Modeling the cis-Oxo-Labile binding site motif of non-heme iron oxygenases: Water exchange and oxidation reactivity of a non-heme iron(IV)-Oxo compound bearing a tripodal tetradentate ligand

Article abstract of DOI:10.1002/chem.201002297

The spectroscopic and chemical characterization of a new synthetic non-heme iron(IV)-oxo species [Fe^IV(O)(^Me,HPytacn)(S)] ²⁺ (2, ^Me,HPytacn=1-(2′-pyridylmethyl)-4,7-dimethyl- 1,4,7-triazacyclononane, S=CH₃CN or H₂O) is described. Complex 2 was prepared by reaction of [Fe^II(CF₃SO ₃)₂(^Me,HPytacn)] (1) with peracetic acid. Complex 2 bears a tetradentate N₄ ligand that leaves two cis sites available for binding an oxo group and a second external ligand but, unlike the related iron(IV)-oxo species with tetradentate ligands, it is remarkably stable at room temperature (t_1/2>2a h at 288a K). Its ability to exchange the oxygen atom of the oxo ligand with water has been analyzed in detail by means of kinetic studies, and a mechanism is proposed on the basis of DFT calculations. Hydrogen-atom abstraction from C-H bonds and oxygen-atom transfer to sulfides by 2 have also been studied. Despite its thermal stability, 2 proves to be a very powerful oxidant that is capable of breaking the strong C-H bond of cyclohexane (bond dissociation energy=99.3a kcala mol^-1). A powerful oxidant: A non-heme oxo-iron(IV) complex (see picture), containing a labile site cis to the terminal oxo group, has been prepared and characterized. The complex is unusually stable (t_1/2>2a h at 288a K). Water exchange, oxo transfer, and Ha abstraction reactions have been studied. Despite its thermal stability, the complex is a very powerful oxidant.

Full text of DOI:10.1002/chem.201002297

DOI: 10.1002/chem.201002297
Modeling the cis-Oxo-Labile Binding Site Motif of Non-Heme Iron
Oxygenases: Water Exchange and Oxidation Reactivity of a Non-Heme
Iron(IV)-Oxo Compound Bearing a Tripodal Tetradentate Ligand
[
a, e]
[a]
[b]
[b]
Anna Company,
Irene Prat, Jonathan R. Frisch, Dr Ruben Mas-Ballestꢀ,
[d] [a] [d]
[
c]
[c]
Mireia Gꢁell, Gergely Juhꢂsz, Xavi Ribas, Dr Eckard Mꢁnck,* Josep M. Luis,*
[
b]
[a]
Lawrence Que, Jr.,* and Miquel Costas*
Abstract: The spectroscopic and chemi-
cal characterization of a new synthetic
a second external ligand but, unlike the
related iron(IV)-oxo species with tetra-
dentate ligands, it is remarkably stable
at room temperature (t1/2 >2 h at
288 K). Its ability to exchange the
oxygen atom of the oxo ligand with
water has been analyzed in detail by
means of kinetic studies, and a mecha-
nism is proposed on the basis of DFT
calculations. Hydrogen-atom abstrac-
tion from CꢀH bonds and oxygen-atom
non-heme
iron(IV)-oxo
Pytacn)(S)]
=1-(2’-pyridylmethyl)-4,7-di-
species
IV
Me,H
2+
[
Fe (O)(
(2,
Me,H
Pytacn
methyl-1,4,7-triazacyclononane,
CH CN or H O) is described. Complex
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
transfer to sulfides by 2 have also been
studied. Despite its thermal stability, 2
proves to be a very powerful oxidant
that is capable of breaking the strong
CꢀH bond of cyclohexane (bond disso-
S=
3
2
II
2
was prepared by reaction of [Fe -
Me,H
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(CF SO ) (
Pytacn)] (1) with perace-
3
3 2
Keywords: bioinorganic chemistry ·
iron · oxygenases · reaction mecha-
nisms · tetradentate ligands
ꢀ
1
tic acid. Complex 2 bears a tetraden-
ciation energy=99.3 kcalmol ).
tate N ligand that leaves two cis sites
4
available for binding an oxo group and
Introduction
[
a] Dr. A. Company, I. Prat, Dr. X. Ribas, Dr. M. Costas
Departament de Quꢀmica
Universitat de Girona
Iron(IV)-oxo compounds are proposed as the active species
Campus Montilivi, E17071 Girona, Catalonia (Spain)
Fax : (+34)972-41-81-50
E-mail: miquel.costas@udg.edu
in the catalytic cycles of several O -activating non-heme
2
iron enzymes, such as isopenicillin N-synthase, pterin-depen-
dent hydroxylases, and 2-oxoglutarate-dependent oxygenas-
[
b] J. R. Frisch, D. R. Mas-Ballestꢁ, Prof. Dr. L. Que, Jr.
Department of Chemistry and Center for Metals in Biocatalysis
University of Minnesota
[1–3]
es.
Quite recently, high-spin (S=2) iron(IV)-oxo inter-
mediates have been trapped and spectroscopically character-
Minneapolis, MN 55455 (USA)
Fax : (+1)612-625-0389
E-mail: larryque@umn.edu
ized in five different non-heme iron enzymes: namely, tauri-
[4,5]
ne:a-ketoglutarate dioxygenase (TauD),
halogenase
[6]
[7]
[8]
CytC3, propyl-4-hydroxylase, tyrosine hydroxylase, and
[
c] Dr. M. Gꢂell, Dr. J. M. Luis
Institut de Quꢀmica Computacional
Universitat de Girona
Campus Montilivi, E17071 Girona, Catalonia (Spain)
Fax : (+34)972-41-83-56
[9]
aliphatic halogenase, SyrB2. Parallel synthetic efforts have
IV
succeeded in the preparation of octahedral S=2 [Fe (O)-
2
+ [10]
IV
A
H
U
G
R
N
U
G
and octahedral S=1 Fe =O species using tet-
2
5
[11,12]
radentate N ligands with available trans-binding sites,
4
E-mail: josepm.luis@udg.edu
[13–17]
tetradentate N ligands with available cis-binding sites,
4
[
d] Dr. G. Juhꢃsz, Prof. D. E. Mꢂnck
Department of Chemistry
[18–22]
pentadentate N5 ligands,
and a pentadentate N4S
[23]
ligand. Very recently, two examples of trigonal bipyrami-
dal S=2 iron(IV)-oxo complexes, which bear structural and
electronic resemblance to the enzymatic non-heme inter-
Carnegie Mellon University
4
400 Fifth Avenue, Pittsburgh, PA 15213 (USA)
Fax : (+1)412-268-5058
E-mail: em40@andrew.cmu.edu
[24–26]
mediates, have been described.
