One of the key solutions to improve engine performance and reduce exhaust emissions of internal combustion engines is direct injection of bio-fuels. A new modified bio-ethanol is produced to be substituted by fossil fuels in gasoline direct injection engines. The key advantages of modified bio-ethanol fuel as an alternative fuel are higher octane number and oxygen content, a long-chain hydro-carbon fuel, and lower emissions compared to fossil fuels. In the present study spray properties of a modified bio-ethanol and its atomization behaviors have been studied experimentally and theoretically. Based on atomization physics of droplets dimensional analysis has been performed to develop a new non-dimensional number namely atomization index. This number determines the atomization level of the spray. Applying quasi-steady jet theory, air entrainment and fuel-air mixing studies have been performed. The spray atomization behaviors such as atomization index number, Ohnesorge number, and Sauter mean diameter have been investigated employing atomization model. The influences of injection and ambient conditions on spray properties of different blends of modified bio-ethanol and gasoline fuels have been investigated performing high-speed visualization technique. Results indicate that decreasing the difference of injection and ambient pressures increases spray cone angle and projected area, and decreases spray tip penetration length. As expected, increasing injection pressure improves atomization behaviors of the spray. Increasing percentage of modified bio-ethanol in the blend, increases spray tip penetration and decreases the projected area as well.