. The H NMR spectra were taken
on a Bruker model dpx 300 (300 MHz) NMR spectrometer.
Chemical shifts (d) are expressed in ppm relative to tetra-
methyl silane (TMS) as an internal standard. Mass spectra
were measured on a Brooker Ion trap (Esquire 3000) mass
spectrometer from Regional Research Laboratory (RRL),
Jammu, India.
The homogeneity of the compounds was checked by thin
layer chromatography (TLC) on silica gel G (Merck) coated
plates by using toluene:ethyl acetate:formic acid (5:4:1) as sol-
vent system. Iodine chamber and UV lamp were used for the
visualization of TLC spots.
From the present study it was concluded that all the pharma-
cophoric elements were present in the titled compounds;
although the HBD is not in accordance of the proposed
pharmacophore model (Fig. 4). All the compounds except 23,
27e29 showed protection in the two animal models of seizures.
Compounds 12, 19, 26 emerged as anticonvulsants with no neu-
rotoxicity and can be claimed to detect compounds possessing
against generalized toniceclonic (grand mal) and generalized
absence (petit mal) seizures, respectively. Different substitu-
tions at the distal aryl ring with halogens and alkyl groups re-
sulted in variations like increase and decrease in the activity.
In the toxicity studies none of the compounds showed neurotox-
icity or liver toxicity. In CNS depressant study, most of the com-
pounds showed the decrease in the immobility time. Dose used
for anticonvulsant and CNS depressant activity are 30 mg/kg,
but we cannot suggest the mechanism of action involved for
both the activities effect. However, to confirm this fact, further
studies are required to be carried out.
5.1.1. 6-Substituted-1,3-benzothiazol-2-amine (1e6)
A mixture of aniline (0.01 mol) and potassium thiocyanate
(0.01 mol) in glacial acetic acid (10%) was cooled and stirred.
To this solution bromine (0.01 mol) was added dropwise at
such a rate to keep the temperature below 10 ꢂC throughout
the addition. Stirring was continued for an additional 3 h
and the separated hydrochloride salt was filtered, washed
with acetic acid and dried. It was dissolved in hot water and
neutralized with aqueous ammonia solution (25%), filtered,
washed with water and dried, recrystallized with benzene to
obtain 6-substituted-1,3-benzothiazol-2-amine.
5. Experimental protocols
5.1. Chemistry
The melting points were determined in open capillary tubes
in a Hicon melting point apparatus and are uncorrected. The
homogen elemental analyses (C, H, N) of all compounds
5.1.1.1. 6-Chloro-1,3-benzothiazol-2-amine (1). Yield 74%;
IR: nmax (cmꢀ1) 3490 (NH), 1580 (C]N); 1H NMR

1118
A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
Table 2
Anticonvulsant, neurotoxicity screening and CNS depression study of compounds 11e30
Compd. no
Intraperitoneal injection in micea
Mean average immobility time (s)c
MES screen
scPTZ
0.5 h
NTb
Mean ꢁ SEM
Untreated
0.5 h
4 h
4 h
0.5 h
4 h
Treated
11
12
300
30
300
30
100
30