[
e] Dr. A. Company
Present address: Institut fꢂr Chemie, Technische Universitꢄt
Berlin (Germany)
These metastable and highly oxidizing complexes have
been characterized by various spectroscopic techniques in-
[
11,22,25–27]
cluding, in selected cases, X-ray crystallography.
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/chem201002297.
IV
The ability of synthetic non-heme Fe (O) species to act as
1622
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Chem. Eur. J. 2011, 17, 1622 – 1634

FULL PAPER
[28,29]
oxidants has been proved
and several studies demon-
strate that they are capable of performing several oxidation
reactions including: oxygen-atom transfer to sulfides and tri-
[
16,30,31]
[13,19,21]
phenylphosphine,
epoxidation
and possible di-
[
21,32]
[33]
hydroxylation
gen-atom abstraction,
of alkenes, alcohol oxidation,
hydro-
[18,34,35]
aliphatic and aromatic hydrox-
[36,37]
II
[38]
ylation, oxo transfer to other Fe complexes, and glu-
tathione and peptide oxidation.
analogy to the better studied heme systems, it is widely
postulated that non-heme iron-oxo species can exchange
their oxygen atom with water through an oxo–hydroxo tau-
tomerism before attack to a substrate (Scheme 1). Thus, in-
[39,40]
On the other hand, by
[41]
IV
Scheme 2. Schematic representation of synthetic Fe (O) complexes rele-
vant to this work. bpmcn= N,N’-bis(2-pyridylmethyl)-N,-N’-dimethyl-
trans-1,2-diaminocyclohexane, tpa=tris(2-pyridylmethyl)amine , bpd=di-
methyl 2,4-bis(2-pyridinyl)-3,7-diazabicyclo-[3.3.1]-nonane-9-on-1,5-dicar-
boxylate, Bn-tpen=N-benzyl-N,N’,N-tris(2-pyridylmethyl)-1,2,diamino-
ethane , Pytmc= 1-(2’-pyridylmethyl)-4,8,11-Tetramethyl-1,4,8,11-tetra-
aza-cyclotetradecane.
Scheme 1. Top: Proposed oxo–hydroxo tautomerism in heme systems.
IV
Bottom: Proposed oxo–hydroxo tautomerism in [Fe (O) AHCUTNGRNENUG( tmc)-
2
+
IV
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(CH
(N4Py)]
amine.
3
CN)]
(tmc=1,4,8,11-tetraaza-cyclotetradecane) and [Fe (O)-
(N4Py=N,N-bis(2-pyridylmethyl)-N-bis(2-pyridyl)methyl-
2
+
ACHTUNGTRENNUNG
hydrogen-atom abstraction reactions. Moreover, the ex-
1
8
direct evidence for the participation of these compounds in
the catalytic cycle of enzymes and/or synthetic models
change of its oxo group with the oxygen atom from H2 O
[42]
[43]
has been studied by means of kinetic and computational
methods.
comes from the incorporation of oxygen from water into
oxidized products. Recently, this has been experimentally
proved in two synthetic non-heme iron(IV)-oxo species;
IV
2+
IV
well-defined [Fe (O)
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
and [Fe (O)-
Results and Discussion
3
2
+
A( N4Py)]
C
H
T
U
N
G
T
R
E
N
N
U
N
G
IV
change with water (Scheme 1).
Preparation and characterization of the Fe (O) species: Re-
II
Me,H
[45]
Because the vast majority of non-heme iron oxygenases
contain cis-labile binding sites, synthetic iron(IV)-oxo com-
plexes with tripodal tetradentate ligands are especially rele-
vant from a biological point of view. However, they have
proved to be less stable than their pentadentate counter-
action of [Fe
A
H
U
G
R
N
U
G
Pytacn)] (1; Scheme 3)
with
3
3 2
two equivalents of peracetic acid (CH CO H) in CH CN at
158C produces, within 20 min, a novel species (2) with a
UV/Vis spectrum showing a band at l =750 nm (e=
200m cm ) and a weaker shoulder around 900 nm (e
3
3
3
max
ꢀ
1
ꢀ1
[28]
ꢀ1
ꢀ1
parts,
thus rendering their study more difficult. For this
ꢁ100m cm ; Figure 1). Complex 2 is relatively stable in
reason most of the studies have been performed by using
solution, with a half-life time (t ) of 2.4 h at 158C. The
1
/2
pentadentate N ligands (N4Py) or equatorial tetradentate
N4 ligands with two labile sites in a trans configuration
nature of the transient species 2 has been determined by a
combination of Mçssbauer, H NMR, and resonance Raman
5
1
(
tmc). Herein, we address this issue and we describe the
spectroscopy, ESI-MS, and DFT calculations. We studied the
Mçssbauer spectra of 2 in acetonitrile between 4.2 and
100 K in parallel applied magnetic fields up to 8.0 T. Two
representative spectra are shown in Figure 2. In zero applied
preparation and spectroscopic and chemical characterization
of a new Fe (O) species bearing the tripodal tetradentate
1
IV
-(2’-pyridylmethyl)-4,7-dimethyl-1,4,7-triazacyclononane
Me,H
ligand (
Pytacn). This species
shows an unprecedented high
stability at room temperature,
which allows us to evaluate sev-
eral biologically relevant as-
pects of its reactivity. These
studies include the evaluation
of its ability to perform oxygen-
atom transfer to sulfides and Scheme 3. Schematic view of formation and decay of 2, along with the proposed chemical structures of 1–3.
