In the course of screening soil organisms for new insecticidal metabolites, strain W719was found to produce a group of metabolites active against the tobacco budwormHeliothis virescens. The active metabolites were purified by a combination of solvent partitioning and chromatographic steps, and the physico-chemical properties and insecticidal activity of the main components were determined. The two main components have MW'sof925 and 939, appear to belong to the macrocyclic lactam family of natural products, and possess significant insecticidal activity.In a program for the discovery of novel natural products for use in crop protection, a tobacco budworm{Heliothis virescens) laboratory diet feeding assay was established to screen for insecticidal metabolite production by 5,000 soil microorganism isolates. Amongthe actives was strain W719, which produced a complexof novel lipophilic insecticidal metabolites. The crude culture extract and purified complex demonstrated moderate residual contact activity in foliar assays against tobacco budwormand beet armyworm,Spodoptera exigua. This paper outlines the purification, spectroscopic properties, and insecticidal activity of these metabolites.
ExperimentalStrain Isolation and Storage Strains used in the insecticidal screening program were isolated at the laboratory of Dr. Jerry Ensign, University of Wisconsin, Madison on colloidal chitin agar with 50^g/ml cycloheximide from various soil and rhizosphere samples1*. Strain W719was not speciated, but is presumed to be a strain of Streptomyces. The strain was subcultured on Oatmeal agar (Difco) with cycloheximide at 50 /ig/ml and passaged at 28°C. Freezer stock cultures were madeby suspending whole cultures or spore preparations from agar slants in 10% glycerol, flash freezing, and storing at -70°C.
CultivationThe 7 day growth of a 15-ml oatmeal agar slant (50^g/ml cycloheximide) was suspended in 5ml sterile deionized water, heat shocked at 45°C for lOminutes1*, and used to inoculate 10 new 15ml slants, one 100 ml slant, or one 500-ml tribaffled flask with a loose fitting plastic cap containing 150 ml of medium (glucose lOg, soluble starch 20g, Amber EHC (Amber Laboratories, Milwaukee, Wisconsin) 2g,Bacto-casitone 2g, Bacto-yeast extract 2g and Bacto-beef extract 2g, CaCO3 3 g, and deionized H2O 1,000ml). Flasks were incubated at 28°C with shaking at 150rpm in a G25 NewBrunswick platform shaker. For screening, 10ml whole culture samples were taken at days 3.5, 5 and 7, lyophilized, and testedPresent address: The Dow Chemical Company,
