Academic Sciences International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 5, Suppl 4, 2013

Research Article

ACUTE ORAL TOXICITY AND HYPOGLYCAEMIC STUDY OF ETHANOLIC EXTRACT OF

PORTULACA OLERACEA (WHOLE PLANT) IN SWISS ALBINO MICE

SABEEHA SHAFI*, NAHIDA TABASSUM

Department of Pharmaceutical Sciences, University of Kashmir, Hazratbal, Srinagar, Kashmir, Jammu & Kashmir India.
Email: [email protected]
Received: 09 Sep 2013, Revised and Accepted: 08 Oct 2013
ABSTRACT
Objective: To carry out a preliminary phytochemical screening, acute oral toxicity and hypoglycemic study of 50% ethanolic extract of Portulaca
oleracea (whole plant) in swiss albino mice.
Methods: Preliminary phytochemical screening was done as per standard procedures. Acute oral toxicity study was conducted as per OECD
423 guidelines, and method of Babu. The doses taken were 500 mg/kg,1000 mg/kg,1500 mg/kg and 2000 mg/kg b.w. The doses were given
only once and the observations were made for 72 hours. Different behavioural parameters like grooming, hyperactivity, sedation,
respiratory arrest, convulsions, increased and decreased motor activity and mortality were observed. The hypoglycaemic activity of the
50% ethanolic extract was also evaluated. In this activity the doses taken were 200 and 400mg/kg b.w. The doses were given once daily for
14 days. The hypoglycaemic activity was assessed by estimating the serum glucose levels. Both the studies were compared with the Normal
Control groups.
Result: Phytochemical screening revealed that the whole plant extract of Portulaca oleracea contain alkaloids, tannins, glycosides carbohydrates,
flavonoids, terpenes, phenolics, saponins, proteins and steroids. In acute oral toxicity study, the animals showed behavioural changes also.50% of
the animals died at the dose level of 500 mg/kg b.w and 100% animals died at the dose levels of 1000,1500 and 2000mg/kg b.w, thereby indicating
that the dose below than 500mg.kg b.w is safe for further studies. In hypoglycemic activity the dose of 400mg/kg b.w showed a highly significant
reduction of serum glucose levels.
Conclusion: These Preliminary tests indicate that the 50% ethanolic extract of Portulaca oleracea (whole plant) at the dose level of 500 mg/kg can
be used for further pharmacological research and at the dose level of 400mg/kg has significant hypoglycemic activity.
Keywords: Ethanolic extract, Acute oral toxicity, Phytochemical screening, Hypoglycemic activity.

