INSECTICIDAL POTENCY OF ASYSTASIA VOGELIANA ON BEAN SEEDS (VIGNA UNGUICULATA) IN STORAGE.

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1 Journal of Science, Engineering and Technology 1(2), September Journal of Science, Engineering and Technology 1(2), December 2012: ISSN INSECTICIDAL POTENCY OF ASYSTASIA VOGELIANA ON BEAN SEEDS (VIGNA UNGUICULATA) IN STORAGE. 1 IBOH, C. I., 2 OKON, O. E., 1 AJANG, R. O., 1 OKOI, A. I., 2 ARONG, G. & 3 EFFIOM, O. E. 1 Department of Biological Sciences, Cross River University of Technology, Calabar 2 Department of Zoology and Environmental Biology, University of Calabar, Calabar 3 Department of Biological Sciences, Veritas University, Abuja, Nigeria. Corresponding author: cletoec@yahoo.com Abstract Investigation on the effect of Asystasia vogeliana powder on bean weevil (Callosobruchus maculatus) was carried out in CRUTECH, Calabar, with a view to protecting Vigna unguiculata and having an alternative means to chemical application on beans (Vigna unguiculata) in storage. The powder of Asystasua vogeliana (Yoseh) was applied at different dosages of 1mg, 3mg and 5mg on stored bean grains (Vigna unguiculata) and tested against the bean weevil, Callosobruchus maculatus, in the laboratory. Adult mortality at 24, 48, 76 and 96 hours were assessed. Adult.emergency and grain damage result showed that 3 and 5 mg powder of the plant leaves were more toxic to C. maculatus and suppressed F1 progeny emergence compared to 1mg powder. The efficacy of the powder on the weevil was dose dependent with higher doses providing greater protection to bean grains. The highest mortality rate of C. maculatus was observed at a concentration of 5mg of A. vogeliana powder after 3 days. The phytochemical screening of the plant revealed the presence of Alkaloids, Flavonoids, Tannins, Glycosides, Saponins, Polyphenol, Steroids, Anthraquinones and reducing compounds. This research work provides the scientific basis for the use of A. vogeliana powder in storedproduct protection at the small-scale farmer level to reduce the application of toxic synthetic insecticides. Keywords: Insecticidal, Asystasia vogeliana, Protection, Beans, Phytochemicals. INTRODUCTION For thousands of years insects have been a problem associated with storing surpluses of dry fruits with evidence from archaeological deposit (Buckholder and Ma, 1985) as well as written records of storage and identification of cereals stretching back to 3,000 B.C. (Livinson and Livinson, 1994). There is need to protect stored food from attack by insects because they can destroy large quantities particularly during long term storage (Pimentel, 1991). Allergic reaction due to arthropods in stored food including Sitophilus grenarious, has been reported (Van Lynden-Van et al., 1996). These arthropods cause contamination of grains with their excreta. The history of farmers battle against stored product pest revealed that they have learnt to exploit natural reserves that would lead to a degree of population suppression of pests. Traditional methods usually provide cheap and feasible ways of postharvest control handling of the crop Weevil damage in stored food grains may amount to 10% - 40% in places where modern storage technologies have not been introduced or are lacking (Shaaya, 2007). Basically, farmers should be fairly aware that hygienic practices are essential for successful storing. Some traditionally used materials are often added to granaries to reduce pest activity. Inert dust for example is added in variable amount to the stored product. Friction of dust particles with insect lead to desiccation and hampers the development of the pest (Golob and Wisley, 1980). Ghaffar and Chauhan (1999), showed that hydrophobic silica dusts resulted in