300
100
300
300
300
300
300
300
30
X
X
42.0 ꢁ 1.8
34.4 ꢁ 1.4
34.0 ꢁ 1.6
35.8 ꢁ 1.6
41.0 ꢁ 2.1
42.0 ꢁ 1.2
49.0 ꢁ 1.2
34.6 ꢁ 1.6
58.2 ꢁ 1.8
50.6 ꢁ 1.3
48.2 ꢁ 2.5
39.8 ꢁ 1.3
39.8 ꢁ 1.3
47.0 ꢁ 1.0
55.0 ꢁ 1.9
40.8 ꢁ 2.5
31.4 ꢁ 1.3
28.8 ꢁ 0.9
26.2 ꢁ 1.7
23.2 ꢁ 0.8
e
29.0 ꢁ 1.9**
24.0 ꢁ 1.6
14.8 ꢁ 1.5***
29.0 ꢁ 1.6
19.8 ꢁ 1.8***
22.6 ꢁ 1.0***
32.6 ꢁ 1.5***
25.0 ꢁ 1.4*
40.4 ꢁ 1.5*
42.6 ꢁ 2.4***
38.0 ꢁ 2.3**
20.2 ꢁ 1.5***
24.6 ꢁ 1.4**
30.0 ꢁ 1.5***
43.0 ꢁ 1.7**
34.2 ꢁ 2.6
31.2 ꢁ 1.2
27.8 ꢁ 0.7
10.2 ꢁ 1.1**
16.8 ꢁ 0.9***
e
(e)
X
(e)
X
13
14
300
300
100
300
300
30
300
300
100
300
300
100
30
300
300
300
300
300
100
30
X
X
15
16
X
X
X
X
17
18
X
X
(e)
(e)
(e)
X
(e)
(e)
(e)
X
19
20
30
30
100
100
100
300
100
300
30
100
300
100
e
100
300
300
e
21
22
100
30
(e)
X
(e)
X
23
24
100
30
100
300
30
100
300
100
e
(e)
X
(e)
X
25
26
300
30
(e)
X
(e)
X
27
28
e
e
e
e
e
e
e
e
e
X
X
29
30
300
300
30
300
300
30
X
X
300
e
300
e
X
X
Phenytoin
Carbamazepine
Fluoxetine
(e)
(e)
e
(e)
(e)
e
30
29.6 ꢁ 0.6
100
14.2 ꢁ 0.9***
100
e
100
e
e
e
e
e
n ¼ 6.
a
Doses of 30, 100 and 300 mg/kg were administered. The data indicates the minimum dose whereby bioactivity was demonstrated in half or more of the mice.
The animals were examined at 0.5 and 4 h.
b
NT indicates neurotoxicity screening using rotorod test. A dash (e) indicates an absence of activity. X denotes not tested.
Dose ¼ 30 mg/kg (p.o); p* < 0.05, p** < 0.01, p*** < 0.001. Data was analyzed by unpaired students ‘t’ test.
c
(DMSO-d6): (d, ppm) 6.12 (s, 2H, NH2, D2O exchangeable),
6.79e7.13 (m, 3H, AreH).
(d, ppm) 6.30 (s, 2H, NH2, D2O exchangeable), 7.17e8.21
(m, 3H, AreH).
5.1.1.2. 6-Fluoro-1,3-benzothiazol-2-amine (2). Yield 81%;
IR: nmax (cmꢀ1) 3318 (NH), 1568 (C]N); 1H NMR
(DMSO-d6): (d, ppm) 6.73e7.06 (m, 2H, AreH), 7.21 (s,
2H, NH2, D2O exchangeable), 7.30 (m, 1H, AreH).
5.1.1.4. 6-Methyl-1,3-benzothiazol-2-amine (4). Yield 80%;
IR: nmax (cmꢀ1) 3240 (NH), 1520 (C]N); 1H NMR
(DMSO-d6): (d, ppm) 2.05 (s, 3H, CH3), 5.21 (s, 2H, NH2,
D2O exchangeable), 6.63e6.94 (m, 3H, AreH).
5.1.1.3. 6-Nitro-1,3-benzothiazol-2-amine (3). Yield 77%; IR:
5.1.1.5. 6-Methoxy-1,3-benzothiazol-2-amine (5). Yield 78%;
IR: nmax (cmꢀ1) 3300 (NH), 1583 (C]N); 1H NMR
1
nmax (cmꢀ1) 3318 (NH), 1568 (C]N); H NMR (DMSO-d6):
Table 3
Enzyme estimation of the selected compounds
Treatment
Alkaline phosphatase ꢁ SEMa, ns
SGOT ꢁ SEMb, ns
SGPT ꢁ SEMc, ns
Total Protein
(g/100 ml) ꢁ SEMd, ns
Control
12
18
13.06 ꢁ 0.25
13.64 ꢁ 0.06
13.30 ꢁ 0.03
13.96 ꢁ 0.07
13.04 ꢁ 0.09
12.98 ꢁ 1.00
13.48 ꢁ 0.20
13.53 ꢁ 0.38
148.6 ꢁ 1.50
149.3 ꢁ 1.23
148.0 ꢁ 0.45
147.9 ꢁ 0.02
148.6 ꢁ 0.08
147.3 ꢁ 0.01
149.1 ꢁ 0.02
146.8 ꢁ 0.40
27.67 ꢁ 0.84
27.89 ꢁ 0.71
29.67 ꢁ 0.60
29.33 ꢁ 0.61
27.98 ꢁ 0.34
27.81 ꢁ 0.62
27.48 ꢁ 0.45
28.49 ꢁ 0.08
1.80 ꢁ 0.01
1.83 ꢁ 0.03
1.71 ꢁ 0.03
1.98 ꢁ 0.75
1.90 ꢁ 0.32
1.88 ꢁ 0.23
1.79 ꢁ 0.08
1.81 ꢁ 0.30
19
20
22
24
26
ns denotes non-significant. Results given as mean ꢁ SEM were calculated using ANOVA followed by Dunnett’s comparison test.
