Oil-in-water nanoemulsion was formulated using sesame oil, non-ionic surfactant (Tween20/Tween80) and water by ultrasound cavitation method. Development of nanoemulsion was optimized for process parameters such as surfactant type, surfactant concentration and emulsification time to obtain lower droplet diameter with greater stability. Increase in surfactant concentration and emulsifi{ligature}cation time resulted in nanoemulsion with minimized droplet diameter. Tween80 was more effective in reducing droplet size when compared to that of Tween20. Selected formulation with optimized process parameter (with oil-surfactant mixing ratio of 1:3 v/v and Tween80 as surfactant) was used for delivery of eugenol. Eugenol-loaded nanoemulsion was formulated with droplet diameter of 13. nm and was stable for more than 1 month. Sesame oil blended eugenol-loaded nanoemulsion demonstrated lower droplet size and higher stability than only-eugenol (without sesame oil) nanoemulsion. Eugenol-loaded nanoemulsion S3E3 exhibited antibacterial activity against Staphylococcus aureus. Inactivation kinetics of S. aureus showed time and concentration killing of bacteria upon treatment with S3E3 nanoemulsion. Fluorescence microscopy results demonstrated that S3E3 nanoemulsion treatment resulted in alteration of membrane permeability. In situ assessment of S3E3 in orange juice exhibited a significant reduction in the native bacteria population. © 2013 Elsevier B.V.