This study compares the structural and thermal properties of silk fibroin (SF) films obtained from different raw materials: silk coccons (SC) and silk fibrous wastes (SW). Samples were processed by solution casting (SFC and SFW), and casting with a subsequent water annealing treatment (WA-SFC and WA-SFW). The presence of crystalline (silk I and silk II) and amorphous structures was evaluated using X-ray diffraction (XRD) and Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR) spectroscopy. Thermal properties of SF films were studied by DSC measurements. Accordingly, to FTIR and XRD results, the presence of silk I structure is greater in SFW that in SFC. Differences in enthalpy of crystallization peaks in DSC curves reveals that SFC presents a higher content of amorphous structures than SFW. The analysis also point that water annealing treatment induces conformational changes in secondary structures of SF films. It can be concluded that the presence of crystalline structures in SF films varies with the raw material and water annealing treatment. This fact represents a progress in the development of processing alternatives to control SF materials structure and properties.