Antrodia camphorata is a fungus native to Taiwan, and it is considered a precious medicinal agent. We analyzed triterpenoids, polysaccharides and 1,3-β-D-glucan, three major effective components in A. camphorata extracts (ACE). ACE exhibited a selective cytotoxic effect on BxPC-3 human pancreatic cancer cells. ACE markedly inhibited the migration ability of BxPC-3 cells. Treatment of BxPC-3 cells with ACE resulted in the increase of cells in the sub-G1 phase and G2/M phase arrest. Apoptosis was confirmed by validating phosphatidylserine externalization, the observation of characteristic chromatin condensation, and nuclear DNA fragmentation. ACE induced apoptosis in BxPC-3 cells through a mitochondria-dependent pathway by triggering an appropriate balance of bax/bcl-2, cytochrome c release, activation of caspase-9 and -3, and poly(ADP-ribose) polymerase cleavage. ACE shows great therapeutic potential due to its cytotoxic effects against BxPC-3 cells which include inhibiting cell migration and inducing mitochondria-mediated apoptosis.