The microstructure and mechanical properties of β-nucleated iPP before and after being annealed at different temperatures (90-160 °C) have been analyzed. Annealing induced different degrees of variation in fracture toughness of β-nucleated iPP samples, namely, slight enhancement at relatively low annealing temperatures (<110 °C) and great improvement at moderate temperatures (120-130 °C), whereas dramatic deterioration at relatively high temperatures (>140 °C) has been observed. The variation of fracture toughness of β-nucleated iPP is observed to be dependent on the content of β-NA. Experiments, including scanning electronic microscope (SEM), wide-angle X-ray diffraction (WAXD), differential scanning calorimetry (DSC), small-angle X-ray scattering (SAXS), and dynamic mechanical analysis (DMA), are performed to study the variations of microstructures as well as the toughening mechanism of the β-nucleated iPP after being annealed. The results indicate that the decreased number of chain segments in the amorphous region and the formation of microvoids, which is easily triggered by the secondary crystallization at 120-130 °C, are mainly responsible for the great improvement of toughness through promoting the lamellae to slip or elongate along the impact direction and inducing the intense plastic deformation during the fracture process.