Biodiesel fuel is one of the fuels that has been and is a potential alternative fuel to fossil diesel, not only helping to reduce dependence on fossil fuels but also being environmentally friendly. Some properties of fuel greatly affect engine performance, such as the cetane number, specific gravity, and viscosity. Meanwhile, biodiesel fuel is produced from many different raw materials and technologies, leading to some different basic properties. This article presents the results of a survey on the impact of biodiesel fuel with a mixing ratio of 0% (B0) and 20% made from fish fat (BF20), waste frying oil (BW20), and palm oil (BP20) on the performance and emissions of a single-cylinder diesel engine AVL-5402. The test was conducted under conditions of 25%, 50%, and 75% load at 1400 rpm and 2200 rpm. The results showed that carbon monoxide (CO), hydrocarbons (HC), and nitrogen oxides (NOx) emissions were affected by cetane number and oxygen content, while viscosity affected smoke level.