ABSTRACT

The study was designed to investigate the immunomodulatory effect of apple cider vinegar in alloxan induced male diabetic rats. Twenty-five (25) albino wistar rats with the body weight between 74-352g were randomly assigned to 5 groups. Group 1 which served as the control group was administered commercial rat chow and distilled water only, group 2 served as the negative control and was administered 150mg/kg body weight of alloxan. Groups 3, 4 and 5 were administered with 150mg/kg body weight of alloxan, thereafter they were treated with 0.5ml/kg body weight, 1ml/kg and 1.5ml/kg body weight of 10% organic apple cider vinegar to every other day for 21 days. All the animals were allowed free access to feeds and water ad libitum. The result for total white blood cell (TWBC) showed a significant decrease (P<0.05) in group 5 when compared to group 1. Neutrophil recorded a significant increase (P<0.05) in all the experimental groups when compared to the negative control. Lymphocytes concentration had a significant decrease (P<0.05) in all the experimental groups when compared to the negative control. The concentration of monocytes revealed a significant decrease (P <0.05) in group 3 and a significant increase (P<0.05) in group 4 when compared to the control. However, there was a significant decrease (P<0.05) in group 3 and a significant increase in group 4 when compared to group 2. Eosinophil count showed a significant increase (P <0.05) in group 3 and 4 when compared to group 1. A significant increase (P <0.05) was observed in group 3 and 4 when compared to the negative control.

Basophil recorded a significance increase (P<0.05) in group 2 when compared to group 1. However, all the experimental groups had a significant decrease (P<0.05) when compared to the negative control. Mean Corpuscular hemoglobin in the study revealed a significant decrease (P<0.05) in group 5 when compared to the negative control. This result indicates that apple cider vinegar has potential benefits in diabetic rats and thus could be used as a nutritional therapy in diabetic patients. Apple cider vinegar may be considered useful in precluding metabolic syndrome by reducing diabetes and its associated complications.

TABLE OF CONTENTS

Title Page    –         –         –         –         –         –         –         –         –         i

Certification –         –         –         –         –         –         –         –         –         ii

Dedication   –         –         –         –         –         –         –         –         –         iii

Acknowledgements         –         –         –         –         –         –         –         iv

Abstract       –         –         –         –         –         –         –         –         –         v

Table of Contents  –         –         –         –         –         –         –         –         vi

List of Tables         –         –         –         –         –         –         –         –         vi

