PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MISTLETOE LEAVES


PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MISTLETOE LEAVES  

ABSTRACT

The sample of Mistletoe leaves was collected from an orange tree at Ururo in Umunze Anambra State. The leaves was extracted by using some solvent like ethanolic solvent, aqueous solvent n-hexane solvent and ethyl acetate solvent. The extracts was further analyzed which helps in determining the presence of phytochemical compounds in the Mistletoe plant leaf, such as alkaloids, flavonoid, phenol, tannins, steroid and saponins. These phytochemical compounds are found to be antimicrobial active agents. The efficiency of this plant in the treatment of various diseases necessitated the present work in order to identify the classes of natural products present in the leaf of the plant. These pharmacological values of this plant in herbal medicine and as base for the development of new drug.

TABLE OF CONTENT

CONTENT                                                                            

Title page                                                                                        

Declaration                                                                                       

Certification                                                                                     

Dedication                                                                                        

Acknowledgment                                                                              

Table of content                                                                            

Abstract                                                                                           

CHAPTER ONE: PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MYSTETOE LEAVES

1.0    INTRODUCTION                                                                       

AIM AND OBJECTIVES                                                      

LITERATURE REVIEW OF COMMON                    

NAME MISTLE TOE                                                            

1.2    PLANT TAXONOMY                                                           

1.3    PHYTOCHEMICALS                                                           

1.3.1 TANINS                                                                                 

1.3.2 STEROIDS                                                                                     

1.3.3 PHENOLS                                                                            

1.3.4 SAPONINS                                                                           

1.3.5 FLAVONOIDS                                                                      

1.3.6 ALKALOIDS                                                                         

1.4    MEDICAL USES OF MISTLE TOE                                             

CHAPTER TWO: PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MYSTETOE LEAVES

2.0    MATERIALS AND METHODS                                           

2.1    REAGENTS                                                                          

2.2    CHEMICALS USED                                                             

2.3    APPARATUS USED                                                            

2.4    PHYTOCHEMICAL METHOD                                            

2.5    PREPARATION OF THE SAMPLE                                   

2.6    ETHANOLIC EXTRACT                                                      

2.7    N-HEXANE EXTRACT                                                                 

2.8    AQEOUS EXTRACT                                                           

2.9    ETHYL ACETATE EXTRACT                                            

2.10  CHEMICAL TESTS                                                             

2.10.1        TEST FOR ALKAKLOID                                            

2.10.2        TEST FOR TANIN                                                      

2.10.3        TEST FOR FLAVONOID                                           

2.10.4        TEST FOR PHENOLS                                                        

2.10.5        TEST FOR STEROID                                                

2.10.6        EMULSION TEST                                                      

2.10.7        TEST FOR SAPONIN                                                

2.11            EXPERIMENTAL                                                       

2.11.1        PREPARATION OF SAMPLE                                  

2.11.2        PREPARATION OF REAGENTS                             

2.11.3        EXTRACTION OF SAMPLE                                               

2.11.4        AQUEOUS EXTRACT OF THE LEAVE                            

2.11.5        N-HEXANE EXTRACT OF THE LEAVES               

2.11.6        ETHYL ACETATE EXTRACT OF THE LEAVE               

2.12            CHEMICAL TESTS                                                    

2.12.1        ALKALOID                                                                            

2.12.3        FLAVONOID                                                               

2.12.4        STEROID (SALKOWSKI TEST)                               

2.12.5        SAPONIN                                                                    

2.12.6        EMIULSION TEST                                                     

2.13            EXTRACTION OF ALKALOIDS                               

2.14            METHOD OF SEPARATION                                              

2.14            METHODS USED FOR THE TEST                         

AREAS FOLLOWS

CHAPTER THREE: PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MYSTETOE LEAVES

