The Generation of the Oxidant Agent of a Mononuclear Nonheme Fe(II) Biomimetic Complex by Oxidative Decarboxylation. A DFT Investigation

The oxidative decarboxylation of the iron(II) α-hydroxy acid (mandelic acid) complex model, biomimetic of Rieske dioxygenase, has been investigated at the density functional level. The explored mechanism sheds light on the role of the α-hydroxyl group on the dioxygen activation. The potential energy surfaces have been explored in different electronic spin states. The rate-determining step of the process is the proton transfer. The oxidative decarboxylation preferentially takes place on the quintet state.


Introduction
In recent years, the employment of biomimetic catalysts considerably increased not only for their related high selectivity, but because they are allowed to operate in very mild conditions and they adapt to different types of substrates [1,2].
Encouraged by the new reactions catalyzed by Rieske dioxygenases [1,3,4], sustainable mononuclear nonheme iron complexes have been proposed for bioinspired oxidation catalysis. Dioxygenases are involved in the degradation of a wide range of natural and synthetic compounds by incorporating both atoms of O 2 into substrates. Bioinspired catalysis using peroxides or peracids for the oxidation of hydrocarbon substrates is more common and better characterized, than that based on dioxygen in catalytic oxidations, which has not been fully explored yet and the relative reaction mechanism is often unknown. This last reaction path requires both electron and proton sources [5][6][7][8][9][10][11][12][13] and cosubstrate-assisted O 2 molecular activation by transition metal containing complexes can be an alternative pathway [14][15][16][17][18][19] resorting to the sacrificial reductants represented a good solution [12,20]. Paine's and other groups have investigated a family of iron complexes supported by the monoanionic facial trinitrogen donor ligand, hydrotris(3,5-diphenylpyrazole-1-yl)borate (Tp Ph2 ligand) that activates O 2 by 2 electrons reduction in the oxidative decarboxylation of the α-hydroxy acids as sacrificial reductants [3,[21][22][23]. These studies provide evidence that the the hydroxyl group in the dioxygen activation is important. The iron complexes mentioned above reacting with O 2 are able to carry out the stoichiometric cis-dihydroxylation of olefins producing the diol product [3], as in that with H 2 O 2 [4,24,25]. The mononuclear non-heme iron(II) complex shown in Scheme 1, the object of the present investigation, represents the first biomimetic complex reproducing the dioxygenase activity of α-ketoglutarate-dependent enzymes. Since, as occurred in similar systems [26] no intermediate was detected in the course of the reaction, the steps up to the oxidative decarboxylation of the keto acid play a key role in the control of the rate of the reaction.
To provide deeper insights on the reaction catalyzed by the biomimetic complex of Scheme 1, we have undertaken a theoretical investigation taking into account the spin state of the analyzed species and building up the potential energy surfaces for the reaction path leading to the formation of an iron−oxygen oxidant upon oxidative decarboxylation of α-hydroxy acid.

Results and Discussion
As a first step of work, we have optimized the naked biomimetic iron(II) α-hydroxy acid complex (represented by model a in Figure S1 of the Supplementary Information) in septet, quintet, triplet and singlet spin states.
The coordination bond distances, which we have obtained together with X-ray counterpart [3] and relative electronic energies are collected in Table 1. Results show that the complex assumes the most stable configuration in the high-spin quintet state in well agreement with the available data for similar systems [3,27,28]. The singlet, triplet and septet spin states lie at 17.2, 13.1, and 65.1 kcal/mol above ground one, respectively. Table 1. Main geometrical parameters for naked complex a at all the spin states. Relative energies provided at M06L level of theory are kcal/mol and compared with the experimental X-ray counterpart [3]. Distances are in Å. See Scheme 1 for labels. Since, as occurred in similar systems [26] no intermediate was detected in the course of the reaction, the steps up to the oxidative decarboxylation of the keto acid play a key role in the control of the rate of the reaction.
To provide deeper insights on the reaction catalyzed by the biomimetic complex of Scheme 1, we have undertaken a theoretical investigation taking into account the spin state of the analyzed species and building up the potential energy surfaces for the reaction path leading to the formation of an iron−oxygen oxidant upon oxidative decarboxylation of α-hydroxy acid.

