The compositions of essential oils (EOs) from Spanish marjoram (Thymus mastichina L.) grown in several bioclimatic zones of Murcia (SE Spain) were studied to determine their absolute and relative concentrations using gas chromatography-mass spectrometry. 1,8-Cineole and linalool were the main components, followed by α-pinene, β-pinene and α-terpineol. (-)-Linalool, (+)-α-terpineol and (+)-α-pinene were the most abundant enantiomers. When the antioxidant capacities of T. mastichina EOs and their compounds were measured by five methods, EOs and linalool, linalyl acetate, α-terpinene and γ-terpinene, among others, showed antioxidant activities. All four T. mastichina EOs inhibited both lipoxygenase and acetylcholinesterase activities, and they might be useful for further research into inflammatory and Alzheimer diseases. Bornyl acetate and limonene showed the highest lipoxygenase inhibition and 1,8-cineole was the best acetylcholinesterase inhibitor. Moreover, these EOs inhibited the growth of Escherichia coli, Staphylococcus aureus and Candida albicans due to the contribution of their individual compounds. The results underline the potential use of these EOs in manufactured products, such as foodstuff, cosmetics and pharmaceuticals.