Abstract: To investigate the effects of sodium selenite on the growth, development, and antioxidant activity of highland barley sprouts, highland barley variety Zangqing 2000 was used as the experimental material. Highland barley seeds were soaked in various concentrations of sodium selenite solutions. The study examined the effects of these sodium selenite treatments on germination rate, germination potential, plant height, fresh weight, total selenium and organic selenium contents, as well as total phenolic and flavonoid contents and antioxidant activity in highland barley sprouts. When treated with 5 mg/L sodium selenite, highland barley sprouts achieved the highest germination rate and potential, while sprout plant height and fresh weight reached maximum values. Treatments with 5~80 mg/L sodium selenite significantly increased total selenium and organic selenium contents. Total phenolic and flavonoid contents peaked at 40 mg/L and 10 mg/L sodium selenite treatments, respectively, increasing by 8.53% and 16.41% compared to the control group. The ferric reducing antioxidant power and ABTS⁺ free radical scavenging ability reached their highest levels at 10 mg/L, with concurrently high DPPH free radical scavenging ability, increasing by 11.85%, 12.85% and 11.90% compared to the control group, respectively. The antioxidant activity of highland barley sprouts showed a positive correlation with total phenolic and flavonoid contents. In summary, a treatment concentration of 10 mg/L sodium selenite was identified as optimal, significantly enhancing the quality of highland barley sprouts by synergistically promoting both sprout growth and antioxidant activity. The findings provide theoretical and technical references for selenium-enriched production of highland barley sprouts in the Tibetan region.