Chem. Eur. J. 2011, 17, 1622 – 1634
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
1623

E. Mꢂnck, J. M. Luis, L. Que, Jr., M. Costas et al.
The spectrum of Figure 2A indicates the presence of an S=
ꢀ
1
0
0
diferric contaminant with DE =1.60 mms
and d=
Q
ꢀ1
.45 mms , which represents approximately 15% of Fe. For
the present chemistry such complexes are decomposition
products that frequently represent the thermodynamic
[13]
sink.
High-resolution ESI-MS establishes the elemental compo-
sition of 2 (Figure 1, bottom) as it reveals a clean spectrum
with a major peak with a mass value at m/z 469.08 and an
IV
isotopic pattern fully consistent with the ion {[Fe (O)-
Me,H
+
A
H
U
G
R
N
N
(CF SO )(
Pytacn)]} . Evidence for the presence of the
3
3
oxo ligand was provided by vibrational spectroscopy. Reso-
nance Raman analysis on frozen acetonitrile solutions of 2
with laser excitation at lexc =407.9 nm shows a resonance-
ꢀ
1
ꢀ1
enhanced feature at 831 cm that downshifts to 788 cm
1
8
when 2 is allowed to react with H2 O. The energy of the
band falls in the range of reported synthetic
enzymatic non-heme Fe=O stretching vibrations, and the
43 cm shift is in reasonable agreement with the shift of
[11,15,22,24,46]
and
[47]
ꢀ
1
ꢀ
ꢀ1
ꢀ
37 cm predicted by Hookeꢆs law for a diatomic Fe–O
ꢀ
1
[48,49]
stretch and ꢀ40 cm predicted by DFT calculations.
In
Figure 1. UV/Vis spectrum of
bottom).
2 (top) and its ESI mass spectrum
conclusion, the spectroscopic data allow us to formulate 2 as
(
IV
Me,H
2+
a
new iron(IV)-oxo species [Fe (O)(
Pytacn)(S)] ,
where solvent S=CH CN or H O. The nature of the sixth
3
2
ligand could not be experimentally determined, but insight
into its nature can be obtained from the DFT calculations
described in the next section. Acetonitrile binding was pre-
viously documented in the crystallographic characterization
IV
2+ [11]
of [Fe (O)
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
.
However, in the present
3
case, the presence of a water ligand is proposed on the basis
of DFT calculations (see below), which indicated that these
species are thermodynamically more stable than the corre-
[13,14]
sponding analogue containing an acetonitrile ligand.
Because of the inequivalence of the two binding positions
Me,H
not occupied by
Pytacn, two isomeric forms are possible
for 2 (2a and 2b, Scheme 3). These two isomers differ not
only in the group disposed trans to the oxo ligand but also
in the relative orientation of the pyridine ring with respect
to the Fe–O axis. However, the H NMR spectrum of 2
(Figure 3) showed a unique set of sharp signals that could
Figure 2. Mçssbauer spectrum of 2 in acetonitrile, recorded at 4.2 K in a
parallel field as indicated. Solid lines are spectral simulations based on
1
[
23]
an S=1 spin Hamiltonian with the zero-field splitting parameters D=
ꢀ
1
57
+
27 cm , E/D=0, Fe magnetic hyperfine tensor Ax,y,z/g
n
b=(ꢀ23.7,
ꢀ
1
ꢀ
20.5, ꢀ4.5) T, quadrupole parameters DE
Q
=0.73 mms , h=1, and
be assigned to the pyridine b (42 ppm), g (10 ppm), and b’
ꢀ1
isomer shift d=0.05 mms
.
(
ꢀ13 ppm) protons. The shift pattern for these signals is
1
reminiscent of that found in the H NMR spectrum of
magnetic field (Figure 2A), 2 exhibits at 4.2 K a quadrupole
doublet, which represents 85(5)% of the total iron, with
ꢀ
1
quadrupole splitting DE =0.73 mms and isomer shift d=
0
Q
ꢀ1
.05 mms . These parameters, together with the fact that 2
has integer electron spin (as witnessed by the observation
that 2 displays a doublet rather than magnetically split spec-
IV
tra in zero field), show that 2 is an Fe complex. Spectra re-
corded in applied magnetic fields (see, e.g., Figure 2B) have
IV
features very similar to those reported for S=1 Fe (O)
IV
complexes; S=2 Fe (O) complexes display quite different
[6,8,10,24]
spectra.
monly used S=1 spin Hamiltonian;
We analyzed the spectra of 2 with the com-
[23]
the parameters ob-
tained are very similar to those obtained for other S=1
Fe (O) complexes and are listed in the caption of Figure 2.
1
Figure 3. H NMR spectrum of 2 at 298 K in CD
3
CN, along with selected
IV
signal identifications.
1624
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ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2011, 17, 1622 – 1634

Modeling the Binding Site Motif of Non-Heme Iron Oxygenases
FULL PAPER
IV
2+
[
Fe (O)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(N4Py)] , in which all four pyridine rings are dis-
computed (Table S7 in the Supporting Information). The
substantially higher energies of these states, when compared
with the triplet (S=1) state, further support the Mçssbauer-
based spin-state assignment of 2. Structural parameters for
2a and 2b are very similar. The computed Fe–O distances
in 2a and 2b are 1.62 and 1.63 ꢇ, respectively, in good
agreement with structurally characterized examples of syn-
posed parallel to the Fe–O axis, and different from the pat-
tern associated with pyridine rings oriented perpendicular to
the Fe=O axis. Thus, in the present case, the NMR spec-
trum indicates that only isomer 2a is present in solution,
with the pyridine ring disposed parallel to the Fe–O axis.
