The aim of this study was to evaluate the long-term (16-years) NITROGEN efficiency after the application of organic and mineral fertilizers at two sites Lukavec (S1) and Suchdol (S2) with different soil and climatic conditions in the Czech Republic (Central Europe) and to determine grain yield and NITROGEN content with regard to the requirements of protein content for baking quality of wheat. After the application of NPK treatment the highest average values from both sites of grain yield (6.22 t ha-1), NITROGEN content (2.01%) and NITROGEN uptake (123.6 kg ha-1) were determined, which means 78%, 26% and 121% increases compared to the unfertilized treatment. At the less fertile S1, located on Cambisol, the significant effect of NITROGEN fertilization on yield was observed. The yield of the NPK treatment was by 144% higher compared to the unfertilized Control treatment. The limit of 11.5% of protein content for bakery wheat was not achieved for any of treatments at S1, at S2 for unfertilized treatment and treatments with organic fertilizers. Lower values of recovery efficiency of NITROGEN and N inputoutput BALANCE were found at S2 situated on Chernozem.