CHEMICAL AND NUTRITIONAL COMPOSITION OF WATER LETTUCE PLANT (Pistia stratiotes) HARVESTED FROM DIFFERENT SOURCES IN CAPE COAST

Emmanuel Nuamah

This study was aimed at assessing the nutritional and chemical compositions of water lettuce plant, to determine its suitability for use as livestock feed. The water lettuce plants were collected from three water sources: residential waste water, saline water and freshwater in the Central Region of Ghana from August to December, 2013. The water lettuce plants collected were apportioned into leaves, roots, stems and whole plants. The sections were subjected to proximate analyses whilst the heavy metal concentrations were determined using the Atomic Absorption Spectrometer and flame photometer. The data obtained from the study were analyzed using the General Linear Model (GLM) of the Analysis of Variance (ANOVA), component of the Minitab Statistical Package, (Version 15). Where significant differences were found, the means were separated using Tukey Pair Wise comparison, at 5% level of significance. From the results, it was realized that, the composition of the leaves was highly nutritious compared to the roots, stems and the whole plants as to copper, zinc, potassium and iron were all below their maximum thresholds, hence may rather serve as important micro-nutrients rather than causing harm to organisms that may feed on it. Considering the nutritive quality of the leaves, those harvested from the saline water had a high crude protein content and lower fibre content compared to those from the waste water and freshwater. The dry matter content of the leaves was low and as a result, more of the water lettuce plants need to be harvested in order to obtain a significant proportion for its potential use as livestock feed. Water lettuce obtained from the three water sources had a high level crude protein and moisture content making it a potential feed resource but may require an efficient and cost-effective method of drying if it is to be used on commercial basis. Further studies need to be conducted to determine the most efficient and cost-effective drying method, the true proteins, amino acids, anti-nutritional factors, minerals and vitamins of the water lettuce plants and also, the other metals not detected in the study, namely; mercury, arsenic, lead and magnesium.

CHAPTER ONE 1.0 INTRODUCTION 1.1 Background of the study Aquatic plants are plants that grow partly or wholly in water, and can be rooted in sediment or free floating on the water surface (Wersal and Madsen, 2012). They are also referred to as hydrophytes or macrophytes. Aquatic plants can only grow in water or in soil that is permanently saturated with water. They are therefore a common component of wetlands. Reported in the highly influential book, “The World’s Worst Weeds: Distribution and biology,” Holm et al., (1977, cited by Charudattan, 2001a) listed 10 aquatic weeds including the two most notorious weeds, water hyacinth (Eichhornia crassipes), and water lettuce (Pistia stratiotes). Holm et al., (1977), realized that, Pistia stratiotes is the most difficult aquatic plant to control in African waters. Invasive aquatic plants are noted for their explosive growth potential (Barrett, 1989) and their ability to grow from a few plants to cover hundreds of acres in a few years (Groth et al., 1996). Due to their growth potential, invasive aquatic plants are capable of blocking navigation channels, irrigation ditches and water intake pipes, impedes wildlife resources, constitute a hazard to life and can also reduce aesthetic and recreational value of water bodies, thus affecting tourism and real estate value (Catling and Dobson, 1985). In some cases, the plants have been found to increase breeding habitat for mosquitoes (Eiswerth et al., 2000). Non-native plants and animals are responsible for economic losses and control cost estimated in one analysis at $137 billion per year in the United States alone (Pimentel et al., 2000). Invasion of water lettuce to the water ecosystems of Ghana is a threat. Okali and Hall (1974) reported that, water lettuce dominates the Volta Lake, the upper bays of the rivers Afram, Pawmpawn and Dayi and also the Barikese dam near 1 Kumasi and the Weija lake in Accra (Ewer, 1996). Unlike water hyacinth, it is not completely restricted to fresh water, it demonstrates a remarkable degree of salinity tolerance with a well-defined conductivities of 1900-2000µS (Gerlach, 1996). It invades both fresh and saline water bodies in the Central region of Ghana and in certain times of the year, it covers a significant portion of rivers, lagoons and drainage channels (Personal observation). Animal nutritionists are in the search for alternative energy sources for use in livestock feed compounding. Feed cost and animal competition with humans for feed items suggest strongly that alternative energy sources be used partially or totally to replace the traditional animal feed component in livestock diets, to reduce cost and enhance cheaper meat production and therefore make available the major food items for human consumption (Ngou and Mafeni, 1983). Feed inaccessibility during the rainy season and feed shortage during the dry period remain the crucial constraints. Currently, socioeconomic, demographic and climatic changes have resulted in the reduction of pasture land, productivity and accessibility (Botoni, 2003). Since aquatic invasive plants are currently managed at considerable costs and coordination of efforts, discovering economic uses for them might serve as a better control measure. 1.2 General Objective This study therefore seeks to assess the nutritional and chemical composition of water lettuce, to determine it suitability for use as livestock feed. 1.3 Specific Objectives 1. To determine the proximate composition of water lettuce (Pistia stratiotes) plant from different water bodies in Cape Coast 2. To determine the presence and levels of some heavy metals in the water lettuce plants 2 3. To assess the heavy metal concentration of water bodies in which the water lettuce plants were harvested. 3 CHAPTER TWO 2.0 LITERATURE REVIEW 2.1 ORIGIN AND DISTRIBUTION OF WATER LETTUCE (Pistia stratiotes) The origin of water lettuce (Pistia stratiotes) is still speculative, being most probably from South America (Cordo et al., 1981), as most of its insect natural enemies are on record from South America (Bennett, 1975). Water lettuce is one of the most widely distributed of all floating macrophytes, occurring in all continents, except Antarctica, being now widespread throughout the tropics and subtropics (Holm et al., 1977). The initial spread is suspected to have taken place through ballast water in ships from South America (Labrada and Fornasari, 2002). Water lettuce is widespread in tropical Africa and is reported as troublesome in the Upper Nile, Zambia, Kenya, Zimbabwe and the littoral regions of Angola and Mozambique. In South Africa, it is recorded almost entirely in the subtropical regions (Henderson and Cilliers, 1991), and it has been reported in West Africa: Benin, Côte d’Ivoire, Nigeria and Senegal (Ajuonu and Neuenschwander, 2003). The weed is common on many waters in West Africa (Pettet and Pettet, 1970). However, most botanists and researchers consider it foreign to Africa. The possibility that it is indigenous in numerous areas is enhanced by its ancient medicinal use. Although exotic aquatic weeds have been reported to be present in Africa since the end of the nineteenth century, they started colonizing African freshwater bodies during the early 1950s and rapidly spread in many countries (Tackholm and Drar, 1950 and Mitchell et al., 1990). The growth of invasive aquatic weeds like water lettuce is extremely fast and this allows them to develop huge infestations in areas where they had not been reported only a few years earlier (Labrada and Fornasari, 2002). 