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CHAPTER ONE

1.0 INTRODUCTION

1.1 BACKGROUND OF THE STUDY

Plants provide an alternative strategy in search for new drugs. There is a rich abundance of

plants reputed in traditional medicine to possess protective and therapeutic properties. It is

likely that plants will continue to be a valuable source of new molecules which may, after

possible chemical manipulation, provide new and improved drugs ( Shah et al., 2006).

Traditional medicinalplants are a therapeutic resource used by the population of the

continent specifically for health care, which may also serve as starting materials for drugs

(Sofowora, 1993). Iwu et al. (1999) reported that infectious diseases account for one-half

of all deaths in the tropical countries. As a result, people of all continents have long

applied poultice and imbibed infusions of indigenous plants dating back to prehistory for

health purposes (Cowan, 1999). It comprises of therapeutics practices in existence for

hundreds of years before the development of modern scientific medicine and is still in use

today without any documented evidence of adverse effects.

According to the World Health Organization (WHO, 1977) a medicinal plant is any plant

which in one or more of its organ contains substances that can be used for the synthesis of

useful drugs. This definition distinguishes those plants whose therapeutics properties and

constituents have been established scientifically and plants that are regarded as medicinal

but which have not yet been subjected to thorough investigation. The term herbal drug

determines the part/parts of a plant used for preparing medicines (for examples: leaves,

flowers, seeds, roots, barks, stems, etc.). Furthermore, WHO (2001) defines medicinal

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plant as herbal preparations produced by subjecting plant materials to extraction,

fractionation, purification, concentration or other physical or biological processes which

may be produced for immediate consumption or as a basis for herbal products.

Medicinal plants contain biologically active chemical substances such as saponins, tannins,

essential oils, flavonoids, alkaloids and other chemical compounds ( Harborne,

1973; Sofowora, 1996) which have curative properties. These complex chemical substances

of different compositions are found as secondary plant metabolites in one or more of these

plants. Tyler (1999) has reported that plants also contain certain other compounds that

moderate the effects of the active ingredients.

Telfairia is classified in the tribe Joliffieae of the subfamily Cucurbitaceae. It comprises 3

species, of which Telfairia pedata (Sm. ex Sims) Hook.The names Telfairia pedata and

oyster nut are often erroneously used for Telfairia occidentalis. Cultivars of Telfairia

occidentalis are distinguished by seed colour, thickness of vine, size of leaf, growing

vigour, days to flowering and succulence. In Nigeria the two main cultivars are ugu-ala,

characterized by succulent, broad leaves, small black seeds, thick stem and slow growth

and ugu-elu which has a high growth rate, large brownish seeds with high viability and thin

stem with small leaves. The large succulent leaves of ugu-ala make this cultivar a

commercial vegetable in high demand, while the fast emergence and high growth rate of

‘ugu-elu’ is preferred by farmers because of quick returns. The seed is often

polyembryonic, which is useful for multiplication and in breeding ( Akoroda, 1990).

Telfairia occidentalis commonly called fluted pumpkin occurs in the forest zone of West

and Central Africa, most frequently in Benin, Nigeria and Cameroon. It is a popular

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vegetable all over Nigeria. It is rare in Uganda and absent in the rest of East Africa. It has

been suggested that it originated in south-east Nigeria and was distributed by the Igbos,

who have cultivated this crop since time immemorial. It is, however, equally possible that

fluted pumpkin was originally wild throughout its current range, but that wild plants have

been harvested to local extinction and are now replaced by cultivated forms. It has different

traditional names; among the Igbos, it is known as Ugu, iroko or aporoko in Yoruba, ubong

in Efik, umee in Urhobo and umeke in Edo ( Akoroda, 1990, Badifu and Ogunsina, 1991).

In this article we intend to only discuss the therapeutic potentials of  Telfairia

occidentalis  which by far outweighs the little possible toxicity in biological systems that

has been reported.

1.2 STATEMENT OF PROBLEM

Synthetic drugs have over time been associated with terrible adverse effects while plants

have been shown to possess protective and therapeutic properties with minimal adverse

effects (shah et al., 2006).

1.3 JUSTIFICATION

Telfairia occidentalis is a widely known vegetable in Nigeria and research as shown that it

contains phytochemicals that act as antioxidants. Hence, this research is aimed at

examining the effect of aqueous Telfairia occidentalis leaf on the liver of rats exposed to

garlic-induced oxidative stress.

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1.4 OBJECTIVE OF THE STUDY

The main objectives of the study are;

 To induced the rats with oxidative stress using garlic.

 To treat the rats with aqueous extracts ofTelfairia occidentalis

 To conduct liver functioning tests (AST, ALT, ALP) on the rat.

1.5 SIGNIFICANCE OF THE STUDY

This study will enable the researchers to know effect of aqueous pumpkin (leaf) extract on the liver of

rats exposed to garlic induced oxidative stress.

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CHAPTER TWO

2.0 LITERATURE REVIEW

2.1 Telfairia occidentalis

2.1.1 DESCRIPTION OF Telfairia occidentalis

Fluted pumpkin occurs in the forest zone of West and Central Africa, most frequently in Benin,

Nigeria and Cameroon. It is a popular vegetable all over Nigeria. It is rare in Uganda, and absent in

the rest of East Africa. It has been suggested that it originated in south-east Nigeria and was

distributed by the Igbos, who have cultivated this crop since time immemorial. It is, however, equally

possible that fluted pumpkin was originally wild throughout its current range, but that wild plants

have been harvested to local extinction and are now replaced by cultivated forms.

Telfairia is classified in the tribe Joliffieae of the subfamily Cucurbitoideae. It comprises 3 species,

of which Telfairia pedata (Sm. ex Sims) Hook.The names Telfairia pedata and oysternut are often

used erroneously for Telfairia occidentalis.

