Pakistan Journal of Pharmacology
Vol.26, No.2, July 2009, pp.41-47
HYPOTENSIVE ACTIVITY OF METHANOLIC EXTRACT
OF BERBERIS VULGARIS (ROOT PULP AND BARK)
AISHA AZMATa, MUHAMMAD AHMEDb, NAVAID UL ZAFARc
AND S.I. AHMADd
a
Umm Al-Qura University, Faculty of Medicine, Department of Physiology,
P.O. Box 7607 Makkah, Saudi Arabia
b
Umm Al-Qura University, Faculty of Pharmacy, Department of Pharmacology,
P.O. Box 7607. Makkah. Saudi Arabia
c
Hamdard Labs(WAQF), Pakistan, Karachi.74600.
d
Dr. HMI Institute of Pharmacology and Herbal Sciences,
Hamdard University, Karachi-74600, Pakistan
ABSTRACT
Many plants have been recommended in alternative systems of medicine for hypotensive
activity although few systematic studies have been done. In this study, the hypotensive
activity was studied in normotensive albino rats at different doses of BRPM and BB. The
receptor activity was assessed by the drugs Ach on rat heart. Administration of different
methanolic extract (BRPM, BB) showed significant reduction in blood pressure
comparable to its respective control. Both methanolic extract completely blocked the Ach
receptors in heart. This study suggests that the methanolic extract (BRPM, BB) have
significant Ach receptor blocking and hypotensive activity and may be recommended for
further investigation.
Keywords: Berberis vulgaris, Hypotensive, methanolic extract of root pulp (BRPM),
methanolic extract of root bark (BB).
INTRODUCTION
Each year heart disease (hypertension)
kills twice as many people as cancer and eight
times as many people as car accidents or
infections (Bapist, 2003). According to
Survey, hypertension and other “vascular
diseases” such as stroke and kidney failure
cause 43 percent of all deaths recorded each
year (Saleem et al., 2003). While in the United
State, Hypertension accounts for more deaths,
one million people die annually or more than
2500 die daily (Rose and Treadway, 1999).
This disease is responsible for over 50 percent
of all death in Pakistani males of working age
and about 90 percent of all sudden deaths.
Because it kills a person in the prime of his
carrier, the economic and social burden of the
disease is immense (Debakey and Gott, 1997).
Correspondence: E-mail:aishaazmatkhan@hotmail.com
Control of blood pressure in patient with
hypertension is necessary for cardiovascular
morbidity and mortality (Walker et al., 2002)
because high blood pressure if untreated,
ultimately leads to the most common heart
diseases like angina pectoris, stroke, heart
attack, congestive heart failure and
atherosclerosis (Ibarrolaaet al., 1996).
Therefore the present work have been
planned to see/study the effect of plant on
blood pressure using new and sensitive
technologies and statistical analysis. Earlier
the aqueous extract from Berberis vulgaris
fruit was tested to evaluate its antihypertensive
effects on DOCA-induced hypertension in the
rats (Fatehi, et al., 2005). Present studies led to
the determination of hypotensive activity of
methanolic extract of Berberis vulgaris root.
42
Hypotensive Activity of Methanolic Extract of Berberis vulgaris
MATERIALS AND METHODS
Plant Material
The roots of Berberis vulgaris were
yellowish brown, cylindrical, more or less
knotty, hard and tough. With the bark intact
they are cut into pieces of varying length and a
maximum diameter of 45 mm. The bark is
internally dark brown and soft. The root in
powdered form is bright yellow with a slight
odour and a bitter taste.
Extraction
1. Uncrushed root pulp of Berberis vulgaris
was extracted with methanol four times at
room temperature. The extracts were
combined together and evaporated on
rotavapour. At the end yellowish brown
colored residue (BRPM) was collected
and used during the whole study.
2.
Root bark of Berberis vulgaris was
extracted with methanol four times at
room temperature. The extracts were
combined together and evaporated on
rotavapour. A blackish brown and thick
(BB) matter was settling down in the
flask. That was used during the whole
study.