Despite the inherently low stability of high-valent oxo-
[
27]
1
[11,17,22,24–27,52]
[53,54]
iron(IV) species, the H NMR spectrum of 2 lacks signals
thetic
and enzymatic
non-heme iron(IV)-
[50]
III
corresponding to 1 and 3 (the dinuclear Fe decomposi-
tion product, see below and the Supporting Information),
thus suggesting that 2 is prepared with good purity, in ac-
cordance with the Mçssbauer analysis (see above).
oxo species. The oxo ligand exerts a trans effect that is indi-
cated by a ꢁ0.1 ꢇ lengthening of the corresponding trans
Fe–N distance (compare d
=2.19 ꢇ in 2b with 2.06 ꢇ in
Fe1–N4
2a, and dFe1–N3 =1.99 ꢇ in 2b with 2.11 ꢇ in 2a). The Fe–Npy
distance is 2.03 ꢇ in 2a and 2.02 ꢇ in 2b, values that appear
DFT analysis of the structures of the two isomers of 2: DFT
somewhat intermediate between the short d
ꢁ1.95 ꢇ of
Fe–Npy
IV
calculations were employed to help clarify the molecular
the pyridine rings cis to the oxo ligand in [Fe (O) ACHTUNGTRENNUNG( N4Py)]-
[48,49]
[27]
IV
and electronic structure of 2.
The nature of the sixth
A
H
U
G
R
N
U
(ClO )
and the longer d
=2.118(3) ꢇ of [Fe (O)-
Fe–Npy
4
2
[22]
ligand that completes the octahedral coordination sphere of
the iron center was first studied by comparing the relative
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
for which the pyridine is bound trans
3
IV
Me,H
2+
energies of the corresponding [Fe (O)(
Pytacn)(S)]
(
S=CH CN and H O) complexes. Computational analysis
Relative
[Fe (O)(
thermal
stability
and
decay
of
2:
3
2
IV
Me,H
2+
for S=H O indicates that hydrogen bonding between the
Pytacn)(S)] (2) joins the growing family of
2
water ligand and a second, external water molecule is very
iron(IV)-oxo species formed with tetradentate ligands
[11]
[16]
strong because of the dicationic nature of 2. This interaction
tmc, tpa, N,N’, dimethyl-N,N’-bis(8-quinolyl)ethane-1,2-
IV
Me,H
2+
[17]
[35]
[14]
stabilizes
[Fe (O)(
Pytacn)
A
H
U
G
R
N
U
G
relative
to
diamine (bqen), bpd, and bpmcn (Scheme 2). In the
case of tmc, the ligand topology gives rise to an iron center
with two available trans sites, one of which is occupied by
2
IV
Me,H
2+
ꢀ1
[Fe (O)(
Pytacn)
A
H
U
G
R
N
U
G
by DG=16.1 kcalmol .
3
Acetonitrile binding has been previously demonstrated in
IV
2+ [11]
IV
the X-ray structure of [Fe (O)
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
,
for
an oxo ligand in the Fe -oxo compound. Instead, ligands
3
Me,H
which the hydrophobic pocket created by the N-Me groups
of tmc isolates the acetonitrile ligand. This is likely to pre-
such as
Pytacn, tpa, bpd, bqen, and bpmcn wrap around
the metal ion, so that two sites in a cis relative disposition
are ready for interaction with exogenous ligands. Thus, in
the corresponding iron(IV)-oxo compounds, the oxo group
binds to one of these positions and the other site is presuma-
bly occupied by a neutral labile molecule, such as acetoni-
trile or water (Scheme 2). Despite the similar coordination
spheres around the metal center, the resulting iron(IV)-oxo
compounds exhibit remarkably different stabilities.
vent intermolecular interactions with additional water mole-
Me,H
cules. In contrast, the tripodal tetradentate
Pytacn ligand
lacks such a sterically restricted hydrophobic pocket, which
may constitute the key structural feature that favors water
binding over acetonitrile in 2.
IV
Me,H
Optimized molecular structures of [Fe (O)(
Pytacn)-
2
+
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(H O)] (2a and 2b) obtained by DFT calculations are
2
IV
2+
IV
2+
IV
shown in Figure 4, with selected structural parameters given
[Fe (O) ACHTUNTGRENNUN(G bpmcn)(S)] , [Fe (O) AHCTUNGTRENNUNG
[51]
2+
in the caption. The relative DFT energies of the two iso-
ACHTUNGTRENNUNG
3
2
meric forms of 2 indicate that 2a is energetically favored by
good yields at ꢀ408C because they easily decompose at
ꢀ
1
[13,14,35] IV 2+
2.1 kcalmol , thus supporting our structural assignment
higher temperatures.
[Fe (O) ACHTUNGTERNNNU(G bqen)(S)] has a life-
[17]
based on NMR spectroscopy. Energies corresponding to sin-
glet (S=0) and quintet states (S=2) of 2a and 2b were also
time of t1/2 ꢁ30 min at 08C. In contrast, 2 can be generat-
ed and remains relatively stable at 158C.
Thermal decay of 2 leads to a new species 3 that has been
III
Me,H
3+
identified as [Fe
A
H
U
G
E
N
N
(m-O)
A
T
N
T
E
N
G
Pytacn)2]
by
2
3
2
means of UV/Vis spectroscopy, ESI-MS, and comparison
with an independently prepared sample characterized by X-
[55]
ray crystallography (Figure 5). The relatively poor stabili-
ty observed for iron(IV)-oxo species with tripodal tetraden-
tate ligands, in comparison with those that contain penta-
dentate or tetradentate ligands with trans topologies, can be
understood by considering that only the latter two offer ef-
fective steric protection of the oxo ligand against decompo-
sition into a diiron complex. In addition, these “stable”
iron(IV) compounds lack a cis-labile site adjacent to the re-
active oxo ligand, thus precluding prebinding of a putative
substrate. Based on these arguments, we suggest that the
surprising thermal stability of 2 derives from a combination
Figure 4. DFT-calculated structures of 2a (left) and 2b (right). Selected
bond lengths [ꢇ] for 2a: Fe1ꢀN2 2.05, Fe1ꢀN3 2.11, Fe1ꢀN4 2.06, Fe1ꢀ
O5 2.06, Fe1
ꢀ
O6 1.62, Fe1ꢀN7 2.03 and 2b: Fe1ꢀN2 2.07, Fe1ꢀN3 1.99,
Fe1ꢀN4 2.19, Fe1ꢀO5 1.63, Fe1ꢀO6 2.03, Fe1ꢀN7 2.02.
