Pharmacological Research 48 (2003) 31–35

Antinociceptive effects of Teucrium polium L. total extract

and essential oil in mouse writhing test
Mohammad Abdollahi a,∗ , Hoornaz Karimpour a , Hamid Reza Monsef-Esfehani b
a Department of Toxicology & Pharmacology and b Department of Pharmacognosy, Faculty of Pharmacy,
Tehran University of Medical Sciences, Tehran 14155-6451, Iran

Accepted 4 February 2003

Abstract

In this study, the effect of Teucrium polium L., a wild-growing flowering plant belonging to the family Labiatae and found abundantly
in Iran was studied on writhing test, a visceral pain model in mice. The total extract and essential oil were administered intraperitoneally
30 min before acetic acid 0.6% injection. Total extract in doses (mg kg−1 ) of 150 (39.13%, P < 0.001), 225 (65.44%, P < 0.001), 300
(21.41%, P < 0.01) induced reduction in writhing response as compared to control with the ED50 67.92 mg kg−1 .
The essential oil in doses (mg kg−1 ) of 9.37 (35.22%, P < 0.001), 18.75 (59.63%, P < 0.001), 37.5 (86.60%, P < 0.001), 75 (90.22%,
P < 0.001) and 150 (78.58%, P < 0.001) induced significant reduction in writhing response when compared to control with the ED50 of
29.41 mg kg−1 . In order to ensure the importance of essence in production of visceral antinociception, the extract free from essential oil
was prepared and injected into mice at a dose of 225 mg kg−1 (the most effective dose of the extract) which in comparison to total extract,
the antinociception, reduced from 65.44 to 49.85% (P < 0.001). It is concluded that essential oil is responsible for analgesic properties
of T. polium. This study confirms the antivisceral pain properties of T. polium comparable to those of hyoscine and indomethacin and
suggests a good place for it in antispasmodic therapies in human. The presence of flavonoids and sterols might be responsible for the
anti-inflammatory activity of this plant.
© 2003 Elsevier Science Ltd. All rights reserved.

Keywords: Teucrium; Visceral pain; Writhing test; Mouse; Plant

1. Introduction folk medicine [5]. Anti-inflammatory [15], antihypertensive

Since the available analgesic drugs exert a wide range of
side effects and are either too potent or too weak, the search
for new analgesic compounds has been a priority of phar-
macologists and pharmaceutical industries [1]. Medicinal
plants are believed to be an important source of new chemi-
cal substances with potential therapeutic effects [2,3]. Thus,
study of plant species that traditionally have been used as
pain killers should still be seen as a logical research strategy,
in search for new analgesic drugs [4].
The Teucrium polium L. is a wild-growing flowering
plant belonging to the family Labiatae and is found abun-
dantly in south western Asia, Europe and North Africa.
Teucrium species have been used as medicinal herbs for
over 2000 years as diuretic, diaphoretic, tonic, antipyretic,
antispasmodic, cholagogic and many of them are used in
∗ Correspondingauthor. Tel.: +98-911-2710860; fax: +98-21-6461178.
E-mail addresses: mohammad.abdollahi@utoronto.ca,
mabdol@yahoo.com (M. Abdollahi).
[6] and anorexic [7] effects are other reported activities of
T. polium L. Among the several models of visceral pain,
writhing test has been mostly used as a standard screen-
ing method. To examine the antivisceral pain effects of T.
polium in more detail, the present experiment was designed
to investigate the effects of Teucrium’s total extract and its
essential oil on the writhing test in mice.
2. Materials and methods
2.1. Plant material
T. polium aerial parts were collected during the flowering
period from Siyahbisheh (around Chalous Road, in the North
of Iran). Samples of the plant were identified by botanist
from the Division of Pharmacognosy, University of Tehran.
The plant material was air dried, powdered and extracted
using hydroethanolic (96◦ ) solvent in percolator. The sol-
vent was removed under vacuum in a rotary evaporator until

1043-6618/03/$ – see front matter © 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S1043-6618(03)00059-8
32 M. Abdollahi et al. / Pharmacological Research 48 (2003) 31–35

dryness. The percentage yields based on the dried starting
material was 14% for dried hydroalcoholic extract (w/w).
Essential oil was prepared from powdered plant using
Clevenger apparatus and hydrodistillation method. The per-
centage yields based on the dried plant was 0.2% (v/w).
In order to ensure the superiority of essence in antinoci-
ception, the extract free from essential oil was prepared by
heating the extract mildly.
2.2. Animals
Male albino N-MRI mice from Pasteur Institute of Tehran
weighing 20–25 g were kept under standardized conditions
(temperature: 22 ± 2 ◦ C, normal lighting) and fed with the
normal laboratory diet.
All treatments were done by i.p. administration, 30 min
before i.p. administration of acetic acid 0.6% (15 mg kg−1 ).
2.4. Writhing test
Mice were i.p. administered with the test preparations.
After 30 min, each animal was injected i.p. with acetic acid
0.6% (15 mg kg−1 ) and individually housed in a glass cylin-
der on a flat glass floor and a mirror was arranged at an
angle of 45◦ under the cylinder.
Antinociception was recorded by counting the number of
writhes after injection of acetic acid during three periods of
10 min. A writhe is indicated by abdominal constriction and
stretching of at least one hind limb [11,12].

