Stability and Catalase-Like Activity of a Mononuclear Non-Heme Oxoiron(IV) Complex in Aqueous Solution

Abstract

Heme-type catalase is a class of oxidoreductase enzymes responsible for the biological defense against oxidative damage of cellular components caused by hydrogen peroxide, where metal-oxo species are proposed as reactive intermediates. To get more insight into the mechanism of this curious reaction a non-heme structural and functional model was carried out by the use of a mononuclear complex [Fe^II(N4Py*)(CH₃CN)](CF₃SO₃)₂ (N4Py* = N,N-bis(2-pyridylmethyl)- 1,2-di(2-pyridyl)ethylamine) as a catalyst, where the possible reactive intermediates, high-valent Fe^IV=O and Fe^III-OOH are known and spectroscopically well characterized. The kinetics of the dismutation of H₂O₂ into O₂ and H₂O was investigated in buffered water, where the reactivity of the catalyst was markedly influenced by the pH, and it revealed Michaelis-Menten behavior with K_M = 1.39 M, k_cat = 33 s^-1 and k₂(k_cat/K_M) = 23.9 M^-1s^-1 at pH 9.5. A mononuclear [(N4Py)Fe^IV=O]²⁺ as a possible intermediate was also prepared, and the pH dependence of its stability and reactivity in aqueous solution against H₂O₂ was also investigated. Based on detailed kinetic, and mechanistic studies (pH dependence, solvent isotope effect (SIE) of 6.2 and the saturation kinetics for the initial rates versus the H₂O₂ concentration with K_M = 18 mM) lead to the conclusion that the rate-determining step in these reactions above involves hydrogen-atom transfer between the iron-bound substrate and the Fe(IV)-oxo species.

Keywords: catalase activity; hydrogen peroxide; iron(IV)-oxo; kinetic studies; oxidation.

Scheme 1

Structures of (1), (2) and (3).

Figure 1

Kinetics of hydrogen peroxide degradation catalyzed by 1 in water: (a) pH dependence of hydrogen peroxide degradation determined by volumetrically measuring the evolved dioxygen in the presence (●) and in the absence (○) of 1. The inset shows the time traces for the reaction of 0.275 mM 1 with 0.35 M H₂O₂ at pH 8, 9.5 and 11 at 20 °C. (b) V_in versus [H₂O₂]₀ at [1] = 2.75 × 10⁻⁴ M, pH 9.5 (borate buffer) and 20 °C. The inset shows the time traces for the reaction of 0.275 mM 1 with H₂O₂ (0.11–1.29 M).

Figure 2

Reaction of 2 with H₂O₂ in acetonitrile: (a) UV-Vis spectra of the reaction of 1.5 mM 2 in CH₃CN with 75 equiv of H₂O₂ at 10 °C (path length, 1 cm). Inset: Time course of the reaction monitored at 705 nm (2) and 535 (3). (b) UV/Vis spectra of the decay of 3 generated based on (a). Inset: Time course of the decay of 3 in CH₃CN and CH₃CN/H₂O (v/v = 1:1) solution at 10 °C.

Figure 3

Time course of the decay of 2 monitored at 697 nm at different pH in the presence (●) and in the absence (○) of H₂O₂ at 10 °C. Conditions: [2] = 1.5 mM; [H₂O₂]₀ = 15 mM in MeCN/H₂O (2 cm³, v/v = 1:1, path = 1 cm). (a) pH 7: 0.1 M KH₂PO₄/0.1 M NaOH. (b) pH 8: 0.025 M Na₂B₄O₇.10H₂O/0.1 M HCl. (c) pH 9: 0.05 M NaHCO₃/0.1 M KOH. (d) pH 9.5: 0.05 M NaHCO₃/0.1 M KOH. (e) pH 10.5: 0.05 M NaHCO₃/0.1 M KOH. (f) pH 11: 0.05 M NaHCO₃/0.1 M KOH. I = 0.15 M KNO₃.

Figure 4

(a) Reaction rates of the decay of 2 monitored at 697 nm at different pH values in the presence (●) and in the absence (○) of H₂O₂ and their normalized values (◊) in buffered CH₃CN/H₂O (v/v = 1:1) solution (pH 7–11) at 10 °C. (b) Reaction of 2 with H₂O₂ in buffered CH₃CN/H₂O: UV-Vis spectra of the reaction of 1.5 mM 2 in buffered CH₃CN/H₂O (pH 10, v/v = 1:1) with 10 equiv of H₂O₂ at 10 °C (path length, 1 cm). Inset: Time course of the reaction monitored at 697 (2) and 490 nm in buffered CH₃CN/H₂O (v/v = 1:1) solution (pH 10) at 10 °C.

Figure 5

Kinetic studies on the reaction of 2 with H₂O₂ in buffered MeCN/H₂O solution at pH 8 and 10 °C. (a) UV-vis spectral change of 1.5 mM 2 upon addition of 10 equiv of H₂O₂. Inset shows time course of the decay of in the absence (○) and in the presence of H₂O₂ in MeCN/D₂O (□) and MeCN/H₂O (●) solution, respectively. (b) Plot of k_obs versus [H₂O₂]₀ at [2] = 1.5 mM, pH 8 and 10 °C.

Scheme 2

Proposed mechanism for the oxoiron(IV)-mediated H₂O₂ oxidation.