ANTI-DIABETIC EFFECTS OF ETHANOL LEAF EXTRACT OF ONIONS (Allium cepa) ON ALLOXAN-INDUCED DIABETIC WISTAR ALBINO RATS

ANTI-DIABETIC EFFECTS OF ETHANOL LEAF EXTRACT OF ONIONS (Allium cepa) ON ALLOXAN-INDUCED DIABETIC WISTAR ALBINO RATS Choji Solomon S. Department of Biochemistry, Faculty of Natural and Applied Sciences, Plateau State University, Bokkos. P.M. B 2012, Jos. Nigeria. [email protected] +2347065752410 Damla Faith U. Department of Biochemistry, Faculty of Natural and Applied Sciences, Plateau State University, Bokkos. P.M. B 2012, Jos. Nigeria Barde Larry A. Department of Biochemistry, Faculty of Natural and Applied Sciences, Plateau State University, Bokkos. P.M. B 2012, Jos. Nigeria Zakka Riyang. Department of Food Science and Technology, Faculty of Agricultural Science, Federal University Wukari. P.M.B 1020 Adegbite Adeshola. Ladoke Akintola University of Technology. Ogbomoso. Oyo State. Abstract. Diabetes is a chronic disease characterised by high blood glucose level and abnormal metabolism of carbohydrates, protein and fat. The condition is characterised by persistent hyperglycaemia. Allium cepa leaf is a functional food used in traditional medicine for the treatment of diabetes mellitus. The use of plants especially vegetable as antidiabetic remedies have added interest of joining two basic diabetes mellitus control factors: food and medication. The ethanol extract of Allium cepa leaf was investigated for antidiabetic effects using alloxan- induced diabetic wistar albino rats. Wistar albino rats were randomly divided into six groups; Group A rats were non-diabetic control. Diabetes was induced in groups B, C, D, E and F by single intraperitoneal injection of alloxan (150mg/kg body weight). Group B were not treated and served as negative control group. Group C were treated with glibenclamide (5mg/kg body weight), thus served as postive control group. Groups D, E and F were treated with 200, 300 and 400 mg/kg body weight of the extract respectively for a period of two weeks through intraperitoneal route. The effect of treatment with the doses of the extract and standard drug were studied on blood glucose level, total serum cholesterol and body weight. Allium Cepa extract produced a dose- dependent significant reduction in the blood glucose level when compared with that of the control group. Significant total serum cholesterol reduction was observed at 300 and 400mg/kg. An observed decrease in body weight of the negative control group was recorded and significant increase for all other groups. The findings from this study indicate that the crude extract of Allium cepa leaf caused a significant hypoglycaemic and hypocholesterolemic activity in alloxan-induced diabetic rats thus, validates its use in ethno – medicine for the control of diabetes mellitus. KEY WORDS: Diabetes mellitus, Allium cepa, Alloxan, Blood glucose, Cholesterol Glibenclamide. 1.0 INTRODUCTION. Diabetes Mellitus (DM) is a group of metabolic disorders associated with disturbances in the metabolism of fuel molecules due to absolute deficiency of insulin, insufficient insulin secretion and / or its secretion [1]. It is a disorder that affects the body’s ability to make or use insulin. Insulin is a hormone produced in the pancreas that helps transport glucose (blood sugar) from the bloodstream into the cells so they can break it down and use it for fuel. People cannot live without insulin [2]. It is also a widespread endocrine disorder that is associated with considerable morbidity and mortality and is found in all population throughout the world [3] Despite the presence of anti-diabetic drugs in the pharmaceutical market, the treatment of diabetes with medicinal plants is often successful. Herbal medicine and plant components with insignificant toxicity and less or no side effect are notable therapeutic options for the treatment of this disease around the world [4]. The most common herbal active ingredients used in treating diabetes are flavonoids, tannins, phenols and alkaloids [5]. The existence of these compounds implies the importance of the anti-diabetic properties of these plants [4]. Allium cepa is one of the recognised medicinal plants known to possess several medicinal properties including lowering of blood pressure, antiseptic, hypoglycaemic and hypocholesterolemic activity [6]. In the rural communities, many people depend solely on medicinal plants for the treatment of diabetes due to its easy accessibility, affordability and availability even when the efficacy of the herbal remedies has not been established [6]. Dietary therapy is unarguably the best treatment for diabetes. The diabetic diet should be carefully monitored to minimize the load placed on the blood glucose regulating mechanism. The use of plants, especially vegetables, by the population as antidiabetic remedies has added interest of joining two basic diabetes mellitus control factors: food and medication [7]. This research is thus geared towards finding a medicinal plant that will not only increase the energy content of diabetics but also lower glycaemic index properties for the management of diabetic pressures in our society. 2.0 MATERIALS AND METHODS. 2.1 Materials. 