Cultivars of Telfairia occidentalis are distinguished by seed colour, thickness ofvine, size of leaf,

growing vigour, days to flowering and succulence. In Nigeria the two main cultivars are ‘ugu-ala’,

characterized by succulent, broad leaves, small black seeds, thick stem and slow growth, and ‘ugu-

elu’ which has a high growth rate, large brownish seeds with high viability, and thin stem with small

leaves. The large succulent leaves of ‘ugu-ala’ make this cultivar a commercial vegetable in high

demand, while the fast emergence and high growth rate of ‘ugu-elu’ is preferred by farmers because

of quick returns. The seed is often polyembryonic, which is useful for multiplication and in breeding.

It is Perennial, dioecious herb climbing by coiled, often branched tendrils to a height of more than 20

m; root system ramifying in the top surface of the soil; stem angular, glabrous, becoming fibrous

when old. Leaves arranged spirally, pedately compound with 3–5 leaflets; stipules absent; petiole

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(2–)4–11(–15) cm long; leaflets with petiolules 0.5–3.5 cm long, central one largest, up to 15(–19)

cm × 10(–12) cm, lateral ones asymmetrical, usually dentate in the upper two-thirds, sometimes

scabrid underneath, 3-veined from near the base. Male inflorescence an auxiliary raceme up to 3(–

5.5) cm long, on a peduncle up to 25 cm long, with at base of peduncle one long-pedicellate flower

flowering long before the others; female flowers solitary in leaf axils. Flowers 5-merous, cream

coloured, pedicel up to 4 cm long, receptacle campanulate, sepals triangular, up to 5 mm long, petals

free, oblong, fringed; male flowers with 3 stamens, two 4-locular and one 2-locular, with large

reddish connective; female flowers similar to male flowers but with inferior, cylindrical, 3-celled

ovary and 3 large, heart-shaped stigmas. Fruit a drooping, ellipsoid berry 40–95 cm × 20–50 cm,

weighing up to 6 kg, with 10 prominent ribs, pale green and covered with white bloom wax, fruit pulp

yellow, many-seeded. Seeds compressed ovoid, up to 4.5 cm long, black or brown-red. Seedling with

hypogeal germination, developing first a taproot and then numerous, spreading auxiliary roots;

epicotyl 5–12 cm long; cotyledons plan convex, fleshy. Telfairia occidentalis is a tropical vine grown

in West Africa as a leaf vegetable and for its edible seeds. Common names for the plant include fluted

gourd, fluted pumpkin, ugu (in the Igbo language), and ikong-ubong (in the Efik and Ibibio

languages). Telfairia occidentalis is member of the family Cucurbitaceae and is indigenous to

southern Nigeria. The fluted gourd grows in many nations of West Africa, but is mainly cultivated in

southeastern Nigeria and it is used primarily in soups and herbal medicines. Although the fruit is

inedible (Ajayi, 1990). The seeds produced by the gourd are high in protein and fat, and can,

therefore, contribute to a well-balanced diet. The plant is a drought-tolerant, dioecious perennial that

is usually grown trellised.

Telfairiaoccidentalis is traditionally used by an estimated 30 to 35 million people indigenous people

in Nigeria, including the Efik, Ibibio, and Urhobo. However, it is predominantly used by the Igbo

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ethnic group, who continue to cultivate the gourd for food sources and traditional medicines. A

recurring subject in the Igbo’s folklore, the fluted gourd is noted to have healing properties and was

used as a blood tonic, to be administered to the weak or ill. It is endemic to southern Nigeria, and was

an asset to international food trades of the Igbo ethnic group.

Telfairia is a small genus of flowering plants in the squash family which are native to Africa. They

include Telfairia occidentalis, the fluted pumpkin, which is an important vegetable in Nigeria and

other African nations. It is also known as the oyster nut, a common name it shares with its relative

Telfairia pedata. These are woody and herbaceous dioecious vines which bear squashlike fruits that

contain large, nutritious oily seeds. The third species is Telfairiabatesii which is very rare and not

cultivated.

These vines grow quickly to lengths of 30 meters or more, using tendrils to scale trees.

Telfairiapedata is grown as an ornamental plant for its attractive foliage. The seeds of

Telfairiaoccidentalis and Telfairia pedata are over 25% protein and 55% oil, making them a good

source of nutrition. They are used as a traditional food source for nursing mothers, the seed flour is

fermented to make baby food, the leaves and shoots, especially of Telfairia occidentalis , are eaten as

vegetables.

One recent concern has been the impact of Telfairamosaic virus, which, along with other problems, is

a threat to the already low genetic diversity of plants of this genus.

The effects of chronic garlic intake on various endogenous antioxidant enzymes and lipid

peroxidation on two major organs, the liver (L) and kidneys (K), were investigated. Wistar albino rats

were fed with fresh garlic homogenate daily by gavage in three different doses (250, 500 and 1000

mg/kg/day) for 30 days. After this period, rats were sacrificed and liver and kidneys were harvested

for biochemical estimation. In comparison to saline-treated rats, the 250 mg/kg/day dose significantly

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(P<0.02) reduced thiobarbituric acid reactive substances (TBARS) (L: 187.48+/-9.23 vs 150.66+/-

11.45; K: 177.38 15.88 vs 120.66+/-9.39 nmol/g wet. weight) and glutathione peroxidase (GPx) (L:

0.2438+/-0.05 vs 0.0046+/-0.0005; K: 0.1459+/-0.034 vs 0.0055+/-0.0003 U/mg protein). There was

no change in catalase and reduced glutathione (GSH) but superoxide dismutase (SOD) increased

significantly (P<0.01) (L: 5.49+/-0.76 vs 18.38+/-2.26; K: 11.47+/-1.48 vs 21.22+/-3.19 U/mg

protein). Both 500 and 1000 mg/kg/day doses significantly (P<0.05) reduced endogenous

antioxidants (catalase and SOD) without altering TBARS. A 1000 mg/kg/day dose of garlic caused

marked histopathological and ultrastructural changes in both liver and kidneys. The results suggest

that garlic in low doses has the potential to enhance the endogenous antioxidant status, although at

higher doses a reversal of these effects is observed. The present study also highlights the potential

ability of a high dose of garlic to induce morphological changes in the liver and kidneys, indicating

the need to identify a safe dose range for garlic.

