analysis of essential oil from watermelon seedssospoly journal of science & agriculture, vol. 2,...
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SosPoly Journal of Science & Agriculture, Vol. 2, (Dec., 2017) ISSN: 2536-7161
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Umaru Ali Shinkafi Polytechnic Sokoto, Nigeria
ANALYSIS OF ESSENTIAL OIL FROM WATERMELON SEEDS
Mansur Ahmad, Riskuwa Faruk, Kasimu Abubakar Shagari and Shehu Umar
Shehu Shagari College Of Education, Sokoto-Nigeria
ABSTRACT
Watermelon belongs to the family of Cucurbitaceae and it well known in Nigerian modern
and traditional system for its modern and traditional uses. The present investigation was
carried out to determine the chemical properties, proximate composition as well as the
possible bioactive components of watermelon seeds using GCMS analysis. The oil from the
seeds was extracted using soxhlet extraction procedure. Moisture was determined directly on
the seeds by oven drying at 105oC for 6 hours. The yield of the dry seeds from the sample was
determined. The ripened seeds and dry seeds were then ground in some blender, separately,
and placed in a vacuum oven at 60oC for 6 hours and finally stored in a desiccator until
analyzed. Proximate analyses were performed in triplicate in accordance with the AOAC
procedures (AOAC, 1990). The ash was determined by heating overnight at 500oC and the
protein content of the seeds by standard Kjeldahl (total %N) procedure. Wiss and Devine
(1961) method was adopted for the chemical analysis. Free Fatty Acid of the oil
(2.46±0.01%) which shows the better quality of the oil. The acid value (7.60mg/NaOH/g) fall
within acceptable limits for edible oils (≤10mg/NaOH/g). The GC-MS analysis was carried
out on a GC-MS-QP 2010 Shimadzu system using NIST database. Seven (7) components from
watermelon seeds were identified. The prevailing elements in the watermelon seeds were
palmitic acid, carbonic acid, oleic acid, Dioctylester, propanedioic acid, linoleic acid
chloride and delta-tocopherol. The presence of various bioactive compounds confirms the
applications of watermelon for multiple ailments by traditional and modern practitioners.
However, isolation of individual phytochemical constitutes may proceed to find a novel drug
or lead compound.
Keywords: Watermelon, Seeds, GC-Ms, Proximate Analysis, Essential Oil
INTRODUCTION
The use and dependence on plants as medicines by man has been in existence since time
immemorial, and man continues to search for plants as drugs for a particular disease within
his reach. Synthetic medicines are safe than herbal medicines because the phytochemical in
the plant extract target the biochemical pathway (Zaidan and Badrul, 2005). Medicinal plants
are an expensive gift from nature to human. The approval of traditional medicines as an
alternative form of healthcare and the improvement of microbial resistance to the existing
antibiotics has lead researchers to scrutinize the antimicrobial compounds (Parvathi, S.,
2010). Medicinal plants have been used all over the world for the treatment and prevention of
various ailments especially in developing countries where infectious diseases are endemic
and modern health facilities, and services are inadequate (Zaida and Badrul, 2005). The
medicinal actions of plants unique to particular plant species or groups are consistent with the
concept that the combination of secondary products for a specific plant taxonomically distinct
(Wink et at., 1999).
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Screening active compounds from the plant have led to the invention of new
medicinal drugs which have protection and treatment roles against various diseases including
cancer, Alzheimer‟s diseases (Sheeja and Kuttan, 2007). Plant remains a vital source of drugs
and nowadays much emphasis has been given to nutraceuticals.
Watermelon (Citrullus lanatus), family Cucurbitaceae is a vine-like flowering plant
originally from southern Africa. Its fruits, which is also called watermelon is a referred to by
botanists as a pepo, a berry which has a thick rind (exocarp) and fleshy center (mesocarp and
endocarp) pepos are derived from an inferior ovary and are characteristic of the
Cucurbitaceae. The watermelon fruit, loosely considered a type of melon-although not in the
genus Cucumis has a smooth exterior rind (green, yellow and sometimes white) and a sweet
interior flesh (usually pink, but sometimes orange, yellow, red and sometimes green if not
ripe). It is also commonly used to make a variety of salads, most notably fruit salad
(Pomeranz and Clifton, 1981).
