Larvicidal potential of extracts of persea americana seed and chromolaena odorata leaf against aedes vittatus mosquito

 

Table Of Contents


  • <p> </p><p><strong>ABSTRACT</strong></p><p><strong>

Chapter ONE

INTRODUCTION

  • </strong><br>
  • 1.0INTRODUCTION<br>
  • 1.1Statement of Research Problem<br>
  • 1.2Justification<br>
  • 1.3Aims and Objectives<br>1.
  • 3.1General Aim<br>1.
  • 3.2Specific Objectives</p><p><strong>

Chapter TWO

LITERATURE REVIEW

  • </strong><br>
  • 2.0LITERATURE REVIEW<br>
  • 2.1Mosquito<br>2.
  • 1.3Life Cycle of Aedes<br>2.
  • 1.4Mosquito Morphology and Feeding Habits<br>2.
  • 1.5Mosquito Born Diseases<br>2.
  • 1.6Mosquito Control Methods<br>2.
  • 1.7Active Ingredients in Plants Responsible for Larval Toxicity<br>2.
  • 1.8Mechanism and mode of action of insecticide/larvicide<br>2.
  • 1.9Toxicity Response Determinant and Variation of plant derived larvicides<br>2.
  • 1.10Scope for isolation of toxic larvicidal active ingredients from plants<br>
  • 2.2Chromolaena odorata<br>2.
  • 2.1Classification of C. odorata<br>2.
  • 2.2Origin and Distribution<br>2.
  • 2.3Traditional Uses of C. odorata<br>2.
  • 2.4Phytochemical Composition of C. odorata<br>2.
  • 2.5Medicinal Values of C. odorata<br>2.
  • 2.6Antibacterial effect of C. odorata<br>2.
  • 2.7Toxicity of C. odorata<br>
  • 2.3Persea americana<br>2.
  • 3.1Classification of Persea americana<br>2.
  • 3.2Biological activities of Persea americana constituents<br>2.
  • 3.3Phytochemical composition of avocado seed<br>2.
  • 3.4Tradomedicinal Uses of Avocado Seed<br>2.
  • 3.5Larvicidal and antimicrobial activities<br>2.
  • 3.6Toxicity of avocado seed</p><p><strong>

Chapter THREE

RESEARCH METHODOLOGY

  • </strong><br>
  • 3.0MATERIALS AND METHODS<br>
  • 3.1Materials<br>3.
  • 1.1Chemicals<br>3.
  • 1.2Plants Collection and identification<br>
  • 3.2Methods<br>3.
  • 2.1Preparation of extracts<br>3.
  • 2.2Mosquito Larvae culture<br>
  • 3.3Phytochemical Analysis<br>3.
  • 3.1Test for Saponins<br>3.
  • 3.2Test for tannins<br>3.
  • 3.3Test for flavonoids<br>3.
  • 4.4Test for sterols<br>3.
  • 4.5Test for Terpenoids<br>3.
  • 4.6Test for Anthracenes<br>3.
  • 4.7Test for cardiac glycosides<br>3.
  • 4.8Test for alkaloids<br>
  • 3.4Preparation of Stock Solutions<br>3.
  • 4.1Preparation of Test Concentrations For Bioassay<br>
  • 3.5Larvicidal Bioassay<br>3.
  • 5.1Determination of Lethal Concentrations<br>
  • 3.6Thin Layer Chromatography (TLC)<br>3.
  • 6.1Column Chromatography<br>
  • 3.7Characterization of Larvicidal Compounds In The Bioactive Fraction<br>3.
  • 7.1Fourier Transform Infra-Red Spectroscopy(FTIR)<br>3.
  • 7.2Gas Chromatography/Mass Spectroscopy (GC/MS)<br>
  • 3.8Statistical Analysis</p><p><strong>