INTRODUCTION is reported that it can be used as a salad and cooked like soups. It is
used to treat burns earache, insect stings, inflammation, skin sores,
Diabetes is silent killer that kills one person every 10 seconds. India ulcers, pruritis (itching skin) eczema and abscesses, in treatment of
is having the highest number of diabetics in the world. It is a cardiovascular disorders, dysuria, haematuria, gonorrhoea,
multifactorial disease which is characterized by hyperglycemia, dysentery, sore nipples and ulcers of mouth. It is used as blood
lipoprotein abnormalities, raised basal metabolic rate, defect in purifier. Roasted seeds are reported to be diuretic and anti-
reactive oxygen species scavenging enzymes and high oxidative dysenteric. Omega 3– fatty acids [9] present are used in the
stress induced damage to pancreatic beta cells It is associated with production of compounds that effect blood pressure clotting, the
long term complications, including retinopathy, nephropathy, immune system, lower cholesterol (LDL) and prevent certain
neuropathy and angiopathy and several others[1]. People with cancers and control coronary spasms. They have positive effect on
diabetes are 2-4 times more likely to develop heart diseases[2]. brain and in such conditions as depression, bipolar disorder,
Efforts are ongoing to understand and manage diabetes mellitus Alzhemiers disease, schizophrenia, hyperactivity and migrane.
because the disease and disease related complications are increasing Reported Pharmacological Activities include antifungal[10],
day by day. In spite of presence of large number of medicines in the antibacterial[11], analgesic, anti-inflammatory [12][13], gastric
pharmaceutical market, remedies from medicinal plants are used antiulcerogenic [14], bronchodilatory [15], skeletal muscle relaxant
with success to treat this disease. There has been great demand for [16],antihypertensive [17], neuropharmacological [18], wound
plant products due to low cost, easy availability and lesser side healing [19], antioxidant [20], antifertility [21] and antitumour
effects. For this plant materials are continuously scrutinized and activities [22].Therefore, with reference to traditional and reported
explored for their effect as antidiabetic and hypoglycaemic activity. uses, the present study was undertaken to investigate the
The acute oral toxicity test aims at establishing the therapeutic index, hypoglycaemic activity of this plant and give a scientific rational for
i.e. the ratio between the pharmacologically effective dose and the its use.
lethal dose on the same strain and species (LD50/ED50). The
greater the index the safer the compound and vice versa. However, MATERIALS AND METHODS
the term acute oral toxicity is most often used in connection to
lethality and LD determinations [3][4]. The evaluation of the toxic Plant Material
action of the plant extracts is important in order to consider a The whole plant of Portulaca oleracea (Family Portulacaceae ) were
treatment safe. collected from Nishat area of the district, Srinagar, during the
Portulaca oleracea, belonging to Family Portulacaceae (Purslane months of April to June and authenticated by a plant taxonomist in
family) is commonly called as Common Purslane in English, as Kurfa the Centre of Plant Taxonomy, University of Kashmir, Srinagar. The
in Mumbai, as Loni, Ghol in Gujrati, as Kursa, Chhota Lunia in identification was done on the basis of the characters described by
Hindi,as Lonak in Punjabi and as Nunar in Kashmiri. It is a Kirtikar and Basu, 1935. A sample of the plant material was
cosmopolitan weed in warm temperate, tropical and subtropical deposited in the herbarium of the Department of Taxonomy,
regions of the world[5][6]. In Srinagar it grows along waste lands University of Kashmir under voucher specimen number
and in cultivated gardens. It contains carboxylic acids, some gums, 1011(KASH) dated 15-09-2008 for future reference. The plant
fatty acids[7] beta-carotene and volatile oil and Portuloside A, a material was dried in a well ventilated room with outside
monoterpene glucoside and phenolic alkaloids[8]. In folk medicine it temperature ranging between 18 to 320 C.
 Sabeeha et al.
Int J Pharm Pharm Sci, Vol 5, Suppl 4, 389-393

Preparation of the extract Hypoglycaemic activity

The dried whole plant was coarsely powdered and 500 gm of the The 50 % ethanolic extract of Portulaca oleracea whole plant (PO)
material was allowed to macerate for 48 hrs with 50% ethanol, with was administered orally once daily to mice at the dose levels of
occasional shaking. After 48 hrs, the ethanolic extract was filtered 200mg/kg b.w and 400 mg/kg b.w respectively and compared with
through Whatmans filter paper. The plant material was then Normal Control group. Before initiation of experiment, the mice
macerated again with fresh 50% ethanol and the filtrate obtained were acclimatized for a period of seven days. To study
from the first and the second maceration was then combined and the hypoglycaemic activity, animals of either sex (20-25 g b w) were
solvent was recovered. After the recovery of alcohol, the extract was divided into three groups of six mice each. The treatment was given
then evaporated to dryness. The process was repeated several times as per the following protocol.
and the yield was noted. The extract was refrigerated at 40 C for
future use in experimental studies. Group I- Normal Control ( 2% aqueous gum acacia)

Phytochemical Screening Group II PO (200 mg/kg b.w)

The extract obtained was subjected to qualitative tests for Group III PO (400 mg/kg b.w)
identification of different constituents like tannins, alkaloids, The treatment was continued for 14 days. During this period, mice of
saponins, glycosides, terpenes, phenolics, flavonoids, carbohydrates, Normal Control group received only 2% gum acacia.
proteins and steroids, by using simple and standard qualitative
methods. [23][24][25]. Assessment of hypoglycaemic activity