HISTORY
Watermelon is thought to have originated in southern Africa, where it is growing wild.
Alphonse de Candolle, in 1882 already considered the evidence sufficient to prove that
watermelon was indigenous to tropical Africa (North Carolina State University). Though
Citrullus cololyn this often supposed to be a wild ancestor of watermelon and is now found a
native in North and West Africa. (Dane and Liu, 2007) suggest by chloroplast DNA
investigations that the cultivated and wild watermelon appears to have diverged
independently from a common ancestor, possibly Citrullus ecirrhosus from Namibia.
It is not known when the plant was first cultivated, but Zohary and Hopf note
evidence of its cultivation in the Nile valley from at least as early as the second millennium
BC. Although watermelon is not depicted in any Egyptian hieroglyphic text nor makes any
ancient writer mention, it finds of the characteristically large seeds is reported in Twelfth
dynasty sites numerous watermelon seeds were recovered from the tomb of Pharaoh
Tutankhamen (Daniel and Maria, 2000). By the 10th
century AD, watermelons were
cultivated in China, which is today the world‟s single largest watermelon producer. By the
13th
century, Moorish invaders had introduced the fruit to Europe according to John Mariana
the Dictionary of American food and Drink.
Charles Fredric Andrus, a horticulturist at the USDA vegetable Breeding laboratory in
Charleston South Carolina, set out to produce a disease-resistant and wilt-resistant
watermelon. The result was that gray melon from Charleston. Its oblong shape and hard rind
made it easy to stack and ship. Its adaptability meant it could be grown over a wide
geographical area. It produced high yields and was resistant to the most severe watermelon
disease Anthracnose and Fusarium wilt.
In Japan, farmers of the Zentsuji region found a way to grow cubic watermelons by
planting the fruits in glass boxes and letting them naturally assume the shape of the
receptacle. The square is designed to make the melons easier to stack and store, but the
square watermelons are often more than double the price of normal ones. Pyramid shaped
watermelons have also been developed, and any polyhedral shape may potentially also be
used.
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VARIETIES
There are more than 1200 varieties of watermelons ranging from less than a pound to more
than two hundred pounds, with flesh that is red, orange, yellow or white (Baker Creek, 2008).
Several notable varieties are as follows:
Carolina cross: - This variety of watermelons produced the current world record melons
weighing 262 pounds (119kg). It has green skin, red flesh and commonly produces fruits
between 65 and 150 pounds (29 and 68 kg). It takes about 90 days from planting to harvest,
(Baker Creek, 2008).
Yellow Crimson: - Variety of watermelon that has a yellow colored flesh. This particular
type of watermelon is “sweeter” and more “honey” flavored than another red flesh
watermelon (Anioleka et al., 2007).
Orangeglo: - This variety has a very sweet orange pulp, and is a significant oblong fruit
weighing 9kg (20-30 pounds). It has a light green rind with Jaggiest dark green stripes. It
takes about 90-100 days from planting to harvest. The Moon and Stars variety of watermelon
has been around since 1926 (Moons and star, 2008). It has many small yellow circles with
purple-black rind, (Stars) and one or two large yellow circles (Moon). The watermelon
weights 9-23kg (20-50 pounds) (Johnson and Sahaya, 2012). The flesh is pink or red and has
brown seeds. The foliage is also spotted. The time from planting to harvest is about 90 days.
Cream of Saskatchewan: - This variety consists of small round fruits around 25cm (10
inches) in diameter. It has a quite thin, light green with dark green striped rind, with sweet
white flesh and black seeds. It can grow well in cold climates. This variety was brought to
Saskatchewan, Canada by Russian immigrants. These watermelons take 80-85 days from
planting to harvest.
