In this paper, prepolymer of waterborne polyurethanes (PLA-WPU-C-pre) were synthesized by the IPDI, PLA-OH and PTMG as monomer, DPA and BDO as chain extender. Then the prepolymer of PLA-WPU were emulsified by bubbling with carbon dioxide (CO2) into the water. In further, the 1,6-diiodohexane was added into the PLA-WPU emulsions for crosslinking, and then the mixtures are pouring into a PTFE mold, the films with crosslinking structure (PLA-WPU-C) were obtained after curing. The effects of PLA-based polyol content, cross-linked structure and annealing time on the mechanical properties, antibacterial properties and biodegradability of PLA-WPU-C were investigated. The results show that PLA-WPU-C has good biodegradation and antibacterial properties, and the cross-linked PLA-WPU has better mechanical properties (27.8 MPa and 384%) when the molar ratio of PTMG to PLA-OH is 3:1 and the annealing time is 1h.