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EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS

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[icon type=”icon-pencil”]: EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS
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This study,EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS contains concise information that will serve as a framework or guide for your project work. The project study is well-researched for academic purposes and are usually provided in complete chapters with adequate References.

Keywords:EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS


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RESEARCH BODY

ABSTRACT

 

Globally, the estimated incidence of diabetes and projection for the year 2030 as given by the International Diabetes Federation (IDF) is 350 million. Kigelia africana is highly used for ethnomedicinal purposes although there is paucity of scientific information on its use. This work was therefore, aimed at evaluating the anti-diabetic and antioxidative potential of the plant. Ethanol, methanol and n- hexane extracts of the leaves of Kigelia africana were used for the study. Alloxan diabetes was induced in a total of 60 adult male albino rats weighing between 90 and 160 g. The alloxan was dissolved in cold normal saline. After 72 hr, diabetes was confirmed and the rats were divided into twelve (12) groups of five (5) rats each. Group 1 served as the normal control, group 2 was the diabetic untreated, group 3 received 2.5 mg /kg b.wt of glibenclamide, groups 4, 6 and 8 received ethanol, methanol and n-hexane leaves extract while group 5, 7 and 9 received ethanol, methanol and n-hexane fruit extract respectively of 500 mg/kg b.wt of the extracts. Groups 10-12 were administered equal combination of the leaves and fruits extracts. The rats were fed orally for 21 days after which some biochemical and oxidative parameters were statistically analysed. Phytochemical screening for different bioactive compounds was done using standard methods and indicated the presence of flavoniods, alkaloids, saponins, soluble carbohydrates, tannin, steroids, glycosides and reducing sugars. Proximate analysis revealed the presence of proteins (13.9%), carbohydrates (63.5%), fats and oil (11.4%) and crude fibre (2.2%). LD50 showed that the extracts were safe. The glucose level decreased while body weight increased in all the treated groups compared with the diabetic rats untreated. Oral administration of 500mg/kg b.w of K. africana extract significantly reduced (p<0.05), the sorbitol, glycohaemoglobin (HbA1c), total protein, and vitamin C concentrations in diabetic rats (groups 4-12) in comparison with the positive control. There were significant differences in glycohaemogolin, sobitol, total protein and vitamin C concentration in diabetic rats fed with a combination of the two parts of the plant extracts (groups 10-12) as against groups 4-9 administered single extracts. Malondiadehyde (MDA) concentration significantly decreased (p < 0.05) in all the test groups compared with the diabetic untreated rats. Low density lipoprotein, total cholesterol, and triacylgycerol levels decreased significantly (p < 0.05) in the treated groups in comparison with the positive control animals (group 3). However, administration of 500 mg/kg b.w of K. africana increased significantly (p<0.05) the high density lipoprotein (HDL) across the test groups as against the diabetic untreated group. Significant decreased (p<0.05) in the lipid profiles (except HDL) was recorded in groups 10, 11 and 12 treated with a combination of two parts (leaf and fruit) of K. africana in comparison with groups 4-9 orally fed with a single plant extract. Furthermore, the data recorded significantly increased (p < 0.05) antioxidant enzymes (SOD, CAT GPX) activities in diabetic treated groups (both combination and single) with reference to the positive control group. Similarly, significant increase (p > 0.05) of SOD and CAT activities and SOD percentage inhibition was observed in group 3 treated with 2.5 mg/kg b.wt of glibenclamide (standard) compared with all the test groups. Significant reduction (p < 0.05) in the activities of ALT, ALT and total bilirubin concentration were observed in the test groups treated with the extracts compared with the diabetic untreated rats. ALT activity and total bilirubin level decreased significantly (p < 0.05) in groups 10, 11 and 12 administered a combination of leaf and fruit extracts as against groups 4-9 treated with either leaf or fruits only. The results suggest that management and prevention of diabetic complications can be achieved by the use of K. africana.