2 96 IBOH, C. I. ET AL: Insecticidal Potency of Asystasia Vogeliana on Bean Seeds... efficient control of Callosobruchus chinensis as no beetle survived after 48 hours at a concentration of 0.1 percent. A similar effect can also be achieved through treatment with wood or ash, collected from burnt tree wood or a farmer s store. In an experiment in India, pretreatment of Vigna radiate seeds with inert clay resulted in 100 percent adult mortality of C. chinensis within 24 hours (Hassanali et al, 1997). In Bangladesh, covering potato with rice husk, sand or wood ash resulted in the control of Phthorimaea operculella in storage (Pantherius, 1999). In Sudan, treating potato tubers with neem leaf extracts and then placing them in jute sacks reduced postharvest loss caused by Phthorimaea operculella. Others extracted oils, such as bean or groundnut oil, have been recognized as toxicants or growth inhibitor of C. maculatus (Adedire and Ayaji, 1996). Asystasia vogeliana is a vascular plant of the Dzanga-Sangha reserve and also found in the Niger Delta region of Nigeria, Lagos, Cameroon, Congo, Guinea etc. It is a fish killer leaf used in other countries like India, Sudan, Ghana and South America. It has been established that in North West Guyana, up to 16 percent of the village fishing still prefer the use of A. vogeliana poison despite the superiority of modern netting materials and prohibition by law of A. vogeliana (Van Andel, 2000). This plant also contains rotenone which is extremely toxic to cold blooded animals and is used as an insecticide (Van Andel, 2000). This plant derived rotenone is synthesized and used for experimental fishing as well as the elimination of undesirable fish population in natural waters (Dolmen et al, 1995). Materials and methods The plant (A. vogeliana) used in this investigation was collected from a damp reserve in Nko village, Yakurr Local Government Area of Cross River State, Nigeria. The plant was transported in a sterile dark polythene bag to the biology laboratory of Cross River University of Technology, Calabar, and authenticated by the herbarium unit of the same University. The plant leaves were spread in the laboratory table to air-dry in a wellventilated area under ambient environmental condition of 32 0 C and 60% Relative Humidity (R.H), for 1-2 weeks to ensure proper drying. The plant leaves were thereafter pulverized using a milling machine (Model ed-5). The powder was then stored in a clean plastic container and preserved in the refrigerator to maintain its freshness until when needed for use. Insect culture Pure culture of Callosobruchus maculatus was collected from established laboratory cultures maintained in the Department of Crop Science of the University of Calabar. These were reared in disinfected bean grains at ambient temperature of 20 0 C to 28 0 C and relative humidity of 75%, in a grain storage research laboratory of the Cross River University of Technology, Calabar. Clean uninfected seeds of beans (V. unguiculata) were collected from the market and kept in the cooled room for about one month for proper disinfection, aeration and to prevent molding in the laboratory (Adedire and Ayaji, 1996). Thirty pairs (male and female) of adult Callosobruchus maculatus were introduced into each container and left for 12 days to breed (sexing). After 12 days the adult Callosobruchus maculatus were withdrawn from the container and left between days for emergence. The newly emerged C. maculatus were used for the experiment. Application of plant powder Disinfected beans (50g) from the freezer were weighed into 8cm diameter transparent plastic containers for each replicate. The leaves powder of A. vogeliana were applied by direct mixing to the bean grains calculated on the weight of plant material to the weight of grain (100g) respectively. The plant powder was applied at three rates of 1mg, 3mg and 5mg in transparent plastic containers, while the control received no plant powder. 15 pairs of 2 day old C. maculatus adult were introduced into each plastic container, containing treatment for mating and oviposition for 6 days. The plastic containers had their covers perforated and covered with nylon mesh to facilitate aeration and confinement of weevils. Each treatment was replicated 4 times and laid out in a Complete Randomized Design (CRD) on the laboratory bench. A mortality count was done every 24, 48, 72 and 96 hours after treatment by sieving out the content into a clean white tray and counting the number of dead insects. Each time a count was done, dead individuals were discarded while live ones were returned to their respective treatment. After 12 days all live and dead adult weevils were removed and discarded and the seeds kept aside for F1 progeny emergence. Calculation of bean weevil mortality was given as: No. of dead insects x 100 Total no. of insects x 1 Weevil emergence test The test was based on the filial generation (F1) progeny emergence, five weeks after the withdrawal of both live and dead adult weevil from the previous experiment (weevil mortality test). The adult weevil (F1) progeny emerging from each treatment were counted and recorded to find a measure of effects of both powder on weevil and reproduction. They were sieved off subse-

