It is greatly desired to develop a high-efficiency demulsifier for the treatment of O/W emulsions in the production fluids of the petroleum industry. In this study, the amphiphilic Janus-SiO2 nanoparticles (Janus-C18) was synthesized and used as a demulsifier for surfactant-stabilized O/W emulsions. The successful preparation of Janus-C18 was verified by transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), thermogravimetric analysis (TGA), and contact angle measurements. Then, Janus-C18's demulsification efficiency was systematically evaluated by the bottle test. Various silica nanoparticles and emulsifier were employed in the control experiments. Based on the Janus-C18's demulsification performance in different conditions and the zeta-potential measurement, the possible demulsification mechanism of Janus-C18 was proposed. Furthermore, Janus-C18 exhibited excellent performance in separating oil from crude oil/water emulsions at a relatively low concentration (150 mg/L), with the oil removal rate exceeding 99%.