Introduction
The major problem facing seeds storage in Nigeria and other African countries are insect pests, leading to loss in weight and reduction in seeds quality (Akinkurolere et al., 2006; Adedire et al., 2011; Akinneye and Ogungbite, 2013; Ashamo et al., 2013; Ojo and Ogunleye, 2013a; b). Crop protection therefore plays a vital and integral role in modern agricultural production. The ever-lasting demands on yield and the intensification of farming practices have increased the problem of pest damage, and hence control (Martin et al., 2012).

The most common means in insect pest control are the use of synthetic insecticides whose effects on the environment can cause water, soil, and atmospheric genetic perturbations related to cancer amongst users of these pesticides. Public awareness of these adverse effects of chemical insecticides especially on the environment has lead researchers to find new avenue of insect control in order to replace these dangerous synthetic chemical insecticides (Sutherland et al., 2002; Zibaee, 2011). So researches has been concentrated on the plant kingdom for solutions leading to the production of a myriad of secondary compounds that can have toxic, growth reducing and antifeedant properties against insects (Zibaee, 2011). In spite of the effectiveness of many botanical insecticides, their insecticidal activity is yet to be comparable to many synthetic chemical insecticides and the once that are believed to be comparable with chemical insecticides have not commanded more than 1% of the global insecticide market (Isman, 2000; Begum et al., 2013) because they are believed to be short live and loss their potency over time (Oruonye and Okrikata, 2010). For these reasons, there is need to search for other plants which could comparably contend with synthetic chemical insecticides since tropical region of the world including Nigeria are well endowed with numbers of plant species that could have insecticidal properties. Therefore, this study investigated the bio-efficacy of crude extracts from Cheese wood, A. boonei as a surface protectant against cowpea bruchid, C. maculatus, attacking cowpea seeds in storage.

1 Material and Methods
Newly emerged adult C. maculatus used for this study were obtained from already existing culture in the Postgraduate Research Laboratory of the Department of Biology, Federal University of Technology, Akure, Nigeria. They were subsequently reared inside 1 litre Kilner jars, on un-infested cowpea seeds Vigna unguiculata variety Ife brown obtained from International Institute for Tropical Agriculture, Ibadan, Nigeria. The culture was placed in an insect rearing cage at ambient temperature of (28+2)℃ and (75+5)% relative humidity.

Leaf, stem bark and root of Alstonia boonei used for this study were sourced fresh from Akola farm at Igbara-Odo Ekiti, Ekiti State, Nigeria. These plant parts were rinsed in clean water to remove sand and other impurities, cut into smaller pieces before air-dried in the laboratory. The cleaned dried plant parts were pulverised into very fine powder using an electric blender (Supermaster ®, Model SMB 2977, Japan). The powders were further sieved to pass through 1mm2 perforation. The powders were packed in plastic containers with tight lids and stored in a refrigerator at 4℃prior to use.

For extraction by the kneading method, 800 g of each of the pulverised plant parts were weighed into a plastic bowl. Pre-boiled water was added gradually, a little at a time, from a kettle and mixed with A. boonei powders. The procedure was repeated until a dough-like material was formed. The extract of A. boonei leaf, stem bark and root were pressed out manually with hand and poured into a 250 ml volumetric flask and stored at room temperature. From this stock, 0.3 ml of A. boonei leaf, stem bark and root extracts were used.

0.3 ml of each extracts of A. boonei was mixed separately with 20 g of un-infested cowpea seeds in 250 ml plastic containers. The extracts and seeds were thoroughly mixed using a glass rod and then agitated for 5~10 min to ensure uniform coating. Control experiments were also set up. Ten pairs of teneral adult C. maculatus were introduced into each of the containers and covered. Four replicates of the treated and untreated controls were laid out in Complete Randomized Design in insect cage. Beetle mortality was observed daily for 4 days. The beetles were confirmed dead when there was no response to probing with sharp pin at the abdomen. The total number of eggs laid per replicate was recorded after 4 days post treatment. Percentage adult mortality was corrected using Abbott (1925) formular thus:
            P_O-P_C       100
P_T =—————×    ——
          100 – P_C       1
Where P_T=corrected mortality (%)
             P_O=observed mortality (%)
             P_C=control mortality (%)

The experimental set up was kept inside the insect rearing cage for further 30 days for the emergence of the first filial (F1) generation. The percentage number of adult beetle emergence was calculated according to the method described by Odeyemi and Daramola (2000).

The percentage reduction in adult emergence of F1 progeny or inhibition rate (IR) was calculated according to the method described by Tapondju et al. (2002).
             C_n-T_n     100
% IR=————×  ——
                 C_n         1

Where Cn is the number of emerged insects in the control and Tn is the number of emerged insects in the treated container.