Results and Discussion
As a first step of work, we have optimized the naked biomimetic iron(II) α-hydroxy acid complex (represented by model a in Figure S1 of the Supplementary Information) in septet, quintet, triplet and singlet spin states.
The coordination bond distances, which we have obtained together with X-ray counterpart [3] and relative electronic energies are collected in Table 1. Results show that the complex assumes the most stable configuration in the high-spin quintet state in well agreement with the available data for similar systems [3,27,28]. The singlet, triplet and septet spin states lie at 17.2, 13.1, and 65.1 kcal/mol above ground one, respectively. Table 1. Main geometrical parameters for naked complex a at all the spin states. Relative energies provided at M06L level of theory are kcal/mol and compared with the experimental X-ray counterpart [3]. Distances are in Å. See Scheme 1 for labels.
The maximum deviation results to be less than 0.06 Å. From the superposition of the optimized structures at the considered spin states, it emerges that the mandelate ring shows major deviations whereas the coordination around the iron does not suffer drastic changes ( Figure 1). This suggests that in the quintet, the carboxylic group of the mandelic acid acts as monodentate ligand.
Molecules 2020, 25, x FOR PEER REVIEW 3 of 10 The maximum deviation results to be less than 0.06 Å. From the superposition of the optimized structures at the considered spin states, it emerges that the mandelate ring shows major deviations whereas the coordination around the iron does not suffer drastic changes ( Figure 1). This suggests that in the quintet, the carboxylic group of the mandelic acid acts as monodentate ligand. For the O2 addition (a-O2 complex), two different binding modes, namely end-on (eo) and side-on (so), have been taken into account. From computations, the eo binding mode appears to be the preferred one in all the considered spin states. Looking at the Figure 2 collecting the optimized structures of the eo obtained adducts, the triplet a-O2 complex represents the lowest energy configuration followed by the septet, quintet and singlet. A support to this finding arises from the analysis of the triplet spin state HOMO frontier orbital (see Figure 3) that mainly involves the metal center and its surrounding with the mandelate ligand less interested in the occurring electron redistribution.
As concerns the geometrical structure, the entry of dioxygen induces a reorganization around the metal center that now appears to be hexacoordinated (octahedral geometry). The O2 occupies the apical position, while the carboxylate moiety (lying on the equatorial plane) acts as bidentate ligand with both the Fe-O bond lengths of 2.17 Å. These values well compare with those present in dioxygenase enzymes [20,26].
The Fe-Oa bond assumes the value of 2.08, 1.93, and 2.21 Å in the triplet, quintet and septet electronic states, respectively. The O-O bond distance in the ground state (1.24 Å) reveals a significant deviation from the corresponding value in the enzymatic environment (1.31 Å). This behavior can be ascribable to the weaker electro-donating character of Tp Ph2 ligand towards the iron(III), present in the a-O2 complex, relative to the usual biological ligands in the enzymes [26].
The considered reaction mechanism (Scheme 2) arises from experimental indications. In fact, biomimetic and enzymatic studies suggested that the iron(III) superoxide species, obtained upon addition of O2 at the iron(II) center, has the ability to abstract hydrogen atom from the O-H group of α-hydroxy acid.
Since the singlet electronic spin state of iron-dioxygen complexes lies at very high energy, only the septet, quintet and triplet potential energy surfaces (PESs) have been considered (see Figure 4). For the O 2 addition (a-O 2 complex), two different binding modes, namely end-on (eo) and side-on (so), have been taken into account. From computations, the eo binding mode appears to be the preferred one in all the considered spin states. Looking at the Figure 2 collecting the optimized structures of the eo obtained adducts, the triplet a-O 2 complex represents the lowest energy configuration followed by the septet, quintet and singlet. A support to this finding arises from the analysis of the triplet spin state HOMO frontier orbital (see Figure 3) that mainly involves the metal center and its surrounding with the mandelate ligand less interested in the occurring electron redistribution.
As concerns the geometrical structure, the entry of dioxygen induces a reorganization around the metal center that now appears to be hexacoordinated (octahedral geometry). The O 2 occupies the apical position, while the carboxylate moiety (lying on the equatorial plane) acts as bidentate ligand with both the Fe-O bond lengths of 2.17 Å. These values well compare with those present in dioxygenase enzymes [20,26].
The Fe-O a bond assumes the value of 2.08, 1.93, and 2.21 Å in the triplet, quintet and septet electronic states, respectively. The O-O bond distance in the ground state (1.24 Å) reveals a significant deviation from the corresponding value in the enzymatic environment (1.31 Å). This behavior can be ascribable to the weaker electro-donating character of Tp Ph2 ligand towards the iron(III), present in the a-O 2 complex, relative to the usual biological ligands in the enzymes [26].
The considered reaction mechanism (Scheme 2) arises from experimental indications. In fact, biomimetic and enzymatic studies suggested that the iron(III) superoxide species, obtained upon addition of O 2 at the iron(II) center, has the ability to abstract hydrogen atom from the O-H group of α-hydroxy acid.       As far as the PES for the septet is concerned, no transition state connecting the hydroperoxide with the final oxidant agent (TS2 of Scheme 2) has been intercepted along the surface. A similar finding has been previously obtained for two enzymatic systems [26,29]. Instead, on the quintet and triplet PESs, both TS1 and TS2 have been characterized confirming that the oxidative decarboxylation is a multistep process.
The addition of O2 molecule starting to the complex (a) leads to the formation of adducts that in all the three considered spin states lie below the reactants' asymptote suggesting as their formation is an exergonic process (see Figure 4).

