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Biochemistry

Phytochemical Analysis Of Carica Papaya Leaf Oil

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Abstract

The phytochemical composition of Carica papaya leaf oil was investigated in this study using a Gas Chromatography machine fitted to a flame ionization detector. The results obtained were0.0002 µg/ml for Spartein, 3.43 µg/ml for Anthocyanin, 19.76 µg/ml for Tannin,  5.02 µg/ml for Phenol, 2.62 µg/ml for Epicatechin, 22.57 µg/ml for Lunamarine,  43.68 µg/ml for Saponin, 4.35 µg/ml for Ribalinidine, 0.29 µg/ml for Phytate, 33.11 µg/ml for Rutin, and 67.39 µg/ml for Catechin. This study has shown that C. papaya contains both beneficial phytochemicals and antinutrients

 CHAPTER ONE

INTRODUCTION/LITERATURE REVIEW

1.0      Introduction

Carica papaya is a tropical fruit, often seen in orange- red, yellow green, and yellow orange hues with a rich orange pulp. Whole plant parts, fruits, roots, bark, peel, seeds and pulp are known to have medicinal properties. It has been used for the treatment of numerous diseases like warts, corns, sinuses, eczema, cutaneous tubercles, blood pressure, dyspepsia, constipation, amenorrhoea, general debility, expel thread worms and stimulate reproductive organs (Aravind et al., 2013). It also effectively treats and improves all types of digestive and abdominal disorders (Jaime et al., 2007). Leaves of papaya, one of the plant part with numerous medicinal value has the history of steaming and eating with spinach in Asia. It has found to have significant effect on various tumor cell lines and the tea extract of leaves found to have antimalarial and antispasmodic activities. It has found to increase the appetite, ease menstrual pain and relieve nausea (Natarajan et al., 2014). Most important traditional use of leaf juice is its capability to increase white blood cells and platelets, normalizes clotting and also repairs the liver (Noriko et al., 2010). Ayurvedic literature reveals that papaya leaf extract has haemostatic properties and recent studies on ability of C. papaya leaf aqueous extract on platelet augmentation in cyclophosphamide induced thrombocytopenia rat model was studied and found significant effects (Indran et al., 2008). Pilot studies done in dengue patients with leaf juice revealed the effect of leaf juice on elevating white blood cells, platelet count and recovery without hospital admission (Hettige, 2008). This thus necessitates the need for the phytochemical profiling of the leaf oil to identify the bioactive constituents attributing significant activity (Neetu and Arun, 2014).

Hence, in the current study, an effort was taken to study the phytochemical profile of papaya leaf oil using advanced chromatographic technique, Liquid Chromatography-Flame Ionization Detection (LC-FID).

1.1      Plant oils

The use of fats and oils by man dates back to antiquity. Their chemical composition and specific properties have allowed them to find use as foods, fuels and lubricants. Their sources are numerous, encompassing plant leaf, animal, and marine sources. As it is with all matter, their usefulness to man is determined by their chemical nature; and all fats and oils have certain characteristics in common. Fats and oils are naturally occurring substances which consist predominantly of mixtures of fatty acid esters of the trihydroxy alcohol or glycerol (Nwobi  et al., 2006). Different fats and oils come about due to the fact that there are numerous fatty acids of various kinds and these can be combined in an infinite number of ways on the hydroxyl centers of glycerol. Moreover, the physical properties of fats and oils are dependent on the nature of fatty acids involved in the ester. Hence the traditional distinction of fats as solids and oils as liquids arises from the fact that due to the different chemical structures of the different fatty acids combined in the esters, the bonding forces in existence vary in strength resulting in different melting points. These differences are manifested in different chain lengths, the presence or otherwise of unsaturation as well as geometric conformations. The present emphasis on conservation and environmental friendliness has brought about renewed interest in the use of these “natural oils” for non edible purposes. Their established superiority in terms of biodegradability(Charley, 1970), when compared with mineral oils, as well as the fact that they are renewable and generally non toxic has focused attention on technologies that would enhance their usefulness as bio fuels and industrial lubricants (Honary, 2004).

