The present invention relates to a 1,2,3,4,6-O-pentagalloylglucose molecular imprinting monolithic column preparation method, which comprises: using 1,2,3,4,6-O-pentagalloylglucose as a template molecule, using a ternary pore forming agent N,N-dimethylformamide, dimethyl sulfoxide and 1-butyl-3-methyl tetrafluoroborate as a solvent, adding a functional monomer, a cross-linking agent, metal ions, a methyl ether methacrylate oligomer and an initiator, carrying out ultrasonic dissolution, inflating nitrogen, removing dissolved oxygen, and carrying out a polymerization reaction in a water bath so as to finally obtain the 1,2,3,4,6-O-pentagalloylglucose molecular imprinting monolithic column having selectivity. According to the present invention, the imprinting factor of the method can be up to 73.9, the method can be used for solid phase extraction, and the selective purification of the 1,2,3,4,6-O-pentagalloylglucose can be achieved; and the printing polymer has characteristics of simple preparation and good durability, and can be adopted as the solid phase extraction filler so as to provide the effect method for the separation and purification of the 1,2,3,4,6-O-pentagalloylglucose (the purity is greater than 98%).