Chem. Eur. J. 2011, 17, 1622 – 1634
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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1625

E. Mꢂnck, J. M. Luis, L. Que, Jr., M. Costas et al.
with thioanisole at different reaction times and analyzing by
GC-MS the percentage of labeled sulfoxide generated.
Within experimental error, the two methods afforded identi-
cal reaction rates, thus supporting the validity of the second
approach. In any case, exchange rate constants were ob-
tained by nonlinear regression fitting of the data to a single
exponential function.
The rate of water incorporation into 2 (kobs) was found to
1
8
be linearly dependent on [H2 O] and independent of [2].
The data are consistent with the rate law in Equation (1), in
1
8
which the use of a relatively large [H2 O] allows for its
analysis as a pseudo-first-order reaction [Eq. (1)]:
Â
Ã
IV
18
d Fe
¼
O
Â
ÃÂ
Ã
Â
Ã
IV
16
18
2
IV
16
¼
kexc Fe
¼
O H O ¼ k Fe
¼
O
obs
III
dt
Figure 5. ORTEP plot (50% probability ellipsoids) of [Fe
2
A
H
U
G
E
N
N
ACHTUNGERTNNUNG( m-
Me,H
CH
3
CO
2
)(
Pytacn)
2
]
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(ClO
4
)
3
·CH
3
CN (3·CH
3
ð1Þ
[
ꢇ] and angles [8]: Fe1ꢀO1 1.801(3), Fe1ꢀO3 2.062(4), Fe1ꢀN1 2.167(4),
Fe1ꢀN3 2.180(4), Fe1ꢀN4 2.197(4), Fe1ꢀN2 2.225(4), O1ꢀFe2 1.805(3),
Fe2ꢀO2 2.017(3), Fe2ꢀN5 2.163(4), Fe2ꢀN8 2.184(4), Fe2ꢀN7 2.196(4),
Fe2ꢀN6 2.247(4); O1-Fe1-O3 95.42(15), Fe2-O1-Fe1 135.2(2).
ꢀ
1
ꢀ1
A bimolecular rate constant of kexc =(0.028ꢂ0.003)m
s
at 295 K was extracted from this analysis. Activation param-
eters for water exchange were also determined by measuring
exchange rates between 275 and 305 K, which afforded
°
ꢀ1
°
DH =(10.2ꢂ0.8) kcalmol
and
DS =(ꢀ32ꢂ
Me,H
of the oxidatively robust nature of the
Pytacn ligand and
ꢀ1
ꢀ1
3
) calK mol (Figure 6). The negative activation entropy
some steric hindrance imposed by the N-Me groups of the
ligand that project into the space surrounding the Fe=O
unit. Consistent with this notion, in the absence of acetate
ion, which facilitates dimerization, the oxidation of 1 with
H O in an aqueous acetonitrile solvent mixture does not
2
2
result in the common rapid formation of oxo-bridged ferric
[56]
dimers, but instead in the formation of the mononuclear
III
Me,H
+
species formulated as {[Fe (OH)(
Pytacn)] ACHTUNTGNERNUG( CF SO )}
3 3
on the basis of its ESI mass spectra. The remarkable stabili-
ty of 2 has allowed us to study some aspects of its biological-
ly relevant reactivity, which are detailed below.
Oxygen-atom exchange with water: Because of the biologi-
Figure 6. Eyring plot for the determination of the activation parameters
of the oxygen-atom exchange of 2 with H O ([H O]=0.2m).
2 2
cal relevance of water exchange reactions at metal cen-
18
18
[
57]
18
ters, the ability of 2 to exchange its oxo ligand with H2
O
was studied by taking advantage of ESI-MS. Complex 2 was
generated by reaction of 1 with two equivalents of
CH CO H in acetonitrile. Subsequently, 600 equivalents of
is consistent with an associative event in the transition state,
most likely a bimolecular collision between the Fe (O) spe-
3
8
3
1
IV
H2 O were added to the reaction solution of 2, and several
ESI mass spectra were taken over time. The initial single
cies and a water molecule. Second-order rate constants for
water exchange at [Fe (O) AHCTUNGTRENNUN(G tpa)(S)] were also measured
(Table 1). The accumulated data indicate that the water ex-
IV 16
IV
2+
peak at m/z 469.1 associated with {[Fe
A
T
N
T
E
N
N
( O)-
Me,H
+
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(CF SO )(
Pytacn)]} decreased gradually, while a new
3
3
IV 18
peak at m/z 471.1 corresponding to {[Fe
A
H
U
G
R
N
N
change rate for 2 is substantially faster than those for
Me,H
+
IV
2+
IV
2+
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(CF SO )(
[Fe (O)
A
H
U
T
E
N
N
(tmc)
A
H
U
G
R
N
N
(CH CN)]
and [Fe (O) AHCTUNGTRENNUN(G N4Py)] , which
3
3
3
lack cis-available sites to the oxo ligand, and also somewhat
IV
2+
3
faster than that of [Fe (O) AHCTUNGTERUNNN(G tpa)(S)] . Nevertheless, consid-
strong s-donating ligand, inhibited the water exchange ex-
hibited by 2, which suggested that the available coordination
site in a cis configuration with respect to the oxo group
plays an important role in facilitating the water exchange
process.
ering the pentadentate nature of N4Py, the higher reaction
rate observed for 2 represents a rather modest enhance-
ment. Furthermore, the smaller reaction rate measured for
IV
2+
[Fe (O)
A
H
U
G
R
N
U
G
suggests that reaction rates are not
solely determined by the presence of a cis-labile site. Analy-
sis of the enthalpy and entropy activation parameters indi-
cates that the faster exchange rate in 2 has an entropic
origin, which compensates for its higher activation enthalpy.