2.1.2 SCIENTIFIC CLASSIFICATION OF Telfairia occidentalis

Kingdom: Plantae

Class: Tracheophytes

Sub class: Rosids

Order: Cucurbitales

Family: Cucurbitaceae

Genus: Telfairia

Species: Telfairiaoccidentalis

2.1.3 ECOLOGY

In the wild, fluted pumpkin occurs in forest fringes and secondary forest, possibly often as a relic of

former cultivation. Fluted pumpkin grows fast in the warm humid tropics, producing edible leaves in

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the rainy season and at the beginning of the dry season, for a period of 6–10 months. In traditional

agriculture, it is a rainfed crop and water deficiency during the dry season reduces its productivity.

Although it is fairly drought tolerant, rainfall appears to be the major factor in its productivity. The

best leaf and fruit yield and highest plant survival rate occur when the plants are irrigated 2–3 times

per week during the dry season. Fluted pumpkin can be grown under a wide range of soil conditions.

It can be managed as a short-term perennial when grown on well-drained soils, slightly shaded and

mulched. On soggy soils and in sunlit spots it can only be grown as an annual.

2.1.4 DISEASE AND PEST

White leaf spot disease, caused by Phoma sorghina, reduces the leaf lamina. It also affects the seed. It

is controlled by biweekly foliar spraying with Dithane M-45 at a concentration of 500 ppm. Fusarium

moniliforme forms a dry powdery mass of mycelia on the fruits. Erwinia aroideae causes soft rot of

the leaves with yellowish ooze; it also affects the fruits. A prevalent virus disease is Telfairiamosaic

virus (Telfairiamosaic potyvirus) (TeMV), causing mottling of the leaves and low leaf yield; it also

causes chlorosis, stunting and abnormal fruit development. It is transmitted by the aphid Aphis

spiraecola and via the seed. Telfairia occidentalis is remarkably resistant to root-knot nematodes

(Meloidogyne spp.).

Rhizopus stolonifer, Aspergillus niger, Botryodiplodia theobromae and Erwinia spp. are diseases of

Telfairia occidentalis fruits in storage. Fungi may cause up to 95% loss, bacteria only 5% loss in

long-term fruit storage.

A common pest ofTelfairia occidentalis is the grasshopper Zonocerus variegatus which feeds on the

foliage and stems. The leaf beetle Copa occidentalis feeds on the leaves, flowers and other plant

parts, while Spodoptera caterpillars feed on leaves and bore into fruits. Pachmola (flower beetles)

and Nezara spp. (green shield bug) feed on leaves, stem and fruits. Margaronia indica defoliates the

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plant, white beetle (Baris spp.) feeds on fruits. Sylepta derogata, Aphis gossypii and Aphis

spiraecola hinder growth by feeding on stem, foliage and flower buds, and transmit viruses. There are

some unidentified predators that feed on the aphids. Thrips of the genus Taeniothrips cause flower

abortion.

2.1.5 ANTIOXIDANT AND FREE RADICAL SCAVENGING PROPERTY

Almost all organisms possess antioxidant defense and repair systems that evolved to protect

them against oxidative damage, these systems are insufficient to prevent them entirely.

However, antioxidant supplements or foods containing antioxidants may be used to help

human body reduce oxidative damage (Yang et al., 2002). In recent years, there has been a

particular interest in the antioxidant and health benefit of phytochemicals in food and

vegetables. This was as a result of their potential effects on human health ( Wei and Shiow,

2001). Many researchers especially in the field of medical sciences have observed  free

radical scavenging ability and antioxidant property in Telfairia occidentalis. The darkish

green leafy vegetable leaves of Telfairia occidentalis and extracts (such as aqueous and

ethanol extracts) from the leaves have been found to suppress or prevent the production

of free radical and scavenge already produced free radical, lower lipid peroxidation status

and elevates antioxidant enzymes (such as superoxide dismutase and Catalase) both in

vitro and in vivo (Oboh et al., 2004, 2006; Nwanna and Oboh, 2007; Adaramoye et al.,

2007; Emeka and Obidoa, 2009; Kayode  et al., 2009; Kayode et al., 2010). Telfairia

occidentalis has also be found to protects and ameliorates oxidative brain and liver

damaged induced by malnutrition in rats (Kayode  et al., 2009, 2010). Nwanna and Oboh

(2007) reported the hepatoprotective property of polyphenol extracts of  Telfairia

occidentalis  leaves on acetaminophen induced liver damaged. Oboh (2005) reported that

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both aqueous and ethanolic extractof Telfairia occidentalis leaves protect the liver cells

against garlic- induced oxidative damage. However, the aqueous extract is more effective

than the ethanolic extracts, which could be attributed to the higher antioxidant activity of

the aqueous extracts of Telfairia occidentalis leaves. Hepatoprotectiveeffects of Telfairia

occidentalis  leaves have been reported by Eseyin et al. (2005b), Emeka and Obidoa

(2009) and Kayode et al. (2010). The use of the leaves in folk medicine in Nigeria in the

treatment of certain diseases in which the participation of  reactive oxygen species have

been implicated could be as a result of the antioxidant and free radical scavenging ability.

2.1.6ANTIPLASMODIAL AND ANTIMICROBIAL PROPERTIES

Malaria is a potentially life-threatening disease in the tropics as it affects over 400 million

people yearly and is responsible for the deaths of an estimated 10,000 women of

reproductive age and over 1 million infants and young children each year ( Barbin,

1989; Mishra et al., 2003). Drug resistance, increases in the production and circulation of

fake drugs and high cost of newer and effective drugs have been a major factor affecting

the poor populace, thus making their choice of herbal remedies inevitable and economical.

