The effects of different concentration of ethanol, ratio of fatty acid to urea, ratio of fatty acid to ethanol, inclusion time, inclusion temperature and times of inclusion on pinolenic acid in pressed pine seed oil were studied. Based on results from single factor test, the optimal experimental design was carried out according to the principle of Box-Behnken central combination test design. The urea inclusion regression model of pinolenic acid was established using the comprehensive score of pinot acid content and loss rate as the response value. The results showed that the fatty acid∶urea =1∶2.88（g/g）, fatty acid∶ethanol =1∶7.98 （g/mL）, inclusion time was 23.44 h, and the inclusion temperature was 4.18 ℃. Under this condition, the content was increased from 39.27% to 53.36%, with 17.24% loss of pinolenic acid, and the comprehensive score was 0.954 3, and the comprehensive evaluation index was the highest.