Download Antihyperglycemic activity studies on methanol extract of Petrea

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Advances in Natural and Applied Sciences, C(C): CC-CC, 2010
ISSN 1995-0772
This is a refereed journal and all articles are professionally screened and reviewed
ORIGINAL ARTICLE
Antihyperglycemic Activity Studies on Methanol Extract of Petrea Volubilis L.
(Verbenaceae) Leaves and Excoecaria Agallocha L. (Euphorbiaceae) Stems
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Mahabuba Rahman, 1Aziza Siddika, 1Bithika Bhadra, 1Shahnaz Rahman, 1Bipasha Agarwala,
Majeedul H. Chowdhury, 1Mohammed Rahmatullah
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Department of Biotechnology & Genetic Engineering, University of Development Alternative, Dhanmondi,
Dhaka-1205, Bangladesh
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Present address: New York City College of Technology The City University of New York 300 Jay Street,
Brooklyn, NY 11201, USA.
Mahabuba Rahman, Aziza Siddika, Bithika Bhadra, Shahnaz Rahman, Bipasha Agarwala, Majeedul
H. Chowdhury, Mohammed Rahmatullah: Antihyperglycemic Activity Studies on Methanol Extract
of Petrea Volubilis L. (Verbenaceae) Leaves and Excoecaria Agallocha L. (Euphorbiaceae) Stems
ABSTRACT
Petrea volubilis L. (Verbenaceae, local name: nilmoni lota) is a vinous plant found in various regions of
Bangladesh. Excoecaria agallocha L. (Euphorbiaceae, local name: gewa) is a mangrove species found in the
Sunderbans forest region in the south-western coastal districts of the country. The objective of the present study
was to evaluate the antihyperglycemic potential of leaves of P. volubilis, and stems of E. agallocha, both plant
parts being used in the folk medicinal system of Bangladesh for treatment of a variety of ailments, including
diabetes. Antihyperglycemic activity was studied by oral glucose tolerance tests in glucose-loaded Swiss albino
mice. The methanol extract of leaves of P. volubilis showed dose-dependent and significant reductions of serum
glucose levels in mice, when evaluated at doses ranging from 50 to 400 mg per kg body weight. At a dose
of 400 mg leaf extract/kg body weight, the reduction in glucose level was comparable to that of the standard
antihyperglycemic drug, glibenclamide, administered at a dose of 10 mg/kg body weight. The methanol extract
of stems of E. agallocha demonstrated more potent antihyperglycemic activity compared to glibenclamide,
when examined at doses of 200 and 400 mg per kg body weight. The results indicate that the leaves of P.
volubilis and stems of E. agallocha can be potential sources of antidiabetic drugs.
Key words: Petrea volubilis, Excoecaria agallocha, antihyperglycemic, mice
Introduction
Petrea volubilis L. (Verbenaceae, local name: nilmoni lota) is a vinous plant found in a scattered manner
in various regions of Bangladesh. The plant is comparatively little known and there is a total absence of any
significant scientific studies on its pharmacological properties or its phytochemical constituents. Excoecaria
agallocha L. (Euphorbiaceae, local name: gewa) is a common mangrove species of the Sunderbans forest
region, which covers portions of the three coastal south-western districts of Bangladesh, namely Bagerhat,
Khulna and Satkhira. Both plants are used by the folk medicinal practitioners of Bangladesh for treatment of
a variety of ailments including diabetes.