1.1.1  Plant leaf oils – general properties

Plant leaf oils are obtained from oil containing leaves by different pressing methods, solvent extraction or a combination of these (Bennion, 1995). Crude oils obtained are subjected to a number of refining processes, both physical and chemical. These are detailed in various texts and articles (Fennema, 1985; Bennion, 1995). There are numerous plant oils derived from various sources. These include the popular plant oils: the foremost oilseed oils – soybean, cottonseed, pea-nuts and sunflower oils; and others such as palm oil, palm kernel oil, coconut oil, castor oil, rapeseed oil and others. They also include the less commonly known oils such as rice bran oil, tiger nut oil, patua oil, kome oil, niger seed oil, piririma oil and numerous others. Their yields, different compositions and by extension their physical and chemical properties determine their usefulness in various applications aside edible uses. Cottonseed oil was developed over a century ago as a byproduct of the cotton industry (Bennion, 1995). Its processing includes the use of hydraulic pressing, screw pressing and solvent extraction (Wolf, 1978). It is classified as polyunsaturated oil, with palmitic acid consisting 20 – 25%, stearic acid 2 – 7 %, oleic acid 18 – 30% and linoleic acid 40 – 55% (Fennema, 1985). Its primary uses are food related – as salad oil, for frying, for margarine manufacture and for manufacturing shortenings used in cakes and biscuits. Palm oil, olive oil, cottonseed oil, peanut oil, and sunflower oil amongst others are classed as Oleic – Linoleic acid oils seeing that they contain a relatively high proportion of unsaturated fatty acids, such as the monounsaturated oleic acid and the polyunsaturated linoleic acid (Dunn, 2005). They are characterized by a high ratio of polyunsaturated fatty acids to saturated fatty acids. They thus, have relatively low melting points and are liquid at room temperature (Gertz  et al., 2000). Iodine values, saponification values, specific compositions and melting points in addition to other physical properties have been determined and are widely available in the literature (Williams, 1966; Oyedeji and Oderinde, 2006). Other oils fall under various classes such as the erucic acid oils which are like the oleic linoleic acid oils except that their predominant unsaturated fatty acid is erucic acid (C22). Rapeseed and mustard seed oil are important oils in this class. Canola oil is a type of rapeseed oil with reduced erucic acid content (Applewhite, 1978). It is a stable oil used in salad dressings, margarine and shortenings. Soybean oil is an important oil with numerous increasing applications in the modern day world. It is classed as a linolenic acid oil since it contains the more highly unsaturated linolenic acid. Other oils include castor oil (a hydroxy-acid oil) which contains glycerides of ricinoleic acid (Erhan  et al, 2006). Also worthy of note is that coconut oil, which unlike most plant leaf oils is solid at room temperature due to its high proportion of saturated fatty acids (92%) particularly lauric acid (Bennion, 1995. Due to its almost homogenous composition, coconut oil has a fairly sharp melting point, unlike other fats and oils which melt over a range (Bennion, 1995). Oils from several sources are the subject of recent researches. Examples include corn oil (Sanchez, 2008); camelina sativa oil (Abramovic and Abram, 2005); Palmarosa oil and Cineole oil (Rodriguez, 2006).