Scheme 2.
Proposed mechanism for the oxidative decarboxylation promoted by the non-heme biomimetic complex model (a).
Since the singlet electronic spin state of iron-dioxygen complexes lies at very high energy, only the septet, quintet and triplet potential energy surfaces (PESs) have been considered (see Figure 4).  As far as the PES for the septet is concerned, no transition state connecting the hydroperoxide with the final oxidant agent (TS2 of Scheme 2) has been intercepted along the surface. A similar finding has been previously obtained for two enzymatic systems [26,29]. Instead, on the quintet and triplet PESs, both TS1 and TS2 have been characterized confirming that the oxidative decarboxylation is a multistep process.
The addition of O2 molecule starting to the complex (a) leads to the formation of adducts that in all the three considered spin states lie below the reactants' asymptote suggesting as their formation is an exergonic process (see Figure 4). As far as the PES for the septet is concerned, no transition state connecting the hydroperoxide with the final oxidant agent (TS2 of Scheme 2) has been intercepted along the surface. A similar finding has been previously obtained for two enzymatic systems [26,29]. Instead, on the quintet and triplet PESs, both TS1 and TS2 have been characterized confirming that the oxidative decarboxylation is a multistep process.
The addition of O 2 molecule starting to the complex (a) leads to the formation of adducts that in all the three considered spin states lie below the reactants' asymptote suggesting as their formation is an exergonic process (see Figure 4).
On the three computed potential energy surfaces, the first transition state (TS1) is located at 18.6 (triplet), 11.1 (quintet) and 13.4 (septet) kcal/mol and their corresponding optimized structures are shown in Figure 5. On the three computed potential energy surfaces, the first transition state (TS1) is located at 18.6 (triplet), 11.1 (quintet) and 13.4 (septet) kcal/mol and their corresponding optimized structures are shown in Figure 5. For the TS1 in the triplet state, the barrier is 7.5 kcal/mol higher than that of quintet and a spin crossing from these two PESs occurs. This is due to the fact that the resulting iron(III) hydroperoxo species prefers to be a quintet (9.1 kcal/mol), while the corresponding in the triplet state is 9.3 higher in energy. The TS1 describes the proton transfer from the α-OH group of the mandelate to the Ob atom of the molecular oxygen, as confirmed by the imaginary frequency (638.1i cm −1 ). This result enhances the role played by the α-hydroxy acid ligand in this process as proposed by Paine et al. [3].
The next formed species, containing the Fe(III)-OOH moiety, is found at 9.1 kcal/mol and 11.0 kcal/mol in the quintet and septet state, respectively. In contrast, the iron(III) hydroperoxo in the triplet state lies at higher energy. This finding reflects the relative geometrical differences (see Figure  6a) In fact, in the quintet state, the carboxylate moiety is monocoordinated to iron (O1-Fe: 1.96 Å, O2-Fe: 3.18 Å) and the C-C bond results to be more elongated (1.71 Å) with respect to the previous steps. The benzaldehyde is prone to be released but it still retained by an H bond between its carbonyl (2.07 Å) and the OOH moiety. From the iron(III) hydroperoxo, the reaction proceeds with the definitive C-C bond breaking (2.05 Å) via TS2 that is characterized by an imaginary frequency related to the C-C stretching mode (211.8i cm −1 ) as shown in Figure 4. In addition, this bond's elongation generates the loss of the H bond since the OH-O=C is now 3.35 Å.
The product in its quintet spin state, at 41.3 kcal/mol below the separated reactants, evidences as the decarboxylation is a highly exergonic process. In the product, the CO2 is already formed (O-C-O angle is 180°) and it lies at 2.67 Å from the metallic center engaged in a sandwich-like topology between two phenyl rings (ligand and benzaldehyde) (see Figure 6b). Some changes occur also in the metallic core due to the elongation of Oa-Ob bond (1.48 Å) and to the Fe-Oa distance of 1.91 Å. In this way, the iron(II) hydroperoxo oxidant agent is formed and ready to act as nucleophilic agent in the next oxidation reactions that are not object of the present study. For the TS1 in the triplet state, the barrier is 7.5 kcal/mol higher than that of quintet and a spin crossing from these two PESs occurs. This is due to the fact that the resulting iron(III) hydroperoxo species prefers to be a quintet (9.1 kcal/mol), while the corresponding in the triplet state is 9.3 higher in energy. The TS1 describes the proton transfer from the α-OH group of the mandelate to the O b atom of the molecular oxygen, as confirmed by the imaginary frequency (638.1i cm −1 ). This result enhances the role played by the α-hydroxy acid ligand in this process as proposed by Paine et al. [3].
The next formed species, containing the Fe(III)-OOH moiety, is found at 9.1 kcal/mol and 11.0 kcal/mol in the quintet and septet state, respectively. In contrast, the iron(III) hydroperoxo in the triplet state lies at higher energy. This finding reflects the relative geometrical differences (see Figure 6a). In fact, in the quintet state, the carboxylate moiety is monocoordinated to iron (O 1 -Fe: 1.96 Å, O 2 -Fe: 3.18 Å) and the C-C bond results to be more elongated (1.71 Å) with respect to the previous steps. The benzaldehyde is prone to be released but it still retained by an H bond between its carbonyl (2.07 Å) and the OOH moiety. From the iron(III) hydroperoxo, the reaction proceeds with the definitive C-C bond breaking (2.05 Å) via TS2 that is characterized by an imaginary frequency related to the C-C stretching mode (211.8i cm −1 ) as shown in Figure 4. In addition, this bond's elongation generates the loss of the H bond since the OH-O=C is now 3.35 Å.
The product in its quintet spin state, at 41.3 kcal/mol below the separated reactants, evidences as the decarboxylation is a highly exergonic process. In the product, the CO 2 is already formed (O-C-O angle is 180 • ) and it lies at 2.67 Å from the metallic center engaged in a sandwich-like topology between two phenyl rings (ligand and benzaldehyde) (see Figure 6b). Some changes occur also in the metallic core due to the elongation of O a -O b bond (1.48 Å) and to the Fe-O a distance of 1.91 Å. In this way, the iron(II) hydroperoxo oxidant agent is formed and ready to act as nucleophilic agent in the next oxidation reactions that are not object of the present study.  [3]. To give a contribution to this aspect, geometry optimizations of the two forms have been performed and the resulting optimized geometries (shown in Figure S2) indicate that the quintet spin state proves to be the most stable one.