2.2 TAXONOMY, MORPHOLOGY AND BIOLOGY OF Pistia stratiotes Water lettuce is a perennial herb of the aroid family (Araceae), a monotypic genus in the subfamily Aroideae (Grayum, 1990). Water lettuce is the only species 4 in the genus Pistia and it is not likely to be confused with any other species. There are several indigenous species in the same family, including arum lily flowers, but all are terrestrial herbaceous plants with tuberous roots (Henderson and Cilliers, 1991). Water lettuce plants consist of a rosette of leaves with a tuft of long, numerous fibrous roots, resembling floating lettuce. The leaves which range from 2 to 35 cm long are pale yellowish-green, closely overlapping, ribbed, with many longitudinal veins radiating from the base, and softly hairy on both surfaces (Henderson and Cilliers, 2002). The rosettes occur singly or in groups connected by short stolons. The flowers are inconspicuous pale green spathes near the center of the rosette. Each spathe is constricted near the middle, with a whorl of male flowers above and a single female flower below the constriction. The fruits are many-seeded green berries, and the mature seed coat is thick, golden brown, and wrinkled (Dray and Center, 2002). The plant reproduces vegetatively through the formation of stolons and daughter plants (Forno and Julien, 2000). The role of sexual reproduction is considered less important than that of vegetative reproduction, although seed germination is an important factor in the dynamics of water lettuce populations (Dray and Center, 1989). P. stratiotes reportedly does not produce fruits and seeds in the United States (Dray and Center, 1989). However, seed production occurs in Africa (Holm et al., 1977), India (Mitra, 1966), South America (Da Silva, 1981), and South-East Asia (Bua-ngam and Mercado, 1975). Water lettuce seeds can remain dormant for months, withstand freezing and drought, and still germinate when favorable conditions become prevalent (Pieterse et al., 1981). Studies in Philippine rice fields (Bua-ngam and Mercado, 1975) indicate that seed production plays an important role in the re-establishing of populations after catastrophic destruction. This evidence has important implications for aquatic weed management schemes. Herbicide application is very effective at eliminating infestations, particularly in waterways closed to re-introduction of the weed from other areas. However, viable seeds on benthic sediments will germinate under 5 favourable conditions. These seedlings provide an initial stock which, through vegetative propagation, will reinfest a water body (Dray and Center, 1989). Growth varies seasonally in density of rosettes, from less than 100 to over 1000 per m 2 in south Florida (Dewald and Lounibos, 1990). There have been numerous reports of Pistia stratiotes ability as a bioaccumulator (Qian et al., 1999; Cecal et al., (2002, cited by Azzam, 2006). Maine et al., (2001, cited by Azzam, 2006) reported that Pistia stratiotes has superior performance and higher average relative growth rate when bred in heavy metal induced stress environment compared to Eichhornia crassipes and Hydromistia stolonifera. 2.3 NUTRIENT COMPOSITION OF WATER LETTUCE According to the study “Nutritive value of water lettuce (Pistia stratiotes) and its possibility in animal feed”, an analysis of the nutritive value of water lettuce (Pistia stratiotes) as animal forage conducted by Rodríguez et al. (2000) reported that, the proximate, structural and mineral composition of water lettuce (Pistia stratiotes) were found to be 8.62% crude protein, 1.32% total N, 1.16% ether extract, 46.62% N-free extract, 21.12% ash and 19.13% crude fibre. The mineral contents are 5.56% K, 3.24% Ca, 1.00% Mg, 0.26% P, 0.61% Na, 0.26% Fe, 0.12% Mn, 42 ppm Cu and 181 ppm Zn. According to the study “Composition of Indian Aquatic Plants in relation to utilization as animal forage”, Banerjee and Mata (1990) reported that, water lettuce has 5.3% dry matter, 20.5% crude protein, 17.0% ash, 3.8% crude fat, 19.1% crude fibre and 39.6% nitrogen free extract. Ayoade et al. (1982), reported from the research, “Trial of Pistia stratiotes L. as animal feed”, Pistia stratiotes contains 6.9% organic matter, 30.1% ash, 13.9% crude protein, 21.9% crude fibre, 3.2% ether extract and 27.8% nitrogen free extract. Ghani et al., (2010) reported from the work “Toxic Effect of Heavy Metals on Plant Growth”, that, heavy metals caused significant decreases in growth and protein content and 6 the higher the concentration of heavy metal in the soil, the greater was the toxic effect on the plant. 2. 4 HEAVY METALS The term heavy metal refers to any metallic chemical element that has a relatively high density and is toxic, highly toxic or poisonous at low concentrations (Bryan & Langston, 1992). Heavy metal is a general collective term which applies to the group of metals and metalloids with an atomic density greater than 4g/cm³ (Duffus, 2002). They are defined by the United Nations Economic Commission for Europe (UNECE) as “those metals or, in some cases, metalloids which are stable and have a density greater than 4.5 g/cm3 and their compounds” (UNECE, 1998). Alloway (1995a) defines heavy metals as “elements which have an atomic density greater than 6 g/cm3.” The term, Potential Toxic Elements (PTEs), are sometimes used for this group of metals (Alloway, 1995). Although it is a loosely defined term, heavy metals are widely recognized and usually applies to the widespread contaminants of terrestrial and freshwater ecosystems. Examples of heavy metals are cadmium, chromium, copper, mercury, lead, zinc, arsenic, boron and the platinum group of metals, which comprises Platinum, Palladium, Rhodium, Ruthenium, Osmium, and Iridium. Unlike almost all organic pollutants, such as organochlorines, heavy metals are elements which occur naturally in the Earth’s crust. They are therefore found naturally in soils and rocks with a subsequent range of natural background concentrations in soils, sediments, waters and organisms. Anthropogenic releases can give rise to higher concentrations of the metals relative to the normal background values. The most important anthropogenic releases of heavy metals to the environment come from metalliferous mining and smelting, agricultural materials (pesticides and fertilisers), irrigation and application of sewage water and sludge, fossil fuel combustion and metallurgical industries (Alloway, 1995b). Heavy metals make 7 significant contribution to environmental pollution as a result of anthropogenic activities such as mining, energy- and fuel production, power transmission, intensive agricultural practices, sludge and industrial effluent dumping and military operations (Foy et al., 1978; Salt et al., 1998; Orcutt and Nilsen, 2000; Cseh (2002, cited by Azzam, 2006); Pilon- Smits, 2005). As they are elements, they cannot be broken down; therefore heavy metals will persist in the environment. Unlike many organic pollutants, which eventually degrade to carbon dioxide and water, heavy metals will tend to accumulate in the environment, especially in lake, estuarine or marine sediments and can be transported from one environment compartment to another (Duffus, 2002). Whether the source of heavy metals is natural or anthropogenic, the concentrations in terrestrial and aquatic organisms are determined by the size of the source and adsorption and/or precipitation in soils and sediments. The extent of adsorption depends on the metal, the absorbent, the physico-chemical characteristics of the environment (e.g. pH, water hardness and redox potential) and the concentrations of other metals and complex chemicals present in the soil water, river or lake. Heavy metals also accumulate in organisms as a result of direct uptake from the surroundings across the body wall, from respiration and from food. Uptake via food is most important in terrestrial organisms and it may also be important in the aquatic environment. Dietary uptake can include heavy metals adsorbed on particulates present on the surface of leaves, which have not been absorbed by the plant (Duffus, 2002). Heavy metals are dangerous because they tend to bioaccumulate. Bioaccumulation means an increase in the concentration of a chemical in a biological organism over time, compared to the chemical’s concentration in the environment. Compounds accumulate in living things anytime they are taken up and stored faster than metabolized or excreted (Wild, 1993). 8 2.4.1 Potential Sources of Heavy Metals in the Environment The amounts of most heavy metals deposited to the surface of the Earth by man are many times greater than depositions from natural background sources. Combustion processes are the most important sources of heavy metals, particularly, power generation, smelting, incineration and the internal combustion engine. Combustion processes cause the released or emission of volatile elements such as arsenic (As), cadmium (Cd), lead (Pb) and mercury (Hg). (Hutton and Symon 1986; Battarbee et al., 1988; Nriagu and Pacyna 1988; Nriagu 1989). The functioning of natural biological systems is increasingly affected by human activities and it is difficult to find a river or other water body whose natural regime has not been modified by man’s activities. An increase in urbanization and industrial activities, and higher exploitation of cultivable land has brought about a huge increase in the quantity of discharges and wide diversification in types of pollutants that reach rivers and other aquatic environments. Many African countries depend on agriculture to boost their economy, thus pesticides are likely to represent an important source of xenobiotic in contaminated rivers. The ultimate sink for many of these contaminants in the aquatic environment is due to discharges or to hydrologic and atmospheric processes (Lagadic et al., 2000). 