ABSTRACT
Indiscriminate use of antibiotics is very common in developing countries, it is a major cause for development of antimicrobial-resistance. Thus it has now become essential to look for newer antibiotics mostly plants. Azadirachta indica is an Indian tree well known for its several pharmacological activities, and has been used against a few diseases in traditional medicine. This study was undertaken to screen for the phytochemical constituents as well as to investigate the antibacterial potential of Azadirachta indica leaf extract on Pseudomonas aeruginosa, Salmonella typhi, Staphylococcus aureus and Escherichia coli. The leaves of Azadirachta indica were collected, dried and was extracted with methanol and water. The susceptibility of these organisms to the methanol and aqueous extracts of Azadirachta indica were evaluated using the agar well diffusion method at various dilutions of 200 mg/ml, 100 mg/ml and 50mg/ml. Cultures of the test organisms were grown on Mueller Hinton agar plates. The agar plates were then incubated at 37°C for 24hours after which the zones of inhibition were measured. The results of this study demonstrated that, for all test organisms screened, the highest concentration (200mg/ml) of the methanol showed maximum antimicrobial action with zones on inhibition in the range of 9.0mm to 14.0mm. The methanol extract of Azadirachta indica demonstrated significant inhibitory activity against S. aureus compared to the other test microorganisms with an MIC value of 12.5mg/ml. Evaluation of the phytochemicals was made using standard methods. The phytochemical screening indicated the presence of alkaloids, saponins, tannins, flavonoids and phenolic content in the leaves. This result indicates that these plant materials have potential medicinal uses and the methanolic and water extracts can protect against some common microorganisms and thus may be a prospective therapeutic agent to combat antibiotic resistant bacteria.
TABLE OF CONTENTS
Title page i
Certification ii
Dedication iii
Acknowledgements iv
Table of Contents v
List of Tables vii
List of Figures viii
Abstract ix
CHAPTER ONE
1.0 INTRODUCTION
1.1 Aim of the Study 3
1.2 Objectives of the Study 3
1.3 Statement of the Problem 3
1.4 Justification of the Study 3
CHAPTER
TWO
2.0
LITERATURE REVIEW 5
2.1 Incidence of Antibiotics Resistance 5
2.2 Antibacterial Natural Products 5
2.3 Mechanisms of Action of Antibacterial agents 6
2.4 Clinical Bacterial Isolates 7
2.4.1 Escherichia coli 7
2.4.2 Staphylococcus aureus 7
2.4.3 Pseudomonas aeruginosa 9
2.5 Description of Azadirachta indica 9
2.6 Pharmacological and Therapeutic properties of
Azadirachta indica 12
2.7 Compounds isolated from Azadirachta indica 12
CHAPTER
THREE
3.0
MATERIALS AND METHODS 14
3.1 Preparation of Media
used 14
3.2 Sterilization 14
3.3 Collection of Plant
Materials 14
3.4 Preparation of Plant
Extracts 14
3.5 Microbial Strains 15
3.6 Antibacterial
Susceptibility Testing 15
3.7 Minimum Inhibitory
Concentration 15
3.8 Quantitative
Phytochemical Analysis 16
3.8.1 Determination of
saponins 16
3.8.2 Determination of
tannins 17
3.8.3 Determination of
alkaloids 18
3.8.4 Determination of
flavonoids 19
3.8.5 Determination of
phenolic content 19
CHAPTER
FOUR
4.0 RESULTS
CHAPTER
FIVE
5.0
DISCUSSION, CONCLUSION AND RECOMMENDATIONS 26
5.1 Discusion 26
5.2 Conclusion 29
5.3 Recommendations 29
REFERENCES 30
LIST OF TABLES
Table 1: Diameter zone of
inhibition (mm) produced by extracts of Azadirachta
indica against the test organisms. 22
Table 2: MIC and MBC values
(mg/ml) of the extract against the test organisms 23
Table 3: Qualitative
Phytochemical Screening of the Plant 24
Table 4: Quantitative
Phytochemical composition of Azadirachta
indica 25
LIST OF FIGURES
Fig. 1:Azadirachta indica leaves
11
CHAPTER ONE
1.0 INTRODUCTION
Rising antibiotic resistance and the
scarcity of new antimicrobials has long been acknowledged. A major challenge in
global health care is the need for novel, effective and affordable medicines to
treat microbial infections, especially in developing countries of the world,
where up to one-half of deaths are due to infectious diseases. However,
indiscriminate use of antibiotics in human and veterinary healthcare systems
has lead to the emergence of multi-drug resistant (MDR) strains of different
groups of microorganisms. The emergence and dissemination of Multi-drug
resistant bacteria has made chemically synthesized antibiotics ineffective for
the treatment of infectious diseases caused by such bacteria. Antimicrobial
resistance (AMR) is the ability of microorganisms such as bacteria and fungi to
grow despite exposure to antimicrobial (antibacterial or antifungal) agent
designed to inhibit their growth. In general, microorganisms exhibit antimicrobial
resistance by innate (e.g., absence of drug target site) and/or acquired (e.g.,
enzymatic degradation of drug) mechanisms conferred by antimicrobial resistance
genes (ARGs) acquired via horizontal gene transfer (HGT) (transformation,
transduction, and conjugation) from other microorganisms. These circumstances have
propelled the researchers and scientists to explore new antimicrobial
substances from various sources such as medicinal plants. The green medicines
are widely believed as safe and dependable in contrast with expensive synthetic
drugs that have undesirable side effects along with beneficial effects. The
plants have been in use in traditional medicine worldwide since long time but
are still understudied, particularly in clinical microbiology.