1.2      Carica papaya

Plate 1: Carica papaya leaves

Carica papaya generally known as papaya, Pawpaw or Papau, Papaya Melon tree, Kapaya, Lapaya, Papyas, Papye, Tapayas, Fan mu gua, is one of the world’s most important fruit and it belongs to the small family Caricaceae. The genus carica linn is represented by four species in Nigeria, of which Carica papaya is most widely cultivated and best-known species (Mohanan  et al, 2007). Papaya is commonly known for its food and nutritional values throughout the world. Originally derived from the southern part of Mexico, Carica papaya is a perennial plant, and it is presently distributed over the whole tropical area. In particular, Carica papaya fruit circulates widely, and it is accepted as food or as a quasi-drug. The different parts of the Carica papaya plant including leaves, seeds, latex and fruit exhibited to have medicinal value. The different parts such as fruit, leaf, stem, latex, flowers obtained from Papaya are used for medicinal and various other purposes. The stem, leaf and fruit of papaya contain plenty of latex. The latex from unripe papaya fruit contains enzymes papain and chymopapain (SheikhFauziya  et al., 2013). The whole plant of papaya contain enzyme; Papain, Lycopene, Isothyocynate, important mineral; (Copper and Magnesium), Vitamins (vitamin A and vitamin C, Vitamin B6, Riboflavin, Thiamin, Vitamin K), Carbohydrates, Carotenoids, Flavonoids and other phenolic compounds are plant derived compounds with antioxidant activities by scavenging free radicals and represent a special group of nutritional supplements. Food rich in these antioxidants plays a key role in the prevention of oxidative stress based diseases.

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1.2.1  Papain

Papain breaks down the fibrin cancer cell wall and protein into amino acid form. Other than papain it also contain lycopene which highly reactive towards oxygen and free radical. Isothyocynate is effective against breast, lung, colon pancreas, prostate as well as leukemia. These enzymes capable of inhibiting both formation and development of cancer cell (SheikhFauziya  et al., 2013; Meshram  et al., 2014; Elisa et al., 2014; Godson, 2012). Fruits are rich in antioxidants such as flavonoids, anthocyanins,carotenoids, and vitamins. Experiments have shown that C. papaya have antioxidant, anthelmintic, antimutagenic, antiprotozoan, antibacterial, antifungal, antiviral, anti-inflammatory, antihypertensive, hypoglycaemic, hypolipidemic, wound healing, antitumor, free-radical scavenging, anti-sickling, neuroprotective, diuretic, abortifacient, and antifertility activities (Meshram, 2014). Epidemiological data available have shown the effectiveness of consuming fresh fruits and their juice on overcoming certain degenerative diseases including cancer, cardiovascular diseases, aging, arthritis, and others (Annegowda  et al., 2014). Medicinal plants play important roles in preventing various diseases and have received much attention from many researchers over the last few decades. Studies on the antioxidant contents of fruits and vegetables are increasing because natural antioxidant consumption has been found to be related with decreased risk for cancer and heart diseases (Zuhair Radhi  et al., 2013; Maisarah  et al., 2013). Cancer is the second leading cause of death and is becoming the leading one in old age. It has been estimated that by 2030 the number of new cancer cases will increase by 70% worldwide due to demographic changes alone. The process of cancer development is due to genetic and epigenetic alterations which lead to disruption in basic biological functions, such as cell division, differentiation, angiogenesis (Stavridi et al., 2010; Chuu  et al., 2011; Hoffman-Censits, 2013; Higano  et al., 2014. Projections indicate that the deaths over the world from cancer will rise to more than 13.1 million in 2030.The purpose of this review is to conduct a literature search to unveil the scientific evidence that C. papaya may be of use in the treatment and prevention of cancer (Thao et al., 2013). Thus, Carica papaya has different properties and has wide appications.

1.3      Chemical Composition of Carica papaya

1.3.1  Antioxidant capacity

Antioxidant activity of Carica papaya decreases the risk of oxidative damage to tissues (Mikhalchik et al., 2004; Mahmood et al., 2005). Antioxidants are the substances that can prevent or retard the oxidation of easily oxidizable materials such as fat, the functions of which are generally based on their abilities to scavenge reactive free radicals in food (Karabhari et al., 2014). The leaves of papaya have been shown to contain many active components that can increase the total antioxidant power in blood (Seigler et al., 2002; Noriko et al., 2010). Fermented papaya preparation (FPP) has defined antioxidant and immune-modulating potentials. The ability of FPP influence signaling cascades associated with cell growth and survival presents a rational for chemopreventive adjunct that can be used in combination with traditional redox based therapies that target oxidative stress in the cancer micro environment. Yoshino et al., (2009) provides ample evidence that FPP is one such antioxidant. Antioxidant functions are associated with decreased DNA damage, diminished lipid peroxidation, maintained immune function and inhibited malignant transformation of cells (Maisarah et al., 2013; Gropper et al., 2009). Result showed that there was considerable variation in the antioxidant activities where it ranges from the lowest of 58% to the highest of 91% where the orders of the antioxidant activity are as follow: α-tocopherol > unripe fruit > young leaves > ripe fruit > seed (Maisarah et al., 2013). Papaya seeds might be used as natural antioxidants (Kaibing et al., 2011).

