Synthesis and anticonvulsant activity of some substituted 1,2,4-thiadiazoles

Article abstract of DOI:10.1016/j.ejmech.2008.06.015

A series of new substituted 1,2,4-thiadiazoles were synthesized by appropriate route and screened for anticonvulsant, neurotoxic and sedative-hypnotic activity. The structures of the synthesized compounds were confirmed by IR spectroscopy, ¹³C NMR and elemental (nitrogen and sulphur) analysis. After i.p. injection of the compounds to mice or rate at doses of 30, 100, and 300 mg/kg, body weights were examined in the maximal electroshock-induced seizures (MES) and subcutaneous pentylenetetrazole (scPTZ)-induced seizure models after 0.5 and 4 h. Rotorod method and phenobarbitone-induced hypnosis potentiation study were employed to examine neurotoxicity and sedative-hypnotic activity, respectively. All the compounds except 4g showed protection against MES screen after 0.5 h. Compounds 3a-c, 4a-c were active at 100 mg/kg dose i.p., whereas remaining compounds showed activity at 300 mg/kg. All 14 compounds except 3 g showed neurotoxicity at 100 and 300 mg/kg after 0.5 h. Compounds 3b and 4b showed NT after 4 h. Two compounds 3b and 4g showed significant (p < 0.05) percentage increase in sleeping time i.e. 67% and 59%, respectively. It may be concluded that the synthesized compounds were potent against MES-induced seizures than ScPTZ induced and showed low potency as sedative-hypnotic agent which is advantageous.

Full text of DOI:10.1016/j.ejmech.2008.06.015

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European Journal of Medicinal Chemistry 44 (2009) 1100e1105
http://www.elsevier.com/locate/ejmech
Original article
Synthesis and anticonvulsant activity
of some substituted 1,2,4-thiadiazoles
Arun Gupta a, Pradeep Mishra a, S.N. Pandeya b, Sushil K. Kashaw a,
Varsha Kashaw a, James P. Stables c
*
,
a Pharmaceutical Chemisrty Division, Department of Pharmaceutical Sciences, Dr. H.S. Gour University, Sagar, India
b Institute of Pharmacy, V.B.S. Purvanchal University, Uttar Pradesh, India
c Preclinical Pharmacology Section, Epilepsy Branch, National Institutes of Health, Bethesda 20892-9020, USA
Received 30 January 2008; received in revised form 17 June 2008; accepted 20 June 2008
Available online 26 June 2008
Abstract
A series of new substituted 1,2,4-thiadiazoles were synthesized by appropriate route and screened for anticonvulsant, neurotoxic and seda-
tive-hypnotic activity. The structures of the synthesized compounds were confirmed by IR spectroscopy, 13C NMR and elemental (nitrogen and
sulphur) analysis. After i.p. injection of the compounds to mice or rate at doses of 30, 100, and 300 mg/kg, body weights were examined in the
maximal electroshock-induced seizures (MES) and subcutaneous pentylenetetrazole (scPTZ)-induced seizure models after 0.5 and 4 h. Rotorod
method and phenobarbitone-induced hypnosis potentiation study were employed to examine neurotoxicity and sedative-hypnotic activity,
respectively. All the compounds except 4g showed protection against MES screen after 0.5 h. Compounds 3aec, 4aec were active at
100 mg/kg dose i.p., whereas remaining compounds showed activity at 300 mg/kg. All 14 compounds except 3 g showed neurotoxicity at
100 and 300 mg/kg after 0.5 h. Compounds 3b and 4b showed NT after 4 h. Two compounds 3b and 4g showed significant ( p < 0.05) percentage
increase in sleeping time i.e. 67% and 59%, respectively. It may be concluded that the synthesized compounds were potent against MES-induced
seizures than ScPTZ induced and showed low potency as sedative-hypnotic agent which is advantageous.
Ó 2008 Elsevier Masson SAS. All rights reserved.