CHAPTER ONE: INTRODUCTION

1.1     Background of the Study –         –         –         –         –         –         1

1.2     Aim and Objectives of the study          –         –         –         –         6

1.2.1  Aim of the Study   –         –         –         –         –         –         –         6

1.2.2  Objectives of the study    –         –         –         –         –         –         6

1.3     Scope and Limitation of the study       –         –         –         –         7

1.4     Significance of the study –         –         –         –         –         –         7

1.5     Definition of terms –         –         –         –         –         –         –         8

CHAPTER TWO: LITERATURE REVIEW

2.1     Description of Apple cider vinegar –    –         –         –         –         9

2.1.1  Nutritional value Apple cider vinegar –         –         –         –         10

2.1.2  Safety concerns of Apple cider vinegar –       –         –         –         12

2.1.3  Health Benefits of Apple cider vinegar          –         –         –         13

2.1.4  Uses of Apple cider vinegar       –         –         –         –         –         14

2.1.5  Pharmacological Effect of Apple cider vinegar       –         –         15

2.2     The Chemistry of blood   –         –         –         –         –         –         22

2.2.1  Composition of Blood     –         –         –         –         –         –         23

2.3     Hematological Indices     –         –         –         –         –         –         24

2.3.1  White Blood cell (WBC) differential   –         –         –         –         24

2.3.2  Mean Corpuscular Hemoglobin (MCH)         –         –         –         35

2.4     Diabetes Mellitus  –         –         –         –         –         –         –         38

2.5     Signs and Symptoms of Diabetes        –         –         –         –         39

2.6     Diagnosis of Diabetes Mellitus  –         –         –         –         –         42

2.7      Alloxan and its Biological Effects      –         –         –         –         44

2.7.1 Mechanisms of Action of Alloxan       –         –         –         –         46

CHAPTER THREE: MATERIALS AND METHODS

3.1     Materials      –         –         –         –         –         –         –         –         49

3.2     Methods       –         –         –         –         –         –         –         –         49

3.2.1 Sample Collection  –         –         –         –         –         –         –         49

3.2.2  Preparation of Samples    –         –         –         –         –         –         50

3.2.3  Experimental Animal       –         –         –         –         –         –         50

3.2.4  Experimental Design and Grouping of Animals       –         –         51

3.2.5  Collection of Blood Samples     –         –         –         –         –         52

3.3     Haematological Assay     –         –         –         –         –         –         53

3.4     Determination of some haematological Parameters  –         –         53

3.5     Statistical Analysis          –         –         –         –         –         –         55

CHAPTER FOUR: RESULTS AND DISCUSSION

4.1     Results         –         –         –         –         –         –         –         –         56

4.2     Discussion   –         –         –         –         –         –         –         –         59

CHAPTER FIVE: CONCLUSION AND RECOMMENDATION

5.1     Conclusion –         –         –         –         –         –         –         –         66

5.2     Recommendations –         –         –         –         –         –         –         66

References

CHAPTER ONE: INTRODUCTION

1.1     Background of the Study

Diabetes mellitus (DM) has been considered as one of the most universal endocrine, metabolic disorders which is characterized by the irregular metabolism of fat, protein and carbohydrate due to decreased efficacy of insulin secretion or altered insulin activity. About 14 million patients were suffering in diabetes mellitus in the United State and this ratio has been increasing by about 700, 000 each year (Jajarm et al., 2008). Diabetes has been categories into 3 major types. Type 1 diabetes known as insulin dependent diabetes mellitus (IDDM) which result in deficiency of insulin, insulin resistance known as type II diabetes, it is also called non-insulin dependent diabetes (NIDDM) and gestational diabetes is known as type III diabetes which results in glucose intolerance during pregnancy (Hassan, 2018).

Diabetes mellitus is a global health issue and one of the 5^th main mortality and morbidity causes in many areas of the world. More than 171 million people were affected by diabetes globally in the year 2000 and this figure is predicted to reach 366 million by the year 2030 (Gispen et al., 2000). Main causes of diabetes are impaired insulin secretion, insulin resistance and overproduction of hepatic glucose. Insulin resistance is not only a major contribution to diabetes development but also play a mere role in hypertension, dyslipidemia and atherosclerosis (Ali, 2018).

A complication of diabetes includes cataract, nephropathy, retinopathy, wound infection and atherosclerosis (Peter et al., 2011).

Diabetes mellitus is associated with premature mortality decreased quality of life and increase health-care expenditures. The result of previous studies showed that diabetes prevalence is increasing with the passage of time in Pakistan. The numbers of people affected with diabetes in Pakistan is 6.9 million and this will grow to 11.5 million according to the International Diabetes Federation by 2025 and urgent measures are required to control (Petsiou, 2014).

Actually, simple inexpensive diet strategies are greatly needed for diabetes control. In the present study, the effect of oral apple cider vinegar consumption on blood biochemical factors in type 2 diabetic and hyperlipidemia patients were investigated.

Apple cider vinegar is vinegar made from fermented apple juice, and used in salad dressings, marinades, vinaigrettes, food preservatives, and chutneys. It is made by crushing apples, then squeezing out the juice. Bacteria and yeast are added to the liquid to start the alcoholic fermentation process, which converts the sugars to alcohol. In a second fermentation step, the alcohol is converted into vinegar by acetic acid-forming bacteria (Acetobacter species). Acetic acid and malic acid combine to give vinegar its sour taste (Hill et al., 2005).

Haematology  refers  to  the  study  of  the  numbers  and  morphology  of  the  cellular  elements  of  the  blood the  red  cells  (erythrocytes),  white  cells  (leucocytes),  and  the  platelets  (thrombocytes)  and  the  use  of  these  results  in  the  diagnosis  and  monitoring  of  disease  (Addass et al., 2012). Haematological  studies  are useful  in  the  diagnosis  of  many  diseases  as  well  as  investigation  of  the  extent  of  damage  to  blood.

