NUTRITIONAL POTENTIAL OF SYNSEPALUM DULCIFICUM PULP AND THE EFFECTOF THE METHANOLIC EXTRACT ON SOME BIOCHEMICAL PARAMETERS IN ALBINO RATS

7

ABSTRACT

The nutritive and antinutritive compositions of S. dulcificum pulp were analysed to augment the available information on the anti-diabetic effect of the plant. Biochemical parameters like liver function enzymes (ALT, AST, ALP) and bilirubin concentrations,serum total protein, serum albumin and globulin, kidney function parameters (creatinine and urea concentrations), blood glucose, serum lipid profile and lipid peroxidation were determined in rats that were administered different concentrations of the methanolic extract to ascertain their effects. The internal organs (liver and kidney) were also removed and used for histopathological studies. From the result of the study, the proximate composition shows that S. dulcificum contains 7.75% protein, 59.55% moisture content, 4.36% ash, 6.24% crude fibre, 3.26% fat and 18.84% carbohydrate.The result of the mineral analysis shows that S.dulcificum pulp contains 100 mg/g calcium, 24.20 mg/g iron, 9.49 mg/g zinc, 6.22 mg/g copper, 0.01 mg/g chromium and 0.01 mg/g cobalt. Vitamin analyses shows that the S. dulcificum pulp contains 0.04% vitamin A, 22.69% vitamin C, 0.01% vitamin D and 0.02% vitamin K. Antinutrient analyses of the pulp show 5.67% oxalate, 0.03% phytates and 0.02% hemagglutanin. Amino acid profile shows that S.dulcificum pulp contains 8.055% tryptophan, 1.35% phenylalanine, 0.7% isoleucine, 0.5% tyrosine, 1.05% methionine, 0.4% proline, 0.69% valine, 1.1% threonine, 0.4% histidine, 0.5% alanine, 1.02% glutamine, 1.6% glutamic acid, 0.7% glycine, 0.3% serine, 1% arginine, 0.1% aspartic acid, 1.23% asparagine, 0.6% lysine and 0.6% leucine. Quantitative phytochemical analysis shows that the pulp contains 3.45% saponins, 57.01%`flavonoids, 7.12% tannins, 0.0001% alkaloids, 0.0001% glycosides, 0.0003% resins, 0.0002% terpenoids, 0.0001% steroids and 0.0003% cyanogenic glycosides.The results of the acute toxicity show that the methanol extract is not toxic to the mice at concentrations up to 5000mg/kg body weight. From the results obtained, the animals receiving 100mg/kg b.w of the methanolic extract showed significantly reduced (p<0.05) serum levels of glucose, bilirubin, low density lipoprotein cholesterol and ALT after the 14 day study compared to the 28 day study. However, no significant difference (p>0.05) was also observed across the groups in their serum ALP, AST, creatinine, urea, cholesterol, TAG, albumin and globulin levels on the 14th day compared with the 28th day. A significant difference (p<0.05) was observed in the malondaldehyde and serum protein concentrations in the 500mg/kg b.w test group while glucose concentration decreased significantly (p<0.05) in the 100mg/kg b.w and 500mg/kg b.w test group after the 14 day study compared with the 28 day study. High density lipoprotein cholesterol level significantly increased (p<0.05) in the 200mg/kg b.w test group. Histopathological examination shows normal liver architecture across the groups at 100mg/kg b.w, 200mg/kg b.w and 500mg/kg b.w. Kidney sections of rats showing normal glomerulus (G) and renal tubules (arrow) at same concentrations.

