The disclosure describes a method of separating and recovering valuable metals from remanufacturing solution of a spent desulfurization catalyst containing vanadium, and more particularly, to a method of separating and recovering a valuable metal from remanufacturing solution of a spent desulfurization catalyst containing vanadium, which includes: adding organic acid to a spent hydrodesulfurization catalyst after collecting the spent hydrodesulfurization catalyst in order to prepare the remanufacturing solution of the spent hydrodesulfurization catalyst; adding an extracting agent and a diluent to the remanufacturing solution to extract molybdenum and extracting vanadium from an extracted filtrate to obtain an organic phase; and adding a stripping agent to the organic phase to strip and recover vanadium.