Dye Decolorization by a Miniaturized Peroxidase Fe-MimochromeVI*a

Abstract

Oxidases and peroxidases have found application in the field of chlorine-free organic dye degradation in the paper, toothpaste, and detergent industries. Nevertheless, their widespread use is somehow hindered because of their cost, availability, and batch-to-batch reproducibility. Here, we report the catalytic proficiency of a miniaturized synthetic peroxidase, Fe-Mimochrome VI*a, in the decolorization of four organic dyes, as representatives of either the heterocyclic or triarylmethane class of dyes. Fe-Mimochrome VI*a performed over 130 turnovers in less than five minutes in an aqueous buffer at a neutral pH under mild conditions.

Keywords: artificial metalloenzyme; bleaching; dye degradation; enzymatic treatment; peroxidase.

Figure 1

MC6*a designed model and its dominant enzymatic activities exhibited with different metals.

Figure 2

Structures of the dyes tested in this work. (a) Neutral Red (NR), Methylene Blue (MB), Xylenol Orange (XO), and Bromophenol Blue (BPB) are shown in one representative resonance/tautomeric structure over several possible ones. The bar plot (b) reports the decolorization percentage for NR (110 μM, red), MB (12.9 μM, cyan), XO (78 μM, orange), and BPB (13.5 μM, blue) when 1 μM FeMC6*a was absent (left, half filled) or present (right, filled) in the reaction mixture (100 mM phosphate buffer, pH 6.5, 0.25 mM H₂O₂).

Figure 3

Decolorization process as followed by UV–Vis spectroscopy. Two spectra are reported for each dye, the first before the addition of H₂O₂ (black dashed line) and the second after the addition of H₂O₂, when no further consumption of the dye was observed (red line). All the reactions were performed in a solution of the dye and 1 μM FeMC6*a in 100 mM phosphate buffer, pH 6.5, to which 0.25 mM H₂O₂ was added. Dye oxidation was monitored over 15 min for NR (60 μM, (a)), MB (12.9 μM, (b)), XO (78 μM, (c)), and BPB (13.5 μM, (d)). Insets show the photographic pictures of the reaction mixtures before (left) and after (right) decolorization.

Figure 4

Decolorization percentage of NR (110 μM (a)) and MB (9.9 μM (b)) as a function of the FeMC6*a concentration. The reaction was performed in 100 mM phosphate buffer, pH 6.5, 3 mM H₂O₂. The dashed line between each point was intended as a guide for the eye, while the error bars defined a 10% confidence level.

Figure 5

Decolorization percentage of NR (110 μM (a)) and MB (9.9 μM (b)) as a function of the H₂O₂ concentration. The reaction was performed in 100 mM phosphate buffer, pH 6.5, 0.5 μM FeMC6*a. The dashed line between each point was intended as a guide for the eye, while the error bars defined a 10% confidence level.

Figure 6

Decolorization percentage of NR (a) and MB (b) as a function of their respective concentrations (26–106 μM and 0.72–10 μM, respectively). The reaction was performed in 100 mM phosphate buffer, pH 6.5, 0.5 μM FeMC6*a, and 3.0 mM H₂O₂. The dashed line between each point was intended as a guide for the eye, while the error bars defined a 10% confidence level.

Figure 7

NR (0.12 mM) degradation followed by UV–Vis absorption spectroscopy. Spectra were acquired every minute for 15 min (colored lines from violet to red) (a). The absorption band decrease at 520 nm is accompanied by the formation of a band at 330 nm (b). The reaction was performed in 100 mM phosphate buffer, pH 6.5, 1.0 μM FeMC6*a, and 0.10 mM H₂O₂.

Figure 8

Oxidation activity of FeMC6*a. (a) Initial rate dependence towards the H₂O₂ concentration. The reaction conditions were FeMC6*a (1 µM) and NR (25 mM) in 100 mM phosphate buffer, pH 6.5. (b) Initial rate dependence towards NR concentration. The reaction conditions were FeMC6*a (1 µM) and H₂O₂ (0.10 M) in 100 mM phosphate buffer, pH 6.5. The data points were fitted using the Michaelis–Menten equation (red lines), and the error bars corresponded to the standard deviation of three repetitions.

Scheme 1

Proposed mechanism for NR oxidation by FeMC6*a.