Chapter FOUR

DATA PRESENTATION AND ANALYSIS

  • </strong><br>
  • 4.0RESULTS<br>
  • 4.1Phytochemical Constituents of Extracts of Persea americana Seed and Chromolaena odorata Leaf<br>
  • 4.2Larvicidal activity of different solvent extracts of Persea americana seed against Aaedes vittatus mosquito<br>
  • 4.3Larvicidal Activity of Different Solvents Extract of Chromolaena odorata Leaf Against Aedes vittatus Mosquito<br>
  • 4.4Larvicidal activity of chromatographic fractions of n-hexane extracts of Persea americana seed against Aedes vittatus<br>
  • 4.5Larvicidal activity of chromatographic fractions of n-hexane extracts of Chromolaena odorata leaf against Aedes vittatus mosquito<br>
  • 4.6GC/MS Characterisation of Most Potent Chromatographic Fraction (nHPa6) of P. americana<br>
  • 4.7GC/MS characterisation of Most Potent Chromatographic Fraction (nHCo6) of C. odorata<br>
  • 4.8Functional group Identification of nHPa6<br>
  • 4.9Functional groups Identification of nHCo6 fraction</p><p><strong>

Chapter FIVE

SUMMARY, CONCLUSION AND RECOMMENDATIONS

  • </strong><br>
  • 5.0DISCUSSION</p><p><strong>CHAPTER SIX</strong><br>
  • 6.0SUMMARY, CONCLUSION AND RECOMMENDATIONS<br>
  • 6.1Summary<br>
  • 6.2Conclusions<br>
  • 6.3Recommendations<br>REFERENCES<br>APPENDICES</p><p><strong>Abbreviations</strong></p><p>LC = Lethal Concentrations</p><p>GCMS = Gas Chromatography Mass Spectrometry</p><p>FTIR = Fourier Transform Infra-Red</p><p>n-Hexane = Normal Hexane</p><p>TLC = Thin Layer Chromatography</p><p>DMSO = Dimethyl Sulfoxide</p><p>JE = Japanese Encephalitis</p><p>WHO = World Health Organisation</p><p>Cx = Culex</p><p>An = Anopheles</p><p>Ae = Aedes</p><p>CHIKV = Chikungunya Virus</p><p>Bti = Bacillus Thuriengiensis</p><p>Bs = Bacillus Sphaericus</p><p>DDT = Dichloro-Diphenyl-Trichloroethane</p><p>IGR = Insect Growth Regulator</p><p>CSI = Chitin Synthesis Inhibitor</p><p>CNS = Central Nervous System</p><p>ACh = Acetyl Choline</p><p>AChE = Acetyl Choline Esterase</p><p>GABA = Gammaamino Butyric Acid</p><p>ATP = Adenosine Triphosphate</p><p>PTTH = Prothoracictropic Hormone</p><p>CCl4 = Tetrachloromethane</p><p>NMR = Nuclear Magnetic Resonance</p> <br><p></p>

Project Abstract

<p> The larvicidal activity of various solvent (ethanol, ethyl acetate and n-hexane) extracts of Persea americana seed and Chromolaena odorata leaves against Aedes vittatus mosquito was analysed. The most potent solvent (n-hexane) extracts of both plants were fractionated using column chromatography and most effective fractions isolated and identified using Gas Chromatography Mass Spectrometry and Fourier Transform Infrared techniques. Phytochemical screening revealed the presence of steroids, cardiac glycosides and terpenoids in all the extracts. The larvicidal bioassay of Persea americana seed gave LC50 values of 0.827ppm, 1.799ppm and 2.732ppm for n-hexane, ethanol and ethyl acetate extracts respectively, while, Chromolaena odorata leaf extract had LC50 values of 1.835ppm, 3.314ppm, and 5.163ppm for n-hexane, ethanol and ethyl acetate respectively. Column chromatographic fractionation of most potent n-hexane (crude) extracts of both plants, showed increased activity in some of the fractions of Persea americana (nHPa6) and Chromolaena odorata (nHCo6) which showed higher mortality, with LC50 values of 0.486ppm and 1.308ppm respectively. GC/MS analysis of components in nHPa6 and nHCo6 showed oleic acid as the most abundant, in fractions of both plants. The FTIR analyses of nHPa6 and nHCo6 showed absorption bands of the functional groups present, which included; alcohol, alkane, alkene, alkyl halide, aldehyde, carboxylic acid and carbonyl ester, thus, supporting the GCMS result. The n-hexane, ethanol and ethyl acetate extracts of P. americana seed and C. odorata leaves have shown good larvicidal activity and should therefore be further exploited for the control of mosquito larvae. <br></p>

