Antidiabetic and Hypoglycaemic Activity of the Crude Extracts of the Plant Leucas Aspera

Abstract

This study indicates that Leucas aspera extracts have good antidiabetic activity. Ethanol and Petroleum ether extracts of Leucas aspera exhibited significant anti-hyperglycemic activities in alloxan induced as well as streptazotocin induced hyperglycemic rats. They can also improve the condition of diabetes as indicated by parameters like body weight along with serum cholesterol and triglyceride levels. The different extracts of the plant Leucas aspera were tested for oral hypoglycaemic and anti-diabetic activity, by glucose oral tolerance test, alloxan induced and streptozotocin induced diabetic rats respectively. The extracts of Leucas aspera have shown significant (P<0.001) increase in glucose tolerance, the maximum effect was given by ethanolic extract. In alloxan-induced diabetic rats also the maximum percentage reduction in blood glucose levels was found to be in ethanol extract. Animals, which received STZ, also showed a significant reduction in body weight, and increase in water and food intake as compared to vehicle control, which is significantly reversed by petroleum ether and ethanolic extracts of Leucas aspera after 21 days of treatment. These results indicate that the plant Leucas aspera possess significant anti – diabetic activity.

Full text

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INTERNATIONAL JOURNAL OF
PHARMACEUTICAL INNOVATIONS RESEARCH ARTICLE
Antidiabetic and Hypoglycaemic Activity of the Crude Extracts of the Plant
Leucas Aspera
Gupta N*a, Subhramanyam EVSb, Sharma R Ka
a NIMS Institute of Pharmacy, Shobha Nagar, Jaipur-303121
b Srinivasa College of Pharmacy, Mangalore, Karnataka, India
Abstract:
This study indicates that Leucas aspera extracts have good antidiabetic activity. Ethanol and Petroleum
ether extracts of Leucas aspera exhibited significant anti-hyperglycemic activities in alloxan induced as
well as streptazotocin induced hyperglycemic rats. They can also improve the condition of diabetes as
indicated by parameters like body weight along with serum cholesterol and triglyceride levels. The
different extracts of the plant Leucas aspera were tested for oral hypoglycaemic and anti-diabetic
activity, by glucose oral tolerance test, alloxan induced and streptozotocin induced diabetic rats
respectively. The extracts of Leucas aspera have shown significant (P<0.001) increase in glucose
tolerance, the maximum effect was given by ethanolic extract. In alloxan-induced diabetic rats also the
maximum percentage reduction in blood glucose levels was found to be in ethanol extract. Animals,
which received STZ, also showed a significant reduction in body weight, and increase in water and food
intake as compared to vehicle control, which is significantly reversed by petroleum ether and ethanolic
extracts of Leucas aspera after 21 days of treatment. These results indicate that the plant Leucas aspera
possess significant anti – diabetic activity.
Key words: Leucas aspera, hypoglycemic activity, antidiabetic activity, diabetes mellitus.
__________________________________________________________________________________
*Corresponding Author:
Nakul Gupta
Email: - nakulmgupta76@rediffmail.com

