Abstract: In this paper, the preparation of Selenized Gastrodia elata polysaccharides (SeGEP) was conducted through HNO₃-Na₂SeO₃ method using GEP and sodium nitrite as the raw materials, and the modification conditions were optimized by single factor experiment combined with response surface method using selenium content as the index. Ultraviolet spectrum (UV), infrared spectrum (IR), nuclear magnetic resonance (NMR), particle size test, zeta potential test, Congo red test, iodine-potassium iodide test, and scanning electron microscope (SEM) techniques were carried out to explore the structure of GEP and SeGEP. Antioxidant tests were conducted to evaluate the effect of selenylation modification on GEP in vitro. The results showed that the optimum selenylation parameters were as follows: Reaction temperature 74 ℃, nitric acid concentration 0.041%, reaction time 8.4 h, and under the optimum conditions, the selenium content could be reached to 3891.05±10.86 μg/g. Structural characterization analysis showed that SeGEP had been successfully accomplished, and SeGEP had reduced the particle size, increased the absolute value of Zeta potential and improved the stability in solution by comparing with GEP. It was also found that both GEP and SeGEP might possess triple helix with long side branched chain structures, whereas selenylation modification could change the micromorphology of GEP. The in vitro antioxidant results showed that the DPPH radical scavenging rate of SeGEP at 10 mg/mL was 98%±1.52% and the maximum iron reducing power was 0.99±0.24, and the ABTS⁺ radical scavenging rate of SeGEP at 1 mg/mL was 97.49%±1.16%, respectively. SeGEP exhibited higher antioxidant activities than that of GEP, indicating that selenylation modification could improve the antioxidant ability of GEP. This study provided a theoretical basis for the development of selenium supplement and functional food related to selenized Gastrodia elata polysaccharides.