Abstract A novel strategy was applied in the oriented immobilization of laccase from Echinodontium taxodii on concanavalin A-activated Fe3O4 nanoparticles (GAMNs-Con A) based on laccase surface analysis. These nanoparticles showed higher enzyme loading and activity recovery compared with conventional covalent binding. Along with the improvement in thermal and operational stabilities, the oriented immobilized laccase (GAMNs-Con A-L) exhibited higher substrate affinity than free laccase. Free laccase and GAMNs-Con A-L were then applied in the removal of sulfonamide antibiotics (SAs). Although both free and immobilized laccase resulted in the rapid removal of SAs, GAMNs-Con A-L showed a higher removal rate of SAs compared with the free counterpart in the presence of S-type compounds present in lignin structure. Syringic acid mediated the fastest removal efficiency of SAs among S-type compounds and resulted in an almost complete removal of these substances after incubation for 5min. The oxidation products of SAs were identified via LC-ESI+-MS. The results suggested the transformation of SAs and S-type compounds were catalyzed by laccase, resulting in the formation of cross-coupled products.