2.4.2 Beneficial Heavy Metals In small quantities, certain heavy metals are nutritionally essential for a healthy life. Some of these are referred to as trace elements (e.g. copper, iron, manganese and zinc). Some form of these, are commonly found naturally in foodstuffs, in fruits and vegetables, and in commercially available multivitamin products (Brown et al., 2004). Diagnostic medical applications include direct injection of gallium during radiological procedures, dosing with chromium in parental nutrition mixtures, and the use of lead radiation shield around x-ray equipment (Kennish, 1992b). Heavy metals are also common in industrial applications in the manufacture of pesticides, 9 batteries, alloys, and electroplated metal parts, textile dyes, steel and so forth (NIOSH, 1999). Many of these products are in homes and actually enhance quality of life when properly used. 2.4.3 EFFECTS OF HEAVY METALS 2.4.3.1 Toxicity to Plants Plants’ responses to heavy metal toxicity include leaf discoloration, chlorosis, necrosis, dwarfism, gigantism, leaf expansion inhibition and root growth inhibition (Othman, 2001). Within plant cells, excessive amount of certain heavy metals can modify the permeability of the plasma membrane, causing leakage of ions and solutes. Several metals like copper have a high affinity for sulphydryl and carboxyl groups, which lead to a decrease in the plasma lemma ATP-ase activity. In addition, some cell components can be damaged by free radicals formed by metal participation (Moolenaar, 1998). Excessive heavy metal contamination also has many negative effects on chlorophyll. Some metals have a high affinity for sulphydryl groups and can cause the inhibition of enzyme and chlorophyll synthesis (Bargagli, 1998). Displacements of these essential elements could decrease the levels of chlorophyll content (Rozema and Verkleij, 1991). 2.4.3.2 Toxicity to Humans The toxicity of a metal is usually defined in terms of the concentration required to cause an acute response (usually death) or a sub-lethal response (Smith, 1986). Predicting the consequences of metal exposure on living organisms is complicated because metals may be essential or non-essential. Very low concentrations of essential metals can be as harmful as high concentrations. Nonessential metals display more conventional toxicity curves, showing a sigmoidal increase in proportion of exposed individuals dying with an increase in metal concentration (Newman and Clements, 2008). Understanding this dichotomy of 10 essential and non-essential metal concentration–effect can still be insufficient for sound prediction of metal effects. For example, WHO (1996, cited by Azzam, 2006) reported that acute exposure of copper and zinc can cause fever, vomiting, nausea, stomach cramps and diarrhea. In a long term, exposure to heavy metals such as copper, cadmium, chromium, zinc, mercury and lead caused carcinogenic effects (Pyatt et al., 2005). The main characteristics of chronic lead toxicity are sterility both in males and females, and abnormal fetal development Johnson (1998, cited by Azzam, 2006). Lead has been reported in inhibiting heme synthesis and in decreasing red cell survival in carcinogenicity and nucleic acid destabilization (Pyatt et al., 2005). According to Godt et al., (2006), the main characteristics of cadmium toxic effects include respiratory impacts (lungs edema, pneumonitis and destruction of the mucous membrane), reproductive effects (testicular necrosis, estrogen-like effects and affecting steroid hormone synthesis), Kidney damage (proteinuria, kidney stones, glomerular and tubular damage) and skeletal system effects (loss of bone density, mineralization and itai-itai disease). The bioactivity of some non-essential elements can also be affected by another element. For example, mercury toxicity is lowered if sufficient concentrations of selenium are also present (Newman and Clements, 2008). 2.4.3.3 Effects on the Environment Heavy metals have long been recognized as one of the most important pollutants in coastal waters (Johnson et al., 2000). It is caused by their toxicity and capacity to accumulate in marine organisms. However at low concentrations, some are essential in many physiological processes for plant, animal and human health (Basile et al., 2005). Heavy metals are a long-term problem, unlike organic pollutant in which heavy metals are not biodegradable and will enter the food chain through a number of pathways causing progressive toxic actions due to their accumulation in different organs during a life span and long term exposure to contaminated environments 11 (Machynlleth, 1998 cited by (Azzam, 2006). Austin (1998, cited by Azzam, 2006) also mention about the capability of vertebrates and invertebrate to accumulate heavy metals from aquatic environment. For instance, cadmium, copper, lead and zinc have been detected using atomic absorbance spectrometry in gill, muscle, vertebrate and viscera of rabbitfish (Siganus oramin) from polluted waters in Hong Kong (Zhou et al., 1998). 2.5 SOME HEAVY METALS AND THEIR MAXIMUM THRESHOLDS 2.5.1 Iron Iron, one of the most abundant metals on Earth, is essential to most life forms and to normal human physiology. Iron is an integral part of many proteins and enzymes that maintain good health (Institute of Medicine, 2001). In humans, iron is an essential component of proteins involved in oxygen transport (Dallman, 1986). It is also essential for the regulation of cell growth and differentiation (Bothwell, 1979, Andrews, 1986). A deficiency of iron limits oxygen delivery to cells, resulting in fatigue, poor work performance, and decreased immunity (Institute of Medicine, 2001, Bhaskaram, 2001). On the other hand, excess amounts of iron in man can result in toxicity and even death (Corbett, 1995). There is considerable potential for iron toxicity because very little iron is excreted from the body. Thus, iron can accumulate in body tissues and organs when normal storage sites are full. For example, people with hemachromatosis are at risk of developing iron toxicity because of their high iron stores. Symptoms of Alzheimer‟s and Parkinson‟s disease may also be iron-related (Corbett, 1995). According to the Agricultural Research Council Party, the maximum tolerable level of iron is 500mg Fe/kg diet or 500ppm. 12 2.5.2 Copper Copper can be released into the environment by both natural sources and human activities. Examples of natural sources are wind-blown dust, decaying vegetation, forest fires and sea spray (Cuzzocrea et al., 2003). Copper can easily form complexes with organic compounds, which are quite stable in the environment (Zhou et al., 1998). It is estimated that, around 3.2 million tonnes of copper was released to the environment from 1910-1990 (WHO, 1996). Copper is essential to life and is found in all body tissues. Copper deficiency can lead to a variety of abnormalities, including anemia, skeletal defects, degeneration of the nervous system, reproductive failure, pronounced cardiovascular lesions, elevated cholesterol, impaired immunity, and defects in the pigmentation and structure of the hair (Smith, 1974; NRC, 1980 cited by Azzam, 2006). However, at high concentrations, copper is also capable of causing toxic effects. The ingestion of excess copper can cause gastrointestinal problems and the exacerbation of vibriosis in human (Austin, 1998 cited by Azzam, 2006); Siegel, 1998). Copper is also reported to be highly toxic against sperms (Wong et al., 2001) and may affect spermatogenesis with regard to motility, production, maturation and fertilizing capacity of the spermatozoa (Skandhan, 1992). According to Council Directive 80/778/EEC, the toxic level of copper is 25ppm. 2.5.3 Zinc Zinc is widely used in modern society, most commonly to coat or galvanize iron to prevent corrosion. It is also mixed with other metals to form alloys such as brass. Particles released from vehicle tires and brake linings are a major source of zinc in the environment (WHO, 2001). Zinc is an essential nutrient for the human body and has an importance for health (Hotz et al., 2003). Zinc acts as a catalytic or structural component in many enzymes that are involved in energy metabolism and in transcription and translation of RNA (Moolenaar, 1998). 