Although carbon nanozymes have attracted great interest due to their good biocompatibility, low cost, and high stability, designing high-active carbon nanozymes still faces great challenges. Herein, ultrathin nitrogen-doped carbon nanosheets with rich defects (d-NC) were prepared through a high-temperature annealing process, using potassium chloride and ammonium chloride as templates. Owing to the large specific surface area, rich defects and the high exposure of active sites, the proposed d-NC nanozymes exhibited excellent peroxidase-like activity. The d-NC nanozymes possess maximal reaction velocity and their specific activity is 9.4-fold higher than that of nitrogen-doped carbon nanozymes, indicating that the induced defects can boost the catalytic performance. Benefited from the good peroxidase-like activities of d-NC nanozymes, the colorimetric sensing platforms were constructed for the detection of urease activity and fluoride ion, exhibiting satisfactory stability and selectivity. This study not only offers a way to synthesize carbon nanozymes with improved enzyme-like activities but also broadens their applications in colorimetric biosensing. Display omitted The colorimetric sensing platform based on defect-rich and ultrathin nanozymes was constructed for the detection of urease activity and fluoride ion.