A study has shown that the ethanol root extract of  Telfairia occidentalis possess

antiplasmodial potential. The blood schizontocidal activity of the root extract is

comparable to that of chloroquine (Okokon et al., 2007). The ethanolic and aqueous

extracts of Telfairia occidentalis  have been reported to show inhibitory effect on growth on

some of the commonly encountered Enterobacteriaceae in Nigeria, namely   Escherichia

coli,  Pseudomonas aeroginosa, Proteus sp. and Salmonella typhii. However, both extracts

did not inhibit the growth of the fungi tested, which are  Aspergillus favus,  Aspergillus

fumigatum,  Peniccillium italium and  Geotrichum albidun  (Oboh et al., 2006). Antibacterial

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activity of the leaves was also reported by Odoemena and Onyeneke

(1998) while Oluwole et al., (2003) reported Telfairia occidentalis  anti-inflammatory

activities. Antibacterial activity of the root extract against  Staphylococcus

aureus, Streptococcus pyogenes, Shiegella dysenterae and  Kliebsiella pneumoniae has

been reported by Odoemena and Essien (1995).

2.1.7 Telfairia occidentalis BOOSTS BLOOD LEVEL AND BEAT DIABETES

In Nigeria, the herbal preparation of the plant has been employed in the treatment of

anaemia, chronic fatigue and diabetes (Alada, 2000; Dina et al., 2006; Aderibigbe et al.,

1999). Anaemia constitutes a serious health problem in many tropical countries because of

the prevalence of malaria and other parasitic infections. In anaemia there is decreased level

of circulating haemoglobin, less than 13 g dL -1  in male and 12 g dL -1  in females (Okochi et

al., 2003). In the tropics, where malaria is endemic, between 10 to 20% of the population

presents less than 10 g dL -1  of Haemoglobin (Diallo et al., 2008). Children are more

vulnerable. The leaves are rich in iron and play a key role in the cure of anaemia, they are

also noted for lactating properties and are in high demand for nursing mothers ( Okoli and

Mgbeogu., 1983).

Type 2 diabetes is associated with increased  oxidative stress, which probably results both

from excess generation of reactive oxygen species and decreased antioxidant defenses

(Baynes, 1991; Tribe and Poston, 1996). In recent years, it has been known that, the most

important factor to increase the free radicals production in diabetes is the hyperglycemic

status, which can induce damage through overproduction of superoxide radical in the

mitochondria (Brownlee, 2001). Superoxide is converted to hydroxyls, which can diffuse

through membranes and initiate lipoperoxidation. The oxidation of unsaturated lipids has

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implications not only for atherosclerosis, but also for stability and integrity of the red cell

membranes (Steinberg et al., 1989). Increased levels of lipoperoxidation as evidenced by

breakdown products like malondialdehyde have been found in erythrocytes and plasma type

2 diabetic patients. Supplementation with antioxidants is therefore, an attractive potential

therapy. Aqueous extracts of Telfairia occidentalis  had been reported to reduce blood

glucose level and also have antidiabetic effects in glucose induced hyperglycemic and

streptozotocin (STZ) induced diabetic mice (Aderibigbe  et al., 1999), while it did not alter

the glucose levels in normoglycemic mice. Salman et al., (2008) also reported

reduced bloodglucose level by Telfairia occidentalis  leaves in male rats. Hypoglycemic

effects have also been reported by many researchers (Aderibigbe   et al., 1999; Eseyin et

al., 2000, 2005c, 2007; Nwozo et al., 2004).

2.1.8 GENERAL HEALTH BENEFITS DERIVED FROM Telfairia occidentalis

Telfairia occidentalis is an important staple vegetable grown in Nigeria. The plant

produces luxuriant edible green leaves, which are rich in iron and vitamins. Stems of the

plants have branching, long twisting tendrils and the leaves are divided into three to five

leaflets with the terminal leaflets up to 15 cm long, while the male plant is grown

principally for leaves and seeds, which are important soup condition. Recent studies have

shown that Telfairia occidentalis leaf is rich in minerals (such as iron, potassium, sodium,

phosphorus, calcium and magnesium), antioxidants, vitamins (such as thiamine, riboflavin,

nicotinamide and ascorbic acid, phyto-chemicals such as phenols. Harvesting of fluted

pumpkin takes place 120-150 days after sowing ( Longe et al., 1983; Okoli and Nyanayo,

1988; Akoroda  et al., 1990; Ladeji et al., 1995; Oboh and Akindahunsi, 2004; Horsfall and

Spiff, 2005; Oboh, 2005; Fasuyi, 2006; Oboh et al., 2006; Ajibade et al., 2006; Kayode et

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al., 2009). The leaves contain essential oils, vitamins; root contains cucubitacine,

sesquiterpene, lactones (Iwu, 1983). The young leaves sliced and mixed with coconut water

and salt are stored in a bottle and used for the treatment of convulsion in ethno medicine

(Gbile, 1986). The leaf extract is useful in the management of cholesterolemia, liver

problems and impaired defense immune systems ( Eseyin et al., 2005a, b).The roots are

used as rodenticide and an ordeal poison ( Gill, 1992). The essential amino acids contents

compared favorably with those of important legumes ( Asiegbu, 1987). The amino

acid profile of Telfairia occidentalis  had also been shown to be very rich and includes

alanine, aspartate, glycine, glutamine, histidine, lysine, methionine, tryptophan, cystine,

leucine, arginine, serine, threonine, phenylalanine, valine, tyrosine and isoleucine ( Tindall,

1968; Fasuyi, 2006). Emeka and Obidoa (2009) study reveals that the long term feeding

of Telfairia occidentalis-supplemented diet caused a significant increase in weight of

animals which may be due to its content of rich nutrients.

