Bombyx Mori fiber consists of two major proteins which are fibroin and sericin. The silk fibroin (SF) is the core structural protein of silk fiber. SF protein structures comprise of primary and secondary structures; where the primary structure contains series of amino acid and secondary structure with Silk I refers to the water-soluble and Silk II, high β sheet extent which is insoluble. This study was conducted to compare the structural and characterization of insoluble Thai Bombyx Mori SF with different types of post-treatement. Thai silk cocoons, which were degummed and dissolved in 9.3 M LiBr solution at 60 °C. The obtained SF solutions were dialyzed and purified. SF films were prepared by solution casting and immersing in methanol and ethanol, followed by water annealing in water saturated vacuum. Post-treatment was purposely done to regenerate and induce of the β sheet structure to enhance the insolubilities and the stabilities properties of the SF films. The SF films structural conformation, characterization and thermal stability were characterized. Attenuated total reflectance-Fourier transformed infrared spectroscopy (ATR-FTIR) showed that SF films were presented in a more stable form after ethanol post treatment, which also supporting by X-ray diffraction (XRD) analysis which indicated the tendency to higher structural organization. Thermal analysis resutls showed that SF was thermally stable and improved after post treatment. The contact angle of post treated SF increased the hydrophobicity of the films. The thai SF films could be the promising candidate for applications in tissue regeneration, optical devices, and flexible electronic displays with the possibility to control the SF structure and properties.