Project Overview

<p> </p><p><strong>1.0 </strong><strong>INTRODUCTION</strong></p><p>Insect-transmitted diseases remain a major cause of morbidity and mortality worldwide. Mosquito species belonging to genera; <em>Anopheles</em>, <em>Aedes</em>&nbsp;and <em>Culex</em>, are vectors (Redouane <em>et al</em>., 2002) for the transmission of malaria, dengue fever, yellow fever, filariasis, schistosomiasis and Japanese encephalitis (JE), transmitting diseases to more than 700 million people annually (Oyewole <em>et al</em>., 2010; Govindarajan, 2009). Mosquitoes also cause allergic responses in humans which include local skin irritation and systemic reactions such as angioedema. <em>Aedes spp</em>&nbsp;are generally regarded as a vector responsible for transmission of yellow fever and dengue fever, which is endemic to Southeast Asia, the Pacific island area, Africa, Central and South America.</p><p>The World Health Organization (W.H.O. 2012) has recommended vector control as an important component of the global strategy for preventing insect-transmitted diseases. The most commonly employed method for the control of mosquito-borne diseases involve the use of chemical-based insecticide, though it is not without numerous challenges, such as human and environmental toxicity, resistance, affordability and availability (Ghosh <em>et al</em>., 2012).</p><p>Extracts from plants has been good sources of phytochemicals as mosquito egg and larval control agents, since they constitute an abundant source of bioactive compounds that are easily biodegradable into nontoxic products. In fact, many researchers have reported on the effectiveness of plant extracts or essential oils against mosquito larvae. They act as larvicides, insect growth regulators, repellents, and oviposition attractants (Pushpanathan, 2008; Samidurai <em>et al.,</em>&nbsp;2009; Mathivanan <em>et al.,</em>&nbsp;2010).</p><p><em>Persea Americana </em>is an evergreen tree belonging to<em>&nbsp;Lauraceae </em>family and its fruits are commonly known as avocado pear or alligator pear. The plant originates from Central America but it has shown easy adaptation to other tropical regions, thus widely cultivated in tropical and subtropical regions. The various parts (leaves, fruits and seed) of this plant have numerous uses from edible pulp as source of nutrients to the seed preparation as remedy (Arukwe <em>et al.,</em>&nbsp;2012).</p><p>The seed extracts of <em>Persea americana</em>&nbsp;has many vital application in traditional medicine, for the treatment of diarrhoea, dysentery, toothache, intestinal parasites, skin infection (mycoses) and management of hypertension and the leaves have been reported to have anti-inflammatory and analgesic activities (Adeyemi <em>et al.,</em>&nbsp;2002; Ozolua <em>et al.,</em>&nbsp;2009). Phytochemical screening of avocado seed shows the presence of fatty acids, Triterpenes, anthocyanin, flavonoids and abscisic acids (Leiti <em>et al.,</em>&nbsp;2009).</p><p><em>Chromolaena odorata </em>is a weed which belongs to<em>&nbsp;Asteraceae </em>family. It is found in tropical and subtropical areas, extending from west, central and southern Africa to India, Sri Lanka, Bangladesh, Laos, Cambodia, Thailand, southern China, Taiwan, and Indonesia. The weed goes by many common names including; Siam weed, devil’s weed, French weed, communist weed (Vaisakh and Pandey, 2012). In Nigeria, the <em>Chromolaena odorata</em>&nbsp;is referred to as <em>Obu inenawall</em>&nbsp;by the Igbo and ―<em>ewe awolowo</em>‖ by the Yoruba. This plant is exploited traditionally for its medicinal properties, especially for external uses as in wounds, inflammation and skin infections. Some studies also demonstrate the efficacy of its leaf extract, as antioxidant, anti-inflammatory, analgesic, anti-microbial and cytoprotective agent (Ajao <em>et al.