Indian Journal of Pharmacology 2002; 34: 251-255 SHORT COMMUNICATION

ANTICONVULSANT ACTIVITY OF G. GLABRA

ANTICONVULSANT ACTIVITY OF ROOTS AND RHIZOMES OF GLYCYRRHIZA

GLABRA

SHIRISH D. AMBAWADE, VEENA S. KASTURE, SANJAY B. KASTURE

Natural Products Laboratory, Division of Pharmacology, N.D.M.V.P. Samaj's College of Pharmacy,

Nashik - 422 002.

Manuscript Received: 14.6.2001 Revised: 8.1.2002 Accepted: 4.3.2002

ABSTRACT Objective: To study the anticonvulsant activity of ethanolic extract of Glycyrrhiza glabra in albino rats
and mice.
Methods: The anticonvulsant activity of ethanolic extract of roots and rhizomes of Glycyrrhiza glabra
(10, 30, 100 and 500 mg/kg, i.p.) in mice was assessed using maximum electroshock seizure (MES) test
and pentylenetetrazol (PTZ) using albino mice. The lithium-pilocarpine model of status epilepticus was
also used to assess the anticonvulsant activity in rats.
Results: The ethanolic extract of G. glabra did not reduce the duration of tonic hindleg extension in the
MES test even in the dose of 500 mg/kg. However, the extract significantly and dose-dependently delayed
the onset of clonic convulsions induced by pentylenetetrazol. The dose of 100 mg/kg afforded protection
to all animals. The extract also protected rats against seizures induced by lithium-pilocarpine.
Conclusion: The ethanolic extract of G. glabra inhibits PTZ and lithium-pilocarpine-induced convulsions
but not MES-induced convulsions.

KEY WORDS Anticonvulsant glycyrrhiza glabra lithium-pilocarpine MES PTZ

INTRODUCTION occurs when glutamatergic excitatory neurotrans-

In the traditional system of medicine, the roots and
rhizomes of Glycyrrhiza glabra (family: Leguminosae)
have been in clinical use for centuries. Roots have
demulcent, antacid, anti-ulcer1, anti-inflammatory,
expectorant, tonic, diuretic, laxative, and sedative
properties2. They also possess antipyretic3, antimi-
crobial, antiherpes 4, and anxiolytic 5 activities.
Glycyrrhizin, a triterpene saponin, possesses antivi-
ral activity 6. In the traditional system of medicine G.
glabra also known as liquorice is recommended for
the treatment of epilepsy 7.
Attempts to find out a common neurochemical basis
for human or experimental epilepsy have been dis-
appointing. An imbalance between the excitatory and
inhibitory neurotransmitters is responsible for
seizures 8,9. At neuronal level, seizure activity often
mitters overrides gamma-aminobutyric acid (GABA)
mediated inhibition 10. Several animal models of con-
vulsions have been developed to evaluate anti-sei-
zure activity. Many drugs that increase the brain con-
tents of GABA have exhibited anticonvulsant activity
against seizures induced by maximum electroshock
(MES), pentylenetetrazol (PTZ) and lithium-pilo-
carpine (Li-Pilo). The MES is probably the best-vali-
dated method for assessment of anti-epileptic drugs
in generalized tonic-clonic seizures 11,12. The PTZ-
induced seizures are similar to the symptoms ob-
served in the absence seizures and drugs useful in
treatment of absence seizures suppress PTZ induced
seizures8. The status epilepticus induced by lithium-
pilocarpine increases brain contents of acetylcho-
line13. The reproducibility of the responses to this
treatment makes this a very useful model to investi-
gate various facets of seizures 14.

Correspondence: S.B. Kasture

e-mail: kasture_sb@hotmail.com
252 S.D. AMBAWADE et al.

