This study investigated the phytochemical compositions of Pinus peuce (Balkan pine) essential oils (EO), total polyphenol, and flavonoid content from different plant parts: leaves, twigs, wood, and cones. Furthermore, this study evaluated the biological activities of the EO. The EO yield varied among different plant parts, and it was the highest in the wood of one to two-year-old twigs (1.08 % – 2.40 %). The predominant compounds of the EOs, α-pinene, β-pinene, and limonene, were found in all analyzed trees from the five populations, but in different amounts. The concentration ranges for α-pinene were 32–56 % in leaves (L), 24–49 % in twigs (T), 26–48 % in tips of twigs (TT), and 10.5–48 % in wood of one to two-year-old twigs. The concentrations of β-pinene were 7.9–18.3 % in L, 5.9 % - 15.7 % in T, 7.9–13.2 % in TT, and 4.2–6.5 % in WT. The concentration of limonene was 2.9–12 % in L, 5.7–45 % in T, and 1.2–60 % in WT. Furthermore, camphene, β-myrcene, bornyl acetate, germacrene D, and β-caryophyllene were found in large quantities in some plant parts. The female cones had the highest concentration of polyphenolic compounds (56.03 mg GAE g−1), followed by the annual wood (24.56 mg GAE g−1) and leaves (22.86 mg GAE g−1). The EO of P. peuce had a high antimicrobial effect against Salmonella enterica subsp. enterica and Escherichia coli. Within 24 h of application, the EOs demonstrated 100 % efficacy against the aphid species Sitobion avenae and Rhopalosiphum padi. P. peuce EO from twigs demonstrated significant bioherbicidal activity against the seeds of weed species Anthemis arvensis and Papaver rhoeas effectively inhibiting sprout and root growth. These results demonstrated the potential of P. peuce EO as a novel ingredient in bioherbicides and plant protection products.