The application of hydrophobic ionic liquids (ILs) as promising alternatives to volatile organic solvents in liquid-liquid extraction processes of biomolecules was evaluated through the determination of the partition coefficients of the aminoacid L-tryptophan, taken as model biomolecule, between aqueous solutions and distinct ILs. Factors affecting the effectiveness of the recovery such as the pH of the aqueous medium, the nature of the IL anion and the nature and the chemical structure of the IL cation were assessed. The results show that the pH of the aqueous phase strongly influences the success of the separation and that the anion/cation hydrophobic characters are main structural factors ruling the extraction efficiency. The evidences gathered in this work suggest that L-tryptophan partitioning between the aqueous and IL phases is ruled by a complex interplay of intermolecular forces between the solute and the IL solvent, such as electrostatic interactions between the cationic form of the aminoacid and the anion of the IL and interactions established at the level of the IL cation. These findings support earlier molecular interpretations of the mechanisms that govern the partitioning of biomolecules between ILs and aqueous phases.