Antifeedant Effects of the Limonoids from Entandrophragma candolei (Meliaceae) on the Gram Pod Borer, Helicoverpa armigera (Lepidoptera: Noctuidae) OPENDER KOUL,* ,² WLODZIMIERZ M. DANIEWSKI, ‡ JATINDER SINGH MULTANI, ² MARIA GUMULKA, ‡ AND GURMEET SINGH ² Insect Biopesticide Research Centre, 30 Parkash Nagar, Jalandhar 144 003, India, and Institute of Organic Chemistry, Polish Academy of Sciences, Kasprzaka 44, 01-224 Warsaw, Poland The biological activity of the limonoids prieurianin and epoxyprieurianin isolated from Entandrophragma candolei (Harms) (Meliaceae) and their respective acetates was assessed using the gram pod borer, Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae). The compounds exhibited strong antifeedant activity in a diet choice bioassay with epoxyprieurianin acetate being most effective with 48.3 ppm deterring feeding by 50% (DI 50 ) and prieurianin the least effective (DI 50 ) 91.4 ppm). The effect on growth of larvae was concomitant with the reduced feeding by neonate and third instar larvae. In nutritional assays, all the compounds reduced growth and consumption when fed to larvae without any effect on efficiency of conversion of ingested food (ECI), suggesting antifeedant activity alone. No toxicity was observed nor was there any significant affect on nutritional indices following topical application, further suggesting that prieurianin-type limonoids act specifically as feeding deterrents. KEYWORDS: Entandrophragma candolei; prieurianin; epoxyprieurianin; prieurianin acetate; epoxy- prieurianin acetate; antifeedants; Helicoverpa armigera INTRODUCTION The Meliaceae family of plants has shown great potential for pest management in terms of secondary plant chemistry or the presence of allelochemicals in its various genera (1). These allelochemicals have considerable potential as antifeedants or biopesticides (2). The idea of using nontoxic feeding deterrents as crop protectants has attracted much attention during the past decade, but many antifeedants also show some post ingestive physiological activity (3). Azadirachtin from Azadirachta indica is the most potent insect feeding deterrent and natural insect growth regulator isolated to date. It occurs at concentrations of 0.1 to 0.9% in the seed kernel and it has been established that 30 to 60 g azadirachtin per hectare is sufficient to combat and repel key pests of various crops (4). As an insecticide, azadirachtin-based products control more than 400 species of insects including those in the insect orders Lepidoptera, Co- leoptera, Homoptera, Diptera, Heteroptera, Caelifera, and Thys- anoptera, etc. It is also well-known now that the biopesticide potential of neem in particular, and the family Meliaceae in general, is due to the presence of characteristic limonoid-type compounds. Many oxidative products in various species impart an edge to Meliaceae plants as their biological activity seems to relate to current concepts of the evolution of the limonoids (5). In searching for antifeedants of natural origin we decided to investigate Meliaceous plants, apart from Melia species, and accordingly selected Entandrophragma candolei (Harms) for investigation against a lepidopteran pest. Extracts from the bark of this tree are known to inhibit feeding in stored grain pests (6). In an earlier preliminary study of prieurianin isolated from Nymania capensis, antifeedant activity was demonstrated in a leaf-disc choice assay against tobacco budworms and Mexican bean beetles (7). MATERIALS AND METHODS Chemistry. The bark of the tree Entandrophragma candolei (Harms) was collected in the primeval forest in Zaire near Kisangani in 1990, shade-dried, and kept in stock for further processing and chemical investigations. Prof. F. Szafranski, University of Kisangani, identified the trees (8). The shade-dried bark was pulverized into a powder and subsequently subjected to sequential solvent extraction. A choloroform extract (1.5 g) of the material, found to possess activity against insects, was subjected to column chromatography on silica gel using a benzene/ acetone gradient mobile system. The chromatography was monitored by TLC (C6H6/Me2CO, 4:1 solvent system) and yielded 380 mg of prieurianin and 910 mg of epoxyprieurianin. To produce analytical samples, final purification was performed using a semi-prep HPLC column (30 cm × 0.8 cm) filled with 10-µm Nucleosil C-18 RP. The mobile phase was a mixture of acetonitrile, methanol, and water (58: 15:27). The main peaks were collected and evaporated to dryness, and finally the residue was subjected to spectral investigation, the results of which compared well with that of Gullo et al. (9) and Lukacova et al. (10). * To whom correspondence should be addressed. E-mail: koul@ jla.vsnl.net.in. ² Insect Biopesticide Research Centre. ‡ Polish Academy of Sciences. J. Agric. Food Chem. 2003, 51, 7271-7275 7271 10.1021/jf0304223 CCC: $25.00 © 2003 American Chemical Society Published on Web 10/29/2003