CHAPTER ONE
1.0
Introduction
1.1
Background to the Study
Plants offer a large range of natural compounds belonging to different
molecular families which have various properties to humans (Hervé et al.,
2008). Phytochemicals are plant derived chemical compounds that can be
used as therapeutic agents. They reduce the risk of cancer due to dietary
fibers, polyphenol, antioxidants and anti-inflammatory effects. (Kinderley,
2006). The phytochemicals are produced via secondary metabolism in
relatively small amounts (Hasler, 1998).
In recent times quite a number of some plant parts i.e. leaves, stems and
roots have been used due to the presence of phytonutrients in them.
Scientifically, research is being undertaken to bring to limelight, the
therapeutic properties of the phytochemicals present in these plants and
also use them as a yardstick in modern medicinal plant uses (Riby et al.,
2006). Some groups of phytochemicals, which appear to have significant
health potentials, are carotenoids, flavonoids, phytoestrogens, nondigestible carbohydrate i.e. dietary fiber and prebiotics (Prior and Cao,
2000).
1
A typical example of this plant is a palm, coconut (Cocos nucifera linn). It
belongs to the family Arecaceae (Palmae). Palmea is a vast family
consisting of about 217 genera and about 2500 species. Cocos nucifera
belongs to the order arecales and it is the sole species of the genus cocos
belonging to the subfamily cocoideae, which includes 27 genera, and 600
species (Evans, 2002). One of the primary natural products from dry
coconut fruit is the coconut fiber which can be used for fuel and are a
source of charcoal. Activated carbon manufactured from coconut shell is
considered superior to those obtained from other sources, mainly because
of small macropores structure which renders it more effective for the
adsorption of gas and vapor and for the removal of color, oxidants,
impurities and odor of compounds (Grimwood et al., 1975).
A dried half coconut shell with husk can be used to buff floors. It is known
as a bunot in the Philippines and simply a "coconut brush" in Jamaica. The
fresh husk of a brown coconut may serve as a dish sponge or body sponge.
In Thailand, the coconut husk is used as a potting medium to produce
healthy forest tree saplings. The process of husk extraction from the coir
bypasses the retting process, using a custom-built coconut husk extractor
designed by ASEAN-Canada Forest Tree Seed Centre (ACFTSC) in 1986.
Fresh husks contain more tannin than old husks. Tannin produces negative
2
effects on sapling growth (Somyos, 1991). In parts of South India, the shell
and husk are burned for smoke to repel mosquitoes.
3
1.2
Statement of the Problem
This study was done to ascertain the phytochemicals in coconut fiber to
account for the use of its charcoal in ulcer management. This study seeks
to fill this knowledge gap by screening for the phytochemicals of coconut
fiber (Cocos nucifera linn) in order to diversify their use as important
ingredient in the life of man.
1.3
Significance of the Study
This research will provide detailed information for pharmaceuticals, foods
and other relevant areas. This is expected to help create job for the
unemployed Nigerians who would go into commercial cultivation of and
processing of coconut.
1.4
Aim and Objectives
The overall aim of this study is assessment of the extracts of coconut fiber
samples for selected active principles responsible for the use of its charcoal
in ulcer management.
In specific terms the objectives of the study were
i. To successively extract the active principles with n-hexane (a nonpolar solvent), ethyl acetate (a mild polar solvent) and methanol (a
strong polar solvent).
4
ii. Phytochemical analysis of the fiber extracts for some secondary
metabolites using simple chemical tests.
iii Document the finding for pharmaceuticals, foods and other relevant
areas.
