PHYTOCHEMICAL AND ANTIMICROBIAL STUDIES OF Spermacoce verticillata (Rubiaceae) G.F.W. Meyer

Title page
Table of content

CHAPTER ONE: Introduction
1.0       General introduction
1.1       Bacterial infection
1.2       Statement of research problem
1.3       Justification of the study
1.4       Aim and Objectives of the study
1.5       Statement of Research Hypothesis

CHAPTER TWO: Literature review
2.1       Spermacoce verticillata
2.1.1    Morphology of Spermacoce verticillata
2.1.2    Geographical distribution
2.1.3    Classification of the plant Spermacoce verticillata
2.1.4    Ethno medicinal uses
2.1.5    Pharmacological actions of Spermacoce verticillata and other species in       the genus and family

3.0       Material and methods
3.1       Materials
3.1.1    Solvents/Reagent
3.2       Methods
3.2.1    Collection, identification and preparation of plant materials
3.2.2    Extraction
3.2.3    Partitioning (liquid-liquid partitioning)
3.3       Preliminary phytochemical screening
3.3.1    Test for sterols/Terpenes
3.3.2    Test for Flavonoids
3.3.3    Test for Alkaloids
3.3.4    Test for Tannins
3.3.5    Test for Anthraquinones
3.3.6    Test for Saponins
3.4       Chromatographic procedures
3.4.1    Thin layer chromatography (TLC)
3.4.2    Column Chromatography
3.4.3    Gel filtration chromatography
3.4.4    Preparative thin layer chromatography (PTLC)
3.4.5    Spectral Analysis
3.5       Chromatographic Separation
3.5.1    Thin layer chromatography
3.5.2    Column chromatographic separation of hexane fraction
3.5.3    Preparative thin layer chromatography (PTLC)
3.5.4    Spectroscopic Analysis
3.5.5    Melting Point
3.6       Antimicrobial screening
3.6.1    Test organisms
3.6.2    Susceptibility studies
3.6.3    Minimum inhibitory Concentration (MIC)
3.6.4    Minimum Bactericidal Concentration (MBC)

4.0       Results
4.1       Phytochemical studies
4.1.1    Preliminary phytochemical Screening
4.1.2    Results of thin layer chromatography
4.2       Result of column chromatographic separation
4.2.1    Isolation of S1
4.2.2    TLC analysis of S1
4.2.3    Solubility of S1
4.3       Spectral analysis of S1
4.3.1    Results of NMR Proton Magnetic Resonance Spectral of S1 Carbon-13 Magnetic Resonance Spectral of S1
4.4       Result of antimicrobial activity studies

5.0       Discussion
5.1       Phytochemical studies
5.2       Antimicrobial activity
5.3       Summary
5.4       Conclusion

The plant Spermacoce verticillata is used in ethnomedicine for the treatment of itches, diarrhoea, sores and other ailments. The hexane soluble fraction of acetone extract of the whole plant Spermacoce verticillata was subjected to phytochemical and antimicrobial screening. The phytochemical techniques employed were dry column vacuum chromatography and preparative thin layer chromatography. The antimicrobial activity was studied using agar diffusion, and broth dilution methods. The result of preliminary phytochemical screening carried out on the crude acetone extract revealed the presence of carbohydrates, Flavonoids, Anthraquinones, terpernoids, Tannis and Steroids; Tannins flavonoids, and Steroids/terpenes were also found to be present in the hexane fraction. Extensive chromatographic separation of the hexane soluble fraction using silica gel, dry column chromatography, followed by preparative thin layer chromatography led to the isolation of stigmasterol. The structure of this compound was established by spectral analysis including 1D and 2D NMR. The result suggest that the hexane fraction of Spermacoce verticillata plant possess phytochemical constituents that may be useful in the management ofmicrobial diseases.

1.0              General Introduction

The use of plants and their preparations to treat infectious diseases is an ageold practice and in the past possibly the only method available (Peach, 1995). However, the systemic study of plants for detecting antimicrobial activity is of comparatively recent origin. These investigations have been triggered by the emergence and spread of antibiotic resistant microorganisms causing the effective lifespan of existing antibiotics limited. Hence the plant kingdom is being screened for newer and effective chemotherapeutic agents. Higher plants can serve both as potential antimicrobial crude drugs as well as a source of new antiinfective agents (Peach, 1995).

It is evident, that even though scientific advances have been made in our quest to understand the physiology of the body, biotechnology and the treatment of diseases, natural products remain a crucial, cheap and uncontroversial component of the comprehensive health care strategy for the future (Patwadhan, 2005). Natural products such as plant extracts, either as pure compounds or as standardized extracts, provide unlimited opportunities for new drugs discoveries because of the unmatched availability of chemical diversity (Cosa et al., 2006).

In the last decades, various studies have demonstrated that plants serve as reservoirs for innumerable microorganisms known as endophytes; these microorganisms live in the host plant at least for one period of their life cycle, without causing apparent harm to them (Petrini, 1991; Bacon and White, 2000). Even with the existence of uncountable epiphytic and soil microorganism, diverse studies have shown the potential of endophytes as a promising source of natural products for the discovery of a variety of different classes of bioactive molecules to be.......

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