In this study, the effect of sodium pyrophosphate (NaPP) on the separation of apatite from dolomite by flotation was systematically investigated. Flotation results revealed that NaPP could selectively depress the flotation of apatite, thus realizing the separation of apatite from dolomite. Further, the selective depression mechanism of NaPP was studied through zeta potential measurements, contact angle measurements, and X-ray photoelectron spectroscopy (XPS) analysis. The results demonstrated that the adsorption of sodium oleate (NaOL) onto apatite surface was depressed by the preferential interaction of NaPP with active Ca sites. For dolomite, while the presence of NaPP hindered the interaction of NaOL with active Ca sites, it appeared to enhance the reactivity with active Mg sites. Thus, the adsorption of NaOL onto dolomite surface was hardly influenced. In this way, the separation of apatite from dolomite was achieved.