Colloidal lignin particles (CLPs) have sparked various intriguing insights toward bio-polymeric materials and triggered many lignin-featured innovative applications. Here, we report a multi-solvent sequential fractionation methodology integrating green solvents of acetone, 1-butanol, and ethanol to fractionate industrial lignin for CLPs fabrication. Through a rationally designed fractionation strategy, multigrade lignin fractions with variable hydroxyl group contents, molecular weights, and high purity were obtained without altering their original chemical structures. CLPs with well-defined morphology, narrow size distribution, excellent thermal stability, and long-term colloidal stability can be obtained by rational selection of lignin fractions. We further elucidated that trace elements (S, N) were reorganized onto the near-surface area of CLPs from lignin fractions during the formation process in the form of -SO42− and -NH2. This work provides a sustainable and efficient strategy for refining industrial lignin into high-quality fractions and an in-depth insight into the CLPs formation process, holding great promise for enriching the existing libraries of colloidal materials.