In this study, the stereoselectivity of flufiprole enantiomers in regards to their bioactivity., acute toxicity and environmental fate is reported for the first time. Four types of representative insects (Plutella xylostella, Nilaparvata lugens, Mythimna separata and Acyrthosiphon pisum) were used to investigate enantioselective bioactivity. Acute toxicities of flufiprole enantiomers toward two non-target organisms were also evaluated. Moreover, stereoselective degradation in four vegetables under field conditions was studied in response to food safety concerns. The bioactivity of (R)-flufiprole was 1.9-5.1 times higher than that of (S)-flufiprole. (R)-flufiprole also showed 3.7-5.7 times higher acute toxicity to Scenedesmus obliquus and Trichogramma japonicum Ashmead than (S)-flufiprole. Opposite stereoselective degradation of the two enantiomers was observed in pak choi, spinach cucumber, and tomato. (S)-flufiprole degraded faster in pak choi and spinach, resulting in an enrichment of (R)-isomer. By contrast, (R) -isomer was preferentially degraded in cucumber and tomato. Molecular simulation technology was used to illuminate the mechanism of enantioselective bioactivity. The Glide Score (-5.82 kcal/mol) for (R) -isomer was better than that (-5.11 kcal/mol) of (S) -isomer and this calculation showed (R)-flufiprole was more effective in pest control. Consequently, significant stereoselectivity of flufiprole enantiomers should be taken into account when assessing the environmental health risk of the pesticide. (C) 2016 Elsevier B.V. All rights reserved.