Bone tissue engineering aims to construct biological substitutes for repairing bone defects. Nanofibrous (NF) scaffolds are commonly utilized to mimic the extracellular matrix (ECM) environment and promote tissue regeneration in tissue engineering process. Poly (lactic acid) (PLA) has attracted much attention in the field of tissue engineering because of its biocompatibility, biodegradability and so on. However, the intrinsic hydrophobicity and the lacking of active functional groups limit its practical application to some extent. In this study, poly(ethylenimine) (PEI) modified PLLA nanofibrous scaffolds were fabricated in a one step process by aminolysis combined with thermally induced phase separation technique for introducing more functional groups, PEI acting as the modifier. The morphology of PEI-modified PLLA scaffolds prepared under different experimental conditions was analyzed by scanning electron microscope (SEM). The suitable conditions to fabricate scaffolds with a homogeneous nanofibrous structure, good hydrophilicity and excellent mechanical properties were determined according to the results of SEM, water contact angle (WCA) and mechanical properties testing. Besides, Fourier transform infrared spectroscopy (FTIR), proton nuclear magnetic resonance spectroscopy (¹H NMR), X-ray Photoelectron Spectroscopy (XPS) and gel permeation chromatography (GPC) were used to confirm the occurrence of the ammonolysis reaction between PLLA and PEI. The in vitro biomineralization study showed that the PEI-modified PLLA scaffolds had a greater ability to induce the formation of apatite in 1.5SBF than PLLA scaffolds, indicating that the bone-bioactivity of PLLA scaffolds was significantly improved after modification with PEI. Furthermore, cell culture assay revealed that MC3T3-E1 osteoblasts exhibited better proliferation performance on the PEI-modified PLLA scaffolds. All the results implied that the synthesized modified PLLA nanofibrous scaffolds may provide promising applications in bone tissue engineering.