1.3.2  Free Radical Scavenging Capacity

Papaya has many phenolic groups which may scavange free radicals. Aqueous extract of papaya leaves shows anti-oxidant activity. The fiber of papaya is able to bind cancer-causing toxins in the colon and keep them away from the healthy colon cells. These nutrients provide synergistic protection for colon cells from free radical damage to their DNA (Aravind et al., 2013; Vijay et al., 2014). Astaxanthin, zeaxanthin, and lutein are excellent lipid-soluble antioxidants that scavenge free radicals, especially in a lipid soluble environment. Carotenoids at sufficient concentrations can prevent lipid oxidationand related oxidative stress.

1.4      Role of phytochemicals in the prevention of cancer

Phytochemicals occur naturally in plants and they are responsible for colour and organoleptic properties, such as the deep purple of blue berries and red for tomatoes. Previous reports have indicated that phytoconstituents in fruits and vegetables may reduce the risk of cancer possibly due to dietary fibers, polyphenols, antioxidants and anti-inflammatory effects (Saidu et al., 2013). The papaya showed that the plants contained some phytochemical compounds which possess good antimicrobial properties on the test clinical isolates used in thestudy. The phytochemical analysis of the plant showed that the flower contain saponin, Tannin, Alkaloids and Flavonoids. This finding can be attested to the work of Sikanda  et al. (2013) who also reported similar finding and also stated the effect of these phytochemical as a good antimicrobial agent on different test organism (Ekaiko et al., 2015; Sikandar  et al., 2013). Cells in humans and other organisms are constantly exposed to a variety of oxidizing agents, some of which are necessary for life. These agents may be present in air, food, and water, or they may be produced by metabolic activity within cells. The key factor is to maintain a balance between oxidants and antioxidants to sustain optimal physiological conditions. Overproduction of oxidants can cause an imbalance, leading to oxidative stress, especially in chronic bacterial, viral, and parasitic infections (Liu, 1995). Oxidative stress can cause oxidative damage to large biomolecules such as lipids, proteins, and DNA, resulting in an increased risk for cancer and Cardio vascular disease (Liu, 1995; Ames, 1991; Ames  et al., 1993). To prevent or slow the oxidative stress induced by free radicals, sufficient amounts of antioxidants need to be consumed. Fruits, vegetables, and whole grains contain a wide variety of antioxidant compounds (phytochemicals), such as phenolics and carotenoids, and may help protect cellular systems from oxidative damage and also may lower the risk of chronic diseases (Sun et al., 2002; Chu et al., 2002; Adom et al., 2002; Wang et al., 1996; Vinson et al., 2001 and Adom et al., 2003). Strong epidemiological evidence suggests that regular consumption of fruits and vegetables can reduce cancer risk. Block et al. (1992) reviewed 200 epidemiological studies that examined the relationship between intake of fruits and vegetables and cancer of the lung, colon, breast, cervix, esophagus, oral cavity, stomach, bladder, pancreas, and ovary. In 128 of 156 dietary studies, the consumption of fruits and vegetables was found to have a significant protective effect.The risk of cancer was 2-fold higher in persons with a low intake of fruits and vegetables than in those with a high intake. Significant protection was found in 24 of 25 studies for lung cancer. Fruits were significantly protective in cancer of the esophagus, oral cavity, and larynx. Fruits and vegetable intake was protective for cancer of the pancreas and stomach in 26 of 30 studies and for colorectal and bladder cancer in 23 of 38 studies. A prospective study involving 9959 men and women in Finland showed an inverse association between the intake of flavonoids and incidence of cancer at all sites combined (Knekt et al., 1997). After a 24-y follow-up, the risk of lung cancer was reduced by 50% in the highest quartile of flavonol intake (Rui Hai,2004). Lycopene Papaya has an abundance of cancer fighting lycopene. It is a key intermediate in the biosynthesis of many important carotenoids, such as beta-carotene and xanthophylls.Men consuming lycopene-rich fruits and vegetables such as papaya, tomatoes, apricots, pink grapefruit, watermelon, and guava were 82% less likely to have prostate cancer compared to those consuming the least lycopene-rich foods (Vijay et al., 2014; Aravind et al., 2013; Karabhari Rekha Bhaskar, 2014Aswani et al., 2012). Papaya is considered a good source of lycopene, with average values ranging from 0.36 to 3.4 mg/100 g FW, being ranked number 4 of overall foods in the USDA nutrient reference database, after red guava, water melon and tomatoes (Emmy et al., 2015).

