To investigate the antioxidant physiological regulatory mechanisms during the germination of naturally aged alfalfa （Medicago sativa） seeds， this experiment used ‘Zhongmu No.1’ and ‘Zhongmu No.2’ alfalfa seeds stored for 4 and 7 years， respectively， as experimental materials. After being imbibed at different stages， seed germination indices， antioxidant enzyme activities of the ascorbic acid-glutathione （AsA-GSH） cycle， and antioxidant contents were measured to analyze the changes in antioxidant indices of alfalfa seeds during storage. The main results are as follows： storage for 7 years significantly reduced the germination capacity of alfalfa seeds， specifically manifested by decreases in germination potential， germination rate， germination index， vitality index， and seedling root length with increasing storage duration. Storage duration had little effect on the initial water absorption rate of seeds but influenced the progression of the imbibition stage. As storage time increased， the lag phase during the imbibition of alfalfa seeds was prolonged. Natural ageing leads to a decline in the function of the AsA-GSH cycle during imbibition. Compared to seeds stored for 4 years， alfalfa seeds stored for 7 years exhibited significantly reduced activities of catalase （CAT）， glutathione reductase （GR）， and monodehydroascorbate reductase （MDHAR） . Ascorbic acid （AsA） and glutathione （GSH） content decreased， and oxidised glutathione （GSSG） accumulated， indicating weakened reactive oxygen species （ROS） scavenging capacity and exacerbated membrane lipid peroxidation. Seeds stored for 4 years exhibited enhanced antioxidant capacity during the mid-to-late stages of imbibition by upregulating the activity of certain antioxidant enzymes， such as peroxidase （POD） and GR， thereby partially delaying the excessive accumulation of ROS. Using the membership function method， the germination capacity of alfalfa seeds was ranked as follows： ‘Zhongmu No.1’ 4 a > ‘Zhongmu No.2’. 4 a > ‘Zhongmu No.1’ 7 a > ‘Zhongmu No.2’ 7 a. Compared to the ‘Zhongmu No.2’ seeds， under natural ageing conditions， Zhongmu No. 1 exhibited relatively stable vigor and physiological status， demonstrating stronger storage tolerance.

Alfalfa seeds  /  Storage years  /  AsA-GSH cycle  /  Seed vigor