a
P ¼ 0.6382.
b
P ¼ 0.2133.
c
P ¼ 0.0847.
d
P ¼ 0.9995.

A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
1119
S
C
O
C
N
C
A
NH
NH
S
D
HBD
Fig. 4. Showing A: hydrophobic aryl ring, C: distal hydrophobic domain, D:
electron donor domain, HBD: Hydrogen bond acceptor/donor moiety in basic
structure of titled compounds 11e30.
The reaction mixture upon filtration yielded substituted ben-
zoyl chloride as a plummy liquid.
Ammonium thiocyanate (0.1 mol) was added to substituted
benzoyl chloride (0.1 mol) and refluxed for 30 min in benzene.
The resultant mixture was filtered and substituted benzoyl iso-
thiocyanates (7e10) were obtained in the form of liquid and
ammonium chloride as a residue. Ammonium chloride was
removed by filtration as a residue from the mixture.
5.1.2.1. Benzoyl isothiocyanate (7). Yield 68%; IR: nmax
1
(cmꢀ1) 2154 (NeCeS), 1628 (C]O); H NMR (DMSO-d6):
(d, ppm) 7.48e8.01 (dd, 5H, AreH).
Fig. 2. Section of liver. Group: control, interference: normal hepatic paren-
chyma with portal triad (PT), central vein (CV). Magnification: 100ꢃ.
(DMSO-d6): (d, ppm) 3.81 (s, 3H, OCH3), 5.33 (s, 2H, NH2,
D2O exchangeable), 6.66e7.07 (m, 3H, AreH).
5.1.2.2. 2-Chlorobenzoyl isothiocyanate (8). Yield 62%; IR:
1
nmax (cmꢀ1) 2100 (NeCeS), 1650 (C]O), 840 (CeCl); H
NMR (DMSO-d6): (d, ppm) 7.30e7.89 (dd, 4H, AreH).
5.1.1.6. 6-Bromo-1,3-benzothiazol-2-amine (6). Yield 82%;
IR: nmax (cmꢀ1) 3210 (NH), 1560 (C]N); 1H NMR
(DMSO-d6): (d, ppm) 6.03 (s, 2H, NH2, D2O exchangeable),
6.99e7.33 (m, 3H, AreH).
5.1.2.3. 4-Chlorobenzoyl isothiocyanate (9). Yield 60%; IR:
1
nmax (cmꢀ1) 2250 (NeCeS), 1670 (C]O), 836 (CeCl); H
NMR (DMSO-d6): (d, ppm) 7.38e7.95 (dd, 4H, AreH).
5.1.2.4. 4-Methoxybenzoyl isothiocyanate (10). Yield 66%;
IR: nmax (cmꢀ1) 2180 (NeCeS), 1634 (C]O); 1H NMR
(DMSO-d6): (d, ppm) 3.63 (s, 3H, OCH3), 7.20e7.79 (dd,
4H, AreH).
5.1.2. 2/4-Substituted benzoylisothiocyanates (7e10)
Substituted benzoic acid (0.1 mol) and thionyl chloride
(0.1 mol) were refluxed in benzene (50 ml) for one hour.
