Biologically active micropollutants that contain diverse phenolic/aromatic structures are regularly present in wastewater effluents and are even found in drinking water. Advanced green technologies utilizing immobilized laccase and/or peroxidase, which target these micropollutants directly, may provide a reasonable alternative to standard treatments. Nevertheless, the use of these enzymes is associated with several issues that may prevent their application, such as the low activity of laccase at neutral and basic pH or the necessity of hydrogen peroxide addition as a co-substrate for peroxidases. In this study, the activity of laccase from Trametes versicolor and horseradish peroxidase was evaluated across a range of commonly used substrates (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), syringaldazine, and guaiacol). Moreover, conditions for their optimal performance were explored along with an assessment of whether these conditions accurately reflect the effectivity of both enzymes in the degradation of a mixture of bisphenol A, 17α-ethinylestradiol, triclosan, and diclofenac in tap drinking water and secondary wastewater effluent. Laccase and horseradish peroxidase showed optimal activity at strongly acidic pH if ABTS was used as a substrate. Correspondingly, the activities of both enzymes detected using ABTS in real waters were significantly enhanced by adding approximately 2.5% (v/v) of McIlvaine's buffer. Degradation of a mixture of micropollutants in wastewater with 2.5% McIlvaine's buffer (pH 7) resulted in a substantial decrease in estrogenic activity. Low degradation efficiency of micropollutants by laccase was observed in pure McIlvaine's buffer of pH 3 and 7, compared with efficient degradation in tap water of pH 7.5 without buffer. This study clearly shows that enzyme activity needs to be evaluated on micropollutants in real waters as the assessment of optimal conditions based on commonly used substrates in pure buffer or deionized water can be misleading.
Keywords: Anthropogenic pollution; Biodegradation; Ecotoxicity; Endocrine-disrupting chemicals; Enzymatic treatment; Wastewater treatment.