AbstractPlant materials have been used in different fields such as therapeutics, cosmetics, and energy for ages. Several studies have investigated seeds, whether common or not, in order to ensure a better valuation of natural resources. Among these, Pistacia atlantica Desf. has been the subject of several works on its characterization and valorization. Among the current trends in sustainable development and environmental protection, valorizing natural wild plants via green chemistry has become prevalent. One of these plants, Pistacia atlantica Desf., is a tree that grows in arid and semi‐arid areas, notably in Tunisia, and produces seeds rich in oil. In this study extracting the oil from its seeds is tried by three methods (supercritical carbon dioxide (CO2), pressure, and hexane), and the efficiency of these extraction processes is compared in order to obtain the best yield and maximize its valorization in a variety of industrial fields. The obtained oil is found to be rich in polyunsaturated fatty acids, namely linoleic and oleic acids, with a similar acidic composition among all extraction methods. The tocopherol composition of the oil is determined using high performance liquid chromatography. The total polyphenol content is determined using the Folin‐Ciocalteu colorimetric analysis method. The results show that the seed oil of Pistacia atlantica Desf. extracted by supercritical CO2 gives the highest extraction yield (25%) and the lowest acidity and peroxide values with a high degree of oxidation at 232 and 270 nm. This method also gives the highest content of alpha, gamma, and delta tocopherol as well as total phenolic content compared to the other extraction methods. The composition of chlorophylls and carotenes in the seed oil of Pistacia atlantica Desf. is determined. Besides, the analysis of the sterol composition reveals that β‐sitosterol is still prevailing. Among all the tested extracts, the supercritical CO2 extract demonstrates the best antioxidant performance against the tested radicals. The oil extracted by supercritical carbon dioxide (CO2) is of a higher quality compared to that extracted by pressure and by hexane.Practical applications: The Pistacia atlantica Desf. oil exhibits interesting physicochemical and biological characteristics. The type of extraction affects the quality of the major and minor components in the seeds. However, the supercritical CO2 method produces a better oil quality with the presence of antioxidants such as tocopherols and polyphenols. In fact, it is found that the oil extracted by the three methods has an important sterolic profile with the predominance of β‐sitosterol. According to the classification of vegetable oils, unsaturated fatty acids of atlas pistachio oil can be categorized as oleo linoleic oil, which enhances its nutritional value. Having these characteristics, the use of this oil can be considered in agrifood products.