Abstract: To identify the effect of different nitrogen fertilizer on grassland, fertilization and grazing treatments were applied to perennial grassland grazed by Wumeng semi-fine wool sheep in Weining County. The results showed that copper content in grasses was relatively low, but was higher than the minimum requirement for sheep. Application of nitrogen significantly increase nitrogen content (P<0.01) in herbage but there were no differences among different forms of nitrogen. Ammonium sulfate significantly increased the sulfur and zinc content (P<0.01) of herbage, but significantly decreased the selenium content (P<0.01). At the end of grazing experiment, copper, iron and selenium concentration in the blood of sheep grazing grassland treated with ammonium sulfate was significantly lower (P<0.01) than in sheep grazing ammonium nitrate treated grassland; blood zinc and sulfur were also significantly higher (P<0.01). Sheep grazing ammonium sulfate treated pasture had significantly lower (P<0.01) blood hemoglobin and hematocrit capacity than sheep grazing pasture treated with ammonium nitrate and the control (no fertilizer); antioxidant enzymes including serum ceruloplasmin, serum superoxide dismutase and serum glutathione peroxidase were significantly lower (P<0.01) than in sheep grazing the pasture treated with ammonium nitrate and the control. Malondialdehyde (MDA) content was also higher (P<0.01). There were no differences in other blood mineral element concentrations, blood indexes and serum biochemical values. It was concluded that fertilizer selection should be based on soil mineral content; grassland with low copper content is not suitable for ammonium sulfate fertilizer. Ammonium sulfate fertilizer significantly affected the antioxidant system function.