Novel strategy for enhancing TAG accumulation by simultaneous utilization of fermentable and non-fermentable carbon sources as substrate for cultivation of oleaginous yeast Rhodosporidium kratochvilovae HIMPA1 were undertaken in this investigation. The yeast strain showed direct correlation between the size of lipid bodies, visualized by BODIPY stain (493-515 nm) and TAG accumulation when examined on individual fermenting and non-fermenting carbon sources and their mixtures. Maximum TAG accumulation (mu m) in glucose (2.38 +/- 0.52), fructose (4.03 +/- 0.38), sucrose (4.24 +/- 0.45), glycerol (4.35 +/- 0.54), xylulose (3.94 +/- 0.12), and arabinose (2.98 +/- 0.43) were observed. Synergistic effect of the above carbon sources ( fermentable and non-fermentable) in equimolar concentration revealed maximum lipid droplet size of 5.35 +/- 0.76 mu m and cell size of 6.89 +/- 0.97 mu m. Total lipid content observed in mixed carbon sources was 9.26 g/l compared to glucose (6.2 g/l). FAME profile revealed enhanced longer chain (C-14:0-C-24:0) fatty acids in mix carbon sources. (C) 2015 Elsevier Ltd. All rights reserved.