The seeds are highly nutritious and are roasted or boiled and eaten like the seeds of

breadfruit (Treculia); they are also sometimes used as soup thickeners ( Okoli and

Mgbeogu, 1983). The seed is very rich in oil, especially unsaturated  fatty acids which form

61% of the oil (Odoemena and Onyeneke, 1998; Okoli and Nyanayo, 1988). Akubue et al.

(1980) and Taylor et al. (1983) have documented that Telfairiaoccidentalisseeds are a good

source of four minerals required in human nutrition. There report showed that the seed

contained 29% oil and 30% protein. Asiegbu (1987) reported Telfairiaoccidentalisseed

contain 47% oil and 31% protein. The protein was said to be markedly deficient in the

sulphur amino acid. Longe et al. (1983) reported that Telfairiaoccidentalisseeds had 53%

fat, 22% protein, 3% fibre, 15% carbohydrates and 2% ash. Oyolu (1980) observed that

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vegetables will continue to remain the primary source of proteins, minerals and vitamins in

African countries, he noted that leaves and edible shoots of Telfairiaoccidentalistogether

contain 85% moisture, while the dry portion of what is usually consumed contains

11% crude protein, 25% carbohydrate, 3% oil, 11% ash and as much as 700 ppm iron. The

results of the study carried out by Christian (2007) shows that the seed contained essential

nutrients in significant amount that can supplement other foods. The levels of  crude

protein (3.47%), crude fat (31.38%), moisture (10.93), Ash (2.02%), carbohydrate

(50.08%), fibre (2.12%), calcium (280 μg g -1 ), phosphorus (2100 μg g -1 ), iron (69 μg g -1 ),

sodium (10.80 μg g -1 ), potassium (1280 μg g -1 ), vitamin A (890 IU) and vitamin C (0.7 μg

g -1 ) detected in the seed were compared with  nutritional composition of some plant foods in

Nigeria. The study shows that the seed of Telfairia occidentalis  Hook F. is high in

carbohydrate, fat and phosphorus. The seed also contained levels of  vitamin A which can

supplement other dietary sources. The oil of Telfairia occidentalis  seeds have a high iodine

values compared to palm oil which indicates that the oil has a high content of unsaturated

fatty acids relative to palm oil. This suggests that it may be used as edible oil for cooking

or manufacturing of margarine (Christian, 2007).

The high protein content in leaves of plants such as  Telfairia occidentalis could have

supplementary effect for the daily protein requirement of the body. The symptoms of

protein energy malnutrition such as Kwashiorkor and Marasmus were rarely observed

among dwellers in region where adequate amount of protein is obtained from fruits/seeds

and leaves of plants rich in proteins such as Telfaira occidentalis  (Dike, 2010; Kayode et

al., 2010; Kayode et al., 2009).

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Fasuyi and Nonyerem (2007) investigations shows that Telfairia occidentalis leaf meal

caused increased growth in birds. Adaramoye et al. (2007) reported that Telfairia

occidentalis  leaves has hypolipdemic effect and may be a useful therapy in

hypercholestolemia. This confirms the research work of Eseyin et al. (2005a). The vitamin

A content and consumption pattern of some green leafy vegetables (which

includes Telfairia occidentalis) among pregnant women in Calabar, Nigeria was

investigated by Williams et al. (2009). They observed that Telfairia occidentalis  has the

highest vitamin A content which is adequate enough to sustain their  vitamin A requirement

(Williams et al., 2009). The fruits of  Telfairia occidentalis have been utilized in the

production of marmalade (Egbekun et al., 1998). The use of Telfairia occidentalis in

reproduction and fertility in traditional medicine is gradually becoming a thing of interest

in medical science. A study carried out by Nwangwu and his colleague shows that   Telfairia

occidentalis  has the potential to regenerate testicular damage and also increase

spermatogenesis (Nwangwa et al., 2007). However, more research work is required to

establish this observation.

2.2 Oxidative Stress

2.2.1 Toxicity Of Reactive Oxygen Specie

Oxygen is used in metabolic reactions. Sometimes it reacts with the metabolic compound to form

a free radical which causes biological damage (Rahman, 2007). Oxygen is a stable product but

when there is gain or loss of an electron in oxygen molecule or its compound it becomes

reactiveoxygen species. Toxicity of oxygen refers to the toxic endpoints of the oxygen

formation.Oxidative stress refers to the distorted balance between the transformation of the reactive

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oxygenspecies from one form to another indulging beneficial component of cell or

biomolecules(Alvarado and Acre, 2016).

Reactive oxygen species (ROS) play a most important role in variousbiological processes, like as the

oxidative burst reaction which is necessary for phagocytes, andare taking part in a variety of cell

signaling pathways. In the cytosol, IκB is a sequester andinhibitory factor of NF-κB. ROS are also

participating in moderating the action of specifictranscription factors, containing activator protein–1

and NF-κB. So ROS can play a veryimportant role in inflammation modulating. Perturbed reduction

state of cellproduces lipid peroxides, protein carbonyls and other adducts in the cell. These adducts in

celldeposits in clumps to inactivate the normal cell functioning processes of the cell that lead

DNAdamage, strand breaks etc.(Giordano, 2005).

2.2.2 Diseases / Disorder Linked To ROS

The reactive oxygen species are the product of cellular metabolism in also normal condition and

readily they are important for cellular signaling pathways. But in some extreme condition,

theaccumulation of ROS up to a limit can cause pathological condition responsible for variousdiseases

(Fanjul-moles, 2016).The reactive oxygen species generated through the process ofoxidative stress are

superoxide radical, hydroxyl radical and hydrogen peroxide. Oxidative DNAdamage is mostly indirect

and adduction of radicals to the DNA can cause mutation so that thecell may become cancerous.

Mitochondrial ETC is one of the types of internal source factor forthe formation of free radicals in

internal cellular metabolism (Shinde et al., 2012).

2.2.2.1 Alzheimer’s Disease

It is a kind of memory loss disease which shatters the memory and other main mental functions.

