Objective The effect of preservation duration on compressive mechanical properties of rabbit patellar was studied with the attempt to provide reference for ligament preservation. Methods The compressive mechanical properties of rabbit patellar ligament preserved at -20°C after different preservation duration (in 36 d) were tested with the universal tensile testing machine. The microscopic morphology of collagen fiber was observed on the scanning electron microscopy (SEM). The enthalpy and denaturation temperature of collagen fibers was measured with differential scanning calorimetry (DSC). Results With the increase of preservation duration, the compressive stress of the patellar ligament at 40% strain increases from 0.019 MPa to 0.112 MPa and then decreases to 0.057 MPa. SEM shows that the cross-linking of collagen fibers is initially strengthened and then weakened. DSC results show that the enthalpy increases from 59.47 J/g to 67.10 J/g and then decreases to 54.43 J/g.The denaturation temperature increases from 67.62°C to 77.28°C and then decreases to 64.10°C. Conclusions With the increase of preservation duration, the rabbit patellar ligament’s compressive stress at strain of 40% increases firstly and then decreases. According to DSC and SEM, the reason for this change may be the variation of cross-linking level of collagen fibers. The stronger the cross-linking of collagen fibers, the stronger the compressive mechanical properties