Scientific Validation of Antidiabetic Activity of Ethanol Extract of Ficus glomerata Barks and Roots in Streptozotocin-Induced Diabetic Rats

Article Information Received 15 December 2013 Received in revised form 19 Feb 2014 Accepted 23 Feb 2014 Abstract The tribal people of Chhattisgarh state of India use Ficus glomerata for the treatment of diabetes therefore we planned to investigate antidiabetic effect of bark and root extract of bark and root in Streptozotocin (STZ)-induced diabetic rats. Oral administration of ethanol extract of bark and root of Ficus glomerata at the doses of 200 and 400 mg/kg body weight was studied in STZinduced diabetic rats. After administration of extract the fasting blood glucose levels were significantly decreased. The oral glucose tolerance was also improved. The effect of extracts on serum lipid profile like total cholesterol, triglycerides, low density lipoprotein (LDL) and high density lipoprotein (HDL) were also measured in the diabetic and non diabetic rats. There was significant reduction in total cholesterol, LDL and improvement in HDL in diabetic rats were observed. The activity of the plant extracts was comparable with glibenclamide, a well-known antidiabetic drug. These results indicated that Ficus glomerata possesses significant antidiabetic and antihyperlipidemic effect.


Introduction
Medicinal plants continue to provide valuable therapeutic agents, both in modern and in traditional medicine.As powders, extracts, decoctions, or infusions, plants are being used in the traditional systems of medicine in many parts of the world, especially in rural communities, for the control, management, and/or treatment of a variety of human and animal ailments.It is considered that herbal medicines are safer than allopathic medicines; hence there is increase in demand of herbal medicines in western countries 1 .
Every year the number of diabetic patients is growing alarmingly all over the World.Diabetes mellitus and its complications are becoming a global burden and have to be dealt with firmly 2 .Diabetes mellitus is characterized by hyperglycemia and disturbances in carbohydrate, protein and fat metabolisms, secondary to an absolute or relative lack of the hormone insulin.Besides hyperglycemia, several other factors including dyslipidemia or hyperlipidemia are involved in the development of micro and macrovascular complications of diabetes, which are the major causes of morbidity and death 3 .Most of the hypoglycemic agents used in allopathic medicines are reported to have side effects in the long run.Currently World Health Organization has recommended the development of herbal hypoglycemic agent.The uses of herbal medicines in management of diabetes are exhibiting more beneficial than allopathic drugs, due to its safe and cost effective 2 .We planned to develop new herbal antidiabetic agents with minimal or no side effects.
Ficus glomerata belongs to the family Moraceae (Mulberry family), and is commonly known as Gular, Umar, Umber, and Udumbara.It is an herb found all parts of India.It is an evergreen tree; common throughout the State near villages and along streams and rivers; also planted along road-side.Phytochemical investigated on Ficus glomerata have reported the presence of cycloartenol, euphorbol, hexacosanate, triacetate, taraxerone, tetratriterpene, glauanolacetate, racemosic acid, glauanol, glucose, hentriacontane.
Plant is commonly used in antihyperlipidemic, diabetic, hepatoprotective, analgesic and wound healing 4,5 .The aim of this investigation was to ascertain the scientific basis for the use of this plant in the management of diabetes and hyperlipidemic, using streptozotocin induced diabetic rats.

UK Journal of Pharmaceutical and Biosciences
Available at www.ukjpb.com 2 Materials and Methods

Plant material
The bark and root of Ficus glomerata was collected from the forest area of Raipur (Chhattisgarh).The plant was authenticated by botanist Dr. Sumeet Gairola plant systematic section, plant biotechnology division, I.I.I.M, Jammu, India and a voucher specimen was deposited in the Herbarium of Department of Botany, IIIM Jammu (Acc.no.22231).After authentification, plant material was dried at room temperature until it was free from the moisture.Finally barks and roots were subjected to size reduction to get coarse powder.

Preparation of Ficus glomerata ethanol extracts
The powder of the bark and root of Ficus glomerata, was packed separately in the Soxhlet apparatus and extracted with ethanol, until the completion of the extraction.The extract was filtered while hot, and the resultant extract was distilled in vacuum under reduced pressure in order to remove the solvent completely, and later dried in a desiccator.After that ethanol extract of bark (EFGB) and root (EFGR) were kept in air tight container for further study.

