[Objective] To compare the translocation and distribution of imidacloprid in tobacco with spray and root irrigation application methods. [Methods] Pot experiment in the greenhouse was carried out, and LC-MSMS was used to determine the con- tent of imidacloprid in different parts of tobacco plants (roots, stems, the upper, middle and lower leaves) at different time. [Results] The imidacloprid could be absorbed by root and could be transported to all parts of the tobacco plant after irrigating root, but the original deposition amount was larger and the transport efficiency was lower after spraying. [Conclusion] The translocation and distribution of imidacloprid by spraying was more uniform and the holding efficiency was better, but imidacloprid with root irrigation could act on leaf directly, and had better readily availability.