Peptide fractions showing anticancer activity were isolated from anchovy sauce, and their abilities to induce apoptosis in a human lymphoma cell (U937) were determined by biochemical and flow cytometric methods. The butanol extract (fraction Aob) from the desalted hydrophobic peptide fraction (Ao) separated from anchovy sauce by HP-20 adsorption chromatography was found to possess strong antiproliferative activity against U937 by inducing apoptosis. Treatment of U937 with Aob resulted in sub-G1 peak representing apoptosis in the cell cycle analysis, and apoptotic DNA fragmentation. Antiproliferative activity of the peptide fraction further purified successively by silica gel chromatography, TLC, and reversed-phase HPLC increased proportionally with the purification fold. The peptide fraction having antiproliferative activity was found to be composed of Ala and Phe, and its molecular weight was estimated to be 440.9 Da.