BIOLOGICAL ACTIVITY OF CANDLE WOOD (ZANTHOXYLUM ZANTHOXYLOIDES (LAM) ZEPERNICK &TIMLER) AGAINST COWPEA BRUCHID, CALLOSOBRUCHUS MACULATUS (FABRICIUS)

Abstract

Botanicals have become the focus for discovery of novel bioinsecticides for protection of crops most especially because of their biodegradability, non-toxicity, target organism specificity and environmental friendliness. Partitioned extracts of Zanthoxylum zanthoxyloides were investigated for biopesticidal effect on Callosobruchus maculatus with aim of identifying the most bioactive components and increase ease of handling when compared to bulk crude plant material used by farmers. Crude powder of rootbark of Z. zanthoxyloides was made as well as partitioned extracts using Kupchan partition extraction methods with methanol, acetone, ethyl acetate and n-hexane as solvents. Analysis of the rootbark of the plant showed 19.87% ash content, 24.00% crude protein, 24.85% crude fibre, 3.13% oil extract, 15.42mg/100g alkaloids, 45.90mg/100g tannins, 1039.14mg/100g saponins, 26.47mg/100g flavonoids, 150.0mg/100g iron, 244.70mg/100g calcium, 36.65mg/100g magnesium, 449.00mg/100g potassium and 128.30mg/100g of phosphorus. The crude powder and extracts were bioassayed on C. maculatus in stored cowpea. Insecticidal activities of the partitioned extracts increased with increasing concentration; toxicity test revealed that n-hexane at 1%, 2% and 3%, acetone extract at 3%, and crude powder at 1.5g all have same effect on the insect as the positive control, achieving 100% mortality within 24 hours post treatment. Number of adult emergence was zero in n-hexane treatment. There was a significant difference (P>0.05) in weight loss by the cowpea in the different fractional treatments while positive and negative controls also differed significantly (P>0.05) with all the partitioned treatments. Partition extracts from Z. zanthoxyloides were more bioactive than the crude powder, and n-hexane fractions contain the most active compounds against C. maculatus. Toxicological studies were carried out to investigate the mammalian toxicity of Z. zanthoxyloides root bark widely used in traditional medical practice and as protectant of stored cereal products in Nigeria, using Wistar albino rats. Serum, kidney and liver were tested for oxidation stress and tissue damage markers; aspartate amino transferase (AST), alanine amino transferase (ALT), urea, bilirubin and creatinine contents, and Kidney and liver glutathione peroxidase (GPx), glutathione transferase (GST), nonprotein sulphadryl (NP-SH), thiobarbituric acid reactive substances (TBARS), total sulphadryl (T-SH) and superoxide dismutase (SOD). These tests were carried out using commercially available kits. Results obtained for all doses (1, 5 and 10%) of the tested parameters did not significantly vary from the control (p>0.05), indicating normal liver and kidney function (even in the face of variation in liver GPx and SOD). Bioassay guided fractionation of n-hexane extract of Z. zanthoxyloides revealed 27 fractions labelled 4-30 and these were tested for toxicity against C. maculatus, fractions 10-14 (fraction A) and 18-24 (fraction B) were found to be most toxic; resulting in mortality rate of 80-100% within 48 hours post treatment. The insects were exposed to 0.00, 1.00, 2.00, 3.00, 4.00 and 5.00 % W/V of the fractions for 24 hours, survivors were collected and homogenised separately, the supernatants obtained served as source of enzymes for bioassay to determine the activity of superoxide dismutase (SOD), Catalase (CAT), glutathione peroxidase (GPx), acetylcholinesterase (AChE), Carboxylesterase (CarEST), glutathione transferase (GST) and Total protein in C. maculatus. There was a general increase in the activities of the enzymes at lower doses and decrease at higher doses in both fractions, except in fraction B where CAT activity increases with increasing concentration and AChE activity decreased significantly (p>0.05) with increasing concentration of the fraction. The fractions were further subjected to Gas Chromatography – Mass Spectrometry (GC-MS) analysis to determine its bioactive compounds and their insecticidal activity. GC-MS result revealed 55 compounds from both fractions out of which Dibutyl phthalate (11.75%), Hexadecenoic acid (5.78%), Heneicosane (3.75%), Heptadecane (3.35%) and Hentriacortane (2.39%) have proven insecticidal activity. This work has proven the insecticidal potential of Z. zanthoxyloides, its mode of action on C. maculatus and revealed some of its bioactive compounds. It showed that n-hexane extract contains most active biomolecules to be explored for bioinsecticide formulation with high potency for development of new biopesticide, eliminate toxic components and increase ease of handling when compared to bulk crude plant material used by farmers. This study has also proven that the plant is safe for human consumption at the rates or dosages tested.