From Leaf Metabolome to In Vivo Testing: Identifying Antifeedant Compounds for Ecological Studies of Marsupial Diets

Identifying specific plant secondary metabolites that influence feeding behavior can be challenging, but a solid understanding of animal preferences can guide efforts. Common brushtail possums (Trichosurus vulpecula) predominantly eat Eucalyptus species belonging to the subgenus Symphyomyrtus, and avoid eating those belonging to the Monocalyptus subgenus (also called subgenus Eucalyptus). Using an unbiased ¹H NMR metabolomics approach, a previous study identified unsubstituted B ring flavanones in most species of monocalypts examined, whereas these compounds were absent from symphyomyrtles. We hypothesised that unsubstituted B ring flavanones act as feeding deterrents for common brushtail possums. In the current study, we tested this hypothesis by comparing how much possums ate of a basal diet, with diets containing one of four structurally related compounds; pinocembrin, flavanone (unsubstituted B ring flavanones), chrysin (the flavone analogue of pinocembrin), and naringenin (a flavanone with B ring substitution). We found that pinocembrin and flavanone deterred feeding relative to the basal diet, but that chrysin and naringenin did not at equivalent concentrations. Thus, unsubstituted B-ring flavanones may explain why brushtail possums avoid eating monocalypt species. Furthermore, small differences in the structure of secondary compounds can have a large impact on antifeedant properties. These results demonstrate that metabolomics can be a valuable tool for ecologists seeking to understand herbivore feeding preferences.