Physical properties of biodiesel play an important role in the injection, atomization and combustion performance. An experiment was carried out to investigate the spray properties of biodiesel. The experimental setup was based on an electronic unit-pump (EUP) bench, a constant volume chamber and a high-speed digital camera. The photographs of spray were dealt with by using an image processing procedure. Then the spray tip penetrations and cone angles were obtained and analyzed. The experimental results indicate that the spray tip penetrations and cone angles of biodiesel increase with increasing injection duration. In addition, with decreasing ambient pressure the spray tip penetrations increase while the cone angles decrease. Furthermore, ambient pressure has a stronger effect on the spray properties of biodiesel than injection pressure. On the macroscopically view, the shape of biodiesel spray is similar to that of diesel. The final tip penetrations and cone angles of biodiesel are greater than those of diesel due to its higher viscosity, density and bulk modulus. Copyright (c) 2008 John Wiley & Sons, Ltd.