PHYTOCHEMICAL AND ANTIMICROBIAL SCREENINGOF THE STEM BARK EXTRACT(S) OFINDIGOFERA ARRECTAHOCHST EX A. RICH(FABACEAE

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ABSTRACT

The Pulverized stem bark of Indigofera arrectawas exhaustively extracted with methanol and concentrated in vacuo using rotary evaporator at 40 0C.The extract was later subjected to solvent partitioning to yield soluble extracts of n-hexane, ethyl acetate, chloroform, and methanol. Genernal phytochemical screening of the fractions revealed the presence of secondary metabolites such as cardiac glycoside,steroid, terpenes flavonoids and tannins. The antimicrobial activity against S. aureus,S. pyogenes,S.feacalis, S.typhii, E.coli C. ulcerans,P. vulgaris and C.albicans was tested using the tube dilution and agar diffusion methods as outlined by the NCCLS. The results of the antimicrobial activity as indicated by the zonesof inhibition of growth of microorganism ranged from 20mm to 40mm for the n-hexane extract, 16mm to 21mm for ethyl acetate extract and 20mm to 27mm for the methanol extract. The MIC result for the n-hexane, ethyl acetate and methanol extracts ranged from 7.5mg/ml to 15mg/ml. The MIC of 15mg/ml exhibited by the n-hexane extract against both gram positive and gram negative bacteria indicates broad spectrum activity of Indigofera arrect. The n-hexane fractions was subjected to Column Chromatography using silica gel to yield 87 fractions, which were combined based on their thin layer chromatography analysis and recrystallized in methanol to give a pure white crystalline powder, which melts at 144oC. The structure of the isolated compound was established by spectroscopic analysis and by direct comparison of the data obtained with those reported in literature to be Stigmasterol (3β,22E-Sigmasta-5,22-dien-3-ol).

TABLE OF CONTENTS

 

Title page

Abstract

Table of contents

List of abbreviations

 

CHAPTER ONE

1.0       Introduction

1.1       Definition of a drug

1.2       Medicinal plants

1.3       Medicinal plant research

1.4       Aim

1.4.1    Objectives

1.5       Scope and limitations of research

1.6       Justification of the research

 

CHAPTER TWO

2.0       Literature review

2.1       Botanical description of the genus Indigofera arrecta

2.1.1    Other botanical information

2.1.2    Origin and geographical distribution

2.2       Chemical constituents of indigofera arrecta

2.2.1    Traditional medicinal uses

2.2.2    Uses of some of the genus

2.3       Production and international trade

2.3.1    Pharmacological properties

2.3.2 Adulterations and substitutes

2.3.3 Growth and development

2.3.4    Ecology

2.3.5 Management

2.3.6    Propagation and planting

2.3.7    Diseases and pests

2.3.8    Harvesting

2.3.9    Yield

2.4       Handling after harvest

2.4.1    General description of Indigofera arrecta (HOCHST  EX.A.RICH )

2.5       Review of some natural products from plants, tests and  their  uses

2.5.1Alkaloids

2.5.2Flavonoids

2.5.3    Saponins

2.5.4    Glycosides

2.5.5    Tannins

2.5.6    Steroids

2.5.7.   Terpenoids

2.6       Factors which can affect the level or the composition of the active ingredients in medicinal plant

2.7       Some microorganisms and their effects on the human body

2.7.1    Staphylococci

2.7.2    Streptococci

2.7.3    Candida

2.7.4    Enterobacteriaceae

2.7.5    Klebsiella

2.7.6    Escherichia coli

2.7.7    Salmonellae

 

CHAPTER THREE

3.0       Materials and method

3.1       Materials/reagents/equipment and analytical procedure

3.1.1    Solvents

3.1.2    Equipment

3.1.3    Reagent

3.1.4    Microbial media, test organisms and equipment for antimicobial test

3.1.5    The identification and preparation of plant material

3.1.6    Extraction procedure for crude extract

3.2       Preliminary phytochemical screening

3.2.1    Test for steroids/terpenes

3.2.2    Test for flavonoids

3.2.3    Test for alkaloids

3.2.4    Test for tannins

3.2.5    Test for anthraquinones

3.2.6    Test for saponins

3.2.7    Test for glycoside (fecl3 test)

3.3       Antimicrobial screening

3.3.1    Preparation of bacterial test organisms

3.3.2    Preparation of fungal test organisms

3.3.3    The stock dilution of the plant extracts

3.3.4    Preparation of the nutrient agar

3.3.5    Preparation of the sabouraud dextrose agar media

3.3.6    The punched agar diffusion method [bryant, 1972]

3.3.7    Preparation of inoculums of test organisms

3.3.8    Sensitivity test of the extract using agar diffusion method

3.3.9    Determination of minimum inhibitory concentration using tube dilution method

3.4       Minimum bactericidal concentration (mbc)

3.4.1    Chromatographic purification of extracts

3.4.2    Thin layer chromatography (TLC)

3.4.3    Column chromatography

3.4.4    Solvents system/elution

3.4.5    Gel filtration chromatography

3.4.6    Thin layer chromatography of the n-hexane extract

3.4.7    Column chromatography of n-hexane fraction

 

CHAPTER FOUR

4.0       Results

4.1       Result of extraction

4.2       Result of phytochemical screening

4.3       Results of antimicrobial activity

4.4       Result of chromatographic separation

4.5       Column chromatography of n-hexane fraction

4.6       Isolation of EB

4.7       TLC analysis of EB

4.8       Result of antimicrobial activity of compound EB

 

CHAPTER FIVE

5.0       Discussion of result

5.1       Extraction

5.2       Phytochemical screening

5.3       Antimicrobial screening

5.4       Physical and chemical properties of EB

5.4.1    Spectral analysis

5.4.2    FTIR

5.4.3    1H NMR

5.4.4    13C NMR

5.4.5    DEPT

 

CHAPTER SIX

6.0       Summary, conclusion and recommendation

6.1.      Summary

6.2       Conclusion

6.3       Recommendation

References

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