TOXIC PROFILE OF THE METHANOL LEAF EXTRACT OF Senna siamea AND ITS ANTIOXIDANT PROPERTIES

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ABSTRACT


Medicinal plants have been assumed to be good in detoxifications because they contain bioactive compounds capable of doing this. However, some plants are very toxic to both humans and animals with the potential of damaging certain organs in the body. Many plant-derived substances, collectively termed “phytochemicals,” are becoming increasingly known for their antioxidant activity. This study is aimed at investigating the toxic and as well as the antioxidant profile of methanol extract of Senna siamea leaves, extensively used in Nigeria folk medicine to manage various ailments. The potent metabolites were quantified using standard methods and the antioxidant scavenging potentials of the extract on DPPH and superoxide radicals were also determined. Both acute and sub-acute toxicity profile were evaluated by determining the LD50, liver and kidney function tests, which was then correlated and confirmed with histopathologic techniques of hepatocytes and kidney cells. Furthermore, haematological parameters of the test rats were determined. The qualitative phytochemical screening of the leaves of S. siamea revealed the presence of proteins, carbohydrates, tannins, alkaloids, steroids, glycosides, flavonoids, reducing sugars, terpenoids and quinones in the leaves. The result of these tests also indicates that tannin, carbohydrates, reducing sugar, terpenoids, alkaloids, total phenols, glycosides and flavonoids were contained in various amounts when quantified. The antioxidant vitamin contents of the extract was also quantified and this showed that vitamin E was significantly higher (p< 0.05), (181.70±2.47 mg/100g) when compared to vitamin A (40.35±13.60 mg/100g) and vitamin C (4.11±0.06 mg/100g). The extract scavenged DPPH and superoxide radicals in concentration dependent manner. The EC50 of DPPH., superoxide radical were 12.59±0.00 and 39.38±0.01 respectively. Acute toxic test shows no death was observed in any group of the mice used, indicating that the extract could be tolerated by the mice at 5000mg/kg bw, the highest concentration used. The liver marker enzymes AST and ALP showed significantly increased (p<0 .05="" activity="" alp="" changes="" in="" no="" p="" serum="" showed="" significant="" their="" while="">0.05) when compared to the control. Furthermore, bilirubin concentration were significantly increased (p<0 .05="" 3="" during="" sup="" the="">rd
and 4th week of administration of extract to the 4th group. The kidney function test showed a significant increase (p<0 .05="" 1="" a="" and="" concentration="" creatinine="" first="" in="" increase="" observed="" p="" second="" serum="" significant="" sup="" the="" urea="" was="" week.="">st week of group 4. The result of the blood electrolytes showed no significant difference (p>0.05) in sodium ion but significant changes (p<0 .05="" and="" any="" blood="" cell="" changes="" chloride="" did="" hematological="" hemoglobin="" hite="" in="" ion.="" levels="" not="" observed="" of="" p="" packed="" parameters="" potassium="" produce="" significant="" the="" volume="" were="">0.05). Significant reduction (p< 0.05) was observed in catalase activities of the 2nd and 3rd weeks of extract administration. These results obtained indicate that the extract could be toxic to hepatocytes and kidney cells at higher concentration.

TABLE OF CONTENTS

Title Page
Abstract
Table of Contents
List of Figures
List of Tables
List of Plates
List of Abbreviations

