PHYTOCHEMICAL AND ANTI INFLAMMATORY PROPERTIES OF METHANOL EXTRACT OF CRATEVA ADANSONII STEM BARK

ABSTRACT

  This research investigated the phytochemical and anti-inflammatory properties of methanol extract of Crateva adansonii stem bark. Although several edible and non-edible plants parts are used in inflammatory treatment, many record has been found of the use of Crateva adansonii stem bark. For this research, fresh stem bark of Crateva adansonii were collected from Asata village in Enugu State. The cuttings were authenticated at the Bioresource development centre. They were then dried at room temperature for one month in an open lab space, grounded into powder and weighed on a beam balance as 460.6g. The powder was soaked for twenty-four to fourty-eight hours in methanol to get a methanol extract and then concentrated into paste at a set temperature range of 30-55⁰C in a water bath. A population of twenty adult wistar Albino rats was used for anti-inflammatory test. The rats were divided into five (5) groups of four (4) albino rats each. They were administered 3% tween-80 mixed with dichloromethane extract of Crateva adansonii and the control was administered with 0.5ml of 3% tween-80. Acute inflammation was induced an hour after test substances were administered by injecting egg albumin in the subplanter region of the right hind paw and edema assessed by mercury displacement for a period of 0-180 minutes. Anti-inflammatory effect was significant within 30 minutes of induced edema with inhibition occurring in three phases of 0-30, 30-60, 60-90. 90-120 to 180 minutes. Inhibition was highest at the third phase. Crateva adansonii barks showed anti-inflammatory effect by inhibiting “premediator

 TABLE OF CONTENTS

CHAPTER ONE

 1.0   Introduction

 1.2   Crateva Adansonii as a Plant

 1.3   Distribution

 1.4   Scientific Classification

 1.5   Properties

 1.6   Research Aim and Objectives

 1.7   Background of Study

 CHAPTER TWO-LITERATURE REVIEW

 2.0   Definition of Inflammation

 2.1   Principle of Inflammation

 2.2   Types of Inflammation

 2.3            Categories of Inflammation Mediated by the Immune  System

 2.4   Mediators of Inflammation

 2.5   Histamine and Serotonin

2.6   The Coagulation Mechanism

 2.7   Fibrinolysis

 2.8   The Kinin –Forming System

 2.9   Inflammation and Diseases

2.10  Ways of Treating Inflammation

 2.11  Immune Selective Anti-inflammatory

 2.12  The Use of Herbs in the Treatment of Inflammation

 2.13  Anti-Inflammatory Drugs

 2.14  Phytochemicals

 2.15  Glycoside

 2.16  Flavonoids

 2.17  Tannins

 2.18  Saponins

3.2.7 Test for Terpenoids

 3.2.8 Test for Anthraquinones

 3.2.9 Test for Coumarins

3.2.10 Test for Phlobatannins

 3.2.11 Test for Flavonoids

 3.2.12 Test for Tannins

 3.2.13 Test for Saponin

 3.2.14 Test for Resins

 3.2.15 Test for Steroids and Terpenoids

3.2.16  Test for Glycoside

3.2.17 Preparation of Reagents for Anti-Inflammatory Test

 3.2.18 Test for Anti –Inflammation Activity

3.2.19 Thin layer Chromatorgaphy (TLC)

 3.2.20 How to Run A TLC Plate

 CHAPTER FOUR

 4.0      Extraction

 4.1            Photochemical analysis of Crateva Adansonii Bark Extract

  CHAPTER FIVE

 5.1 Discussion

 5.2 Conclusion

REFERENCES

 APPENDIX

CHAPTER ONE

  1.0            INTRODUCTION

Inflation is a Latin word (inflammare) which is translated means to set on fire. It is a complex biological response of vascular tissue to harmful stimuli such as pathogens, damaged cells irritant. Inflammation is the reaction of vascularized tissue to local injury caused by certain stimuli like infections, chemicals and biochemical agents, thermal or other physical trauma, antigen-antibody interaction etc (Carol, 1994). Without inflammatory response, wounds will not heal and minor infections would be over weening. Though inflammation aims at limiting damage and restoring function, some enzymes and toxic products within phagocytic cells are released to the extend of damaging the tissue. The advent of anti-inflammatory agents has made inflammation which as been a threat to human life due to its complex, multicontent, to loose its power. These anti-inflammatory agents or drugs help reduce, pain by inhibiting inflammation as opposed to opioids, which affects the central nervous system. It also prevent repairs, prevent and stop the consequences of inflammation by acting on the body responses without directly antagonizing the causative agent (Stedman, 2000). These anti-inflammatory process involves the process of balancing pro-inflammatory acute-phase reactants (Russell et al. 2000), altering biochemical pathway forming prostaglandins by inhibiting cyclooxygenase enzyme from catalyzing the reaction, as a result, suppress, compensate and correct the mechanical and structural abnormalities by assistive device. (Masumoto et al.2009).