TABLE OF CONTENTS

CHAPTER ONE: INTRODUCTION

1.1       Kigelia africana

1.1.1    Description of Kigelia africana

1.1.2    Taxonomy of Kigelia africana

1.1.3    Traditional uses of Kigelia africana

1.1.4    Chemical constituents of Kigelia africana

1.1.5    Antibacteria and antifungi

1.2       Diabetes

1.2.1    Diabetes mellitus

1.2.2    Diabetes Type 1 and 2

1.2.3    Insulin resistance

1.2.4    Diabetic complications

1.3       Hyperglycemia and diabetic complication

1.4       Mechanism of tissue damage mediated by hyperglycemia

1.4.1    Aldose reductase pathway

1.4.2    Non-enzymatic glycation

1.4.3    Carbonyl stress in diabetes

1.4.4    Activation of protein kinase C isoforms

1.5       Oxidative stress

1.5.1    Mechanism of increased oxidative stress in diabetes mellitus

1.5.2    Glucose autoxidation

1.5.3    Free radicals

1.5.4    Reactive oxygen species and oxidative stress

1.6       Antioxidant system

1.6.1    Scavenging properties of antioxidants

1.6.2    Positive and negative effects of free radicals

1.7       Lipid peroxidation

1.8       Antioxidant supplementation in diabetes mellitus

1.9       Alloxan

1.9.1    Alloxan diabetes and streptozotocin diabetes

1.9.2    Alloxan: Mechanism of action

1.9.3    Beta cell toxicity and diabetogenicity of alloxan

1.9.4    Streptozotocin: Mechanism of action and beta cell selectivity

1.9.5    Beta cell toxicity of streptozotocin

1.9       Rationale for the study

1.10     Aim and objectives of the study

1.10.1  Aim of the study

1.10.2  Specific objectives of the study

CHAPTER TWO: MATERIALS AND METHODS

2.1       Materials

2.1.1    Chemicals

2.1.2    Instrument/Equipment

2.1.3    Drug

2.1.4    Plant material

2.2       Methods

2.2.1    Animal management

2.2.2    Preparation of plant extracts

2.2.3    Design of the experiment

2.2.4    Yield of extracts

2.2.5    Phytochemical analysis of the crude extracts

2.2.5.1 Test for the presence of alkaloids

2.2.5.2 Test for carbohydrates

2.2.5.3 Test for reducing sugar

2.2.5.4 Test for protein

2.2.5.5 Test for fats and oil

2.2.5.6 Test for glycosides

2.2.5.7 Test for acidic substances

2.2.5.8 Test for the presence of flavonoids

2.2.5.9 Test for the presence of steroids

2.2.5.10  Test for tannins

2.2.5.11 Test for resins

2.2.5.12 Test for saponins

2.2.3.13  Test for terpenoids and steroids

2.2.6    Proximate Analysis

2.2.6.1 Crude protein

2.2.6.2 Crude fat

2.2.6.3 Moisture

2.2.6.4 Ash /Mineral matter

2.2.6.5 Crude fibre

2.2.6.6 Carbohydrate or nitrogen free extract (NFE)

2.2.7    Acute toxicity test

2.2.7.1 Determination of LD50 of the extract

2.2.8    Induction of diabetes

2.2.9    Determination of fasting and random glucose concentrations

2.2.10  Determination of sorbitol concentration

2.2.11  Determination of total protein concentration

2.2.12  Determination of haemoglobin glycosylation

2.2.13  Determination of malondialdehyde concentration

2.2.14  Determination of vitamin C concentration

2.2.15  Assay of catalase activity

2.2.16  Assay of superoxide dismutase (SOD) activity

2.2.17  Assay of glutathione peroxidase activity

2.2.18  Determination of total cholesterol concentration

2.2.19  Determination of high density lipoprotein (HDL) cholesterol concentration

2.2.20  Determination of low density lipoprotein (LDL) cholesterol concentration

2.2.21  Determination of triacylglycerol concentration

2.2.22  Assay of aspartate aminotransferase (AST) activity

2.2.23  Assay of alanine aminotranferase (ALT) activity

2.2.24  Determination of total bilirubin concentration

2.3       Statistical analysis

CHAPTER THREE: RESULTS

3.1       Qualitative phytochemical composition of ethanol, methanol and n-hexane leaf and fruit extracts of Kigelia africana

