International Journal of Research in Pharmaceutical and Biomedical Sciences
ISSN: 2229-3701
___________________________________________Research Article
Aerial parts of Aqueous extract of Cynodon dactylon shows
hypotensive effect in high fructose treated Wistar rats
Deepak Bharti1, Pradnya Jagtap2*, vaishali Undale2 and Ashok Bhosale2
1SND
College of pharmacy, A/P- Babhulgaon, Tal-Yeola, Dist-Nasik, Maharashtra, India.
2Seth
Govind Raghunath Sable College of pharmacy, Saswad, Tal- Purandar, Dist- Pune,
Maharashtra, India.
_____________________________________________________________________________________
ABSTRACT
The purpose of the current study was to investigate the effect of the Aq. Extract of C. dactylon (100, 200mg/kg) on
mean arterial pressure (MABP) and plasma glucose, insulin, cholesterol, triglycerides and Creatinine levels in rats
with fructose induced hypertension. Chronic fructose treatment in rats has repeatedly been shown to elevate
blood pressure in association with increased glucose level and triglyceridemia. The Wistar rats who drank a 10%
fructose solution for 3&/6 weeks showed significant increase not only in plasma glucose and cholesterol levels but
also in MABP. Chronic administration of Aq. Extract of C. dactylon(100, 200mg/kg) in hypertensive rats significantly
reduced mean arterial BP(p≤0.01). The extract also shows significant decrease in pressor response to NA (0.5, 1.0
µg/kg) and Adrenalin (0.5, 1.0 µg/kg). These results suggest that the Aq. Extract of C. dactylon exerts is
antihypertensive effect by partly improving peripheral vascular resistant.
Key Words: C. dactylon, fructose, hypertension, pressor response.
INTRODUCTION
A number of Indian medicinal plants have been used
for thousands of years in the traditional system of
medicine (Ayurveda). Cynodon dactylon (L.) Pers.
(Family: Poaceae) commonly known as “Doob”,
”Durva” in India, is a weed and has been regarded to
possess various medicinal properties.1 The plant
posses antimicrobial, and antiviral activity and has
also been used to treat urinary tract infection, calculi
and prostatitis. The aqueous plant extract is used as
anti-inflammatory, anti-epileptic, diuretic, antiemetic and purifying agent. It also has significant
application in treating dysentery, dropsy and
secondary syphilis.2 Cynodon dactylon has been used
as an antidiabetic agent in traditional system of
medicine in India. The many investigation reported
the
hypoglycemic,
hypolipidemic
and
antihyperglycemic activity of the aqueous extract of
Cynodon dactylon. Although Cynodon Dactylon
(Cyn. Dac) has been used in the folk medicine of
many countries, however it is reported the plant
effect against ischemia/reperfusion (I/R)-induced
arrhythmias. In that study, probable anti-arrhythmic
effect of Cyn.Dac was investigated in isolated rat
hearts and finding showed marked protective effects
of Cyn.Dac against I/R-induced arrhythmias in
isolated rat hearts. Regarding the presence of
Vol. 3 (2) Apr – Jun2012
flavonoid
glycosides
confirmed
during
phytochemical screening of the extract and their role
in the scavenging of oxygen free radicals, it seems
one of the potential cardioprotective mechanism of
the plant is anti-inflammatory properties. Probably,
metabolic and/or direct mechanism(s) may also
involve.3
It has been shown that flavonoid compounds present
in various plants may produce beneficial effects in
cardiovascular diseases such as atherosclerosis,
coronary artery disease and hypertension4. Diseases
of the cardiovascular system implicate lifestyle. Nongenetic environmental factors, such as diet can
produce hypertension. Fructose is widely present in
numerous foods. It has been commonly used as a
sweetener and promoted as being useful for weight
reduction, exercise endurance, and diabetes 5. It has
been demonstrated that hypertension develops when
normal rats are fed a fructose-enriched diet 6 - 9.
