Single cell oils (SCOs) accumulated by oleaginous yeasts have emerged as potential alternative feedstocks for biodiesel production. As microbial lipid accumulation is species and substrate specific, selection of an appropriate strain is critical. This study evaluates the biomass of newly isolated yeast Scheffersomyces coipomoensis CB1 for biodiesel production based on its lipid content and fatty acid profile. Yeast biomass was produced by batch fermentation in shake flasks under a high carbon/nitrogen (C/N) ratio to induce intracellular lipid accumulation using D-xylose as sole carbon source. Biomass was harvested by centrifugation, heat-dried and pulverized. Total cellular lipids were extracted as fatty acid methyl esters (FAME) by in situ transesterification and estimated gravimetrically. The chemical composition of fatty acids was characterized by GC-FID analyses. The key fuel properties – density, kinematic viscosity, cetane number, cold flow properties and oxidative stability were calculated from the fatty acid composition. Yeast strain generated 5 g/L biomass with 25 % lipid (w/w) on dry weight basis under un-optimized conditions. The yeast lipids showed the major presence of monounsaturated fatty esters (18:1,16:1) suitable for better ignition quality, oxidative stability, and cold‐flow properties of the biodiesel. Analyzed fuel properties of the yeast oil were in good agreement with international biodiesel standards. The use of less toxic co‐solvents or environment-friendly lipid extraction method together with solvent recovery and recycling would help improve process economics for sustainable production of biodiesel from the high-oleic yeast oil using the hemicellulosic fraction of agro-wastes.