The inflammatory reaction is phylogenetically and ontogenetically the oldest defense mechanism. The cells of immune system are widely distributed throughout the body, but if an infection or tissue damage occurs. It is necessary to concentrate them and their products at the site of damage.

Three major events occurring during this response:

1.                 An increased blood supply to the damaged tissue . It is performed by vasodilation. The inflamed tissue look like containing greater number of vessels.

2.                 Increased capillary permeability caused by retraction of the endothelial cells. This permits larger molecules than usual to escape from the capillaries, and thus allows the soluble mediators of immunity to reach the site of inflammation.

 3.                 Leukocytes migrates out of the capillaries into the surrounding tissues. In the earliest stages of inflammation, neutrophils are particularly

prevalent, but later monocytes and lymphocytes migrate towards the site of infection (Ashcroft et al.1999).

For the possibility of surrounding tissue damage, inflammatory responses must be well ordered and controlled. The body must be able to act quickly in some situations, for example to reduce or stop the lost of blood, whereas tissue repair can begin later. Therefore a wide variety of interconnected cellular and humoral (soluble) mechanisms are activated when tissue damage and infections occur. On the other hand, if injury is negligible, the body must have mechanisms which are able to stop tissue damage when the agent is removed. The development of inflammatory reactions is controlled by cytokines, products of plasma enzymes (complement, the coagulation clotting, kinin and fibrolytic pathways), by lipid mediators (prostaglandin and leukotrienes) released from different cells/ and by vasoactive mediators from the mast cells, basophils and platelets. These anti-inflammatory reactions differ. Fast-acting mediators such as vasoactive amines and the product of the kininsystem, modulate the immediate response. Later, newly synthesized mediators such as leukotrienes are involved in the accumulation and activation of other cells. However, in inflammatory reactions initiated by the immune system, the ultimate control is exerted by the antigen itself in the same ways it controls the immune response itself. For this reason, the cellular accumulation at the site of chronic infection or auto-immune reactions where antigen cannot ultimately be eradicated, is quiet different from the sites were antigenic stimulus can be rapidly cleared.

 The nervous system can also participate in the control of inflammation especially axon reflexes, but inflammation may be realized in non-nervous tissues as well.Inflammation may become chronic in certain settings where the acute process characterized by neutrophil infiltration and swelling gives way to predominance of mononuclear phagocytes and lymphocytes.This probably occurs to some degree with the normal healing process but becomesexaggerated and chronic when there’s materials as in certain infections (e.g tuberculosis) or following introduction

of foreign bodies (example asbestos) or deposition of crystals (example urate crystals). Chronic inflammation is often associated with fussion of mononuclear cells to form multinucleated gigantic cells, which eventually become granuloma. Chronic inflammation is seen under of delayed hypersensitivity (Nathan, 2002).

1.2            CRATEVA ADANSONII AS A PLANT.

The flowering tree crateva religiosa (syn crateva adansonii) is called the sacred garlic pear and temple plant, and many other names in a variety of dialets, including Balai, lamok, abiyuch, barna, varuna and bidasi. The tree is sometimes called the spider tree because the showy flower bear long, spidery stamens. It is native to Japan, Australia, much of South East Asia and several South east Asia and several South pacific islands. It is grown elsewhere for fruit especially in part of African continent.

The crateva adansonii plant is a moderate sized, spreading unarmed, deciduous tree growing to a height of 15 meters. Bark is grey, the wood yellowish-white turning light brown when old. Leaves are clustered at the end of the branchlets with a common petiole 5-10 centimeter long, at the summit of which are three leaflets. Leaflets are ovate-lanceolate or ovate 7.5-12 centimeter long, 4-6 centimeter wide. Pointed at the base rather slender pointer at the tip. Flower occur in terminal corymbs, about 5 centimeters in diameter, greenish yellow, and at length purplish.

 Petals are ovate or oblong with the claw haft as long as the limb. Fruit is ovoid or rounded, 3-5 centimeter in diameters, with a hard and rough rind.

 Seeds  are  about  10  centimeter  in  length,  numerous  kidney-shaped,  and embedded in a yellow pulp.

 1.3            DISTRIBUTION

1.                 In waste places, along streams and in thickets near the sea.

 2.                 Occurs in India, Myanmar, Sri lanka, Malasia, Indonesia and China. (nature serve. 2011).

3.                 Sometimes planted as ornamentals tree for its beautiful flower.

1.4            SCIENTIFIC CLASSIFICATION

Kingdom:                            Plantae

 Division:         Angiosperrrrms

 Class:  Eudicots

 Sub- class:      Rosids

 Order: Brassicales

 Family:           Capparaceae

 Genues:          Crateva

  Species:         C.reliigiosa

 Binomial name

 Crateva religiosa  Frost .F.