The objective of the present study was to investigate Table 1: Effect of ethanolic extract of Glycyrrhiza glabra (EE)
anticonvulsant activity of ethanolic extract of G. glabra on pentylenetetrazol-induced seizures in mice.
(EE) against the seizures induced by MES, PTZ and
combination of lithium sulphate and pilocarpine nitrate. Treatment Onset of clonic convulsions Incidence of
(mg/kg) in sec (mean±SEM) convulsions (%)
MATERIALS AND METHODS
Vehicle 140.3 ± 13.8 100
Preparation of extract: The roots and rhizomes of EE 10 664.5 ± 93.0* 66.6
Glycyrrhiza glabra were purchased from commer-
cial source. The roots and rhizomes (1.0 kg) were 30 780.0 16.6
crushed to a coarse powder and extracted with 100 A 0
ethanol (70% v/v) using Soxhlet's extractor for 24 h.
Diazepam 2 A 0
The extract was concentrated under reduced pres-
sure and then dried in air (yield -170 g). The extract F 3.9
was stored in a refrigerator and reconstituted in wa- d.f. 2,15
ter for injection just before use.
P <0.001
Animals: Male Albino Swiss mice weighing 22-25 g
and male Albino Wistar rats weighing 125-150 g were n = 6 in each group. *P<0.001 when compared to the vehicle
obtained from National Institute of Toxicology, Pune. treated group (ANOVA followed by Dunnett's test).
Animals were housed in groups of 6-8 per cage at a
A = Absence of convulsions.
temperature of 25o ±1oC and relative humidity of
45-55%. A 12:12 dark:light cycle was followed
during the experiments and the experiments were Pentylenetetrazol-induced seizures: The EE was
carried out during 1200-1400 h. Animals had free administered i.p. in varying doses (10-100 mg/kg)
access to food and water however, food but not water 30 min before the subcutaneous injection of PTZ (80
was withdrawn 8 h before and during the experiments. mg/kg) and mice were observed for onset of
The Institutional Animal Ethics Committee approved myoclonic spasm and clonic convulsions 15. One
the protocol of the study. group received vehicle while other group received
Drugs: Pentylenetetrazol (Sigma, USA), lithium sul- diazepam (2.0 mg/kg, i.p.) as a reference standard.
phate (Glenmark Pharmaceuticals, India), The animals were observed for onset of convulsion
Pilocarpine nitrate (FDC Limited, India), diazepam upto 30 min after PTZ. Each group contained six
(Calmpose inj. Ranbaxy, India), were used in this animals.
study. The drugs were dissolved in water for injec- Lithium-pilocarpine-induced status epilepticus:
tion and administered in a volume of 5 ml/kg to both Albino rats were divided randomly into five groups
rats and mice. each containing six animals. Status epilepticus was
Acute toxicity: The ethanolic extract was adminis- induced by administration of pilocarpine (30 mg/kg,
tered in doses of 600 and 1000 mg/kg, i.p., to groups i.p.) 24 hr after lithium sulphate (3 mEq/kg, i.p.). The
of mice, each containing ten animals and mortality effect of EE (10, 30 and 100 mg/kg, 30 min before
was observed after 24 h. injection of pilocarpine nitrate) was studied on the
severity of status epilepticus. One group received only
Assessment of anticonvulsant activity vehicle while the other group received diazepam (1.0
mg/kg). The severity of status epilepticus was ob-
Maximum electroshock-induced seizures: The EE served every 15 min till 90 min and thereafter every
was administered to groups of mice (n = 6) in doses 30 min till 180 min, using the scoring system de-
ranging from 10-500 mg/kg, i.p. 30 min before appli- scribed earlier16: No response-stage 0, fictive scratch-
cation of electric shock (42 mA, 0.2 sec) using corneal ing-stage 1, tremors-stage 2, head nodding-3,
electrodes15. The duration of tonic hindleg extension forelimb clonus-stage 4, rearing and falling back-
was noted. stage 5.
 ANTICONVULSANT ACTIVITY OF G. GLABRA 253

Table 2: Effect of ethanolic extract of Glycyrrhiza glabra (EE) on lithium-pilocarpine-induced status epilepticus in rats.