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1.4.1  Benzyl Isothicyanate

Papaya is one of the few examples known of a plant containing both glucosinolates and cyanogenic glucosides (Williams et al., 2015). It is rich in benzyl isothiocyanate (BITC) which may provide potential for use in chemoprevention of cancer. It has been suggested that the anti-carcinogenic effects of isothiocyanates are related to their capacity to induce phase II enzymes such as glutathione S-transferase, nicotinamide adenine dinucleotide phosphate and quinine reductase (Cavell et al., 2011; Nakamura et al., 2000). The glucosinolates are known to be degraded into isothiocyanates by enzymatic action of plant-specific myrosinase or intestinal microbiota in the human body (Basu et al., 2008). It suggested that the extract containing BITC and other phytochemical(s) may potentially provide the means for the treatment and prevention of selected human diseases such as cancer, and may also serve as immunoadjuvants for vaccine therapy (Emmy et al., 2015; Noriko et al., 2010).

1.4.2  Beta Carotene

The fruit is an excellent source of beta carotene that prevents damage caused by free radicals that may cause some forms of cancer (Aravind et al., 2013).

1.4.3  Saponin

Saponins can recognize cancer cells, because cancer cells have cell membranes and structures are different from normal cells. Cancer cell membranes contain more compounds such as cholesterol. Saponins can bind cholesterol contained in the membrane of cancer cells, thereby disrupting membrane permeability (Marline Nainggolan et al., 2015; Sung and Rao, 1995). Saponins also reduce the occurrence of reactive oxygen species such as H2O2 and inhibit signaling pathways phosphatidyl inositol-3 kinase which may be the reason for the prevention of damage chromosome (Marline and Kasmirul, 2015; Pawar et al., 2001).

1.4.4  Flavonoid

Flavonoid compounds inhibit cell proliferation in various human cancer cells through the inhibition of oxidative processes that can lead to cancer initiation. This mechanism is mediated decrease xanthin oxidase enzyme, Cyclooxygenase (COX) and Lipooxygenase (LOX) required in the process prooxidation thereby delaying cell cycle.Flavonoids also inhibit the expression of topoisomerase I and II enzymes that play a role in catalyzing DNAscreening. Topoisomerase enzyme inhibitor complex will stabilize DNA topoisomerase and cause cuts and damage (Marline Nainggolan and Kasmirul, 2015; Ren et al., 2003). Proteolytic enzymes: 1.4.5   Papain and chymopapain

