Biotransformation of diclofenac by isolated super-degrader Pseudomonas sp. DCα4: Postulated pathways, and attenuated ecotoxicological effects

Abstract

Significant concentrations of emerging xenobiotics, like diclofenac (DCF), possessing severe irreversible eco-toxicological threats, has been detected in aquatic systems worldwide, raising the concerns. This present investigation is intended to explore an efficient solution to support the existing wastewater treatment policies to handle DCF contamination by bacteria-mediated biotransformation. DCF-tolerant bacterial strains were isolated from pharmaceutical wastewater and selected based on their non-virulence nature and degradation ability. Among those, Pseudomonas sp. DCα4 was found to be the most dominant DCF degrader exhibiting 99.82% removal of DCF confirmed by HPLC after optimization of temperature at 30.02 °C, pH at 6.9, inoculum of 4.94%, and time 68.02 h. The degradation kinetics exhibited the process of DCF degradation followed a first-order kinetics with k of 0.108/h and specific degradation rate of 0.013/h. Moreover, the enzyme activity study indicated predominant hydrolase activity in the DCF treatment broth of DCα4, implying hydrolysis as the main force behind DCF biotransformation. HRMS analysis confirmed the presence of 2-hydroxyphenylacetic acid, 1,3-dichloro,2-amino, 5-hydroxybenzene, and benzylacetic acid as major intermediates of DCF biodegradation indicating non-specific hydrolysis of DCF. Whole genome analysis of most related strains which were confirmed by near full 16S rRNA gene sequence homology study, predicted involvement of different N–C bond hydrolase producing genes like puud, atzF, astB, nit1, and nylB. The ecotoxicological study using Aliivibrio fischeri exhibited 47.51% bioluminescence inhibition by DCF-containing broth which was comparable to the same caused by 1 mg/mL of K2Cr2O7 whereas remediated broth exhibited only 0.51% inhibition implying reduction of the ecotoxic load caused by DCF contamination. Cost analysis revealed that possible integration of the process with existing ones would increase per litre expense by $0.45. These results indicated that the described process of DCF biodegradation using the super-degrader DCα4 would be an advancement of existing pharmaceutical wastewater treatment processes for DCF bioremediation.