The biodiesel becomes one of the favorite alternative fuel applied to diesel engines. This research aims to understand the physics of spray and combustion characteristics of a biodiesel fuel in a constant volume chamber. For spray visualization, biodiesel was injected into a combustion chamber and a high speed camera was applied at various combustion conditions. To investigate heat-release rates and flame propagations, spark was ignited on a hydrogen fuel for the premixed combustion and then biodiesel was injected directly. In addition, parametric study was made by various geometries of combustion chambers and temperatures of fuels and injection pressures. This technology may contribute to improve the performance of bio-diesel engine and reduce emissions in future.