13 However, like other metals, it can be toxic in high concentrations (ANZECC, 2000). Although uncommon, gastrointestinal distress and diarrhea have been reported following ingestion of beverages stored in galvanized cans or prepared using galvanized utensils (WHO, 2001). Other symptoms of zinc toxicity are slow reflexes, paralyzation of extremities, anaemia, metabolic disorder, terratogenic effects and increased mortality (Klaassen, 1996). According to the European Commission Regulation (ECR) 237790 (consolidated), the maximum permissible level of zinc is 50mg/kg or 50ppm. 2.5.4 Potassium Potassium has been recognized as an essential nutrient in animal nutrition since its importance was pointed out by Ringer, S. (1883). It occurs naturally in the form of several mineral salts but does not occur as metallic potassium. Potassium in foods is associated with salts of weak organic acids. Young animals will fail to grow and will die within a few days when the diet is extremely deﬁcient in potassium. Potassium deficiency can develop as a consequence of increasing losses from the gastrointestinal tract and kidneys, e.g. during prolonged diarrhea or vomiting, and in connection with use of laxatives or diuretics. Potassium deficiency due to low dietary intake only is very uncommon, due to the widespread occurrence of potassium in foods. Symptoms of potassium deficiency are associated with disturbed cell membrane function and include muscle weakness, disturbances in heart function, which can lead to arrhythmia and heart seizure. Mental disturbances, e.g. depression and confusion, can also develop (EFSA, 2006). According to Chemical Analysis of Ecological Material (1974), the concentration range of sodium is 0.5-10ppm. 2.5.5 Sodium Sodium is an essential nutrient involved in fluid and electrolyte balance and is required for normal cellular function. The major adverse effect of increased sodium 14 intake is elevated blood pressure. Higher blood pressure is an acknowledged risk factor for ischaemic heart disease, stroke and renal disease which are major causes of morbidity and mortality. The effect of sodium on blood pressure is linked to that of chloride. The regulation of total body content is closely related to the regulation of total body potassium, the main intracellular cation, and the regulation of total body water. The membrane bound sodium-potassium pump (the sodiumpotassium-activated adenosine triphosphate Na+-K+ ATPase) plays a fundamental role in maintaining the partitioning of sodium and potassium between the extracellular and intracellular compartments respectively, and the energy required for this process represents a significant component of the metabolic rate. (EFSA, 2006). According to Chemical Analysis of Ecological Material (1974), the concentration range of sodium is 2-100ppm. 2.6 NUTRIENT COMPOSITION OF FEEDS/FEED COMPONENTS USED FOR MOST LIVESTOCK’S 2.6.1 Maize Maize, like the other cereals grains, has certain limitations as a food for farm animals. Though an excellent source of digestible energy, it is low in protein and the proteins present are of poor quality. Maize contains about 730g starch/kg DM which is very low in fibre and has a high metabolizable energy value. The starch in maize is more slowly digested in the rumen than that of other grains, and at high levels of feeding a proportion of the starch passes into the small intestine, where it is digested and absorbed as glucose. The oil content of maize varies from 40-60g/kg DM and is high in linoleic acid. The crude protein content of maize is very variable and generally ranges from about 90 to 140g/kg DM, although varieties have been developed recently containing even higher content (McDonald et al., 2002). 15 2.6.2 Millet The composition of millet is very variable, the crude protein content being generally within the range 100-120g/kg DM, the other extract 20-50g/kg DM and the crude fibre 20-90g/kg DM. Millet has a nutritive value very similar to that of oats and contains a high content of indigestible fibre oweing to the presence of hull (McDonald et al., 2002). 2.6.3 Rice The two main by-products obtained from rice milling, namely hulls and rice meal. The hulls are high in fibre content and can contain up to 210g/kg DM silica. Rice meal or rice bran comprises the pericarp, the aleurone layer, the germ and some of the endosperm and is valuable product containing about 120-145g/kg DM and 110-180g/kg DM (McDonald et al., 2002). Table 2.1: Nutrient Composition of Feeds/Feed components used for most Livestock’s Feedstuff class Dry Matter (%) CP (% DM) ME (MJ/Kg DM) Straw stovers 88-92 3-4 5.5-7.5 Cereals 82-91 9-11 12-14 Grasses 20 10-22 9-12 Oilseed cakes 89-91 22-50 12-14 Green Legumes 15-27 17-24 10-12 Source: McDonald et al., 2002 ` 16 CHAPTER THREE 3.0 MATERIAL AND METHODS 3.1 Study Area The study was conducted in 3 surrounding communities of the University of Cape Coast, namely: Apewosika, Bakaano and Kwaprow. The water sources in these communities were selected based on the level of invasion of the water bodies by the water lettuce plant and their proximity with the University of Cape Coast. The water source at Apewosika was a channel which was made up of waste water from different areas. The water source at Bakaano was the Fosu lagoon which had a high level of salinity due to its merging with the sea and the water source at Kwaprow was a freshwater which was mostly used by the people of the community for domestic purpose. The water lettuce samples were collected from these location to demonstrate its survivability in fresh water bodies as well as its tolerance in saline water waste (different water conditions). The water body at Bakaano is saline whiles that of kakum is fresh water. 3.2 Description of the study area The study area and its surrounding are shown in Fig. 1. It is bounded on the south by the Gulf of Guinea, on the west by the Komenda – Edina – Aguafo Abrem District and on the north by the Twifo Hemang lower Denkyira District. It has a total land area of 9826 km2 (Faanu et al., 2011). 17 Res. J. Environ. Earth Sci., 3(3): 269-274, 2011 Fig. 1: Map of the Study area showing the University of Cape Coast and its surroundings Geologically, the area is dominated by batholiths and is generally undulating with steep slopes. There are valleys of various streams between the hills, with Kakum being the largest stream. The minor streams end in the Fosu lagoon at Bakaano. The metropolis has double maxima rainfall and the major rainy season occur between May to November and minor rainy season falls between November January. It is a humid area with mean monthly relative humidity varying between 85 and 99%. The sea breeze has moderating effect on the local climate. The soil profile shows a top soil of about 0.33mm Faanu et al., 2011). 3.3 Sampling and Sample Preparation for Laboratory Analysis The samples (water lettuce and their respective water samples) were collected for analysis from August to December. The water lettuce from the water sources were harvested with a scoop net into a transparent polythene bags and the 18 water samples were collected into plastic bottles. The samples were coded as A, B, and K: A= Sample from residential waste water B= Sample from Fosu Lagoon K= Sample from Kakum river These samples were transported to the Animal Science Department (School of Agriculture) Laboratory (University of Cape Coast). At the laboratory, some of the water lettuce were randomly selected and apportioned in to leaves, stem, root and whole plant; whiles the water samples were stored in freezer for subsequent analysis of moisture content, dry matter, ash, crude protein, crude fibre and the level of heavy metals in both the water lettuce and water samples. 3.4 Laboratory Analysis of Feed Samples Analysis for nutritive quality of the feed sample were conducted following the procedure and methods described by (AOAC, 1990). 3.4.1 Analysis for Moisture and Dry Matter Content The weight of the apportioned water lettuce samples were taken with an electronic balance (ADP 2100), and put into coded envelopes. The samples were then dried in an oven at a temperature of 600C for 48 hours until a constant weight was obtained. After drying, the samples were allowed to cool for 30 minutes in a desiccator and the dried weight was taken with the balance. After the weighing of the cooled samples, percentage moisture and percentage dry matter were determined using the formula below: 19 Percentage Moisture = � � ℎ Percentage Dry matter = 3.4.2 ℎ − � � ℎ � � ℎ � � � ℎ � ℎ ℎ � ×100 ×100 ℎ Analysis for Ash Content (total minerals and Organic Matter) The ash content which represents the total minerals, was estimated by burning away the organic material. About 0.5g of the sample was weighed using a balance (ae ADAM AAA 250LE) into a previously dried and weighed crucible and the reading were recorded. The sample was then charred over a hot plate (Kjeldahl apparatus) for 4 hours. After the charring of the sample, it was then cooled for 30 minutes in a desiccator and weighed. The percentage ash content was then determined using the formula: Percentage Ash Content = 3.4.3 � � ℎ ℎ � � ℎ ×100 Analysis for Crude Fibre About 0.4g of the sample was weighed into a conical flask using a balance (ae ADAM AAA 250LE) and 100ml of the acid was added and boiled for 30 minutes on a hot plate (Kjeldahl apparatus). After boiling, the remnant in the conical flask was filtered using Buckner funnel and suction pump. The residue was then transferred back into the conical flask and 100ml of sodium hydroxide was added and boiled on a hot plate for 30 minutes. It was then filtered using the Buckner funnel and suction pump and what was obtained was the indigestible portion. The residue was then dried at a temperature of 600C and afterwards, cooled in a desiccator for 30 minutes. The weight of the residue plus the crucible was obtained and recorded. The residue was then charred on the Kjeldahl apparatus (hot plate) for 3 hours and cooled in desiccator for 30 minutes. The sample was then 20 