ABSTRACT
Phytochemistry of Acalypha wilkisiana maginata leaf was studied as well as an evaluation of antimicrobial activities of the leaf extracts in different solvents, against Methycilin Resistant Staphylococcus aureus (MRSA) from clinical samples. Extracts were obtained in water, ethanol and ethyl acetate and their yield, phytochemical content and antimicrobial activity were determined. Results showed yields of 11.62, 16.43 and 8.97% in water, ethanol and ethyl acetate respectively. Phytochemical composition showed Tannin (0.05% to 0.96), alkaloid (0.97 to 1.97), phenol (0.07 to 0.33), saponin (0.15 to 0.55) and flavonoid (0.15 to 0.27) percent. Hydrogen cyanide (HCN, 0.00 – 2.48mg/kg) was highest in the water extract. While the other phytochemicals had varied highest concentrations in the different solvents. Variations were also recorded in the antimicrobial activity of extracts against the staphylococcus aureus isolates. Water extract recorded the highest mean diameter of inhibition of 15.17mm (19.0mm to 20.3mm) while ethyl extract had the least inhibition of 13.1mm (9.0 to 16.7mm). All the extracts had lower inhibition relative to the standard drug (vancomycin) which recorded 18.28mm. Average resistance of 15% and 10% were exhibited by the isolates against the water extract whereas no resistance was recorded with ethanol extract and the standard drug (Vancomycin). Also 75% susceptibility was recorded for ethanol extract as against 65% each for water and ethyl acetate extracts. The minimum inhibition concentrations (MIC) was 100mg/ml for ethyl acetate extract 50mg/ml for water and ethanol extracts each. Again, the minimum bactericidal concentrations (MBC) of the extracts 250mg/ml, 300mg/ml and 350mg/ml for ethanol, water and ethyl acetate extracts respectively.
TABLE
OF CONTENTS
Title Page i
Certification ii
Dedication iii
Acknowledgments iv
Table of Contents v
List of Figures viii
List of Tables ix
Abstract x
CHAPTER ONE
INTRODUCTION 1
Aims
and Objectives 5
Objectives 5
CHAPTER TWO
LITERATURE REVIEW
2.1 The Plant Acalypha Wilkisiana 6
2.1.1
Phytocomponents of Acalypha wilkisiana 6
2.1.2 Phytochemical Studies and Isolated
Constituents from Acalypha wilkesiana 8
2.1.3
Botanical Features 8
2.1.4
Pharmacological Properties 8
2.1.5
Therapeutic Potentials of Acalypha wilkisiana 10
2.1.6 Research Commenced on Acalypha wilkesiana 11
2.2
Toxicity of Acalypha wilkesiana 13
2.2.1
Medicinal value of Acalypha wilkisiana 14
2.2.2
Traditional use of Acalypha
wilkesiana 16
2.3 Phytochemistry 18
2.3.1 Useful Phytochemicals 18
2.3.2 Alkaloids 18
2.3.3 Flavonoids 19
2.3.4 Phenol 19
2.3.5 Tannins 20
2.3.6 Other Phytochemicals 20
2.4 Plants and Antimicrobial Activity 20
2.5 Antibacterial
Agents from Flowering Plants 22
2.5.1
Infectious Diseases 22
2.5.2
Infections Due to Staphylococcus
aureus 23
2.5.2.1 Staphylococcal Food Poisoning 25
2.5.2.2 Skin and Soft-tissue Infections 25
2.6 Methicillin Resistant Staphylococcus aureus 27
2.6.1 Risk Factors of MRSA 27
CHAPTER THREE
3.0 MATERIALS AND
METHODS
3.1 Source of Materials 28
3.2 Sample and Medium Preparations 28
3.2.1 Media Preparation 28
3.2.2 Plant Preparation 28
3.2.3 Extraction of Plant Extract 29
3.3.0 Methods of Analysis in Qualitative Phytochemical Screening 29
3.3.1 Test for Alkaloids 29
3.3.2 Test for Flavonoids 29
3.3.3 Test for Saponin 30
3.3.4 Test for Tannin 30
3.3.5 Test
for Phenols 30
3.3.6 Confirmation of MRSA 31
3.3.6.1 Colony 31
3.3.6.2 Microscopic Examination 31
3.3.6.3
Biochemical Tests 31
3.3.6.4 Sugar
Utilization Tests 31
3.4.0 Reconstitution of Extract 32
3.4.1 Antibiotic Sensitivity Test 32
3.4.2 Determination of MIC 33
3.4.3 Determination of MBC 33
CHAPTER FOUR
4.0 RESULTS 34
CHAPTER FIVE
DISCUSSION AND CONCLUSION
5.1
Discussion 43
5.2
Conclusion 46
References 47
LIST OF FIGURES
Fig. 2.1: Morphology of A. wilkesiana a)
Leaf morphology b) Details of the leaf morphology
showing the indumentum c) Pistillate bract morphology (Sagun
et al., 2010). 9
Fig. 2.2:
Acalypha wilkesiana var. macafeana
hort. Shrub 12
LIST OF TABLES
Table 4.1: Extract
yield of Acalypha wilkisiana maginata in different solvents 37
Table 4.2 Phytochemical
composition of Acalypha wilkisiana maginata extracts (%) 38
Table
4.3: Antimicrobial Activity of Acalypha wilkisiana maginata leaf extract
against
MRSA from clinical sources 39
Table
4.4: Antibacterial