2.1.1 Chemicals and Reagents. Baker Ltd Dagenham, England, BDH Chemicals Ltd; Poole England, Sigma Chemicals, St Louis, USA, Emzor Pharmaceuticals Industry Ltd, Nigeria and Randox Laboratories. London, UK. 2.1.2 Plant The Allium cepa leaves used for the experiment was bought from Barkin Ladi Market, Plateau State, Nigeria. The plants were identified by Professor Pob Poppva in the Department of Botany, University of Jos, Plateau State. A voucher specimen was deposited in the herbarium unit of the department. 2.1.3 Experimental Animals. A total of thirty-six (36) adult male Wistar albino rats weighing 80 to 150g and twelve (12) mice were used for the experiment. The experimental animals were purchased from Chris Animal Farm, G.R.A. Awka. They were housed six (6) rats per cage at the experimental Animal House of Biochemistry Department, Nnamdi Azikiwe University, Awka, Anambra State. They were acclimatized for two weeks under standard laboratory conditions and were maintained on water and Guinea growers mash pellet (Vital Feed Grand Cereals Nigeria Ltd, Jos, Nigeria) that was obtained from Eke Market, Awka, Anambra State. 2.2 Methods 2.2.1 Preparation of ethanol leaf extract of Allium cepa . The leaves of Allium cepa were properly washed with distilled water and dried at room temperature for three weeks. The dried leaves were then pulverised using corona manual grinding machine. The powdered samples of Allium cepa was weighed and exactly 1475g was extracted in 5 litres of 80% ethanol for 24 hours with occasional stirring, sieved and filtered using filter paper (Whatman number 1). The filtrate was then concentrated using a rotary evaporator at 600C and appeared as a dark brown gel solid. The extracts were kept in a labelled glass container and stored in a refrigerator until when required for reconstitution and administration. 2.2.2 Phytochemical Screening of Secondary metabolites(Constituents) The qualitative phytochemical screening of the ethanol leaf extract of Allium cepa was carried out using standard procedures as outlined by [8], [9]. 2.2.3 Acute toxicity and Median Lethal Dose (LD50) test of ethanol leaf extract of Allium cepa. The median Lethal Dose (LD50) was determined using Wistar albino mice as described by the modified method of [10]. Test animals were divided into six (6) groups. The first 3 groups which contain 3 animals each were given 10mg/kg, 100mg/kg and 1000mg/kg body weight of the ethanol extract of Allium Cepa leaves. The Allium Cepa extract was administered orally and was monitored for 24 hours. The last 3 groups which contain one animal each per group were then given 1600mg/kg, 2900mg/kg and 5000mg/kg body weight of the ethanol extract of Allium Cepa leaves and were observed for 24 hours. 2.2.4 Induction of Diabetes. Alloxan was prepared and induced by adopting the method of [11]. All rats, except for the normal control group were intraperitoneally injected with 150mg/kg body weight of the prepared alloxan dissolved in normal saline solution. The blood glucose levels of the rats were checked before the administration of alloxan using one touch glucometer (Fine touch, USA) and test strips. The rats were then fasted for 16 hours, but with free access to water after which they received an intraperitoneal injection of alloxan 150mg/kg body weight. The rats were orally given 20ml each of 10% glucose solution after 2 hours to prevent hypoglycaemia. The animals were allowed free access to food and water after alloxan administration. After 48 hours of the alloxan administration, blood was collected orbito-rectally and their glucose levels were checked using one touch glucometer and test strips. Diabetes was confirmed to have been induced if the glucose level was observed to be far much higher than normal (above 140mg/dl). 2.2.5 Experimental Design This study was carried out on alloxan –induced diabetic rats for two (2) weeks. A total of thirty-six (36) Wistar albino rats were used for the experiment. The albino rats were randomly divided into six (6) groups with six (6) rats in each group. The extract and the reference drug were administered intraperitoneally to the animals. Group A – Normal (non-diabetic control) Group B – Diabetic (negative) control group Group C – Diabetic (positive) control – this group received 5mg/kg body weight of glibenclamide. Group D – This group received 200mg/kg body weight of the extract. Group E – This group received 300mg/kg body weight of the extract. Group F – This group received 400mg/kg body weight of the extract The weights of the animals were carefully monitored before the induction and throughout the duration of the experiment. 2.2.6 Biochemical Assay 2.2.6.1 Blood glucose level determination Determination of the blood glucose level was done by the glucose-oxidase principle [12] using the one touch instrument and results were reported as mg/dl [13]. 2.2.6.2 Determination of total serum cholesterol. The cholesterol of the serum was oxidised to tetraene derivative by ferric ions derived from ferric perchlorate using four different test tubes that were marked test, control, standard and blank. The absorbance was measured (using spectrophotometer) at 590nm wavelength and compared with that of a pure solution of cholesterol [14]