weighed and recorded. The percentage crude fibre was then determined using the formula: Percentage Crude Fibre = �� ℎ ×100 Where Fibre content = Weight of residue (ash + fibre + crucible) – Weight of (ash + crucible) 3.4.4 Analysis for Crude Protein Digestion About 0.2g of the sample was weighed into a labelled Kjeldahl flask. About 4.5ml of prepared digestion mixture was added to each of the flasks containing the sample. The samples were digested at a temperature of 3600C for 2 hours after which the solution became colorless, and was allowed to cool. A blank digestion was also carried out. About 20ml of distilled water was added to each flask and mixed thoroughly to dissolve and allowed to cool. It was then transferred into a 100ml volumetric flask and topped –up with distilled water to the mark. Distillation and Titration The steam distillation apparatus was set up and steam was passed through for 20 minutes. After the flushing out of the apparatus, 100ml conical flask containing 5ml of boric acid indicator solution was placed under the condenser of the distillation apparatus. About 25 ml of the digested sample was then pipetted into a flask and transferred into reaction chamber and 10 ml of the alkali was added to the mixture. Distillation commenced immediately and 50 ml of distillate was collected. The distillate was titrated with 0.1 M H2SO4 to a pink end point. The percentage nitrogen (%N) and protein (% P) were calculated using the formula: % N = (T-B) × M × % CP = %N × F � 4. 7 � ℎ ×5 × 21 Where T= Sample titre value B= Blank titre value = 0.2ml M= Molarity of acid = 4 M 3.4.5 Analysis for Ether Extract (Fat) About 5g of the sample was weighed into thimble and plugged with cotton wool. It was transferred into the soxhlet extractor by dropping the whole thimble into it. The soxhlet was allowed to sit on the round bottom flask and the ether was poured and allowed to siphon. This was done twice and connected to a condenser. The set-up was allowed to stand for 4 hours and the flask was removed and put into a Gallenkamp oven at 500C and heated for the ether to dry off. The weight was taken and the fat content was calculated from the formula: % Fat = � � × 100 3.5 Determination of Copper, Zinc, Potassium, Sodium and Iron in samples and water from the study site The preparation of sample solutions suitable for elemental analysis of copper, zinc, potassium, sodium and iron involved an oxidation process which was necessary for the destruction of the organic matter, through acid oxidation before a complete elemental analysis was carried out. 3.5.1 Determination of Potassium and Sodium using Flame Photometer Potassium and sodium in the digested samples were determined using a flame photometer. In the determination, the following working standards of both K and Na were prepared: 0, 2,4,6,8 and 10ppm. The working standards as well as the sample solutions were aspirated individually into the flame photometer and their emissions (readings) recorded. A calibration curve was plotted using the 22 concentrations and emissions of the working standards. The concentrations of the sample solutions were extrapolated from the standard curve using their emissions. CALCULATION = %K & Na/g Sa C e e × e Where C= Concentration of the sample (Stewarte et. al., 1974) 3.5.2 Determination of Iron, Copper and Zinc using Atomic Absorption Spectrophotometer Standard solutions of 1, 2 and 5�g/mL solutions of Fe, Cu and Zn were prepared. The standard solutions were aspirated into the atomic absorption spectrophotometer (AAs) and the respective calibration curves were plotted on the AAS. As the sample solutions were aspirated their respective concentrations were provided. CALCULATIONS % Fe = � % Cu= � % Zn = � × � × � × � × × × � ℎ � ℎ × �� ℎ 23 �� 3.6 Data Analysis The data obtained from the study were analyzed using the General Linear Model (GLM) of the Analysis of variance (ANOVA), components of the Minitab Statistical Package, Version 15 (Minitab, 2007). Where significant differences were found, the means were separated using Tukey Pair Wise comparison, at 5% level of significance. 24 CHAPTER FOUR 4.0 RESULTS AND DISCUSSION Table 4.1 presents the Proximate Composition of the leaves of water lettuce plant Table 4.1: Proximate Composition of the leaves of water lettuce plant Composition Waste Lagoon Freshwater SED Significance water Moisture 93.19b 96.16a 92.13b 0.4454 *** Dry Matter 6.81b 3.84c 7.87a 0.4517 *** Crude Protein 22.31b 30.34a 16.16c 0.6978 *** Fibre 15.96a 12.10b 15.10a 0.2995 *** Fat 2.27b 4.35a 2.68b 0.1994 *** Ash 10.34a 4.28b 10.83a 0.6995 *** SED= Standard error of difference, Means in the same row with different superscripts are significant, ***= Significant (p<0.001) The analysis of water lettuce sample from all the water bodies were significantly higher among leaves, roots, and stems harvested from waste water, freshwater and the saline water. Ash content of the stems on the other hand had no significant difference among the plants from all the water bodies. The levels of heavy metals detected in the study were also significantly higher among the plants from all the water bodies except sodium which had no significant difference. The moisture content of the leaves had a high significantly difference among the treatments. The moisture content of the leaves of plants obtained from the saline water had a higher significant difference to that of the freshwater and waste water. The leaves of plants harvested from waste water and freshwater were slightly lower than earlier findings by Banerjee and Mata (1990) who reported 94% moisture content. The moisture content of leaves of plants harvested from the saline water were slightly higher than that of the earlier findings by Banerjee and Mata (1990) who reported 94% moisture content. The leaves of plants from the waste 25 water was similar to earlier findings of Ayoade et al., (1982) who reported 93.1% moisture content whereas that of the freshwater was slightly lower than earlier findings of Ayoade et al., (1982). The variation in the findings (present and previous) may be due to the time and method of harvesting, climate, location, pollution and the type and duration of storage (Dangour et al., 2009). It may also be due to irradiance, nutrient and water availability, temperature, salinity and compaction (Poorter et al., 2009). The dry matter (DM) had a higher significance (p<0.001) among all the water bodies. Generally, the leaves harvested from the freshwater had the highest DM followed by that harvested from the waste water and those of the saline water. Compared to that of literature, DM of leaves harvested from the waste water and freshwater were slightly higher than earlier works by Banerjee and Mata (1990) who reported 5.3% DM whilst those harvested from the saline water was slightly lower than earlier findings by Banerjee and Mata (1990) who reported 5.3% DM. This may be due to similar reasons pointed out for the moisture content of the leaves. The crude protein (CP) ranged between 16.16% and 30.34%. Generally, the CP content obtained in the leaves were statistically different among all the water bodies. The present findings among all the water bodies were higher than earlier work of Ayoade et al., (1982) who reported 13.9% CP. The leaves obtained from the waste and saline water were higher than earlier findings by Banerjee and Mata (1990) who reported 20.5% and that of the freshwater was slightly lower to earlier findings by Banerjee and Mata (1990). The variations in the crude protein content may be due to environmental difference (temperature, rainfall, humidity and solar radiation) since nutrient concentration in aquatic environment are known to affect the crude protein of the water plant (Hutchinson, 1975). The findings from the present study revealed that, the fibre content of the leaves ranged between 12.10 and 15.96% and therefore, might be too high for monogastrics to handle (Johnston et al., 2003) who reported that addition of bulky, 26 fibrous ingredients to the diet of livestock will dramatically change the handling characteristics of the final feed. There was an insignificant difference between the fibre content in the leaves of plants harvested from waste water as well as freshwater but the fibre content of the plants harvested from waste water and freshwater were significantly higher than that of saline water. The present findings were higher than previous findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 19.13%, 19.1% and 21.9% respectively. This may be due to the stage of growth at which the water lettuce were harvested (Dangour et al., 2009). The fresher the plant is, the higher it’s nutritional content and lower its fibre content. Wolverton and McDonald (1976) suggested that, the higher fibre content in older aquatic plant could be a result of increased cellulose content. The fat content ranged between 2.27% and 4.35%. There was an insignificant difference between fat content in the leaves of plants obtained in the waste water and freshwater but those of the waste and freshwater were significantly lower than that of saline water. Fat content obtained among all the treatment agreed with earlier findings that, aquatic plants contain 1.18% to 5.42% crude fat (Boyd, 1986). Generally, the ash content of the leaves were highly significantly different but the ash content obtained in the leaves harvested from waste water and freshwater were significantly higher than that of saline water. Ash content among all the treatments were lower than earlier findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 21.12%, 17.0% and 30.1% ash content respectively. The higher content of ash in water lettuce could indicate availability of minerals which may probably be essential for plant growth and development and also important in livestock feed formulation. 