A small molecular organic depressor glycerine-xanthate was synthesized. The effect of glycerine-xanthate on the flotation of sulfide minerals was investigated based on a function of pH value and concentration of glycerine-xanthate through flotation experiments in the presence and absence of Cu²⁺. The results show that glycerinee-xanthate has a strong dressing effect on marmatite at pH>6 and on arsenopyrite in weak acid and base conditions with butyl-xanthate as collector. In the presence of glycerine-xanthate, marmatite is activated by addition of Cu²⁺, but arsenopyrite cannot be activated and remains unfloatable. So the selective separation can be achieved for two minerals. The depression of glycerine-xanthate on sulfide minerals was discussed based on the radical electronegative calculation and the theory of HSAB. Infrared spectrum shows that there are some —OH and —CSS— in glycerine-xanthate molecule, which competes with butyl-xanthate on the mineral surface. As a result of many hydrophilic groups in glycerine-xanthate, the surfaces of marmatite and arsenopyrite become hydrophilic, thus the flotation of marmatite and arsenopyrite is depressed. The collector is adsorbed preferentially on the surface of marmatite and it shows a better floatability in the presence of Cu²⁺, whereas, the surface of arsenopyrite absorbs glycerine-xanthate and the flotation of arsenopyrite is depressed by glycerine-xanthate.