Percentage weight loss of the cowpea seeds was determined by re-weighing after 35 days and the % loss in weight was determined as follows:
                                Difference in weight        100
% weight loss= ———————————×           ——
                                     Initial weight                  1

After re-weighing, the numbers of damaged cowpea seeds were evaluated by counting wholesome seeds and seeds with bruchid emergent holes. Percentage seed damaged was calculated as follows:
                                       No. of seeds damaged with emergent hole     100
% seed damaged= ————————————————————×                  ——
                                                     Total number of seeds                          1

Beetle Perforation Index (BPI) used by Fatope et al. (1995) was adopted for the analysis of damage. Beetle perforation index (BPI) was defined as follows:
            % treated cowpea seeds perforated       100
BPI=——————————————————×             ——
             % control cowpea seeds perforated         1

BPI value exceeding 50 was regarded as enhancement of infestation by the weevil or negative protectability of the extract tested.

Cowpea seeds weighing 20 g each that have been previously treated with A. boonei extracts for 30, 60 and 90 at rate 0.3ml was used for this experiment. Similar parameters described were assessed for the persistent toxicity A. boonei extracts of to cowpea bruchid.

Data were subjected to analysis of variance (ANOVA) and treatment means were separated using the New Duncan’s Multiple Range Test. The ANOVA was performed with SPSS 16.0 software (SPSS, Inc. 2007).

2 Results
Table 1 shows and the effects of contact toxicity of A. boonei leaf, stembark and root extracts obtained by kneading method on adult mortality of C. maculatus after 1, 2, 3 and 4 days of infestation and 30, 60 and 90 days of post treatment. The stem bark extract completely evoked 100% mortality of adult C. maculatus at concentration tested after 3 days of application. This is followed by leaf extract causing 100% mortality of adult cowpea bruchid while the least toxic was root extract evoking 87.50% adult C. maculatus after 4 days of application (Table 1).
 

Table 1 Mortality of adult C. maculatus in cowpea seeds treated with 0.3 v/w A. boonei extracts as contact insecticide obtained by Kneading method

Stem bark extract caused 70% adult mortality of C. maculatus at rate 0.3 ml/ 20g of cowpea seeds after 30 days of post treatment. This is followed by leaf extract causing 57% mortality of cowpea bruchid at rate 0.3ml per 20g of cowpea seeds after 30 days of post treatment. The least toxic extract was root evoking 37.5% adult C. maculatus at rate 0.3ml per 20g of cowpea seeds after 30 days of post treatment (Table 2).
 

At 60 days of post treatment, 60% adult mortality of cowpea bruchid was recorded on cowpea seeds treated with stem bark extract. At 90 days of post treatment, 42% adult mortality of cowpea bruchid was recorded on cowpea seeds treated with stem bark extract at rate 0.3 ml. Generally, percentage adult mortality of C. maculatus decreases with length of storage period (Table 2). This shows that A. boonei stem bark had the greatest insecticidal potential, while A. boonei the least effective was A. boonei root.
Tables 3 and 4 shows the effects of A. boonei leaf, stem bark and root extracts obtained by kneading method on oviposition, adult emergence and reduction in progeny development of adult C. maculatus after 4 days of infestation, 30, 60 and 90 days of post treatment. Stem bark extract completely protected cowpea seeds from being damaged and 100% reduction in progeny development after 4 days of application (Table 3). Leaf extract of A. boonei completely protected cowpea seeds and 100% reduction in progeny development at rate 0.4 ml after 4 days of application. The results obtained on cowpea seeds treated with root extract were significantly different from untreated cowpea seeds (Table 3).
 

 
At 30 days of post treatment, A. boonei stem bark extract reduced the number of eggs laid by cowpea bruchid and 100% reduction in progeny development of adult C maculatus (Table 4). Generally, the number of eggs laid, % adult emergence and reduction in progeny development of adult C. maculatus increases with increase in storage periods but significantly different (P<0.05) from untreated cowpea seeds (Tables 4).
 

Percentage seeds damaged, weigh loss and Beetle Perforation Index caused by C. maculatus in cowpea seeds treated with leaf, stem bark and root extracts of A. boonei obtained by Kneading method were shows in Table 5 and Table 6. There was neither seed damage nor weight loss recorded in the cowpea seeds treated with stem bark extract and BPI was zero after 4 days of application. The BPI of 1.42 was obtained on seeds treated with A. boonei leaf extract. However, these values were not significantly different from BPI of the treated cowpea seeds with root extract after 4 days of application but significantly different (P<0.05) from untreated (Table 5). 
 

At 30 days of post treatment, stem bark extract completely protected the seeds from being damaged by C. maculatus at the concentration tested. There was neither seed damage nor weight loss recorded in the cowpea seeds treated with stem bark extract and BPI was zero for the concentration tested after 30 days of application (Table 6). At 60 days of post treatment, stem bark extract completely protected the seeds from being damaged by cowpea bruchid, C. maculatus. Generally, % seeds damaged, % weight loss and Beetle Perforation Index by adult C. maculatus increases with increase in storage periods (Table 6). As a result of the feeding activity of C. maculatus larvae on the cowpea seeds, the % seeds damaged, % weight loss and Beetle Perforation Index of the untreated cowpea seeds was significantly (P<0.05) reduced compared with the treated seeds.
 

