Diclofenac (DCF) is a highly consumed non-steroidal anti-inflammatory drug that is excreted partially metabolized and is poorly removed during wastewater treatment. Previous findings demonstrated that DCF in wastewater treatment plants (WWTP) is partially removed to nitro/nitroso compounds. The reactive nitrogen species, that are microbially produced during nitrification in the activated sludge of WWTP, were suspected to be involved in the transformation of DCF. Therefore, here, we investigated the molecular features governing such biotransformation and the role played by nitrifying bacteria by biodegradation experiments at lab scale in enriched nitrifying sludge bioreactors spiked with DCF and other structurally related non-steroidal anti-inflammatory drugs (NSAIDs). We provided evidence of the incorporation of NO/NO2 groups into DCF originated from ammonia by isotopically labelled biodegradation experiments. Nitroso and nitro-derivatives were tentatively identified for all NSAIDs studied and biotransformation mechanisms were proposed. Our findings from biodegradation experiments performed under different incubation conditions suggested that biotransformation of DCF and its related NSAIDs might not only be microbially mediated by ammonia oxidizing bacteria, but other nitrifiers co-occurring in the activated sludge as ammonia oxidizing archaea and nitrite oxidizing bacteria. Follow-up studies should be conducted to disentangle such complex behaviour in order to improve removal of these contaminants in WWTPs.
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