The invention provides a microbial oil preparation method. The microbial oil preparation method comprises the steps that fermentation is conducted to obtain oil-producing microbial fermentation liquor; the fermentation liquor is concentrated to obtain concentrated fermentation liquor with the solid content higher than 10%; the PH value of the fermentation liquor is regulated between 5 and 8.1, phospholipase A is added into the fermentation liquor, the temperature is controlled between 38 DEG C and 51 DEG C, and the fermentation liquor is stirred and sheared for 2-6 hours; the oil phase and the water phase are separated to obtain microbial oil with the phosphorus content lower than 5 ppm. The microbial oil preparation method has the advantages that the degumming process and the microorganism wall breaking process are combined, so that oil preparation processes are reduced, and production cost is lowered; furthermore, the degumming process is conducted before the crude oil preparation process, so that produced crude oil is higher in quality.