This study evaluated the antifungal, physical, and mechanical properties of tissue conditioner incorporated with different amounts of zinc oxide nanoparticles (ZnOnps) at different storage times (0, 7, and 14 days). Specimens of 0, 5, 10, 15 wt% ZnOnps, or 15 wt% nystatin incorporated into tissue conditioner were fabricated (control, 5Zn, 10Zn, 15Zn, and Nys). The direct contact test (n=6) was performed to evaluate the antifungal effect against C. albicans suspension. The penetration depth (n=6) and tensile bond strength (n=8) were evaluated following ISO 13139. The 15Zn significantly reduced C. albicans cell number compared with control at all storage times (p<0.001). The penetration depths and tensile bond strengths of the 5Zn, 10Zn, 15Zn, and Nys were not significantly different compared with control at all storage times (p>0.05). In conclusion, the 15Zn provides antifungal effect up to 14 days without adverse effects on penetration depth and tensile bond strength.