Alkylphenol polyethoxylates (APEOn) are the second-largest category of commercial nonionic surfactants, which are difficult to degrade naturally in the environment. This study examined the degradation of octylphenol polyethoxylate (OPEOn) by laccase and its laccase-mediated systems. The results showed that OPEOn was poorly degraded by laccase alone. 2, 2'-azino-bis [3-ethylbenzothiazoline-6-sulphonic acid] (ABTS), 1-hydroxybenzotriazole (HBT), and 2, 2, 6, 6-tetramethylpiperidine-1-oxyl (TEMPO) were selected as the redox mediators. Experimental results also indicated that 52.4% of the initial OPEOn amount was degraded by laccase in the presence of TEMPO. The degradation efficiency was analyzed using high-performance liquid chromatography. Furthermore, the structural characteristics of the degradation products were measured using matrix-assisted laser desorption/ionization-time of flight mass spectrometry and nuclear magnetic resonance spectroscopy, and it could be found that the laccase-TEMPO system could gradually shorten the ethoxylate chain by oxidizing the primary hydroxyl group of OPEOn, thereby degrading the OPEOn of the macromolecule into small molecules. The maximum of the ion peak distributions of OPEOn decreased from n = 8 finally down to 3. The novel enzymatic system introduced by this study will become a promising alternative method for high-efficiency APEOn conversion and had great potential value in wastewater treatment. © 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.