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Advances in Natural and Applied Sciences, 8(4) April 2014, Pages: 203-207
AENSI Journals
Advances in Natural and Applied Sciences
ISSN:1995-0772 EISSN: 1998-1090
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Methanol Extract of Allium sativum l. Leaves Show Antinociceptive Activity in Acetic
Acid-Induced Writhing Tests in Mice
Sohana Afrin Nirjhor, Humayra Jannat, Mousumi Ahmed, Diponcor Ghosh, Indrani Mandal, Ummay Kawchur
Trisha, Mohammed Rahmatullah
Faculty of Life Sciences, University of Development Alternative, Dhanmondi, Dhaka-1209, Bangladesh.
ARTICLE INFO
Article history:
Received 25 January 2014
Received in revised form 8
March 2014
Accepted 10 March 2014
Available online 5 May 2014
Keywords:
Allium sativum, antinociceptive, mice
ABSTRACT
The antinociceptive effect of methanolic extract of Allium sativum leaves were studied
through the intraperitoneally administered acetic acid-induced pain model in Swiss
albino mice. At doses of 50, 100, 200, and 400 mg methanolic extract of Allium
sativum leaves (MEAS) per kg body weight, MEAS caused dose-dependent and
significant reductions in the number of abdominal constrictions (writhings) induced in
mice by intraperitoneal administration of acetic acid. At the afore-mentioned four
doses, the number of abdominal constrictions (writhings) was reduced, respectively, by
24.0, 40.0, 44.0, and 52.0%. A standard antinociceptive drug, aspirin, when
administered at doses of 200 and 400 mg per kg body weight, reduced the number of
writhings by 32.0 and 60.0%, respectively. Thus MEAS at doses of 100, 200 and 400
mg per kg body weight produced antinociceptive effects and caused alleviation of pain,
which was greater than that of the pain-alleviating drug, aspirin, when the latter was
used at a dose of 200 mg per kg body weight.
© 2014 AENSI Publisher All rights reserved.
To Cite This Article: Sohana Afrin Nirjhor, Humayra Jannat, Mousumi Ahmed, Diponcor Ghosh, Indrani Mandal, Ummay Kawchur Trisha,
Mohammed Rahmatullah., Methanol extract of allium sativum l. Leaves show antinociceptive activity in acetic acid-induced writhing tests in
mice. Adv. in Nat. Appl. Sci., 8(4): 203-207, 2014
INTRODUCTION
Allium sativum L. (Liliaceae, English: garlic, Bengali: roshun) is a plant native to Central Asia but is now
grown all over the world as a spice crop. Its close relatives are onion and shallot. Garlic has since antiquity
being regarded as a medicinal plant, the part used for medicinal purposes being the bulb or cloves of the plant,
which remains underneath the soil while growing. There have been reports in Egyptian papyruses dating about
1550 BCE of the use of garlic for a variety of ailments. Hippocrates, Pliny and Aristotle also have mentioned the
use of garlic for therapeutic purposes.
Modern science has recognized the many beneficial effects of garlic. It has been reported to improve
learning and memory retention (Borek, 2006). Garlic has been shown to suppress the nuclear factor-kappaB
activation pathway and so reduce inflammation (Aggarwal and Shishodia, 2004). The beneficial effects of garlic
in diabetes and cardiovascular disorders have been well established. S-allyl cysteine, a compound present in
garlic has been shown to restore erectile function in diabetic rats through inhibition of generated oxygen species
(Yang et al., 2013). Garlic extract has been shown to decrease blood glucose level and improve lipid profile in
alloxan-diabetic rabbits (Sher et al., 2012; Nasri, 2013). The renoprotective effect of aged garlic in
streptozotocin-induced diabetic rats has been demonstrated (Shiju et al., 2013). The nephroprotective effect of
aqueous extract of Allium sativum bulbs has been attributed to attenuation of vascular endothelial growth factor
and extracellular signal-regulated kinase-1 expression in diabetic rats (Shiju et al., 2013). In fructose-fed male
rats, garlic has been observed to attenuate oxidative stress and insulin resistance (Sivaraman et al., 2013).