Melitopolski: - This variety has small round fruits roughly 28-30cm (11-12 inches) in
diameter. It is an early ripening variety that originated from the Volga River region of Russia
are known for the cultivation of watermelons. This watermelon is seen piled high by vendors
in Moscow in summer. This variety takes around 95 days from planting o harvest.
Densuke: - This variety has round fruit up to 25 16 (11 kg). The rind is black with no stripes
or spots. It can only grow on the Island of Hokkaido, Japan, where up to 10,000 watermelons
are produced every year. In June 2008, one of the first harvested melons is being sold at an
auction for 650,000 yen (6300 USD) making the most expensive watermelon ever sold. The
average selling price is generally around 25,000 yen (250 USD) (Vohra and Kaur, 2011).
LITERATURE REVIEW
Definition of Fats and Oils
Fats and oils can be defined by layman as the substances that are clearly fatty in nature,
greasy in texture, and are immiscible with water biochemically , fats and oils are defined as
substances that are insoluble in water and can be extracted by organic solvents of low polarity
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such as petroleum ether chloroform, methanol, n-hexane, etc. Chemists defined fats and oils
as the triglycerides of fatty acids in which one molecule of glycerol combined with three
molecules of long chain fatty acid with the elimination of water molecule.
Sources of Fats and Oils
Fats and oils from plants are found mostly in fruits and seeds while in animals it‟s stored as
fatty tissues which protect certain parts of the body and as insulation against cold. Synthetic
fats and oils are produced by oxidation of hydrocarbons to fatty acids, which are then
esterified with glycerol, or synthesis of fatty acids from carbohydrates by microorganisms.
Essential Oils
Essential oils are highly volatile substances isolated by physical process from an odoriferous
plant of a single botanical species. Therefore the fragrance of buds flowers leaves or bark,
and terminal branches of individual plants is due to the presence of these volatile oils. They
bear the name of the plant from which it is derived from example peppermint oil or rose oil.
Such essentials oil was thought to represent their very essence of odor and flavor. After many
years of research, it is now known that these essential oils are a mixture of up to twenty or
more fragrance chemical compounds of highly aromatic substances mostly benzene straight
chain hydrocarbons compounds of intermediate molecular lengths and terpene derivatives.
ECONOMIC IMPORTANCE OF WATERMELON
Nutritive Values
Watermelon raw (edible part) has the total energy of about 127kj (30kcal), Carbohydrates
content 7.55g, Sugars (6.2g), Dietary fiber (0.4g) Fat (0.15g) Protein (0.61g) Water (91.45g),
Vitamin A equivalent to 28µg (3%), Thiamin (Vit.B1) 0.033mg (3%), Riboflavin
(vit.B2)0.021mg (1%), Niacin (Vit.B3) 0.178mg (1%), Pantothenic acid (B5) 0.221mg (4%),
Vit.B6 0.045mg (3%), Folate (Vit. B9)3.4g (1%), Vit.C 8.1mg (14%), Calcium 7mg (1%) Iron
0.24mg (2%), Magnesium 10mg (3%), Phosphorus 11mg, Potassium 112mg (2%), Zinc
0.10mg (1%), (Iakshmi and Kaul, 2011).
Sources of Other Compounds/Substances
Watermelons contain a significant amount of citrulline, and after consumption of several
kilograms of watermelon, an elevated concentration is detected in the blood plasma, this
could be mistaken for citrullinaemia or other urea cycle disorders (Martin and Negro, 2007).
Watermelon rinds usually a light green or white color is also edible and contains
many hidden nutrients that most people avoid eating due to its unappealing flavor. They are
sometimes used as a vegetable (Martin and Negro, 2007)
In China, watermelons are stir-fried, stewed or more often pickled. When stir-fried the
skinned and fruited rind is cooked with olive oil, garlic, chili peppers, scallions, sugars, and
rum. Pickled watermelon rind is also commonly consumed in the southern us. Watermelon
juice can also be made into wine. Watermelon is also mildly diuretic and contains significant
amounts of beta-carotene. Watermelon with red flesh is a significant source of lycopene,
(Hdider and Lenucci, 2011).