Kinetic analyses of water exchange in 2 were performed
18
by monitoring O incorporation into 2 directly by ESI-MS,
1
8
and indirectly by quenching of a solution of 2 and H2
O
1626
www.chemeurj.org
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2011, 17, 1622 – 1634

Modeling the Binding Site Motif of Non-Heme Iron Oxygenases
FULL PAPER
[
50]
Table 1. Kinetic parameters for the water exchange reaction at selected
oxo-iron(IV) complexes.
sian 09. The dispersion energy has been computed using
[60]
the DFT-D3 method and program of Grimme et al. Ac-
cording to our theoretical calculations, a key aspect of the
reaction is the presence of an exogenous water molecule at
close distance to the oxo group and the water ligands. In the
first step the exogenous water assists the transfer of one H
ion from the bound H O to the oxo ligand, via a transition
state (TS1), with a barrier of DG =9.3 kcalmol . After the
first proton-transfer event, an Fe (OH) species (intermedi-
ꢀ
3
°
°
[58]
Compound
k
[m
exc ꢈ10
s
DH
[kcalmol
DS
Ref.
ꢀ
1
ꢀ1 [a]
ꢀ1
ꢀ1
ꢀ1
A
H
U
G
R
N
N
]
]
[calK mol ]
2
28ꢂ3
10.2ꢂ0.8
4.1ꢂ0.6
ꢀ32ꢂ3
ꢀ57ꢂ8
–
2
+
[b]
[Fe(O)
[Fe(O)
[Fe(O)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(tmc)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(CH
3
+
CN)]
2.0
[44]
[44]
–
+
2
[b]
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(N4Py)]
2.4
4.0ꢂ0.6
ꢀ57ꢂ8
2
+
[c]
[c]
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(tpa)(S)]
8ꢂ2
2
°
ꢀ1
[
[
a] At 295 K. [b] Calculated from experimental data in ref. [44].
c] Eyring analysis could not be performed because of limited thermal
IV
2
stability.
ate I1) is formed which is energetically favored compared to
ꢀ
1
the starting complex 2a by 2.2 kcalmol . Our spectroscopic
data do not provide evidence for a significant accumulation
of I1 in solution, but a dihydroxomanganese(IV) species has
Computational analysis of the water exchange mechanism:
An obvious goal arising from these studies is to establish the
fundamental chemical steps by which 2 can exchange its
oxygen atom with water. The answers to this question may
be of fundamental interest due to the structural similarity
between 2 and biologically relevant non-heme oxo-iron(IV)
species, in which the oxo group binds in a cis configuration
with respect to one or more available coordination sites. In
the heme paradigm, the incorporation of oxygen from water
is explained by the so-called oxo–hydroxo tautomerism pro-
recently been described for a related macrocyclic N
4
[61]
IV
ligand, and analogous LFe (OH) species have been pro-
2
posed on the basis of DFT methods for a bispidine-based
[32,62]
complex (Scheme 2).
It is possible that the energy of I1
is underestimated in our calculations, and that it is slightly
higher in energy than the corresponding oxo-aqua isomeric
species (see also below for a discussion based on computed
Mçssbauer parameters). In I1, the exogenous water mole-
cule has a H-bond interaction with the O5H hydroxide
ligand (oxo group in 2a). In the next step, the exogenous
water changes its interaction from the O5H hydroxo group
to the second hydroxo ligand (O6H, aqua in 2a) and the in-
termediate I2 is formed. This step is endothermic with a
[
41]
posed by Meunier and Bernadou, which entails the bind-
ing of labeled water trans to the oxo group and then tauto-
merization of this species to a symmetric trans-dihydroxo-
ACHTUNGTRENNUNGi ron(IV) intermediate that scrambles the label (Scheme 1).
ꢀ
1
This pathway clearly cannot be operative for 2, since there
is no binding site trans to the oxo group available for coordi-
nation with water, and an alternative mechanism must apply
in the present case. The mechanism depicted in Figure 7
starts with the iron(IV)-oxo complex 2a, with a coordinated
water molecule in a cis position with respect to the oxo
free energy cost of 4.2 kcalmol . When the free energy cor-
rections are included, TS2 and I2 are essentially isoenerget-
ic. Close inspection reveals that TS2 and I2 differ in the ori-
entation of the O5ꢀH bond, which is not involved in hydro-
gen bonding with the external water molecule. I2 evolves
+
through a second H transfer, passing through TS3 and
[49,58]
group. In the DFT study
of the water exchange mecha-
forming 2b, in which the oxo ligand is cis to the NCH py
2
nism, the solvent effects and the dispersion correction have
been taken into account in the single-point calculations. Sol-
vent effects for acetonitrile were computed using the polar-
ligand. Considering that TS2 and I2 are nearly isoenergetic,
the larger barrier along the overall process is TS2+TS3
°
ꢀ1
with DG =13.0 kcalmol , and corresponds to the second
proton transfer. Proton transfer not assisted by the second
water molecule has an activation enthalpy barrier about
three times higher. Isomer 2b could not be experimentally
[59]
izable continuum model (PCM) as implemented in Gaus-
ꢀ
1
observed because it is 2.11 kcalmol higher in energy than
a, and because the activation barriers associated with each
2
of the elemental steps that connect the backwards conver-
sion of 2a to 2b are small. In this scenario, water ligand ex-
change at 2b and microscopic microreversibility lead to the
formation of 2a, in which the oxygen atom of the oxo group
(
trans to a NCH Py unit) comes from water. Interestingly,
2
the mechanism for water exchange at 2 bears resemblance
IV
2+
to the DFT-computed conversion of [Fe (OH) L]
(in
2
which L is a tetradentate bispidine ligand related to bpd,
IV
2+ [32,62]
Scheme 2) to [Fe (O)
A
H
U
G
R
N
U
G
.
