Rapeseed meal, a major byproduct of biodiesel production, has been used as a low-cost raw material for the production of a generic microbial feedstock through a consolidated bioconversion process. Various strategies were tested for the production of a novel fermentation medium, rich in free amino nitrogen (FAN): commercial enzymes (CEs) (2.7 mg g(-1) dry meal), liquid state fungal pre-treatment (LSF) using Aspergillus oryzae (4.6 mg g(-1)), liquid state fungal pre-treatment followed by fungal autolysis (LSFA) (9.13 mg g(-1)), liquid state pre-treatment using fungal enzymatic broth (EB) (2.1 mg g(-1)), but the best strategy was a solid state fungal pre-treatment followed by fungal autolysis (34.5 mg g(-1)). The bioavailability of the nitrogen sources in the novel medium was confirmed in fed-batch bioreactor studies, in which 82.3 g dry cell L-1 of the oleaginous yeast Rhodosporidium toruloides Y4 was obtained with a lipid content of 48%. The dry cell weight obtained was higher than that obtained using conventional yeast extract, due to a higher total nitrogen content in the novel biomedium. The fatty acids obtained from the microbial oil were similar to those derived from rapeseed oil. (C) 2012 Elsevier Inc. All rights reserved.