Understanding the progression of periodontal destruction is at the forefront of periodontal research. Therefore, we aimed to capture the dynamics of gingival tissue proteome during initiation and progression of an experimental periodontitis developed by placing silk ligatures on the second molars of the specific pathogen-free C57BL/6 mice. Pressure cycling technology (PCT), a recently developed platform that uses ultra-high pressure to disrupt tissues, was utilized to achieve an efficient and reproducible protein extraction for quantitative shotgun proteomics. Lysis and protein digestion of gingival tissue was performed using PCT and analysed by Orbitrap Fusion. The proteomics data were quantitatively assessed with the Progenesis QI software.