The constituents of E. agallocha and some of their pharmacological properties have been well described
in the scientific literature. A novel phorbol ester, 12-deoxyphorbol 13-(3E,5E-decadienoate), with anti-human
immunodeficiency virus properties, has been described from leaves and stems of the plant (Erickson et al.,
1995). From the resinous wood of the plant, a number of diterpenes have been reported, of which several
Corresponding Author: Dr. Mohammed Rahmatullah, Pro-Vice Chancellor and Dean, Faculty of Life Sciences
University of Development Alternative House No. 78, Road No. 11A (new) Dhanmondi,
Dhaka-1205 Bangladesh
Telephone: +88-01715032621; Fax: +88-02-815739
Email: [email protected]
Adv. in Nat. Appl. Sci., C(C): CC-CC, 2010
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exhibited anti-tumor promoting activity in vivo in a two-stage carcinogenesis test of mouse tumor using 7,12dimethylbenz[a]-anthracene (DMBA) as an initiator and 12-O-tetradecanoylphorbol-13-acetate (TPA) as a
promoter (Konishi et al., 1998; Konoshima et al., 2001). Antioxidant activity has been reported for methanolic
extract of leaves (Masuda et al., 1999). Triterpenoids, including taraxerone, b-amyrin acetate, taraxerol,
acetylaleuritolic acid, as well as steroids like b-sitostenone and b-sitosterol have been reported from stems and
twigs of the plant (Tian et al., 2008). From the stems and twigs of the plant, diterpenoids have further been
reported with properties of tumor necrosis factor-a inhibitory expression induced by lipopolysaccharide in
mouse macrophages Raw 264.7 cells (Li et al., 2010). Extract of leaves of the plant reportedly demonstrated
anti-plasmodial activity against Plasmodium falciparum (Ravikumar et al., 2010).
Considering the use of the two plants, P. volubilis and E. agallocha in the folk medicinal system of
Bangladesh for treatment of diabetes, we decided to evaluate the antihyperglycemic potential of the plants in
oral glucose tolerance tests in glucose-challenged mice. The leaves of the former and stems of the latter were
selected for antihyperglycemic studies, because these are the plant parts used in folk medicine for their antidiabetic effects.
Materials and Methods
Plant material and extraction
The leaves of P. volubilis were collected from Dhaka district in August 2010. The plant was
taxonomically identified by Bangladesh National Herbarium at Dhaka (Accession Number 35,048). The leaves
of P. volubilis were air-dried in the shade for 120 hours, grounded into a fine powder, and were extracted with
methanol at a ratio of 1:3 (w/v). After 24 hours, the mixture was filtered; filtrate was collected and the residue
was again extracted with methanol at a ratio of 1:2 (w/v) for 24 hours. Filtrates were combined and evaporated
to dryness. The initial weight of dried leaf powder used for extraction was 100g; the final weight of the extract
was 3.94g.
The stems of E. agallocha were collected from the Sunderbans forest region in Satkhira district during
January 2010. The plant was taxonomically identified by Bangladesh National Herbarium at Dhaka (Accession
Number 34,971). Stems were sliced into small pieces, dried and pulverized to a fine powder. 100g of the stem
powder was extracted for 24 hours with methanol at a ratio of 1:4 (w/v). After 24 hours, the mixture was
filtered; filtrate was collected and the residue was again extracted with methanol at a ratio of 1:4 (w/v) for
24 hours. Filtrates were combined and evaporated to dryness. The final weight of the extract was 7.04g.
Chemicals and Drugs
Glacial acetic acid was obtained from Sigma Chemicals, USA; glibenclamide and glucose were obtained
from Square Pharmaceuticals Ltd., Bangladesh. All other chemicals were of analytical grade.
Animals
In the present study, Swiss albino mice (male), which weighed between 14-20g were used. The animals
were obtained from International Centre for Diarrhoeal 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 one week prior to actual experiments. The study was conducted following approval by the
Institutional Animal Ethical Committee of University of Development Alternative, Dhaka, Bangladesh.
Antihyperglycemic activity
Glucose tolerance property of methanol extract of P. volubilis leaves and E. agallocha stems was
determined as per the procedure previously described by Joy and Kuttan (1999) with minor modifications. In
brief, fasted mice were grouped into eight groups. The various groups received different treatments like Group-I
received vehicle (1% Tween 80 in water, 10 ml/kg body weight) and served as control, Group-II received
standard drug (glibenclamide, 10 mg/kg body weight), Groups III-VI received the methanol extract of P.
volubilis leaves at four different doses of 50, 100, 200 and 400 mg/kg body weight; Groups VII and VIII were
administered methanol extract of E. agallocha stems at doses of 200 and 400 mg per kg body weight,
respectively. Each mouse was weighed and doses adjusted accordingly prior to administration of vehicle,
standard drug, and test samples. All substances were orally administered. Following a period of one hour, all
Adv. in Nat. Appl. Sci., C(C): CC-CC, 2010
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mice were orally administered 2 g glucose/kg of body weight. Blood samples were collected two hours after
the glucose administration through puncturing heart. Serum glucose levels were measured by glucose oxidase
method (Venkatesh et al., 2004).
