In the present work, cagaite (Eugenia dysenterica DC.) seed oil was studied as a potential inedible raw material for biodiesel production. The oil was extracted using a Soxhlet extractor and the fatty acids acyl esters that make up biodiesel were obtained by alkaline transesterification using sodium hydroxide. The influence of reaction parameters was also evaluated: molar ratio, time, and amount of catalyst. The conversion of fatty acids into fatty acid methyl esters (FAME), which make up biodiesel, was calculated using Nuclear Magnetic Resonance Spectroscopy (1HNMR) spectra. In a 30 min reaction period, the molar ratio of oil to methanol was 1:4, resulting in a conversion of 63.57%. However, when the molar ratio was increased to 1:8, the conversion reached 81.74% during the same 30 min reaction period. After 60 min of reaction, additional increases in conversion were observed when molar ratios of 1:4, 1:6, and 1:8 were used. Under these conditions, the conversions achieved were 92.08%, 98.24%, and 98.78%, respectively. The physico‐chemical properties were evaluated and the results showed that cagaite seed oil biodiesel was similar to soybean biodiesel, which is the most commonly produced biodiesel in Brazil. It was thus an important substitute for soybean biodiesel.