5.1.3. N-{[(6-Substituted-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}-2/4-substituted benzamides (11e30)
The final compounds 11e30 were obtained when 6-
substituted benzothiazoles 1e6 (0.02 mol) and substituted
benzoyl isothiocyanate 7e10 (0.02 mol) were refluxed in ben-
zene for 5 h, solid material obtained was filtered and recrystal-
lized from the benzene. The spectral data of 11e30 are given
below. The physico-chemical properties compounds are
presented in Table 1.
5.1.3.1. N-{[(6-Chloro-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}benzamide (11). IR: nmax (cmꢀ1) 3379, 3180 (NH),
3072 (CHeAr), 1673 (C]O), 1551 (C]N), 1090 (C]S),
1
803 (CeCl); H NMR (DMSO-d6): (d, ppm) 7.47e7.95 (m,
8H, AreH), 8.17 (s, 1H, NHC]O, D2O exchangeable),
12.98 (bs, 1H, NHC]S, D2O exchangeable).
5.1.3.2. N-{[(6-Fluoro-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}benzamide (12). IR: nmax (cmꢀ1) 3290, 3185 (NH),
3070 (CHeAr), 1674 (C]O), 1562 (C]N), 1321 (CeF)
Fig. 3. Section of liver. Group: 2, interference: normal hepatic parenchyma
with portal triad (PT), central vein (CV). Magnification: 100ꢃ.
1
1092 (C]S); H NMR (DMSO-d6): (d, ppm) 7.32e7.96 (m,

1120
A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
8H, AreH), 8.14 (s, 1H, NHC]O, D2O exchangeable), 12.80
(bs, 1H, NHC]S, D2O exchangeable); mass (EI) m/z: 331
(Mþ).
(m, 7H, AreH), 7.80 (s, 1H, NHC]O, D2O exchangeable),
12.71 (bs, 1H, NHC]S, D2O exchangeable).
5.1.3.10.
2-Chloro-N-{[(6-methoxy-1,3-benzothiazol-2-yl)-
5.1.3.3.
N-{[(6-Nitro-1,3-benzothiazol-2-yl)amino]carbono-
amino]carbonothioyl}benzamide (20). IR: nmax (cmꢀ1) 3363,
3190 (NH), 3040 (CHeAr), 2940 (CHeAliph), 1700
(C]O), 1460 (OCH3), 1578 (C]N), 1050 (C]S), 835 (Ce
Cl); 1H NMR(DMSO-d6): (d, ppm) 3.85 (s, 3H, OCH3),
7.06e7.69 (m, 7H, AreH), 7.78 (s, 1H, NHC]O, D2O ex-
changeable), 12.61 (bs, 1H, NHC]S, D2O exchangeable);
mass (EI) m/z: 377 (Mþ).
thioyl}benzamide (13). IR: nmax (cmꢀ1) 3223, 3174 (NH),
3083 (CHeAr), 1684 (C]O), 1556 (C]N), 1349 (Ce
1
NO2), 1128 (C]S), 806 (CeCl); H NMR (DMSO-d6): (d,
ppm) 7.36e8.33 (m, 8H, AreH), 9.09 (s, 1H, NHC]O,
D2O exchangeable), 13.27 (bs, 1H, NHC]S, D2O exchange-
able); mass (EI) m/z: 358 (Mþ).
5.1.3.4. N-{[(6-Methyl-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}benzamide (14). IR: nmax (cmꢀ1) 3357, 3185 (NH),
3056 (CHeAr), 2921 (CHeAliph), 1676 (C]O), 1509
(C]N), 1466 (CH3), 1090 (C]S); 1H NMR (DMSO-d6):
(d, ppm) 2.43 (s, 3H, CH3), 7.14e7.96 (m, 8H, AreH), 9.84
(s, 1H, NHC]O, D2O exchangeable),12.81 (bs, 1H,
NHC]S, D2O exchangeable); mass (EI) m/z: 327 (Mþ).
5.1.3.11. N-{[(6-Bromo-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}-4-chlorobenzamide (21). IR: nmax (cmꢀ1) 3390, 3210
(NH), 3075 (CHeAr) 1790 (C]O), 1555 (C]N), 1150
(C]S), 812 (CeCl), 563 (CeBr); 1H NMR (CDCl3): (d,
ppm) 7.36e8.16 (m, 7H, AreH), 8.42 (s, 1H, NHC]O,
D2O exchangeable), 12.73 (bs, 1H, NHC]S, D2O exchange-
able); mass (EI) m/z: 426 (Mþ).
