ABSTRACT
Antioxidant potentials of the methanol leaf extract of Piper guineense (Uzuza) leaves was evaluated using standard analytical procedures. Properties such as phytochemicals, proximate composition, mineral composition and total antioxidant potentials were determined. The result of the phytochemical analysis showed the presence of alkaloid (20.67 ± 0.85 %), flavonoid (0.70 ± 0.02 %), phenol, (7.21 ± 0.21 %), tannins (0.72 ± 0.01 %), saponins (14.82 ± 0.36 %) and HCN (126.00 ± 10.58 mg/kg). The proximate composition assay revealed that the plant contains moisture content (11.40 ± 0.37 %), crude fat (1.90.00 ± 0.07 %), ash content (17.30 ± 0.46 %), crude fibre (21.64 ± 0.80 %), crude protein (13.62 ± 0.57 %) and carbohydrate (45.54 ± 1.00 %). The mineral elements present were copper (75.1 ± 0.44 mg/kg), iron (33.80 ± 0.53 mg/kg), magnesium (23.60 ± 0.60 mg/kg), manganese (74.00 ± 1.00 mg/kg) and zinc (124.00 ± 10.21 mg/kg). The total antioxidant potentials of Piper guineense leaf extract was 16.5mg equivalent. A graph change of change in absorbance against concentration of the standard (ascorbic acid) (mg/L) and the graph of change in absorbance against the concentration of the extract (mg/L) was plotted. The result showed that piper guineense leaf aside from having valid nutritional and therapeutic potentials, is also a good source of antioxidant that is capable of enhancing human nutraceutical needs.
TABLE OF CONTENTS
Cover
page i
Title
page ii
Certification
iii
Declaration iv
Dedication v
Acknowledgement vi
Table of Contents vii
List of Tables
viii
List of Figures ix
List of Plates x
Abstract xi
CHAPTER 1: INTRODUCTION
1.1 Background to the Study 1
1.2 Aims and Objective of the Study 4
1.3 Statement of Problem 5
1.4 Justification of the Study 5
1.5 Scope and Lamination of the Study 6
CHAPTER 2: LITERATURE REVIEW
2.1 Morphology and Description of Piper guineense 7
2.2 Phytochemicals lsolated from Piper guineense 8
2.3 Nutritional
value 12
2.4 Uses
of Piper guineense 13
2.5 Antioxidants 15
2.5.1 Natural
and synthetic antioxidants 17
2.6 Phytochemicals
22
2.6.1 Alkaloids
22
2.6.2 Saponin 23
2.6.3 Phenols
24
2.6.4 Tannins
25
2.6.5 Flavonoids
25
2.6.6 Hydrogen
cyanide (HCN) 26
2.6.7
Cardiac glycosides 27
2.7 Proximate
Composition 27
2.7.1 Moisture
content 28
2.7.2 Ash
residue 28
2.7.3 Total
carbohydrates 29
2.7.4 Crude
fat 29
2.7.5 Crude
protein 30
2.7.6 Crude
fibre 30
2.8 Mineral
Elements 31
2.8.1 Copper
(Cu) 32
2.8.2 Iron
(Fe) 32
2.8.3 Manganese
(Mn) 33
2.8.4 Zinc
(Zn) 33
2.8.5 Magnesium
(Mg) 33
CHAPTER 3: MATERIALS
AND METHOD
3.1 Materials
Used 34
3.2
Plant materials 34
3.3 Sample
Preparation 34
3.4
Extraction 34
3.5 Phytochemical
Analysis 35
3.5.1 Determination
of saponins 35
3.5.2 Determination
of tannins 35
3.5.3 Determination
of alkaloids 36
3.5.4 Determination
of phenol 37
3.5.5 Determination
of flavonoids 37
3.5.6 Determination
of cyanogenic glycoside (HCN) 38
3.6 Proximate
Composition 38
3.6.1 Determination
of ash residue 38
3.6.2 Moisture
of content determination 39
3.6.3 Fat
determination 39
3.6.4 Crude
fibre determination 40
3.6.5 Protein
determination 40
3.6.6 Determination
of carbohydrate 42
3.7 Determination of Minerals 42
3.7.1 Preparation of ferric reducing antioxidant
power (FRAP) reagent 42
3.7.2 Serial dilution of the stock solution of
ascorbic acid 43
3.7.3 Measurement
of total antioxidant potentials of the methanol exract
using FRAP method 44
3.8 Statistical
Analysis 44
CHAPTER 4: RESULTS
AND DISCUSSION
4.1 Phytochemical 46
4.2 Proximate
Composition 47
4.3 Mineral
Element 49
4.4 Result
of FRAP Assay 50
CHAPTER 5: CONCLUSION
AND RECOMMENDATION
5.1 Conclusion
54
5.2 Recommendation
54
Reference
LIST OF TABLES
2.1 Ethanomedicinal uses of Piper guineense 15
4.1 The results of qualitative and
quantitative phytochemical
analysis 45
4.2 The results of proximate analysis 47
4.3 The results of qualitative and
quantitative mineral element
Analysis 49
4.4
Results of FRAP Assay
50
4.5
Concentration of Standard and change in absorbance 51
4.6 Concentration of Methanol extract and
change in absorbance 52
LIST OF FIGURES
Page
2.1 Chemical structure of some Phytochemcials
Isolated
from Piper
guineense 12
2.2 Different types of Antioxidant compounds
(Rajani, 2004) 16
2.3 (a) Natural Antioxidant (Tocopherol), (b)
Synthetic
antioxidant (BHA) 18
2.4 Antioxidant reaction
19
2.5
DPPH Radicals Scavenging Capacity: Transformation of
DPPH form to non-radical form 20
2.6
Transformation of ABTS radical form to reduced
non-radicals form 21
2.7
FRAP antioxidant reaction with ferric salt 22
2.8 The structure of Alkaloid – Piperine 23
2.9 Structure of Saponin – Glycoside 24
