The invention belongs to the technical field of preparation of light-emitting semiconductor nanoparticles, and relates to a non-toxic Cu2S/ZnS core/shell quantum dot and a preparation method thereof.The preparation method comprises the following steps: adding zinc salt, water, cuprous salt, an oil-soluble organic solvent and sulfur salt into a container in sequence, sealing the container, separating and removing the water phase after a constant temperature hydrothermal reaction, carrying out centrifugal separation of impurities to obtain a solution A; adding an alkaline reagent and a sulfydryl ligand into the solution A to remove the oil-soluble organic solvent, separating to remove the oil phase so as to obtain an aqueous phase solution B, namely the water-soluble Cu2S/ZnS quantum dot. The obtained oil-soluble and water-soluble Cu2S/ZnS quantum dots both have the characteristic of fluorescence luminescence, different wavelengths of luminescence and low cytotoxicity.