CHAPTER ONE: INTRODUCTION
1.1       Senna siamea
1.1.1    Taxonomy and nomenclature
1.1.2    Botanical description, ecology and cultivation
1.1.3    Uses of senna siamea
1.1.3.1 Therapeutic Uses
1.1.3.2 Economical and industrial Uses
1.2       Toxicity: an overview
1.2.1    Liver: morphology, functions and toxicity
1.2.1.1 Alanine aminotransferases (ALT)
1.2.1.2 Aspartate aminotransferase (AST)
1.2.1.3 Alkaline phosphatase (ALP)
1.2.1.4 Total bilirubin
1.2.2    Kidney: morphology, functions and toxicity
1.2.2.1 Creatinine
1.2.2.2 Blood urea nitrogen (BUN)
1.2.2.3 Blood electrolytes
1.2.2.3.1   Chloride ion
1.2.2.3.2          Sodium ion
1.2.2.3.3          Potassium ion
1.3       Antioxidants and free radicals
1.3.1    Antioxidant
1.3.1.1 Enzymatic antioxidants
1.3.1.1.1          Superoxide dismutases (SODs)
1.3.1.1.2          Catalase
1.3.1.2 Antioxidants vitamins
1.3.1.2.1          Vitamin E
1.3.1.2.2          Vitamin C
1.3.1.2.3          Beta-carotene
1.3.2    Free radicals
1.3.2.2 Hydroxyl radical (•OH)
1.3.2.3 Nitric oxide radical (No•)
1.3.2.4 Superoxide radical (•O2–)
1.3.2.5 Diphenylpicrylhydrazyl (DPPH) radical
1.4       Haematological parameters
1.4.1    Packed cell volume (PCV)
1.4.2    White blood cell (WBC)
1.4.3    Haemoglobin (Hb)
1.5       Phytochemicals
1.5.1    Tannins
1.5.2    Glycoside
1.5.3    Quinones
1.5.4    Flavonoids
1.5.5    Terpenoids
1.5.6    Alkaloids
1.6       Aim and objectives
1.6.1    Aim of the study
1.6.2    Objectives of study

CHAPTER TWO:  Materials and Methods
2.1       Materials
2.1.1    Collection of plant material
2.1.2    Animals
2.1.3    Chemicals/reagents
2.1.4    Instruments/equipment
2.2       Methods
2.2.1    Preparation of senna siamea leaves extract
2.2.2    Experimental design for Median Lethal dose
2.2.3    Sub-acute toxicity profile
2.2.4    Qualitative phytochemical analysis
2.2.4.1 Qualitative determination of protein (millon’s test)
2.2.4.2 Qualitative determination of alkaloids (wagner’s test)
2.2.4.3 Qualitative determination of glycosides (fehling’s test)
2.2.4.4 Qualitative determination of reducing sugar
2.2.4.5 Qualitative determination of saponins
2.2.4.6 Qualitative determination of flavonoids
2.2.4.7 Qualitative determination of tannins
2.2.4.8 Qualitative determination of cardiac glycosides
2.2.4.9 Qualitative determination of resins
2.2.4.10 Qualitative determination of carbohydrates (molisch’s test)
2.2.4.11 Qualitative determination of steroids
2.2.4.12Qualitative determination of acidic content
2.2.5 Quantitative phytochemical analysis
2.2.5.1 Quantitative determination of alkaloid content
2.2.5.2 Quantitative determination of carbohydrate content
2.2.5.3 Quantitative determination of reducing sugar content
2.2.5.4Quantitative determination of flavonoid content
2.2.5.5 Quantitative determination of tannin content
2.2.5.6 Quantitative determination of total phenolic content
2.2.5.7 Quantitative determination of terpenoid content
2.2.5.8 Quantitative determination of steroid content
2.2.6 Determination of antioxidant vitamin contents
2.2.6.1 Vitamin A
2.2.6.2 Vitamin C
2.2.6.3 Vitamin E
2.2.7    Radical scavenging activity
2.2.7.1 DPPH scavenging activity
2.2.7.2 Superoxide radical-scavenging assay
2.2.8    Liver function tests
2.2.8.1 Alanine aminotransferase (ALT)
2.2.8.2 Aspartate aminotransferase (AST)
2.2.8.3 Alkaline phosphatase (ALP)
2.2.8.4 Bilirubin
2.2.9    Renal function tests
2.2.9.1 Sodium ion
2.2.9.2 Potassiumion
2.2.9.3 Chlorideion
2.2.9.4 Urea
2.2.9.5 Creatinine
2.2.10  Antioxidant Enzyme
2.2.10.1 Catalase assay
2.2.11  Haematology
2.2.11.1 Total white blood cell (WBC) count
2.2.11.2 Packed cell volume (PCV)
2.2.11.3 Haemoglobin estimation (Hb)
2.2.12  Histopathological examination
2.2.13  Statistical analysis

