• Sugarcane molasses was used as substrate for M. circinelloides growth. • Cultivation on sugarcane molasses supplied satisfactory lipid content (29 wt. %). • Lipids are suitable for nutraceuticals and biofuel production. • Concentrated solutions of PUFA were produced via enzymatic hydrolysis of lipids. • High FAEE yield (>96.5 %) was obtained by direct transesterification. Sugarcane molasses (SM) a low-cost substrate was used for single-cell oil production by Mucor circinelloides. Cultivations were carried out with diluted molasses at a concentration of 40 g sugars L−1 with and without nutrients supplementation. The best performance in regard to the lipid productivity (0.66 g L−1 day−1), biomass (11.2 g L−1) and lipid content (29 wt.%) was attained in a medium containing SM supplemented with minor nutrients. The profile of fatty acid composition revealed suitable properties for biodiesel synthesis, although high levels of polyunsaturated fatty acids (PUFA) were also found, which may suggest the possibility of using the oleaginous biomass from Mucor circinelloides URM 4182 as feedstock to yield both products. Concentrated solutions of polyunsaturated fatty acids (PUFA) were obtained by enzymatic hydrolysis of the acylglycerol structures, while biodiesel was prepared by direct transesterification of the oleaginous biomass using acid heterogeneous catalyst. [ABSTRACT FROM AUTHOR]