Characterization of the catabolic pathway of diclofenac in Raoultella sp. KDF8

The removal of diclofenac, which is a widely used non-steroidal anti-inflammatory drug and a dangerous environmental contaminant of surface water, was studied. Its complete degradation by Raoultella sp. KDF8 to primary metabolic products was described with individual metabolites identified by mass spectrometry. The Raoultella strain KDF8 was cultivated with unlabelled and (13)C6-labelled diclofenac. A total of 32 compounds were detected as intermediates in diclofenac metabolism by high resolution tandem mass spectrometry. Furthermore, proteomic analysis showed that the presence of diclofenac resulted in the upregulation of 24 enzymes associated with the benzoate, catechol, protocatechuate and ketoadipate pathways. The presence of genes encoding specific enzymes, such as catechol 1,2-dioxygenase, protocatechuate 3,4-dioxygenase and quercetin 2,3-dioxygenase, has been verified by sequence analysis of PCR products. The description of diclofenac metabolism by strain KDF8 provides a detailed view of the degradation of xenobiotic aromatic compounds by microorganisms in general.