Magnolol, a hydroxylated biphenyl agent isolatedfrom herbal planet Magnolia officinalis, is a componentof traditional Asian herbal teas. It has been reported tohave anti-microbial, anti-inflammatory, and anti-canceractivity. Non-small cell lung cancer (NSCLC) cell lines(A549, H441 and H520) and normal human bronchialepithelial cells (HBECs) were used to evaluate the cytotoxiceffect of magnolol. We show that magnolol inhibitedcellular proliferation, increased DNA fragmentation, anddecreased mitochondrial membrane potential in all NSCLCcells, but had no cytotoxic effect on HBECs. Magnololtriggered the release of pro-apoptotic proteins: Bid, Bax and cytochrome c from mitochondria, but did not activatethe caspase-3, -8, and -9, suggesting that magnolol inducesapoptosis of NSCLC cell lines via a caspase-independentpathway. The caspase-independent pathway is mediatedthrough the activation of nuclear translocation of apoptosis-inducing factor, endonuclease G and cleaved poly(-ADP-ribose) polymerase, which played important roles inmediating cell death. Furthermore, magnolol inhibitedPI3K/AKT and ERK1/2 activity, but up-regulated p38 andJNK activity in A549 cell lines. The results of this studyprovided a basis for understanding and developing magnololas a novel treatment of NSCLC.