ABSTRACT
Piliostigmathonningii is one of the African plants with numerous medicinal values in ethnomedicine. It belongs to the family of Fabaceae and can be used for treating ailments like loose stooling in teething children, wound dressing, ulcer treatment, worms’ infestation, hemorrhoids, inflammations, bacterial infections, gonorrhea, stomach ache and headache. This study probed the chemical constituents and antimicrobial properties of the leaf and seed of P. thonningii. Qualitative phytochemical screening of the leaf and seed extracts of the plant showed the presence of alkaloids, flavonoids, tannins, saponins, terpenes, steroids, glycosides, and phenols.The quantitative aspect of the leaf showed that the alkaloid content was 15.5 %, tannins 10.2 %, saponins 9.6 %, flavonoids 4.3 %, terpenes 15.2 %, steroids 16.9 % and Phenols 12.9 %. The seed contained alkaloid 6.2%, tannins 4.2 %, saponins 4.8 %, flavonoids 2.5 %, terpenes 15.9 %, steroids 13.3 % and phenols 7.9 %. The vitamins detected in the leaf included Beta-carotene (10.428±0.000 mg/100g), ascorbic acid (13.432±0.020 mg/100g), riboflavin (3.412±0.035 mg/100g), niacin (0.941±0.000 mg/100g), thiamin (2.424±0.002 mg/100g) while that of the seed included Beta-carotene (5.349±0.013 mg/100g), ascorbic acid (8.4±0.040 mg/100g), riboflavin (0.640±0.000 mg/100g), niacin (0.0131±0.000 mg/100g) and thiamin (0.431±0.0031 mg/100g). The mineral compositions detected in the leaf included calcium (2413.02±0.001 mg/100g), magnesium (908±0.003 mg/100g), potassium (1103.04±0.023 mg/100g), sodium (53.01±0.018 mg/100g), phosphorus (46.24±0.000 mg/100g), nitrogen (1842.03±0.002 mg/100g), iron (16.31±0.000 mg/100g), zinc (0.243±0.001 mg/100g) and copper (0.001±0.000 mg/100g), while that of the seed included calcium (2010.34±0.006 mg/100g), magnesium (728.84±0.023 mg/100g), potassium (1004.02±0.011 mg/100g), sodium (38.03±0.061 mg/100g), phosphorus (20.02±0.001 mg/100g), nitrogen (785.01±0.003 mg/100g), iron (2.02±0.001 mg/100g), zinc (0.01±0.000 mg/100g) and copper (0.01±0.000 mg/100g). The GC-MS results of the chloroform extract of the leaf at different retention times showed a promising result of 17 probable structures with 14 of them, having medicinal values. In the same vein, the seed showed 9 probable structures with 7 of them having medicinal values. The antimicrobial activities of the chloroform extract of the leaf and seed were studied using hospital isolates of five pathogenic microorganisms. Its activity at a varying concentrations of 150 mg/ml, 75 mg/ml, 50mg/ml, 25 mg/ml, 12.5 mg/ml and 6.25 mg/ml was investigated againstS.aereus, B. cereus, E. coli, S. typhiand P.aeruginosa using agar well diffusion method. The result showed that the extract of the leaf exhibited activity against three micro-organisms in a concentration-dependent pattern, with the highest growth inhibition seen in S. aereus and the lowest growth in S. typhi with MIC of 25 mg/ml and 75 mg/ml respectively while the seed showed inhibition activity against two microorganisms in a concentration-dependent manner, with the highest growth inhibition seen in S. aereus and the lowest growth in E. coli with MIC of 50 mg/ml and 75 mg/ml respectively. The result of this research work revealed that P. thonningii has a lot of phytochemicals which might be responsible for its therapeutic potentials in ethnomedicine.