Proteolytic enzymes have a long history of use in cancer treatment.Proteolytic enzymes have been promoted by numerous alternative cancer practitioners for many years, but most recently by those who are evaluating the benefit of proteolytic enzymes in patients with advanced pancreatic cancer in a large-scale study, funded by the National Institute of Health’s National Center for Complementary and Alternative Medicine, with collaboration from the National Cancer Institute. This larger trial is a follow-up to a smaller study that showed dramatic improvements in these patients.Once absorbed the body prevents digestion of proteins in blood and other body tissues by producing anti-proteases. The production of these anti-proteases is critical to the mechanism of action of proteolytic enzymes.These antiproteinases block the invasiveness of tumor cells as well as prevent the formation of new blood vessels (angiogenesis).Proteolytic enzymes exert a number of other interesting anticancer mechanisms including the mechanism of metastasis (the spread of cancer) and the enhancement of the immune response.The Papain enzyme is similar to pepsin, a digestive enzyme in our body. Both papain and chymopapain can help lower inflammation and improve healing from burns (Aravind et al., 2013; Michael, 2001; Gonzalez  et al., 1999).

1.4.6  Fibrin

Another useful compound not readily found in the plant kingdom is Fibrin. It reduces the risk of blood clots and improves the quality of blood cells, optimizing the ability of blood to flow through the circulatory system. Fibrin is also important in preventing stoke (Aravind et al., 2013). Anticancer activity Papaya is one of the few examples known of a plant containing both glucosinolates and cyanogenic glucosides (Emmy et al., 2015; Williams et al., 2015). It is rich in benzyl isothiocyanate (BITC) which may provide potential for use in chemoprevention of cancer. A study on the anticancer effect of Carica papaya in experimentally induced mammary tumours in rats showed that showed that administration of aqueous leaf extract of Carica papaya at a dosage of 200 mg/kg body wt showed anticancer effect (Gurudatta et al.,2015). Petroleum ether (40-600 C), Chloroform, ethyl acetate and methanol 80% extracts of C. papaya aerial parts were tested for their anticancer activity on three cancer cells TK10 (renal), UACC62 (melanoma) and MCF7 (breast) cancer cells using a Sulforhodamine B (SRB) assay. Petroleum ether of C. papaya at the concentration of 100μg/ml has shown a significant anticancer effect for MCF7 (breast) cancer cells and showed less anticancer effect for the other two cancer cells while the other extracts have mild anticancer effect on the three cancer cells (Khaled et al., 2013; Bhadane Vishal et al., 2014).Various parts of Carica papaya Linn. (CP)have been traditionally used as ethnomedicine for a number of disorders, including cancer. Study was conducted to examine the effect of aqueousextracted CP leaf fraction on the growth of various tumor cell lines and on the anti-tumor effect of human lymphocytes. Result showed significant growth inhibitory activity of the CP extract on tumor cell lines. In PBMC, the production of IL-2 and IL-4 was reduced following the addition of CP extract, whereas that ofIL- 12p40, IL-12p70, IFN- γ and TNF- α was enhanced without growth inhibition (Noriko et al., 2010; Bhadane Vishal et al., 2014). Recent research on papaya leaf tea extract has demonstrated cancer cell growth inhibition. It appears to boost the production of key signaling molecules called Th1-type cytokines, which help regulate the immune system (Aravind et al., 2013). Papaya leaf juice has been consumed by people living on the Gold Coast of Australia, withsome anecdotes of successful cases being reported for its purported anticancer activity (Noriko et al., 2010). A recent study found that papaya leaf extract could prevent growth of cancer cells, including pancreatic cancer – one of the most devastating forms of cancer (Noriko et al., 2010; Scarlett et al., 2011). This result suggests that papaya leaf may contain compounds that limit the proliferation of pancreatic cancer cells (Quan et al., 2013). The leaf tea or extract has a reputation as a tumor destroying agent (Godson et al., 2012; Ayoola et al., 2010 and Walter Last, 2008).Carica papaya extract inhibited the proliferative responses carcinoma,breast adeno carcinoma,pancreatic epithelioid carcinoma, lung adeno carcinoma, pancreatic epithelioid carcinoma andmesothelioma in a dose-dependent manner (Noriko et al., 2010). Epidemiological studies have shown that increased consumption of fruits is associated with a reduced risk of developing cancer (Rajarajeswaran et al., 2011; Block et al., 1992).