Animals
Male Wistar albino rats having a weight of 170 -220 g were kept in quarantine for 10 days under standard husbandry conditions (27.3 o C, Relative humidity 65 ± 10%) for 12 hours in dark and light cycle, respectively, and were given standard food and water ad libitum.The study was permitted by the Institution Animal Ethical Committee.

Oral glucose tolerance test (OGTT)
The oral glucose tolerance test 6 was performed in overnight fasted (18 hours) normal rats.The rats were divided into seven groups (n = 6).Group I served as normal control rats, administered drinking water daily; Group II had glucose control rats; Group III rats were administered standard drug Glibenclamide (0.5 mg/kg); Group IV rats were administered EFGB (200 mg/kg); Group V rats were administered EFGB (400 mg/kg); and Group VI rats were administered EFGR (200 mg/kg); and Group VII rats were administered EFGR (400 mg/kg).Glucose (2 g/kg) was fed to rats of Group II to Group VII, 30 minutes prior to the administration of the extracts and standard drug.Blood was withdrawn from the retroorbital sinus after 0, 30, and 90 minutes of extract and standard drug administration, and the plasma obtained after centrifugation at 3000 rpm was estimated for fasting plasma glucose levels using a glucose oxidase-peroxidase glucose estimation kit.

Induction of non-insulin dependent diabetes mellitus (NIDDM)
Non-insulin dependent diabetes mellitus was induced 7,8 in overnight fasted adult Wistar strain albino male rats weighing 170 -220 g by a single intraperitoneal injection of 60 mg/kg Streptozotocin, 15 minutes after i.p. administration of 120 mg/kg of nicotinamide.
Streptozotocin (STZ) was dissolved in a citrate buffer (pH 4.5) and nicotinamide was dissolved in normal saline.Hyperglycemia was confirmed by the elevated glucose levels in plasma, determined at 72 hours and then on day 7, after injection.The threshold value of fasting plasma glucose to diagnose diabetes was taken as > 126 mg/dl.Only those rats that were found to have permanent NIDDM were used for the study.

Experimental design
The animals were segregated into five groups of six rats each.The extract was administered for 28 days.Group I served as normal control rats, administered drinking water daily for 28 days; Group II had diabetic control rats, administered drinking water daily for 28 days; Group III diabetic rats were administered standard drug Glibenclamide (0.5 mg/kg); Group IV diabetic rats were administered EFGB (200 mg/kg); Group V diabetic rats were administered EFGB (400 mg/kg); and Group VI diabetic rats were administered EFGR (200 mg/kg); and Group VII diabetic rats were administered EFGR (400 mg/kg) for 28 days.The fasting glucose levels were determined on days 0, 7 th , 14 th and 28 th of extract administration.During the experimental period, the rats were weighed daily and the mean change in body weight was calculated.

Estimation of biochemical parameters
The biochemical parameters were determined on day 12 after the animals were sacrificed by cervical dislocation.Total cholesterol, triglycerides (TGL), high-density lipoprotein (HDL) and low-density lipoprotein (LDL), were determined by the glucose oxidase method, using an auto-analyzer 9 .

Statistical analysis
The results are expressed as mean ± SEM of six independent experiments.Statistical significance between the groups was evaluated by one-way analysis of variance (ANOVA) followed by Dunet's test.A P < 0.05 value was considered as statistically significant.

Oral glucose tolerance effects
The effects of ethanol extracts of Ficus glomerata on the plasma glucose level are shown in Table 1.After administration of glucose in rats the rise in glucose level was observed in glucose control, extract treated and standard group.In rats treated with barks and root extracts of Ficus glomerata, there was a significant reduction in plasma glucose level, while in glucose control rats the plasma glucose level increased.Meanwhile same results were observed in glibenclamide treated group.2.

Effect on non-insulin dependent diabetes mellitus
The animals treated with streptozotocin namely Group II, a significant increase in serum glucose level was observed on 0, 7 th , 14 th and 28 th day when compared with normal group rats (Group I).
The Group III received glibenclamide (0.5 mg/kg p.o.) showed significant decrease in serum glucose level when compared with diabetic control rats.After the oral administration of EFGB in diabetic control rats, a significant reduction in blood glucose level was observed when compared with diabetic control rats.Moreover the administration of EFGR in diabetic control rats, also significantly decreased the serum glucose level compared with diabetic control rats.The outcomes exhibited that EFGB and EFGR at dose of 400 mg/kg body weight significantly decreases the blood glucose level of diabetic rats on 7 th day.While the EFGB and EFGR at dose of 200 mg/kg body weight significantly decreases the blood glucose level of diabetic rats on 14 th day.From results it has been observed that the bark extracts showed maximum activity as compared to root extract.

