Silphinene Derivatives: Their Effects and Modes of Action on Colorado Potato Beetle Azucena Gonza ´ lez-Coloma,* ,† Carmen Gutie ´rrez, † Raimundo Cabrera, ‡ and Matı ´as Reina § Centro de Ciencias Medioambientales, CSIC, Serrano 115-dpdo., 28006 Madrid, Spain, UDI Fitopatologı ´a, Facultad de Biologı ´a, Universidad de la Laguna, Tenerife, Spain, and Instituto de Productos Naturales y Agrobiologı ´a, CSIC, Avda. Astrofisı ´co F. Sanchez, 38206 La Laguna, Tenerife, Spain The silphinene derivatives 11-hydroxy-5R-(angeloyloxy)silphinen-3-one and 11,5R-dihydroxy- silphinen-3-one were generated by means of chemical hydrolysis of the natural antifeedant and toxic silphinene 11-acetoxy-5R-(angeloyloxy)silphinen-3-one and bioassayed against Colorado potato beetle larvae (Leptinotarsa decemlineata). Both compounds showed significant antifeedant activity against this insect in choice and no-choice assays. Futhermore, exposure of larvae to these compounds over a 24 h period resulted in reduced feeding and growth rates. To distinguish between antifeedant and toxic effects, growth efficiencies were calculated as the slope of the regression of relative growth rate on relative consumption rate. The comparison of these results with those of antifeedant simulation bioassays indicates that strong feeding inhibition is associated with the 11- acetate substituent, while negative effects on larval growth through contact toxicity are related to the lipophilicity of the compounds. These substances were also bioassayed against three species of the plant pathogen Fusarium. The natural silphinene and the two derivatives showed mild antifungal activity inversely related to their polarity. Keywords: Leptinotarsa decemlineata; Coleoptera Chrysomelidae; antifeedant; toxic; silphinene derivatives INTRODUCTION The silphinene 11-hydroxy-5R-(angeloyloxy)silphinen- 3-one is a sesquiterpene which is relatively abundant in the plant Senecio palmensis Chr. Sm. (Compositae), an endemic species to the Canary Islands. This com- pound is a potent antifeedant against the Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say.) (Coleoptera Chrysomelidae), with an additional toxic effect (Gonza ´ lez-Coloma et al., 1995). Antifeedants are gaining importance as potential components of integrated pest management strategies for insect control. In addition to reduced feeding on plants, some of these substances have biological effects such as toxicity, oviposition deterrence, and growth- regulating activity (Zehnder and Warthen, 1988; Liu et al., 1989, 1990). They can also enhance the activity of other insect control agents (Murray et al., 1993). The CPB is a major potato pest in North America and Europe. The chemical control of this pest with synthetic insecticides has induced a rapid development of resist- ance of the beetle populations to most of these chemi- cals, including some natural insecticides such as aver- mectins and the δ-endotoxin of Bacillus thuringiensis (Brattsten, 1991; Whalon et al., 1993). One approach to this problem will be to use insect antifeedants that exert both behavioral (antifeedants) and physiological (toxins) effects (Jermy, 1990). Some examples of such substances are the citrus limonoids limonin, obacunone, and nomilin (Mendel et al., 1991). When dealing with natural substances, compound availability is a major problem. The synthetic trans- formation of natural product models can lead to a better understanding of their mode of action and can also help to overcome the economic problem of compound avail- ability. Model antifeedants have been synthesized based on limonoids such as azadirachtin, salanin, and limonin or the triterpene betulin (Ley et al., 1987; Yamasaki and Klocke, 1989; Bentley et al., 1990; Huang et al., 1995). In this work, we describe the preparation of two structurally related derivatives of a natural CPB anti- feedant and toxic silphinene isolated from Senecio palmensis (Gonza ´ lez-Coloma et al., 1995), and we study their mode of action on CPB larvae. We also describe their toxicity against a different biological system, the plant pathogen Fusarium, and we compare their struc- tures and activity with the parent compound. MATERIALS AND METHODS General. IR spectra were recorded in CH2Cl2 solutions using a Perkin-Elmer spectrometer. 1 H and 13 C NMR spectra in CDCl3 were measured with Bruker spectrometers: WP-200 SY (200 MHz) and AMX (400 MHz) (chemical shifts reported are relative to residual CDCl3, 7.26 ppm for 1 H and 77.0 ppm for 13 C). Mass spectra and elemental analysis were obtained on Hewlett Packard 5995 and Fisons EA 1108 instruments, respectively. Hydrolysis of 11R-Acetoxy-5-(angeloyloxy)silphinen- 3-one (1). A 400 mg portion of compound 1, isolated from S. palmensis (Gonza ´ lez-Coloma et al., 1995), was dissolved in 25 mL of distilled MeOH. After the addition of 10 mL of 35% methanolic KOH, the reaction mixture was agitated for 48 h at room temperature and monitored with TLC chromatogra- phy. The resulting mixture was poured on a 2 N H2SO4 solution and extracted with CH2Cl2. The organic layer was then extracted with NaCl and H2O, dried on Na2SO4, and vacuum concentrated. The crude residue (315 mg) was then chromatographed on a silica gel column eluted with a gradient of n-hexane:ethyl acetate (81:15-0:100), to give the following compounds: the substrate compound 1 (50 mg), a partially * Author to whom correspondence should be ad- dressed (e-mail, azu@cc.csic.es). † Centro de Ciencias Medioambientales. ‡ Universidad de la Laguna. § Instituto de Productos Naturales y Agrobiologı ´a. 946 J. Agric. Food Chem. 1997, 45, 946-950 S0021-8561(96)00574-2 CCC: $14.00 © 1997 American Chemical Society