Bioactive conformation of drugs is one of the key points for understanding the ligand-receptor interactions. In the present study, by combining density functional theory-based (DFT-based) conformation analysis with quantitative structure-activity relationship analysis (QSAR), we developed successfully a new approach (DFT/QSAR) to carry out bioactive conformation analyses for a series of 25 cyclic imide derivatives as protoporphyrinogen oxidase (PPO) inhibitors. Further potential energy surface scan, molecular docking and molecular dynamic simulation calculations validated that the DFT/QSAR-derived conformation is indeed very similar to the 'real' bioactive conformation. We believe the DFT/QSAR approach provides a simple alternative for the bioactive conformation of small molecules, especially in the case that the three-dimensional structure of protein is unknown.