5.1.3.5. N-{[(6-Methoxy-1,3-benzothiazol-2-yl)amino]carbo-
nothioyl}benzamide (15). IR: nmax (cmꢀ1) 3270, 3110 (NH),
3070 (CHeAr), 2930 (CHeAliph), 1670 (C]O), 1545
5.1.3.12. 4-Chloro-N-{[(6-chloro-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (22). IR: nmax (cmꢀ1) 3239, 3177
(NH), 3077 (CHeAr), 1673 (C]O), 1562 (C]N), 1055
1
1
(C]N), 1085 (C]S); H NMR (DMSO-d6): (d, ppm) 3.82
(C]S), 804 (CeCl); H NMR (DMSO-d6): (d, ppm) 7.37e
(s, 3H, OCH3), 7.05e7.96 (m, 8H, AreH), 8.13 (s, 1H,
NHC]O, D2O exchangeable), 12.81(bs, 1H, NHC]S, D2O
exchangeable); mass (EI) m/z: 343 (Mþ).
7.99 (m, 7H, AreH), 8.17 (s, 1H, NHC]O, D2O exchange-
able), 12.61 (bs, 1H, NHC]S, D2O exchangeable).
5.1.3.13. 4-Chloro-N-{[(6-fluoro-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (23). IR: nmax (cmꢀ1) 3285, 3172
(NH), 3072 (CHeAr), 1675 (C]O), 1562 (C]N), 1322
(CeF), 1095 (C]S), 802 (CeCl); 1H NMR (DMSO-d6):
(d,ppm) 7.28e8.17 (m, 7H, AreH), 8.36 (s, 1H, NHC]O,
D2O exchangeable), 12.81 (bs, 1H, NHC]S, D2O
exchangeable).
5.1.3.6. N-{[(6-Bromo-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}-2-chlorobenzamide (16). IR: nmax (cmꢀ1) 3290, 3280
(NH), 3062 (CHeAr), 1690 (C]O), 1583 (C]N), 1048
(C]S), 802 (CeCl), 545 (CeBr); 1H NMR (CDCl3): (d,
ppm) 7.32e7.87 (m, 7H, AreH), 9.19 (s, 1H, NHC]O,
D2O exchangeable), 11.78 (bs, 1H, NHC]S, D2O
exchangeable).
5.1.3.14. 4-Chloro-N-{[(6-methyl-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (24). IR: nmax (cmꢀ1) 3390, 3205
(NH), 3081 (CHeAr), 2950 (CHeAliph), 1675 (C]O), 1560
(C]N), 1094 (C]S), 806 (CeCl); 1H NMR (DMSO-d6):
(d, ppm) 2.27 (s, 3H, CH3), 7.36e7.80 (m, 7H, AreH), 8.19
(s, 1H, NHC]O, D2O exchangeable), 13.07 (bs, 1H,
NHC]S, D2O exchangeable).
5.1.3.7. 2-Chloro-N-{[(6-chloro-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (17). IR: nmax (cmꢀ1) 3310, 3298
(NH), 3060 (CHeAr), 1690 (C]O), 1599 (C]N), 1049
(C]S), 808 (CeCl); H NMR (DMSO-d6): (d, ppm) 7.46e
7.79 (m, 7H, AreH), 8.14 (s, 1H, NHC]O, D2O exchange-
able), 12.92 (bs, 1H, NHC]S, D2O exchangeable).
1
5.1.3.8. 2-Chloro-N-{[(6-fluoro-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (18). IR: nmax (cmꢀ1) 3249, 3135
(NH), 3100 (CHeAr), 1638 (C]O), 1548 (C]N), 1045
5.1.3.15. 4-Chloro-N-{[(6-methoxy-1,3-benzothiazol-2-yl)ami-
no]carbonothioyl}benzamide (25). IR: nmax (cmꢀ1) 3448,
3174 (NH), 3094 (CHeAr), 2932 (CHeAliph) 1675
1
1
(C]S), 801 (CeCl), 1300 (CeF); H NMR (DMSO-d6): (d,
(C]O), 1574 (C]N), 1091 (C]S), 805 (CeCl); H NMR
ppm) 7.30e7.81 (m, 7H, AreH), 7.97 (s, 1H, NHC]O,
D2O exchangeable), 13.02 (bs, 1H, NHC]S, D2O exchange-
able); mass (EI) m/z: 365 (Mþ).
