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N2O degradation

Published on 24 April 2018

Dr Stéphane Torelli
CNRS Researcher
Laboratoire Chimie et Biologie des Métaux
17 avenue des Martyrs
38 054 Grenoble Cedex 09
Phone: 33 (0)4 38 78 91 07
Fax: 33 (0)4 38 78 91 24

Nitroux oxide activation using transition metal complexes

N2O is a major greenhouse gas that is 300 times more powerful than CO2[1] and also a subsequent ozone depleting agent [2]. Additionally, this molecule is an appealing oxidant owing to its thermodynamic potency and environmentally friendly nature, as only N2 is the sole by-product. However, efforts to utilize N2O as an oxygen transfer oxidant are disturbed by its high kinetic stability [3]. The literature is not really documented in this field.
2O is also involved in biological processes since it is reduces, during the bacterial denitrification process, into N2 and H2O. The enzyme that performs this reaction, Nitrous oxide reductase (N2Or) has been identified and its X-Ray crystal structure solved. A way to prepare molecules activie towards N2O could consequently consist in preparing copper complexes bearing a similar environment than that of N2Or. Driven by our work in the domain on copper-sulphur chemistry, we investigate a bio-inspired approach of N2Or [4]. Several motivations arise from this approach. This metalloprotein has been studied for more than 30 years [5-10] and two types are identified up to now: Copper-Iron N2Or [11-13] and Copper-only N2Or. The latter are of interest here and consist in head-to-tail homodimers and several X-ray crystal structures are available (Figure 1) [14-17].

Figure 1. From left to right: Overall view of the crystal structure of the N2Or dimer from Achromobacter cycloclastes. One monomer is uniformly coloured grey and the other violet; Structures of Cuz and CuA clusters. Protonation states have been assigned independently.

In all cases the protein crystallizes as a homodimer (~66 kDa each) with two different copper sites, CuA and CuZ. CuA is well-known in biology for promoting rapid electron transfer processes. These electrons are needed at the “real” CuZ active site in order to recover a catalytic redox form. CuA can accommodate both CuI and CuII and exists as a delocalized mixed valent form Cu1.5+Cu1.5+ in the resting form. The oxidized Cu2II state generated upon electron release is reduced by electrons coming by example from c-type cytochrome.
The CuZ center is our principal target here because it is the location of N2O reduction. Depending on the genomic background and on the purification procedures, different redox forms have been identified. These forms were named according to colour exhibited by the isolated protein and reflect the redox state of both CuA and CuZ sites [4]. Globally, this novel structural motif consists of a µ4-sulfide-bridged tetranuclear copper cluster with a distorted tetrahedral shape. Three of the copper ions are coordinated by two histidine ligands, whereas the fourth one has only one histidine ligand. The first published X-Ray crystal structure shows a OH- residue coordinated to one Cu and a water molecule at a Cu2 edge. More precisely, the CuZ centre has been characterized as the so-called CuZ or CuZ* forms. The former is redox active while the latter is redox inactive. The resting form has been characterized by spectroscopy and contains a 1CuII/3CuI redox state with a total spin of 1/2. Electron Paramagnetic Spectroscopy (EPR) and Density Functional Theory (DFT) indicate that the spin density is delocalized over the entire cluster, leading to a mixed-valence system [18-21]. N2O reduction is supposed to occur at the fully reduced state (CuI4), and calculations argue for a mechanism involving a bent binding of the substrate to two copper centres (Figure 2). Only two of the four copper might be involved in the process [20, 23].

Figure 2. Proposed mechanism for N2O reduction.

A key point in all these structures is the presence of an exogenous ligand at one copper edge. The nature of this ligand has been controversial for several years and remains unclear since a recent report in Nature by Pomowski et al [23]. Even if an hydroxo group was suspected so far, they identify a sulphur residue (Figure 8, left). As the solved structure comes from a different isolated organism, it is difficult to rationalize these observations. More importantly, under pressurized-N2O conditions, crystals have been obtained with a N2O molecule trapped close to the active site (Figure 3, right).

Figure 3. CuZ site from Pseudomonas stutzeri with selected bond distances.

Even if it appears that N
2O interacts with quite all the atoms of the active site, it does not mean that this depiction is relevant to the substrate fixation. Consequently, driven by the proposal of Gorelsky et al, we decided to investigate the chemistry of {Cu2S}-containing edifices towards N2O-activation. To closely resemble the active site, we used pyridine groups as N-donating atom and the sulphur was introduced using thiophenol derivatives. The first generation of ligand prepared harbours bis-pyridylmethylamine as coordinating unit. For storage purposes, the isolation of the disulphide intermediate is of interest (Figure 4).
Concomitantly with trials for reducing the disulphide and engage the Cu
I chemistry, we investigated the reactivity of our disulphide derivative with CuI. We basically expected the formation of a tetranuclear copperI complex that could structurally highly mimic N2Or active site. The reaction of Me(BPA)LS-S with adapted amount of CuI salt allowed the isolation of a paramagnetic species. This result was not consistent with the presence of diamagnetic CuI centres. X-ray crystallography measurements, together with a complete spectroscopic and theoretical work (in collaboration with M. Orio, University of Lille 1, LASIR laboratory) allow to refine the nature of this new compound that was finally identified as a dinuclear mixed-valent species containing a {Cu2S}2+ core. Additionally, a striking 2.57 Å Cu-Cu bond was proposed according to the crystallographic structure (Figure 4a). The relevance of this intermetallic bond was also confirmed by calculations through Natural Bond Order (NBO) analysis (Figure 4b). A clear overlap between two 4s/3d hybrid orbitals is observed. This bond appears to be, from the best of our knowledge, the longest Cu-Cu bond ever reported in the literature [24].