Keywords: 1,2,4-Thiadiazole; Anticonvulsant activity; Maximal electroshock-induced seizures; Subcutaneous pentylenetetrazole-induced seizure; Neurotoxicity
1. Introduction
the available antiepileptic drug is ideal as these can be associated
withchronicandtoxicsideeffects[4]. Thusthesearchforthenew
anticonvulsant drugscontinuetobeanactiveareaofinvestigation
in medicinal chemistry. The MES test is a proven method of
generalized toniceclonic seizures and identifies clinical candi-
dates that prevent seizure spread [5,6]. At present scientists are
working at various nucleus to explore suitable pharmacophore
responsible for anticonvulsant potential. As a part of our ongoing
research program [7,8] to find novel anticonvulsants, herein we
describe synthesis, anticonvulsant, neurotoxicity and sedative-
hypnotic activity of some new 1,2,4-thiadiazoles.
Epilepsy is one of the most common diseases of the brain
affectingatleast50millionpersonsworldwide. Thereiscurrently
a need for improved agents for the treatment of seizures, since
currently available drugs are effective only in 60e80% of epilep-
tic patients [1]. Polytherapy with antiepileptic drugs (AEDs) is
necessary in clinical practice because of the limited efficacy of
monotherapy [2,3]. In recent years many new chemical entities
(NCEs) have been designed that were structurally dissimilar
from many common anticonvulsant containing dicarboximide
function (CONRCO), which contributes to toxic side effects.
Duringthe pastdecadeseveralnew drugs wereapproved(Rufina-
mide, Retigabine, Pregabaline, Remacemide). However, none of
2. Chemistry
The synthesis of 1,2,4-thiadiazoline was accomplished as
showed in Fig. 1. Aryl thiourea (1) was prepared by reacting
* Corresponding author. Tel.: þ91 7582240757; fax: þ91 7582265457.
E-mail address: sushilkashaw@gmail.com (S.K. Kashaw).
0223-5234/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.06.015

A. Gupta et al. / European Journal of Medicinal Chemistry 44 (2009) 1100e1105
1101
S
Heat
Ar-NH2 + NH4SCN + HCl
Ar-NH-C-NH2
(1)
H2O2
Ar
N
Ar
N
N
NaOH
HCl
Ar
N
H
NH
Ar
N
H
O
Ar
N
H
N
H
Ar
N
S
(2)
N
S
(3)
N
S
(4)
(3a-3g)
(4a-4g)
N
N
F
N
c
e
g
a
N
OC2H5
N
f
d
b
Fig. 1. Scheme for the synthesis of title compounds.
aromatic/hetero aromatic amines with ammonium thiocyanate
in the presence of concentrated hydrochloric acid. Compounds
1 were oxidatively cyclized into 3-aryl amino-4-aryl-5-imino-
D2-1,2,4-thiadiazoline (2). Compounds 2 on refluxing with
concentrated hydrochloric acid gave 3-aryl amino-4-aryl-5-
oxo-D2-1,2,4-thiadiazoline (3aeg). Compounds 2 isomerised
to 3,5-diarylamino-1,2,4-thiadiazole in the presence of sodium
hydroxide solution (4aeg).
kg after 0.5 h. NT level of reference drugs phenytoin and
chlorpromazine is 100 mg/kg at 0.5 h. Only 3b and 4b showed
pronged NT level (after 4 h). Some of the synthesized com-
pounds were challenged in phenobarbitone-induced hypnosis
potentiation test to see their effect on general behavior. Com-
pounds 3b and 4g showed significant percentage increase in
sleep level i.e. 67 and 59.5%, respectively. Percentage increase
in sleep by remaining compounds was insignificant. Above re-
sults and discussion indicates that the new compounds are
more selective towards MES screen than ScPTZ screen. Pres-
ent research will guide our future development of potent and
selective anticonvulsant drugs.
3. Pharmacology
Initial anticonvulsant evaluation was undertaken by the an-
ticonvulsant drug development (ADD) program protocol. Male
albino mice (CF-1 strain, 18e25 g) and male albino rats (Spra-
gueeDawley, 100e150 g) were used as experimental animals.