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Introduction:
Diabetes Mellitus is the name given to a group
of disorders characterised by chronic
hyperglycemia, polyuria, polydipsia,
polyphagia, emaciation, and weakness due to
disturbance in carbohydrate, fat, and protein
metabolism associated with absolute or relative
deficiency in insulin secretion and/or insulin
action.[1] In the recent past many hypoglycaemic
agents are introduced, still the diabetes and the
related complications continue to be a major
medical problem not only in developed
countries but also in developing countries.
Many Indian medicinal plants are reported to be
useful in diabetes.[2,3] Leucas aspera belonging
to the family Labiate is used as anti-
inflammatory, stimulant, in jaundice, cough,
asthma, conjunctivitis, diabetes, malaria,
headache, otalgia, skin diseases, snake bite,
toothache, and wound healing etc. [4] The
present study was undertaken to verify the claim
and evaluate the anti-diabetic activity of the
plant Leucas aspera.
Experimental:
Plant Material
The plants of Leucas aspera were collected
from the local areas around the Mangalore,
Karnataka, India, during the period August-
September and authenticated by botanist of St.
Agnes College, Mangalore. A voucher specimen
(NIMS/2010/NLA) is being maintained in the
laboratory of Phytochemistry and
Pharmacognosy, NIMS Institute of Pharmacy,
Shobha Nagar, Jaipur, India. The whole plant
then, including root, stem, leaves and flower
were shade dried and chopped into small pieces.
Preparation of extracts
The shade dried plant was powdered (300g) and
extracted with three different solvents petroleum
ether, ethanol (99.99%), and water in three
different soxhlet extractors exhaustively for 20-
24 hours. The extracts were concentrated to
dryness under reduced pressure and controlled
temperature (40-500 C) using flash evaporator.
Test animals
Male wistar albino rats (160 – 200 g) were used
in the experiment. Animals maintained under
standard environmental conditions, were fed
with a standard diet (Hindustan Lever, India)
and water ad libitum. The animals were fasted
for 16h before experimentation but allowed free
access to water. Institutional animal Ethics
Committee’s permission was obtained before
starting the experiments on animal.
Effect of Leucas aspera extracts on oral
glucose tolerance in rats:
Male wistar albino rats (160 – 200 g), fasted for
16h before experimentation but allowed free
access to water and were divided into 4 groups
of 6 rats each. Group l control and received
distilled water, Group II, III, and IV received
petroleum ether, ethanol and aqueous extracts
respectively at a dose of 200 mg/kg body weight
as a fine aqueous suspension orally. All groups
were given glucose (2 g/kg body weight, p.o) 30
min after administration of the drug. Blood
samples were collected from the tail vein just
prior to glucose administration and at 30 and 90
min after the glucose loading. Serum was
separated and glucose levels were measured
immediately by glucose-oxidase method. [5]

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Effect of the Leucas aspera extracts on
alloxan-induced diabetic rats
Male wistar rats (180-200g) were made diabetic
by a single i.p. injection of 120mg/kg body
weight of alloxan monohydrate in sterile normal
saline (Insulin dependent diabetes mellitus)[6],
The rats were maintained on 5 % glucose
solution for next 24h to prevent hypoglycaemia.
Five days later blood samples were drawn from
tail vein and glucose levels were determined to
confirm the development of diabetes.
The diabetic rats were divided into five groups,
each containing six animals. Controls rats
(Group I) were given distilled water orally,
while Leucas aspera petroleum ether, ethanol,
and aqueous extracts were given to groups II,
III, and IV respectively, at a dose of 200 mg/kg,
orally. Group V received glibenclamide at dose
of 10 mg/kg. Blood samples were collected
from the tail vein just prior to and of 1h, 3h and
5h after drug administration.[7,8]
Effect of the Leucas aspera extracts on
streptozotocin induced diabetes
Sprague Dawley rats were used, to induce Non-
insulin dependent diabetes mellitus, a single
dose of injection of streptozotocin (90 mg/kg:
i.p.) was given to the 2 days old pups. Another
group of pups received only saline. The animals
were weaned at 30 days and after a period of 3
months; they were checked for fasting glucose
levels to confirm the status of NIDDM. The
animals showing fasting glucose levels > 140
mg/dl were considered as diabetic. The pups
that received saline were considered as control
animals.
The experimental animals were divided in six
groups, six animals in each group. Group I
vehicle control, Group II, Group III and Group
IV NIDDM treated with Leucas aspera
petroleum ether, ethanolic and aqueous extract
200 mg/kg respectively, and group VI treated
with standard drug glibenclamide (10 mg/kg per
day p.o), Treatment was given daily for 3
weeks. At the end of 3 weeks treatment, the
animals were kept on 12 h fasting and the blood
samples were collected from the tail vein.
Serum samples were analysed
spectrophotometrically for glucose, cholesterol
and triglycerides. Body weight, Water intake
(ml/rat/day) and food intake (grams/rat/day)
were also recorded.[9,10]
Histopathological studies
For the histopathological studies, animals were
sacrificed at the end of 3 weeks treatment; after
the collection of blood samples, pancreas, liver
and kidney were removed, washed with cold
saline and preserved in 10% formalin in
buffered form, processed and stained with
hematoxylin and eosin.
Statistical analysis
The results are expressed as mean S.E.M. the
significant of various treatments was calculated
using students t-test.
Results
The extracts of Leucas aspera have shown
significant (P<0.001) increase in glucose
tolerance (table 1). Petroleum ether and ethanol
extracts reduced the glucose levels
approximately near to normal within 90 minutes
of the drug administration but the maximum
effect was given by ethanolic extract, in contrast
to these two groups aqueous extract group did
not reduce the glucose levels significantly.
In alloxan-induced diabetic rats also, Leucas
aspera petroleum ether and ethanol extracts