3 Journal of Science, Engineering and Technology 1(2), September Table 1: Effect of Asystasia vogeliana powder on adult Callosobruchus maculatus mortality at 24, 48, 72, and 92 hours. Plant Con. mg Time A. vogeliana 24hours 48 hours 72 hours 96 hours Control (untreated) No. of Mean ± SE ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 0.57 quently daily to prevent mating and subsequent oviposition by F1 as mating in C. maculatus does not occur before weevils are 3 days old (Walgenbach and Buckholder, 1987). Recording of emergence of C. maculatus was terminated on 11 th week. Weevil perforation The extent of weevil damage was assessed using the exit-hole counted as a measure of damage to the grains after 11 weeks treatment. Grains that were riddle with exit-hole were counted, the percentage damage (PD) was calculated using the method in Adedire and Ayaji (1996). The percentage damage (PD) = Total no. of treatment grain perforated x 100 Total no. of grain x 1 Photochemical screening of plant material Several standardized extraction procedures for plant metabolites have been carried out by various researchers such as Audu et al (2007), Alinnor (2008), Njoku and Obi (2009), Roy (2010), Wang et al (2010), Obasi et al (2010) and Prashant et al (2011). The phytochemical screening of A. vogeliana material was done in Cross River University of technology using the different standardized methods. The methods of Sofowara (1993), Harbone (1973) and Trease and Evans (1989) were used for the identification of Alkaloids, Flavonoids, Tannin, Glycosides, Saponins, Polyphenol, anthraquinones, Steroids and reducing compounds present in the plant. Table 2: Effect of Asystasia vogeliana powder on C. maculatus emergence after 77 days. Treatment Conc. (mg) Number of Adult emergence Percentage Emergence Mean adult emergence ± ± ± ± 0.09 Results The effect of the plant powder on adult Callosobruchus maculatus mortality is presented in table 1. At 24 hours exposure and 1mg of plant powder, adult C. maculatus died, giving a mean death rate of and a standard error of The recorded after 48 hours were adults of C. maculatus at the same concentration, with a mortality rate of and a standard error of Also at 72 hours exposure, were recorded, with a mean death rate of and a standard error of However, at 96 hours exposure of C. maculatus in 1mg powder 400 adults died, giving a mean death rate of 100 and a zero standard error. At 3mg concentration of powder, 240 adults of C. maculatus died in 24 hours, giving a mean death rate of 60, with a standard error of At 5mg concentration, the same order of was observed. Adult C. maculatus mortality rate significantly increased in concentration according to the number of days of exposure. The least mean mortality of ±0.3 (mean ± SE) was observed in the first day of exposure of concentration of 1.0mg. The highest mean mortality value of ± 0.00 (mean ± SE) was observed in the third and fourth days in 5mg concentration at the plant powder. There was no mortality recorded among Callosobruchus maculatus during the first day of their exposure in the control experiment. Table 2 illustrates the effect of Asystasia vogeliana powder on C. maculatus emergence after eleven weeks. At zero concentration of A. vogeliana, one hundred and eight (108) adults of C. maculatus emerged with a mean adult emergence of ± 0.27 (mean ± SE). It was observed that at 3mg concentration 21 adults of C. maculatus emerged, having a mean adult emergence of 5.23 ± 0.41 (Mean ± SE). However, only 10.6 adult of C. maculatus emerged at 5.0mg concentration of A. vogeliana powder, and had a mean adult emergence of 2.65 ± 0.09 (Mean ± SE). The number of emerged adult decreased with increase in concentration of powder. The least number of 10.6 adults emergence was recorded at 5.0mg concentration of A. vogeliana. Discussion The Efficacy of Asystasia vogeliana powder to control Callosobruchus maculatus has been estab-