2.3. Administration of reference and test compounds 3. Statistical analysis

Hyoscine (1 mg kg−1 ) [8] and indomethacin (5 mg kg−1 )
[9] were dissolved in normal saline and administered in-
traperitoneal (i.p.) reference drugs for comparison. The
respective controls received only saline.
The extract was suspended in a solution of carboxy-
methylcelulose, Tween 80, water (0.5:3.03:96.97, w/v/v)
[10]. Respective controls received only that suspension
as vehicle. An emulsion of essential oil was prepared us-
ing Tween 80 (2.5%, v/v) and water as solvent. Controls
received that emulsion as vehicle.
The doses of 75, 150, 225, 300 mg kg−1 of total extract;
9.37, 18.75, 37.5, 75, 150 mg kg−1 ; of the essential oil and
225 mg kg−1 of the extract free essential oil, were prepared.
Comparison between groups was made by one-way anal-
ysis of variance (ANOVA) followed by Tukey’s multiple
comparison test. Differences with P < 0.05 and lower
between experimental groups were considered statistically
significant.
4. Results
Antinociceptive effects induced by different doses of to-
tal extract on the writhing test in mice are shown in Table 1.
Total extract in doses (mg kg−1 ) used, induced significant
reduction in writhing response when compared to control

Table 1
Effect of intraperitoneal administration of total extract on abdominal writhing induced in mice by intraperitoneal injection of acetic acid
Dose (mg kg−1 ) N Number of abdominal Reduction in P value
writhings in 30 min writhing response
(mean ± S.D.) (%)
Control
Vehicle (10 ml kg−1 ) 6 56.50 ± 7.220 – –
Total extract
75 6 48.17 ± 5.913 11.61 <0.001 vs. hyoscin
<0.001 vs. indomethacin
150 6 33.17 ± 4.622 39.13 <0.001 vs. control
<0.001 vs. hyoscin
<0.001 vs. indomethacin
225 6 18.83 ± 5.037 65.44% <0.001 vs. control
300 6 42.83 ± 8.704 21.41% <0.01 vs. control
<0.001 vs. hyoscin
<0.001 vs. indomethacin
Hyoscine
1 6 14.17 ± 2.137 74.00% <0.001 vs. control
Indomethacin
5 6 13.17 ± 3.312 75.83% <0.001 vs. control
N: Number of animals per group. Inhibition percent was obtained in comparison to control. ANOVA results: F = 60.09, d.f. = 41. The ED50 for
antinociceptive effects of total extract was 67.92 mg kg−1 . The treatment compounds were suspended in a solution of carboxymethylcelulose, Tween 80,
water (0.5:3.03:96.97, w/v/v). Control animals received this suspension as vehicle. All treatments were done by intraperitoneal injection 30 min before
intraperitoneal administration of acetic acid 0.6% (15 mg kg−1 ). Antinociception was recorded by counting the number of writhes after injection of acetic
acid during 30 min. A writhe is indicated by abdominal constriction and stretching of at least one hind limb.
 M. Abdollahi et al. / Pharmacological Research 48 (2003) 31–35 33

Table 2
Effect of intraperitoneal administration of essential oil on abdominal writhing induced in mice by intraperitoneal injection of acetic acid
Dose (mg kg−1 ) N Number of abdominal Reduction in P value
writhings in 30 min writhing response
(mean ± S.D.) (%)
Control
Vehicle (10 ml kg−1 ) 6 51.81 ± 6.01 – –
Essential oil
9.37 6 35.330 ± 9.158 35.22 <0.001 vs. control
<0.001 vs. hyoscine
<0.001 vs. indomethacin
18.75 6 22.000 ± 2.366 59.63 <0.001 vs. control
37.5 6 7.333 ± 4.761 86.60 <0.001 vs. control
75 6 5.333 ± 2.875 90.22 <0.001 vs. control
150 6 11.670 ± 4.937 78.58 <0.001 vs. control
Hyoscine
1 6 14.170 ± 2.137 74.00 <0.001 vs. control
Indomethacin
5 6 13.170 ± 3.312 75.83 <0.001 vs. control
N: Number of animals per group. Inhibition percent was obtained in comparison to control. ANOVA results: F = 54.88, d.f. = 47. The ED50 for
antinociceptive effects of essential oil was 29.41 mg kg−1 . Treatment compounds were emulsified using Tween 80 (2.5%, v/v) and water. The control
animals received only this emulsion as vehicle. All treatments were done by intraperitoneal injection 30 min before intraperitoneal administration of
acetic acid 0.6% (15 mg kg−1 ). Antinociception was recorded by counting the number of writhes after injection of acetic acid during 30 min. A writhe
is indicated by abdominal constriction and stretching of at least one hind limb.