All these extracts (BRPM and BB) and
chemical were dissolved in distilled water for
in vivo experiments. All extracts and
chemicals were freshly prepared before each
experiment.
Animals
Adult Sprague dawley rats (200-225g) of
either sex were obtained from Dr. Hafiz
Muhammad Ilyas Institute of Pharmacology
and Herbal Sciences (Dr. HMIIPHS) and were
housed in groups of 6 per cage for seven days
prior to experimentation with free access to
food and tap water ad libitum and kept on a
12h light/dark cycle. Each experimental group
consisted of six animals. All animals were
housed in an air-conditioned room at 23+1ºC
during the quarantine period.
Instruments for the extraction and recording
of blood pressure parameters
Rotavapour (R114 Buchi) was used for
the extraction of BRPM and BB. The arterial
blood pressure was recorded by using research
grade blood pressure transducer (Harvard, 603003) coupled with four channels Harvard
Universal Oscillograph (Curvilinear, 50-9307)
(United kingdom).
Hypotensive activity
Normotensive Sprague-Dawley rats of
either sex (200-250 g) were anaesthetized with
Pentothal sodium (40 mg/kg i.p.) as described
by Ulicna et al., 2003. Then the trachea was
exposed and cannulated with polyethylene
cannula to facilitate spontaneous respiration.
Drugs were injected (vol. 0.2-0.25 ml) through
a polyethylene cannula inserted into the
external jugular vein followed by a saline flush
(0.2 ml). The arterial blood pressure was
recorded from the carotid artery via arterial
cannula connected to a research grade blood
pressure transducer (Harvard, 60-3003)
coupled with four channels Harvard Universal
Oscillograph (Curvilinear, 50-9307) (UK). The
temperature of the animals was maintained at
37oC by using the overhead lamp. Animals
were allowed to equilibrate for at least 15 min
before administration of any drug.
Measurements
Mean arterial blood pressure was calculated as
sum of the diastolic blood pressure plus onethird-pulse width (Adebouye et al., 1999).
Changes in blood pressure were expressed as
the percent of control values, obtained
immediately before the administration of test
substance
(Saleem,
et
al.,
2003).
Acetylcholine used as positive control caused
57.61 + 2.31% (mean + SEM, n=15) fall in
mean arterial blood pressure as the dose of 10-6
M/kg. The hypotensive studies were carried
out on different doses of BRPM and BB.
Methods for the determination of receptor
activity
Normotensive Sprague-Dawley rats of
either sex (200-250 g) were anaesthetized and
their blood pressure was recorded through
Azmat et al.
43
carotid artery as described earlier. Atropine 104
M/kg were injected through a polyethylene
cannula inserted into the external jugular vein
followed by a saline flush (0.2 ml), to block
the muscarinic receptors. The arterial blood
pressure was continuously monitored from the
carotid artery via arterial cannula connected to
a research grade blood pressure transducer
(Harvard, 60-3003) coupled with four channels
Harvard Universal Oscillograph (Curvilinear,
50-9307) (UK). Animals were allowed to
equilibrate for at least 5 min than
Acetylcholine was administered to check the
blockade of receptor than BRPM and BB
administered one by one and change in blood
pressure was monitored.
Statistical analysis
Changes in blood pressure and serum
biochemical levels were compared using
student’s t-test Values of P<0.05, P<0.01 and
P<0.001 were considered to be significant.
RESULTS
I. Effect of various doses of BRPM on
Various Blood Pressure Parameters
The effect of intravenous administration
of various doses of methanolic extract of root
(BRPM) on various blood pressure parameters
has been presented in Table 1.
a) Effect of 0.5mg/kg BRPM on various
Blood Pressure Parameters
BRPM at the dose of 0.5mg/kg was found
to reduce the systolic pressure, diastolic and
mean arterial blood pressure by 31%, 36% and
32% respectively in comparison with their
controls respectively as shown in Fig. 1.
