A preparation method of paeoniflorin and albiflorin, which comprises the following steps: a) percolating and extracting Paeonia Lactiflora by percolation extraction process to obtain Paeonia Lactiflora extract; and b) carrying out macroporous resin column chromatography to the Paeonia Lactiflora extract to obtain macroporous resin column separation mixture; c) carrying out alumina column chromatography to the macroporous resin column separation mixture, the alumina column chromatography comprises: c-1) dissolving the macroporous resin column separation mixture; adding aluminum oxide into the macroporous resin column separation mixture solution, and drying to obtain alumina chromatography sample; c-2) arranging the alumina chromatography sample at the top of the alumina column; eluting the alumina column with ethyl acetate as eluent; collecting fluid from the alumina column after eluting; combining paeoniflorin and/or albiflorin flows/flow in the fluid, to obtain alumina chromatography eluting liquid; and c-3) drying the alumina chromatography eluting liquid to obtain alumina chromatography mixture; d) carrying out silica gel column chromatography to the alumina chromatography mixture, wherein the silica gel column chromatography comprises: d-1) dissolving the alumina chromatography mixture, adding silica gel into the alumina chromatography mixture solution, and then drying to obtain silica gel chromatography sample; d-2) arranging the silica gel chromatography sample at the top of the silica gel column; eluting the silica gel column with ethyl acetate as eluent; collecting fluid from the silica gel column after eluting; combining paeoniflorin or albiflorin flow in the fluid respectively; and d-3) evaporating the flows containing paeoniflorin or albiflorin to dryness respectively, to obtain two separate products paeoniflorin and albiflorin. a) percolating and extracting Paeonia Lactiflora by percolation extraction process to obtain Paeonia Lactiflora extract; and b) carrying out macroporous resin column chromatography to the Paeonia Lactiflora extract to obtain macroporous resin column separation mixture; c) carrying out alumina column chromatography to the macroporous resin column separation mixture, the alumina column chromatography comprises: c-1) dissolving the macroporous resin column separation mixture; adding aluminum oxide into the macroporous resin column separation mixture solution, and drying to obtain alumina chromatography sample; c-2) arranging the alumina chromatography sample at the top of the alumina column; eluting the alumina column with ethyl acetate as eluent; collecting fluid from the alumina column after eluting; combining paeoniflorin and/or albiflorin flows/flow in the fluid, to obtain alumina chromatography eluting liquid; and c-3) drying the alumina chromatography eluting liquid to obtain alumina chromatography mixture; c-1) dissolving the macroporous resin column separation mixture; adding aluminum oxide into the macroporous resin column separation mixture solution, and drying to obtain alumina chromatography sample; c-2) arranging the alumina chromatography sample at the top of the alumina column; eluting the alumina column with ethyl acetate as eluent; collecting fluid from the alumina column after eluting; combining paeoniflorin and/or albiflorin flows/flow in the fluid, to obtain alumina chromatography eluting liquid; and c-3) drying the alumina chromatography eluting liquid to obtain alumina chromatography mixture; d) carrying out silica gel column chromatography to the alumina chromatography mixture, wherein the silica gel column chromatography comprises: d-1) dissolving the alumina chromatography mixture, adding silica gel into the alumina chromatography mixture solution, and then drying to obtain silica gel chromatography sample; d-2) arranging the silica gel chromatography sample at the top of the silica gel column; eluting the silica gel column with ethyl acetate as eluent; collecting fluid from the silica gel column after eluting; combining paeoniflorin or albiflorin flow in the fluid respectively; and d-3) evaporating the flows containing paeoniflorin or albiflorin to dryness respectively, to obtain two separate products paeoniflorin and albiflorin. d-1) dissolving the alumina chromatography mixture, adding silica gel into the alumina chromatography mixture solution, and then drying to obtain silica gel chromatography sample; d-2) arranging the silica gel chromatography sample at the top of the silica gel column; eluting the silica gel column with ethyl acetate as eluent; collecting fluid from the silica gel column after eluting; combining paeoniflorin or albiflorin flow in the fluid respectively; and d-3) evaporating the flows containing paeoniflorin or albiflorin to dryness respectively, to obtain two separate products paeoniflorin and albiflorin.