Fossil-based fuels are limited and will be consumed in the near future, the petroleum crises and price increases, the increase of air pollution caused by the vehicles necessitate the investigation of new and renewable energy sources. The energy source, which is the greatest technical potential, is biodiesel among these sources. In this study, yellow mustard (Sinapis alba L.) seeds which are not commercially produced in our country were used as a feedstock in the biodiesel production. The biodiesel production carried out from yellow mustard seed crude oil obtained from yellow mustard seeds with using a screw press by single-step transesterification method. A commercial diesel fuel and six different fuel blends prepared with 2% biodiesel, 20% biodiesel, 2% biodiesel and 5% ethanol, 2% biodiesel and 10% ethanol, 20% biodiesel and 5% ethanol, and 20% biodiesel and 10% ethanol used in the tests. Some important physical and chemical properties of these fuels have been measured using standard methods. Afterwards, the performance, combustion and exhaust emission characteristics were examined in a single-cylinder, four-stroke, water-cooled, direct-injection diesel engine fueled with the test fuels at different engine speeds. As a result, calorific value, kinematic viscosity, water content, density, acid value and cold flow properties adversely affected by biodiesel addition into the diesel fuel, and the improvement of the flash point value was determined. The kinematic viscosity, density and cold flow properties improved by ethanol addition to the blended fuels, however water content, flash point and calorific value deteriorated. The addition of 2% biodiesel into the diesel fuel improved performance, and increasing the rate of biodiesel and addition to alcohol were negatively affected the performance. The combustion characteristics of the test fuels were similar to those of diesel fuel. Furthermore, it has been found that alcohol additions reduced exhaust gas temperature, smoke opacity, CO, CO2 and NOx emissions and increased O-2 emissions. Engine noise emissions were fluctuated depending on the engine speed.