TABLE OF CONTENTS

Title Page

Certification

Dedication

Acknowledgement

Abstract

Table of Contents

List of Figures

List of Tables

List of Abbreviations

 

CHAPTER ONE: INTRODUCTION

1.1                   Sweeteners

1.1.1                Common Sweeteners and Their Production

1.1.1.2             Natural Sweeteners

1.1.1.2.1          Honey

1.1.1.2.2          Maple Syrup

1.1.1.2.3          Molasses

1.1.1.2.4          Stevia

1.1.1.2.5          Sucrose

1.1.1.3             Artificial Sweeteners

1.2                   Synsepalum dulcificum

1.3                   Nutrients

1.3.1                Carbohydrates

1.3.2                Proteins

1.3.3                Fats

1.4                   Phytochemicals

1.5                   Antinutrients

1.6                   Vitamins

1.6.1                VitaminA

1.6.2                Vitamin C

1.6.3                Vitamin D

1.6.4                Vitamin E

1.6.5                Vitamin K

1.7                   Antioxidant

1.8                   Some Minerals and Their Biological Functions

1.8.1                Calcium (Ca)

1.8.1.1             Metabolic Functions and Deficiency Symptoms of Calcium

1.8.2                Magnesium (Mg)

1.8.2.1             Metabolic Functions and Deficiency Symptoms of Magnesium

1.8.3                Zinc (Zn)

1.8.3.1             Metabolic Functions and Deficiency Symptoms of Zinc

1.8.4                Iron (Fe)

1.8.4.1             Metabolic Functions and Deficiency Symptoms of Iron

1.8.5                Copper (Cu)

1.8.5.1             Metabolic Functions and Deficiency Symptoms of Copper

1.9                   Blood Glucose

1.9.1                Blood Glucose Regulation

1.10                 Lipids

1.10.1              Lipoproteins: Types and Functions

1.10.1.1           Chylomicrons

1.10.1.2           Very Low Density Lipoprotein (VLDL)

1.10.1.3           Low Density Lipoprotein (LDL)

1.10.1.3.1        Metabolism of Low Density Lipoprotein via LDL Receptor

1.10.1.3.2        Regulation of LDL Receptor

1.10.1.4           High Density Lipoprotein (HDL)

1.11                 Total Cholesterol andCholesterol Balance in Tissues

1.11.1              Diet and Cholesterol Regulation

1.12                 Liver Function Biomarkers

1.12.1              Alanine Aminotransferase

1.12.2              Aspartate Aminotransferase

1.12.3              Alkaline Phosphatase

1.12.4              Clinical and Diagnostic Significance of Liver Function Enzymes

1.12.5              Bilirubin

1.12.6              Serum Protein

1.12.7              Serum Albumin

1.13                 Renal Function Biomarkers

1.13.1              Blood Urea Nitrogen (BUN)

1.13.2              Creatinine

1.14                 Lipid Peroxidation

1.14.1              Initiation

1.14.2              Propagation

1.14.3              Termination

1.14.4              Types of Lipid Peroxidation

1.14.4.1           Non- Enzymatic Lipid Peroxidation

1.14.4.2           Enzymatic Lipid Peroxidation

1.15                 Research Objectives

1.15.1              General Objectives

1.15.2              Specific Objectives

 

