The effects of different ratios of NO3--N/NH4+-N (1/7,1/3,3/5,5/5,5/3,3/1,7/1) on alfalfa (Medicago sativa cv. Gannong No.3) in terms of its nutritional quality and feeding values were studied in an outdoor rain-protected chamber. Alfalfa plants were cultivated in sand with an optimum nitrogen supply (210 mg/L). The results indicated that under the ratio of NO3--N/NH4+-N (5/3), the contents of crude protein (CP), total protein (TP), digestible dry matter (DDM) and dry matter intake (DMI) were significantly higher than their respective values in the other treatments (P<0.05), while the contents of neutral detergent fiber (NDF) and acid detergent fiber (ADF) were significantly lower than their respective values in the other treatments (P<0.05). Thus, the 5/3 ratio was determined to be the best ratio for nutrient accumulation. From the perspective of microelement nutrition analysis, mixed nitrogen mainly with NH4+-N was more beneficial for the accumulation of iron, manganese, and molybdenum, and the adaptation ratios were 1/3, 1/7, and 3/5, respectively. Mixed nitrogen mainly with NO3--N was more beneficial for the accumulation of zinc, and the adaptation ratio was 3/1. In the relative feeding values analysis, the RFV value in the 5/3 treatment was significantly higher than those in the other treatments (P<0.05), and the RFV of all treatments was greater than 100. Referring to the classification standard of alfalfa grass products in America, the nutritional quality in all treatments was at least level 2, with that in the 5/3 and 7/1 treatments reaching level 1. When alfalfa was grown in an environment of available nitrogen mainly with NO3--N and the ratio of NO3--N/NH4+-N was closer to 5/3, the alfalfa crop had good nutritional quality.