Table 6 Percentage seeds damaged, weigh loss and Beetle Perforation Index caused by C. maculatus in cowpea seeds treated with 0.3 v/w extract of A. boonei obtained by Kneading method after 30, 60 and 90 days of post-treatment

 3 Discussion
Previous studies by Ileke and Oni (2011); Ileke et al. (2012); Ileke et al. (2013); Ileke et al. (2014) have shown insecticidal activity of A. boonei powders against Sitophilus zeamais, A. boonei stem bark oil obtained by cold extraction using five different solvents on the mortality of cowpea bruchid, C. maculatus and A. boonei latex against C. maculatus. In this study, A. boonei stem bark extract was the most toxic to adult C. maculatus. The insecticidal potential of this plant extracts on the cowpea bruchid could be as a result of the presence of some chemical compounds of the triterpenoids, indole and alkaloid group such as alstonine, astondine, and porphine that have been identified from A. boonei (Phillipson et al., 1987; Anonymous, 1992; 2001; Moronkola and Kunle, 2012; Ojo and Ogunleye, 2013a; b). A. boonei stem bark extract completely protected cowpea seeds against C. maculatus infestation and also prevented oviposition, adult emergence and reduction in F1 progeny of C. maculatus. The effect of the extract on oviposition could be due to respiratory impairment which probably affects the process of metabolism and consequently other systems of the body of the bruchid (Osisiogu and Agbakwuru, 1978; Onolemhemhem and Oigiangbe, 1991; Adedire et al., 2011; Ileke et al., 2013a; b; Ojo and Ogunleye, 2013b). Extracts have been reported to inhibit locomotion (Adedire et al., 2011; Ileke et al., 2013); hence, the beetles were unable to move freely thereby affecting mating activities and sexual communication (Adedire et al. 2011; Ileke et al., 2012). The inability of the eggs to stick to the cowpea seed due to the presence of the extracts also reduced adult emergence arising from egg mortality (Adedire et al., 2011; Ileke et al., 2013a). It has been reported that one of the main mechanisms of action of plant extracts is their ability to penetrate the chorion of bruchid eggs via the micropyle and cause the death of developing embryos through asphyxiation (Don–Pedro, 1989a; b). Reduction in progeny development may be due to early mortality and partial or complete retardation of embryonic development (Dike and Mbah, 1992).

The protectability of the extracts was highly remarkable. The contact toxicity bioassay of A. boonei extracts on adult C. maculatus observed at 30, 60 and 90 days after treatment revealed the insecticidal potency of A. boonei stem bark extract. This shows that the A. boonei stem bark extract can protect stored cowpea conveniently for 90 days. Some botanicals have been tested for long term protectability on cereals and legumes (Shaaya et al., 1997; Pereira, 1983; Ajayi and Adedire, 2003; Oni, 2009). Shaaya et al. (1997) tested the toxicity of palm kernel and rice bran oils on chick peas and mung beans against C. maculatus infestation at rates 1.5 g and 3 g/kg cowpea seeds and reported full protection for a period of 4 to 5 months. Pereira (1983) studied the effectiveness of six vegetable oils as protectants of cowpea seeds and bambara groundnuts against C. maculatus at rate 8 ml/kg. He reported a good protection for a period of 3 to 6 months. Ajayi and Adedire (2003) studied the protectability of sub-ledal doses of Hura crepitans seed oil as cowpea protectant against C. maculatus at concentrations 0.25, 0.5 and 1% v/w protected seeds from been damage by C. maculatus for a period of 3 months. Oni (2009) reported the effectiveness of three local cultivars of pepper, Capsicum species as cowpea protectants at concentrations 0.5 ml/ 20g, 0.1 ml/ 20g, 2.0 ml/ 20g, 5.0 ml/ 20g and 10 ml/ 20g of cowpea seeds. She reported a good protection for a period of 6 months. All tested concentrations of A. stem bark extract gave protection for a period of 90 days. The insecticidal potential of A. boonei leaf and root oil extracts was greatly reduced on mortality of adult C. maculatus at 30, 60 and 90 days of post treatment. This was in agreement with Adedire and Ajayi (1996) who reported that the toxicity of some plant extracts used for control of weevils did not persist indefinitely, some extracts lost their effectiveness within 95 days of treatment. Oni (2009) also reported that the oil extract of three local cultivars of pepper, Capsicum species used in protecting maize grains against Sitophilus zeamais, lost their potency after 6 months in storage.

4 Conclusions
This study has further revealed the insecticidal potential of A. boonei extracts obtained by kneading method as cowpea protectant against C. maculatus and could serve as an alternative to synthetic insecticides for the protection of stored cowpea seeds against C. maculatus or other related insect pests by local farmers that could not afford or have access to soxhlet extraction apparatus and rotary evaporator. The plant is medicinal, readily available, safe, eco – friendly, biodegradable and they have not been reported to be toxic to man.

Acknowledgment
Thanks are due to the Government of Ekiti State, Nigeria for financial support through the PhD scholarship awarded to the first author, Kayode D. Ileke tenable at the Federal University of Technology, Akure, Nigeria in 2009.

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