CHAPTER TWO : MATERIALS AND METHODS

2.1                   Materials

2.1.1                Plant materials

2.1.2                Animals

2.1.3                Chemicals and Reagents

2.1.4                Equipment /Instruments

2.2                   Methods

2.2.1                Experimental Design

2.2.2                Extraction of Plant Material

2.2.3                Determination of the Extract Yield

2.2.4                Toxicological studies

2.2.4.1             Acute Toxicity Studies and Lethal Dose (LD50) Test

2.2.5                Proximate Analysis

2.2.5.1             Moisture

2.2.5.2             Crude Protein

2.2.5.3             Crude Fat

2.2.5.4             Crude Fibre

2.2.5.5             Ash/Mineral Matter

2.2.5.6             Carbohydrate or Nitrogen Free Extract (NFE)

2.2.6                Estimation of Vitamins

2.2.6.1             Determination of Vitamin A

2.2.6.2             Determination of Vitamin C

2.2.6.3             Determination of Vitamin D

2.2.6.4             Determination of Vitamin E

2.2.6.5             Determination of Vitamin K

2.2.7                Determination of Mineral Content of S. dulcificum Pulp

2.2.7.1             Determination of Phosphorus

2.2.8                Determination of Amino Acid Profile

2.2.8.1             Defatting of the Pulp

2.2.8.2             Hydrolysis of the Pulp

2.2.8.3             Nitrogen Determination

2.2.8.4             Loading of the Hydrolysate into TSM Analyzer

2.2.8.5             Method of Calculating Amino Acid values using Chromatogram Peaks

2.2.9                Qualitative Phytochemical Studies on Synsepalum dulcificum Pulp

2.2.9.1             Test for Alkaloids

2.2.9.2             Test for Glycosides

2.2.9.3             Test for Cyanogenic Glycosides

2.2.9.4             Test for Tannins

2.2.9.5             Test for Saponins

2.2.9.6             Test for Flavonoids

2.2.9.7             Test for Resins

2.2.9.8             Test for Terpenoids and Steroids

2.2.10              Quantitative Phytochemical Analysis of S.dulcificum Pulp

2.2.10.1           Determination of Alkaloids

2.2.10.2           Determination of Cyanogenic Glycosides

2.2.10.3           Determination of Saponins

2.2.10.4           Determination of Flavonoids

2.2.10.5           Determination of Tannins

2.2.10.6           Determination of Steroids

2.2.10.7           Determination of Terpenoids

2.2.11              Antinutrient Analysis of S. dulcificum Pulp

2.2.11.1           Determination of Oxalates

2.2.11.2           Determination of Phytates

2.2.11.3           Determination of Haemagglutanins

2.2.12              Blood Sample Collection for Biochemical Analysis

2.2.13              Biochemical Assays

2.2.13.1           Assay of Alanine Aminotransferase (ALT) Activity

2.2.13.2           Assay of Aspartate Aminotransferase Activity

2.2.13.3           Assay of Alkaline Phosphatase (ALP) Activity

2.2.13.4           Determination of Bilirubin Concentration Using Colorimetric Method

2.2.13.4.1        Determination of Total Bilirubin (TB) Concentration

2.2.13.5           Total Serum Protein Assay

2.2.13.6           Serum Albumin Concentration

2.2.13.7           Creatinine

2.2.13.8           Urea

2.2.13.9           Blood glucose Assay

2.2.13.10         Estimation of Serum Lipid Concentrations

2.2.13.10.1      Estimation of Total Cholesterol Concentration

2.2.13.10.2      Estimation of Low Density Lipoprotein-Cholesterol Concentration

2.2.13.10.3      Estimation of High Density Lipoproteins (HDL)–Cholesterol Concentration

2.2.13.10.4      Estimation of Triacylglycerol

2.2.13.11         Estimation of Lipid Peroxidation

2.2.14              Histopathological Examination

2.2.15              Statistical Analysis

 

 