Anti-hyperlipidaemic activity
The outcomes of lipid profiles in control and experimental rats are exhibited in table 3. The rats of diabetic control showed significant increase in serum TGL, total cholesterol and LDL while increase in HDL when compared with normal.The rat treated with glibenclamide also reduced TGL, total cholesterol, LDL, and increased HDL when compared with diabetic control group.The EFGB and EFGR showed significant decrease in total cholesterol, LDL, Triglycerides and significant increase in HDL when compared with diabetic control group.All these effects were observed on day 28 th .From result of lipid profile it has been observed that the barks extract exhibited maximum antihyperlipidaemic activity on compared with roots extract.The present experimental result indicated that ethanol extracts of barks and roots exhibited a potent blood glucose lowering properties in STZ diabetic rats.
During the study, the body weights of rats before and after induction of diabetes, and after treatment were measured (Table 4).The results exhibited that decreased in body weight of rats after induction of diabetes, and increased in body weight of rats after treatment with extracts.

Discussions
Ficus glomerata plant has long been used as a folk medicine for treating diabetic, jaundice, analgesic and wound healing.The In the present study, ethanol extract of bark and root of Ficus glomerata had significantly decreased total cholesterol, triglycerides

Samyal
et al.Antidiabetic Activity UK J Pharm & Biosci, 2014: 2(1); 44 Induction of diabetes in experimental rats was confirmed by the presence of a high fasting plasma glucose level.The effect of bark and root extracts of Ficus glomerata, on serum glucose levels of normal and Streptozotocin-induced rats are shown in Table present study was designed to evaluate the effects of ethanol extracts of bark and root in STZ-induced-diabetic rats.In light of the results, our study indicates that EFGB and EFGR have good antidiabetic activity.The EFGB and EFGR significantly reduced the blood glucose level in STZ-induced-diabetic rats as compared to the diabetic control group.Moreover the EFGB and EFGR increased the body weight of diabetic rats.The possible mechanism by which Ficus glomerata brings about its hypoglycemic action in diabetic rat may be by potentiating the insulin effect of plasma by increasing either the pancreatic secretion of insulin from the existing beta cells or by its release from the bound form.Generally, it has been observed that hyperlipidemia is a complication associated with hyperglycemia.During study it was observed increase in total cholesterol, triglycerides, LDL, and decrease in HDL in streptozotocin induced diabetic rats as compared to normal animals.The EFGB and EFGR showed significant reduction in total cholesterol, LDL, Triglycerides and significant rise in HDL when compared with diabetic control group.The potent antidiabetic effect of the plant extract suggests the presence of potent antidiabetic active principles, which produced antihyperglycemic effect in diabetic rats.The outcomes of lipid profile confirmed the potent antidiabetic activity of bark and root of Ficus glomerata.In recent years, considerable interest has been directed towards the investigation of plasma lipids and lipoproteins pattern in diabetes mellitus due to the fact that abnormal lipid level leads to the development of coronary artery disease in diabetic patients 6 .Samyal et al.Antidiabetic Activity UK J Pharm & Biosci, 2014: 2(1); 45

Table 2 : Effect of ethanol extracts of Ficus glomerata on fasting plasma glucose level in rats Group Fasting plasma glucose concentration (mg/dl) Day 0 Day 7 th Day 14 th Day 28 th
Ethanol extract of Ficus glomerata of bark (EFGB), Ethanol extract of Ficus glomerata of root (EFGR), Values are expressed as mean ± SEM (Number of animals, n=6); significantly different at a P<0.05 when compared with normal control group, *P<0.05 when compared with diabetic control group

Table 4 : Effect of ethanol extracts of Ficus glomerata on changes in body weight in rats
Computation of in vivo antidiabetic activity of Holarrhena antidysenterica seeds extracts in streptozotocin-induceddiabetic rats.UK Journal of Pharmaceutical and Biosciences.2013; 1(1); 11-17.9. Maurya AK, Tripathi S, Kahrana M, Ahmed Z, Sahu RK.