Three new compounds, (24R, S)-3alpha-acetoxy-24-hydroxy-5alpha-lanosta-8,25-di en-21-oic acid, named tsugaric acid C (1); 3alpha-acetoxy-5alpha-lanosta-8, 24-diene-21-O-beta-D-xyloside, named tsugarioside B (2); and 3alpha-acetoxy-(Z)-24-methyl-5alpha-lanosta-8,23,25-tr ien-21-oic acid ester beta-D-xyloside, named tsugarioside C (3), and a mixture of two known steroids were isolated from the fruit bodies of Ganoderma tsugae. The structures of 1-3 were determined by spectral and chemical methods. The cytotoxic activity of the lanostanoid constituents of this fungus was evaluated against several different cancer cell lines.