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1.5      Phytochemicals and treatment of specific conditions or diseases

The basis for the use of therapeutic drugs in modern day medicine is the history of natural product use in ancient times and in folk medicine around the world. Primitive cultures used plants as a source not only of medicines but also of toxic substances for killing animals, and for stimulants and hallucinogens used in religious rites. Traditionally, natural plant products have been the source for the search for new drugs, by pharmaceutical companies. Plant sources of herbal medicines rich in polyphenols are being studied in detail to find active molecules with healing properties. Haslam, (1998) itemized some of the medicinal plants that contain polyphenolic metabolites and the disorders for which they have been used historically. Several tests of medicinal efficacy of phytochemicals in ethnobotanicals from various indigenous cultures have been reported. Hammond  et al., (1998) studied 33 species of medicinal plants from north-eastern Peru. Tona  et al., (1998) reported studies of 45 Congolese plant extracts used to treat diarrhea in traditional medicine. Halberstein, (1997) made a descriptive survey of 18 medicinal plants on Grand Caicos Island in the West Indies. The phytochemical constituents in the plants suggest pharmacological and/or physiological efficacy in the ethnomedical treatment of various disorders. Traditional procedures to prepare the plant preparations may enhance the chemotherapeutic value of the plant derivatives, while at the same time reducing their potential toxicity. Nigerian medicinal plants were extracted and tested for in vitro anti-plasmodial active against chloroquine-sensitive and chloroquine-resistant strains of Plasmodium falciparum. Out of 16 extracts, 12 were active against the resistant strains and seven were active against the sensitive strain (Omulokoli  et al., 1997).

1.6                  Phytochemicals in clinical applications, animal studies, cells in culture or in vitro

1.6.1              Analgesic and anti-inflammatory effects in animals

The analgesic properties of phytochemical constituents isolated from a methanolic extract of C. papaya (paw-paw) fruits were tested with mice by intraperitoneal route in an acetic acid-induced abdominal constriction model (Gaertner  et al., 1999). The compounds isolated were moretenone, glutinol, b-sitosterol and stigmasterol. Glutinol and moretenone exhibited marked analgesic action, being 16- 26-fold higher in efficacy than aspirin or paracetamol. The authors suggested that the analgesic compounds in C. papaya justify, at least partially, the popular use of this plant for the treatment of urinary problems. Also, the flavone titonine (7,4′-dimethoxy-3′-hydroxy- flavone) equally isolated from the rind of C. papaya, was methylated and acetylated; (Carvalho  et al., 1999) the native compound and the methylated and acetylated compounds were evaluated for anti-inflammatory activity following intraperitoneal injection of 10mg/kg in rats using the paw edema test with carrageenin (Jones  et al., 1999). The analgesic test using the writhing test method showed a dose-dependent response. A hexane extract of C. papaya used in folk medicine for treatment of several anti-inflammatory disorders, was chromatographically fractionated (Garcia et al., 1999). The extract contained a number of terpenoid compounds. The topical anti-inflammatory activity of the hexane extract was evaluated in the mouse by auricular edema induced by 12-O-tetradecanoylphorbol acetate. In both a chronic and acute model, oedema was reduced. The identity of the active ingredient(s) was not determined, although more than one bioactive component was probably involved in the anti-inflammatory activity. Other studies have reported the anti-inflammatory activity of C. papaya against cobra venom-induced acute inflammation in mice (Philipov et al., 1998). Based on in vitro studies with fractions of various plant extracts, the investigators hypothesized that the anti-inflammatory mechanism involved the inhibition of complement activation. C. papaya species also displayed significant anti-ulcer and cytoprotective activity (Fernandez  et al., 1997). Latha  et al., (1998) tested the anti-arthritis effects of administering 100mg of an alcoholic extract from the flower of C. papaya  per kg body weight to adjuvant arthritic rats. The major histopathological changes in the hindpaws of the rats were reversed, thus showing that anti-inflammatory compounds were among the alkaloids, saponins, steroids and flavonoids in the extract.