Four animals constitute one group. The compounds were sus-
pended in 0.5% methyl celluloseewater mixture or in polyeth-
ylene glycol (PEG).
Table 1
Physical constants of the synthesized 3-arylamino-4-aryl-5-oxo-D2-1,2,4-thia-
diazolines
Ar
N
NH
O
Ar
N
S
4. Results and discussions
Code M.p. Yield Molecular
(ꢀC) (%)
Molecular Element
weight
formula
(found %/calculated %)a
All synthesized compounds except 4g showed protection
against MES seizures after 0.5 h of drug administration. Com-
pounds 3aec and 4aec were active at 100 mg/kg, whereas re-
maining compounds showed activity at 300 mg/kg. These
synthesized compounds exhibited their ability to diminish
the magnitude of toniceclonic seizures. Only six compounds
namely 3a,b and 4aec exhibited prolonged duration of action
at 300 mg/kg after 4 h. Mentioned five compounds showed
rapid and long duration of action. Only one out 14 compounds
(3b) showed protection against ScPTZ. All other compounds
were inactive in ScPTZ screen which indicates their ineffec-
tiveness against absence seizures. None of the compounds
were active after 4 h in ScPTZ screen. NT screening revealed
that all 14 compounds except 3g showed NT at 100e300 mg/
3a
3b
3c
3d
3e
3f
3g
a
170 71
C22H15N3SO
369.45
N (11.40/11.39)
S (8.67/8.66)
200 68
210 74
190 60
184 69
180 79
174 43
C18H19N3SO3 357.43
C14H9N3SOF2 305.11
N (11.73/11.75)
S (8.91/8.95)
N (13.83/13.75)
S (10.53/10.48)
N (25.85/25.80)
S (11.80/11.79)
N (25.75/25.80)
S (11.75/11.79)
N (25.85/25.80)
S (11.75/11.79)
N (35.86/35.70)
S (11.70/11.80)
C12H9N5SO
C12H9N5SO
C12H9N5SO
C10H7N7SO
271.3
271.3
271.3
273.28
Elemental analyses for N and S were within 0.4% of the theoretical values.

1102
A. Gupta et al. / European Journal of Medicinal Chemistry 44 (2009) 1100e1105
Table 2
IR and 13C NMR spectroscopic data of the title compounds
Code
Structure
IR
13C NMR
4"
5"
9"
3"
2"
6"
7"
10"
163 (C-3), 165 (C-5), 140.8 (C-10), 109.4
(C-20, C-200), 126 (C-30, C-60, C-300, C-600),
119 (C-40, C-400), 128.6 (C-50, C-500), 125
(C-70, C-70), 121 (C-80, C-800), 134.3 (C-90,
C-900), 124.7 (C-100, C-1000)
1"
8"
O
3'
2'
N
1'
3366 (NH), 1722 (C]O),
1261 (CN), 1630 (C]C)
4
3
4'
NH
5
3a
2
1
10'
9'
5'
N
S
8'
6'
7'
a"
O
4"
b"
5"
6"
3"
2"
163 (C-3), 165 (C-50), 136 (C-10), 132.9
(C-100), 116.9 (C-20, C-60, C-200, C-600), 115.2
(C-30, C-50, C-300, C-500), 147.5 (C-40, C-400),
14.8 (Ca0, Ca00), 64 (C-b0, C-b00)
3432 (NH), 1713 (C]O),
1310 (CN), 683 (CS), 1624
(C]C)
3b
1"
5' 6'
N
a'
1'
4
3
O
NH
5
O
4'
b'
2
1
N
S
2'
3'
F
4"
5"
6"
3"
2"
1630(C-3), 165 (C-5), 140 (C-1), 1170.9 (C-20,
C-6 ), 116.3 (C-30, C-50), 152.9 (C-4 ), 128.4
(C-100), 123.2 (C-200, C-600), 115.7 (C-300, C-500),
158.5 (C-400)
3424 (NH), 1711 (C]O),
1318 (CN), 733 (CS), 1643
(C]C), 1167 (CF)
1"
3c
5'
6'
N
4
3
NH
O
F
5
4'
1'
2
1
N
S
2'
3'
5"
6"
4"
3"
N
1"
3412 (NH), 1719 (C]O),