CHAPTER THREE: RESULTS

3.1       Proximate Composition of S. dulcificum Pulp

3.2       Mineral Composition of S. dulcificum Pulp

3.3       Vitamin Content of S.dulcificum Pulp

3.4       Amino Acid Profile of S. dulcificum Pulp

3.5       Phytochemical Composition of S. dulcificum Pulp

3.6       Antinutritional Composition of S.dulcificum Pulp

3.7       Acute toxicity (LD50) Studies

3.8       Mean Body Weights of Animals

3.9       Effect of S. dulcificumMethanolic Extract Administration on Alkaline Phosphatase (ALP) Activity in Rats

3.10     Effect of S. dulcificumMethanolic Extract Administration on Alanine Aminotransferase (ALT) Activity in Rats

3.11     Effect of S. dulcificumMethanolic Extract Administration on Aspartate Aminotransferase (AST) Activity in Rats

3.12     Effect of S. dulcificumMethanolic Extract Administration on Bilirubin levels in Rats

3.13     Effect of S. dulcificumMethanolic Extract Administration on Total Serum Protein concentration in rats

3.14     Effect of S. dulcificumMethanolic Extract Administration on Serum Albumin Concentration in Rats

3.15     Effect of S. dulcificumMethanolic Extract Administration on Serum Globulinin Rats

3.16     Effect of S. dulcificumMethanolic Extract Administration on Creatinine Level in Rats

3.17     Effect of S. dulcificumMethanolic Extract Administration on Urea Level in Rats

3.18     Effect of S. dulcificumMethanolic Extract Administration on Blood Glucose Concentration in Rats

3.19     Effect of S. dulcificumMethanolic Extract Administration on Cholesterol Concentration in Rats

3.20     Effect of S. dulcificumMethanolic Extract Administration on High Density Lipoprotein Cholesterol Concentration in Rats

3.21     Effect of S. dulcificumMethanolic Extract Administration on Low Density Lipoprotein Cholesterol Concentration in Rats

3.22     Effect of S. dulcificumMethanolic Extract Administration on Triacylglycerol Concentration in Rats

3.23     Effect of S. dulcificumMethanolic Extract Administration on Malondialdehyde Concentration in Rats

3.24     Effect of S. dulcificumMethanol Extract Administration on the Histopathology of Rat Liver [14 days duration]

3.25     Effect of S. dulcificumMethanol Extract Administration on the Histopathology of Rat Liver [28 days duration]

3.26     Effect of S. dulcificumMethanol Extract Administration on the Histopathology of Rat Kidney [14 days duration]

3.27     Effect of S. dulcificumMethanol Extract Administration on the Histopathology of Rat Kidney [28 days duration]

CHAPTER FOUR: DISCUSSION

4.1       Discussion

4.2       Conclusion

4.3       Suggestions For Further Studies

REFERENCES

APPENDICES

LIST OF FIGURES

Figure 1           Structure of Sucrose

Figure 2           Syvsepalum dulcificum Fruit

Figure 3           Synsepalum dulcificum Tree

Figure 4           Structure of Cholesterol

Figure 5           Mechanism of Non-Enzymatic Lipid Peroxidation

Figure 6           Proximate Composition of S. dulcificum Pulp

Figure 7           Amino Acid Analyses of S. dulcificum Pulp

Figure 8:          Effect of S.dulcificum Methanolic Extract Administration on Alkaline phosphatase Activity in Rat

Figure 9           Effect of S.dulcificum Methanolic Extract Administration on Alanine Aminotransferase Activity in Rat

Figure 10         Effect of S.dulcificum Methanolic Extract Administration on Aspartate Aminotransferase Activity in Rat

Figure 11         Effect of S.dulcificum Methanolic Extract Administration on Bilirubin Concentration in Rat

Figure 12         Effect of S.dulcificum Methanolic Extract Administration on Total Serum Protein in Rat

Figure 13         Effect of S.dulcificum Methanolic Extract Administration on Serum Albumin in Rat

Figure 14         Effect of S.dulcificum Methanolic Extract Administration on Serum Globulin in Rat

Figure 15         Effect of S.dulcificum Methanolic Extract Administration on Creatinine Level in rat

Figure 16         Effect of S.dulcificum Methanolic Extract Administration on Urea Level in Rat

Figure 17         Effect of S.dulcificum Methanolic Extract Administration on Blood Glucose Concentration in Rat

Figure 18         Effect of S.dulcificum Methanolic Extract Administration on Total Cholesterol in Rat

Figure 19         Effect of S.dulcificum Methanolic Extract Administration on High-Density Lipoprotein Cholesterol Concentration in Rat

Figure 20         Effect of S.dulcificum Methanolic Extract Administration on Low-Density Lipoprotein Cholesterol Concentration in Rat

Figure 21         Effect of S.dulcificum Methanolic Extract Administration on Triacylglycerol Concentration in Rat

Figure 22         Effect of S.dulcificum Methanolic Extract Administration on Malondialdehyde Concentration in Rat

Figure 23         Photomicrograph of Liver Sections of Rats 14 days Post Administration With S.dulcificum Methanolic Extract