Nature has been a source of medicinal
treatment for thousands of years and plant - based systems continue to play an
essential role in the primary health care of 80% of the world's underdeveloped
and developing countries. Some of these
plants contain secondary metabolites which can affect humans and mammals.
Plants have formed the basis of traditional medicine system that has been the
way of life for thousands of years for the treatment of conditions such as
malaria, ulcers, cardiovascular diseases and skin problems. Despite the limited
existence of clinical trials to support therapeutic claims, its use has
expanded over time. Mostly, herbs and species contain polyphenols which are
most powerful natural antioxidants and are highly valued for their antioxidant,
and antimicrobial effects. Antioxidant compounds like phenolic acids,
polyphenols and flavoniods inhibit the mechanism that leads to degenerative
diseases (Hamid et al., 2010).
Knowledge of herbs has been handed down
from generation to generation, many natural products could serve as the
starting point in the development of modern medicines because of their
pharmacological and therapeutic activities, and during the last decade, an
increase in their use has been observed in metropolitan areas of developed
countries (Dhaliya et al., 2013).
However, some of them are known to carry
antibacterial properties as well.
The screening of plants for medicinal
purposes represent a serious effort to discover newer, safer and possibly more
effective drugs with the potential of fighting pathogenic bacterial and fungi.
(Abubakar, 2010).
Products derived from Azadirachta indica is widely recommended in a variety of biological
uses such as anti-allergic, antimicrobial, anti-malaria, anti- ulcer, anti
-tumor, incesticidal and pesticidal .
1.1 AIM OF THE STUDY.
The aim of the study is to evaluate the
antibacterial activity of Azadirachta
indica against clinical bacterial isolates
in order to know whether it has any prospect as a future antibiotics.
1.2 OBJECTIVES OF THE STUDY.
This project work was designed to
determine the followings;
1.
To determine the effects of Methanol and aqueous
extract of Azadirachta indica on some clinical bacterial
isolates.
2.
To identify the major phytochemicals in Azadirachta indica.
1.3 STATEMENT OF THE PROBLEM.
The use of antibiotic in treatment of
infections have resulted in the development of resistant strains which have
rendered antibiotic treatment ineffective. Currently, multiple antibiotic
resistance have been reported in a wide range of human infections. Most records
of morbidity and mortality are due to the fact that most of the bacteria
pathogens have developed resistance to antibiotics.
Azadirachta
indica has been used for decades as an antibacterial agent but it lack
specific scientific data to prove its efficacy.
1.4
JUSTIFICATION OF THE STUDY.
Since there is a continual need for new
antimicrobial agents to combat drug resistance, the quest for newer and potent
antimicrobial agents has ventured into studies on plants since plants are known
to produce varied chemicals for defence purposes against microorganisms, also
they produce cytotoxic compounds some of which have been successfully utilized
as chemotherapeutic agents and hence give a laudable reason to continue with
search for new treatment protocols for man. Thus the extracts from the leaves
of Azadirachta indica, if proven
effective will be a better option in combating antimicrobial resistance on some
bacteria-causing infections. Moreso, this research work will enable us
understand the efficacy of medicinal plants and its beneficial properties used
for the management of various ailments.
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