A preventive effect of garlic has been shown in changes in serum biochemical parameters and
histopathology of pancreas and liver in streptozotocin-induced diabetic rats (Masjedi et al., 2013). In
experimental diabetes, garlic has been shown to inhibit rat alpha-glucosidase and enhanced pancreatic insulin
and cardiac Glut-4 mRNAs expression (Moradabadi et al., 2013). Alliin,a component of garlic, reportedly
prevented LPS-induced inflammation in 3T3-L1 adipocytes (Quintero-Fabián et al., 2013). Diallyl trisulfide,
another component of garlic has been shown to demonstrate protective effect against vascular endothelial
impairment (Liu et al., 2014). Protective effects of garlic have also been noted in Alzheimer’s disease,
Corresponding Author: Mohammed Rahmatullah, Faculty of Life Sciences, University of Development Alternative,
Dhanmondi, Dhaka-1209, Bangladesh.
Telephone: +88-01715032621 Fax: +88-02-815739; E-mail: [email protected]
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Sohana Afrin Nirjhor, et al, 2014
Advances in Natural and Applied Sciences, 8(4) April 2014, Pages: 203-207
atherosclerosis, hyperlipidemia, hypertension, thrombosis, cancer, skin diseases, and fungal infections
(Bongiorno et al., 2008).
We had been screening Bangladeshi medicinal plants for their potential antidiabetic, antinociceptive, and
cytotoxic activities (Anwar et al., 2010; Jahan et al., 2010; Rahman et al., 2010; Rahmatullah et al., 2010;
Shoha et al., 2010; Ali et al., 2011; Barman et al., 2011; Hossan et al., 2011; Jahan et al., 2011; Rahman et al.,
2011; Sutradhar et al., 2011; Ahmed et al., 2012; Arefin et al., 2012; Haque et al., 2012; Sathi et al., 2012). As
part of that process, the objective of the present study was to evaluate the antinociceptive potential of
methanolic extract of Allium sativum leaves in Swiss albino mice in acetic acid-induced gastric pain model
experiments.
MATERIALS AND METHODS
Plant material:
Leaves of Allium sativum were collected from Mullatagor in Kushtia district, Bangladesh during February
2013. The plant was taxonomically identified at the Bangladesh National Herbarium at Dhaka (Accession
Number 38,573). The sliced and air-dried leaves of Allium sativum were grounded into a fine powder and 100g
of the powder was extracted with methanol (1:5, w/v) for 48 hours. The extract was evaporated to dryness at
40oC. The final weight of the extract was 5.49g.
Chemicals:
Glacial acetic acid was obtained from Sigma Chemicals, USA; aspirin was obtained from Square
Pharmaceuticals Ltd., Bangladesh.
Animals:
In the present study, Swiss albino mice (male), which weighed between 13-18g were used. The animals
were obtained from International Centre for Diarrheal Disease Research, Bangladesh (ICDDR,B). All animals
were kept under ambient temperature with 12h light followed by a 12h dark cycle. The animals were
acclimatized for three days prior to actual experiments. The study was conducted following approval by the
Institutional Animal Ethical Committee of University of Development Alternative, Dhaka, Bangladesh.
Antinociceptive activity:
Antinociceptive activity of the methanol extracts of Allium sativum leaves (MEAS) was examined using
previously described procedures (Shanmugasundaram and Venkataraman, 2005). Briefly, mice were divided
into seven groups of six mice each. Group 1 served as control and was administered vehicle only. Groups 2 and
3 were orally administered the standard antinociceptive drug aspirin at a dose of 200 and 400 mg per kg body
weight, respectively. Groups 4-7 were administered methanolic leaf extract (MEAS) at doses of 50, 100, 200
and 400 mg per kg body weight, respectively. Following a period of 60 minutes after oral administration of
standard drug or extract, all mice were intraperitoneally injected with 1% acetic acid at a dose of 10 ml per kg
body weight. A period of 15 minutes was given to each animal to ensure bio-availability of acetic acid,
following which period, the number of abdominal constrictions (writhings) was counted for 10 min. The
following formula was used for calculation of percent inhibition of the number of writhings in aspirin and
MEAS administered animals compared to control mice,
Percent inhibition = (1 – We/Wc) X 100
where We and Wc represents the number of writhings in aspirin or MEAS administered mice (Groups 2-7),
and control mice (Group 1), respectively.