27 Table 4.2 shows the Proximate Composition of the root of water lettuce plant Table 4.2: Proximate Composition of the root of water lettuce plant Composition Waste Lagoon Freshwater SED Significance water Moisture 94.36b 96.06a 92.67c 0.3418 *** Dry Matter 5.64b 3.94c 7.33a 0.3418 *** Crude Protein 20.10b 24.33a 12.03c 0.7034 *** Fibre 12.12c 13.88b 15.60a 0.4501 *** Fat 1.41a 1.53a 0.35b 0.0987 *** Ash 16.15b 22.99a 23.40a 1.1514 *** SED= Standard error of difference, Means in the same row with different superscripts are significant ***= Significant (p<0.001) The moisture content of the roots had higher significant difference among the treatments. The moisture content obtained from the roots of plants harvested from the waste water was similar to earlier work by Banerjee and Mata (1990) who reported 94% moisture content. The MC of root of plants harvested from freshwater was slightly lower than earlier findings by Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 94% and 93.1% moisture content respectively whereas those obtained in the saline water being slightly higher than earlier findings by Banerjee and Mata (1990) who reported 94% moisture content. The difference in the present and previous study may probably be due to similar reasons pointed out in the leaves of the water lettuce plant. The dry matter (DM) of the roots obtained from the present findings had a significant difference among all the treatments. The dry matter content of the roots harvested in the waste water was similar to earlier findings by Banerjee and Mata (1990) who reported 5.3%. The DM obtained in the roots harvested from the freshwater was slightly higher than earlier findings by Banerjee and Mata (1990) who reported 5.3% whereas the root of plants harvested from the saline water was 28 slightly lower than earlier works of Banerjee and Mata (1990) who reported 5.3%. The difference in DM in the present and previous study may probably be as a result of similar reasons pointed out for the leaves. There was a statistical difference among all the treatments with the highest CP obtained in the root of plants harvested from the saline water followed by the waste water and that of the freshwater. The CP obtained in the root of plants harvested from waste water and saline water were slightly higher than earlier work of Ayoade et al., (1982) who reported 13.9% CP whereas that of the freshwater was lower than earlier findings by Banerjee and Mata (1990) who reported 20.5%CP. The variations in the crude protein content may be due to similar reasons given for the leaves of the plant. The finding from the present study revealed that, the fibre content of the roots ranged between 12.12 and 15.6% and therefore, might be too high for monogastrics to handle (Johnston et al., 2003). There was a higher significant difference (p<0.001) among the fibre content obtained in all the treatments. The present findings were higher than previous findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 19.13%, 19.1% and 21.9% respectively. The variation between the present and previous findings may be due to similar reasons pointed out for the leaves. There was an insignificant difference between fat content obtained in the root of plants harvested from waste water and saline water but were significantly higher and agreed with earlier findings that, aquatic plants contain 1.18% to 5.42% crude fat (Boyd, 1986). The fat content of the roots of plants harvested from the freshwater was slightly lower than earlier findings by Boyd, 1986 who reported 1.18% to 5.42% crude fat content. The ash content of the roots analyzed were highly significant. The ash content of roots of plants harvested from saline water and freshwater were slightly higher than previous findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 21.12%, 17.0% and 30.1% 29 respectively whereas roots of plants harvested from the waste water was slightly lower than previous findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 21.12% and 17.0% respectively. Table 4.3 indicates the Proximate Composition of the stem of water lettuce plant Table 4.3: Proximate Composition of the stem of water lettuce plant Composition Waste Lagoon Freshwater SED Significance water Moisture 94.20a 86.20c 87.85b 0.5939 *** Dry Matter 5.79b 13.79a 12.15c 0.5939 *** Crude Protein 18.62b 21.99a 13.01c 0.4757 *** Fibre 19.01 18.27 20.20 0.6137 NS Fat 1.70b 2.68a 2.45c 0.1300 *** Ash 13.22 14.03 13.63 0.8024 NS SED= Standard error of difference, Means in the same row with different superscripts are significant ns= not Significant, **= significant (p<0.01), ***= Significant (p<0.001) The moisture content obtained for the stem had a high significant difference among all the treatments. The stem of plants harvested from the waste water had the highest moisture content followed by those from the freshwater and then the saline water. The moisture content obtained from the stem of the plants from the waste water was similar to earlier work by Banerjee and Mata (1990) who reported 94% moisture content whereas that of the freshwater and saline water were lower than earlier findings by Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 94% and 93.1% moisture content respectively. The stem of plants harvested from the waste water was slightly higher than earlier findings of Ayoade et al., (1982) who reported 93.1% moisture content. The variation in the findings 30 (present and previous) may be due to similar reasons pointed out in the leaves of the plant. The dry matter (DM) obtained from the present findings was significantly higher (p<0.001) among the treatments. The DM the stem of plants harvested from freshwater and saline water were higher than earlier findings by Banerjee and Mata (1990) who reported 5.3% whiles that of the waste water was similar to earlier findings by Banerjee and Mata (1990) who reported 5.3%. There was a higher significant difference (p<0.001) among all the treatments with the highest CP obtained in the stem of plants harvested from the saline water, followed by those from the waste water and freshwater. The crude protein content of the stem of plants harvested from the waste water and saline water were higher than earlier work of Ayoade et al., (1982) who reported 13.9% CP and Banerjee and Mata (1990) who reported 20.5%CP whereas the CP obtained in the stem of plants from freshwater was similar to earlier findings by Banerjee and Mata (1990) who reported 13.9%CP. The finding from the present study revealed that, there was an insignificant difference in fibre content of the stem of plants among the treatments. The highest fibre content was obtained in the stem of plants harvested from the freshwater followed by that of the waste water and saline water. The present findings of the stem of plants among all treatments were slightly lower than earlier findings of Ayoade et al., (1982) who reported 21.9%CP whereas that of the waste water was similar to earlier findings by Rodríguez et al., (2000) and Banerjee and Mata (1990) who reported 19.13% and 19.1% respectively. There was a higher significant difference among fat content obtained in the stems of all the water bodies. The fat content obtained in the stem of the plants in all treatments were in agreement with earlier findings that, aquatic plants contain 1.18% to 5.42% crude fat (Boyd, 1986). 31 The present findings from all the water bodies were lower to earlier findings of Rodríguez et al., (2000), Banerjee and Mata (1990) and Ayoade et al., (1982) who reported 21.12%, 17.0% and 30.1% ash content respectively. Table 4.4 presents the Proximate Composition of the whole water lettuce plant Table 4.4: Proximate Composition of the whole water lettuce plant Composition Waste Lagoon Freshwater SED Significance water Moisture 93.91a 92.86a 92.39b 0.7265 * Dry Matter 6.09b 7.12b 7.60a 0.7503 * Crude Protein 19.03b 28.22a 13.36c 0.8589 *** Fibre 19.53a 12.31b 20.30a 0.5614 *** Fat 2.27b 3.71a 0.75c 0.1905 *** Ash 14.36b 11.55a 10.56a 1.2820 ** SED= Standard error of difference, Means in the same row with different superscripts are significantly *= significant (p<0.05), **= significant (p<0.01), ***= Significant (p<0.001) There was statistical difference among the moisture content of the entire plants but the moisture content of entire plants harvested from waste water and saline water were significantly higher than those of freshwater. The moisture content of plants among all the treatment were slightly lower than earlier findings by Banerjee and Mata (1990) who reported 94% moisture content whereas that of the waste water was similar to earlier findings of Ayoade et al., (1982) who reported 93.1% moisture content. The variation in the findings (present and previous) may be due to similar reasons given for the leaves of the plant. 