Six triterpenoids, two sterols, and two phenolic constituents were isolated from the methanol extract of sclerotia of Inonotus obliquus (Hymenochaetaceae). Their chemical structures were identified as lanosterol (1), 3 beta-hydroxylanosta-8,24-dien-21-al (3), inotodiol (5), trametenolic acid (6), lanosta-8,25-dien-3,22,24-triol (7), beta-sitosterol (2), ergosterolperoxide (4), betulin (8), 2,5-dihydroxy-benzaldehyde (9), and 3,4-dihydroxybenzalacetone (10) on the basis of spectroscopic methods and comparison with the literature. Among these compounds, 9 was isolated for the first time from this mushroom. To the best of our knowledge, the present study marks the first chemical investigation of I. obliquus from Alaska. Among the isolated compounds, compounds 1, 3, 4, 5, 6, 8, 9, and 10 were evaluated for in vitro cytotoxic activity against A 549 (human alveolar basal epithelial carcinoma cells), L1210 (mouse lymphocytic leukemia cells), COLO 205 (colorectal adenocarcinoma cells), MCF-7 (breast cancer cells), and HL-60 (human leukemia) cancer cell lines. 3 beta-Hydroxylanosta-8,24-dien-21-al (3) and tramentenolic acid (6) exhibited modest cytotoxic effects against L1210 cells with IC(50)s of 62.5 and 34.4 mu M, and betulin (8) showed weak cytotoxic effects against A549 and HL-60 with IC(50)s of 81.2 and 87.5 mu M, respectively. In addition, 3,4-dihydroxybenzalacetone (10) exhibited the strongest cytotoxic activities against A549 and HL-60 cells with IC50 values of 23.6 and 21.7 mu M, respectively.