CHAPTER ONE
1.0 INTRODUCTION
Plants show enormous versatility in synthesizing complex materials which have no immediate obvious growth or metabolic functions. These complex materials are referred to as secondary metabolites (Cowan, 1999; Cheesebrough, 2000). Plants secondary metabolites have recently been referred to as phytochemicals. Phytochemicals are naturally occurring and biologically active plant compounds that have potential disease inhibiting capabilities (Kasaikina, 1997). It is believed that phytochemicals may be effective in combating or preventing disease due to their antioxidant effect (Halliwell and Gutteridge, 1992; Farombi et al., 1998). Antioxidants protect other molecules from oxidation when they are exposed to free radicals and reactive oxygen species which have been implicated in the aetiology of many diseases and in food deterioration and spoilage (Halliwell and Gutteridge, 1992; Kasaikina, 1997; Farombi, 2000; Koleva et al., 2000).
Medicinal plants have been used for centuries before the advent of orthodox medicine. Leaves, flowers, stems, roots, seeds, fruit, and bark can all be constituents of herbal medicines. The medicinal values of these plants lie in their component phytochemicals, which produce definite physiological actions on the human body. The most important of these phytochemicals are alkaloids, tannins, flavonoids and phenolic compounds (Hill, 1952; Charlot, 1964).
Ocimum gratissimum Linn (Labiatae) is grown for the essential oils in its leaves and stems. Eugenol, thymol, citral, geraniol and linalool have been extracted from the oil (Sulistiarini, 1999). Essential oils from the plant have
been reported to possess an interesting spectrum of antifungal properties (Dubey et al., 2000). The antinociceptive property of the essential oil of the plant has been reported (Rabelo et al., 2003). The whole plant and the essential oil are used in traditional medicine especially in Africa and India. The essential oil is also an important insect repellant. O. gratissimum is germicidal (Nakamura et al., 1999; Pessoa et al., 2003; Holets et al., 2003) and has found wide use in toothpastes and mouth washes as well as some topical ointments. It is used as an excellent gargle for sore throats and tonsillitis. It is also used as an expectorant and a cough suppressant. The plant extract is used against gastrointestinal helminths of animals and man (Fakae, 2000; Chitwood, 2003). In addition, O. gratissimum carminative properties make it a good choice for upset stomach. It is used as an emetic and for hemorrhoids. The plant is also used for the treatment of rheumatism,
paralysis, epilepsy, high fever, diarrhea, sunstroke, influenza, gonorrhea and mental illness (Dhawan et al., 1977; Abdulrahman, 1992; Charlot, 1957; Sofowora, 1993; Sulistiarini, 1999). In addition, the plant is used as a spice and condiment in the southern part of Nigeria.
In traditional medicine the leaves of O. gratissimum have been used as a general tonic and anti-diarrhea agent and for the treatment of conjunctivitis by instilling it directly into the eyes; the leaf oil when mixed with alcohol is applied as a lotion for skin infections and taken internally for bronchitis and sinusitis (Iwalokun et al., 2003). The pulped foliage is believed to have antiseptic properties. The leaves are also chewed traditionally for all tooth and gum disorders and popularly used for treatment of diseases such as upper respiratory tract infection, diarrhea, pile, cough, fever, pneumonia, surface wound and stomach ache (Burkill, 1995; Iwalokun et al., 2003; (Cheesebrough, 2000). The leaves are also used to reduce blood glucose levels (Gill, 1992; Mohammed et al., 2007). It plays an important role in blood coagulation and renal function (James and Kuipers, 2003; Madamba et al., 2006). O. gratissimum leaves are used to prepare soups and porridge for women after delivering among the Igbo’s of Nigeria (Ijeh et al., 2004). Also the dried leaves are snuffed to alleviate headaches and fever among other uses (Effraim et al., 2001). Although conventional antibiotics have been very useful in orthodox medicine, it has been argued by many that its concomitant use with herbal extracts is not desirable as one normally antagonizes the activity of the other (Madamba et al., 2006). Considering the fact that ocimum gratissimum is used in most local dishes to achieve a variety of purposes, there is need to ascertain if its extract antagonizes or acts as a synergy when used together with conventional antibiotics (Effraim et al., 2001). In addition, despite the fact that the various extracts of ocimum gratissimum have been tested in-vitro and shown to be active against some bacterial and fungal isolates, specific strain differences supposes that a lot more strains of bacteria and fungi across other regions be tested to ascertain their in-vitro activity against this spice (Cheesebrough, (2000).
1.1 Aim of Research
This research is aimed at determining the phytochemical constituents (quantitative and qualitative) of hexanolic extract of Ocimum gratissimum (Scent leaf).
1.2 LITERATURE REVIEW
Phytochemicals are chemical compounds that occur naturally in plants. Some phytochemicals are responsible for colour and others organoleptic properties, such as the deep purple colour of blue berries and the smell of garlic (Dillard and German, 2000). The term is generally used to refer to those chemicals that may have biological significance, for example antioxidants, but are not established as essential nutrients. Scientists estimate that there may be as many as 10,000 different phytochemicals having the potential to affect diseases such as cancer, (Burkill, 1995) stroke or metabolic syndrome. Phytochemicals refers to a wide variety of compounds made by plants but is mainly used to describe those compounds that may affect human health. Phytochemicals are found in plant based foods such as fruits, vegetables, beans and grains (Nwosu and Okafor, 1995). Only a few phytochemicals have been studied closely. Some of the better known phytochemicals include beta-carotene and other carotenoids, ascorbic acid (vitamin C), flavonoids, tannins, alkaloids, saponins, terpenoids and glycosides. Phytochemicals are promoted for the prevention and treatment of many health conditions, including cancer, heart disease, diabetes and high blood pressure (Okuda et al., 1983). There is some evidence that certain phytochemicals may help prevent the formation of potential carcinogens, block the action of carcinogens on their target organs or tissue, or act on cells to suppress cancer development. Researchers are looking for specific compounds in these foods that may account for these healthful effects in humans (Pessoa et al., 2002).
