Biotransformation Changes Bioaccumulation and Toxicity of Diclofenac in Aquatic Organisms

Biotransformation plays a crucial role in regulating the bioaccumulation potential and toxicity of organic compounds in organisms but is, in general, poorly understood for emerging contaminants. Here, we have used diclofenac as a model compound to study the impact of biotransformation on the bioaccumulation potential and toxicity in two keystone aquatic invertebrates: Gammarus pulex and Hyalella azteca . In both species, diclofenac was transformed into several oxidation products and conjugates, including two novel products, that is, diclofenac taurine conjugate (DCF-M403) and unexpected diclofenac methyl ester (DCF-M310.03). The ratios of biotransformation products to parent compound were 12-17 for DCF-M403 and 0.01-0.7 for DCF-M310.03 after 24 h exposure. Bioconcentration factors (BCFs) of diclofenac were 0.5 and 3.2 L kg ww -1 in H. azteca and G. pulex , respectively, whereas BCFs of DCF-M310.03 was 164.5 and 104.7 L kg ww -1 , respectively, representing a 25- to 110-fold increase. Acute toxicity of DCF-M310.03 was also higher than the parent compound in both species, which correlated well with the increased bioconcentration potential. The LC 50 of diclofenac in H. azteca was 216 mg L -1 , while that of metabolite DCF-M310.03 was reduced to only 0.53 mg L -1 , representing a 430-fold increase in acute toxicity compared to diclofenac. DCF-M403 is less toxic than its parent compound toward H. azteca , which may be linked to its slightly lower hydrophobicity. Furthermore, the transformation of diclofenac to its methyl ester derivative was explored in crude invertebrate extracts spiked with an S -adenosylmethionine cofactor, revealing possible catalysis by an S -adenosylmethionine-dependent carboxylic acid methyltransferase. Methylation of diclofenac was further detected in fish hepatocytes and human urine, indicating a broader relevance. Therefore, potentially methylated metabolites of polar contaminants should be considered for a comprehensive risk assessment in the future.