Hypertension develops in normal rats fed a high
fructose diet as early as 2 weeks after initiation of the
diet. The hypertension is accompanied by the
metabolic abnormalities of hyperinsulinemia, insulin
resistance and hypertriglyceridemia.
The aqueous extract of aerial parts of C. dactylon has
also been shown to reverse the high blood pressure
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International Journal of Research in Pharmaceutical and Biomedical Sciences
and hypertriglyceridemia developed due to fructose
feeding in rats.
Thus the present study was aimed to evaluate the
effects of aqueous extract of aerial parts of C
dactylon and to determine whether the extract could
prevent the development of hypertension and some
related abnormalities induced by a high fructose diet
in normal rats.
MATERIAL AND METHODS
Animals
Male Wistar rats weighing 200-250 g were used in
this study. The rats were housed in an
environmentally controlled room with a 12-h light:
12-h dark cycle and free access was allowed to
normal rat and tap water. The rats were in
stabilization to acclimatize to the housing conditions
for 1 week. After that, the rats were randomly divided
into groups of six rats each. Fructose treated rats
were given 10% fructose to drink ad libitum for 3, 4
or 6 weeks. Fructose solution was prepared everyday
by dissolving the fructose in tap water. Ordinary tap
water was given to control animals to drink
throughout the whole experimental period.
Rats were weighed previously to dietary
manipulation and at the end of the study. The rats
were trained to the procedure of systolic blood
pressure measurement at 1300 h, twice a week, at
week 5 and 6, and the mean of three to four
consecutive readings was used as the reported value
of the systolic blood pressure for each rat. Indirect
systolic blood pressure (SBP) was measured by
means of the tail-cuff method using a MLT-125R
pulse transducer connected to ML125- NIBP
controller (model S/N N10337). At week 3 & 6 and
also after acute treatment of Aq. Extract of C.
dactylon, blood samples were collected from the
retroocular plexus in fasting conditions (5 h.), and
centrifuged at 4 °C( Remi’s CM 12 cooling
microfuge, Remi Instruments division, Vasai, India).
Plasma glucose, cholesterol and triglyceride levels
were measured by means of spectrophotometry (UV
spectrometer, V-630, Jasco) and commercial kits
(One touch Horizon blood glucometer, Lifscan
Inc,Militias,CA,95035,USA, Cholesterol-wybenga &
pileggi, bioLab Diagnostic Pvt. Ltd, Batch
no.2034T,triglyceride GPO-PAP KIT, Batch no2257T, BioLab Diagnostic Pvt. Ltd, Biosar, MS.).
Preparation of extract
Cynodon dactylon was collected from the available
sources. It was identified and authenticated by
Agharkar Research Institute; Pune.11 The green plant
up to 500 g was extracted with boiling water for 48 h.
The resulting extract was filtered and concentrated in
Vol. 3 (2) Apr – Jun2012
ISSN: 2229-3701
rota-vapour under reduced pressure. Resulted extract
was collected and stored at cool temperature.
Toxicological Study
Acute Oral Toxicity of aqueous extract
Acute Oral Toxicity was carried out according to
OECD guidelines. Six albino mice (female) weighing
between 25 to 40 was used. No any symptoms occur
in 4 hr after drug administration. No mortality occurs
after 24 hr. above study was done at dose 2000 mg/kg
and 5000 mg/kg body weight.
Preliminary Pharmacognostical Study
Standard screening test of the aqueous extract was
carried out for various plant constituents. The powder
was screened for the presence or absence of
secondary metabolites such as alkaloids, flavonoids,
tannins, glycosides, saponins etc. using standard
procedures
General procedure
SBP and pulse rate in conscious rats were measured
every week by the indirect tail-cuff method using an
MLT-125R
pulse
transducer
ML125-NIBP
controller. The animals were placed in the restrainers
(MLA 5022 rodent restrainer for rats) 15 min prior to
the readings being taken. In this method, the
reappearance of pulsation on a digital display of the
blood pressure cuff is detected by a pressure
transducer and is amplified and recorded digitally as
the SBP. During the measurement, seven individual
readings were obtained in a rapid sequence. The
highest and the lowest readings were not considered,
and the average of the remaining five readings was
retained. SBP and heart rate were measured in acute,
preventive and curative experiments.
