Abstract: The inadvertent dissolution of gangue minerals is frequently detrimental to the flotation of valuable minerals. We investigated the effect of conditioning time on the separation of brucite and serpentine by flotation. By analyzing the Mg²⁺ concentration, relative element content, and pulp viscosity, we studied the effect of mineral dissolution on brucite flotation. The results of artificially mixed mineral flotation tests (with −10 μm serpentine) showed that by extending the conditioning time from 60 to 360 s, a large amount of Mg²⁺ on the mineral surface gradually dissolved into the pulp, resulting in a decreased brucite recovery (from 83.83% to 76.79%) and an increased recovery of serpentine from 52.12% to 64.03%. To analyze the agglomeration behavior of brucite and serpentine, we used scanning electron microscopy, which clearly showed the different adhesion behaviors of different conditioning times. Lastly, the total interaction energy, as determined based on the extended DLVO (Derjaguin–Landau–Verwey–Overbeek) theory, also supports the conclusion that the gravitational force between brucite and serpentine increases significantly with increased conditioning time.