Influences of different nitrogen applications on photosynthesis and utilization of excitation energy were explored by comparing two field-grown wheat (Triticum aestivum L.) cultivars with high or low grain protein content [High protein cultivar ‘8901’ (HC) and low protein cultivar ‘1391’ (LC)]. High nitrogen application significantly decreased both CO2 assimilation and photorespiration in both cultivars during the early stages after anthesis. However, the actual photosystem II (PSII) efficiency (φPSII) was not significantly different between high, moderate and low nitrogen applications in the HC. As a result, the ratio of VPSII to the quantum yield of carbon metabolism (φCO2) measured under non-photorespiratory conditions in the HC was higher under high nitrogen application than under low or medium nitrogen application. The grain protein content of the HC was also increased by high nitrogen application. In contrast, high nitrogen application decreased the actual PSII efficiency in the flag leaves of the LC in the early stages after anthesis and different nitrogen applications did not significantly alter the φPSII/φCO2 ratio or grain protein content in the LC. No significant difference was detected in the activity of superoxide dismutase or ascrobate peroxidase between different nitrogen treatments in either cultivar throughout the entire experimental period. These results indicate that more excitation energy is partitioned to nitrogen metabolism in the flag leaves of the HC under high nitrogen application whereas the partitioning of excitation energy in the LC was not affected by nitrogen application.