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

Abstract

BackgroundPeroxidases 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.ResultsIn 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 H2O2 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.ConclusionsDetailed 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.

Differential Degradation and Detoxification of an Aromatic Pollutant by Two Different Peroxidases

“…However, SRB dye degradation by CPO was optimal at pH 2 and further increases in pH caused the degradation to decrease dramatically until it reached almost 25% at pH 4 ( Figure 3A). These results are in agreement with previously published studies with SBP, where maximum degradation of Trypan Blue [35], Crystal Ponceau 6R [36], Amido Black [37], and Remazol…”

“…As can be seen from Figure 3C, there was no significant change in the degradation of the SRB dye by SBP at H2O2 concentrations ranging from 0.2 mM to 16.5 mM. These results, along with those previously reported, suggest that SBP is a remarkably stable enzyme, compared to CPO, as it can tolerate high concentrations of strong oxidants, such as H2O2 [35,36,42].…”

“…However, when 1-hydroxybenzotriazole (HOBT), one of the most commonly used redox mediators, was added to the reaction mixture, rapid and efficient SRB dye degradation was observed ( Figure 2A and Figure 2B). This absolute requirement of HOBT was not observed in the SBP-degradation of many other dyes [27,35], but is consistent with peroxidase-mediated degradation of Crystal Ponceau 6R and ThT dyes [27,36]. Optimization of HOBT concentration for efficient SRB dye degradation showed that 50 μM HOBT was the optimum amount of redox mediator for SBP-based degradation, whereas the CPO-based reaction showed the best degradation of SRB dye at 167 μM HOBT ( Figure 2B).…”

“…The dye degradation assays were carried out in 3 mL total volume by incubating the specific concentration of the dye with either SBP or CPO in the appropriate buffer (pH 4 for SBP and pH 2 for CPO) and different concentrations of hydrogen peroxide and monitored using a 4 ml quartz cuvette in a Spectrophotometer (Cary 60, USA) as previously described [35,36]. When needed, the redox mediator, 1-hydroxybenzotriazole (HOBT), was added in the reaction mixture to help with the dye degradation assay.…”

fficient Aerobic Degradation of Various Azo Dyes by a Sphingomonas sp Isolated from Petroleum Sludge

“…High performance liquid chromatography (HPLC) was carried out as previously described [17]. Briefly, an Agilent PH 1100 liquid chromatography system, (Agilent, USA) with an Agilent Zorbax SB-C18 column 150 mm x 4.6 mm packed with 5 µm particle size, coupled to a diode array detector was used (Agilent, USA).…”