5
CHAPTER TWO
2.0
Literature Review
2.1
Phytochemicals
Phytochemicals are chemical compound that naturally occur in plants
(phyto-plant in Greek). We are surrounded with hundreds of fruits,
vegetables, whole grains and spices rich in vitamins, minerals and other
essential substance called Phytochemicals. They are critical for
maintaining health and preventing disease they are found in plants and have
been used alone or in combination to prevent cancer, heart disease, diabetes
etc. Some phytochemicals with varying physiological properties may be
elements rather than complex molecules e.g. Selenium (abundant in many
fruits and vegetables) is a dietary mineral involved with major metabolic
pathways including thyroid hormone metabolism and immune function
(Brown and Arthur, 2001). Particularly it is an essential nutrient and
cofactor for the enzymatic synthesis of gluthathione, an endogenous antioxidant (Khanna et al., 2007).
American Cancer Society (ACS) rejects the claim that taking
phytochemicals supplement is good for long term health benefits. Rather
consuming phytochemicals in the fruits, vegetables, spices, beans for
which they are taken provide a better alternative (NIH, 2014).
6
2.2 Primary Metabolites
Proximate constituents of plant like Carbohydrate, Protein, Lipid (fat and
oil) are the primary metabolites which are responsible for growth, energy
and development.
2.2.1 Carbohydrate
Carbohydrates are produced through a process known as photosynthesis in
the presence of sunlight in the chlorophyll of plant by water and carbon
dioxide. Carbohydrates have the molecular formula C6H12O6, sometimes
called Saccharide (Sugar). Classified based on number of saccharides
molecules:
Monosaccharide,
Oligosaccharide,
Polysaccharide.
Carbohydrate serves various functions that include sweetening agent,
reducing agent, food for in fact, raw material (in wine, syrup, jellies,
alcohol etc.) among others (Khanna et al., 2007).
2.2.2 Protein
Proteins carry on most or all vital life processes in the human system, is the
most important compound that is found or made up off living organism,
Protein is main organic constituent of the body such as skin, hair, nails,
muscles etc. Thus, protein may be defined as condensation polymer of αamino acids and having peculiar overall structure which determine their
7
specific physiological functions in the living organism (Bahl and Bahl,
1980). Protein may be classified either by chemical composition or by
molecular shape.
 By chemical composition categories includes;
Simple protein and Conjugate protein
 By molecular shape they include;
Fibrous protein and Globular protein
Amino acid is a bifunctional compound containing both an amine and
carboxylic acid, they are the basic units that form protein molecule. Base
on essentiality Histidine, Isoleucine, Leucine, Lysine, Methionine,
Phenylalanine, Theonine, Tryptophan, and Valine are essential amino
acids and Alanine, Serine, Asparagine, Aspartic acid, Glutamate, Glutamic
acid, Glycine, Arginine, Proline, Tyrosine and Cystidine are non-essential
amino acid (Umar, 2005).
2.2.3 Lipids
Lipids are biologically active compounds that are insoluble in water
(Philip, 1997), but soluble in organic solvent like benzene, ether,
chloroform (McDonald et al., 1995). In plants lipid are of two types i.e.
Structural and Storage lipid.
8
 Structural lipid is present as constituent of various membranes and
protective surface layers and make up about 7% of leaves of higher
plants (Umar, 2005).
 Storage lipids occur in fruits and seed and are predominantly oil
(McDonald et al., 1995).
According to Umar, 2005 lipid can be classified based on backbone
structure i.e.
 Glycerol which further sub-classified into;
Simple and Compound or Complex
 Non-glycerol
Natural fats and oils are the tri-ester of glycerol with long chain carboxylic
acids (12-20 carbons), known as triglycerides or triacylglycerides. The
distinctions between fat and oil are based on their difference state at room
temperature. Natural occurring fatty acid are unbranched and have even
number of carbon atoms (Umar, 2005), it may be saturated and unsaturated
(sub-classified into Monoenoic are those with fewer double bond per
molecule and Polyenoic or Poly unsaturated fatty acid (PUFA) and exist in
Trans and Cis configuration) (McDonald et al., 1995). Some fatty acid are
essential (Polyenoic) e.g. linoleic acid, linolenic acid, arachidonic acid,
their primary sources are terrestrial and marine plants, phytoplanktons,
marine animals (McDonald et al., 1995).