In this disease, brain cells lose connections, degenerate and finally cells die which cause slowly

17
memory loss. Recent studies reveal that the neuron mitochondria from aging in the rats, are

moresusceptible to produce reactive oxygen species that cause neurodegenerative dementia

likeAlzheimer's disease. One of the common forms of dementia thataffect our older population

characterized by degeneration of neurons within the brain. It affectshippocampus region of the brain

which is the responsible part of the brain for memory formationso it causes loss of memory and

premature aging(Parihar and Brewer, 2007).

2.2.2.2 Parkinson’s Disease

. Parkinson’s disease is a common neurodegenerative disease induced bysome stressors from the

endoplasmic reticulum and oxidative damage It is a kind of movement disorder which affects the

central nervous system, responsible for bodymovement so in this disease, patient’s lose body balance

and movements. Over expression of α-synuclein and oxidative stress has been implicated in the

neuronal cell death in the Parkinson’s disease. In Parkinson’s disease, selective degeneration of

dopaminergicstimuli of neurons occurs (Xiaotian et al., 2013).

2.2.2.3 Amyotrophic Lateral Sclerosis

It is a nervous system related disease which causes muscle weakness and loss of the movement or

control caused by the motor neurons degeneration. ALS probably leads to a new disease, skeletal

muscledystrophy in infected patients (Kong and Xu,1998). The brain has two types of receptors

glutamate and GABA receptors. Glutamate is achemical mediator which stimulates motor neurons so

its excess formation, is possiblyresponsible for motor neuron destruction in ALS. The GABA receptor

acts to lessen the effectsof glutamate. Gabapentin and Baclofen named medicines act as a GABA

modulators whichincrease GABA activity, are the probable cure for ALS (Amyotrophic lateral

sclerosis) (Diana et al., 2017).

2.2.2.4 Cardiac Disease

18
Oxidative stress and ROS are most factors of cardiac diseases like congestive heart

failure,hypertension, and atherosclerosis (Sugamura and Keaney, 2011). Smooth muscles and

cardiacmuscles, on the regulation of metabolic process, release free oxygen radicals in the form of

byproducts of the process. This leads to myocardial infarction. Some phagocytescells utilize the LDL

cholesterol and oxidize LDL that lead to the formation of artery cloggingplaque and thus responsible

for the stroke. ROS have been involving inthe important processes which have notable effects on

cardiac functions like hypertrophy, ionflux (calcium), extracellular matrix (ECM) configuration,

metabolism, signaling of variousgrowth factors, cytokines and gene expression. Thioredoxin and

thioredoxin reductase(enzymatic pathways of formation of ROS) together form an additional enzymatic

antioxidantand redox regulatory system that has been implicated in a wide variety of ROS-related

processes.Thioredoxin and thioredoxin reductase can catalyze the regeneration of many

antioxidantmolecules, including ubiquinone, lipoic acid, and ascorbic acid, and as such constitute

animportant antioxidant defense against ROS. Deletion of thioredoxin reductase results

indevelopmental heart abnormalities and in cardiac death secondary to a severe dilatedcardiomyopathy

(Giordano, 2005).

2.2.2.5 Cancer

Free radicals play an important role as the carcinogenic molecule for DNA damage whichinhibits DNA

repair. Attack of free radicals on the hydroxyl group of nucleic acids makes and mutation

fragmentation of nucleic acid thus giving rise to cancer. ROS cause DNA damage andleads to DNA

strand break or fragmentation of the bases. Oxidative products of DNA induceactivation of some

transcription factor like NF-κ B (nuclear factor κ B) that causes abnormal cell

19
growth. Post-transcriptional modification by phosphorylation and nitration by free radicals alsoinduces

cancer risk (Simone et al., 2010). Endogenous and exogenous stimuli participate incancer

developmental process. Formation of Oxygen-free-radicals (OFR), which attacks not only

on the bases but also the deoxyribosyl backbone of DNA, is one kind of endogenous damage

stimuli which is format through the intermediate oxygen reduction. OFR is a class of important

carcinogens for DNA-damage (Valko et al., 2004).

2.2.2.6. Ageing Bones

Remodeling of bone is a natural phenomenon where the bone absorption and formation takes place

during entire life. Less formation of bones and more breaks down will causedeteriorating bone health in

aging people. With the aging of human, could occur the deficienciesof both primary and secondary

Vitamin D which may be participated in the pathogenesis processof senile osteoporosis and also take

part in the development of osteomalacia in an adult humanbeing. Although a mild deficiency could lead

to a stage of secondary hyperparathyroidism, withresults in the development of osteoporosis (Lau and

Adarchi, 2011). Decreased mineralcontent and fragility are the symptoms of aging bones. Loss of bone

density may be related toosteoporosis where the bones are more prone to fractures. In the bones, the

rate of repair andregain their mechanical adeptness (power) after fracture, the rate of revascularization,

cellproliferation & differentiation of the cells and bones healing process, decreases with aging(Boskey

and Coleman, 2010).

2.2.3 Pathophysiological Significance Of Free Oxygen Radicals

Free radical plays significant role in controlling the blood pressure and also fight for the cures

ofinfection. Formation of reactive oxygen species is a physiological process, which increases

theproduction of free radicals and also its lead to the dysbalance between the production of radicalsand

antioxidants which could lead to oxidative stress with the changes of various biologicalfunctions and

20
structural changes in the cells (Halliwell, 1991). Immune cells participate indefense system by

destroying foreign molecules such as virus or bacteria by producing oxidantsor free radicals. Some

phagocytic cells like macrophages, eosinophils, neutrophils as well as Tand B lymphocyte contain

NADH oxidase which forms superoxide free radicals responsible fordamaging the micro-organisms.

The process is known as the oxidative or respiratory burst. It is the defense mechanism used by the

immune cells of the body (Abheri et al., 2010).