Figure 4. Structure of the S-based ligand and its reactivity with CuI;
a) X-ray crystal structure of the mixed-valent dicopper complex exhibiting the {Cu2S} core;
b) NBO analysis of the complex showing the relevance of the Cu-Cu bond.

The disappointing result was that this compound is unreactive towards N
2O. The hypothesis was that more exchangeable positions at the metal centres could be necessary to allow the fixation of N2O which is a poorer ligand compared to O2 was the starting point of the preparation of a new ligand. The initial L(Me)S-S structure was modify by replacing one pyridylmethyl group on each tripodal N atom with a methyl substituent. The new Me(MAM)LS-S ligand was thus prepared and its copper chemistry investigated (Figure 5). Depending on the solvent and the Cu
I salt, two compounds were isolated. In each case a MV compound was obtained, with a Cu-Cu bond but with completely different coordination spheres. Except the thiophenolate, an exclusive N-containing one is present in [2-(CH3CN)2] a mixed N,O environment is achieved with [2-(H2O)(CF3SO3)].

Figure 5. Mixed-valent dicopper complexes isolated starting from Me(MAM)LS-S depending on experimental conditions.

[2-(CH3CN)2] and [2-(H2O)(CF3SO3] were fully characterized by UV-Vis, EPR, electrochemistry and elemental analysis.
While [1-(CF3SO3)2] was totally unreactive, [2-(H2O)(CF3SO3)] and [2-(CH3CN)2] are sensitive to N2O. UV-Vis and EPR changes upon N2O exposure are depicted in Figure 6.

Figure 6. Evolution of the UV-Vis spectra of [2-(H2O)(CF3SO3)]
(A) and [2-(CH3CN)2](C) upon exposure to N2O in acetone;
(B) modification of the X-Band EPR (10K) spectrum of [2-(H2O)(CF3SO3)] upon N2O bubbling (final spectrum recorded after 6 min).

These results clearly indicate an interaction between both complexes and the substrate. In UV-Vis, the disappearance of the IVCT transition is related to changes in the redox states of the copper ions. The LMCT band also vanished, concomitantly with the apparition of new transitions around 420 nm. In EPR, we observed the loss of around 70 % of the signal upon exposure of
[2-(H2O)(CF3SO3)] to N2O for 6 minutes. The new species is thus EPR silent and we postulated the formation of cupric species in which the metal ions are relatively bly antiferromagnetically coupled. This hypothesis was comforted by the isolation of single crystals of the final product (Figure 7, left).

Figure 7.

The structure shows the presence of a dinuclear copper
II complex in which the metal ions are bridged with the sulphur atom from the starting ligand and by a hydroxo ion. The coordination sphere is complete by the nitrogen atoms from the ligand and by two triflates counter-ions. All the copper ions being at a (+II) oxidation state, we then assumed that N2O was reduced by our complexes and that N2 was released. GC-MS experiments were then performed and unambiguously shown that extra N2 was present in the headspace gas of the experiment compared to the blank and to the stable amount of O2 contamination (Figure 7, right).
This result gave the irrefutable proof that a minimalist {N
4Cu2S} core can perform N2O reduction and validates our starting hypothesis. Current work is in progress in order to get mechanistic insights, structure of putative intermediates and to prepare more reactive copper complexes.


Esmieu C, Orio M, Mangue J, Pécaut J, Ménage S and Torelli S
Valence localization at a bio-inspired mixed-valent {Cu2S}2+ motif upon solvation in acetonitrile: Effect on nitrous oxide reductase (N2Or) activity.
Chemistry - A European Journal, 2018, 24(20): 5060-5063

Esmieu C, Orio M, Le Pape L, Lebrun C, Pécaut J, Ménage S and Torelli S
Redox-innocent metal-assisted cleavage of S-S bond in a disulfide-containing ligand.
Inorganic Chemistry, 2016, 55(12): 6208-6217

Mangue J, Dubreucq Q, Pécaut J, Ménage S and Torelli S
Unexpected migration and O to S benzylic shift of thiocarbamate-containing salicylaldehyde derivatives.
ChemistrySelect, 2016, 1(20): 6345-6348

Dufour F, Pigeot-Remy S, Durupthy O, Cassaignon S, Ruaux V, Torelli S, Mariey L, Mauge F and Chaneac C
Morphological control of TiO2 anatase nanoparticles: What is the good surface property to obtain efficient photocatalysts?
Applied Catalysis B-Environmental, 2015, 174: 350-360

Iali W, Lanoe PH, Torelli S, Jouvenot D, Loiseau F, Lebrun C, Hamelin O and Ménage S
A Ruthenium(II)-Copper(II) dyad for the photocatalytic oxygenation of organic substrates mediated by dioxygen activation.
Angewandte Chemie - International Edition, 2015, 54(29): 8415-8419

Esmieu C, Orio M, Torelli S, Le Pape L, Pécaut J, Lebrun C and Ménage S
2O reduction at a dissymmetric {Cu2S}-containing mixed-valent center.
Chemical Science, 2014, 5: 4774-4784

Martin F, Torelli S, Le Paslier D, Barbance A, Martin-Laurent F, Bru D, Geremia R, Blake G and Jouanneau Y
Betaproteobacteria dominance and diversity shifts in the bacterial community of a PAH-contaminated soil exposed to phenanthrene.
Environmental Pollution, 2012, 162: 345-353

Torelli S, Orio M, Pécaut J, Le Pape L, Jamet H and Ménage S
A {Cu2S}2+ mixed-valent core featuring a Cu-Cu bond.
Angewandte Chemie International Edition, 2010, 49(44): 8249-8252


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A {Cu2S}2+ mixed-valent core featuring a Cu--Cu bond.
Angewandte Chemie International Edition, 2010, 49(44): 8249-8252