32 The dry matter (DM) obtained in the entire plant had a lower significant difference (p<0.05) among the treatments. There was insignificant difference between the DM of the plants from the waste water and the saline water whereas those from the freshwater was significantly higher than that of waste water and saline water. The DM obtained in plants from all treatments were all slightly higher than earlier findings by Banerjee and Mata (1990) who reported 5.3%. The difference in DM in the present and previous study may probably be as a result of similar reasons pointed out in the leaves. There was a higher statistical difference (p<0.001) among all the treatments with the highest CP obtained in the plants harvested from the saline water followed by waste water and that of the freshwater. The crude protein content obtained in plants harvested from waste water and saline water were higher than earlier findings of Ayoade et al., (1982) who reported 13.9% CP whereas CP of plants harvested from the freshwater was similar to Ayoade et al., (1982) earlier findings. The crude protein content of entire plants harvested from the saline water was higher than earlier work of Banerjee and Mata (1990) who reported and 20.5% CP. The variations in the crude protein content may be due to similar reasons given for the leaves of the plant. The finding from the present study revealed that, the fibre content of the entire plants were highly significant different. There was no significant difference between the fibre content of the waste water and freshwater but the fibre content obtained in the plants harvested from freshwater and waste water were significantly higher than that of the saline water. The fibre content of plants harvested from freshwater and waste water were slightly lower than previous findings of Ayoade et al., (1982) who reported 21.9% whereas that of plants harvested from the waste water was similar to earlier findings by Rodríguez et al., (2000) and Banerjee and Mata (1990) who reported 19.13% and 19.1% respectively. There was a significant difference (p<0.001) among fat content obtained in the plants in all the treatments. The fat content obtained in whole plants harvested 33 from waste water and saline water were in agreement with earlier findings that, aquatic plants contain 1.18% to 5.42% crude fat (Boyd, 1986) but the fat content of plants harvested from the freshwater was slightly lower than earlier findings of (Boyd, 1986) who reported 1.18% to 5.42% crude fat. There was a significant difference (p<0.01) among all the treatments. The ash content obtained in the plants harvested from freshwater and saline water were significantly lower than that of the waste water. The present findings were lower than previous findings of Rodríguez et al., (2000) who reported 21.12%, Banerjee and Mata (1990) who reported 17.0% and Ayoade et al., (1982) who reported 30.1%. SUMMARY Comparing the results obtained in the study in terms of the apportioned parts of the plant, it was realized that, the proximate composition of the leaves was highly nutritious compared to the roots, stems and the whole plants. Considering the nutritive quality of the leaves, those harvested from the saline water had a high crude protein content and lower fibre content compared to those from the waste water and freshwater. The dry matter content of the leaves was low and as a result, more of the water lettuce plants need to be harvested in order to obtain a significant proportion for its potential use as livestock feed. It is therefore important to adopt a cost-effective and efficient drying method if it is to be used on commercial basis. 34 Table 4.5 shows the Levels of some heavy metals in the leaves of the lettuce plants Table 4.5: Levels of some heavy metals in the leaves of the water lettuce plants Metal(ppm) Waste Lagoon Freshwater SED Significance water Cu 4.70 × 10-4a 4.47 ×10-4a 2.10 ×10-3b 2.0 × 10-4 *** Zn 4.20 ×10-3b 3.41 ×10-3b 6.90 ×10-3a 6.9 × 10-4 *** K 1.532a 1.502a 1.162b 0.0540 *** Na 0.703 0.842 0.810 0.0370 NS Fe 4.129a 2.514b 4.063a 0.2650 *** SED= Standard error of difference, Means in the same row with different superscripts are significantly different ns= not Significant, ***= Significant (p<0.001) The leaves harvested from the waste water contained higher concentrations of the heavy metals compared with the freshwater and saline water. Results of this study revealed that, there were significant differences (P<0.001) in the concentration of heavy metals in all treatments except sodium. The copper in the leaves harvested from waste water and saline water were significantly higher than that of the freshwater. Also, it was revealed that, zinc in the leaves of plants obtained from waste water and freshwater were significantly higher than that of the saline water. For potassium, leaves harvested from waste water and saline water were significantly higher than that of the freshwater whereas for iron, roots of plants harvested from waste water and freshwater were significantly higher than that of the saline water. The higher concentrations of heavy metals may be due to the activities carried out at the location including washing of cars around the water bodies, farming and dumping of waste which served as a means of releasing heavy metals into the environment. Surface runoff water through the gutters constructed 35 into the water bodies may have also contributed to higher levels of heavy metals. According to Duffus (2002), concentrations of the heavy metals in saline water are higher than that of freshwater and this might be due to the fact concentrations in terrestrial and aquatic organisms are determined by the size of the source and adsorption and/or precipitation in soils and sediments. The extent of adsorption depends on the metal, the absorbent, and the physico-chemical characteristics of the environment (e.g. pH, water hardness and redox potential). Other factors such as changes in temperature, pH as well as effluent from domestic, municipal and agricultural waste including pesticides might have contributed to the higher level of heavy metals in the water body (Okweye et al., 2009). Generally, the maximum threshold of heavy metals detected in the leaves from the three water bodies were below their maximum thresholds. The concentration of copper, zinc, potassium, sodium and iron were below their maximum limit of 25ppm, 50ppm, 10ppm, 100ppm and 500ppm respectively. Table 4.6 shows the levels of some heavy metals in the root of the water lettuce plant Table 4.6: Levels of some heavy metals in the root of water lettuce plant Metal(ppm) Waste Lagoon Freshwater SED Significance water Cu 3.19 ×10-3b 1.66 ×10-3c 4.73 ×10-3a 2.44 ×10-4 *** Zn 1.15 ×10-2b 1.65 ×10-3a 5.41 ×10-3c 4.53 ×10-4 *** K 1.132b 1.338a 0.766c 0.0290 *** Na 0.654 7.066 0.697 0.0371 NS Fe 4.129a 2.514b 4.062a 0.2654 *** SED= Standard error of difference, Means in the same row with different superscripts are significant ns= not Significant, ***= Significant (p<0.001) 36 The results detected in the roots were significantly different (P<0.001) in concentration of heavy metals except sodium which had insignificant difference among the treatments. The concentration of copper in the roots of plants harvested from waste water and freshwater were significantly higher than that of the saline water whereas the concentration of Zinc in the roots of plants harvested from freshwater and saline water were significantly higher than that of the waste water. Also, the concentration potassium detected in the roots of plants harvested from waste water and saline water were significantly higher than that of the freshwater but that of the saline water was higher than the waste water whereas the concentration of iron of the roots of plants harvested from waste water and the freshwater were significantly higher than that of the saline water. The variation in the concentration of the metals in the study may be due to the fact that, concentrations in terrestrial and aquatic organisms are determined by the size of the source and adsorption and/or precipitation in soils and sediments. The extent of adsorption depends on the metal, the absorbent, and the physico-chemical characteristics of the environment (e.g. pH, water hardness and redox potential) Duffus, (2002). The variation in the concentration of the metals detected in the study may probably be due to industrial activities, and higher exploitation of cultivable land which has brought about a huge increase in the quantity of discharges and wide diversification in types of pollutants that reach aquatic environments. The ultimate sink for many of these contaminants in the aquatic environment may be due to discharges or hydrologic and atmospheric processes (Lagadic et al., 2000). The concentration of copper, zinc, potassium, sodium and iron were below their maximum limit of 25ppm, 50ppm, 10ppm, 100ppm and 500ppm respectively. 