3.0              RESULTS                                                                   

3.1              DISCUSSION ON PHYTOCHEMICAL RESULTS 

TABLE 1:   MINIMAL INHIBITION AND MINIMAL BATERICIDAL

CONCENTRATION OF THE FOUR EXTRACTS

TABLE 2:   MEAN DIAMETER OF ZONE OF INHIBITION (IN MM)

CHAPTER FOUR: PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MYSTETOE LEAVES

DISCUSSION                                                                                 

CONCLUSION                                                                               

RECOMMENDATION                                                                   

REFERENCES                                                                              

CHAPTER ONE

1.0    INTRODUCTION

The use of the leaves, stem and bark of the plants for medicinal purposes is an age long practice. Also in the modern times, and among civilized people, the medicinal use of herbs and shrubs in the treatment of so many diseases both physiological and otherwise is an important breakthrough in pharmocognacy. Mistle toe is a flowering plant which lives on and obtains its nutrition from other plants by parasitizing on its host.

          Mistle toe grows as a partial parasite on a variety of trees, such as apples, oranges, breadfruits across Northern Europe, Asia and Africa. Mistle toe falls under the category of Medicinal plants. Mostly found in tropical and subtropical areas which has proved to contain phytochemicals and other constituents.[1]

          The Mistle toe white berries are potentially poisonous and toxic ad should be avoided. It has been discovered that it is capable of treating cancer, lowering of blood pressure, internal bleeding, convulsion s and so on. Despite the wide use of Mistle toe plant of medicine, the use of mistle toe cannot be routinely recommended. In that case, it should be used under the supervision of a licensed health care professional because it has its indications and side effects.

The efficiency of this plant in the treatment of various diseases necessitated the present work in order to identify the natural product present in the leaf of the plant.

AIM AND OBJECTIVES

The aim of this project is

1.   To determine the class of phytochemical compounds existing in Mistle Toe leaf.

2.   To determine the physiochemical characteristics of Mistle leaf.

3.   To extract and partition the active principle of Mistle toe and ascertain  their antimicrobial activity.

1.1    LITERATURE REVIEW OF COMMON NAME MISTLE TOE

          The literature review of the past work on plant mistle toe showed its distribution an morphology life form and seasonal changes, seasonal changes, growth rate, vegetative reproduction and the controlling environmental factors (biotic and abiotic) affecting the distribution growth and abundance.

This review has shown Viscum L (Synomyns Viscum Mistle toe) is common bushy plant of the family lanrenthecea which grows as an epiphyte on the branches of the deciduous trees. This relationship was observed across the European continent by ancient people who were so impressed with these plants that Mistle toe becomes interwoven into their legend Myths and religious belief.

Extracts produced from Mistle toe as well as certain isolated components are able to stimulate different functions of the immune system. The natural killer cells have been suggested as a candidate for direct tumor cell destruction when one of the Mistle Toe comes into contact with the back of a tree generally through the agency of birds after a few days, it sends forth a thread like root, flattened at the extremity like the proboscus of fly. It finally pierces the back and roots itself firmly in the growing wood from which it has the power of selecting and appropriating to its own use, such juice are filled for its substance. The stems of mistle toe is yellowish and smooth, the leaves are tongue shaped, broader towards the end and is one to three (1 to 3) inches long, very thick and leathery, the leave colour is yellow green and arranged in pairs, with very short footstalks.

          Many popular writers on herbs recommend mistletoe as a treatment for conditions from anxiety to cancer, because of the relatively high price of coffee, some people have even advocated it as a pleasant tasting substitute.

          Over centuries, herbalists through Europe, have relied on a tea and tincture of the berries to treat some of the system associated with rising  blood pressure, headaches, dizziness, loss of energy irritability, etc. European herbalists have a couple of different ways of using mistle toe as a heart sedative and antihypertensive. One take equal parts about two table spoons each of mistle toe and hawthorns berries and lemon balm leaves and steep the two points of boiling water for 25 minutes. One half of the warm tea is taken morning and evening the other way is four teaspoonful of chopped mistle toe in 1-1 four pints of cold water overnight and taking one cup of the cool beverage first thing next morning.

