Ore flotation tests were successfully conducted in the laboratory and at the Dexing Copper Mine by applying sodium humate as a pyrite depressant. By adding 40–60 g/t of sodium humate to the pulp and adjusting the pH to 10–10.5 with CaO, a concentrate with a Cu content of 24% was obtained without reducing the Cu recovery rate. In addition, the dosage of CaO was reduced, and the recovery of Au, Ag and Mo in the copper concentrate was enhanced due to the reduced pH of the pulp. The zeta potential, adsorption of xanthate and contact angle of the mineral surface were measured, and the results from surface measurements indicated that there was a strong hydrophilic interaction between sodium humate and the surface of pyrite. Moreover, the results revealed that the interaction between sodium humate and chalcopyrite was weak. Infrared (IR) spectra of pyrite and sodium humate were obtained, and the results indicated that sodium humate was chemically adsorbed on the surface of pyrite.