【Objective】Phenotypic and physiological characteristics of Qingshui alfalfa in fertility variation material (Medicago sativa cv. Qingshui) were studied. The objective of this study was to breed male sterile material and support alfalfa hybrid seed production.【Method】 One single plant was selected from six strains of materials with different fertility (GN-A1、GN-A2、GN-A3、GN-A4、GN-A5 and GN-A6). Polens were collected at the full flowering stage for I2-KI fertility staining,and SS,SP,Pro,CAT,POD,SOD,MDA,chlorophyll content determination of buds and leaves were measured.【Result】 GN-A1,GN-A2,GN-A3 and GN-A6 were identified as sterile by I2-KI staining. GN-A4 and GN-A5 were partially fertile. The number of buds per branch,style length and filaments length of sterile plants were significantly lower than those of fertile plants,while the Alae petals length,keel petals length and calyx length were significantly higher than those of fertile plants. The SS content,flower bud and leaf of male sterile plants were significantly higher than those of partially fertile and fertile plants. The contents of SP, Pro and bud of male sterile plants were significantly lower than those of fertile plants. With the SOD activity,male sterile plants was higher than that of fertile and partially fertile plants,and SOD activity in leaves of male sterile plants was higher than that of fertile plants. The CAT activity of flower buds of sterile and partially fertile plants was significantly lower than that of fertile plants. The correlation analysis of pollen activity and flower bud physiological indexes showed that the dyeing rate was significantly positively correlated with CAT,SP and Pro,but negatively correlated with SOD and SS.【Conclusion】 CAT,SP,Pro,SOD and SS were closely correlated with fertility. The SP content, Pro content and CAT activity of flower organs of sterile plants were insufficient,and the physiological metabolic reaction of sterile plants was not obvious in leaves.