Figure 24         Photomicrograph of Liver Sections of Rats 28 days Post Administration With S.dulcificum Methanolic Extract

Figure 25         Photomicrograph of Kidney Sections of Rats 14 days Post Administration With S.dulcificum Methanolic Extract

Figure 26         Photomicrograph of Kidney Sections of Rats 28 days Post Administration With S.dulcificum Methanolic Extract 

INTRODUCTION

The worsening food crisis and the consequent widespread prevalence of malnutrition in developing and under-developed countries have resulted in high mortality and morbidity rates, especially among infants and children in low-income groups (Enujiugba and Akanbi, 2005). Food has been defined as any substance containing primarily carbohydrates, fats, water, protein, vitamins and minerals that can be taken by an animal or human to meet its nutritional needs and sometimes for pleasure. Items considered as food may be sourced from plants, animals or fungus as well as fermented products like alcohol. Food is also anything solid or liquid that has a chemical composition which enables it provide the body with the material from which it can produce heat or any form of energy, provide material to allow for growth, maintenance, repair or reproduction to proceed and supply substances, which normally regulate the production of energy or the process of growth, repair or reproduction. Food is therefore, the most basic necessity of life (Turner, 2006).

 

Nutrition is the science that deals with all the various factors of which food is composed and the way in which proper nourishment is brought about. The average nutritional requirements of groups of people are fixed and depend on such measurable characteristics as age, sex, height, weight, degree of activity and rate of growth. Good nutrition requires a satisfactory diet which is capable of supporting the individual consuming it, in a state of good health by providing the desired nutrients in required amounts. It must provide the right amount of nutrients and fuel to execute normal physical activity. If the total amount of nutrients provided in the diet is insufficient, a state of under- nutrition develops.

 

Plants are primary sources of medicines, food, shelters and other items used by humans everyday. Their roots, stems, leaves, flowers, fruits and seeds provide for humans (Amaechi, 2009; Hemingsway, 2004). Fruits are sources of minerals, fibre and vitamins which also provide essential nutrients for the human health (Anaka et al., 2009). Some fruits are also known to have antinutritional factors such as phytate and tannins,that can diminish the nutrient bioavailability if they are present at high concentrations (Baum, 2007). It has been reported that these anti-nutritional factors could also help in the treatment and prevention of certain important diseases like the anti-carcinogenic activities reported for phytic acid which has been demonstrated both invivo and invitro (Anaka et al., 2009).

 

The reliance on starchy roots and tubers and certain cereals as main staples result in consumption of non-nutritious foods. The insufficient availability of nutrient rich diets and the high cost of available ones have prompted an intense research into harnessing the potentials of the lesser known and underutilized crops, which are potentially valuable for human and animal foods to maintain a balance between population and agricultural productivity, particularly in the tropical and sub-tropical areas of the world. The challenge of improper nutrition especially in developing countries which include Nigeria, is indeed alarming. The World Health Organization (WHO, 2007) reported that poor nutrition contributes to one out of two deaths associated with infectious diseases among children within five yearsand the aged. Poor diet can have an injurious impact on health, causing deficiency diseases such as scurvy, beriberi and kwashiokor, health-threatening conditions such as obesity, metabolic syndrome, and such common other diseases as cardiovascular diseases, diabetes and osteoporosis. Under-nutrition among pregnant women in developing countries leads to one out of six infants being born with low birth weight, which is a risk factor for neonatal deaths, learning disabilities, mental retardation, poor health and premature death. One out of three people in developing countries is affected by vitamin and mineral deficiencies making them prone to infectious diseases and impaired psycho intellectual development. Under and chronic nutrition problems and diet related chronic diseases account for more than half of the world’s diseases (WHO, 2007). In most of these side effects or diseases, the biochemical and haematological parameters are usually altered. For a food to be considered safe for human and animal consumption, its effect on these parameters need to be investigated to understand the nutritional potentials and safety of such foods with a view to determining their acceptability.

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