9
2.2.4 Minerals
Minerals are large family of nutrient essential to human body although
some are present in the body but in small percentage. They are components
of hormones and certain factors with special physiological functions
(Tianshi, 1997).
These elements (Sodium, Potassium, Phosphate, Zinc, Iron, Copper,
Magnesium and Phosphorus) not only take part in various metabolic
processes but also play vital role in growth, development, immunity,
regulation, mitotic cell division, genetic expression in the body. They differ
from the proximate constituent because they cannot be synthesize by the
body i.e. they are ingested.
i.
Calcium
It’s one of the basic constituents in which 99% is present in the skeleton
(bones), teeth and 1% in the blood cell and soft tissues of human body. It
plays key role in neuromuscular reflexes, blood coagulation, cell adhesion,
heart rhythm regulation among others. The recommended daily allowance
(RDA) of calcium by US National Institute of Health (NIH) is 80mg for
average adult not lower than 150mg for pregnant women, nursing mothers
and elderly, and 1000mg for children (Tianshi, 1997). Modern medical
studies proved that more than 100 diseases are closely related to calcium
10
deficiency such as late growth of hair, osteoporosis, heart, digestive
system, arthritis, rickets (Adeyeye, 2002).
ii.
Sodium
Sodium is classified as major mineral nutrient and essential to human
health, it is necessary for water-acid balance, cell permeability, for glucose
absorption, nerves and muscles functioning. Its deficiency can cause
dehydration of body due to lowering of osmotic pressure (McDonald et al.,
1995). The RDA is 200-500mg for adult.
iii.
Potassium
Potassium is primarily an intracellular cation (Umar, 2005) and is very
essential in the formation of glycogen, protein synthesis, regulation of fluid
pressure balance, neuromuscular excitability and muscles contraction
(Hegarty, 1998). RDA is 200mg per day but only 8% is retain in the body
(NIH, 2014). Potassium deficiency causes weakness, paralysis, growth
retardation and rarely death (McDonald et al., 1995).
iv.
Phosphorus
The most abundant minerals in human body are phosphorus (700mg) and
1200mg of calcium in man, about 80-85% of phosphorus is found in
skeleton. Phosphorus is present in cell, blood as soluble phosphate ion, in
11
lipids, protein, carbohydrate, nucleic acid and nucleoproteins (responsible
for cell division, reproduction and transmission of heredity traits)
(Adeyeye, 2002).
v.
Magnesium
Activator of many enzymes in carbohydrates, protein and lipid metabolism
is magnesium (McDonald et al., 1995). Magnesium serve as intracellular
fluid, function in stabilizing some structures and energizing others in all
types of biopolymers e.g. DNA, RNA, Protein, Lipid (Garba, 1999).
Magnesium deficiency causes uncontrollable twisting of muscles leading
to convulsion, even death (Hegarty, 1988). RDA intake is 150mg per day
for children, 200-400mg per day for men and 300mg per day for women.
2.3 Secondary metabolites
They are phytochemicals that have significant pharmacological and
biological effect on living system and are often used for drugs and
pharmaceuticals. Examples are Alkaloid, Steroid, Tannin, Glycoside,
Cardiac glycoside, Saponin, Flavonoid etc.
2.3.1 Alkaloid
12
Alkaloid are the basic substance which contain one or more nitrogen atoms,
usually in combination as part of cyclic system (Harbone, 1973). They are
single class of secondary plants metabolites of which about 5,500 are
known. Alkaloid may be defined as plant derivative compound that is toxic
or physiologically active, contain nitrogen in heterocyclic ring, is basic and
limited distribution in plant kingdom (Bahl and Bahl, 1980).