Calculations indicate
2
that this formal tautomerization takes place with a small
ꢀ1
energy barrier of 8.3 kcalmol . Unlike in 2, the reaction in-
IV
volves a spin crossover from the S=1 Fe (OH) to the ther-
2
ꢀ
1
IV
modynamically more stable (4.4 kcalmol ) S=2 Fe (O)-
(OH ) tautomer, without the assistance of a second water
AHCTUNGTRENNUNG
2
molecule. In contrast, all iron(IV) species in our calculations
Figure 7. DFT mechanism for water exchange at 2.
remain on the S=1 surface.
Chem. Eur. J. 2011, 17, 1622 – 1634
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
1627

E. Mꢂnck, J. M. Luis, L. Que, Jr., M. Costas et al.
Taking this computed mechanism into consideration, the
first-order kinetic dependence on water concentration is at-
tributed to the initial equilibrium step involving the binding
of a labeled water molecule, which replaces the initially
bound unlabeled water. Nam, Que, and co-workers have
proposed that a seven-coordinate cis-oxo-aqua intermediate
is formed in the first step of the reaction between [Fe(O)-
lower-energy species in the DFT-calculated mechanism, the
Mçssbauer parameters favor the oxo-iron(IV) formulation,
in agreement with the resonance Raman evidence for an
Fe=O unit.
Oxidation of sulfides: oxygen-atom transfer: The oxygen-
atom transfer ability of 2 was evaluated for the oxidation of
2
+
2+
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(N4Py)]
and [Fe(O)
A
T
N
T
E
N
N
A
H
U
G
R
N
U
(CH CN)]
with water
sulfides. Complex 2 reacted rapidly at 273 K in CH CN with
3
3
[
34]
(
10 equivalents of thioanisole affording the corresponding
sulfoxide (methylphenyl sulfoxide) in 98% yield. The reac-
tion was monitored with UV/Vis spectroscopy by following
the decrease of the band at 750 nm characteristic of the
iron(IV)-oxo complex 2. Under conditions of excess sub-
strate (5–50 equiv with respect to 2), the reactions showed
pseudo-first-order behavior so that the observed reaction
rates (kobs) were linearly dependent on substrate concentra-
2
+
and [Fe(O)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(tpa)(S)] , but owing to the presence of the
labile water/acetonitrile ligand, a six-coordinate intermedi-
ate 2b formed via a ligand-exchange reaction is favored.
The breaking of the FeꢀOH bond is likely to be the origin
2
of the higher activation enthalpy which, however, is entropi-
cally compensated because of the dissociative nature of the
event.
An obvious consequence of the present mechanism is
that, owing to the asymmetry of the two cis-labile sites,
oxygen incorporation from water into the oxo ligand re-
quires two oxo–hydroxo tautomerism reactions. That consti-
tutes a fundamental mechanistic difference from oxygen-
tion. From this analysis, a second-order rate constant (k ) of
H
ꢀ1
ꢀ1
(1.0ꢂ0.1)m
s
was obtained for the oxidation of thioani-
sole (see the Supporting Information). Additional informa-
tion about the mechanism of this transformation was gained
through the initial observation that the reaction rates were
highly dependent on the para substituent of the aromatic
ring. Thus, we measured the second-order rate constants
[41]
atom incorporation from water in metalloporphyrins.
DFT computation of the Mçssbauer parameters of the spe-
cies implicated in the water exchange mechanism: Consider-
ing that the DFT-computed water exchange mechanism pre-
dicts that tautomeric bis-hydroxo-iron(IV) (I1 and I2) and
oxo-iron(IV) (2a and 2b) are close in energy, the corre-
sponding Mçssbauer parameters were calculated with DFT
methods and the results are collected in Table 2 (see the Ex-
(k ) for a series of para-substituted methyl phenyl sulfides,
X
p-X-thioanisoles (X=CN, Cl, CH , OCH ). Plotting of log-
3
3
AHCTUNGTNERNUG( k /k ) for the different substrates against the correspond-
X H
ing Hammett parameters (s ) afforded a good linear corre-
p
lation, and a Hammett value of 1=ꢀ1.5 was obtained
(Figure 8). The negative 1 value reflects the electrophilicity
[
a]
Table 2. DFT-calculated Mçssbauer parameters of oxo-iron(IV) (2a
and 2b) and bis-hydroxo-iron(IV) (I1 and I2) tautomeric species.
ꢀ
1
[a]
ꢀ1
Compound
DE
Q
[mms
]
d [mms ]
2
I1
I2
2
Exp.
a
0.57
2.28
2.44
0.83
0.73
0.18
0.12
0.18
0.17
0.05
b
[
a] See experimental section for details of the DFT analysis.
Figure 8. Hammett plot representing log
X H
ACHTUNGTRENNGU( k /k ) against the Hammett pa-
rameter (s ) for the reaction of 2 with p-X-thioanisoles at 273 K.
p
perimental Section for details of the DFT analysis). The cal-
culated isomer shifts for 2a and 2b are a bit too high rela-
tive to the experimental values. The Pittsburgh group, using
Gaussian with the B3LYP functional (calibration of Vrajma-
of the oxo group in 2. On the other hand, a plot of log(kX)
against the one-electron oxidation potentials of each p-X-
[14a]
su et al. ), observed that the calculated d values for tmc
IV
come out quite well whereas those of most Fe =O com-
thioanisole (E8 ) afforded a linear correlation with a slope
ox
plexes with pyridine and amine ligands are generally too
high, by as much as 0.09 mms . For the question at hand it
is significant that the DEQ values of I1 and I2 are much
of ꢀ3.0 (see the Supporting Information). As slope values
ꢀ1
around 10 would have been expected for a process initiated
[63,64]
by one-electron transfer,
the observed slope of 3 indi-
larger than those of 2a and 2b. In agreement, large DE
values have been reported for [Fe (OH) ACHUTNRGNENUG( OOtBu)-
cates that the oxidation of sulfides by 2 does not occur by
such a process but rather through a direct oxygen-atom
transfer mechanism. Further evidence for a direct oxygen-
atom transfer between the terminal oxo ligand and the sul-
Q
IV
2
+ [14b]
ꢀ1
IV
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(bpmcn)]
(OH)(LOMe)Fe
thoxy-3,5-dimethylpyridin-2-yl)d -methyl]amine,
(DE =1.75 mms ) and the Fe -OH site of
Q
IV
IV
3+
[
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(m-O)Fe (O)
A
H
U
G
R
N
U
G
(LOMe =tris[(4-me-
1
8
DE =
fide came from labeling of the oxo group with O. This was
2
Q
ꢀ
1
[14c]
18
1
.96 mms ).