Decreases in various blood pressure
parameters were statistically significant
(p<0.005). Hypotensive effect remained for
73.33+10.18 sec than it returned to normal.
b) Effect of 1mg/kg BRPM on various
Blood Pressure Parameters
Intravenous administration of 1mg/kg
dose of BRPM showed significant decrease in
systolic, diastolic and mean arterial blood
pressure by 42%, 44% and 43% respectively
as shown in Fig. 1. These reductions in various
Blood Pressure parameters were statistically
significant (p<0.005) when compared to their
control. At the dose of 1mg/kg hypotensive
effect returned to normal after 157.73+6.84
sec.
c) Effect of 10mg/kg BRPM on various
Blood Pressure Parameters
BRPM (10 mg/kg) also showed a decrease
in the systolic, diastolic and mean arterial
blood pressure by 48%, 51% and 50%
respectively. The reductions in various blood
pressure
parameters
were
statistically
Table 1
Effect of methanolic extract of root (BRPM) obtained at different doses
on various BP Parameters
Parameters
Systolic BP
Diastolic BP
MABP
0.5mg/kg
Control
Treated
109 ±
76.83 ±
8.63
12.3
(6)
(6)
103.83 ±
72 ±
8.71
11.93
(6)
(6)
105.53 ±
73.61 ±
8.68
12.05
(6)
(6)
1mg/kg
Control
Treated
122.75 ±
69.25 ±
7.11
9.04
(6)
(6)
121.75 ±
66.25 ±
7.11
10.18
(6)
(6)
122.08 ±
67.25 ±
7.11
9.79
(6)
(6)
10mg/kg
Control
Treated
135.36 ±
70.09 ±
4.75
5.54
(11)
(11)
133.45 ±
65.55 ±
4.64
5.65
(11)
(11)
134.09 ±
67.06 ±
4.67
5.56
(11)
(11)
30mg/kg
Control
Treated
126.14 ±
77 ±
9.24
11.34
(7)
(7)
123.43 ±
71.71 ±
9.09
11.81
(7)
(7)
124.33 ±
73.48 ±
9.14
11.63
(7)
(7)
Hypotensive Activity of Methanolic Extract of Berberis vulgaris
44
Table 2
Effect of methanolic extract of root Bark (BB) obtained at different doses
on various BP Parameters
Parameters
Systolic BP
Diastolic BP
MABP
10mg/kg
Control
116.33±2.48
(6)
115±2.83
(6)
115.44±2.71
(6)
Treated
77.33±0.41
(6)
74.33±0.41
(6)
75.33±0.24
(6)
The values have been presented as mean+S.E.M (n)
significant (p<0.005) (Fig. 1). Hypotensive
effect at 10mg/kg remained effective for a
longer time period of 977.20+237.42 sec. It
tries to return to control values but failed and
the blood pressure dropped to 10.53+2.54%.
d)
Effect of 30mg/kg BRPM on various
Blood Pressure Parameters
BRPM at the dose of 30mg/kg was found
to reduce the systolic pressure (126.14+9.24 to
77+11.34),
diastolic
(123.43+9.09
to
71.71+11.81) and mean arterial blood pressure
(124.33+9.14 to 73.48+11.63) that was 40%,
43% and 42% respectively in comparison with
their controls as shown in Fig. 1. Decreases in
various blood pressure parameters were
statistically significant (p<0.005). At the dose
of 30mg/kg hypotensive effect remained for
851+441.17 sec than it tries to return to normal
but failed and the blood pressure dropped to
11+4.16%.
In all the dosage of BRPM tested
maximum reduction in various blood pressure
parameters was observed at 10mg/kg (Fig. 1)
being the optimal dose.
II. EFFECT OF VARIOUS DOSES OF BB
ON VARIOUS BLOOD PRESSURE PARAMETERS
The effect of intravenous administration
of Methanolic extract of root bark (BB) on
various blood pressure parameters has been
presented in Table 2.
a)
Effect of 10mg/kg BB on various
Blood Pressure Parameters
Methanolic extract of root bark BB (10
mg/kg) also showed a decrease in the systolic
(116.33+2.48 to 77.33+0.41), diastolic
(115+2.83 to 74.33+0.41) and mean arterial
blood pressure (115.44+2.71 to 75.33+0.24)
by 33%, 35% and 34% respectively. The
reductions in various blood pressure
Azmat et al.
