The essential oil (1.83% yield, w/w) of Amomum tsao-ko was obtained by hydrodistillation method in Clevenger apparatus. 38 constituents were detected in essential oil of Amomum tsao-ko by GC-MS analysis. 32 compounds representing 98.393% were identified, of which 1,8-cineole (40.891%), α-phellandrene (9.769%), 4-propyl-benzaldehyde (6.988%), and (E)-citral (4.949%) were major compounds. In particular, 1,8-cineole (48.03%) was quantified accurately by GC-FID using external standard method. As a reference to penicillin sodium, antibacterial activity of the essential oil and 1,8-cineole were evaluated against the bacteria and fungi using agar disc diffusion respectively. The order of antimicrobial activitiy of the essential oil against the bacteria was as follows: Bacillus subtilis.> Staphylicoccus albus.> Escherichia coli. The order of antimicrobial activity of the essential oil against the fungi was: Aspergillus oryzae> Rhizopus sp.> Penicillium sp.The 1,8-cineole showed weak inhibiting effect to the bacteria and ineffective against the fungi.