4 98 IBOH, C. I. ET AL: Insecticidal Potency of Asystasia Vogeliana on Bean Seeds... lished in this study. It was observed that there was no mortality of C. maculatus recorded in the first day of the control experiment. This was so because the plant powder appeared to be very potent especially at higher concentration because of their choky odours, which may have exerted a toxic effect, discouraging feeding by the weevil. This might be due to the effect of alkaloids, flavonoids and saponins present in the powder (Obamanu et al, 2005). Saponins and alkaloids inhibits phosphorylation and blocks the mitochondrial enzymes, hence impairing oxygen consumption (Singh et al, 2005). It was held that the powder potency to kill might have stemmed from the fact that their offensive odours acted as an antifeedant. This could explain what might have led to the starvation of some of the weevils as the day passed by. The application of A. vogeliana powder on grains, covered the outer layer of these grains and this might have influenced the suppression of oviposition and larva development. A. vogeliana is the second plant discovered to have antifeedant, bacteriocides, fungicides and nematicide properties, which have positive impact on usage by man (Singh et al, 2005). These properties might be responsible for the oviposition and developmental suppression of these weevils. The effectiveness of the tropical plant powder to control C maculatus which is the most economically important insect pest of stored bean in Africa and Nigeria in particular has been established in this investigation. The study revealed that the potency of the plant powder was dependent on the dosages applied and the number of days C. maculatus was exposed. The authors suggest for more investigation to ascertain these observation as well as the health implications of using the powder in storing food products. ACKNOWLEDGEMENT We are grateful to Dr. Okoi, A. I. (the immediate past HOD of Biological Sciences, CRUTECH), and Curators Ekpiken, Emmanuel E. and Akiba, Ekpenyong E. in the Herbarium Unit of CRUTECH, who identified the plant A. vogeliana used in this investigation. The phytochemical screening of A. vogeliana in the Chemistry Department of CRUTECH by Mr. Iboh, Iwara Edet (Senior Technologist) is highly appreciated. REFERENCES Adedire, C. O. and Ayaji, T. (1996). Toxicity and oviposition deterrency of some plant extract as grain protection against the bean weevil Callosobruchus maculatus (mots). Nigeria Journal of Entomology, 13: Alinnor, I. J. (2008). Prelimanry phytochemical and antibacterial activity screening of leaves of Varnonia amygdaliana. Journal of Chemistry and Sociology, Nigeria. 33 (1): Audu, S. A., Mohammed, I., Kaita, H. A. (2007). Phytochemical Screening of the leaves of Laphira lanceolata (Ochanaceae). Life Science Journal 4(4): Buckholder, W. E. and Ma, M. (1985). Pheromones for monitoring and control of stored product insects. Annals Review of Entomology. 30: Dolmen, D. Arnkleiv, J. V. and Haukebo, T. (1995). Rotenone tolerance in the freshwater pearl mussel Margaritifera margaritifer. Nordic Journal of Freshwater Research. 70: Ghaffar, M. A. and Chanhan, Y. S. (1999). Polarization to protect pigeonpea seed from bruchid damage during storage. International chickpea and pigeonpea newletter. 6: Golob, P. and Wisley, O. O. (1980). The use of plant and minerals as protectants of stored product. Tropical Product Institute G 138. Postharvest test and avidity section natural resource institute, Lanthan, Uk pp. 32 Harbone, J. B. (1973). Phytochemical method, Chapman and hall Ltd, London pp Hassanli, A. Bekele, A. J. and Obeng-Ofori, D. (1997). Evolution of Ocimum kenyense (Agobangiral) as source of repellents, toxicants and protectant in storage against three major stored product insect pests. Journal of Applied Entomology. 121: Livinson, H. and Livinson, A. (19994). Origin of grain storage and insect species consuming defecated food. Anzifurschsdi. 67:47-59 Njoku, O. V. and Obi, C. (2009). Pytochemical constituents of some selected medicinal plants. African Journal of Pure and Applied Chemistry 3(11): Obasi, N. L., Egbuonu, A. C. C., Ukoha, P. O., and Ejikeme, P. M. (2010). Comparative phytochemical and antimicrobial screening of some solvent extracts of Samanea saman pods. African Journal of Pure and Applied Chemistry. 4 (9): Obomanu, F. G. Fekarurhobo and Howard, I. C. (2005). Antimicrobial activities of extracts of leaves of Asystasia vogeliana (Vahl). Journal of Chemical Society of Nigeria. 30: P Pantherius, C. U. (1999) Storage waste in traditional bean granaries in Togo. Insect sciences and it Application. 9:

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