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Health Benefits of Watermelons
Watermelon has plenty of potassium that is useful in cleansing the toxic depositions of
kidneys. Additionally, it‟s helpful in lowering amount of uric acid in the body, thus reducing
the risk of kidney problems and development of renal calculi. Besides these, being loaded
with water contents, it encourages frequent urinating; again it‟s useful for kidney cleansing.
Furthermore, the antioxidants in the watermelons ensure the well-being of kidneys.
A great deal of magnesium and potassium found in the watermelons are incredibly
effective in reducing the blood pressure level. The carotenoids contained in the fruit protect
walls of arteries and veins from hardening, thus helping to lower blood pressure level,
(Poduri et al., 2012).
Alongside tomatoes, watermelon has moved up to the front of the line in recent
research studies on high-lycopene foods. Lycopene is a carotenoid phytonutrient that is
especially important for our cardiovascular health, and an increasing number of scientists
now believe that lycopene is an important for bone health as well. Among whole, fresh fruits
that are commonly eaten in the U.S., watermelon now accounts for more U.S. intake of
lycopene than any other fruit, (Saha and Poduri, 2012).
Health scientists are becoming more and more interested in the citrulline content of
watermelon. Citrulline is an amino acid that is commonly converted by our kidneys and other
organ systems into arginine (another amino acid). The flesh of a watermelon contains about
250mg of citrulline into arginine. In particular, if a person‟s body is not making enough
arginine, higher levels of arginine can help improve blood flow and another aspect of our
cardiovascular health, (Azevedo et al., 2007).
There is also some preliminary evidence from animal studies that greater conversion of
citrulline into arginine may help prevent excess accumulation of fat in fat cells due to a
blocked activity of an enzyme called alkaline phosphatase, or TNAP, (Saha et al., 2012).
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MATERIALS AND METHODS
MATERIALS: The material used includes the fruits of fresh watermelons.
CHEMICALS
REAGENTS MANUFACTURER %PURITY
Petroleum ether B.O.H Chemical Ltd. England 99.8%
Sulphuric acid B.O.H Chemical Ltd. England 97%
Sodium hydroxide B.O.H Chemical Ltd. England
Conc. HCl B.O.H Chemical Ltd. England 99%
Phenolphthalein B.O.H Chemical Ltd. England
Pyridine B.O.H Chemical Ltd. England 96%
Ethanol B.O.H Chemical Ltd. England 99.9%
Starch C. Gerhard Gmbu Company
Potassium iodide B.O.H Chemical Ltd. England 97%
Iodine James Borough Puy Ltd. London 99%
Chloroform B.O.H Chemical Ltd. England 99.4%
Sodium thiosulphate B.O.H Chemical Ltd. England 98%
Glacial acetic acid B.O.H Chemical Ltd. England 99.8%
Potassium chloride B.O.H Chemical Ltd. England 97%
Ammonium sulfate B.O.H Chemical Ltd. England 98%
Boric acid indicator B.O.H Chemical Ltd. England
Sodium sulfate B.O.H Chemical Ltd. England 96%
Copper sulfate B.O.H Chemical Ltd. England 95%
APPARATUS
APPARATUS TYPE/MODEL COMPANY
Spatula Glass Pyrex glass England
Volumetric flask Glass Pyrex glass England
Measuring cylinder Glass Pyrex glass England
Burette Glass Pyrex glass England
Conical flask Glass Pyrex glass England
Tripod Metallic Pyrex glass England
Soxhlet set
Hot air oven Metallic, HQ4400 Muttle, Switzerland
GC-MS machine QP 2010 Shimadzu, Japan
Crucibles Ceramic Muttle, Switzerland
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Sampling
Watermelon fruits were purchased from Ramen-Kura, a local market for vegetables in Sokoto
state, in the northern part of Nigeria. The flesh is removed and the seeds collected were
washed and dried for easy removal of the epicarp.