</em>, 2011). The oil from <em>C. odorata</em>&nbsp;also had been exploited as insecticide, ovicide and larvicide (Noud Agbessi <em>et al.</em>, 2006). Previous phytochemical studies of the leaf extracts of <em>C. odorata</em>&nbsp;show the presence of alkaloid, cardiac glycosides, anthocyanin, tannin, and flavonoids (Ngozi <em>et al</em>., 2009).</p><ul><li><strong>Statement of Research Problem</strong></li></ul><p>An estimated 3.3 billion people are at risk of malaria globally, with populations living in sub-Saharan Africa having the highest risk (WHO, 2012) and two-fifths of the world‘s population is at risk of dengue fever (WHO, 2003). Malaria alone accounts for about 50 per cent of out-patient consultation, 15 per cent of hospital admission, and also among the top three causes of death in the country.</p><p>In recent years, the use of many synthetic insecticides in mosquito control programme has been limited, due to many challenges such as, high cost of synthetic insecticides, environmental sustainability, toxic effect on human health (immune suppression), and other non-target organisms, environmental persistence, higher rate of biological accumulation and magnification through ecosystem, as well as increasing insecticide resistance on large scale (Srivastava and Sharma, 2000; Raghvendra and Subbarao, 2002). These challenges have resulted in an urge to search for environmentally sustainable, biodegradable, affordable and target selective insecticides against mosquito species (Saxena and Sumithra, 1985; Kumar and Dutta, 1987; Chariandy <em>et al</em>., 1999; Markouk <em>et al</em>., 2000; Tare <em>et al</em>., 2004).</p><p>Consequently, the application of eco-sustainable alternatives such as biological control of vectors has become the main focus of the control programme to replace the synthetic chemical insecticides (Gosh <em>et al.,</em>&nbsp;2012). One of the most effective alternative approaches under the biological control programme is to utilise the plants biodiversity as a reservoir of safer insecticides of botanical origin as a simple, affordable and sustainable method of mosquito control.</p><p>Mosquito larvae is the easiest stage to target in its life cycle and several studies have documented the efficacy of plant extracts as a reservoir pool of bioactive toxic agents against mosquito larvae. Furthermore, evolution of the resistance to plant-derived compounds has rarely been reported (Sharma <em>et al.,</em>&nbsp;2006).</p><p>However, the main reasons for the failure in laboratory to field utilisation of bioactive phytochemicals are poor characterization and inability to determine the active toxic components responsible for larvicidal activity (Ghosh <em>et al.,</em>&nbsp;2012). Hence, there is a need for the characterisation, of various plant extracts to determine the active (larvicidal) components of locally available plants for mosquito control. This will help to reduce dependence on expensive and mostly imported products, and stimulate local efforts to enhance the general public health.