In this study, we investigate the effect of pretreatment method on lipid extraction from Enteromorpha intestinalis using physical, thermo-chemical, and enzymatic process such as ultrasonication, high temperature treatment, freezing, microwave irradiation, osmotic shock, pH shock, homogenizing, and enzymatic treatment. In pretreatment with separated lipid extraction, the high extraction yield was obtained by high temperature treatment ($121^{circ}C$ for 5 min) with 0.1 N HCl, which is 1.4 times higher than that of control. In pretreatment with direct lipid extraction, the high extraction yields were obtained by 0.1 N HCl pretreatment, microwave irradiation (700W, 1 min with twice), and 10% NaCl pretreatment, which is 1.45 times higher than that of control. In the result of enzymatic pretreatment with 17 kinds of enzymes, Cellic CTec II showed the high extraction yield of 5.3%, and which is 1.9 times higher than that of control. Moreover, the extraction yield was increased by the increase of enzyme amounts. In 10% enzyme amount, about 5.8% yield was obtained.