The anti-obesity effects of extracts of Inonotus obliquus and roasted I. obliquus were investigated through the measurement of the differentiation of 3T3-L1 pre-adipocytes to adipocytes. Cytotoxic activity was determined by MTT assays on 3T3-L1 pre-adipocyte. Adipocyte differentiation was assessed through the evaluation of lipid accumulation (by using Oil Red O staining) and the expression of adipogenic proteins (by using western blotting). Compared with the control, the treatment of differentiating adipocytes with extracts of I. obliquus , and roasted I. obliquus significantly inhibited differentiation. In the presence of an adipogenic cocktail (isobutylmethylxanthine, dexamethasone, and insulin), I. obliquus and roasted I. obliquus extracts significantly decreased the expression of adipogenic proteins, peroxisome proliferator-activated receptor-γ (PPARγ), and CCAAT/enhancer binding protein alpha (C/EBP-α), relative to the expression level of N-acetyl-L-cystein (NAC, negative control). These results indicated that the extracts of I. obliquus and roasted I. obliquus were potential candidates for the prevention obesity.