Volatile compounds from fruits of Peucedanum alsaticum L. were isolated by hydrodistillation (HD) and headspace solid-phase microextraction (HS-SPME), and then analyzed by GC-MS. The predominant volatile compounds were α-pinene, sabinene, limonene, and β-phellandrene (more than 60% of the oil). The β-pinene, β-myrcene, camphene, β-caryophyllene, bornyl acetate, and germacrene D content was also high. The in-vitro antibacterial activity of the essential oil and some of its components against ten reference strains of Gram-positive and Gram-negative bacteria was evaluated by the agar dilution method. The essential oil and the compounds were more active against Gram-positive than Gram-negative bacteria.