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have shown considerable reduction in blood
glucose levels. The maximum percentage
reduction in blood glucose levels was found to
be in ethanol extract. The results are graphically
represented in figure 1.
Administration of the STZ (90 mg/kg: i.p.) led
to significantly elevated levels of Serum
glucose, cholesterol and triglycerides in
experimental animals in comparison to vehicle
control. Treatment with Leucas aspera
petroleum ether, ethanol extracts and standard
drug has shown significant reduction in levels of
these parameters as compared to diabetic control
rats (Table 2).
Animals, which received STZ, also showed a
significant reduction in body weight, and
increase in water and food intake as compared
to vehicle control, which is significantly
reversed by petroleum ether and ethanolic
extracts of Leucas aspera after 21 days of
treatment (Table 3).
Discussion
This study indicates that Leucas aspera extracts
have good antidiabetic activity. Ethanol and
Petroleum ether extracts of Leucas aspera
exhibited significant anti-hyperglycemic
activities in alloxan induced as well as
streptazotocin induced hyperglycemic rats. They
can also improve the condition of diabetes as
indicated by parameters like body weight along
with serum cholesterol and triglyceride levels.
The number of functionally intact β-cells in the
islet organ is of decisive importance for the
development course and outcome of diabetes.
The renewal of β-cells in diabetes has been
studied in several animal models. The total β-
cell mass reflects the balance between the
renewal and loss of these cells. It was also
suggested that regeneration of islet β-cells
following destruction by alloxan may be the
primary cause of the recovery of alloxan-
injected guinea pigs from the effects of the
drug.[11] In alloxan-induced diabetes, (-
)Epicatechin[12] and Vinca rosea extracts[13] has
also been shown to act by -cell regeneration.
Similar effects in streptozotocin-treated diabetic
animals were reported by pancreas tonic [14],
ephedrine[15], and Gymnema sylvestre leaf
extracts[16]. In the current studies, the damage of
pancreas in streptazotocin-treated diabetic
control rats and regeneration of β-cells by
glibenclamide was observed. The comparable
regeneration was also shown by ethanolic and
petroleum ether extracts of Leucas aspera.
Conclusion
The data obtained from this study indicates that
the extracts of the plant Leucas aspera are
capable of exhibiting significant
antihyperglycemic activities in diabetic rats.
These extracts showed improvement in
parameters like body weight and lipid profile as
well as regeneration of β-cells of pancreas and
so might be valuable in diabetes treatment.