3.2       Quantitative phytochemical composition of ethanol, methanol and n-hexane leaf and fruit extracts of Kigelia africana

3.3       Percentage proximate compositions of ethanol, methanol and n-hexane leaf and fruit extracts of Kigelia africana

3.4       Percentage yield of leaf and fruit samples of Kigelia Africana

3.5       Acute toxicity studies

3.6       Effect of ethanol, n-hexane and methanol extracts of leaves and fruits of Kigelia africana on sugar level of diabetic rats

3.7       Body weights of diabetic rats treated with ethanol, n-hexane and methanol extracts of leaves and fruits of Kigelia africana before and after experiment

3.8       Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on sorbitol concentration in alloxan-induced diabetic rats

3.9       Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigeria africana on total protein in alloxan-induced diabetic rats

3.10     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on glycosylated haemoglobin concentration in alloxin-induced diabetic rats

3.11     Effect of ethanol, methanol and n-hexane extracts of leaf and fruit of Kigelia africana on malondialdehyde (MDA) concentration in alloxan-induced diabetic rats

3.12     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on vitamin C concentration in alloxan-induced diabetic rats

3.13     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on catalase activity in alloxan-induced diabetic rats

3.14     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on superoxide dismutase (SOD) activity in alloxan- induced diabetic rats

3.15     Effect of ethanol methanol and n-hexane leaf and fruit extracts of Kigelia africana on percentage inhibition of superoxide dismutase in alloxan-induced diabetic rats

3.16     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on glutathione peroxidase activity in alloxan-induced diabetic rats

3.17     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on cholesterol concentration in alloxan-induced diabetic rat

3.18     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana high density lipoprotein in alloxan-induced diabetic rats

3.19     Effect of ethanol, methanol and n-hexane leaf and fruit extracts of Kigelia africana on low density lipoprotein concentration in alloxan-induced diabetic rats

3.20     Effect of ethanol, methanol and n-hexane leaf and fruit extracts of Kigelia africana on triacylglycerol (TAG) concentration in alloxan-induced diabetic rats

3.21     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on aspartate aminotranferase (AST) in alloxan-induced diabetic rats

3.22     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on alanine aminotransferase (ALT) in alloxan-induced diabetic rats

3.23     Effect of ethanol, methanol and n-hexane leaf and fruit extract of Kigelia africana on total bilirubin concentration in alloxan-induced diabetic rats

CHAPTER FOUR: DISCUSSION

4.1       Discussion

4.2       Conclusion

REFERENCES

APPENDICES

CHAPTER ONE

INTRODUCTION

Diabetes mellitus is a metabolic disorder resulting from a defect in insulin secretion, insulin action or both. Insulin deficiency in turn leads to chronic hyperglycemia with disturbances of carbohydrate, fat and protein metabolism (Kumar et al., 2011).