N
CH3
N
Fig. 1: Nicotine structure
2.3.2 Saponin
Structurally, two types of Saponin are recognized i.e. Steroidal (common
tetracyclic triterpenoids C27) and Triterpenoid (pentacyclic triterpenoids
C30), both have glycosidal linkage at C3.
O
RO
13
Fig. 2: Tetracyclic triterpenoid
14
2.3.3 Steroids
Are lipid fractions compound characterized by cyclopents (α)
phenanthrene (fig. 4). Steroid are compounds based on combination of
three cyclohexane rings and one five members ring, they play some part in
biochemical systems (Balbao, 1976).
R
Fig. 3: Steroid general structure
Cholesterol, sex hormones (androgen-testosterone, estrogen-estroidiol and
progestin’s- progesterone), bile acid and Vit. D are commonest and
biologically crucial steroids.
CH3 CH3
CH3
O
Fig. 4: Testosterone
15
2.3.4 Cardiac glycoside
Glycosides are colourless, non-volatile, crystalline and bitter testing solid
compound. Chemically they are group of organic compounds which can be
resolved by hydrolysis into sugar component (glycone) and other organic
substances (aglycone).
O OH
OR
O
Fig. 5: Glycoside structure
2.3.5 Anthraquinone
Is a tricyclic structure heaving weak reducing properties which may
account for the use of anthranols and anthrones as antiseptic in certain skin
diseases. They are seldom found among Monocotyledon, only in Liliaceae
and in form of usual c-glycoside. Among Dicotyledon, they occur in
Rubiaceae, Leguminoceae, Polygonaceae, Ericaceae etc also in certain
fungi and linchens. Anthraquinone is applicable in vegetable drugs
(Balbao, 1976).
O
O
O16
OH
Fig. 6: Alizarine structure
17
2.3.6 Flavonoid
Largest group of natural occurring phenols and occur in plant both in free
state and as a glycoside. Flavonoid may be described as a series of C6-C3C6 compound.
Fig. 7: Flavonoid structural skeleton
2.4 Coconut Tree
The coconut (Cocos nucifera L. family Arecaceae) is a well distributed
fruit tree all around the world, providing food, especially in the tropical and
subtropical regions and for its many uses it is often called the “tree of life”.
Chan E and Elevitch C.R. (2006). There are 12 different crops of nuts under
the name of coconut palm (DebMandal and Mandal, 2011). C. nucifera is
widely distributed over the Brazilian northeastern coast, where is known
as “Coco-da Bahia”.
18
Popular medicinal uses (against arthritis and diarrhea) of coconut husk
fiber have been reported (Esquenazi et al., 2002; Alviano et al., 2004;
Rinaldi et al., 2009), but the knowledge of its potential benefit or adverse
effects in human beings is still very preliminary. Previous studies showed
that aqueous C. nucifera husk fiber extracts present important biological
activities
such
as antimicrobial, antiviral, antinociceptive, anti-
inflammatory, antioxidant and antineoplastic properties (Esquenazi et al.,
2002; Alviano et al., 2004; Rinaldi et al., 2009; Akinyele et al., 2011; Dua
et al., 2013). Coconut husk fiber is rich in polyphenolic compounds. The
C. nucifera husk fiber aqueous extracts are mainly composed by catechin,
epicatechin and condensed tannins (B-type procyanidins) (Esquenazi et al.,
2002). Plant phenols represent an important group of natural antioxidants
and some of them are potent antimicrobial compounds (Chakraborty and
Mitra, 2008). In general, polyphenols can prevent chronic diseases by their
antioxidant, free radical scavenger and metal chelator properties (Daglia,
2012).
In Nigeria, the coconut tree (Cocos nucifera L) is produced in about 22
states, it is called “Aki Oyibo” or “Aki Bekee” in Igbo, “Kwakwa” in
Hausa and “Agbon” in Yoruba. It is common in the south-east, south-south,
south west north east and north central geographical zones of the country
19
where its parts have many uses such as the leaves for roof thatch, garden
fencing etc. (Yakubu, 2006).