1421 (C]N), 763 (CS), 1310
(CN), 1623 (C]C)
163 (C-3), 165 (C-5), 158.6 (C-20), 147.7
(C-200), 109.9 (C-30, C-300), 138.3 (C-40C-400),
113.3 (C-5,C-500), 148.2 (C-60,C-600)
2"
6'
N
3d
1'
2'
N
4
3
O
NH
5
5'
2
1
N
S
4' 3'
6"
N
5"
4"
1"
2"
3428(NH), 1709 (C]O),
1421 (C]N), 1316 (CN),721
(CS),1624 (C]C)
163 (C-3), 165 (C-5), 137.6 (C-20, C-200),
134.4 (C-30), 145.1 (C-300), 122.8 (C-40,C-400),
124.7 (C-50,C-500), 138.9 (C-60,C-600)
3"
1' 2'
N
3e
N
3'
4
3
NH
O
5
6'
2
1
N
S
5'
4'
1"
4"
N
6"
5"
2"
3"
3446 (NH), 1732 (C]O),
1416 (C]N), 1302 (CN),
732 (CS), 1653 (C]C)
163 (C-3), 165 (C-5), 150.3 (C-20, C-60, C-200,
C-600), 109.1 (C-30, C-50, C-300, C-500), 155.3
(C-40, C-400)
2' 3'
6' 5'
3f
N
4
2
1'
N
4'
O
3
NH
5
1
N
S

A. Gupta et al. / European Journal of Medicinal Chemistry 44 (2009) 1100e1105
1103
Table 2 (continued)
Code
Structure
IR
13C NMR
5"
4"
3"
6"
3426 (NH), 1744 (C]O),
1402 (C]N), 1284 (CN),
693 (CS), 1614 (C]C)
163 (C-3), 165 (C-5), 158 (C-20), 169.3 (C-200),
157.9 (C-40C-400), 110.3 (C-50,C-500), 157.9
(C-60,C-600)
N
N
1"
3g
2"
6'
N
N
1'
2'
N
4
3
O
NH
5
5
2
1
N
S
4' 3'
5. Experimental protocol
and the mixture was heated. The mixture was poured on
crushed ice, the precipitate thus obtained was filtered off by
suction and recrystallized from ethanol. Spectral analysis,
m.p. and elemental analysis confirm the formation of the cor-
responding thiourea.
5.1. Chemistry
Melting points were determined in open capillary tubes in
a Thomas Hoover melting point apparatus and are uncor-
rected. The purity of the compounds was confirmed by thin
layer chromatography using silica gel glass plates and solvent
system of benzene:ethanol (9:1). The spots were developed in
iodine chamber and visualized under ultraviolet lamp. Infrared
(IR) and 13C nuclear magnetic resonance (13C NMR) spectra
were recorded for the compounds in Testcan Shimadzu FTIR
8000 (KBr) 13C Avance Bruker 300 MHz spectrophotometer,
respectively. Chemical shifts are reported in parts per million
(ppm) using tetramethylsilane (TMS) as an internal standard.
Elemental analysis (N and S) was undertaken with Perkin El-
mer-2400 instrument and the measured values agreed within
ꢁ0.4% with the calculated. Mass spectra were obtained on
Joel SX 102/M-6000 mass spectrometer applying FAB
method.
5.1.2. Synthesis of 3-amino-4-aryl-5-imino-D2-1,2,4-
thiadiazoline (2aeg)9
Corresponding aryl thiourea (1aeg, 0.5 mol) was taken in
a conical flask equipped with separating funnel and condenser
and was dissolved in a warm 10 mL of HCl. Hydrogen perox-
ide (60e70 mL) was added dropwise from the separating fun-
nel with continuous stirring and the mixture was kept aside for
2 h. The oxidized mixture was then diluted with water and was
neutralized with dilute ammonia. The precipitate thus obtained
was collected and recrystallized from ethanol (95%).