Sample Preparation
The melon seeds were then subjected to sun-drying the melon seeds of 100g were dry-milled,
and the oil content was extracted by Soxhlet extraction method before being subjected to
chemical, proximate and GC- MS analyses respectively.
Extraction of Oil from Watermelon Seeds
From the grounded sample of watermelon seeds, 50g was weighed into an empty thimble
which was placed in a Soxhlet extraction set. 250ml of n-hexane solvent was then poured into
the extraction flask. The bottom of the extractor containing the sample was connected to the
extraction flask and placed on the heating mantle. As temperature increased steadily n-hexane
boiled and formed a vapor which was condensed by the condenser formerly attached to the
extractor and then dropped into the thimble dissolving and extracting oil present in the
powdered seeds. The extraction was carried out continuously for 6 hours.
After the extraction procedure, the thimble was removed then the n-hexane formed a
vapor which condensed and was collected in the receiver of the Soxhlet extractor while the
oil remained in the flask allowed to cooled and weighed. The procedure was repeated for the
remaining grounded sample of the watermelon seeds, and the oil was obtained.
Reagents Preparation
Na2S203.5H20 (0.1M)
Six (6.0g) of sodium thiosulphate was dissolved in a volumetric flask and made up to mark
250cm3 with distilled water.
Starch Indicator (1%)
One (1.0g) of starch was dissolved in 5sm3
distilled water and then made up to 100cm3
volumetric flask with hot distilled water refluxed for 5minutes and then allowed to cool.
KI (10%)
Ten (10g) of KI was dissolved in a beaker containing 90ml distilled water and then made up
to 100cm3
mark volumetric flask with distilled water.
KOH (0.1M)
One and a half i.e. (1.5g) of KOH was dissolved in distilled water and made up to mark
250cm3
distilled water.
Alcoholic KOH (0.1M)
A pellet of (1.4g) of KOH was dissolved in 50cm3 of ethanol and made up to mark 250cm
3
with distilled water.
HCl (0.5M)
About 10.6cml of concentrated HCl was poured into a beaker containing distilled water and
made up to 250cm3 with distilled water.
Phenolphthalein (1%)
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One gram (1.0g) of phenolphthalein was dissolved in 100cm3 of ethanol.
Solvent Mixture
The solvent mixtures were prepared by mixing 125cm3 of each of toluene and isopropyl
alcohol and 4cm3
of 1% phenolphthalein in isopropyl alcohol.
Na0H (0.25M)
One gram, i.e. (1.0g) of Na0H was dissolved in a 1dm3 volumetric flask and made up to mark
with distilled water.
ICI (Iodine Monochloride)
Pyridine 8.2ml by volume and 6ml of conc. H2S04 was mixed in 20ml cooled glacial acetic
acid, and 26ml of bromine was added to this solution. The mixture was then diluted to Idm3
with glacial acetic acid and was kept in the dark.
ANALYSIS AND DETERMINATIONS: PROXIMATE ANALYSIS
Determination of Moisture
The moisture content of melon seeds was determined by putting freshly obtained seeds in an
oven at 1050C. The moisture content was also determined by drying the rind and finding the
loss in weight.
Procedure
An empty crucible was weighed; 2g of fresh seeds were considered into it and placed in an
oven at 1050C for 24 hours. After 24 hours they were removed allowed to cool and weighed.
2g of the fresh seeds were weighed and shed dried and reweighed.
Determination of Ash Contents
Ash is the non-volatile inorganic matter of the sample that remains after subjecting it to high
temperature.
Procedure: - An empty crucible was weighed. 2g of the fresh sample was weighed and
transferred into the pre-weighed crucible. It was then heated in a muffle furnace at a
temperature of 6000C for s2 hours. It was allowed to cooled and weighed again.
Determination of Crude Protein
The Kjeldahl method was used for crude protein determination.
Principle
The method is based on the transformation of protein nitrogen and other nitrogenous
compounds other than nitrate and nitrite into ammonium sulfate by acid digestion with strong
acid usually concentrated H2S04 i.e.
