The Janus nanoparticles stabilized Pickering emulsions showed great application potential in various fields, including enhanced oil recovery (EOR). In this study, the amphiphilic Janus-SiO2 nanoparticles (Janus-C12) are successfully synthesized by the Pickering emulsion method and characterized by Fourier transform infrared spectroscopy (FT-IR), thermogravimetric analysis (TGA), ζ-potential and contact angle measurements. The phase inversion from oil-in-water (O/W), multiple oil-in-water-oil (O/W/O) emulsion, to water-in-oil (W/O) emulsions was observed with the delicate control for the modification degrees of Janus-C12. The possible emulsification mechanisms for various Janus-C12 stabilized emulsions were proposed. The multiple O/W/O emulsion stability in harsh conditions and the influence factors (nanoparticles concentration, water/oil volume ratio and NaCl concentration) were studied systematically. Moreover, the Janus-C12–79% stabilized multiple O/W/O Pickering emulsions exhibited excellent EOR performance (additional oil recovery of 27.2%) in core flooding tests, and the possible EOR mechanism was explored via microscopic visualization experiments. Display omitted •Several amphiphilic Janus SiO2 nanoparticles (Janus-C12) were synthesized.•The hydrophile-lipophile balance of Janus-C12 was accurately controlled.•The individual Janus-C12-79% could generate multiple O/W/O Pickering emulsions.•The O/W/O Pickering emulsions showed high displacement and great sweep efficiency.•Remarkable EOR performance was achieved by the O/W/O Pickering emulsion flooding.