The use of mushrooms has immense potential in many diverse applications. Until now, more than 3,000 species are consumed around the world, and more than 100 have shown promising clinical activity against cancer and other chronic diseases. Sparassis latifolia (formerly S. crispa) is an edible mushroom that harbors β-glucan reported to possess immunostimulatory and anticancer properties. However there have been no reports on the anticancer activity of hydrophobic fractions of S. latifolia. In this study, the anticancer activities of S. latifolia extract and hydrophobic fractions were investigated using AGS (stomach cancer), A529 (lung cancer), and HepG2 (liver cancer) cell lines. In cytotoxicity results of A529 cells, fractions of A2, A3, A4, A6, A7, A8, A9, and A10 in all 12 fractions show low IC