Results and Discussion
The methanol extract of leaves of P. volubilis, when administered to glucose-challenged mice demonstrated
dose-dependent and significant antihyperglycemic activity. The results are shown in Table 1. The concentration
of serum glucose in control mice was 97.01 ± 5.13 mg/dl (mean ± SEM). The standard antihyperglycemic
drug, glibenclamide, administered at a dose of 10 mg per kg body weight, reduced serum glucose levels to
39.80 ± 1.84 mg/dl. The highest reduction in serum glucose levels with methanol extract of P. volubilis leaves
was observed at a dose of 400 mg per kg body weight. At this dose, serum glucose level in mice was
observed to be 41.98 ± 1.52, which was nearly equivalent to that observed with glibenclamide.
Dose for dose, the methanol extract of stems of E. agallocha exhibited greater potency in the reduction
of serum glucose levels. At a dose of 200 mg extract per kg body weight, serum glucose levels in mice was
reduced to 26.25 ± 3.16 mg/dl, which was significantly lower than the level observed with glibenclamide. At
the highest dose evaluated of 400 mg per kg body weight, serum glucose level in mice was further reduced
to 23.29 ± 2.22 mg/dl (Table 1). It appears from the results obtained in the present study that the mangrove
species plant E. agallocha possess considerable antidiabetic potential and can form the basis for further
scientific studies for isolation of antihyperglycemic constituent(s).
Reduction of serum glucose levels by a plant extract can occur by several mechanisms. The extract may
increase the pancreatic secretion of insulin, or the extract may increase the uptake of glucose (Nyunai et al.,
2009; Farjou et al., 1987). It is also possible that the extract may inhibit glucose absorption in gut, which in
turn will reduce the presence of glucose in serum (Bhowmik et al., 2009). The actual mechanism through
which the crude extracts of leaves of P. volubilis and stems of E. agallocha lowered serum glucose levels in
glucose-loaded mice needs further investigations. It is to be noted in this regard that E. agallocha has been
reported to contain b-amyrin acetate (Tian et al., 2008). The compound a-amyrin acetate has been shown to
possess antihyperglycemic properties in several studies (Singh et al., 2009; Narender et al., 2009). Whether
b-amyrin acetate has similar properties like a-amyrin acetate is not clear but needs to be elucidated.
The results obtained in the present study also validate the folk medicinal uses of both plants for treatment
of diabetes in Bangladesh. Indigenous use of plants can form and have often in the past formed the basis of
discovery of modern allopathic drugs. Our study highlights the importance of scientifically evaluating
indigenous uses of antidiabetic plants of Bangladesh. Diabetes is a debilitating disease for which no modern
cure is known, and which affects millions of people in all countries of the world. As such, it is important that
further research is done on medicinal plants that have been used for centuries by indigenous peoples or folk
medicinal practitioners for treatment of diabetes.
Table 1: Effect of methanol extract of P. volubilis leaves and E. agallocha stems on serum glucose level in hyperglycemic mice.
Treatment
Dose (mg/kg body weight)
Serum glucose level (mg/dl)
Control
10 ml
97.01 ± 5.13
Glibenclamide
10 mg
39.80 ± 1.84*
P. volubilis
50 mg
72.87 ± 1.56*
P. volubilis
100 mg
59.60 ± 4.12*
P. volubilis
200 mg
58.73 ± 2.63*
P. volubilis
400 mg
41.98 ± 1.52*
E. agallocha
200 mg
26.25 ± 3.16*
E. agallocha
400 mg
23.29 ± 2.22*
All administrations were made orally. Values represented as mean ± SEM, (n=6 for P. volubilis, n = 7 for E. agallocha); *P < 0.05;
significant compared to hyperglycemic control animals.
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