TABLE OF CONTENTS
Title Page i
Declaration ii
Certification iii
Dedication iv
Acknowledgement v
Table of Content vi
List of Tables ix
List of Plates x
Abstract xi
CHAPTER 1
INTRODUCTION
1.1 Background of the Study 1
1.2 Statement of the Problem 3
1.3 Scope of the Study 3
1.4 Justification of Research 4
1.5 Aim and Objectives 4
CHAPTER 2
LITERATURE REVIEW
2.1 Origin and geographical distribution of piliostigmathonningii 5
2.2 Botanical Classification 5
2.3 Description of the Plant 6
2.4 Economic and cultural values of P. Thonningii 10
2.5 The Family ‘Fabaceae’ 10
2.6 Phytochemicals 11
2.6.1. Phenols
11
2.6.2 Saponins 12
2.6.3. Steroids 13
2.6.4 Tannins 14
2.6.5 Flavonoids 15
2.6.6
Some of the compounds
isolated from p.thonningi are as follows. 16
2.8 Ethnomedical uses of PiliostigmaThonningii 19
2.9 Pharmacological activities Of Piliostigmathonningii 21
2.9.1 Anti-malaria activity of P.thonningii 21
2.9.2 Anti-Ulcer potential of piliostigmathonningii 21
2.9.3 Piliostigmathonningii as a Remedy to
Cardiovascular Complications and Associated Conditions 22
2.9.4 Anti-diabetic Property of
Piliostigmathonningii Leaf Extract
of Piliostigmathonningii 22
2.10 Mass
spectrometry 23
2.11 Gas
chromatography 23
2.12 Gas
chromatography-mass spectrometry 24
CHAPTER
3
MATERIALS AND
METHODS
3.1 Sample Collection and Identification 26
3.2 Sample Preparation 26
3.3 Sample Extraction 26
3.4 Qualitative Phytochemical Analysis 27
3.4.1 Test for alkaloids (Mayer’s test) 28
3.4.2 Test for tannins 28
3.4.3 Test for saponins 28
3.4.4 Test
for flavonoids 28
3.4.5 Tests
for steroids and terpenoids (Liebermann-Burchard test) 29
3.4.6 Test
for glycosides (Kella-Killian's test) 29
3.5 Quantitative phytochemical analysis 29
3.5.1
Determination of Alkaloids 29
3.4.2. Determination of flavonoids 30
3.4.3 Determination of Phenols 21
3.4.4 Determination of saponins 32
3.4.4 Determination
of tannins 32
3.4.6 Determination of Terpenes 33
3.4.7 Determination of Steroids 34
3.5 Determination of Vitamin content 35
3.5.1 Determination of ascorbic acid 35
3.5.2 Determination of
niacin 36
3.5.3 Determination of riboflavin 37
3.5.4 Determination of
thiamin 37
3.5.5 Determination of β-carotene 38
3.6 Determination of mineral content 38
3.6.1 Determination of
nitrogen content 38
3.7 GC/MS Analysis
3.8 Antimicrobial
screening of piliostigmathonningii leaf and seed chloroform
extract 40
3.8.1 Obtaining and confirming the test organism 40
3.8.2 Anti-bacterial susceptibility test 41
3.8.3 Determination of minimum inhibitory
concentration (MIC) 41
CHAPTER 4
RESULTS AND DISCUSSION
4.1 Phytochemical
screening results 44
4.2 Vitamin
content results 48
4.3 Mineral
content results 50
4.4 GC/MS results
4.5 Antimicrobial assay results 58
CHAPTER 5
CONCLUSION AND RECOMMENDATION
5.1 Conclusion 62
5.2 Recommendations
62
REFERENCES
APPENDIX
LIST OF TABLES
4.1 Qualitative phytochemical screening of
chloroform leaves extract of P.
thonningii 44
4.2 Qualitative phytochemical screening of
chloroform Seed extract of P. thonningii 45
4.3
Quantitative phytochemical screenings of chloroform leave extract of P.thonningii 45
4.4
Quantitative phytochemical screenings of chloroform seed extract of P.thonningii 46
4.5 Quantitative
determination of vitamin content of P.
thonningii leaf extract 47
4.6 Quantitative determination of vitamin
content of P. thonningii Seed extract 48
4.7 Quantitative determinations of mineral
content of the leaves and seeds of
P.thonningii 49
4.8 GCMS analysis Result on the chloroform Leaf
extract of P.thonningii 50
4.6 GCMS analysis
Result on the chloroform Seed Extract of P.thonningii 50
4.8
Zones of growth inhibition at varying concentrations of chloroform
extract of P. thonningiileaf against
the test organism. Zones of inhibition 51
4.9 Susceptibility test in mm on the standard
drugs on the microorganisms 52
4.10
Zones of growth inhibition at varying concentrations of chloroform
extract of P. thonningiiseed against
test organism zones of inhibitions. 53
4.11 Minimum inhibitory concentration of
chloroform leaf extract of P.thonningii 54
4.12 Minimum inhibitory
concentration of chloroform seed extract of P.thonningii 55
LIST OF PLATES
1: Whole plant of Piliostigmathonningii(https:en.wikipedia.org/wiki/P.thonningii) 7
2: Leaves of Piliostigmathonningii (https:en.wikipedia.org/wiki/
P. thonningii) 8
3: Fruit and seeds of Piliostigmathonningii
(https:en.wikipedia.org/wiki/P. thonningii) 9
CHAPTER 1
INTRODUCTION
1.1 BACKGROUND OF THE STUDY
The
use of plants as drugs can be seen as being as old as man himself. The
medicinal plants include various types of plants used in herbalism and some of
these plants have medicinal activities. Medicinal plants are considered as rich
sources of ingredients that can be used in drug development and synthesis.
Besides, these plants play crucial roles in the development of human and
cultural values around the world. Also, some of these plants are considered as
important sources of nutrients and as a result of that, are recommended for
their therapeutic values (Rasool, 2012).
Many
cultures throughout the world still rely on indigenous medicinal plants for
their primary health care needs (Franswort et al., 1985). To date,
25% of modern medicines are derived from plants that have been used by
traditional medicine practitioners. About 200 years ago, the first
pharmacological compound, “morphine “was produced from the poppy flower. Since
then, scientists have been studying plants to create the pharmaceutical
products that we have been using today (Cragg anred Newman, 2005).