The industrial use of this plant generates large amounts of husk fiber as
industrial reject, featuring an environmental problem. Based on our interest
in searching for medicinal plants with antimicrobial activity and in
expanding the knowledge about the phytochemical profile of C. nucifera,
the purpose of this study was to investigate the photochemical activity of
coconut fiber.
2.5 Taxonomical Classification
Kingdom: Plantae
Division: Magnoliophyta
Class: Liliopsida
Subclass: Arecidae
Order: Arecales
Family: Arecaceae
Genus: Cocos
Species: C. nucifera
Binomial Name: Cocos nucifera Linnaeus
2.6 Botanical Description
20
Cocos nucifera (L.) is an important member of the family Arecaceae (palm
family) popularly known as coconut, coco, coco-da-bahia, or coconut-ofthe-beach (Aragão, 2002). The plant is originally from Southeast Asia
(Malaysia, Indonesia, and the Philippines) and the islands between the
Indian and Pacific Oceans. From that region, the fruit of the coconut palm
is believed to have been brought to India and then to East Africa. After the
discovery of the Cape of Good Hope, this plant was introduced into West
Africa and, from there, dispersed to the American continent and to other
tropical regions of the globe (Purseglove, 1972). The plant is an
arborescent monocotyledonous tree of around 25 m in height (giant
coconut) with a dense canopy. The root of the coconut system is
fasciculate. The stem is an unbranched type, and at its apex, a tuft of leaves
protects a single apical bud. The pinnate leaves are feather-shaped, having
a petiole, rachis and leaflets. Under favorable environmental conditions,
the giant adult coconut emits 12-14 inflorescence spikes per year, while the
adult dwarf coconut can emit 18 spikes in the same period. The axillary
inflorescence has globular clusters of female flowers. The plant is
monoecious (male and female reproductive organs on the same plant)
(Passos, 1998). The coconut fruit comprises an outer epicarp, a mesocarp,
and an inner endocarp. The epicarp, which is the outer skin of the fruit, and
the mesocarp, which is heavy, fibrous, and tanned when dry, have many
industrial uses. The endocarp is the hard dark core. Inside is a solid white
21
albumen of varied thickness, depending on the age of the fruit, and with an
oily pulp consistency and a liquid albumen called coconut water that is
thick, sweet, and slightly acidic (Passos EEM., 1998; Andrade AM et al.,
2004).
Fig. 8: Coconut Fibre
2.7 Constituent of Coconut Fibre
Coconut fiber, obtained from coconut, is a natural fiber extracted from the
husk of coconut. The coconut is steeped in hot seawater, and subsequently,
the fibers are removed from the shell by combing and crushing, the same
process as jute fiber. The individual fiber cells are narrow and hollow with
thick walls made of cellulose, and each cell is about 1 mm long and 10–
20 μm in diameter. The raw coconut fibers show length varying from 15 to
35 cm and diameter from 50 to 300 μm. When they are immature and then
22
become hardened and yellowed because a layer of lignin is deposited on
their walls. Coconut fiber shows a good stiffness and is used in products
such as floor mats, doormats, brushes, mattresses, coarse filling material,
and upholstery (Y. Yan, 2016).
CHAPTER THREE
3.0
Materials and Method
3.1
Materials
1. Coconut Fiber
2. 10% Sodium hydroxide
3. Ammonium hydroxide
4. 0.33M Ferric chloride
5. 90% Ethanol
6. 3M Sulphuric acid
7. Fehling solutions A & B
8. Wagner’s reagent
9. Dragendoff’s reagent
10. Mayer’s reagent
3.2
Apparatus
1. Weighing balance
23
2. Measuring cylinder
3. Beakers
4. Soxhlet extractor
5. Evaporating dish
6. Test tube/test tube rack
3.3
Method
3.3.1 Sample Collection and Preparation
The coconut fiber was collected from Agbani in Nkanu West Local
Government Area of Enugu State on 29th September, 2022. The samples
were separated from the outer cover and put in a plastic container pending
analysis.