          Other authors also reported some flavournones and chalcones as constituents of mistle toe [2] cuticular waxes of mistle toe show a high content of linolenic acid, with aliphatic constrincts such as alkenes, esters, aldehydes, primary alcohols and free fatty acids present in much lesser amount [3].

          In local application, it eases reheumatic aches and is very effective for acute attacks of Lumbage. It is employed in menstrual disorder excessive menstruation and uterine haemarhage dueto[4]. The detoxication of the leaves is one of the most effective extra used in treatment of high blood pressure in man, epilepsy and other nervous condition[4].

1.2    PLANT TAXONOMY

Botanical   –       Viscum album (LINN)

Family        -        N.O Loranthaceae

Genus        -        viscum, meaning stick

          Mistle toe is a type of plant that grows in tropical and subtropical areas including partly Africa, Asia, America and European countries. It also serves as medicine.

          The well known mistle toe is an ever green parasitic plant growing on the branches of trees where it forms pedent bushes, it is two to five feet in diameter. It has been found that most of the deciduous trees, especially those with soft bark such as old apple, orange , kolanut and bread fruits trees.

These are approximately 3000 parasite angiosperms in 15 plant families and almost all are dicotyledonous. Although many parasite plants contain functional chlorophyll, they depend on their plant hosts for most or at least some of their grab on requirement and for all their nutrients and water needs. By parasitizing other higher plants, they have a competitive advantage over other forms of life because they do not have to compete in soil for their water and nutrient needs.

          The family branthaceae is large and contains at least nine gene, most of which are abundant in the tropic, most species are large showing flowers and attached variety of tree hosts. The family vicarage contains several general but only pharedndron and viscum are important in the legends and myths relating to the mistle toes. On the European continent, viscum album was the major species which primitive man interacted and which formed the basis for many myths legends and religious beliefs.

          In Britain where the Mistle toe grows so abundantly, the plant is called herb de la crop because churching to an old legend, the crose was made from its wood on an account of which it was degraded to be a parasite.

          The English name is said to be derived from the Anglonsaxon mistle toe, than signifying twigs and line, sequel to Prof. Skeat, mistle toe means “birds and twig” a reference to the fact that  the berries line have been used for making bird line. Dr. Prior here defines the word from tan a twig and miste, meaning different from its being unlike the tree it grows on. In the fourteenth century, it was termed mistidine and also lignum cruces, an illusion to the legend just mentioned. The latin name of the genes, Viscum signifying sticky was assigned to it from the lutinous juice of its berry.

1.3    PHYTOCHEMICALS

          These are chemicals found in a plant, its analysis is a process of identifying the types of natural products that is in the plant. These phytochemicals includes alkaloids, phenols, steroids, saponins, and flavonoids.

1.3.1 TANINS

Tannins are esters which can be hydrolyzed by boiling with dilute acid to yield a phenoilic compound usually a derivative of garlic acid [5]. There are two types of tannins, the hydrolysable and nonhydrolyzable or condensed tannins. Hydrolysable ones are broken down by acids or enzymes into simple groups. They are yellow brown amorphous substances which dissolves in hot water to form colloidal dispersion. Nonhydrolyzable tannins are polymers of phenolic compounds and are not soluble in water. They are converted to more complex components when hydrolyzed by acid or enzymes.

1.3.2 STEROIDS

          Steroids are a class of natural product with multiple aestrogenic centre, with high formular weight aliphatic compound whose molecule includes the characteristic steroid nucleus. Several steroids are very active, physiologically and their effects varies widely by ranging from vitamin active to the action of sex hormone. They contain the sense cyclopentophenantrene ring and atomic number system with cholesterol with the exception of vitamins. They occur as glycoside, acylglycocides as esters and in free form most plants are of such compounds having estrogenic centers is choric acid which can be obtained from bile.