Sample nitrogen + H2S04 (NH4)2 SO4
The ammonia present is digested with the help of sodium hydroxide and distilled out and
collected into receiving flask containing a boric acid indicator which changed color from
pink to green.
(NH4)2S04 + Na0H 2NH3 + H20 + NaS04
NH3 + H3BO NH+
4 + H2B03
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The nitrogen content was thus estimated by titration the content in the receiving flask with
standard acid (H2 S04).
H+
H2B03 H3B03
The amount of H+ consumed in the reaction is equivalent to the amount of nitrogen present
in the sample.
The amounts of protein were then estimated by multiplying the concentration of
nitrogen in the sample by a conversion factor of 6.6 which is equivalent to 16gN/100g
protein.
Procedure
From the dried grounded sample, 2grams was weighed into Kjeldahl digestion flask,
and a catalyst mixture was added (NaS04, CuS04 and Selenium oxide in the ratio 10:5:1).
The content was heated in the sulfuric acid (H2S04). The content was heated in the Kjeldahl
digestion unit until the digestion was completed (approximately 30minutes). The flask was
cooled, diluted volumetric flask and made up to the mark with distilled water. 10ml of the
aliquot was then taken into the digestion flask, and 20mls of 45% Na0H solution was added.
The content was diluted to about 200ml with distilled water and distilled using Kjeldahl
distillation apparatus.
The distillates were received into receiving flask containing 10mls boric acid solution
indicator. After distillation, the distillate was titrated with standardized 0.1M HCl to the
endpoint. The blank was determined in the same way without the sample.
DETERMINATION OF CRUDE FIBRE (AOAC 1990)
Procedure
From the grounded sample, 2grams was weighed and placed in a conical flask containing
200ml of 1.15% H2S04 sulphuric acid and boiled gently for 39 minutes. The content was
filtered, and the residues were scraped into the container with a spatula. The oxidant 1.25%
sodium hydroxide Na0H was added and allowed to boil gently for 30minutes. The content
was then filtered and washed thoroughly with hot distilled water.
The precipitate was allowed to dry, and the residue was scraped into a crucible and
dried overnight at 105OC in a hot air oven. It was then removed and cooled in a desiccator.
The sample was then weighed and ashed at 6000C for ninety minutes in a furnace. This was
finally cooled in a desiccator and weighed.
DETERMINATION OF CRUDE LIPID
Soxhlet extractor used for crude lipid determination. 2grams each of the dried grounded
samples were weighed into a porous thimble and its mouth covered with cotton. The thimble
was then placed in an extractor chamber, which was suspended above a receiving flask
containing petroleum ether (boiling point 40-600C). The flask was heated on a hot mantle and
the oil extracted.
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The extraction continues for about eight hours (8) after which the thimble was
removed from the soxhlet, and the apparatus was reassembled and heated on water-bath
apparatus for solvent recovery. The flask containing the crude oil was then disconnected,
cleaned up and placed in an oven at 1000C for thirty minutes (30). The flask was then cooled
in a desiccator and weighed.
DETERMINATION OF CARBOHYDRATES
To obtain the % carbohydrate content, all the above proximate analysis parameters have to be
summed and subtracted the value obtained out of 100. The remaining value obtained after
substitution is our percentage carbohydrate content in the sample. %CH0= 100- (% Ash +%
crude protein +% crude fiber + % crude lipid).
ANALYSIS AND DETERMINATION
PHYSICO-CHEMICAL ANALYSIS
DETERMINATION OF PEROXIDE VALUE
Peroxide value is an index of lipid deterioration expressed as peroxide per kg of fat or mili-
equivalent. It is the first products of unsaturated oil when peroxide value reaches a certain
level; complex change occurs with formation of ketone, aldehyde and hydroxyl group all
these are volatile and responsible for odor and flavor.
Procedure
From the extracted oil, 2g was weighed into a conical flask, and 25ml of glacial acetic acid
and chloroform was added. 1ml of KI solution was added and allowed to stand in the dark for
1 minute so that the solution becomes straw yellow. 35 ml of distilled was also added and
then followed with ten drops of 1% starch indicator. This was titrated against 0.004M
Na2S203.5H20 till blue color disappeared. The procedure was done in triplicate and blank was
estimated without adding oil.