Conclusions
On the basis of our density functional theory investigation on the oxidative decarboxylation promoted by a biomimetic nonheme complex, the following conclusions can be drawn: 1. The favored reaction path occurs in the quintet spin state as in the corresponding Rieske oxygenase enzymes; 2. A spin cross takes place between the triplet and quintet electronic spin states after the adduct formation; 3. The α-hydroxy group plays an important role in the oxidative decarboxylation; 4. The rate determining step is the proton transfer; 5. The overall reaction results to be more exergonic in the quintet PES.

Computational Details
All density functional theory (DFT) based calculations have been performed by using Gaussian09 code [30]. Following previous suggestions available in literature about the reliable results of the M06L [31,32] exchange-correlation functional in describing electronic structure and  [3]. To give a contribution to this aspect, geometry optimizations of the two forms have been performed and the resulting optimized geometries (shown in Figure S2) indicate that the quintet spin state proves to be the most stable one.

Conclusions
On the basis of our density functional theory investigation on the oxidative decarboxylation promoted by a biomimetic nonheme complex, the following conclusions can be drawn:

1.
The favored reaction path occurs in the quintet spin state as in the corresponding Rieske oxygenase enzymes; 2.
A spin cross takes place between the triplet and quintet electronic spin states after the adduct formation; 3.
The α-hydroxy group plays an important role in the oxidative decarboxylation; 4.
The rate determining step is the proton transfer; 5.
The overall reaction results to be more exergonic in the quintet PES.

Computational Details
All density functional theory (DFT) based calculations have been performed by using Gaussian09 code [30]. Following previous suggestions available in literature about the reliable results of the M06L [31,32] exchange-correlation functional in describing electronic structure and properties of metal containing systems also in various spin states [33][34][35][36], this functional has been adopted for the geometry optimizations.
The crystal structure (X ray) of the biomimetic catalyst [3] has been adopted as starting structure for our computations. In order to reduce the computational time, five phenyl rings of the Tp Ph2 ligand have been replaced by methyl groups (see a of Figure S1). The phenyl group of the Tp Ph2 ligand closely oriented to the iron center has been retained to better evaluate its role on the activated dioxygen species during the catalytic process.
Geometry optimizations and vibrational frequencies have been performed employing differentiated basis set. In particular, for O, N and H atoms in the active center the 6-31+G(d,p) set has been used while for the remaining ones the 6-31G(d,p) one has been chosen. The LANL2DZ effective core potential and the related basis set has been employed for iron.
In order to improve the energetic values, single point calculations with the larger basis set 6-311+G(d,p) for all atoms except for metal ion, has been performed on all the previously optimized structures. The solvent corrections were evaluated using the polarizable continuum model [37,38] as implemented in Gaussian09 taking into account the dielectric constant (ε = 24.93) that better reproduces the polarity of the solvents mixture used in the experimental work [3].
Owing to the unrestricted formalism of all calculations, the spin contamination and its effect on the calculated energetics was monitored as done for similar systems [35,39] and the <S2> values are collected in Supplementary Information section (Table S1).