2.10 Structure of phenol – flavone 24
2.11 Structure of Tannins Theaflavin 25
2.12 Structure of Flavonoids - Flavan 26
2.13 Structure of Cyanide 27
2.14 Structure of Cardia glycosides 27
4.1 Absorbance against concentration of the
Standard (ascorbic acid) 51
4.2 Absorbance against concentration of Standard
(Mg/L) and
that of the extract. (µg/L) 52
LIST OF PLATES
Page
2.1 Piper guineense showing (A) Ripe seeds
(B) Leaves
(C) Unripe seeds 8
CHAPTER 1
INTRODUCTION
1.1 BACKGROUND TO STUDY
An antioxidant is a chemical substance that inhibits the
oxidation of other chemicals. They preserve the vital key cell constituents in
the human body by counteracting the harmful effects of free radicals, which are
natural byproducts of cell metabolism (Miller et al., 2000). The oxidative stress (OS) as influenced by reactive
oxygen species (ROS) can be described as a dynamic imbalance between the
amounts of free radicals generated in the body and levels of antioxidants to
douse and/or scavenge them and safeguard the body against their harmful
consequence (Shirwaikar et al.,
2006).
The World Health Organization (WHO) estimated that 80% of the populations
of developing countries in Asia and Africa rely on traditional medicines,
mostly plant drugs, for their primary health care needs. (WHO, 2008).
Recently, there has been increase on the use of medicinal plants
in the developed countries and this has been linked to the distillation and
production of various medicines and chemotherapeutics from these plants as well
as from locally used herb remedies. The medicinal properties of these plants
could be hinged on the antioxidant, antimicrobial, antipyretic impact of the secondary
metabolites present in them (Soetan and
Iyelaagbe, 2009).
Rephrage, plant extracts and plant-derived drugs have
contributed immensely to the overall health and wellbeing of man (Anyanwu and
Nwosu, 2014).
In 2008, World Health Organization highlighted the significance of
scientific research into alternative medicine. Herbs are widely evaluated for
their attributes as food as well as medicine. Many under developed nations in
the world rely on herbs as a possible addition to World Health Organization collection
of “important drugs” once their benefit has been clinically proven.
Of the 265,000 species of
flowering plants that have been identified on planet earth, only 0.5% of them
have been studied in detail for chemical structure and therapeutic value (Omodamiro and Ekeleme, 2013).
Infact, modern scientists only know the chemical composition of
less than 5% of the flora in the rainforest (Jackson, 1989). At least 12,000
such compounds have been isolated and this number is estimated to be less than
10% of the total (Lai and Roy, 2014; Tapsel et
al., 2006).
Chemical composites in plants regulate their outcome in the
human body via procedures similar to the ones that are well known for the
chemical compositions in orthodox drugs. Therefore, herbal medicines are not
different from orthodox ones in terms of their curative effect. This makes herbal
medicines to be as efficient as orthodox medicines but also present them with
similar capacity to engender damaging consequence.
Research into uses of plants has been accepted as a result of efficient
means to determine future medicines. The use of these plants in medicine is due
to the availability of phytochemical compounds such as phenols, flavonoids,
tannins and alkaloids, present either in the seeds, leaves, stems and roots
(Tapsel et al., 2006).
Medicinal plants have been used since medieval times as origin of
medicine for the treatment of all kinds of ailments. Traditional medicine and
herbs remain functional in the primary healthcare systems of most cultures.
About 25% of the drugs prescribed worldwide and found in modern
pharmacopoeias come from plants, 121 such active compounds being in current use.
Of the 252 medicines regarded as fundamental and necessary by the World Health
Organization (WHO), 11% are sourced from plant and a significant percentage are
artificial medicines derived from natural sources. Examples of essential
medicines derived from plants include; digoxin from Digitalis spp. used
for managing congestive cardiac heart failure (CCF), quinine (antimalarial) and
quinidine (antiarrythmic) from Cinchona spp., vincristrine and
vinblastine (anticancer agents) from Catharanthus roseus, atropine from Atropa
belladonna and morphine and codeine from Papaver somniferum. It is calculated
that 60% of drugs use in treating tumour and infections are currently in the
market or undergoing clinical trial are from plants. (Rates, 2001).
