Abstract: In order to improve the yield of DHA of Crypthecodinium cohnii LS1057, the fermentation medium was optimized.Plackett-Burman design was used to evaluate the effects of medium components on the yield of DHA by C.cohnii LS1057.The results showed that glucose, yeast extract and KH2PO4were the most important factors that influenced the yield of DHA. Based on the experimental results, the path of steepest ascent was undertaken to approach the optimal region of these factors, central composite design and response surface analysis were subsequently employed for further optimization.The optimal medium for C.cohnii LS1057 was obtained, which was composed of 79.76g /L glucose, 14.0g /L yeast extract, 0.5g /L KH2PO4, 20.0g /L sea salt, 5.0g /L MgSO4·7H2O, 8.0g /L KNO3, 0.2g /L FeSO4·7H2O and solution M ( 0.6g /L V B1, 0.1mg /L V B12) 1% ( V/V) . Under the optimum conditions, the maximum yield of DHA reached 1.833g /L in theory and 1.811g /L in the experiment, with an increase of 71.33% compared to the original medium components.The DHA yield of 70 L fermentor reached 2.112g /L under such optimum conditions.The experimental results showed that the conditions optimization by the response surface method were better for enhanced yield of DHA by C.cohnii LS1057.