1.2.1 Phytochemical Screening
Phytochemical screening is a process of identifying the presence of a certain phytochemical constituents in a plant. This process can give scientists the knowledge of the desirable constituents in plants (Silvaet al., 2005). It tells not only the presence of therapeutic agent but also gives information on the presence of commercially important compounds such as tannins ,oils, gums, precursors for the synthesis of complex compound (Thompsonet al., 1985).The major chemical substances in plants have been the alkaloids and steroidal sapogenins, however, other diverse group naturally occurring phytochemicals such as flavonoids, tannins, tepernoid and so on.
1.2.1.1 Flavonoids
Flavonoids are phenolic substances isolated from a wide range of vascular plants, with over 8000 individual compounds known. They act in plants as antioxidants, antimicrobials, photoreceptors, visual attractors, feeding repellants, and for light screening (Arai et al., 2000). Many studies have suggested that flavonoids exhibit biological activites, including antiallergenic, antiviral, anti-inflammatory, and vasodilating actions. However, most interest has been devoted to the antioxidant activity of flavonoids, which is due to their ability to reduce free radical formation and to scavenge free radicals (De-whallely et al., 1990).
The capability of flavonoids to act as antioxidants in vitro has been the subject of several studies in the past years and important structure-activity relationships of the antioxidant activity have been established. The antioxidant efficacy of flavonoids in vivo is less documented presumably because of the limited knowledge on their uptake in humans (Pessoa et al., 2002).
Most ingested flavonoids are extensively degraded to various phenolic acids, some of which still possess a radical-scavenging ability (Fellows and Axtell, 2001). Both the absorbed flavonoids and their metabolites may display an in vivo antioxidant activity, which is evidenced experimentally by the increase of the plasma antioxidant status, the sparing effect on vitamin E of erythrocyte membranes and low-density lipoproteins, and the preservation of erythrocyte membrane polyunsaturated fatty acids (Burkill, 1995).
1.2.1.2 Saponins
Saponins are phytochemicals with foamy characteristics. They are glucosides which are found in most vegetables, beans and herbs. The best sources of saponins are peas, soybeans, and some herbs with names indicating foaming properties such as soapwort, saoproot, soapbark and soapberry (Kalpana et al., 1996). Saponins consist of a polycyclic aglycones attached to one or more sugar side chains. The aglycone part, which is also called sapogenin, is either steroid or a tri-terpene. The foaming ability is caused by the combination of a hydrophobic (fat-soluble) sapogenin and a hydrophilic (water-soluble) sugar part (Li et al., 2006). Saponins have a bitter taste and some saponins are toxic and are known as sapotoxin. Commercially, saponins are extracted mainly from Yucca schidigera and Quillaja Saponaria. Saponins have many health benefits. Studies have illustrated the beneficial effects on blood cholesterol levels, cancer bone health and stimulation of the immune system (Middleton et al., 1994). Most scientific studies investigate the effect of saponins from specific plant sources and the result cannot be applied to other saponins. In cholesterol reduction, saponins bind with bile salt and cholesterol in the intestinal tract. Bile salts from small micelles with cholesterol facilitating its absorption. Saponins cause a reduction of blood cholesterol by preventing its re-absorption (Muldoon and Kritchevsky, 1996). In reduction of cancer risk, studies have shown that saponins have antitumor and anti-mutagenic activites and can lower the risk of human cancers, by preventing cancer cells from growing (Muldoon and Kritchevsky, 1996). Saponins seem to react with the cholesterol rich membranes of cancer cells, thereby limiting their growth and viability. In a previous study, it was found out that saponins may help prevent colon cancer (AOAC, 1994). Some studies have shown that saponins can cause apoptosis of leukemia cells by inducing mitotic arrest. As immune booster, plants produce saponins to fight infections by parasites. When ingested by humans, saponins also seem to help our immune system and protect against viruses and bacteria (Rabelo et al., 2003). As an anti-oxidant, the non-sugar part of saponins have also a direct antioxidant activity, which may result in other benefits such as reduced risk of cancer and heart diseases. Also the detergent properties of saponins have led to their use in shampoos, facial cleansers and cosmetic creams (Iwu, 2003; Šobotník et al., 2008).
1.2.1.3 Tannins
Tannins are non-crystallizable compounds that with water form colloidal solutions their solubility varies with their degree of polymerization) possessing an acid reaction and a sharp “puckering” taste (Funatogawa et al., 2004). They are soluble in alcohols and acetone and insoluble in chloroform, benzole, petroleum ether. The molecular weight of tannins able to transform fresh hides into leather varies from 1000 to 20000. Tannins are amorphous substances of pale yellow to slight brown colour (Iwu, 2003). Numerous tannins are optically active and tend to oxidation by air. Flobaphenes are oxidized condensed tannins. Tannins are generally extracted with hot water.
1.2.1.4 Alkaloids
Alkaloids are often potent as medicines because of their high interactivity with the human body chemistry. They have a long history as local medicines known to the native people (Lapornik et al., 2005). Alkaloids are a very diverse group of chemicals produced by vegetables which are highly reactive substances and have an effect on biological systems in low doses. They are produced by amino acids within the plant as the basic unit in the biogenesis of the true alkaloids is amino acids (Madeira et al., 2002). Most alkaloids are well defined crystalline substances which unite with acids to form salts.