Pharmacological Study[26][27]28] After 14 days of treatment, the mice were fasted overnight and on
the 15th day the mice were anaethesized with diethyl ether and
Animals and Exposure conditions blood sample from each animal was collected by cardiac puncture, in
Swiss albino mice weighing about 20-25 gm were taken for sterilized centrifuge tubes. The blood samples were allowed to
conducting acute oral toxicity study and for hypoglycaemic activity. coagulate at 30oC. Serum was separated by centrifugation at 2500
The animals were procured from Central Animal House, IIIM (Indian r/min at 30oC for 15 min and subjected to biochemical investigation
Institute of Integrative Medicine) Jammu & were housed in clean using standard test kit to assess serum glucose level [29][30].
polypropylene cages. Before initiation of experiment, the mice were Glucose kit was obtained from Crest Biosystems, Goa, India.
acclimatized for a period of 7 days. Standard environmental Statistical analysis
conditions such as temperature ranging from 18 to 32o C, relative
humidity (70%) and 12 hrs dark/light cycle were maintained in the All the results were expressed as mean ± SEM. One way analysis of
quarantine. All the animals were fed with rodent pellet diet variance (ANOVA) was used for the statistical analysis of data.
(Ashirwad Industries) and water ad-libitum under strict hygienic Students “t” test was used for determining the significance. A
conditions. All procedures were performed in accordance to CPCSEA probability value of p > 0.05 was considered as non significant, *p<
guidelines after approval from the Institutional Animal and Ethics 0.05 – significant, **p< 0.01- highly significant and ***p<0.001 as
Committee (IAEC) of the Department of Pharmaceutical Sciences, very highly significant.
University of Kashmir [No. F-IAEC (Pharm.Sc) APPROVAL / 2008/ 4
RESULTS
Dated Oct 23rd, 2008].
Phytochemical screening (Table 1)
Acute Oral Toxicity study
The phytochemical screening of 50% ethanolic extract of Portulaca
50% ethanolic extract of Portulaca oleracea (whole plant) (PO)
oleracea (PO) carried out by standard procedures revealed the
was screened for acute oral toxicity study. The animals were
presence of tannins, alkaloids, saponins, glycosides, terpenes,
distributed into five groups. Group I, II, III, IV and V.
phenolics, flavonoids, carbohydrates, proteins and steroids.
Group I served as Normal Control and received 2% gum acacia.
The results obtained were comparable and satisfied the standard
Group II received 500 mg/kg b.w (PO), literature.

Group III received 1000 mg/kg b.w (PO) Table 1: Results of Phytochemical screening of Portulaca
oleracea (whole plant)
Group IV received 1500 mg/kg b.w (PO)
Phytoconstituents Results
Group V received 2000 mg/kg b.w (PO). 1 Tannins +
All extracts were given in 2% gum acacia. After the extract 2 Alkaloids +
administration, food was withheld for 2 hours. The extracts were 3 Saponins +
administered in a single dose by using specially designed mice oral 4 Glycosides +
feeding needle. The observations that were recorded during 72 5 Terpenes +
hours were grooming, hyperactivity, sedation, respiratory arrest, 6 Phenolics +
convulsions, increased and decreased motor activity and mortality if 7 Flavonoids +
any. 8 Carbohydrates +
9 Proteins +
Observations made during acute oral toxicity studies 10 Steroids +
Grooming- Clearing the fur and skin of itself or another animal
Acute Oral Toxicity Tests ( 72 hour study) (Table 2)
Hyperactivity- Abnormality and excessive activity; unable to relax
Portulaca oleracea administered at four dose levels (500, 1000,
Sedation- Calm and composed animals without any stress 1500 and 2000 mg/kg b.w) revealed the following effects during the
Respiratory arrest- Rising of head acute toxicity studies conducted in mice for 72 hours. Control mice
which had received 2% of gum acacia showed normal behaviour
Convulsions- Tremor in the tail or paddling of the feet
i) Grooming: After, 48 and 72 hours, no grooming was observed
Increased motor activity at all the four dose levels.
Decreased motor activity ii) Hyperactivity: PO extract at the administered dose levels had
Mortality. no effect on the activity of mice which remained normal.

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Int J Pharm Pharm Sci, Vol 5, Suppl 4, 389-393

iii) Sedation: After 48 hours, sedation was seen in 50 % of animals v) Convulsions: After 48 and 72 hours 50 % of animals in the
at the dose of 500 mg/kg b.w while 100 % of animals in the dose range of 500 mg/kg b.w and 100% animals in the dose
dose range of 1000, 1500 and 2000 mg/kg b.w showed range of 1000,1500 and 2000 mg/kg b.w showed convulsions
sedation,as the animals remained calm and composed without as tremor in the tail and the paddling of the feet.
any stress. vi) Motor Activity: After 48 hours, 50 % of animals at the dose of 500
mg/kg b.w and. 100 % of animals in the dose range of 1000,1500
iv) Respiratory arrest: After 48 hours, 50 % of animals in the
and 2000 mg/kg b.w showed decreased motor activity.
dose range of 500 mg/kg b.w and 100% animals in the dose
range of 1000, 1500 and 2000 mg/kg b.w showed respiratory vii) Mortality: 50 % of animals died at the dose of 500 mg/kg b.w
arrest indicated by raising of head in both animals of these after 72 hours while 100 % of animals in the dose range of
groups. 1000, 1500 and 2000 mg/kg b.w died after 24 hours.

