Alangium salvifolium is a medicinal plant, used by herbalist for treating various diseases, one of which is diabetes mellitus in Central India.

However, its antidiabetic activity has not been scientifically validated so far. The aim of the present study was to evaluate the antidiabetic activity of ethanol extracts of leaves and barks of Alangium salvifolium against strepozotocin (STZ)-induced diabetic rats. The ethanol extracts of leaves and barks of Alangium salvifolium at doses of 200 and 400 mg/kg body weight was administered orally to diabetic rats. The blood glucose levels were monitored at specific intervals and found significantly lowered the blood glucose level. Glibenclamide was used as a standard drug at a dose of 0.25 mg/kg. The effect of extracts on induced hyperlipidemia was analyzed where the extracts significantly lowered the elevated total cholesterol, triglycerides (TGL) and low density lipoprotein (LDL) level while increased the high density lipoprotein (HDL). Moreover, the decreased in body weight of rats after induction of diabetes, and increased in body weight of rats after treatment with extracts was observed. The experimental data exhibited that extract of leaves and barks of Alangium salvifolium has significant antidiabetic activity in streptozotocin-induced rats compared to standard drug. The ethanol extracts of leaves exhibited maximum antidiabetic activity as compared to barks extract.

1

Diabetes mellitus often simply referred to as diabetes is a group of metabolic diseases in which a person has high blood sugar. Diabetes was first identified as a disease associated with “sweet urine,” and excessive muscle loss in the ancient world. Elevated levels of blood glucose (hyperglycemia) lead to spillage of glucose into the urine, hence the term sweet urine. It is a chronic disease that requires long-term medical attention both to limit the development of its devastating complications and to manage them when they do occur.Insulin is a hormone produced by the pancreas to control blood sugar. Diabetes can be caused by too little insulin in which pancreas does not make enough insulin, resistance to insulin that is muscle, fat, and liver cells do not respond to insulin normally, or both1-4. As the disease progresses tissue or vascular damage ensues leading to severe diabetic complications such as retinopathy5,6 neuropathy7,8 nephropathy9,10 cardiovascular complications11,12 and ulceration13,14. Thus, diabetes covers a wide range of heterogeneous diseases.

Diabetes was recognized with complete details and its types (Type 1and Type 2 diabetes – that is insulin dependent and non insulin dependent) in the year, 195915. According to W.H.O estimates, by 2025 total 300 million of the worldwide population will be affected by diabetes. For every 21 seconds, someone is diagnosed with diabetes, an estimation given by American Diabetes Association. And, there are 20.8 million diabetics in US at present, which is roughly estimated as 7% of US population, out of this figure about 6.2 millions are unaware of the diabetes existence in there life16, 17. As we know this is one of the old diseases, existing in many individuals and still on rising charts. Hence, scientists are continuously working to relieve us from it, by discovering the relevant drugs and making new researches. The medicinal plants imparts chief role in controlling the diabetes, associated with minimum side effects compared to synthetic drugs.

Alangium salvifolium, commonly known as Sage Leaved Alangium, stone mango, hill sack tree and ancolah18. Alangium salvifolium belongs to genus Alangium, family Alangiaceae. This family consists of twenty-two species out of which Alangium salvifolium is mainly used as medicine in India, China and Phillipines. Itis the most versatile medicinal plants having a wide spectrum of biological activity.

Alangium salviifolium showed potent anticancer, diuretic, anti-inflammatory, antimicrobial, laxative, astringent, emollient, anthelmintic and antiepileptic activities. The plant was also reported for its anti fungal activity, anti microbial activity, cardiac activity and anti fertility activity19-22. In Ayurveda almost all parts of the tree use for medicinal purposes. The roots and the fruits are used for the treatment of rheumatism, leprosy and hemorrhoid. Externally, it is used for the treatment of bites by rabbits, rats, and dogs. Root bark is an antidote for several poisons. Fruits are sweet, cooling and purgative and used as a poultice for treating burning sensation and haemorrhage23. In the present study the antidiabetic activity of leaves and barks of Alangium salvifolium was explored to identify its medicinal properties.

