The paper aims to study process conditions of ultrasonic-microwave synergistic extraction of sea buckthorn seed oil and its influences on the fatty acid composition of the sea buckthorn seed oil. Single factor test was carried out by selecting six factors: solvent type, extraction time, liquid-solid ratio, ultrasonic power, microwave power and extraction temperature. Based on the single factor test, N-hexane was selected as the extraction solvent, and the extraction rate was taken as the response value. Four factors of extraction time, liquid-solid ratio, ultrasonic power and extraction temperature were used to carry out response surface optimization experiment. The fatty acid composition of sea buckthorn seed oil extracted under these conditions and extracted by supercritical carbon dioxide extraction method was analyzed. The optimal extraction conditions were determined as follows: extraction time was 22 min, liquid-solid ratio was 9 mL/g, ultrasonic power was 628 W, microwave power was 200 W, extraction temperature was 57 ℃. Under these conditions, the extraction rate of sea buckthorn seed oil could reach 13.97%. The analysis of fatty acid composition of sea buckthorn seed oil extracted by ultrasonic-microwave synergistic method showed that the content of unsaturated fatty acid was as high as 88.93%, the highest content was linoleic acid, which was 39.60%; followed by linolenic acid, which was 32.60%. Therefore, the ultrasonic-microwave synergistic extraction is an effective method for extracting sea buckthorn seed oil.