Abstract: This study investigated the effects of dynamic high-pressure microfluidization (DHPM) on the structural characteristics and bioactivity of polysaccharides extracted from Dictyophora indusiata (DIP). The physicochemical properties, molecular weight, monosaccharide composition, and Fourier transform infrared (FTIR) spectral features of DIP were systematically analyzed following DHPM treatment. Antioxidant and anti-inflammatory activities were evaluated using DPPH·, ABTS⁺·, and ·OH radical scavenging assays, ferric ion reducing antioxidant power (FRAP), and an lipopolysaccharide-induced RAW264.7 macrophage inflammation model. Compared with the group that did not undergo DHPM treatment, DHPM treatment significantly increased the polysaccharide yield and total polysaccharide content, by 4.87-fold and 1.23-fold, respectively (P<0.01). It also markedly reduced the weight-average molecular weight and significantly increased the glucose content in the monosaccharide composition (P<0.001), without altering the primary structure, which remained pyranose ring-based. Additionally, DHPM-treated DIP demonstrated enhanced antioxidant activity, including improved scavenging of DPPH·, ABTS⁺·, and ·OH radicals and greater ferric ion reducing power. The DHPM treatment also significantly downregulated IL-6 and TNF-α expression in LPS-stimulated RAW264.7 cells (P<0.001), indicating stronger anti-inflammatory activity. In conclusion, DHPM-assisted extraction effectively improves the yield and functional properties of DIP polysaccharides by reducing molecular weight and modifying monosaccharide composition. These findings provide valuable insights for optimizing the extraction and functional utilization of DIP.