as follow: 150 (39.13%, P < 0.001), 225 (65.44%, P <
0.001), 300 (21.41%, P < 0.01). No significant difference
was observed in animals treated by dose of 75 mg kg−1 of
total extract on writhing test when compared to control.
The ED50 for antinociceptive effects of total extract was
67.92 mg kg−1 .
Antinociceptive effects induced by different doses of es-
sential oil on writhing test in mice are shown in Table 2.
Essential oil in all doses (mg kg−1 ) used, induced significant
reduction in writhing response when compared to control
as follow: 9.37 (35.22%, P < 0.001), 18.75 (59.63%, P <
0.001), 37.5 (86.60%, P < 0.001) 75 (90.22%, P < 0.001)
and 150 (78.58%, P < 0.001). The ED50 for antinociceptive
effects of essential oil was 29.41 mg kg−1 .
Antinociceptive effect induced by extract free from es-
sential oil on the writhing test in mice is shown in Table 3.
Extract without essential oil in dose of 225 mg kg−1 induced
significant reduction in writhing response 225 (49.85%, P <
0.001) when compared to control total extract.
5. Discussion
Teucrium species have been used as medicinal herbs for
over 2000 years. Different properties have been reported
by Teucrium various species. T. polium contains a potent
volatile oil, consisting of mono- and sesqui-terpenes, which
was found to inhibit the spontaneous movements of the
rabbit segmented jejunum by 5 mg ml−1 suspension for
an organ bath. The inhibitory effect of T. polium on the
calcium influx through the potential-dependent calcium
channels was proposed as its mechanism of action [14].
The ethanolic extract of T. polium at a dose of 500 mg ml−1
also produced significant inhibition of carrageenin-induced
inflammation and cotton-pellet granuloma. In addition, it
significantly decreased blood glucose level in treated an-
imals. They referred to presence of flavonoids and sterols
as active components of these effects [15]. In another
study, the aqueous extract of T. polium given i.p. at doses
of 50–150 mg kg−1 markedly reduced the serum levels of
cholesterol and triglycerides in hyperlipidemic rats. They
suggested that this effect is related to presence of flavonoids
in this plant [16]. The antihyperlipidemic and inhibitory ac-
tion on lipid peroxidation has been reported for flavonoids
and terpenoids [17,18]. Reduction of rat’s body weight, wa-
ter and food intake was also reported after oral (10 mg ml−1 )
and i.p. (200 mg kg−1 ) administrations of T. polium decoc-
tion in chronic use [7]. Investigations about other species
of Teucrium have also indicated anti-inflammatory and an-
tiulcer properties. The presence of terpenic compounds and
flavonoids was proposed as a possible mechanism [5,10].
The present results demonstrate the significant antinoci-
ceptive effect of essential oil at doses of 18.75, 37.5, 75
and 150 mg kg−1 (ED50 of 29.41) while the total extract
showed weaker efficacy in reduction of visceral pain (ED50
of 67.92). When dose of essential oil increased more than
75 mg kg−1 , the antinociceptive effect was decreased. We
suggest that essential oil has different components which
when the dose is increased, interact together negatively
in induction of antinociceptive effect that is unknown for
34 M. Abdollahi et al. / Pharmacological Research 48 (2003) 31–35

Table 3
Effect of intraperitoneal administration of extract free from essential oil on abdominal writhing induced in mice by intraperitoneal injection of acetic acid
Dose (mg kg−1 ) N Number of abdominal Reduction in P value
writhings in 30 min writhing response
(mean ± S.D.) (%)
Control
Vehicle (10 ml kg−1 ) 6 50.09 ± 5.984 – –
Total extract
225 6 18.83 ± 5.037 65.44% <0.001 vs. control
Extract without essential oil
255 6 27.33 ± 4.885 49.85% <0.001 vs. hyoscine
<0.001 vs. hyoscine
<0.001 vs. indomethacin
Hyoscine
1 6 14.17 ± 2.137 74.00 <0.001 vs. control
Indomethacin
5 6 13.17 ± 3.312 75.83 <0.001 vs. control
N: Number of animals per group. Inhibition percent was obtained in comparison to control. ANOVA results: F = 94.23, d.f. = 29. The treatment
compounds were suspended in a solution of carboxymethylcelulose, Tween 80, water (0.5:3.03:96.97, w/v/v). Control animals received this suspension
as vehicle. Hyoscine and indomethacin were dissolved in saline. All treatments were done by intraperitoneal injection 30 min before intraperitoneal
administration of acetic acid 0.6% (15 mg kg−1 ). Antinociception was recorded by counting the number of writhes after injection of acetic acid during
3o min. A writhe is indicated by abdominal constriction and stretching of at least one hind limb.

us presently. The data demonstrated in Table 3, indicate References

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