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References:
[1] Deb L, Dutta A. Diabetes mellitus its
possible pharmacological evaluation
techniques and naturopathy. Int J Green
Pharmacy 2006; 1(7):28.
[2] Kirithikar KR, Basu BD. Indian Medicinal
Plants: Vol.1, International book
distributors, Dehradun, India, 1995; 371 –
372
[3] Nadkarni KM, Nadkarni AK. Indian
Materia Medica: Vol.1, Popular
Prakashan, Bombay, India, 1996; 615-
616.
[4] Khory RN, Katrak NN. Materia and
medica of India and their therapeutics:
Komal Prakashan, New Delhi,
India,1996; 484-485
[5] Li Y, Wen S, Kota BP, Peng G. Punica
granatum flower extract, a potent α-
glucosidase inhibitor, improves
postparandial hyperglycemia in Zuker
diabetic fatty rats. J of Ethnopharmacol
2005; 99:239-44.
[6] Babu V, Ganga DT, Subramoniam A,
Antidiabetic activity of ethanol extract of
Cassia Kleinii leaf in Streptozotocin-
induced diabetic rats and isolation of an
active fraction and toxicity evaluation of
the extract. Indian J of Pharmacol 2003;
35:290-96
[7] Prasanna GS, Patil PA. Effect of Aqueous
Extract of Momordica charantia linn. on
blood glucose level in diabetic rats treated
with and without glibenclamide. Indian
Drugs 2005; 42(11):718-22.
[8] Gupta MP, Solis NG, Avella ME, Sanchez
C. Hypoglycemic activity of Neurolaena
lobata (L.). J of Ethnopharmacol
1984;10:323-327
[9] Akhani SP, Goyal RK. Antidiabetic
activity of Zingiber officinale in
Streptozotocin- induced Non-insulin
Dependent Diabetic rats. Indian J Pharm
Sci 2005; 67(5):553-557.
[10] Nagappa AN, Thakurdesai PA, Venkat
RN, Singh J. Antidiabetic activity of
Terminalia catappa Linn fruits. J of
Ethnopharmacol 2003; 88: 45–50.
[11] Gorray KC, Baskin D, Brodsky J,
Fujimoto WY. Responses of pancreatic b
cells to alloxan and streptozotocin in the
guinea pig. Pancreas 1986; 1:130–138.
[12] Chakravarthy BK, Gupta S, Gode KD.
Functional beta cell regeneration in the
islets of pancreas in alloxan induced
diabetic rats by (-)-epicatechin. Life
Sciences 1982; 31: 2693–2697.
[13] Ghosh S, Suryawanshi SA. Effect of
Vinca rosea extracts in treatment of
alloxan diabetes in male albino rats.
Indian J of Exp Bio 2001; 39: 748–759.
[14] Rao RM, Salem FA, Gleason JI.
Antidiabetic effects of a dietary
supplement ‘Pancreas Tonic’. J of
National Med Assoc 1998; 90: 614–618.
[15] Xiu LM, Miura AB, Yamamoto K,
Kobayashi T, Song QH, Kitamura H,
Cyong JC. Pancreatic islet regeneration by
ephedrine in mice with streptozotocin-
induced diabetes. American J of Chinese
Med 2001; 29: 493–500.
[16] Shanmugasundaram ER, Gopinath KL,
Radha Shanmugasundaram K, Rajendran
VM. Possible regeneration of the islets of
Langerhans in streptozotocin- diabetic rats
given Gymnema sylvestre leaf extracts. J
of Ethnopharmacol 1990; 30: 265–279.

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Table 1: Effect of Leucas aspera extracts on glucose tolerance in rats
Group
Treatment
(Dose/Kg Body Weight)
Blood Glucose (mg/Dl)
Fasting
30 Minutes
90 Minutes
I
Glucose; 2g
77.25 ± 0.907
146.90 ± 1.76
114.71± 1.60
II
LAPE 200 mg + Glucose
78.99 ± 0.83
102.86± 0.87*
82.17±1.02*
III
LAEE 200 mg + Glucose
79.03±0.91
93.81±1.17*
81.52±1.02*
IV
LAAE 200 mg + Glucose
77.65±0.87
116.32±0.89
97.34±1.21
Values are means ± S.E.M., n = 6, * P<0.001 vs. group I,
LAPE: - Leucas aspera petroleum ether extract
LAEE: - Leucas aspera alcoholic extract
LAAE: - Leucas aspera aqueous extract
L. aspera extracts were given orally 30 min before glucose loading
Table 2: Effect of extracts of Leucas aspera on serum profile in diabetic rats (STZ) after treatment
of 21 days
Group
Treatment
S. Glucose
S. Cholesterol
S. Triglycerides
I
Vehicle Control
74.3 ± 1.37
47.08 ± 2.2
46.83 ± 5.5
II
Diabetic Untreated
192 ± 6.58*
79.33 ± 1.5*
122.83 ± 7.1*
III
Diabetic Treated
With LAPE 200 mg
123.6 ±
2.34**
61.16 ± 1.6**
68.33 ± 6.0**
IV
Diabetic Treated
With LAEE 200 mg
112.3 ±
4.17**
56.83 ± 2.9**
60.26 ± 7.9**
V
Diabetic Treated
With LAAE 200 mg
145.8 ± 3.73
68.50 ± 1.4
92.50 ± 6.1
VI
Diabetic Treated
With 10 mg/Kg Of
Glibenclamide
94.6 ±
2.13**
51.83 ± 2.1**
54.00 ± 6.1**
Each value is mean ± SEM (n=6), *significantly different from vehicle control (P < 0.05),
**significantly different from diabetic untreated (P < 0.05)
LAPE: - Leucas aspera petroleum ether extract
LAEE: - Leucas aspera alcoholic extract,
LAAE: - Leucas aspera aqueous extract

