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Nitrous oxide (N2O) is an important greenhouse gas. Emissions of this gas are generated by the combustion of fossil fuels, mainly in the transportation sector worldwide. Biofuel is a promising alternative instead of diesel considering that it allows a reduction on the emissions of polluting gases. In Brazil, there is a large program on the use of biofuels, which according to the Intergovernmental Panel on Climate Change (IPCC) reduces greenhouse gas emissions. This work shows the N2O emissions resulting from combustions of diesel/beef tallow biodiesel blends of B7, B15, B20, B25, B35, and B50. In parallel, another experiment was done with fuel sample adopted by a bus fleet from Sao Paulo using blends comprising 10% of sugarcane diesel in diesel (CD10) and blends with beef tallow biodiesel of CD10: CD-B15, CD-B20, CD-B25, CD-B35, and CD-B50. The photoacoustic spectroscopy technique coupled to a quantum cascade laser (QCL) was used to detect N2O. Concentrations of the emissions were detected in the range of 5.3 to 7.4 ppmv for diesel/beef tallow biodiesel blends combustion and in the range of 4.3 to 11.0 ppmv for diesel/sugar cane diesel/beef tallow biodiesel blends combustion. The technique used was very sensitive and selective to identify and determine the emission concentrations of N2O.