Unveiling the biological methylation of 2,4,6-trichlorophenol and the formation of 2,4,6-trichloroanisole in a municipal wastewater treatment plant

Odor problems in treated municipal wastewater are a concern, yet the sources and formation dynamics of these compounds within sewerage systems remain unclear. 2,4,6-trichloroanisole (2,4,6-TCA) is a key odorant in the effluents of municipal wastewater treatment plants (WWTPs). This study investigates the formation of 2,4,6-TCA through the conversion of its precursor, 2,4,6-trichlorophenol (2,4,6-TCP). The concentrations of both compounds were quantified across a step-feed anaerobic-anoxic-oxic (A₂O) process over multiple seasons. Effluent concentrations of 2,4,6-TCA consistently exceeded its odor threshold. Inverse correlations between 2,4,6-TCP and 2,4,6-TCA concentrations within the aerobic zones supported the biological transformation. In the anaerobic zones, 2,4,6-TCA also increased only during winter, suggesting a seasonal contribution to elevated 2,4,6-TCA concentrations in effluent during this time. Amplicon sequencing of the activated sludge samples revealed the presence of the bacterial and fungal genera previously associated with methylation of 2,4,6-TCP. Additionally, wastewater samples were collected from multiple WWTPs and pump stations to determine the source of 2,4,6-TCP within the sewerage system. Wastewater from commercial areas exhibited higher 2,4,6-TCP concentrations compared to that from residential areas, suggesting greater contributions from commercial sources. Furthermore, batch experiments suggested that reactions between chlorine and phenolic compounds in wastewater contribute to 2,4,6-TCP formation. These findings highlight the importance of source control of phenolic compounds and chlorine, as well as operational optimization, such as dynamic aeration, to mitigate 2,4,6-TCA formation in wastewater treatment systems.