After induction of hypertension in selected animals
by 10% fructose solution for 6 weeks, animals are
divided into different groups i.e. control(vehicle
treated), losartan(10mg/kg) Aq. Extract of C.
dactylon (100, 200 mg/kg) .The aqueous extract of
aerial parts of C. dactylon or vehicle were
administrated daily by p.o route for 15 days.
Chronic treatment
At the end of 6 weeks of chronic fructose treatment,
the first assessment of the effect of acute C. dactylon
treatment on SBP and heart rate was done. Twentyfour rats were divided into four equal groups two
control groups were given either distilled water p.o.
or 10 mg/kg of losartan i.p and two test groups were
given 100 or 200 mg/kg of extract per kg body
weight p.o. The level of SBP and heart rate were
measured before and 1 & 2 week after a single dose
of the plant extract. At the end of 15 days chronic
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International Journal of Research in Pharmaceutical and Biomedical Sciences
treatment of extract, animals were prepared for
invasive assessment of BP.
Animal preparation
On the day of the experiment, rats were fasted for 5
h,
weighed
and
anaesthetized
with
urethane(1.25gm/kg).The left carotid artery was
cannulated with a polyethylene PE-50 cannula loaded
with heparinized saline solution (15U/ml).The intraarterial cannula was connected to a MLT-844 grass
physiological pressure transducer ( ADInstruments,
unit 13, 22 Lexington rive, Bella Vista, NSW 2153,
Australia) coupled to a ML221 ADInstruments
Bridge Amplifier (ADInstruments, NSW 2153,
Australia) with MLAC11 Grass adapter cable from
ADInstruments for mean arterial pressure (MAP) and
heart rate (HR) record to 8 channel powerlab with
labchart provision(model S/N 830-1905). Also
jugular vein was cannulated with heparinized
polyethylene cannula for the administration of drugs.
After 30min of stabilation of BP, pressor response to
adrenalin (0.5, 1.0 µg/kg) and noradrenalin (0.5, 1.0
µg/kg) was recorded.
Statistical analysis
All the data were expressed as mean±/S.E.M.
Statistical differences were calculated for each of the
above mentioned parameters using the Student’s t test for multiple comparisons. A probability level of
P≤/0.01 was taken to indicate a significant difference
between means.
RESULTS AND DISCUSSION
The preliminary phytochemical tests were showing
the presence of Alkaloids, Cardiac Glycosides, and
Steroids.
General parameters
Basal values of body weight, triglycerides, glucose,
SBP in conscious rats and MAP and HR in
anaesthetized rats are shown in Table 2. As
previously observed, body weight and HR did not
differ between C and F groups, meanwhile SBP,
MAP, plasma triglyceride and glucose levels were
increased in the F group (p≤0.05).
Effect of Aq. Extract on hypertensive rats
The chronic administration 100mg/kg, 200mg/kg of
Aq. Extract of C. dactylon and losartan 10 mg/kg
produced a significant (p ≤0.01) hypertensive effect.
(fig 1)
The pressor response to adrenalin (0.5, 1.0 µg/kg)
(fig 2) was significantly reduced while pressor
response to NA (0.5, 1.0 µg/kg) was reduced in
animals treated with chronic treatment with extract
and losartan compared with control group (fig 3). The
Vol. 3 (2) Apr – Jun2012
ISSN: 2229-3701
pressor response to NA and adrenaline was
significantly increased in fructose treated group as
compared with control group.
High blood pressure is a major risk factor for stroke,
coronary heart disease & renal vascular diseases. The
current & common method for controlling
hypertension is the use of long term treatment drug
therapy. It is well known that drugs have many side
effects which May complicate the patient’s medical
condition. This is why physicians as well as medical
professionals & even patients prefer & trend to use
antique traditional herbal medicines.