DETERMINATION OF ACID VALUE
It is the percentage of free fatty acid expressed as oleic acid.
Principle
Acid value of oil is determined by titration of known weight of oil against 0.25N Na0H using
phenolphthalein as indicator
C17H33COOH + Na0H C17H33C00Na +H20
Procedure
From the extracted watermelon seed oil, 1g was weighed in a conical flask 50ml of denatured
alcohol was then added and shaken and two drops of phenolphthalein indicator were added to
the mixture then titrated against 0.25M Na0H with vigorous shake until the permanent light
pink color was observed.
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SAPONIFICATION VALUE
William and Devine's method was used (1961). Saponification number is the amount of
milligram of potassium hydroxide (KOH) required to completely saponify (I.0g) of oil.
Principle
When oils are treated with excess alcoholic (KOH), the oil gets saponified, and a definite
amount of (KOH) is used up. The excess KOH left unused may then be found by titrating
against 0.5M HCl.
Procedure
From the oil extracted 0.5g was weighed in a quick-fit-reflux flask, and 25ml alcoholic KOH
was then added. It was refluxed for 30minte so that it gets simmered. The flak was allowed to
cool, and 1ml of phenolphthalein indicator was then added and titrated against 0.5M HCl.
IODINE VALUE
The method described by Devine and Williams (1961) was used. This is the percentage of
iodine monochloride (ICI) regarding iodine absorbed by the oil.
Principle
The method is based on the treatment of a known weight of oil or fat with a known volume of
a standard solution of iodine monochloride (ICl). Estimation of ICI is by titrating iodine
liberated by adding excess potassium iodide. Titration was done against sodium thiosulphate
(Na2S203.5H20) with starch as indicator.
ICI + KI KCI +I
2Na2S203+ I2 Na2S406+2NaI
Procedure
From the oil extracted, 0.3g was weighed, and 10ml of carbon tetrachloride (CCl4) was then
added. Also, 25ml of Wiss solution was added the flask was shaken and allowed to stand in
the dark for one hour. 15ml of 10% potassium iodide and 100ml of distilled water was added
to the mixture. The mixture was titrated against (0.1M) Na2S203.5H20 until the blue
coloration disappeared which indicated an endpoint. The blank solution was also titrated at
the same time without oil.
DETERMINATION OF PERCENT FREE FATTY ACID
From the oil extracted, 10g was weighed and then boiled with 50ml ethanol, allowed to cool
and two drops of phenolphthalein indicator were added. The mixture was then titrated against
0.1M Na0H until the pink color was obtained.
Gas Chromatography-Mass Spectrometry (Gc-Ms) Analysis
The GC-MS analysis was carried out on a GC-MS-QP 2010 plus Shimadzu system and gas
chromatograph interfaced to a mass spectrometer (GC-MS) instrument employing the
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following conditions: column Elite – 1 fused silica capillary column (30m x 0.25mm 1D x µI
composed of 100% (dimethyl-polysiloxane).
For GC-MS detection, an electron ionization system with ionization energy of 70 eV
was used. Helium gas (99.999%) was used as the carrier gas at constant flow rate 1ml/min,
and an injection volume of 2µl was employed (split ratio of 10:1) injector temperature-
2500C ion-source temperature 280
0C the oven temperature was programmed from 110
0C
(isothermal for 2min.) to 280oC/min, ending with a 9min isothermal at 280
0C. Mass spectra
were taken at 70 eV; a scan interval of 0.5 sec and fragments from 40 to 550 Da. Total GC
running time was 36minutes. The relative percentage amount of each component was
calculated by comparing its average peak area to the entire areas; software adapted to handle
mass spectra and chromatogram was a turbo mass. The detection employed the NIST Ver. 2.0
the year 2009.