3.4 Reagent Preparation
3.4.1 Sodium Hydroxide (10 %)
Ten grams of Sodium Hydroxide was dissolved in 90 ml of distilled water.
3.4.2 Ammonium Hydroxide
Ten milliliters of ammonia were added to 90 ml of distilled water.
3.4.3 Ferric Chloride (0.33 M)
24
Exactly 5.4 g Ferric Chloride was dissolved in distilled water and the
volume made up to 500 ml with distilled water.
3.4.4 Ethanol (90 %)
This was prepared by adding 5 ml of distilled water to 95 ml of 95 %
ethanol.
3.4.5 Sulphuric acid (3 M)
Fifty-four milliliters (54 ml) of concentrated Sulphuric acid were added to
half filled 1000 ml volumetric flask with distilled water and the volume
made up to 1000 ml mark with distilled water.
3.4.6 Fehling Solutions A & B
(i)
Fehling solution A was prepared by dissolving 69.2 g of Copper
Sulphate in distilled water.
(ii)
Fehling solution B was prepared by dissolving 35.2 g of
Potassium Sodium Tartrate in distilled water then 15.4 g Sodium
hydroxide was dissolved in the solution and the volume made up
to 1 liter.
3.4.7 Wagner’s Reagent
25
Potassium iodide (2 g) was dissolved in distilled water, then 1.3 g of iodine
crystal was added and shirred to dissolve. The volume was made up to 100
ml with distilled water.
3.4.8 Dragendoff’s Reagent
This was prepared by dissolving 3.2 g potassium iodide and 1.7 g bismuth
nitrate in 50 ml distilled water, then 20 g tartaric acid was added and
dissolved and the volume made up to 100 ml.
3.4.9 Mayer’s Reagent
Exactly 1.36 g of mercuric chloride was dissolved in 60 ml of distilled
water, then 5 g of potassium iodide was dissolved in 20 ml distilled water.
Both solutions were mixed and the volume made up to 100 ml.
3.5 Successive Extraction of Active Principles for Phytochemical
Analysis
The active principles in 25 g of the coconut fiber samples were
exhaustively extracted with 250 mL n-hexane in a 500mL capacity Soxhlet
extractor using heating mantle. The extract was concentrated to half the
volume and labeled n-hexane extract of coconut fiber. The same procedure
was repeated with 250 ml of ethyl acetate and methanol and labeled
extracts of coconut fiber respectively.
26
3.6 Screening for Active Principles in the Sample Extract
The active principles in the sample were determined using the n-hexane,
ethyl acetate and methanol extracts. Standard methods were followed to
determine the presence of saponins, glycosides, tannins, flavonoids,
steroids, anthracene, alkaloids, resins, amino acids, carbohydrate and
volatile oils etc. in the non-polar n-haxane, slight polar ethyl acetate and
strong polar methanol extracts.
3.6.1 Test for Tannins
a) A mixture of 4mL of each extract and 4mL of water was stirred very
well and three drops of 0.33 mol/dm3 ferric chloride solution was
added and the mixture observed for immediate green colouration and
result recorded.
b) One milliliter of the extract was treated with few mL of gelatin
solution; a white precipitate is formed revealing the presence of
tannins and phenolic compounds.
c) One milliliter of the extract was treated with few ml of lead acetate
solution. A precipitate production shows the presence of tannins and
phenolic compounds.
27
3.6.2 Test for Hydrolysable Tannins
Four milliliters of 10% ammonia solution was added to 4mL of each extract
and shaken very well and observed for the formation of an emulsion and
the result recorded.
3.6.3 Test for Pseudo Tannins
A match stick was dropped into 3mL of each extract and two drops of
concentrated hydrochloric acid (HCl) was added. The match stick was left
undistorted for 5 minutes and observed for a dark purple colouration on it
and the result recorded.
