<p>Fertilization can increase potential yield of maize, but little is known about the impact of different combinations of fertilizers on grain quality, especially under long-term located fertilization. To explore the effects of long-term nutrient applications on nutritional quality of maize grain, the maize variety Luyu 16 was treated with 12 different fertilization treatments based on 34&#xa0;years of located fertilization under a maize–wheat rotation in this study. The results showed that long-term fertilization can increase iron (Fe) content but decrease sodium (Na) and potassium (K) contents in maize grain. The contents of calcium (Ca) and iron (Fe) in maize grain were highest under high nitrogen (N₂) treatment, while the content of Ca was lowest under high organic manure (M₂) treatment. Under the combined application of nitrogen fertilizer and organic fertilizer, the contents of magnesium (Mg) and phosphorus (P) increased significantly, and K content also increased, although not significantly. In addition, fertilization increased protein content and decreased the content of starch and oil. Furthermore, the contents of total amino acids and lysine were increased under fertilization. The X-ray diffraction spectrum of the starch was of the typical “A” type, and none of the fertilization treatments changed the starch crystal type. The results also indicated that pasting temperature, trough viscosity and final viscosity increased after fertilization treatment, while peak viscosity and breakdown reduced. High inorganic nitrogen (N₂) fertilizer could significantly improve setback, while high organic fertilizer (M₂) significantly reduced setback. These results suggested that rational use of fertilizers, especially the combined use of organic and inorganic nitrogen fertilizers, plays an important role in improving maize grain quality.</p>