(DMSO-d6): (d, ppm)3.82 (s, 3H, OCH3), 7.36e8.15
(m, 7H, AreH), 8.31 (s, 1H, NHC]O, D2O exchangeable),
12.90 (bs, 1H, NHC]S, D2O exchangeable).
5.1.3.9. 2-Chloro-N-{[(6-methyl-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (19). IR: nmax (cmꢀ1) 3300, 3170
(NH), 3045 (CHeAr), 2970 (CHeAliph), 1650 (C]O),
1556 (C]N), 1433 (CH3), 1126 (C]S), 815 (CeCl); 1H
NMR (DMSO-d6): (d, ppm) 2.45 (s, 3H, CH3), 7.15e7.68
5.1.3.16. N-{[(6-Chloro-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}-4-methoxybenzamide (26). IR: nmax (cmꢀ1) 3271, 3228
(NH), 3093 (CHeAr), 2932 (CHeAliph), 1634 (C]O), 1533
(C]N), 1049 (C]S), 812 (CeCl); 1H NMR (DMSO-d6):
(d, ppm) 3.38 (s, 3H, OCH3), 7.20e7.77 (m, 7H, AreH),

A. Rana et al. / European Journal of Medicinal Chemistry 43 (2008) 1114e1122
1121
8.34 (s, 1H, NHC]O, D2O exchangeable), 12.39 (bs, 1H,
NHC]S, D2O exchangeable).
Subcutaneous pentylenetetrazole seizure threshold test:
scPTZ was conducted by administering PTZ dissolved in
0.9% sodium chloride solution in the posterior midline of
the animals. A minimal time of 30 min subsequent to sc ad-
ministration of PTZ was used for seizure detection. Protection
was referred to as the failure to observe an episode of clonic
spasms of at least 5 s duration during this time period.
5.1.3.17. N-{[(6-Fluoro-1,3-benzothiazol-2-yl)amino]carbono-
thioyl}-4-methoxybenzamide (27). IR: nmax (cmꢀ1) 3271,
3185 (NH), 3069 (CHeAr), 2925 (CHeAliph), 1687
1
(C]O), 1561 (C]N), 1303 (CeF), 1058 (C]S); H NMR
(DMSO-d6): (d, ppm) 3.80 (s, 3H, OCH3), 7.09e7.58 (m,
7H, AreH), 9.65 (s, 1H, NHC]O, D2O exchangeable), 9.88
(bs, 1H, NHC]S, D2O exchangeable).
5.2.2. Neurotoxicity screening
The rotorod test was used to evaluate neurotoxicity. The an-
imal was placed on a 3.2 cm diameter knurled rod rotating at
6 rpm. Normal mice can remain on a rod rotating at this speed
indefinitely. Neurological toxicity is defined as the failure of
the animal to remain on the rod for 1 min. Results are ex-
pressed as number of animals exhibiting toxicity/number of
animals tested.
5.1.3.18. 4-Methoxy-N-{[(6-nitro-1,3-benzothiazol-2-yl)amino]-
carbonothioyl}benzamide (28). IR: nmax (cmꢀ1) 3411, 3287
(NH), 3099 (CHeAr), 2916 (CHeAliph), 1674 (C]O),
1
1561 (C]N), 1325 (NO2), 1125 (C]S); H NMR (DMSO-
d6): (d, ppm) 3.58(s, 3H, OCH3), 7.40e8.25 (m, 7H, AreH),
8.69 (s, 1H, NHC]O, D2O exchangeable), 12.46 (bs, 1H,
NHC]S, D2O exchangeable).