 The constituents of the plant have been assessed overtime and it has been found in various parts of the plants to contain:

 Bark yield tannin.

 Phytochiemicals screening of extract of dried leaves yielded allcaloids, carbohydrates, tannin, flavonoids, resins, proteins, oils, steroids and terpenoids.

1.5            PROPERTIES

 Generally considered diuretic, anti-inflammatory, laxative, anti-oxidant, hepatoprotective, antilithics, antirheumatic, antiperiodic, contraceptive, anthelmintic.

Bark has a disagreeable smell, the taste slightly bitter, bitting and pungent. Leaves considered stomachic and tonic. 

Roots and bark considered laxative, lithotriptic and alternative, promoting appetite and increasing biliary sections.

Leaves are rubifacient, tonic and febrifugal.

1.6            RESEARCH AIM AND OBJECTIVES

This research aim at investigating the anti-inflammatory properties of methanol extract of crateva adasonii stem bark. And as objectives, to compare the therapeutic potential, (i.e. anti-inflammatory effect ) of the methanol extract of crateva adansonii bark against the experimental standard indomethacin. 

1.7            BACKGROUND OF STUDY

The plant crateva adansonii also known as or also called sacred garlic pear and temple plant. Apart from its medicinal properties which allows the plants to be used as laxative, it is also an edible plant especially the fruits,

berry and young shoot. The plant is used in herbal treatment of tympanites, convulsion, treatment of fever, rheumatism, urinary calculi etc.

 Crateva adansonii is capable of suppressing the free radicals, production, it is been suggested to have anti-inflammatory properties which counter the synthesis of inflammatory mediators thus forming the background of this research.

CHAPTER TWO

LITERATURE REVIEW

 2.0      DEFINITION OF INFLAMMATION

Inflammation (Latin word “inflammo,”I setmeaninglight,“ the body attempt at self-protection; the aim being to remove harmful stimuli, including damaged cells; irritants (Stedman, 1990). Inflammation is part of the body’s immuneInitially,it is beneficialresponsewhen,for.example, your knee sustains a blow and tissues need care and protection. However, sometimes inflammation can cause further inflammation; it can become self-perpetuating. More inflammation is created in response to existing inflammation (Guyton, 1990).

2.1            PRINCIPLE OF INFLAMMATION

 Inflammation is a complex stereotypical reaction of the body expressing the response to damage of its cells and vascularized tissues, e.g. in normal cornea, the true inflammation does not occur (Smith, 2000).

The discovery of the detailed processes of inflammation has revealed a close relationship between inflammation and the immune response.

The five basic symptoms of inflammation – redness (rubor), swelling (tumor), heat (calor), pain (dolor) and deranged function (funtiolaesa) have been known since the ancient Greek and Roman era (Nathan, 2002). These signs are due to extravasation of plasma and infiltration of leukocytes into the site of inflammation. Early investigators considered inflammation a primary host defence system. From this point of view inflammation is the key reaction of the innate immune response but in fact, but inflammation is more than this, since it can lead to death, as an anaphylactic shock or delibitating diseases, asin arthritis and gout (Lentsch & Ward, 2000).

 According to different criteria, inflammatory responses can be divided into several categories. The criteria includes:

1.                 time- hyperacute (peracute), acute, sub acute, and chronic inflammation;

2.                 the main inflammatory manifestation- alteration, exudation, proliferation;

3.                the degree of tissue damage –superficial, profound (bordered, not bordered);

4.                 characteristics picture- nonspecific, specific;

5.                 Immunopathological mechanisms allergic (reaginic) inflammation,

inflammation mediated by cytotoxic antibodies, inflammation mediated by immune complexes, delayed- type hypersensitivity reactions. (Serhan, 2008)

2.2 TYPES OF INFLAMMATION

 There are two basic types of inflammation- acute and chronic inflammation.

Acute inflammation: is of short duration, which could be anything from a few minutes to a few days. Such inflammation is caused by foreign substance entering the body, or by physical damage. A viral infection may also precipitate acute inflammation.

Chronic inflammation, on the other hand, is long lasting. It may persist for weeks, months or even years. Chronic inflammation may be brought on by acute inflammation or maybe the result of an auto immune disease.

Two other, less common types of inflammation are sub-acute inflammation and granulomatous chronic inflammation.

Sub- acute inflammation has clinical features of acute and chronic inflammation- it is an intermediary stages between the two. Granulomatous chronic inflammation: This is a special type of chronic inflammation that is associated with tuberculosis and some lesser known diseases