In this study, infrared spectroscopy, high‐performance liquid chromatography, and matrix‐assisted laser desorption ionization‐time‐of‐flight‐mass spectrometry (MALDI‐TOF‐MS) technology were applied to systematically explain the Schisandra chinensis’s polysaccharide transformation in configuration, molecular weight, monosaccharide composition, and anti‐ulcerative colitis (UC) activity after vinegar processing. Scanning electron microscopic results showed that the appearance of S. chinensis polysaccharide changed significantly after steaming with vinegar. The MALDI‐TOF‐MS results showed that the mass spectra of raw S. chinensis polysaccharides (RSCP) were slightly lower than those of vinegar‐processed S. chinensis polysaccharides (VSCP). The RSCP showed higher peaks at m/z 1350.790, 2016.796, and 2665.985, all with left‐skewed distribution, and the molecular weights were concentrated in the range of 1300–3100, with no higher peak above m/z 5000. The VSCPs showed a whole band below m/z 3000, with m/z 1021.096 being the highest peak, and the intensity decreased with the increase of m/z. In addition, compared to RSCPs, VSCPs can significantly increase the content of intestinal short‐chain fatty acids (SCFAs). This study showed that the apparent morphology and molecular weight of S. chinensis’s polysaccharides significantly changed after steaming with vinegar. These changes directly affect its anti‐UC effect significantly, and its mechanism is closely related to improving the structure and diversity of gut microbiota and SCFA metabolism.