2 Materials and methods

2.1 Plant material

The leaves and barks of Alangium salvifolium were collected from the forest area of Raipur (Chhattisgarh). The plant was authenticated by Taxonomist, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh, India. A voucher specimen of the plant was preserved in the herbarium for further reference.

2.2 Preparation of extracts

500 gram of powdered of leaves and barks were successively extracted on a Soxhlet apparatus, employing petroleum ether, ethanol and distilled water respectively. The solvents of petroleum ether extract, ethanol extract and distilled water extract were removed by distillation and the last traces of solvent being removed under reduced pressure.The ethanol extract of leaves and barks of Alangium salvifolium were used for further pharmacological activity. The ethanol extract of leaves and barks of Alangium salvifolium were denoted as EASL and EASB respectively.

2.3 Acute toxicity study

Limit test at 2000mg/kg body weight was selected to perform acute toxicity study on selected according to the guidelines of the Organization for Economic Cooperation and Development (OECD)24. The purpose of the study was to determine the dose needed to perform different pharmacological activities.

2.4 Antidiabetic activity

2.4.1 Oral glucose tolerance test (OGTT) of Alangium salvifolium extract

The oral glucose tolerance test 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 EASL (200 mg/kg); Group V rats were administered EASL (400 mg/kg); Group VI rats were administered EASB (200 mg/kg); and Group VII rats were administered EASB (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 retro-orbital 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 kit25,27.

2.4.2 Induction of non-insulin dependent diabetes mellitus (NIDDM)

Non-insulin dependent diabetes mellitus was inducedin 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.

2.4.3 Evaluation of antidiabetic activity of Alangium salvifolium extracts

The animals were segregated into seven 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 rats were administered EASL (200 mg/kg); Group IV rats were administered EASL (400 mg/kg); Group V rats were administered EASB (200 mg/kg); Group VI rats were administered EASB (400 mg/kg); and Group VII rats were administered standard drug Glibenclamide (0.5 mg/kg) for 28 days. The fasting glucose levels were determined on days 0, 7th, 14th and 28th of extract administration. During the experimental period, the rats were weighed daily and the mean change in body weight was calculated28-31.

2.4.4 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 determinedby the glucose oxidase method, using an auto-analyzer29-32.

2.5 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. 3 Results 3.1 Acute toxicity study Limit test at 2000 mg/kg body weight was selected to perform acute toxicity of ethanol extracts of leaves and barks of Alangium salvifoliumon laboratory animals. In LD50 studies, it was found that the animals were safe up to a maximum dose of 2000 mg/kg body weight. There were no changes in normal behavior pattern and no signs and symptoms of toxicity and mortality were observed. 3.2 Selection of dose The LD50 of ethanol extracts of leaves and barks of Alangium salvifolium as per OECD guidelines falls under class four values with no signs of acute toxicity at 2000 mg/kg. The pharmacological evaluations were carried out at doses of 200 to 400 mg/kg body weights. 3.3 Oral glucose tolerance effects of Alangium salvifolium The effects of ethanol extracts of Alangium salvifolium 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 leaves and barks extracts of Alangium salvifolium, 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. 3.4 Effect on non-insulin dependent diabetes mellitus of Alangium salvifolium 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 Alangium salvifolium, on serum glucose levels of normal and Streptozotocin-induced rats are shown in table 2. The animals treated with streptozotocin namely Group II, a significant increase in serum glucose level was observed on 0, 7th, 14th and 28th 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 EASL in diabetic control rats, a significant reduction in blood glucose level was observed when compared with diabetic control rats. Moreover the administration of EASB in diabetic control rats, also significantly decreased the serum glucose level compared with diabetic control rats. The outcomes exhibited that EASL and EASB at dose of 400 mg/kg body weight significantly decreases the blood glucose level of diabetic rats on 7th day. While the EASL and EASB at dose of 200 mg/kg body weight significantly decreases the blood glucose level of diabetic rats on 14th day. From results it has been observed that the ethanol extracts of leaves exhibited maximum antidiabetic activity as compared to barks extract.