Corn stover (CS) hydrolysate was used as the fermentation feedstock of Trichosporon cutaneum CX1 for production of microbial lipid as the potential raw material of biodiesel. Two major technical barriers of the lipid fermentation were investigated: one was the strong inhibition of lignocellulose degradation compounds generated in the CS pretreatment; the other was the low carbon-to-nitrogen molar ratio (C/N ratio) of the CS hydrolysate. The newly established biodetoxification method was applied to remove the inhibitors in the pretreated CS. The enhancement of the pretreatment severity and the biodetoxification intensity on the lipid fermentation was investigated. The results show that the biodetoxification not only efficiently removed the inhibitor substances, but also led to the reduction of nitrogen content and the increase of C/N ratio. The cell lipid content of T. cutaneum CX1 using the biodetoxified CS hydrolysate reached 23.5%, which was doubled than that using the non-detoxified value. (C) 2011 Elsevier Ltd. All rights reserved.