2.2.4 Antioxidants And Their Mechanism Of Action

Antioxidants are the compounds that can stabilize ROS. These molecules are the scavengers offree

radicals and get easily oxidized. Vitamins are the most important class of non-enzymatic antioxidants.

There are two classes water soluble like vitamin C and fat soluble vitamin A(retinoic acid or retinol)

and vitamin E. Vitamin E ( α-tocopherol ) is a predominant scavengerthat has its significant activity in

the protection of biomolecules of biomembranes, which areattacked by free radicals(Irshad and

Choudhary,2002). Supplement of vitamin E, and probablyvitamin C, being able to significantly lower

lipid oxidative damage in both smokers andnonsmokers, so we can conclude that antioxidants vitamins

supplementation reduces oxidativedamage in humans [McCall and Frei, 1998].

2.3 Liver

The liver has a particularly intriguing immunological milieu consisting ofa largest reticulo-endothelial

cell network in the body and being a major source of many components of the innate immune response

including acute phase and complement proteins as well as inflammatory cytokines and chemokines.

The organ is also a significant site of immune-mediated damage initiated by infectious, autoimmune

and malignant stimuli. Recent studies have demonstrated that the liver is also an important site of innate

immune system. The innate or natural immune system is the first rapid line-defense against

environmental threats such as microbial infection and physical or chemical injury. Sequential activation

21
of innate and adaptive immune response is crucial for elimination of microorganisms and for immune

response orchestrated by dendritic cells linking innate and adaptive arms of immune system. Unique

repertoires of dendritic and lymphoid cells including NKT cells and regulatory T lymphocytes modify

the immune response in the liver. Non-immune cells of the liver including endothelial cells,

hepatocytes and biliary epithelial cells also contribute to local immunological potential. All of these

elements play roles, together and independently, determining the outcome of immunological

stimulation within the liver. In addition, immune response upon exposure to exogenous or autogenic

agents varies depending on the host genetic backgrounds. The genetic basis of immune response will

offer new approaches to understanding the pathophysiology, diagnosis and management of patients

with liver diseases.

2.3.1 Liver Enzymes

2.3.1.1 Alanine Aminotransferase (ALT)

Liver cells produce the ALT enzyme. ALT levels increase when liver cells are damaged or are dying.

The higher the ALT levels, the more cell death or inflammation of the liver is occurring. However,

ALTs are not always good indications of how well the liver is functioning; only a liver biopsy can

reveal that. ALT levels can remain low even when the liver is inflamed or is developing scar tissue, or

during a child's immune tolerant stage of the disease.

2.3.1.2 Aspartate Aminotransferase (AST)

Like the ALT enzyme, AST is an enzyme produced by liver cells, but AST is also produced by muscles

and it can be elevated in conditions other than liver disease. For example, AST is often elevated during

a heart attack. In many cases of liver inflammation, the ALT and AST levels are equally elevated. In

some conditions, such as alcoholic hepatitis, AST levels may be higher than ALT levels. AST levels

22
can be normal, yet there can be liver damage Occurring. This test adds one more perspective to the

picture of liver disease.

2.3.1.3 Alkaline Phosphatase

Alkaline phosphatase is an enzyme produced in the bile ducts, intestines, kidney, placenta and bone.

This enzyme is measured to help doctors determine if a disease could be concentrated in the bile duct or

in the liver. When this enzyme level is high, and when ALT and AST levels are fairly normal, there

may be a problem in the bile duct, such as an obstruction. Some bone disorders can also cause alkaline

phosphatase levels to rise.

2.3.1.4 Gamma-Glutamvltranspeptidase (GG)

This enzyme, like alkaline phosphatase, is produced in the bile ducts and may become elevated when

there is a bile duct disorder. Elevations in GGT and alkaline phosphatase usually suggest bile duct

disease. Measurement of GGT s an extremely sensitive test; it can be elevated when there is any liver

disease. Elevated GGT levels are also caused by drugs (even when taken at prescribed levels) and they

are often elevated in heavy drinkers, even when there IS no liver disease.

23
CHAPTER THREE

3.0 METHODOLOGY

3.1 SAMPLE COLLECTION AND PREPARATION

Telfairia occidentalisleaves and Garlic were collected from Ishara Market, Ishara Remo, Ogun State.

Nigeria.TheTelfairiaoccidentalis leaves were sorted out and washed under running water to

remove dirt. They were chopped into small pieces, oven-dried, and milled into a fine

powder.

3.2 PREPARATION OF THE EXTRACT

The aqueous extract was prepared using Soxhlet extracting method. 65g of the powdered

sample was extracted with 325ml of distilled water.Telfairia occidentalis extraction was done at

the biochemistry laboratory, The Gateway(ICT) polytechnic Sapaade, Ogun state. Nigeria.

3.3 APPARATUS AND REAGENT

3.3.1 APPARATUS

Apparatus used for the experiments are : Soxhlet apparatus, Heating mantle, 250ml beaker, Metal

bead cannula, 2ml syringe, Hand gloves, Distilled water, Whatman filter paper, Nose mask, 5ml

syringe, Universal bottles, EDTA bottles and Dissecting tools.

3.3.2 REAGENTS

All reagents used were obtained from recognised dealers and were ensured to be of analytical grade.

3.4 EXPERIMENTAL ANIMALS

Twenty five adult, male Albino rats of approximate age (6-8 weeks) and weight (weighing

200 ± 50 g) were used in the experiment. Animals were kept in clean cages and given

24
normal diet and clean water ad libitum. Animals were acclimated for two weeks before

induction.

