Soybean peroxidase-mediated degradation of an azo dye- A detailed mechanistic study

Background: Peroxidases are emerging as an important class of enzymes that can be used for the efficient degradation of organic pollutants. However, detailed studies identifying the various intermediates produced and the mechanisms involved in the enzyme-mediated pollutant degradation are not widely published. Results: In the present study, the enzymatic degradation of an azo dye (Crystal Ponceau 6R, CP6R) was studied using commercially available soybean peroxidase (SBP) enzyme. Several operational parameters affecting the enzymatic degradation of dye were evaluated and optimized, such as initial dye concentration, H₂O₂ dosage, mediator amount and pH of the solution. Under optimized conditions, 40 ppm dye solution could be completely degraded in under one minute by SBP in the presence of H₂O ₂ and a redox mediator. Dye degradation was also confirmed using HPLC and TOC analyses, which showed that most of the dye was being mineralized to CO₂ in the process. Conclusions: Detailed analysis of metabolites, based on LC/MS results, showed that the enzyme-based degradation of the CP6R dye proceeded in two different reaction pathways- via symmetric azo bond cleavage as well as asymmetric azo bond breakage in the dye molecule. In addition, various critical transformative and oxidative steps such as deamination, desulfonation, keto-oxidation are explained on an electronic level. Furthermore, LC/MS/MS analyses confirmed that the end products in both pathways were small chain aliphatic carboxylic acids.