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The purpose of this study was to characterize the polysaccharides from <i>Athyrium multidentatum</i> (Doll.) Ching (AMC) rhizome and explore the protective mechanism against <span style="font-variant: small-caps;">d</span>-galactose-induced oxidative stress in aging mice. Methods: A series of experiments, including molecular weight, monosaccharide composition, Fourier transform infrared (FT-IR) spectroscopy, and ¹H nuclear magnetic resonance (¹H NMR) spectroscopy were carried out to characterize AMC polysaccharides. The mechanism was investigated exploring <span style="font-variant: small-caps;">d</span>-galactose-induced aging mouse model. Quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) and western blotting assays were performed to assess the gene and protein expression in liver. Key findings: Our results showed that AMC polysaccharides were mainly composed of mannose (Man), rhamnose (Rha), glucuronic acid (Glc A), glucose (Glc), galactose (Gal), arabinose (Ara), and fucose (Fuc) in a molar ratio of 0.077:0.088:0.09:1:0.375:0.354:0.04 with a molecular weight of 33203 Da (Mw). AMC polysaccharides strikingly reversed <span style="font-variant: small-caps;">d</span>-galactose-induced changes in mice, including upregulated <i>phosphatidylinositol 3-kinase</i> (<i>PI3K</i>), <i>Akt</i>, <i>nuclear factor-erythroid 2-related factor 2</i> (<i>Nrf2</i>), <i>forkhead box O3a</i> (<i>FOXO3a</i>), and <i>hemeoxygenase-1</i> (<i>HO-1</i>) mRNA expression, raised <i>Bcl-2</i>/<i>Bax</i> ratio, downregulated <i>caspase-3</i> mRNA expression, enhanced Akt, phosphorylation of Akt (p-Akt), Nrf2 and HO-1 protein expression, decreased caspase-3, and Bax protein expression. Conclusion: AMC polysaccharides attenuated <span style="font-variant: small-caps;">d</span>-galactose-induced oxidative stress and cell apoptosis by activating the <i>PI3K/AKT</i> pathway, which might in part contributed to their anti-aging activity.