During diabetes, failure of insulin-stimulated glucose uptake by fat and muscle cause glucose concentration in the blood to remain high, consequently glucose uptake by insulin-independent tissue increases. Increased glucose flux both enhances oxidant production and impairs antioxidant defenses by multiple interacting non-enzymatic, enzymatic and mitochondrial pathways (Klotz 2002; Mehta et al., 2006). These include activation of protein kinase C isoforms (Inoguchi et al., 2000), increased hexosamine pathway (Kaneto et al., 2001), glucose autoxidation (Brownlee, 2001), increased methylglyoxal and advanced glycation end-product (AGEs) formation (Thornalley, 1998) as well as increased polyol pathway flux ( Lee and Chung, 1999). These seemingly different mechanisms are the results of a single process-that is, overproduction of superoxide by the mitochondrial electron transport system (Tushuizen et al., 2005). This hyperglycaemia-induced oxidative stress ultimately results in modification of intracellular proteins resulting in an altered function and DNA damage, activation of the cellular transcription (NFK B), causing abnormal changes in gene expression, decreased production of nitric oxide, and increased expression of cytokines, growth factors and pro-coagulant and pro-inflammatory molecules (Palmer et al., 1988; Evans et al., 2002; Klotz, 2002; Taniyama and Griendling, 2003). Oxidative stress is responsible for molecular and cellular tissue damage in a wide spectrum of human diseases (Halliwell, 1994), amongst which is diabetes mellitus. Diabetes produces disturbances of lipid profiles, especially an increased susceptibility to lipid peroxidation (Lyons, 1991), which is responsible for increased incidence of atherosclerosis (Guigliano et al., 1996), a major complication of diabetes mellitus . An enhanced oxidative stress has been observed in these patients as indicated by increased free radical production, lipid peroxidation and diminished antioxidant status (Baynes, 1991).

Globally, the estimated incidence of diabetes and projection for year 2030, as given by International Diabetes Federation is 350 million (Ananda et al., 2012). Currently available pharmacotherapies for the treatment of diabetes mellitus include oral hypoglycemic agents

and insulin. However these drugs do not restore normal glucose homeostasis and they are not free from side effects (Bandawane et al., 2011). In view of the adverse effects associated with the synthetic drugs and as plants are safer, affordable and effective, conventional antidiabetic plants can be explored (Kumar et al., 2010). Over 400 traditional plants have been reported for the treatment of diabetes (Ramachandran et al., 2011). Furthermore, following World Health Organization recommendations, investigation of hypoglycemic agents from medicinal plants has become more important (Kumar et al., 2010). Also diabetes has been treated orally with several medicinal plants based on folklore medicine since ancient times.

Kigelia africana (Lam.) Benth (Family: Bignoniaceae) plant has many medicinal properties due to the presence of numerous secondary metabolites. These compounds include iridiods, flavonoids, naphthoquinones and volatile constituents (Houghton, 2002; Gorman, 2004; Asekun et al., 2006). Experimentally, the plant has shown antibacterial, antifungal, antineoplastic, analgesic, anti-inflammatory and antioxidant properties (Saini et al., 2009). Crude extracts of herbs and spices and other materials rich in phenolics are of increasing interest in the food industry because they retard oxidative degradation of lipids and thereby improving the quality and nutritional value of food. Flavonoids, are groups of polyphenolic compounds with known properties, which include free radical-scavenging and anti-inflammatory activities (Frankel, 1995).

An enhanced oxidative stress has been observed in diabetic patients as indicated by increased free radical production, lipid peroxidation and diminished antioxidant status (Baynes, 1991). In diabetes mellitus, alterations in the endogenous free radical scavenging defense mechanisms may lead to ineffective scavenging of reactive oxygen species, resulting in oxidative damage and tissue injury. Oxidative stress is currently suggested as mechanism underlying diabetes and diabetic complications. Oxidative stress may cause oxidative damage of cellular membranes and changes in the structural and functional integrity of sub- cellular organelles and many produce effects that result in the various complications in diabetic disease (West, 2000). Recently, there has been an upsurge of interest in the therapeutic potentials of plants, as antioxidants in reducing free radical induced tissue injury. Although several synthetic antioxidants, such as ascorbic acid, butylated hydroxyanisole and butylated hydroxytoluene are commercially available, they are quite unsafe and toxic (Viny, et al.,2010). A survey of literature revealed that there is no experimental evidence of the antidiabetic effect of Kigelia africana. Therefore, the present work explores this and will in

addition study its potential effect on the oxidative and biochemical parameters of alloxan-induced diabetic rats.

Keywords: EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS


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Keywords: EFFECTS OF ETHANOL, METHANOL AND N-HEXANE LEAF AND FRUIT EXTRACTS OF Kigelia africana ON SOME OXIDATIVE AND BIOCHEMICAL PARAMETERS IN ALLOXAN-INDUCED DIABETIC RATS