3.6.4 Test for Flavonoids
a) Magnesium Ribbon Test (Shinoda Test): A small quantity of
magnesium ribbon was dropped into 2mL of each extract and 5
drops of concentrated hydrochloric acid (HCl) added the formation
of reddish colouration is positive result and it was recorded.
b) Alkaline Test (NaOH and Acid Test): Addition of increasing
amount of NaOH to the alcoholic extracts shows colouration which
decolourises after addition of acid.
c) Lead Acetate Test: To small quantity of residue, 0.5mL of 1% Lead
acetate solution was added and observed for yellow colour ppt.
formation.
28
3.6.5 Test for Resins
Two milliliters of acetic anhydride were added to 2mL of each extract and
2 drops of concentrated surphuric acid added. It was observed for violet
colouration and the result recorded.
3.6.6 Test for Alkaloids
a. Dragendoff’s Test: Two drops of Dragendoff’s reagent was added
to 2mL of each extract and observed for dip brown precipitate and
the result recorded.
b. Wagner’s Test: Two drops of Wagner’s reagent was added to 2mL
of each extract and observed for a dip brown precipitation and the
observation recorded.
c. Mayer’s Test: Three drops of Mayer’s reagent was added to 2mL
of each extract and observed for a reddish precipitation or
colouration.
d. Kraint’s Test: Two drops of Kraint’s reagent was added to 2mL of
each extract and observed for white precipitate.
3.6.7 Volatile Oil Test
29
Six (6) drops of Ferric Chloride (0.33 mol/dm-3) solution was added to a
mixture of 2mL of each extract and 2mL of 90% (v/v) ethanol was added
the resulting mixture was observed for green colouration and the result
recorded.
3.6.8 Test for Amino Acids and Proteins
a) To 1 mL of extract, 2 drops of freshly prepared 0.2% Ninhydrin
reagent was added and heated. Development of purple color
indicates the presence of proteins.
b) The extract was treated with one mL of 40% Sodium Hydroxide
solution and two drops of 1% copper Sulphate reagent. Appearance
of violet color indicates the presence of proteins.
3.6.9 Test for Carbohydrates
a) Fehling’s Test: The extract was treated with 5 mL of Fehling’s
solution A precipitate indicates the presence of reducing sugar.
b) Benedboilin Test: To 1 mL of the extract, added 5 mL of Benedict’s
solution and kept at boiling water bath for 5 min. Red, yellow or
green precipitate indicates the presence of reducing sugars.
3.6.10 Test for Saponins
30
Two and half milliliter of each extract was vigorously shaken with 10 mL
of water for 2 minutes in a test tube. Then 2 mL of olive oil was added and
observed for persistent frothing and emulsion formation and result
recorded.
3.6.11 Test for Saponin Glycoside
Two and half milliliter of mixed Fehling’s solutions A and B was added to
2.5mL of each extract in a test tube and observed for development of bluish
green precipitate and observation recorded.
3.6.12 Test for Steroids and Triterpenoids (Libermann Burchaed)
Two and half milliliter of acetic anhydride was added to 2mL of each
extract in a test tube and cooled well in ice block. Three milliliters of
concentrated Sulphric acid was carefully added and a change from violet
to blue to green colour was observed and recorded.
3.6.13 Test for Glycosides (General)
Dilute Sulphuric acid (2.5mL) was added to 5ml of each extract in a test
tube and boiled for 15 minutes. Then 2mL of 10% NaOH and 5ml of mixed
Fehling’s solution A & B were added. The formation of brick red
precipitate is positive test.
3.6.14 Test for Digital Glycosides
31
A drop of ferric chloride was added to 2ml of each extract in a test tube.
Two milliliter of glacial acetic acid (glacial means no H 2O) and 2mL of
concentrated Sulphuric acid were added. The resulting solutions was
observed for the formation of blue layer and the result recorded.
