Para-aminopropiophenone (PAPP) is a potent methaemoglobin (MetHb) forming agent used for the lethal control of exotic carnivores and mustelids. To assess the sensitivity of Australian wildlife to PAPP we developed an in vivo assay that did not use death as an endpoint. Sub-lethal dose-response data were modelled to predict PAPP doses required to achieve an endpoint set at 80% MetHb (MetHb80). The comparative sensitivity of non-target mammals referenced to this endpoint was found to be highly variable, with southern brown bandicoots (Isoodon obesulus) the most sensitive species (MetHb80 = 6.3 mg kg−1) and bush rats (Rattus fuscipes) the most tolerant (MetHb80 = 1035 mg kg−1). Published LD50 estimates were highly correlated with PAPP doses modelled to achieve the MetHb80 endpoint (r2 = 0.99, p < 0.001). Most dose-response data for native mammals were collected in the field or in semi-natural enclosures, permitting PAPP and placebo dosed animals to be fitted with tracking transmitters and transponders and released at their point of capture. A protracted morbidity and mortality was observed only in Australian ravens (Corvus coronoides). The combination of sub-lethal dose-response assay and survival data collected in the field provided more relevant information about the actual hazard of pest control agents to non-target wildlife species than laboratory-based lethal-dose bioassays. We discuss the need to replace lethal-dose data with biologically meaningful insights able to define a continuum of toxicological hazards that better serve the needs of conservation and veterinary scientists and wildlife managers.