45
parameters were statistically significant
(p<0.005) in comparison with their respective
controls
(Fig. 2). Hypotensive effect at
10mg/kg remained effective for a time period
of 50.5+6.36 sec.
III.
Result of receptor activity
The receptor activity was determined in
rats results demonstrate that BRPM
(10mg/kg), and BB (10mg/kg) has been found
to decrease the mean arterial blood pressure
that was 50% and 34% respectively when
compared with their respective controls (Fig.
1, 2). While, on pre-treatment with Atr (10-4
M) the BRPM (10mg/kg), and BB (10mg/kg)
did not produce any reduction in Blood
pressure. These behavior matches exactly to
that of Acetylcholine. The use of Ach (10-6 M)
has resulted in significant fall in MABP that
was 46% than its respective control. Further,
on pre-treatment with Atr (10-3 M) the Ach
(10-6 M) did not show decline in MABP as
shown earlier without Atr (10-3 M) pretreatment.
DISCUSSION
In present study methanolic extracts of
Berberis vulgaris has been tested for its
hypotensive effects in rats. The acute
intravenous administration of different extract
of Berberis vulgaris (BRPM, BB) as well as
Ach used as positive reference drug (Lahlou et
al 2002) in this study, all exerted immediate
and significant fall in systolic, diastolic and
mean arterial blood pressure (MABP) in
normotensive
anaesthetized
rats.
Physiologically, the reduction in blood
pressure has been reported to be either due to
their direct or indirect effect on pumping
ability of heart (Kimura et al., 1986; Ebeigbe
and Ezimokhai, 1988) or associated with
decrease in peripheral resistance (vascular
muscle tone leading to vasodilation) (Katzung,
1998). On these basis it is difficult to pin point
the exact mechanism of extracts. Earlier it is
suggested by Kamanyi et al., (1993) that the
fall in blood pressure immediately after the
administration of the plant extract was due to a
possible direct relaxing effect of extract on
vascular smooth muscle. It is reported that
Ach cause a generalized vasodilation that is an
indirect effect mediated by release of nitric
oxide from vascular endothelial cells
(Furchgott and Zawadzki 1980). That’s why it
is hypothesized that these extracts might be
acting like Ach and Keeping these
assumptions the receptor activity of extracts
has also been tested in the present study by
using cholinergic agonist Ach and cholinergic
competitive antagonist Atr. The results
demonstrate that Ach and different extracts of
Berberis vulgaris, when administered alone
have reduced the blood pressure. On the other
hand the use of Ach and different extracts of
Berberis vulgaris, on Atr pre-treated animal
did not show such decline in blood pressure as
shown earlier without atropine pre-treatment.
These results clearly indicate that effect of
Berberis vulgaris is mediated through same
receptor and mechanism as established for Ach
indicating a role of cardiac muscarinic receptor
in this response.
It is suggested that different extracts of
Berberis vulgaris stimulates the muscarinic
receptors located on the endothelial cells of the
vasculature. This stimulation results in the
release of endothelium-derived relaxing
factors (EDRFs) or nitric oxide (Moncada et al
1991), that diffuse to smooth muscle cells and
Hypotensive Activity of Methanolic Extract of Berberis vulgaris
46
cause their relaxation (Furchgott 1983;
Vanhoutte et al 1995). It may therefore be
concluded that muscarinic M2 receptors on
cardiac muscles as well vascular dilation by
endothelium derived relaxing factor may
contribute to the cholinergic hypotensive
effect of these extracts.
The present study suggests that different
extracts of Berberis vulgaris possess
significant hypotensive.
CONCLUSION
From the above discussion it is concluded
that BRPM and BB is a physiologically and
pharmacologically active drug because
systolic, diastolic and mean arterial blood
pressure is influenced by BRPM and BB used
in the present study. Different methanolic
extract of berberis vulgaris used in this study
caused hypotensive effect by stimulating nonselective muscarinic receptors.
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