Haematological studies are of ecological and physiological interest in helping to understand the  relationship  of  blood  characteristics  to  the  environment  and  so  could  be  useful  in  the  selection  of  animals  that  are  genetically  resistant  to  certain  diseases  and  environmental conditions (Chung, 2002).

Haematological parameters are  good  indicators  of  the  physiological  status  of  animals  (Isaac et al., 2013).  Haematological parameters  are  those  parameters  that  are  related  to  the  blood  and  blood  forming  organs. Blood act as a  pathological  reflector  of  the  status  of  exposed  animals  to  toxicant  and  other  conditions.

Changes in haematological parameters are often used to determine various status of the body and to determine stresses due to environmental, nutritional and/or pathological factors (Afolabi et al., 2010).

One of the most potent methods to induce experimental diabetes mellitus is chemical induction by Alloxan. It is a well-known diabetogenic agent that is used to induce type I diabetes in experimental animals. Alloxan is a urea derivative which causes selective necrosis of the β-cells of pancreatic islets. In addition, it has been used widely to produce experimental diabetes in animal such as rabbits, rats, mice and dogs with different grades of disease severity by varying the dose of Alloxan used (Etuk, 2010; Iranloye et al., 2011). As it has been widely accepted that alloxan selectively destroys the insulin-producing beta-cells found in the pancrease, hence it is used to induced diabetes in laboratory animals. The toxic action of Alloxan on pancreatic beta cells involve oxidation of essential sulphydryl, inhibition of glucokinase enzyme, generation of free radicals and disturbances in intracellular calcium homeostasis (Dhanesha et al., 2012). The underline mechanism involved the selective uptake of the compound due to its structural similarity to glucose as well as a highly efficient uptake mechanism of the pancreatic beta cells (Lenzen, 2008; Viswanathaswamy et al., 2011). 

1.2     Aim and Objectives of the Study

1.2.1  Aim of the Study

The aim of this research project is to determine the immunomodulatory effect of organic extract of apple cider vinegar in alloxan induced diabetic albino wistar rats.

1.2.2  Objectives of the Study

1. To determine the effect of organic apple cider vinegar on Total White Blood Cells (TWBC) in diabetic rats.
2. To determine the effect of organic apple cider vinegar on neutrophil counts in albino diabetic rats treated with apple cider vinegar.
3. To determine the effect of organic apple cider vinegar on lymphocyte counts in albino diabetic rats treated with apple cider vinegar.
4. To determine the effect of organic apple cider vinegar on monocytes counts in albino diabetic rats treated with apple cider vinegar.
5. To determine the effect of organic apple cider vinegar on eosinophil counts in albino diabetic rats treated with apple cider vinegar
6. To determine the effect of organic apple cider vinegar on basophil counts in albino diabetic rats treated with apple cider vinegar
7. To determine the effect of organic apple cider vinegar on Mean Corpuscular Hemoglobin (MCH) in albino diabetic rats treated with apple cider vinegar.

1.3     Scope and Limitations

This research work focuses on the Immunomodulatory effect of Apple Cider Vinegar in Alloxan-induced diabetic Albino wistar rats due to time constraint, lack of facilities and financial constraint.

1.4     Significance of the Study

The significance of this research work lies in it potential to provide valuable insights into the potential therapeutic benefits of Apple Cider vinegar in the management of diabetes. Also, understanding the effect of Apple cider vinegar on some hematological parameters in diabetic rats.

1.5     Definition of Terms

Hematological Parameters: Are those parameters that are related to the blood and blood forming organs. Also, they are good indicators of the physiological status of animals (Isaac et al., 2013).

Apple Cider Vinegar: Is the fermented juice from crushed apples. It contains acetic acid and nutrients such as B vitamins and vitamin C. acetic acid and malic acid combine to give vinegar it sour taste (Ulbricht, 2010).

Hematology: R`efers to the study of the numbers and morphology of the cellular elements of the blood. The red cells, white cells and the platelets and the use of these results in the diagnosis and monitoring of disease (Addass et al., 2012).

Diabetes: Is chronic diseases that occur either when the pancreas does not produce enough insulin or the cells of the body becoming unresponsive to the hormones effect (Saedi et al., 2016).

Alloxan: Is a white crystalline pyrimidine derivative, C₄ H₂ O₄ N₂, used in biomedical research to induce diabetes in laboratory animals.