Green separation of protein (e.g., bovine serum albumin (BSA)) by low-melting mixture solvents (LoMMSs) depends on the underlying mechanism between BSA and LoMMSs. Here, we for the first time find that eco-friendly biomass-derived LoMMSs could be potentially used for the efficient and green purification of BSA protein by enthalpy-driven interactions. Biomass-derived LoMMSs possess the merits of high biocompatibility, high degradability, high abundance, and low cost. A single high-affinity binding site via hydrogen bonding and van der Waals forces is observed between BSA and LoMMSs by fluorescence and thermodynamic analysis. Experimental results from circular dichroism and infrared spectra demonstrate that the addition of LoMMSs stabilizes the secondary structure of the BSA protein. This work provides a valuable indication for the design of eco-friendly and cost-effective LoMMSs for the purification of protein.