5.1.3. Synthesis of 3-aryl amino-4-aryl-5-oxo-D2-1,2,4-
thiadiazolines (3aeg)
Required compound 2 (0.0l mol) was taken in a round-bot-
tomed flask. After adding 10 mL of hydrochloric acid in
25 mL of ethanol, they were refluxed for an hour on water
bath. On cooling the product, it was filtered and recrystallized
from ethanol. Physico-chemical and spectral data of the com-
pounds are presented in Tables 1 and 2, respectively.
5.1.1. Synthesis of aryl thiourea (1)
Aryl amine (0.1 mol) was taken in 250 mL of beaker con-
taining 100 mL distilled water. HCl (10 mL) was added and
the contents were warmed to dissolve the amine. Ammonium
thiocyanate (7.6 g, 0.1 mol) was added to the amine solution
Table 3
Physical constants of the synthesized 3,5-diarylamino-1,2,4-thiadiazoles
N
Ar
Ar
NH
NH
N
S
Code
M.p. (ꢀC)
178
% Yield
68
Molecular formula
C22H16N4S
Molecular weight
368.48
Element (found %/calculated %)a
4a
N (15.21/15.19)
S (8.73/8.59)
4b
4c
4d
4e
4f
4g
a
145
150
240
248
270
243
51
73
54
68
85
59
C18H20N4SO2
C14H10N4SF2
C12H10N6S
C12H10N6S
C12H10N6S
C10H8N8S
356.4
N (15.74/15.71)
S (8.89/8.85)
304.21
270.32
270.32
270.32
272.29
N (18.43/18.40)
S (10.67/10.51)
N (31.20/31.07)
S (11.73/11.83)
N (31.28/31.07)
S (11.73/11.83)
N (31.20/31.07)
S (11.73/11.83)
N (41.09/41.13)
S (11.70/11.75)
Elemental analyses for N and S were within 0.4% of the theoretical values.

1104
A. Gupta et al. / European Journal of Medicinal Chemistry 44 (2009) 1100e1105
Table 4
IR and 13C NMR spectroscopic data of the titled compounds
Code
Structure
IR
13C NMR
6' 7'
5'
8'
10'
163 (C-3, C-5), 142 (C-10, C-100),
121 (C-20, C-200), 125 (C-30, C-300),
2" 3"
9'
N
1"
4
3410 (NH), 1243
(CN), 731 (CS), 1619
(C]C)
3
NH
5
0
126 (C-4 , C-400), 128.6 (C-50, C-500),
NH
4"
4'
1'
2
1
4a
119 (C-60, C-600), 126.6 (C-70, C-700),
109.4 (C-8, C-800), 134.3 (C-9, C-
900), 124.7 (C-100, C-1000)
N
S
10"
8"
9"
2'
3'
5"
7" 6"
2"
3"
5' 6'
163 (C-3, C-5), 134.7 (C-10, C-100),
116.9 (C-20, C-60, C-200, C-600),
115.2 (C-30, C-50, C-300, C-500), 147.5
(C-40, C-400), 64.7 (C-a0, C-a00), 14.8
(C-b0, C-b00)
a'
a"
b"
N
4
3398 (NH), 1296
(CN), 743 (CS), 1627
(C]C), 1441 (CeO)
3
NH
O
b'
O
NH
NH
5
1
1"
4'
1'
4"
4b
2
N
S
6" 5"
2'
1'
3'
2" 3"
5' 6'
N
163 (C-3, C-5), 138.7 (C-10, C-100),
117.9 (C-20, C-60, C-200, C-600),
116.3 (C-30, C-50, C-300, C-500),
152.2 (C-40, C-400)
4
3409 (NH), 1293
(CN), 738 (CS), 1611
(C]C), 1153 (CF)
NH
F
F
4'
3
5
1"
4"
2
N
1
4c
4d
4e
S
5"
6"
2'
3'
3" 4"
N
3'
6'
5'
1'
2'
N
4
3427 (NH), 1477
