Application of dispersants is one of the widely used oil spill response strategies. With the growing concern for particulate emulsifiers, applying amphiphilic Janus particles to emulsify oil in seawater has been examined in this paper. Amphiphilic Janus particles with a combination of colloidal particles and amphiphilic characteristic are found to show a highly effective emulsification for various oils in seawater and overcome several drawbacks of conventional chemical dispersants. We report the synthesis of amphiphilic Janus silica particles (JSPs) as a novel oil spill dispersant and confirm their amphiphilicity by various techniques. The oil-in-seawater emulsion stabilized by JSPs presents small and stable oil droplets in one month, even at a higher volume ratio of oil to seawater and a wide temperature range. It is believed that the small size and high stability of droplets are very important to oil dispersion and eventual biodegradation. Through confocal laser scanning microscopy and scanning electron microscope observations, JSPs were found to anchor tightly at droplet surface and form a dense interfacial particle film to prevent droplets coalescence. The appropriate wettability of JSPs and the reduced interfacial tension contribute to the high interfacial adsorption energy of JSPs. Additionally, JSPs show weak inhibitory effect on oil-degrading bacteria, which is confirmed by bacterial growth curve, resazurin test and oil biodegradation experiment. This study reveals that amphiphilic Janus silica particles hold an impressive capability for environmentally friendly treatment of oil spills.