This resurgence of interest in medicinal plants and traditional
medicine as a whole is owing to the fact that, conventional drugs could be ineffective
(e.g. side effects and ineffective therapy), wrong dosage of chemically
formulated medicines may cause harmful effects and related problems, a greater
part of the global population do not use synthetic drugs for treatment, perhaps
due to inaccessibility of healthcare centers and the ever increasing cost of
synthetic drugs. Finally, traditional medicine and environmental consciousness
imply that herbal derivatives are harmless. (Rates, 2001).
Medicinal plant extracts comprise of diverse bioactive substance
with diversities of biological activities of valued beneficial guides. The
protective effects of plant are attributed to the activities and potentials of
bioactive substances (Etim et al.,
2013), which are the non-nutrient compounds. They are classified into three
main groups namely alkaloids, isoprenoids (terpenoids and steroids) and phenols
(phenolic acid, flavonoids, tannins and coumarin). The plant derived
phytoconstituents are capable of terminating free radical reactions and hinder
the body or biological systems from oxidative damage and protect against
chronic diseases (neurodegenerative and cardiovascular (Etim et al., 2013).
Oxidative stress is defined as a disruption in the balance
between the making of reactive oxygen species (ROS), free radicals and
antioxidant defenses (Eboh, 2014). This imbalance leads to damage of important
molecules such as proteins, lipids, nucleic acids and is also involved in the
pathogenesis of various forms of diseases including cardiovascular diseases;
cancer etc. Moreover, antioxidants are different group of chemicals that
inhibit the oxidation of other molecules, neutralizing free radicals and
protect specific organs against deleterious effect of dangerous xenobiotics
(Durackova, 2010; Shirwaikar et al.,
2006).
The deleterious impacts of reactive
oxygen species (ROS) are eradicated by antioxidants and are of two kinds, namely,
enzymatic and non-enzymatic (Eboh, 2014).
Halliwell and Gutteridge, (1989); Huang et al., (2005), defined antioxidant in food science as a constituent
in foods when present at low concentration compared to those of oxidizable
substrate greatly reduces or inhibits negative impacts of reactive species,
such as reactive oxygen and nitrogen species (ROS/RNS), on normal biological
roles in humans. Ndiamaka, (2019), reported that, “antioxidants are substance that
are able to prevent the oxidation of easily oxidization materials.”
Natural and artificial antioxidants are extensively applied in conventional
medicine. Some of them have been proven to be effective protectors. That is,
they prolong the life span of laboratory animals when applied to food or
drinking water on a regular basis. In vitro, antioxidants restrain free
radical chain oxidation reactions, culminating in oxidation of fatty acids,
edible fats etc. More so, their effectiveness as scavengers of oxygen free
radicals in cells and tissues is insignificant in relationship to natural
antioxidant enzymes (Koltover, 2010).
1.2
AIMS AND
OBJECTIVE OF THE STUDY
This Project work aims to extract and
isolate antioxidant principles from methanol leaf extract of Piper guineense (Uziza) using the
following specific objectives:
i)
To obtain the methanol leaf extract by cold maceration.
ii)
To subject the methanol leaves extract to phytochemical screening (qualitative
and quantitative), proximate and mineral element composition.
iii)
To determine the total antioxidant potentials in Piper guineense leaves extract.
iv)
To draw conclusion and offer suggestions on the possible therapeutic
utilization of the Piper guineenses leaves
based on the result obtained.
1.3 STATEMENT OF PROBLEM
While a lot has been reported about nutritional and health
values of Piper guineense (Uziza)
leaves, there is very little scientific evidence for verifying the efficacy of
the antioxidant properties, possible health and nutritional values of Piper guineense leaves. Lack of information on the economic
values of Piper guineense leaves resulted in the under cultivation and
utilization of its products. Hence, this research aimed at evaluating the
phytochemical, micro nutritional and antioxidant characteristics of Piper guineense leaf.
1.4 JUSTIFICATION OF THE STUDY
The paucity of knowledge of the phytochemical constituents,
antioxidant and nutritional benefits of Piper
guineense (Uziza) leaves has resulted in its neglect and
under-utilization. It is envisioned that
the outcome of this research will motivate the extraction of the preservative,
nutraceutical and healing capacities of these cooking herbs. Nigeria is home to great biological diversity
including indigenous plants from which potential lead structures for use in
formulation of novel drugs can be obtained. Piper
guineense is used in traditional medicine to boost uterine compression for the
removal of placenta during child birth, regulation of weight and treatment of
pains. Anti-parasitic, antimicrobial and antifungal activities of Piper guineense leaf has also been
reported. Such knowledge will inform pharmaceutical industry on the therapeutic
importance of these plants, and will also validate indigenous knowledge as
valuable resource for medicine and pharmaceutical explorations.
1.5 SCOPE AND LIMITATIONS OF THE STUDY
The experiments will focus mainly on determining the phytochemical
constituents, antioxidant and nutritional benefits of Piper guineense (Uziza) leaves. The study covers nutritional
benefits and antioxidant benefits in order to establish the indigenous claim
that Uziza has some medicinal and pharmaceutical potentials.
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