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Low-grade and high-reserve scheelite, which is associated with calcite, has similar surface properties that cause difficulty in separation. In this study, sulfomethylated kraft lignin (SMKL) was used as a novel eco-friendly inhibitor for the flotation separation of scheelite and calcite. The flotation test results showed that 60 mg/L SMKL had a significant influence on depressing calcite flotation, while it had a slight effect on scheelite flotation. Furthermore, it enhanced the WO3 grade of the concentrate in the artificial mixed ore to 62.02% with a recovery rate of 80.37%. The contact angle and zeta potential showed that SMKL could effectively decrease the surface floatability of calcite and caused the negative shift of minerals’ surface potential. XPS and DFT calculations revealed that the sulfonic acid group of SMKL had an electron-donating ability and was easily adsorbed on the positively charged surface of calcite, which hindered the adsorption of sodium oleate on calcite. SMKL could separate calcium-bearing minerals with a high efficiency and selectivity, providing a new method for industrial production.