The invention belongs to the technical field of food detection and in particular relates to a method for determining aflatoxin B1 in gardenia yellow. The method comprises the following steps of: weighing a certain amount of gardenia yellow powdery sample; extracting, concentrating, purifying and filtering the gardenia yellow powdery sample by trichloromethane; and then determining the aflatoxin B1 by means of ultra-high performance liquid chromatograph-tandem mass spectrum. By means of the characteristic that the aflatoxin B1 is insoluble in water but soluble in trichloromethane while gardenia yellow is soluble in water but insoluble in trichloromethane, liquid-liquid extraction is performed on a gardenia yellow aqueous solution with trichloromethane, so that interference with a target object due to a sample matrix is overcome, the influence of matrix effect is reduced, a false positive or false negative result can be effectively avoided, and the method is accurate to quantify.