The root bark essential oil of Periploca sepium Bunge (Asclepiadaceae/ Apocynaceae) obtained by hydrodistillation was investigated by GC and GC-MS. 2-Hydroxy-4-methoxybenzaldehyde was found to be the main component (78.8% of the total) among 17 identified compounds. 2-Hydroxy-4-methoxybenzaldehyde was separated and purified by preparative thin layer chromatography, and was further identified by means of physicochemical and spectrometric analysis. Both the essential oil and 2-hydroxy-4-methoxybenzaldehyde exhibited antimicrobial activities on all test bacteria and fungi, with MBC/MFC values ranging from 125 microg/mL to 300 microg/mL, MIC values from 80 microg/mL to 300 microg/mL, and IC(50) values from 63.29 microg/mL to 167.30 microg/mL. They also showed moderate antioxidant activity in the DPPH free radical scavenging, beta-carotene bleaching and ferrozine-ferrous ions assays. The results indicated that the major portion of these antimicrobial and antioxidant activities was due to the presence of 2-hydroxy-4-methoxy-benzaldehyde in the oil, which could potentially be developed as an antimicrobial and antioxidant agent in the future.