Statistical analysis:
Experimental values are expressed as mean ± SEM. Independent Sample t-test was carried out for statistical
comparison. Statistical significance was considered to be indicated by a p value < 0.05 in all cases.
RESULTS AND DISCUSSION
At doses of 50, 100, 200, and 400 mg methanolic extract of Allium sativum leaves (MEAS) per kg body
weight, MEAS caused dose-dependent and significant reductions in the number of abdominal constrictions
(writhings) induced in mice by intraperitoneal administration of acetic acid. At the afore-mentioned four doses,
the number of abdominal constrictions (writhings) was reduced, respectively, by 24.0, 40.0, 44.0, and 52.0%. A
standard antinociceptive drug, aspirin, when administered at doses of 200 and 400 mg per kg body weight,
reduced the number of writhings by 32.0 and 60.0%, respectively. Thus MEAS at doses of 100, 200 and 400 mg
per kg body weight produced antinociceptive effects and caused alleviation of pain, which was greater than that
of the pain-alleviating drug, aspirin, when the latter was used at a dose of 200 mg per kg body weight. The
results are shown in Table 1.
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Sensation of pain has been attributed to increase in the expression of prostaglandins (Reynolds, 1982; Rang
and Dale, 2003). Intraperitoneal administration of acetic acid releases prostaglandins like PGE2 and PGF2alpha
and their levels are increased (Deraedt et al., 1980). As such, the antinociceptive activity exhibited by crude
methanolic extract of the leaves of Allium sativum may be due to the extract’s ability to block any further
expression of prostaglandins, which may be mediated through inhibition of cyclooxygenase and/or
lipooxygenase activities. A similar mechanism has been proposed for antinociceptive activity of Ficus deltoidea
Jack (Moraceae) aqueous extract in acetic acid-induced gastric pain model (Sulaiman et al., 2008), and this can
also be the mechanism operating in the present study. However, more studies are warranted to find out the
mechanism behind the observed antinociceptive effects.
Table 1: Antinociceptive effect of crude methanol extract of Allium sativum leaves in the acetic acid-induced gastric pain model mice.
Treatment
Dose (mg/kg body
Mean number of writhings
% inhibition
weight)
Control (Group 1)
10 ml
5.00 ± 0.32
Aspirin (Group 2)
200 mg
3.40 ± 0.24
32.0*
Aspirin (Group 3)
400 mg
2.00 ± 0.45
60.0*
MEAS (Group 4)
50 mg
3.80 ± 0.58
24.0*
MEAS (Group 5)
100 mg
3.00 ± 0.55
40.0*
MEAS (Group 6)
200 mg
2.80 ± 0.58
44.0*
MEAS (Group 7)
400 mg
2.40 ± 0.40
52.0*
All administrations (aspirin and extract) were made orally. Values represented as mean ± SEM, (n=5); *P < 0.05; significant compared to
control.
Allium sativum leaves are known to contain beta-sitosterol (Duke, 1992). The compound isolated from the
leaves of Mentha cordifolia Opiz. has been shown to demonstrate analgesic activity in acetic acid-induced
writhing tests (Villaseñor et al., 2002). Chlorogenic acid, another constituent present in the plant also has been
shown to have analgesic and anti-inflammatory potential (Azza et al., 2011). Sinapic acid has also been reported
to be present in the plant. The compound has been shown to give anti-inflammatory effects through the
suppression of inducible nitric oxide synthase, cyclooxygenase-2, and proinflammatory cytokines expression via
nuclear factor kappaB inactivation (Yun et al., 2008). Thus these compounds can account for the observed
antinociceptive effects.
Allium sativum plants are cultivated and can be commonly found in Bangladesh. Thus this plant can prove
useful in alleviation of pain, which is common in people from all over the world, including Bangladesh. Modern
doctors and hospitals are particularly lacking in the rural areas of the country. Thus the leaves of Allium sativum
can be a useful folk remedy for treatment of common pain.
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