Table 2: Observations during acute toxicity studies of different doses of 50% Ethanolic extract of Portulaca oleracea (whole plant)
S. No. Treatment Observations (Hours)
Grooming Hyperactivity
1 2 3 4 24 48 72 1 2 3 4 24 48 72
1 Normal control + + + + + + + - - - - - - -
2% aqueous gum acacia
2 + + + + + + + - - - - - - -
1 500 mg/kg b.w + + + + - - - - - - - - -
2 + + + + - - - - - - - -
1 1000mg/kg b.w + + + + - - - - - -
2 + + + + - - - - - -
1 1500mg/kg b.w + + + - - - - - - -
2 + + + - - - - - - -
1 2000mg/kg b.w + + + - - - - - - -
2 + + + - - - - - - -
S. No Treatment Observation (Hours)
Sedation Respiratory Arrest
1 2 3 4 24 48 72 1 2 3 4 24 48 72
1 Normal control - - - - - - - - - - - - - -
2% aqueous gum acacia
2 - - - - - - - - - - - - - -
1 500mg/kg b.w - - - - - - - - - - - _ -
2 - - - - + - - - - + -
1 1000mg/kg b.w - - - + + - - - - + -
2 - - - + + - - - - + -
1 1500mg/kg b.w - - - + + - - - - + -
2 - - - + + - - - - + -
1 2000mg/kg b.w - - - + + - - - - + -
2 - - - + + - - - - + -
S. No Treatment Observations (Hours)
Convulsions Increased motor activity
1 2 3 4 24 48 72 1 2 3 4 24 48 72
1 Normal control - - - - - - - - - - - - - -
2% aqueous gum acacia
2 - - - - - - - - - - - - - -
1 500mg/kg b.w - - - - - - - - - - - -
2 - - - - + - - - - - -
1 1000mg/kg b.w - - - - + - - - - -
2 - - - - + - - - - -
1 1500mg/kg b.w - - - + + - - - - -
2 - - - + + - - - - -
1 2000mg/kg b.w - - - + + - - - - -
2 - - - + + - - - - -
S. No Treatment Observation (Hours)
Decreased Motor Activity Death
1 2 3 4 24 48 72 1 2 3 4 24 48 72
1 Normal control - - - - - - - - - - - - - -
2% aqueous gum acacia
2 - - - - - - - - - - - - - -
1 500mg/kg b.w - - - - - - - - - - - - -
2 - - - - + - - - - - - Dead
1 1000mg/kg b.w - - - - + - - - - Dead
2 - - - - + - - - - Dead
1 1500mg/kg b.w - - - + + - - - - Dead
2 - - - + + - - - - Dead
1 2000mg/kg b.w - - - + + - - - - Dead
2 - - - + + - - - - Dead

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Int J Pharm Pharm Sci, Vol 5, Suppl 4, 389-393

Table 3: Results of hypoglycaemic activity of different doses of 50% ethanolic extract of Portulaca oleracea (whole plant)
Group Treatment Serum Glucose Levels (mg/dl)
I Normal Control 91.50± 6.80
II PO 200 mg/kg 60.00± 9.83*
III PO 400 mg/kg 46.83± 5.84**
The observations are mean ± SEM of 6 animals,*p<0.05 and **p<0.01 as compared to that of Normal Control group (One way ANOVA followed by
students “t “test).
*p < 0.05 – Significant, **p < 0.01 – Highly Significant

DISCUSSION ACKNOWLEDGEMENT
Diabetes mellitus is a metabolic disorder characterized by resistance We are highly thankful to University Grants Commission, New Delhi
in the action of insulin, insufficient insulin secretion or both. It is for financial assistance. The facilities provided by the Department of
becoming one of the most common diseases of the world. Type II Pharmaceutical Sciences University of Kashmir for carrying out this
diabetes in young has increased 30 fold over the last 20 years work also need appreciation.
concomitant with increase in obesity. Studies have revealed that all
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