Topoisomerases are essential enzymes involved in all processes of DNA metabolism, and their inhibitors have been identified as potential anti-cancer agents. The present study examined the effect of linoelaidic acid (C18 polyunsaturated fatty) compounds derived from Gardenia jasminoids Ellis extract on the activity of eukaryotic topoisomerases inhibition. The present study identified linoelaidic acid compounds using open column fraction, HPLC, NMR and LC/MS which have effects on cell death in oral cancer cell line, FaDu, but not in immortalized normal cell line, HaCaT. Subsequent studies revealed linoelaidic acid-induced autophagy through LC3 activation. Finally, its inhibition of topoisomerase I and selectively induction of oral cancer cell death possibly implies that linoelaidic acid can be a role as potenial agents in the prevention and therapy of oral cancer.