5.2.3. CNS depressant study
The forced swim pool method described earlier was fol-
lowed, Wistar rats were placed in a chamber (diameter:
45 cm, height: 20 cm) containing water up to a height of
15 cm at 25 ꢁ 2 ꢂC. Two swim sessions were conducted, an
initial attempt of 15 min pre-test, followed by 5 min test ses-
sion 24 h later. The animals were administered an i.p. injection
(30 mg/kg) of the test compounds 30 min before the test ses-
sion. The period of immobility (passive floating without strug-
gling, making only those movements which are necessary to
keep its head above the surface of water) during the 5 min
test period were measured.
5.1.3.19.
4-Methoxy-N-{[(6-methyl-1,3-benzothiazol-2-yl)-
amino]carbonothioyl}benzamide (29). IR: nmax (cmꢀ1) 3400,
3255 (NH), 3126 (CHeAr), 2943 (CHeAliph), 1674
1
(C]O), 1544 (C]N), 1464 (CH3), 1099 (C]S); H NMR
(DMSO-d6): (d, ppm) 2.30 (s, 3H, CH3), 3.44 (s, 3H,
OCH3), 6.99e7.43 (m, 7H, AreH), 9.81 (s, 1H, NHC]O,
D2O exchangeable), 12.79 (bs, 1H, NHC]S, D2O exchange-
able); mass (EI) m/z: 357 (Mþ).
5.1.3.20. 4-Methoxy-N-{[(6-methoxy-1,3-benzothiazol-2-yl)-
amino]carbonothioyl}benzamide (30). IR: nmax (cmꢀ1) 3391,
3298 (NH), 3103 (CHeAr), 2943 (CHeAliph), 1641
5.2.4. Assessment of liver function
1
(C]O), 1550 (C]N), 1054 (C]S); H NMR (DMSO-d6):
The animals were divided into groups of six, and the con-
trol group received a basal diet and vehicle. Other groups
were administered the test drug in a dose of 30 mg/kg/day
po (in methylcellulose) for two weeks. After the stipulated pe-
riod, each animal was anesthetized by anesthetic ether, and
blood was collected from the liver to assess the biochemical
parameters such as serum glutamate oxaloacetate transami-
nase (SGOT), serum glutamate pyruvate transaminase
(SGPT), alkaline phosphate, total protein and total albumin
were measured according to the reported methods.
(d, ppm) 3.73 (s, 6H, 2OCH3), 6.79e7.61 (m, 7H, AreH),
8.46 (s, 1H, NHC]O, D2O exchangeable), 12.40 (bs, 1H,
NHC]S, D2O exchangeable).
5.2. Pharmacology
The anticonvulsant evaluations were undertaken by the Na-
tional Institute of Health, using their reported procedures.
Male albino mice (CF-1 strain, 25e30 g) and male rats
(Sprague-dawley, 100e150 g) were used as experimental ani-
mals. The test compounds and standard drug were adminis-
tered intraperitoneally suspended in Tween 80 (1%) or in
0.5% methyl celluloseewater mixture.
5.2.5. Histopathological study of liver
The histopathological studies were carried out by a reported
method. The rats were scarified under light ether anesthesia af-
ter 24 h of last dosage, the livers were removed and washed
with normal saline. Small pieces of liver tissue were processed
and embedded in paraffin wax. Sections of 5e6 mm in thick-
ness were cut, stained with haematoxylin and eosin, and
then studied under an electron microscope.
5.2.1. Anticonvulsant screening
Initially all compounds were administered i.p. at doses of
30, 100, 300 mg/kg to one to four mice. Activity was estab-
lished using the MES, scPTZ tests.
Maximal electroshock test: maximal electroshock seizure
were elicited with a 60 cycle altering current of 50 mA inten-
sity delivered for 0.25 s via ear clip electrodes. The maximal
seizure typically consists of a short period of tonic extension
of the hind limbs and a final clonic episode. Abolition of the
hind limb tonic extensor component of the seizure is defined
as protection.
Acknowledgments
Authors are thankful to Dr. A. Mukherjee MD, Pathologist,
All India Institute of Medical Sciences (AIIMS), New Delhi
for the histopathological studies and to the University Grants

1122
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R. Kerphirique, S. Gontier, P. Hobert, J. Leblevec, M. Meunier,
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