C. dactylon the presence of reveals the presence of
flavanoids, sterols & cardiac glycosides. The cardiac
glycosides have been also proved for its
cardioprotective & cardiotonic activity, thus the
significant antihypertensive activity of aq. extact of
C. dactylon may be due to the presence of cardiac
glycosides.
Recent studies have shown that a high fructose diet is
associated with increased blood pressure in rats12 - 15.
Several studies have demonstrated that chronic
fructose feeding leads to insulin resistance, glucose
intolerance,
hyperinsulinmia
and
hypertriglyceridemia in a relatively short time in
normal rats16 - 19. These metabolic changes lead to
essential hypertension20. In the present study, the
effect of the aq. Extract of aerial part of C. dactylon
on hypertension induced by a high-fructose diet given
in drinking water was exanimate.
It is generally agreed that fructose loading induces
insulin resistance and glucose intolerance. However,
the effects of fructose treatment on plasma glucose
levels reported by different researchers varied from
no change transient elevation, to a significant. The
present study showed that there was significant
change in plasma glucose levels in Wistar rats
following fructose treatment during 3 or 6 weeks
Hypertriglyceridemia in fructose-treated rats has
been demonstrated by several workers. In this study,
plasma triglyceride and cholesterol levels were
significantly decrease, respectively, in preventive
treatment with the plant extract at 100 and 200
mg/kg. Hypertriglyceridemia has been proposed to be
caused by either increased hepatic secretion of verylow-density lipoprotein-triglyceridc (VLDL-TG) or a
decreased removal of triglyceride rich lipoprotein
from the circulation. In the present study, the
combination of fructose and C. dactylon extract or
losartan produced a marked decrease in plasma
triglyceride levels.
The present study showed that C.dactylon attenuated
the decrease in plasma insulin levels induced by the
high fructose diet; inhibit the increase in plasma
triglycerides and cholesterol level. The present study
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International Journal of Research in Pharmaceutical and Biomedical Sciences
does not rule out cleanly the effect of the extract on
metabolic alterations induced by high fructose-diet.
Systolic blood pressure measuring by tail cuff
method in high fructose treated rats and preventive
and curative treated rats with C.dactylon shows
significant decrease in systolic BP in C.dactylon
treated rats compared with high fructose treated rats.
The effect of C.dactylon on high fructose treated and
preventive and curative treated rats were studied by
invasive method. It was found that chronic preventive
treatment of C, dactylon in high fructose treated rat’s
shows significant decrease in mean arterial BP
compared with high fructose treated rats.
Furthermore, pressure effect of Adrenaline and
Noradrenalin was also studied in high fructose treated
rats and curative treated groups.
In Fructose model, the present study shows that after
drinking 10% fructose solution for six weeks, normal
rats exhibited significant increase in BP.
Treatment with aq. Extract of C.dactylon blocked the
continued elevation of BP. Vascular reactivity to
different known agonists i.e. Adr and NA shows
depressor response with of may be alpha or beta
ISSN: 2229-3701
receptor blocked. The vascular reactivity of NA &
Adr was enhanced in hypertensive rats. The
difference in the vascular reactivity to Adr & NA
may be due to predominance f the beta action of Adr,
which may mask the presser response of the alpha
action. But the predominant beta action of Adr can
only see at high doses. The chief vascular action of
Adr is exerted on small arterioles and precapillary
sphincters, although veins & large arteries also
respond to the drug. Vascular hyper responsiveness
to NA & Adr has been reported in rats with fructose
induced hypertension. AECD treatment prevents the
development enhanced vascular responsiveness to
NA & Adr. Since NA may increase cytoplasmic Ca++
levels by both increasing the influx of extracellular
Ca++
through receptor operated channels & by
causing the release of intracellular Ca++ there may be
an alteration in the coupling of the receptor to the
mobilization of Ca++ .
Further studies are needed to explain the effect of the
extract on the metabolic changes induced by a high
fructose diet.
