One of the most common strategies in the treatment of cognitive disorders is enhancing the acetylcholine level in the brain through the inhibition of acetylcholinesterase. Despite the effectiveness of current modern drugs, more attention has been paid for finding new anticholinesterase agents from medicinal plants. Zatraia multiflora Boiss. is an endemic plant to Iran which has different uses in traditional medicine as anti-inflammatory, antimicrobial, anti spasmodic. We intended to evaluate the in vitro anticholinesterase and free radical scavenging activity of the essential oil and methanolic extract of Z. multiflora. The essential oil and methanolic extract of the plant were evaluated for anticholinesterase activity using modified Ellman method. The free radical scavenging effect of the samples were studied by using of the diphenylpicrylhydrazyl (DPPH). IC50 and the percent of inhibition of acetylcholinesterase was calculated from regression equation. The results showed that both the essential oil and methanolic extract of the plant exhibited high anticholinesterase activity (95.3 +/- 3.4 and 87.9 +/- 2.2% inhibition, respectively) which was similar to eserine (96.2 +/- 1.7% inhibition). The IC50 value of essential oil was determined as 0.97 +/- 0.12 microg mL(-1) in comparison to eserine (0.13 +/- 0.02 microg mL(-1)). The results of antioxidant assay showed that both the essential oil and methanolic extract potentially inhibit DPPH free radical (94.8 +/- 2.4 and 93.2 +/- 1.7% inhibition, respectively). The essential oil and methanolic extract of Z. multiflora have beneficial effect in health promotion and this plant would be good candidate for further studies.