The current results of NOx emission tests of diesel engines fuelled with unconventional biofuels are not unambiguous. Therefore, the aim of the study was to analyze the influence of temperature of methyl esters from animal fats (AFME) on the level of NOx emission. The test was carried out on an experimental stand equipped with a diesel engine with a power of 9.5 kW. The engine was controlled by an electric generator from which the generated energy was fed into the electric network. The test was carried out in accordance with the D1 standard for power generators. As a result of the statistical analysis, significant differences were found for the values of average unit emissions of nitrogen oxides obtained for individual phases of D1 tests, i.e. engine loads amounting respectively to 50%, 75% and 100% of the nominal value of engine torque. On the basis of the test results, a significant influence of fuel temperature on NOx reduction can be found. Increase of fuel temperature by 50°C resulted in reduction of emissions by 6% on average.