Objective: To evaluate the dietary structure of night shift medical staff and provide a theoretical basis for improving their health status. Methods: A total of 375 night shift medical staff from a tertiary hospital in Lanzhou were selected as the study subjects. The food frequency method was used to assess their dietary structure. Results: A total of 375 night shift medical staff were surveyed, including 87 males and 288 females. The intake of vegetables, dairy products, and soy and nuts was insufficient among the night shift medical staff, while the average intake of fruits slightly exceeded the recommended amount. Conclusion: Although the medical staff have a high level of health knowledge, their ability to take health actions is insufficient, resulting in an unreasonable dietary structure. Night shift medical staff generally have high work intensity, and attention should be paid to their dietary structure to reduce the frequency of eating out and improve their health status.