3.5 EXPERIMENTAL DESIGN

These were divided into 5 groups with 5 animals each. The groups are as follow

GROUP TREATMENT
GROUP 1 Normal control Feed + H 2 O
GROUP 2 Negative control 4% Garlic + Feed + H 2 O
GROUP 3 Positive control 20mg/kg Standard drug (Vitamin C) + 4%

Garlic + Feed + H 2 O
GROUP 4 Telfairia occidentalis 200mg/kg extract + 4% Garlic + Feed +

H2O
GROUP 5 Telfairia occidentalis 200mg/kg extract + 4% Garlic + Feed +

H2O
3.6 COLLECTION OF BLOOD SAMPLE

The animals were fasted overnight and blood was collected through cardiac puncture during

the dissecting process. Samples of blood were collected into EDTA bottles and centrifuged

at 4000rpm for 5 minutes to obtain the plasma.

3.7 Determination of ALT, AST and ALP

Serum AST, ALT and ALP activities were estimated by the method of Reitman and Frankel, using the

Randox test kit. The activity of alkaline phosphatase (ALP) was assayed using the method of Kochmar

and Moss (2016).

CHAPTER FOUR

4.0 RESULT

4.1 Aspartate Transaminase Activity

25
400 **
***
****
Plasma AST (U/L)

300

***
200
**
*
100

Figu

re 4.1: Aspartate transaminase activity of rats induced with garlic and treated with pumpkin

4.2 Alkaline Phosphatase Activity

26
800
Plasma ALP (U/L)

600 ***
***
400 ** **
*
200

Fig

ure 4.2: Alkaline phosphatase activity of rats induced with garlic and treated with pumpkin

27
4.3 Alanine Transaminase Activity

800
***
Plasma ALT (U/L)

600

400 ** **
**
*
200

Fig

ure 4.3: Alanine transaminase activity of rats induced with garlic and treated with pumpkin

28
CHAPTER FIVE

5.0 DISCUSSION ANDCONCLUSION

5.1 DISCUSSION

Figure 4.1 showed that the Aspartate transaminase concentration significantly (P>0.05) increased in

rats given garlic only when compared with control. It significantly (P>0.05) decreased in rats given

garlic + pumpkin extracts and garlic + vitamin C when compared with rats given garlic only.

Comparison between rats given pumpkin extracts only and rats given garlic + aqueous pumpkin

extracts showed that the aqueous pumpkin extract significantly reversed the oxidative stress induced by

garlic thereby having a hepatoprotective effect on the rats . This is accompanied by the study of the

long term comsumption of Telfairia occidentalis - supplemented diet by Iweala and Obidoa (2009),

which also showed a non significance of serum protein. This could be associated with the fact that

serum proteins are largely produced in the liver and Oboh (2005) suggested that there could be some

level of effects of Telfairia occidentalis - supplemented diet on rats. The concentration of circulating

triglycerides may be an early and reliable indicator of he patotoxicity in rat and so, its assessment has

been recommend as a part of the core list of liver parameters in preclinical studies. Hepatic damage is

routinely monitored by assaying for specific liver enzyme biomarkers such as AST, ALT and ALP in

the liver and serum of experimental animals (Shivaraj et al., 2009).

Fig 4.2 showed that Alkaline phosphate (P>0.05) increased in rats given garlic only when compared

with control. It significantly (P>0.05) increased in rats given garlic + aqueous pumpkin extract when

compared with rats given garlic + vitamin C. Comparison between rats given aqueous pumpkin extracts

only and rats given garlic + pumpkin extract showed that pumpkin significantly reversed the oxidative

29
stress induced by garlic thereby having a hepatoprotective effect on the rats . This is accompanied by

the study of the long term consumption of Telfairia occidentalis - supplemented diet by Iweala and

Obidoa (2009), which also showed a non significance of serum protein. This could be associated with

the fact that serum proteins are largely produced in the liver and Oboh (2005) suggested that there

could be some level of hepatoprotective effects of Telfairia occidentalis - supplemented diet on rats.

The concentration of circulating triglycerides may be an early and reliable indicator of hepatotoxicity in

rat and so, its assessment has been recommend as a part of the core list of liver parameters in preclinical

studies. Hepatic damage is routinely monitored by assaying for specific liver enzyme biomarkers such

as AST, ALT and ALP in the liver and serum of experimental animals (Shivaraj et al., 2009).

Figure 4.3 showed that the Alanine Transaminase concentration significantly (P>0.05) increased in rats

given garlic only when compared with control. It significantly (P>0.05) decreased in rats given garlic +

pumpkin extracts and garlic + vitamin C when compared with rats given garlic only. Comparison

between rats given pumpkin extracts only and rats given garlic + pumpkin extracts showed that the

aqueous pumpkin extract significantly reversed the oxidative stress induced by garlic thereby having a

hepatoprotective effect on the rats. This is accompanied by the study of the long term consumption of

Telfairia occidentalis - supplemented diet by Iweala and Obidoa (2009), which also showed a non

significance of serum protein. This could be associated with the fact that serum proteins are largely

produced in the liver and Oboh (2005) suggested that there could be some level of hepatoprotective

effects of Telfairia occidentalis - supplemented diet on rats. The concentration of circulating

triglycerides may be an early and reliable indicator of hepatotoxicity in rat and so, its assessment has

been recommend as a part of the core list of liver parameters in preclinical studies. Hepatic damage is

routinely monitored by assaying for specific liver enzyme biomarkers such as AST, ALT and ALP in

the liver and serum of experimental animals (Shivaraj et al., 2009).

30
5.2 CONCLUSION

The medicinal potential of the leaf and of Telfairia occidentalis is no longer in doubt. It is now obvious

that the plant has been proven topossess beneficial antioxidant, antidiabetic, hepatoprotective,

haematological, antiplasmodial, antimicrobial, testiculoprotective, anticancer, antiinflammatory,

anxiolytic and sedative properties. Many of these activities confirm the claims of herbal medical

practitioners, although clinical tests, using human subject, still need to be done. Many researchers have

also attributed some of the medicinal activities of the plant to the high level of antioxidant components

(such asphenolics, flavonoids, Vitamins C and E) and antioxidant property of the plant. However, the

presence of other phytochemicals such as saponins may also play valuable roles in the activities of the

plant.

31
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