1.6.2              Antibacterial, antiparasitic and antiviral effects

A water extract of C. papaya significantly inhibited the replication of herpes simplex virus Type 1 and Type 2 as shown by the reduction of virus induced cytopathogenic effect and protection of cells (Serkedjieva  et al., 1999). In preliminary experiments, the extract delayed the development of herpetic vesicles following infection with HSV1 in albino guinea pigs. No mechanism of action was reported, but the inhibitory effect on virus replication was reported to be related to the content of polyphenol compounds (flavonoids, catechins, a polyphenolic acid and condensed tannins). Two isoprenylflavones present in methanolic extracts from C. papaya showed intensive activity as antibacterial and cariogenic plaque-forming streptococci (Sato  et al., 1996). Among 13 flavanones tested in one study, tetrahydroxyflavanones from C. papaya and Echinosopohora koreensis actively inhibited the growth of methicillin-resistant Staphylococcus aureus (Tsuchiya  et al., 1996).

1.6.3              Antimestastatic effects of phytochemicals

1.6.3.1           Anti-mutagenic testing

Ito  et al., (1998) isolated and identified ten phytochemicals from seeds of oranges, Water melons, Sour-Sop and Paw-paw, and proposed that the ethyl acetate extract from these seeds inhibited mutagenicity induced by 7,12-dimethylbenz[1]anthracene with Salmonella typhimurium strain TM677. The extract completely inhibited DMBA-induced preneoplastic lesions in vitro in mouse mammary gland organ culture. Cell culture studies of promyelocytic leukemia cells. Resveratrol is a triphenolic stilbene present in grapes, lime, Garden egg rind, paw-paw seeds and other plants. The antioxidant and anti-inflammatory activities of resveratrol have been hypothesized to be responsible for the beneficial effects of red wine on coronary heart disease. However, the molecular mechanisms that underlie anti-tumourigenic or chemopreventive activities are unknown. Surh  et al., (1999) reported that resveratrol inhibits growth and has anti-proliferative properties in cultured human promyelocytic leukemia (HL60) cells. These effects appear to be related to the induction of apoptotic cell death by resveratrol, as determined by morphological and ultrastructural changes and other indices. Thus, this phytochemical may be protective against coronary heart disease and also have cancer therapeutic activity. Gehm  et al., (1997) reported that resveratrol increased the expression of native estrogen-regulated genes, and stimulated the proliferation of estrogen-dependent T47D breast cells. Gehm  et al., (1997) and also Calabrese, (1999) concluded that resveratrol is a phytoestrogen receptor agonist, and suggested that this finding may be relevant to the reported cardiovascular benefits of drinking wine. However, the concentrations of resveratrol necessary to elicit these effects in vitro may be unachievable in vivo by consuming natural commodities, even those extremely rich in resveratrol.

In animal and human studies Surh  et al., (1998) reviewed evidence from animal studies to support the anti-carcinogenic and anti-mutagenic effects of capsaicin, the pungent ingredient present in red pepper and ginger. In humans, curcumin, another polyphenolic phytochemical from paw-paw and Turmeric, is under preclinical trial evaluation as an anti-inflammatory and cancer preventive drug (Chan et al., 1998). In human breast cancer cells in culture, genistein has anti-proliferative effects on mitogen-stimulated growth. Plants isoflavonoid conjugates have chemopreventive activity in carcinogen-induced rat models of breast cancer (Barnes, 1997). In rats, the mechanism of the preventive action is in part dependent on its estrogenic activity, which causes rapid differentiation of cells of the mammary gland. The authors point to the importance of future studies to examine the interaction of plants isoflavonoids with other phytochemical components and to test effects in newly developed animal models of breast cancer in which specific genes have been activated or inactivated. As it is important to remember that food phytochemicals are not consumed in isolated, purified form, but in combination with other phytochemicals and food components, this type of approach should apply to studies of the health benefit effects of all food phytochemicals.


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