【Purposes】 Deep eutectic solvent (DES) can be used for the extraction of lignin from lignocellulosic biomass owing to its simple preparation, good solubility of lignin, low cost, and easy recycling. In order to further improve the delignification effect of DES under mild conditions, a multi-stage treatment process with gradually increasing temperature is proposed. 【Methods】 Poplar powder was treated at 80, 100, and 120 ℃, and each treatment lasted for 4 h. 【Findings】 The lignin extraction ratio of poplar powder after three treatments is 89.6%, which is 22.2% higher than that of a single treatment at 120 ℃ for 12 h. The effects of biomass fractionation condition on the structure of cellulose and on the molecular weight of regenerated lignin were determined by Fourier transform infrared spectroscopy, scanning electron microscopy, X-ray diffraction, and gel permeation chromatography. Compared with single treatment, multi-stage treatment by DES can retain high crystallinity cellulose better, and efficiently extracts lignin from poplar powder while avoiding lignin repolymerization, which is conducive to the graded and targeted utilization of different components of lignocellulose.