(C]N), 1239 (CN),
723 (CS), 1624
(C]C)
163 (C-3, C-5), 154.7 (C-20, C-200),
109.9 (C-30, C-300), 138.3 (C-C-40,
C400), 113.3 (C-50, C-500), 148.2
(C-60, C-600)
2"
1"
5"
3
NH
NH
NH
NH
NH
NH
NH
NH
5
2
1
N
6"
N
S
4'
4" 5"
1' 2'
N
6'
N
4
3418 (NH), 1456
(C]N), 1326 (CN),
742 (CS), 1635
(C]C)
163 (C-3, C-5), 137.6 (C-20, C-200),
134.1 (C-30, C-300), 122.8 (C-40,
C-400), 124.7 (C-50, C-500), 138.9
(C-60, C-600)
3"
6"
3'
3
5
2
1
N
N
S
2" 1"
5'
4'
2'
3'
6"
5"
3"
N
4
1'
N
3418 (NH), 1456
(C]N), 1326 (CN),
742 (CS), 1635
(C]C)
4'
1630(0 C-3, C-5), 150.3 (C-20, C-60,
C-2 , C-600), 109.1 (C-30, C-50,
C-300, C-500), 155.3 (C-40, C-400)
3
N
1"
5
4"
2
1
4f
N
S
2"
6'
6'
5'
3425 (NH), 1412
(C]N), 1298 (CN),
725 (CS), 1626
(C]C)
4"
5"
N
N
163 (C-3, C-5), 163.3 (C-20, C-200),
157.9 (C-40, C-60, C-400, C-600),
110.3 (C-50, C-500)
N
3"
2"
1'
2'
N
4
4g
3
5
5'
4'
2
1
N
3'
S
N
1" 6"
5.1.4. Synthesis of 5-diaryl amino-1,2,4-thiadiazole (4aeg)
5.2. Pharmacology
Compound 2 (0.01 mol) was dissolved in warm ethanol
(20 mL). Aqueous sodium hydroxide (10%, 10 mL) was added
and mixed. The contents were refluxed for 4 h on water bath. On
cooling the crystal of the product separated out. The crystals
were filtered and recrystallized from ethanol. The melting point
was determined. Physico-chemical and spectral data of the
compounds are presented in Tables 3 and 4, respectively.
5.2.1. Anticonvulsant screening
In the preliminary screening, each compound was adminis-
tered an i.p. injection at three dose levels (30, 100 and 300 mg/
kg) and the anticonvulsant activity was assessed after 30 min
and 4 h interval of administrations [10,11]. The anticonvulsant
efficacy was evaluated by the maximal electroshock-induced

A. Gupta et al. / European Journal of Medicinal Chemistry 44 (2009) 1100e1105
1105
Table 5
Anticonvulsant activity and minimal motor impairment of substituted-1,2,4-
thiadiazoline
Table 6
Phenobarbitone-induced hypnosis potentiation test for some selected
compounds
Code
Intraperitoneal injection in micea
Code
Duration of sleep
in minutes ꢁ SEM
% Increase
in sleep
MES
ScPTZ
Neurotoxicity
screen
3a
3b
92 ꢁ 4.80
132 ꢁ 5.30***
93 ꢁ 6.40
16.4
67.1
17.7
22.7
36.7
59.5
e
0.5 h
4 h
0.5 h
4 h
0.5 h
4 h
3c
4a
3a
3b
100
100
100
300
300
300
300
100
100
100
300
300
300
e
300
300
e
e
300
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
300
300
300
100
100
100
e
e
300
e
97 ꢁ 6.20*
108 ꢁ 4.20**
126 ꢁ 6.50***
79 ꢁ 4.87
4b
4g
3c
3d
e
e
e
PBS # only (control)
3e
e
e
e
e
e
e
PBS #: Phenobarbitone sodium.