Activity of Acalypha wilkisiana maginata extracts
against
Staphylococcus aureus 40
Table 4.5: MIC
of Acalypha wilkisiana Leaf Extracts
against Staphylococcus Aureus 41
Table
4.6: MBC of Acalypha wilkisiana maginata Extracts against Staphylococcus
Aureus
42
CHAPTER ONE
INTRODUCTION
The work of plants for control, management and cure of diseases is
as old as humanity on earth. Presently, a greater population of the world's
people in the developing countries, rely on herbal medication for their
healthcare needs (WHO, 2001). Plants have been a source
of medicine in the past centuries and today scientists and the general public
recognize their value as a source of new or complimentary medicinal products
(Premanath and Lakshmideri, 2010). This plant-based, traditional medicine
system continues to play an essential role in health care, with about 80% of
the world’s inhabitants relying mainly on traditional medicines for their
primary health care (Owolabi et al.,
2005). Long before mankind discovered the existence of microbes, the idea that
certain plants had healing potential and that they contained what we would
currently characterize as antimicrobial principles, was well accepted. Since
antiquity, man has used plants to treat common infectious diseases and some of these
traditional medicines are still included as part of the habitual treatment of
various maladies. For example, the use of bear-berry (Arctostaphylos uva-ursi) and cranberry juice (Vaccinium macrocarpon) to treat urinary tract infections is
reported in different manuals of phytotherapy, while species such as lemon balm
(Melissa officinalis), garlic (Allium sativum) and tea tree (Melaleuca alternifolia) are described as
broad-spectrum antimicrobial agents (Rios and Recio, 2005).
During
the last two decades, there has been a considerable increase in the study and
use of medicinal plants all over the world especially in advanced countries.
Medicinal plants have been used in Africa before the introduction of
antibiotics and other modern drugs (Kabir et
al., 2009). According to World Health Organization, medicinal plants would
be the best source to obtain a variety of drugs (Nascimento et al., 2000). Therefore, such plants
should be investigated to better understand their properties, safety and
efficacy. The success story of chemotherapy lies in the continuous search for
new drugs to counter the challenge posed by resistant strains of
microorganisms. The investigation of certain indigenous plants for their
antimicrobial properties may yield useful results. Many studies indicate that
some plants have substances such as peptides, unsaturated long chain aldehydes,
alkaloids, essential oils, phenolics, as well as different ethanol, chloroform,
methanol and butanol soluble compounds. These plants have emerged as plants
with compounds possessing significant therapeutic potential against human
pathogens, including bacteria, fungi or virus (El astal et al., 2005).
Nigeria
has a great variety of natural vegetation, which is used in trado-medicine to
cure various ailments. Among the plants used for medicinal purpose in Africa,
particularly in Nigeria is Acalypha wilkesiana. The genus “Acalypha”
comprises about 570 species. Acalypha
wilkesiana Mull. Arg. belongs to
the family Euphorbiaceae and grows as an annual bedding plant. This fast
growing, evergreen shrub provides a continuous splash of color in the landscape
with the bronze red to muted red, 4 to 8 inch long, ear-shaped leaves available
in varying mottled combinations of green, purple, yellow, orange, pink or white,
depending upon cultivar (Egwaikhide and Gimba, 2007). Investigation is ongoing
on almost all the available cultivars within Nigeria with respect to their
phytochemicals and antimicrobial action against medically inclined and
agriculturally related pathogens (Ogbo and Oyibo, 2008). Consequently, this
plant has been reported to have antibacterial and antifungal properties as the
expressed juice or boiled decoction is locally used within Nigeria and some
other parts of West Africa for the treatment of malaria, dermatological and
gastrointestinal infections. Seeds from Acalypha
wilkesiana are essential components of a complex plant mixture used by
traditional healers in southwest Nigeria in the treatment of breast tumors and
inflammation (Udobang et al., 2010).
