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• The dominant components of the essential oil from Atractylodes lancea (Thunb.) DC. were β-eudesmol and hinesol. • The essential oil showed an excellent antioxidant activity correlated well with increasing dose in vitro and in vivo. • The essential oil showed a wide antibiogram against both Gram-positive and Gram-negative bacteria. • The antibacterial target of the essential oil was cell membrane. Atractylodes lancea (Thunb.) DC. is an important traditional herb used eastern Asia in folks for centuries. The antioxidant and antibacterial activities of Atractylodes lancea essential oil were investigated. An essential oil was extracted by steam distillation method from rhizomes of Atractylodes lancea. Chemical components of the essential oil were determined by gas chromatography-mass spectrometry. Antioxidant activities of the essential oil were investigated by DPPH− radical scavenging, inhibition of lipid peroxidation, and effect on the total antioxidant capacity in mice. Antimicrobial activities of the essential oil were determined by agar disc diffusion assay and double-broth dilution method. Antimicrobial mechanisms were investigated based on cell membrane permeability, fluorescence probe-membrane integrity assay, and electron microscopy. The results showed that the essential oil was composed of β-eudesmol (36.5 %), hinesol (29.4 %), elemol (4.21 %), and atractylone (4.10 %). The essential oil showed antioxidant activities in vitro and in vivo , and exhibited antibacterial activities against both Gram-positive and Gram-negative bacteria. The antibacterial mechanism of the essential oil involved the simultaneous disruption of the cell membrane. Therefore, Atractylodes lancea essential oil might be considered as a natural antioxidant and antibacterial agent with potential application in food and pharmaceutical industries. [ABSTRACT FROM AUTHOR]