3.6.15 Test for Anthracenes (Born Traggers Test)
Two milliliters of chloroform were added to 2mL of each extract and was
allowed to separate, to the chloroform layer, 2mL of 10% ammonium
solution was added and vigorously shaken and kept to separate, the
observation of brick red precipitate is a positive result and recorded.
CHAPTER FOUR
4.0
Results and Discussion
4.1
Results
This chapter deals with data presentation, analysis of the data and
discussion of findings. The results of this analysis are represented below in
32
tables presenting phytochemical screening of coconut fiber extracts gotten
from Agbani in Nkanu West Local Government area of Enugu State.
Table 4.1: Results of Phytochemical Screening of Coconut Fiber
Extracts
Coconut fiber extracts
Parameters
n-Hexane Ethyl acetate Methanol
Saponin
-
+
-
Saponin Glycoside
-
+
-
Tannins
+
-
-
Hydrolysable Tannins
+
+
-
Pseudo Tannin
-
-
-
Test of Tannin (Using Gelatin)
-
-
-
Digital Glycoside
-
-
-
Glycoside General
-
-
+
Anthracene (Born Tragger Test)
-
-
-
Resins
-
-
-
(Libemann -
-
-
Volatile Oil Test
-
-
-
Alkaloid
+
-
-
b. Wagner’s Test
+
-
-
c. Mayer’s Test
+
-
-
d. Kraint’s Test
+
-
-
Flavonoid Test
-
-
-
Steroid
and
Triterpenoids
Buchard) Test
a. Draggendoff Test
33
(a) Magnesium ribbon test
(b). Alkaline test
-
-
-
(c). Lead Acetate test
-
-
-
Carbohydrate Test
-
+
+
Amino acid (Protein)
-
-
-
Interestingly, coconut fiber is not rich in secondary metabolites but found
to contain alkaloids in n-hexane extract, testing positive with all the four
test reagents (Wagner’s, Dragendoff’s. Mayer’s and Krant’s reagents).
This probably may be that alkaloids are pronounced in coconut fiber.
The identification of carbohydrate in the n-hexane, ethyl acetate and
methanol extracts suggest that coconut fiber might be rich in carbohydrate.
4.2 Discussion
Screening for active principles (Table 1) in the n-hexane (non-polar
solvent), ethyl acetate (slightly polar solvent) and methanol (very polar
solvent) extracts of coconut fiber revealed the presence of alkaloids,
carbohydrates, tannins, saponin and glycoside. Alkaloids was found to be
present in n-hexane extract and detected by (Drangedoff’s, Wagner’s,
Mayer’s and Krant’s) the four test reagents. Alkaloids were not detected
in ethyl acetate and methanol extracts.
34
The result of active principles screening of coconut fiber extracts revealed
the presence of carbohydrate in the n-hexane, ethyl acetate and methanol
extracts using Fehling’s reagent.
The results of the screening for active principles of coconut fiber extracts
(Table 1) showed tannins to be present in only n-hexane extract while
hydrolysable tannins were present in n-hexane and ethyl acetate extracts.
Saponin and saponin glycoside were found to be present in ethyl acetate
extract only while glycoside was present in only methanol extract.
Interestingly, flavonoids, resins, amino acids, and anthracene were not
present in coconut fiber.
CHAPTER FIVE
5.0 Conclusion
35
The screening of n-hexane, ethyl acetate and methanol extracts for
phytochemicals revealed that coconut fiber is not rich in phytochemicals.
Meanwhile n-hexane extract were found to be richer in phytochemicals
with tannins, alkaloids and carbohydrates present, while ethyl acetate
contain
saponin,
saponin
glycoside,
hydrolysable
tannins
and
carbohydrate. Methanol with only glycoside, and carbohydrate. The study
recommends that the alkaloid be isolated and studied to find its use.
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