Time after Treatment

Pilocarpine EE in mg/kg, i.p.
in min.
Vehicle 10 30 100 Diazepam H P
(1 mg/kg)

0 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 - -
15 1.5 ± 0.2 1.0 ± 0.4 0.3 ± 0.2* 0.8 ± 0.4 0.5 ± 0.2* 7.88 0.01
30 3.0 ± 0.0 2.8 ± 0.4 1.5 ± 0.5 1.0 ± 0.4* 0.7 ± 0.2* 17.7 0.001
45 3.0 ± 0.0 2.3 ± 0.4 2.1 ± 0.5 1.5 ± 0.4* 0.8 ± 0.1* 15.7 0.003
60 3.3 ± 0.2 1.5 ± 0.5* 1.0 ± 0.4* 1.6 ± 0.4 1.1 ± 0.3* 13.1 0.01
75 4.0 ± 0.0 0.5 ± 0.2* 0.1 ± 0.1* 0.6 ± 0.2* 1.4 ± 0.4 19.6 0.0001
90 4.3 ± 0.2 0.5 ± 0.2* 0.5 ± 0.2* 0.8 ± 0.3* 1.3 ± 0.2 19.1 0.0001
120 4.6 ± 0.2 0.3 ± 0.2* 0.6 ± 0.5* 1.0 ± 0.3* 0.8 ± 0.4* 16.3 0.002
150 1.6 ± 0.4 0.0 ± 0.0* 0.3 ± 0.2* 0.5 ± 0.2 1.0 ± 0.0 18.4 0.001
180 0.5 ± 0.2 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 0.5 ± 0.2 10.8 0.02

n = 6 in each group. H and P are values of Kruskal Wallis ANOVA and Probability respectively (df = 4). The values are mean ± SEM
of the scores indicating the severity of seizures.
*P<0.05 as compared to vehicle treated group (Dunn's test).

