AbstractLignins are important biopolymers that can be converted into value-added materials by enzymatic treatments. However, the heterogeneity of the lignin polymer makes it a challenging material to modify. Thus, chemical fractionation was used to obtain lignins with high homogeneity in order to assess their biotechnological utilization. Commercial Alcell, birch organosolv lignins, and steam-exploded pine and eucalypt lignins were sequentially fractionated by ether, ether/acetone 4:1 (v:v), and acetone. All fractions were structurally characterized prior to treatments with Thielavia arenaria, Trametes hirsuta, and Melanocarpus albomyces laccases. The reactivities of the enzymes towards the lignins were determined by oxygen consumption measurements, and the degree of polymerization was confirmed by size exclusion chromatography. Field emission scanning electron microscopy revealed that the surfaces of the lignin nanoparticles were dispersed in the enzyme treatment, suggesting an increase in hydrophilicity of the surfaces detected as loosened morphology. Hence, it was concluded that enzyme-aided valorization is an attractive means for lignin modification, provided that optimum reaction conditions are employed.