*p < 0.5; **p < 0.01; ***p < 0.005.
3f
3g
e
e
e
4a
4b
300
300
300
e
e
300
300
100
100
100
100
300
100
100
e
e
e
300
e
Thirty minutes after drug administration, animals were in-
jected phenobarbitone sodium. The animals fell asleep on their
back. Sleeping time of each rat was taken as the interval be-
tween the loss and return of the righting reflex as indicated
by inability or ability, respectively, of the rat to right itself
in three successive trials when placed on its back. The time
taken by animals to awake was noted. The results are reported
in Table 6. A control was also performed after pre-treatment
with test substance vehicle.
4c
4d
e
e
4e
e
e
e
e
e
e
4f
4g
e
e
e
Phenytoin
Carbamazepine
30
30
100
e
e
100
100
300
a
Doses of 30, 100 and 300 mg/kg were administered. The figures in the ta-
ble indicate the minimum dose whereby bioactivity was demonstrated in half
or more of the mice. The animals (number of animals ¼ 4) were examined 0.5
and 4 h after injections were made. The dash (e) indicates an absence of ac-
tivity at maximum dose administered (300 mg/kg).
Acknowledgement
The authors deeply appreciate the assistance of the Antiep-
ileptic Drug Development Program, Epilepsy Branch, Preclin-
ical Pharmacology Section, NIH, USA.
seizure (MES) and subcutaneous pentylenetetrazole (scPTZ).
The data are presented in Table 5.
5.2.2. Neurotoxicity screen
References
The test is used to evaluate the activity of drugs interfering
with motor coordination. The skeletal muscle relaxation in-
duced by a test compound could be evaluated by testing the
ability of mice or rats to remain on a revolving rod. Minimal
motor impairment was measured in mice by the rotorod test.
The mice were trained to stay on an accelerating rotorod
that rotates at 10 revolutions/min. The rod diameter was
3.2 cm. Trained animals were given i.p. injection of the test
compounds in doses of 30, 100 and 300 mg/kg. Neurotoxicity
was indicated by the inability of the animal to maintain equi-
librium on the rod for at least 1 min in each of the three trials.
[1] T.R. Brown, G.L. Holmes, N. Engl. J. Med. 344 (2001) 1145e1151.
[2] A. Nicolson, J.P. Leach, CNS Drugs 15 (2001) 955e968.
[3] S.J. Czuczwar, K. Przemycki, Pol. J. Pharmacol. 53 (2001) 65e68.
[4] C.L. Deckers, P. Genton, G.J. Sills, D. Schmidt, Epilepsy Res. 53 (2003)
1e17.
[5] J.P. Stables, H.J. Kupferberg, in: G. Avanzini, P. Tanganelli, M. Avoli
(Eds.), Molecular and Cellular Targets for Antiepileptic Drugs, John Li-
bey, London, 1997, p. 191.
[6] H.S. White, J.H. Woodhead, M.R. Franklin, in: R.H. Levy, R.H. Mattson,
B.S. Meldrum (Eds.), Antiepileptic Drugs, fourth ed. Raven, New York,
1995, p. 99.
[7] N. Aggarwal, P. Mishra, J. Pharm. Pharm. Sci. 7 (2004) 260e264.
[8] N. Aggarwal, P. Mishra, J. Zhejiang Univ. 6B (2005) 617e621.
[10] R.L. Krall, J.K. Penry, B.G. White, H.J. Kupferberg, E.A. Swinyard,
Epilepsia 19 (1978) 409e428.
5.2.3. Phenobarbitone-induced hypnosis potentiation test
The drug solution in polyethylene glycol (PEG 100) was
administered in a dose of 100 mg/kg to a group of six animals.
The animals were grouped into groups of six animals each.
[11] R.J. Porter, J.J. Cereghino, G.D. Gladding, B.J. Hessie, H.J. Kupferberg,
B. Scoville, B.G. White, Cleve. Clin. Q. 51 (1984) 293e305.