1.3.3 PHENOLS

          Phenols are any compound that contains a six membered aromatic ring bonded to a hydroxyl group (OH) phenols are a class of organic  compounds, the chemical name is hydroxyl benzene. The phenol molecules have weak tendencies to base the heat ion from the hydroxyl group releasing in the highly water soluble phenolate anion compared to aliphatic alcohols.

1.3.4 SAPONINS

          These are glycoside with a distinctive foaming characteristic. They are found in many plants but obtained their name soap-plant work.[6] the root which was used historically as a soap, they consist of a polycyclic, a glycone that is either a chloric syteroid or trite. Periods attached us. C3 or ether bond to a sugar side chain, the aglycone is referred to as the saprogenic and steroids. Saponins are called saraparins. The ability of a saponin to foam is caused by combination of non-polar sapogenins and water soluble side chain. They are also known as steroidal glycosides.

1.3.5 FLAVONOIDS

Flavonoids are naturally occurring colouring matter in vegetables. They occur in three main groups namely; chlorophyll, arythenoids and flavnoids, which are involved in photosynthesis, they add flavor or aroma to vegetable spices and food, flavonoid pigments are related compounds which are widely distributed in plant tissues and also found in cell saps. They have a basic carbon selection chain that is together with oxygen. The occurring flavonoid have glycoside, implicating the presence of sugar moiety. All flavounoids are structurally related to flavonoid.[7].

          They are polyphenolic compounds that are ubiquitous in nature and are categorized according to chemical strictness into flavonoids, Isoflavones, capelins, anthocynaides and chalcogenes, they are antioxidants and also used as antiviral, antinallergy, anti inflammatory. Its antioxidant properties protects cells against damaging effects of reactive oxygen species such as singlet oxygen super oxide, neurodecenerative disease[8] epoider mycological studies have showed that flavonoids intake is inversely related to mortality from coronary heat disease to the incidence of heart attack.[9].

1.3.6 ALKALOIDS

          These are group of hetrocyclic nitrogen containing organic base which are producers of amino acid metabolism. They are physiologically active basic substances which are hetrocyclic.

          Alkaloids are toxic to man and may have dramatic physiological activities which are widely used in medicine.

1.4    MEDICAL USES OF MISTLE TOE

The plant can be used in curing fresh wounds quickly. The laboratory studies of this plant has proven that mistle toe kills cancer cells and stimulate the immune system.

          It can be used in treatment of internal bleeding, convulsion, gout, hysteric, whooping cough, ashma, dizziness, loss of menstrual circle, diarrhea, chores (rapid, jerkey monument) and rapid heart beat, due to its calming effects, mistle toe is used as a tranquilizer for various nervous conditions and for the treatment of mental and physical exhaustions. It is also used as long term therapy to prevent hardening of the arteries.

          Nervine antispasmodic atomic and narcotic has a great reputation for curing falling sickness (epilepsy) and other conclusive nervous disorders. It has also been employed in checking  internal haemorrhage.

          The physiological effect of the plant is to lessen and temporarily benumbs such nervous action as is reflected to distant organs of body from some control organ which is the spasm of epilepsy and of other convulsive distempers are alloyed large doses of the plants or of its berries would on the contrary, aggravate these convulsive disorders. Young children have been attacked with convulsions after eating freely of the berries. In a French work on domestic remedies 1682, mistle toe (qui de chene) was considered of a great curative power in epilepsy[10]. Regarding it as a specific for this disease, he procured the parasite from the line trees at Hamiton court, and recommended the powdered leaves as much as would lie on a sixpence, to be given in black cherry water every morning. He was followed in the treatment by others who have testifies to its efficiency as a tonic in nervous disorders, considering the specific herb for St. Vista Dance, it has been employed in convulsious delirium, hysteria, neuralgia, nervous debility, urinary disorders, heart diseases and many others complaining from a week-end and disorder state of nervous system.

.

PHYTOCHEMICAL AND ANTIMICROBIAL ANALYSIS OF MISTLETOE LEAVES



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