Statistical analysis: The data are presented as
mean±SEM. The data of PTZ test were analyzed by
one-way analysis of variance (ANOVA) followed by
Dunnett's test and the data of lithium-pilocarpine- in-
duced seizures were analyzed using Kruskal- Wallis
ANOVA followed by Dunn's test. The differences were
considered significant at 5% level.
RESULTS
Acute toxicity: The EE was found to be safe in the
doses used and there was no mortality in a dose of
1 g/kg, i.p.
Assessment of anticonvulsant activity
Maximum electroshock test: The duration of tonic
hindleg extension in mice treated with vehicle was
15.66±1.33 sec. The EE in doses of 10, 30, 100 and
500 mg/kg did not protect animals from seizures and
duration of hindleg extension was not reduced. The
vehicle treated mice showed tonic hindleg extension
for 15.66±1.33 sec whereas mice treated with the
EE (10, 30, 100, and 500 mg/kg) exhibited hindleg
extension for 14.33±1.14, 18.33±1.89, 16.33±1.43,
and 18.8±2.45 sec respectively.
Pentylenetetrazol-induced seizures: In animals
treated with vehicle, clonic convulsions appeared
140.3±13.8 sec after PTZ and all animals died after
seizures. The EE significantly and dose dependently
inhibited the onset and incidence of convulsions. The
convulsions were completely abolished by the dose
of 100 mg/kg. The EE in a dose of 10 mg/kg exhib-
ited seizures in 66.6% of mice and all animals exhib-
iting seizures died within 30 min. No mortality was
observed in the groups treated with EE 30 and 100
mg/kg even after 24 h. Diazepam (2 mg/kg) inhibited
seizures completely (Table 1).
Lithium-pilocarpine-induced status epilepticus:
Rats treated with lithium and pilocarpine showed
stage 4 seizures in all animals 75 min after pilo-
carpine. The EE diminished the intensity of seizures
and none of the animals exhibited stage 4 seizures
254 S.D. AMBAWADE et al.
and the animals were normal in behavior after 180
min. The EE in a dose of 100 mg/kg was more effec-
tive than other two doses in inhibiting the seizures till
45 min. Thereafter the effects of the three doses were
almost similar. The observations are given in Table 2.
DISCUSSION
The observations emanated in the present study in-
dicated that the EE was without any lethal effect in a
dose upto 1 g/kg and possessed anticonvulsant ac-
tivity against seizures induced by PTZ and lithium-
pilocarpine in a dose dependent way. However it was
not effective against MES induced seizures. Since
inhibition of the MES test predicts activity against
generalized tonic-clonic and cortical focal seizures,
lack of activity against MES induced seizures sug-
gests that the EE is not useful in suppressing gener-
alized tonic-clonic seizures. Several drugs are
thought to inhibit seizures by regulating GABA-
mediated synaptic inhibition through an action at dis-
tinct sites of the synapse17.
Although lithium does not possess general procon-
vulsant action in rats, its pretreatment provokes limbic
seizures following administration of sub-convulsant
doses of pilocarpine and other cholinergic agonist18.
The combined treatment with lithium and pilocarpine
results in accumulation of acetylcholine and inositol
monophosphate and reduction in cortical inositol that
are about ten times greater than the effects obtained
with either drugs alone13,18,19. Phenobarbitone, sodium
valproate, diazepam and trimethadione prevent the
limbic seizures induced in rats by pilocarpine, how-
ever, phenytoin and carbamazepine are ineffective20.
Studies from our laboratory have shown that block-
ade of post-synaptic 5-HT receptors and inhibition
of serotonergic transmission inhibited lithium-pilo-
carpine-induced seizures21 and also suppressed
PTZ-induced seizures (unpublished data).
Thus, in conclusion, G. glabra possesses anticonvulsant
property against the PTZ and lithium and pilocarpine
induced seizures. Further research is in progress to iso-
late the compound responsible for the activity.
ACKNOWLEDGEMENTS
This work was supported in part by the All India Coun-
cil for Technical Education, New Delhi, India. Authors
are grateful to Prin. J.S.Shete for providing labora-
tory facilities.
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COL. R.N. CHOPRA

(17.8.1882 - 13.6.1973)

Fifty paise stamp issued on his 101st birth anniversary

Dr. R.N. Chopra, hailing from Jammu & Kashmir had his early education at Lahore. He took Natural
Sciences Tripos from Cambridge University. After completing his medical studies at St. Bartholomew's
Hospital, London he returned to Cambridge to work with Prof. W.E. Dixon on his thesis on "ciliary
movements" and in 1908 he was awarded an M.D. degree. In 1909, he joined the Indian Medical
Service with the 19th Punjab Regiment in North West Frontier Province and served in East Africa in
World War I. He was awarded ScD (Cantab) in 1927. After several years in general medical duties he
joined the Calcutta School of Tropical Medicine (STM) and simultaneously was Professor of
Pharmacology at the Calcutta Medical College. He initiated several programs on clinical and
experimental research including that on Indian indigenous drugs. His pioneering publications include
Indigenous Drugs of India, Glossary of Indian Medicinal Plants, Poisonous Plants of India and Drug
Addiction in India. He introduced research on Rauwolfia Serpentina. He was a Director of STM and
Drug Research Institute, Srinagar. From 1958 onwards he held the post of Scientific Advisor, RRL,
Jammu. He was made Brevet Colonel in 1935 and knighted by the Queen in 1941.
The birth of the Indian Pharmacological Society in 1969 received his blessings. He was the first
president of IPS. Dr. R.N. Chopra oration which was started as a memorial tribute by IPS following his
demise, is an integral part of the Annual Conference of IPS each year.

Dr. Syed Ziaur Rahman

Department of Pharmacology,
J.N. Medical College, Aligarh Muslim University,
Aligarh-202 002.