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The effective, energy-saving and green subcritical fluid extraction (SFE) technology was applied to obtain the oil from <i>Lycium ruthenicum</i> seeds (LRSO). The optimal conditions of extraction parameters were found using response surface methodology with Box-Behnken experimental design. The maximum extraction yield of 21.20% was achieved at raw material particle size of 0.60 mm, extraction pressure of 0.63 MPa, temperature of 50 °C and time of 48 min. Other traditional extraction technologies were comparatively used. The physicochemical property of LRSO was analysed and the chemical compositions indicated that they were rich in unsaturated fatty acid, β-carotene, tocopherols and total phenolics. Furthermore, the antioxidant activity of LRSO was evaluated by scavenging activity of three kinds of radicals (DPPH·, ·OH and O₂⁻·) and lipid peroxidation <i>in vitro</i>. And its results showed the oil had the potential to be a novel antioxidant agent for using in the field of food, pharmaceuticals and cosmetics.<br>Lycium ruthenicum seeds oil (LRSO) was obtained by subcritical fluid extraction (SFE), and the process of SFE was optimized using response surface methodology. LRSO was evaluated by determination of physicochemical property, lipophilic compositions and antioxidant activity. The study revealed the possibility of LRSO as a potential source of valuable product for commercial ventures (food, pharmaceuticals or cosmetics).