Table 1: Chemical tests for chemical constituents of plant extract
Plant
Cynodon
dactylon
Tannins
Saponins
Test
Flavonoids
Steroids
-
-
-
+
Cardiac
glycosides
++
Meyer’s
test
Alkaloids
Wagner’s
Dragendorff’s
test
test
++
As per the chemical tests performed on the Aq. Extract of C. dactylon. It was found that, Aq. Extract of contains steroids, cardiac glycosides, and alkaloids.137
Table 2: Body weight, Triglyceride, Glucose, Cholesterol level in conscious rats
after 6 wks fructose treatment and 15 days chronic preventive treatment
Experimental
group
Body Weight
(gm)
Triglyceride
(mg ml-)
Glucose
(mg ml-)
Cholesterol
(mM)
Systolic BP
After 6 wks
CONTROL
FRUCTOSE
275.5+2.52
After 15 days chronic treatment
275.3+4.02*
FRUCTOSE +
EXR-100
259+6.56#
FRUCTOSE + EXR200
252.16+7.51##
Losartan
253.5+7.65##
0.48+0.01
1.09+0.02*
0.64+0.05#
0.5+0.03##
0.53+0.03##
1.21+0.01
1.54+0.01*
1.40+0.02#
1.24+0.01##
1.25+0.02##
1.26+0.01
1.74+0.02*
1.16+0.01#
1.05+0.008##
1.17+0.01##
99.5+0.76
132.5+1.2
106+1.48
97.5+0.67
99+0.96
Values are mean + S.E.M., N-6 rats per group; * indicates a significant difference (P≤0.01) vs. control group;
#,## indicates a significant difference(P≤0.05) vs. fructose only group.
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International Journal of Research in Pharmaceutical and Biomedical Sciences
ISSN: 2229-3701
* P < 0.01 as compared with control group, # P < 0.01 as compared with Fructose hypertensive rats. Vertical lines represent SEM, n=6.
Fig. 1: Mean arterial blood pressure (mm Hg) after completion of treatment schedule in (1)control, (2) 10%
Fructose, (3) 10% Fructose + 100 extract and (4) 10% Fructose + 200 extract (5) losartan treated groups
* P < 0.01 as compared with control group. # P < 0.01 as compared with Fructose hypertensive rats. Vertical lines represents SEM, n=5.
Fig. 2: Mean change in blood pressure to Adrenalin in (1) control, (2) 10% Fructose,
(3) 10% Fructose + EXT 100 and (4) 10% Fructose + EXT 100 ( 5)LOSARTAN treated groups
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International Journal of Research in Pharmaceutical and Biomedical Sciences
ISSN: 2229-3701
* P < 0.01 as compared with control group. # P < 0.01 as compared with Fructose hypertensive rats. Vertical lines represent SEM, n=6.
Fig. 3: Mean change in blood pressure to Noradrenalin in (1) control, (2) 10% Fructose,
(3) 10% Fructose + 100 extract and (4) 10% Fructose + 200 extract (5) losartan treated groups
CONCLUSION
Findings of this study suggest that C. dactylon is able
to reduce Blood pressure in Fructose induced
Hypertension. Chronic fructose treatment in rats was
administered to elevate blood pressure in association
with increased glucose level and triglyceridemia. The
Wistar rats who received 10% fructose solution for
3&/6 weeks showed significant increase not only in
plasma glucose and cholesterol levels but also in
MABP. Chronic administration of Aq. Extract of C.
dactylon(100, 200mg/kg) in hypertensive rats
significantly reduced mean arterial BP(p≤0.01). The
extract also shows significant decrease in pressor
response to NA (0.5, 1.0 µg/kg) and Adrenalin (0.5,
1.0 µg/kg). These results suggest that the Aq. Extract
of C. dactylon exerts is antihypertensive effect by
partly improving peripheral vascular resistant.
ACKNOWLEDGMENTS
Authors are thankful to Seth Govind Raghunath
Sable College of pharmacy, Saswad for providing
research facilities.
4.
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