On the basis of single factor experiment, response surface methodology was used to optimize the low-temperature pressing process of basil seed oil, and physicochemical indexes, including color, water content and volatile matter, acid value, peroxide value, oxidation induction time and fatty acid composition of basil seed oil, were detected. The results showed that the preheating temperature and the time of preheating and pressing had significant effects on the oil yield of Basil seeds under certain pressing pressure and temperature. The optimum preparation conditions of basil seed oil were pressing with pressure 41 MPa at 50 ℃ for 1.7 h, and with preheating at 73 ℃ for 3.0 h. Under the controlling of these condition, the oil yield reached 20.33%. The prepared basil seed oil had good color, water content, volatile matter, peroxide value and acid value wthich met the Chinese plant oil standard. Its fatty acid composition was similar to that of flaxseed oil, with better quality and stability. It could be used as a new type of healthy oil in the future.