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Table 3: Effect of various extracts of Leucas aspera in streptozotocin induced diabetic albino rats
after 21 days of treatment
Each value is mean ± SEM (n=6), *significantly different from vehicle control (P < 0.05)
**significantly different from diabetic untreated (P < 0.05)
385.2
380.87
381.21
375.21
375.32
290.34
284.18
287.14
378.12
278.76
264.81
270.45
388.04
320.87
308.45
315.11
385.42
262.32
251.44
221.18
210
230
250
270
290
310
330
350
370
390
410
0 hr
1 hr
3 hr
5 hr
diabetic untreated
diabetic treated with LAPE
diabetic treated with LAEE
Figure 1- Effect of Leucas aspera extracts on blood glucose levels in Alloxan induced diabetic rats
Group
Treatment
Average Body
Weight
(Gm/Rat)
Water Intake
(Ml/Rat/Day)
Food Intake
(Gm/Rat/Day)
I
Vehicle Control
204.2 ± 3.01
37.91 ± 1.22
16.75 ± 0.45
II
Diabetic Untreated
151.6 ± 3.84*
51.42 ± 1.51*
33.83 ± 0.71*
III
Diabetic Treated
With LAPE 200 mg
175.2 ± 3.14**
42.16 ± 0.36**
23.14 ± 0.56**
IV
Diabetic Rats treated
With LAEE200 mg
182.3 ± 4.17**
41.15 ± 2.9**
21.34 ± 0.92**
V
Diabetic Rats treated
With LAAE 200 mg
154.8 ± 3.73
46.50 ± 1.4
29.35 ± 1.10
VI
Diabetic Rats treated
With 10 mg/Kg of
Glibenclamide
194.6 ± 2.13**
40.83 ± 2.1**
18.13 ± 1.31**

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Figure-2 Microscopy of pancreas stained with haematoxylin and Eosin (H & E)
(magnification × 200)
Vehicle
control
Diabetic
treated
(Glibenclamid
e)
Diabetic
untreated
Diabetic treated
LAEE
Diabetic treated
LAPE
Diabetic treated
LAAE
Vehicle control islets with normal round
and structural intactness with their
nucleus.
Diabetic untreated rat islets damaged and
shrunken in size.
Diabetic rats treated with glibenclamide
orally at a dose of 10 mg/kg, islets
resemble vehicle control rat islets with
normal round and structural intactness
with their nucleus.
Diabetic rats treated with Leucas aspera
petroleum ether extract orally at a dose
of 200mg/kg, islets resemble normal rat
islets but islets enlarged in size.
Diabetic rats treated with Leucas aspera
ethanolic extract orally at a dose of
200mg/kg, islets resemble normal rat
islets and glibenclamide treated group.
Diabetic rats treated with Leucas aspera
petroleum ether extract orally at a dose
of 200mg/kg, islets resemble Group 2 rat
but little improvement is observed.
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