Biomass like filamentous algae, waste cooking oil and fish byproducts are considered as suitable materials for biodiesel production due to its readily-availability and cost effective. The studies were conducted to investigate the optimum conditions for biodiesel production using alkaline transesterification bioprocess in combination with methanol, ethanol and 1-butanol. Molar ratio of alcohol to oil (3:1, 6:1, 1:1, 1:2, 1:3, 1:4) and type of catalyst were selected to compare biodiesel fuel produced from algae, waste cooking oil and fish byproducts. The biodiesel was of good quality obtained from waste corn cooking oil and fish byproducts implemented in the engine at 2000 rpm for 2 hours and reduced fuel consumption, engine emission like HC, NOx and CO. The highest biodiesel yield was obtained using 1:3 volumetric oil-to-methanol ratio at 1% heterogenous mixture of catalysts compared to homogenous catalyst. The fatty acid methyl ester, viscosity, total acid number and metal (Pb, Na, Ca, Mg, Cu. Fe) content were under ASTM standard. Biodiesel obtained from algae, waste cooking oil and fish byroducts was of ASTM standard and could be used as a source of diesel fuel in diesel engine.