37 Table 4.7 presents the levels of some heavy metals in the stem of water lettuce plant Table 4.7: Levels of some heavy metals in the stem of water lettuce plants Metal(ppm) Waste Lagoon Freshwater SED Significance water Cu 3.35 ×10-3a 4.60 ×10-4b 6.85 ×10-4b 2.36 ×10-4 *** Zn 6.27 ×10-3c 8.68 ×10-3b 1.11×10-2a 8.62 ×10-4 *** K 1.818a 1.460b 1.180c 3.64 ×10-2 *** Na 0.882b 0.990a 0.793c 0.0366 *** Fe 1.983a 0.782b 3.182c 0.1466 *** SED= Standard error of difference, Means in the same row with different superscripts are significant ***= Significant (p<0.001) The results detected for the stems revealed that, the heavy metals were of higher significance (P<0.001) in concentration for all the treatments. The concentration of Cu in the stems harvested from saline water and freshwater were significantly higher than that of the waste water whereas the concentration of zinc in stem of plants harvested from waste water and the saline water were significantly higher than that of the freshwater whereas that of the saline water was higher than those of the waste water. The concentration of potassium in stems harvested from waste water and the saline water were significantly higher than that of the freshwater but that of waste water was higher than that of saline water. The concentration of sodium detected in stems of plants harvested from waste water and saline water were significantly higher than those of the freshwater whereas that of the saline water was higher those from the waste water. Also, the concentration of iron in stem of plants harvested from freshwater and waste water were significantly higher than that of the saline water whereas that of the freshwater was also significantly higher than that of the waste water. Generally, the maximum threshold of heavy metals detected in the leaves from the various sources were below their maximum thresholds. The concentration of copper, zinc, potassium, sodium and 38 iron were below their maximum limit of 25ppm, 50ppm, 10ppm, 100ppm and 500ppm respectively. Table 4.8 indicates the Levels of some heavy metals in the entire plant of water lettuce Table 4.8: Levels of some heavy metals in the entire plant of water lettuce plants Metal(ppm) Waste Lagoon Freshwater SED Significance water Cu 2.06 × 10-3a 9.75 × 10-4b Zn 6.43 × 10-3c 1.46 × 10-2a 4.60 × 10-4c 7.16 × 10-3b 1.07 × 10-4 3.52 × 10-4 *** *** K 1.651a 1.574a 1.062b 0.078 *** Na 0.759 0.853 0.827 0.055 NS Fe 2.139a 2.253a 1.056b 0.108 *** SED= Standard error of difference, Means in the same row with different superscripts are significant ns= not Significant, ***= Significant (p<0.001) The heavy metals detected from the study revealed that, the lettuce plants among the treatments were of higher significance (P<0.001) in concentration except for sodium which had insignificant difference among the treatments. For copper, the plants harvested from saline water and freshwater were significantly higher than that of the waste water but that of saline water was higher than that of freshwater. Concentration of zinc detected also revealed that, those harvested from waste water and freshwater were significantly higher than that of saline water but that of the freshwater was significantly higher than that of waste water. In the potassium and iron analysis, it was detected that the concentration of potassium and iron in waste and saline water were significantly higher than that of the freshwater. 39 SUMMARY Comparing the results obtained in this study with the European Commission Regulation (ECR), Agricultural Research Council Party (ARCP), Chemical Analysis of Ecological Material and Council Directive 80/778/EEC permissible levels of heavy metals in aquatic plants, the concentrations of Cu, Zn, K, Na and Fe were below their maximum limits. This suggests that, although the apportioned parts of the water lettuce plants and the water bodies had witnessed some level of contamination, currently, they are within acceptable limits of European Commission Regulation (ECR), Agricultural Research Council Party (ARCP), Chemical Analysis of Ecological Material and Council Directive 80/778/EEC recommended guidelines. Copper may serve as an essential element by serving as a cofactor in a number of enzymes system and necessary for the synthesis of hemoglobin (Sivaperumal et al., 2007), zinc may also serve as a catalytic or structural component in many enzymes that are involved in metabolism, transcription and translocation of RNA. Also, potassium contribute to the regulation of acid-base sodium and participates in respiration via the chloride shift whiles sodium promotes pH distribution. Iron regulates cell growth and differentiation and also play a significant role in metabolic process. Generally, the level of contamination of the heavy metals detected in the study were higher in the leaves compared to the roots, stems and the whole plants. This may probably be due to the fact that, the plants absorbed these metals through their leaves (Al Jassir et al., 2005). Level of the Cu, Zn, K, Na and Fe detected in the stems, whole plants and the roots followed the leaves in respective order in contamination. 40 Table 4.9 presents the Levels of some heavy metals in three water bodies Table 4.9: Levels of some heavy metal in the three water bodies Metal(ppm) Waste water Lagoon Freshwater Cu 1.0 × 10-2 1.0 × 10-2 2.0 × 10-2 Zn 2.0 × 10-2 4.0 × 10-2 3.0 × 10-2 K 15 17 12 Na 18.28 22.58 19.36 Fe 1.46 1.56 1.72 The levels of heavy metal pollution in the water bodies were below their maximum thresholds except potassium. Copper level range between 1.0 × 10-2 and 2.0 × 10-2ppm whereas zinc level in the water range between 2.0 × 10-2 and 4.0 × 10-2ppm. The range of zinc was below European Commission Regulation (ECR) 237790 (consolidated) maximum threshold of 50ppm.The level of potassium in the water sources ranged between 12 and 17ppm with the detected level of potassium in the lagoon slightly higher than that of waste water and freshwater. The level of potassium detected in the study was above the Chemical Analysis of Ecological Material, (1974) maximum threshold of 10ppm.The level sodium in the water sources ranged between 18.28 and 22.58ppm and was below the Chemical Analysis of Ecological Material, (1974) recommended maximum threshold of 100ppm. The level being below the maximum threshold might not impede the growth of the plant. The level of iron range between 1.46 and 1.72ppm. The variations in the levels of Cu, Zn, K, Na and Fe detected from the water sources may probably be as a result of the degree of pollution of the water sources, that is, the more polluted the water source is, the higher the tendency of the plant intake. It was realized that, the saline water (Fosu lagoon) is more polluted than the freshwater as well as the residential waste water. This may probably be due to the extensive use of pesticide, car washing activities and dumping of domestic waste in the water bodies (Dankwah, 2011) as well as effluent from domestic, municipal and agricultural waste might 41 have contributed to the higher level of heavy metals in the water body (Okweye et al., 2009). 42 CHAPTER FIVE 5.0 CONCLUSIONS AND RECOMMENDATIONS 5.1 CONCLUSIONS 1. Water lettuce from all the three water bodies had high level of crude protein, making it a potential feed resource for compounding livestock feed. 2. Moisture content of the water lettuce was high, hence different methods of drying needs to be used if the water lettuce is to be used on commercial basis. 3. The heavy metals detected in this study (water lettuce) were below their maximum thresholds, and therefore may not cause any harm to the animal if it is utilized as feed resource. 4. The heavy metals detected from the water bodies namely; copper, zinc and iron were below their maximum thresholds, and therefore water lettuce harvested from the water bodies may not cause any harm to the animal if it is utilized as feed resource. 5.2 RECOMMENDATIONS Based on the conclusions, the following recommendations were made: 1. Based on the moisture content from the present study, further studies should be carried out to compare different methods of drying the water lettuce plants to determine the one which is cost-effective and efficient 2. Further studies should be conducted to assess the other metals that were not detect in this study, namely; mercury, arsenic, lead and magnesium to ensure that heavy metals does not have any effect on the water lettuce 3. 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