CHAPTER THREE: RESULTS
3.1       Phytochemical analysis of senna siamea
3.1.1    Qualitative phytochemical analysis
3.1.2    Quantitative phytochemical analysis
3.2       In-vitro antioxidant properties
3.2.1    Effect of mess on DPPH radical scavenging activity
3.2.2    Effect of mess on superoxide radical scavenging activity
3.3       Effect of MESS on in-vivo catalase activity
3.4       Antioxidant vitamins  content of MESS
3.5       Toxicological studies
3.5.1    Lethal Median dose determination
3.5.2    Liver function tests
3.5.2.1 Effect of MESS on Aspartate aminotransferase (AST)
3.5.2.2 Effect of MESS on Alanine aminotransferase (ALT)
3.5.2.3 Effect of MESS on Alkaline phosphatase (ALP)
3.5.2.4 Effect of MESS on Total bilirubin
3.5.2.5 Effect of MESS on Direct bilirubin
3.5.3    Kidney test
3.5.3.1 Effect of MESS on Urea
3.5.3.2 Effect of MESS on creatinine
3.5.3.3 Effect of MESS on chlorideion
3.5.3.4 Effect of MESS on Potassium
3.5.3.5 Effect of MESS on sodium
3.5.5    Haematological parameters
3.5.5.1 Packed cell volume (PCV)
3.5.5.2 Haemoglobin (Hb) concentration
3.5.5.3 White blood cell count (WBC)
3.6       Histopathological examination
3.6.1    The liver
3.6.1.1 Histology of group 1 liver
3.6.1.2 Histology of group 2 liver
3.6.1.3 Histology of group 3 liver
3.6.1.4 Histology of group 4 liver
3.6.1.5 Summary of histology of liver cells
3.6.2 The kidney
3.6.2.1 Histology of group 1 kidney
3.6.2.2 Histology of group 2 kidney
3.6.2.3 Histology of group 3 kidney
3.6.2.4 Histology of group 4 kidney
3.6.2.5 Summary of histology of kidney cells

CHAPTER FOUR: DISCUSSION
4.1       Discussion
4.2       Conclusion
4.3       Recommendation
References
Appendices


CHAPTER ONE

INTRODUCTION


Medicinal plants are believed to be important sources of new chemical substances with potential therapeutic effects. Their activities depend on its phytochemical contents (Meyer et al., 1997). Presently, it is estimated that about 80% of the world’s population are dependent on medicinal plants or plant-derived products for their health needs (Shri, 2003). The use of medicinal plant is presently on the increase due to its availability, affordability, accessibility, and promising efficacy comparable to the often high cost and adverse effects associated with standard synthetic drug agents (Kwada and Tella, 2009). One therefore expect bioactive compounds obtained from such plants to have low animal and human toxicity. However, people are often unaware of important similarities and differences between medicinal herbs and approved medications (Williamson et al., 1996). Some mistakenly think of herbs as natural alternatives to chemicals, failing to recognize that herbs are composed of bioactive chemicals, some of which may be toxic (Fabricant and Farnsworth, 2001). Medicinal plants have been documented to have advantage in toxicity considerations based on their long term use (Zhu et al., 2002).Some plants are very toxic to both humans and animals with the potential of damaging certain organs in the body. (Edoga et al., 2005). This calls for caution in the use of medicinal plants. Toxicity studies on medicinal plants or extracts usually determine the level of safety particularly during the development of drugs (Jaijoy et al., 2010).The use of traditional and herbal medicine is widely practiced in Nigeria and Senna siamea is one of such plants used in Nigerian folklore medicine. It is hence necessary to evaluate the safety of the use of S. siamea in wistar rats.......


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