Identification Of Components
The interpretation of mass spectrum of GC- MS was done using the database of National
Institute of Standard and Technology (NIST) having more than 62,000 patterns. The mass
spectrum of the unknown components was compared with the mass of the known elements
stored in the NIST library. The name, molecular weight, and structure of the parts of the test
material were ascertained.
RESULTS
Proximate Analysis
The moisture content of the seed is quite low (4.56+ 0.04%) and falls within the range of
moisture content of similar seeds as shows from the table below. The ash content (4.20
+0.02%) obtained is higher than the established value for animal feed. The fat, crude protein,
and nitrogen-free extract are also high.
TABLE: 1 PROXIMATE COMPOSITION OF WATERMELON SEEDS
Parameters Composition
Moisture 4.50+0.04
Crude lipid 41.46+0.03
Crude Protein 25.50+05
Crude Fiber 6.34+0.01
Ash 4.20+0.02
Free Nitrogen 14.97+0.08
CHEMICAL ANALYSIS
Free Fatty acid (FFA) value for the watermelon seeds oil is (2.46 + 0.01%). The
saponification value and iodine value were all slightly higher in the oil.
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TABLE 2: PHYSIC-CHEMICAL PROPERTIES OF WATERMELON SEEDS OIL
Iodine value %Acid value saponification value free fatty acid peroxide
value
55.42+0.07 7.60+0.3 112.58+0.5 2.46+0.01 17.47+0.4
GC-MS ANALYSIS
The components present in the watermelon seeds oil were identified by GC.MS analysis as
shown in the table (3) below.
The active compounds with their molecular formula, molecular weight, retention time
(RT) and their percentage compositions are presented in table (3). Seven (7) compounds were
identified in the oil and the prevailing compounds are Palmitic acid (Hexadecanoic acid)
3.80%, Octyl propyl ester (1.03% Delta-Tocopherol (5.16%), Oleic acid (9-octadecenoic
acid) (17.84%), Propanediol acid (Dicyclohexyl ester) 3.74% Dioctyl ester (1, 2-
Benzenedicarboxylic acid) (0.98%) and Linoleic acid chloride 13.77% the spectrum profile
of seven major components with their respective retention time were shown in the table. The
mass spectrum of the compound with retention time 22.50 and 13.77 gave the primary peak.
Table 3.Detected components in watermelon seeds oil using GCMS technique.
SN Name of compound Mole formula Mole weight RT % composition
1 Palmitic acid C16H3202 256 18.H 3.80
2 Carbonic acid (Octadecylpropy ester C22H4403 356 6.51 1.03
3 Delta-Tocopherol C27H 4602 402 5.7 5.16
4 Oleic acid (9 octadecenoic acid ) C18H3404 282 19.8 17.84
5 Propanedioc acid C15H2404 268 6.18 3.74
6 Dioctyl ester C24H3804 390 23.10 0.98
7 Linoleic acid chloride (19,12-
octadecadienoyl chloride)
C18H31Cl0 298 22.50 13.77
unidentified compounds 53.68
Total % composition 100
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Umaru Ali Shinkafi Polytechnic Sokoto, Nigeria
The Chromatogram of the compounds detected from the watermelon seed oil
Structures of some of the identified compounds using GC-MS analysis
Oleic acid
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Umaru Ali Shinkafi Polytechnic Sokoto, Nigeria
DISCUSSION
The low moisture content of watermelon seed is advantageous regarding the shelf life of the
grain, with less moisture content seeds able to be preserved for a more extended period. From
literature, ash content for nuts, seeds, and tubers should fall within acceptable limits for
edible oils (1.5-2.5%) to be suitable for animal feed. The high quantity of oil content is an
indication that watermelon seed is another ready source of oil like the peanuts and soybean
seeds. The high protein content of the oil has a pleasant implication a society with high
protein deficiency. Fatty acid value of oil is highly essential in considering the quality of oil
because of the lower the free fatty acid, the better the quality of the oil which is (2.46%) and
acid value (7.60mg/Na0Hg) which fall within acceptable limits for edible oils
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Umaru Ali Shinkafi Polytechnic Sokoto, Nigeria
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