Millions
of dollars have recently been invested in looking for promising medicinal
herbs. Even though there have been substantial investments in traditional
herbal medicine, it is still relatively modest when compared to the overall
pharmaceutical industries, but it proves that researches are beginning to steer
away from conventional drug development and is now looking inwards to
alternative and natural forms of treatments (Lai and Roy, 2004).
Natural
products have been used throughout human history for various purposes. Today,
plants are being used to treat several health concerns and these herbs are less
expensive and safer means of treatment than some conventional medications,
which is why so many people are choosing to go back to this traditional idea of
medications. A major hypothetical advantage of herbal medicines is the presence
of multiple active compounds that together can provide a potentiating effect
that may not be achievable by any single compound. This advantage presents a
unique challenge for the separation and identification of active constituents.
Most of the compounds that are identified by (activity guided) fractionation
must be tested in appropriate animal or other scientific models to confirm
their activities (Rousseaux and Schachter, 2003).
Herbal
medicine is now globally accepted as a valid alternative system of therapy in
the form of pharmaceuticals, food supplements, nutraceuticals, complementary and
alternative medicines. These medicinal herbs contain many active principles
that are used to treat diseases and relieve symptoms. Herbal medicine uses the
plants' seeds, berries, roots, leaves, barks or flowers for medicinal purposes.
The biological properties of these plants have beneficial effects. Other
factors are also responsible for their benefits as well, such as the type of
environment in which they are harvested and how they are processed. The plants
are either sold raw or as extracts, where they are macerated with water,
alcohol or other solvents to extract some of the chemicals. The resulting
products contain dozens of the chemical, like fatty acids, steroids, alkaloids,
flavonoids, glycosides, saponins, etc (Khanna et al., 1986).
Secondary
metabolites are organic compounds that are not directly involved in normal
growth, development or reproduction of an organism. Secondary metabolites often
play an important role in defense against herbivores (Stamp, 2003).
Human
uses secondary metabolites as medicines, flavoring and recreational drugs
(Prajapati et al., 2003). Nigeria is one of the countries in
the world that is blessed with a lot of medicinal plants in which Pilliostigmathonningii is
one of them.
1.2 STATEMENT OF THE
PROBLEM
Different
parts of Piliostigma thonningii have been used for medicinal
purposes for the treatment of ailments like dysentery, fever, respiratory
ailments, snake bites, hookworm and skin infections in Eastern parts of
Nigeria. The leaves and seeds have been reported in the literature to have
age-long folkloric use in traditional medicine especially in the treatment of
malaria, wounds, ulcer, gingivitis, fever, hemorrhoids, and backache.
This
plant could be used for the treatment of the above ailments because of the
bioactive compounds present in it. But from literature, little work has been
done to identify all the bioactive components available in the leaves and seeds
of the plant. Hence, the chemical studies on the leaves and seeds of the plant
as well as their antimicrobial potencies are herein reported.
1.3 SCOPE OF THE STUDY
Piliostigma
thonningii from literature has been proven to
have medicinal values. The leaves, seeds, root and the stem have phytochemical
components in them, but this work is limited to research on the leaves and the
seeds of P. thonningii.
1.4 JUSTIFICATION OF
RESEARCH
This
study will provide a database for the phytochemicals, vitamins, and minerals
present, as well as the antimicrobial activities of the leaves and seeds
of P. thonningii.
1.5 AIM AND OBJECTIVES
This study aims to identify the
bioactive components available in the leaves and seeds of Piliostigmathonningii.
This aim will be achieved through the following specific objectives.
i.
To isolate and
spectroscopically characterize the bioactive components available in the leaves
of P. thonningii..
ii.
To identify some of
the compounds present in the leaves and seeds of the plants using GC- MS.
- To quantitatively determine some of the
phytochemical constituents of the leaves and seeds of the plant.
- To determine some vitamins and minerals present
in the leaves and seeds of the plant.
- To establish the anti-microbial properties of the
leaves and seeds of the plant.
1.6
SIGNIFICANCE OF STUDY
Piliostigma
Thonningi, a commonly grown plant in Nigeria has
been known for its ethno-medicinal use by different cultural groups in Nigeria
and some other parts of Africa and Asia. This claim have been supported by
various researches done on the plant to extract and isolate some of its
bioactive constituents via various solvent media such as water, ethanol and
methanol. The bioactive properties of these constituents have been tested
against some pathogenic microorganisms such as Staphylococcus Aereus, Escherichia Coli, Salmonella Typhii, Vibrio Cholera,
and Candida Spp and so on.
This study investigates the chemical
composition and the antimicrobial studies of the chloroform extracts of the
leaves and seeds of P. Thonningi. The
isolation of a possible novel compound from the chloroform extracts of both
leaves and seed is expected and its bioactive properties studies.
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