Several research work have
reported the use of plants on the maintenance of good health and in recent
times, most pharmaceuticals used in the treatment of various ailments have
their basis in plant based products (Kamba and Hassan, 2010). It is also known
that researchers have discerned that many indigenous plants in different
ethnicities contain bioactive compounds that exhibit physiology activities
against microorganisms and as such are used as precautions for the synthesis of
useful drugs (Osuagwu and Ihenwosu, 2014).
According to (Bishnu et al.,
2009), the usefulness of plant products in medicine is a function of
bioactive substances which they contain. Plant phytochemicals such as
alkaloids, tannins, phenolic compounds, flavonoids, steroids and many other
secondary metabolites are capable of producing definite physiological action in
the body.
The important medicinal roles
of phytochemicals in the body is well documented. Alkaloids are toxic and can
have powerful effects on body physiology (Harbone, 1973) and this perhaps
explains fear wide application in medicines. Ezeocha et al (2013)
observed that alkaloids are currently being recognized more on the basis of
their pharmacological properties rather than toxicity. They are also reported
to play vital metabolic roles and control development in living system. (Edeoga
et al., 2001) and also are after extensively as starting materials in
the manufacture of steroidal drugs.
Flavonoids are plant
antioxidants and are known to have diverse health benefits which includes the
inhibition of initiation, promotion, production and progression of tumors
(Okwu, 2004), modification and hormones in humans (Okwu and Omodamiro, 2005) an
reduction of incidences of cardiovascular diseases cancers, hyper lipidemia and
many other chronic diseases (de Pascal et al, 2008), Similarly, phenolic
compounds are associated with many health benefits such as toxicity to
pathogens, protect antioxidant activity, and a wide range of pharmacological
activities (Rein et al, 2000). Also, the health benefit of other
phytochemicals have been reported such as the use of saponin as emulsifying
agent and expectorant as well as their antifugal activities. Tannins have also
been associated with many human physiological attitudes like stimulation of
phagocytic cells and host mediated tumor activity. The roles of steroids in the
hormones are well known (Okwu, 2001). Their association with reduction of
canary heart and neurodegenerative disease is also documented (Perella et
al., 2003).
It is therefore established
fact that the efficacy of plant and their products in the management and cure
for diseases as witnessed in disease ethno medical formulas, stems from their
ride phytoclinicals which are responsible for potencies. Many plants have been
found to the useful in medications in addition to their other inhibit values
such as aesthetic uses as ornamentals erosion control etc. One such plant that
has found use in both ornamental and therapeutics is Acalypha.
It is a member of the plant family Euphorbioacaea and has many common names
depending on the species and genera including copper leaf fire dragon, Joseph's
coat etc (Markoshi et al., 2016).
While the plant is reported to have originated in the pacific islands, it grows
very well in most tropical and subtropical regions of the world.
Acalypha wilkisiana maginanta, is one of the species of the planet that has been put to many
uses in Nigeria and elsewhere. Akinyemi et al (2005) reported it is used
in medications in southern Nigeria for ailments such as fungal skin infectious,
gastrointestinal disorders, hypertension and management of diabetes. Research
reports (Golep et al, 2010) show cased the antinutrient activity of the
plant extracts in ethanol against a wide range of pathogens including, staphylococcus, Salmonella typhi,
Escherichia cocci, Klebsiella aerogenes, Pseudomonas Aerugines and Yersina enterocoletica. Research
(Ikewuchi and kewuchi, 2010) conformed the efficacy of the plant extracts on
lowering blood sugar and cholesterol. There has also been reports of the
structure antioxidant activities of the plant extract (Anokwuru et al., 2015), its bateriocidal
activities (Din et al., 2013) and inhibition of methicillin resistant staphylococcus aureus (Santiago et al.,,
2015).
Notwithstanding, some
medicinal plants have been reported to possess, in addition, some levels of
toxicity on consumers and there is paucity of data on toxicity studies of most
of their plants (Makoshi et al, 2016). The authors has established a
dose dependent increase in their function enzymes which suggested possible
hypertoxicity of Acalypha extracts at
low concentrations. While the plant and other plants have established medicinal
values, (Forcados et al., 2016)
called for exercise of caution in their use as therapeutics as high doses may
pose dangers of deleterious side effects.
This project therefore seeks to studying the phytochemistry
evaluation of Acalypha wilkisiana maginata and the antimicrobial activity of the plant extracts in
different solvents against MRSA from clinical sources.
Aims and Objectives
The aim of the project is to
study the phytochemicals present in Acalypha
wilkesiana and the antibiotics
activity of the extracts against MRSA from clinical samples.
Objectives
The objective of this study
includes the following:
1. Phytochemical
screening of Acalypha wilkesiana.
2. Invitro
antimicrobial activity of extract of Acalypha
wilkesiana (in three different solvent) against the MRSA form clinical
samples.
3. Determination
of MIC and MBC.
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