Proteolytic activity of protease XXIII on Silk Fibroin (SF) films was studied. The films were prepared from the SF solution by casting on the polystyrene plates and used as substrate for enzymatic degradation. The SF films were incubated with 1.0 mg mL^-1 protease XXIII at 37°C up to 21 days. After incubation, those of secondary structure and thermal behavior of the SF films were investigated. FT-IR spectra indicated that the SF films predominantly β-structure. There was found that secondary structure of the films did not change even at 21 days of incubation limes. However, slightly decreased of FTIR spectra were also observed by shoulder absorption peaks. The result suggested that some crystalline regions might be digested by the enzyme. This related to the thermal stability from thermogravimetric analysis since the SF films gradually decreased their thermal stability followed the increasing of time exposure to protease XXIII. It is a promising that protease XXIII could be digested SF and will be used this enzyme as a model system for enzymatic study on SF.