Abstract: To elucidate the structural characteristics of acidic polysaccharides from Wolfiporia extensa and their protective effects against H₂O₂-induced oxidative damage in PC12 cells, crude polysaccharides were extracted with a deep eutectic solvent and purified sequentially by DE-52 cellulose and Sephadex gel column chromatography, yielding the acidic polysaccharide fraction WEP-2A. Its structural features and neuroprotective mechanisms were systematically investigated using an H₂O₂-induced oxidative stress model in PC12 cells. Results showed that WEP-2A had a purity of 94.68% and was composed of mannose, galactose, glucuronic acid (GlcUA), glucose, and fucose in a molar ratio of 44.03:23.56:15.42:9.84:7.15. Structural analysis revealed a β-configuration pyranose backbone containing linear glycosidic linkages such as →3)-Glcp-(1→ and →4)-Glcp-(1→, with glucose residues as the major component. GlcUA was identified in the form of →6)-GlcUAp-(1→, a characteristic feature of this acidic polysaccharide. In the PC12 cell model, WEP-2A exhibited a dose-dependent protective effect. The high-dose group showed a cell viability of 83.61%, significantly increased by 102.21% compared to the model group (P<0.05). The activities of antioxidant enzymes SOD, CAT, and GSH-Px were significantly elevated by 93.46%, 125.46%, and 151.63%, respectively (P<0.05), while the MDA content was significantly reduced by 60.47% (P<0.05). Furthermore, WEP-2A significantly upregulated the mRNA expression of Nrf2, HO-1, NQO1, and GCLC by 104.44%, 140.54%, 141.67%, and 109.52%, respectively (P<0.05). In conclusion, WEP-2A can exert antioxidant neuroprotective effects by activating the Nrf2 signaling pathway, and this result provides an experimental basis and theoretical support for its development and application in the field of functional foods.