</p><ul><li><strong>Aims and Objectives</strong></li><li><strong>General Aim</strong></li></ul><p>The aim of this study was to investigate the larvicidal potential of extracts of <em>Persea</em>&nbsp;<em>americana </em>seed and<em>&nbsp;Chromolaena odorata </em>leave against<em>&nbsp;Aedes vittatus </em>larvae</p><ol><li>Phytochemical analysis (qualitative) of the crude extracts of <em>persea americana</em>&nbsp;seed and <em>Chromolaena odorata</em></li><li>Determination of the most potent solvent extracts with larvicidal activity against <em>Aedes vittatus </em>larvae</li><li>Determination of the lethal concentration (LC) of the crude extracts for 50% and 90% mortality (LC50 and LC90).</li><li>Fractionation of the most potent crude extracts and isolation of the most effective (larvicidal) fractions using column chromatography;</li><li>Characterisation of the bioactive (larvicidal) fractions using FTIR and GC/MS techniques.</li></ol><h2>REFERENCES</h2><p>Abbott, W.S. (1925). A method of computing the effectiveness of insecticide. Journal of Economic Entomology, 18, 265–7.</p><p>Adeyemi, O. O., Okpo, S. O. and Ogunti, O. O. (2002). Analgesic and anti-inflammatory effects of the aqueous extract of leaves of Persea americana Mill (Lauraceae). Fitoterapia, 73, 375-380.</p><p>Afolabi, C., Akinmoladun, E.O., and Dan-Ologe, I.A. (2007). Phytochemical Constituents and Antioxidant properties of extracts from the leaves of Chromolaena odorata. Scientific Research and Essay, 2 (6), 191-194.</p><p>Ajao, A. T., Ajadi, T.S. and Oyelowo, M.S. (2011). Evaluation of Multiplicative Killing Effect of C.odorata extracts and β-lactam antibiotics against β-lactamase Producing bacteria, isolated from Selected Hospitals in Ilorin Metropolis. Annals of Biological Research Scholars Research Library, 2, (4), 76-84.</p><p>Alaa, E. B., Balaña-Fouce, R., Sobeih, A. K. and Hussein E. M. K. (1998). The biological activity of some chitin synthesis inhibitors against the cotton leafworm Spodoptera littoralis (Boisduval), (Lepidoptera: Noctuidae). Bulletin of plant health, Pest, 24, 499-506.</p><p>Alaba, A.O. (2005). Malaria and Rural Household productivity in Oyo State. (Doctoral dissertation, University of Ibadan, Nigeria).</p><p>Alisi, C., Ojiako, S. O. A., Osuagwu, C. G., and Onyeze, G. O. C. (2011). Free Radical Scavenging and In-vitro Antioxidant Effects of Ethanol Extract of the Medicinal Herb Chromolaena odorata Linn. British Journal of Pharmaceutical Research, 1(4), 141-155.</p><p>Al-Rajhy, D.H., Alahmed A.M., Hussein H.I. and Kheir, S.M. (2003). Acaricidal effects of cardiac glycosides, azadirachtin and neem oil against the camel tick, Hyalomma dromedaril (Acari: Ixodidae) Pest Management Science, 59(11): 1250-1254.</p><p>Amalraj, D. and Das, P.K. (1998). Estimation of predation by the larvae of Toxorhynchites splendens on the aquatic stages of Aedes aegypti. Southeast Asian Journal of Tropical Medicine and Public Health; 29, 177-83.</p><p>Anees, A.M. (2008). Larvicidal activity of Ocimum sanctum Linn (Labiatae) against Aedes aegypti (L.) and Culex quinquefasciatus (Say). Parasitology Research, 101, 1451-3.</p><p>Anon, (1983). Important Weeds of the World (3rd Edition). Leverkusen, Germany: Bayer A.G.</p><p>Anyasor, G.N., Aina, D.A.; Olushola M., and Aniyikaiye, A.F. (2011). Phytochemical constituent, proximate analysis, antioxidant, antibacterial and wound healing properties of leaf extracts of Chromolaena Odorata. Annals of Biological Research; 2(2): 441-451</p><p>Arukwe, U., Amadi, B.A., Duru, M.K.C., Agomuo, E.N., Adindu, E. A., Odika, P.C., Lele, K.C., Egejuru, L., and Anudike, J. (2012). Chemical Composition of Persea americana Leaf, Fruit and Seed International Journal of Research and Review in Applied Sciences; 11 (2).</p> <br><p></p>

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