628
We conclude that, despite I1 being the
achieved by oxygen-atom exchange of 2 with H2 O and sub-
1
www.chemeurj.org
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2011, 17, 1622 – 1634

Modeling the Binding Site Motif of Non-Heme Iron Oxygenases
FULL PAPER
IV 18
Me,H
Table 3. C H bond dissociation energies (BDEs) and rates for the reac-
tion of 2 with different substrates.
ꢀ
sequent reaction of the resulting [Fe
A
H
U
G
E
N
N
( O)(
Pytacn)-
[
a]
2
+
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(H O)] with thioanisole. Analysis of the resulting sulfox-
ide by GC-MS indicated that 80% of the product was la-
2
ꢀ
1
[67,68]
ꢀ1 ꢀ1
Substrate
BDE [kcalmol
]
2
k [M s ]
18
beled with O.
10-methyl-9,10-dihydroacridine
73.7
86ꢂ19
(
AcrH
2
)
Despite the similarities between the oxygen-atom transfer
process in 2 and in other synthetically prepared iron(IV)-
oxo species, reaction rates are highly dependent on the spe-
cific structure of the ligand. In particular, the oxidation of
xanthene
9
1
75.5
8.1ꢂ0.8
5.7ꢂ0.9
4.2ꢂ0.3
1.2ꢂ0.2
,10-dihydroanthracene (DHA) 77
,4-cyclohexadiene (CHD)
78
80
84
93
fluorene
IV
2+
IV
2+
ꢀ2
thioanisole by [Fe (O)
A
H
U
G
R
N
U
G
A
H
U
G
E
N
N
2,3-dimethyl-2-butene
tetrahydrofuran (THF)
cyclohexane
A
H
U
G
R
N
U
G
ꢀ
3
ACHTUNGTRENNUNG
(
(
[
b]
99.3
ACHTUNGTRENNUNG
ꢀ
1
ꢀ1
273 K) and 0.075m
s
IV
2+
[a] Reactions were run in acetonitrile at 258 K, with 2 generated in situ
from the reaction of 1 (1 mm) with 2 equiv peracetic acid at 158C.
was even smaller for [Fe (O) ACHUTNTGERNNUG( tmc) CAHTUNGTRENNNUG
value of 0.029m
3
ꢀ
1
ꢀ1
[31]
s
at 358C was obtained. Comparison
[
b] 298 K.
IV
2+
with the reactivity of [Fe (O) ACHUTNTGNRENUNG( tpa)(S)] is hampered due
to its inherent instability, which prevents proper measure-
ment of the second-order rate constant at relatively high
formation of brown–dark blue intense chromophores, indi-
cative of alkylphenolate-bound iron(III) species. Consider-
ꢀ
1
ꢀ1
temperatures (a value of 0.44m
action with thioanisole at 228 K has been reported). Nev-
s
corresponding to the re-
ACHTUNGTRENNUNG
[31]
ing the high bond dissociation energy (BDE) of the CꢀH
bond in aromatic substrates, such reactions could not occur
through H-atom abstraction but instead they originate from
ertheless, from these studies it can be concluded that 2 and
IV
2+
[
Fe (O) ACHTUNGTRENNUNG( tpa)(S)] , both of which bear tripodal tetraden-
[37]
tate-based ligands, are much more reactive in the oxygen-
atom transfer reaction than the corresponding complexes
bearing tmc (tetradentate planar) or N4Py and Bn-tpen
electrophilic attack of the aromatic ring by 2.
For this
reason, these substrates were not further addressed in the
present study.
(
pentadentate) ligands.
Complex 2 proved to be a very powerful oxidant in C–H
oxidation reactions (Tables 3 and 4). Compared to related
dicationic iron(IV)-oxo complexes under analogous condi-
Oxidation of activated CꢀH bonds: hydrogen-atom abstrac-
tion: The reactions of 2 with a series of substrates for H-
atom abstraction were studied. Complex 2 reacted with
[
a]
Table 4. DHA oxidation rates of various iron-oxo complexes.
5
equivalents of 9,10-dihydroanthracene (DHA) at 258 K af-
ꢀ
1 ꢀ1
Complex
k
2
[M
s
]
T [K]
Ref.
fording anthracene as the only product detected by GC-MS.
When the reaction was run under N , 0.45 mmol anthracene
per mmol of 2 was measured. The UV/Vis spectra at the
end of the reactions did not show formation of 1. In addi-
tion, the ESI mass spectrum of the resulting solution was
dominated by ions corresponding to mononuclear Fe spe-
cies. Therefore, considering that Mçssbauer spectra indicate
a 85(5)% Fe content in our preparations, on the basis of
stoichiometric considerations we conclude that in this reac-
2
5.7ꢂ0.9
2.8
8.0
258
258
238
243
298
193
193
193
193
258
243
2
IV
2+
A
H
U
G
R
N
U
G
A
H
U
T
E
N
N
[75]
[35]
[24]
[66]
[65]
[65]
[65]
[65]
[76]
[24]
IV
IV
III
2+
2+
A
H
U
G
R
N
U
G
A
H
U
T
N
U
G
A
H
U
G
R
N
U
G
3
CN)]
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
A
T
N
T
E
N
N
(CH CN)]
3
0.016
2+
ꢀ3
A
H
U
G
R
N
U
G
A
H
U
G
R
N
N
A
H
N
T
E
N
N
5.0ꢈ10
28
III
IV
2+
3+
III
[(OH)
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
A
H
U
G
E
N
N
(LOMe)]
III
IVA
3+
ꢀ5
[
Fe Fe
C
T
N
T
E
N
G
2
A
T
N
T
E
N
N
(LOMe
)
2
]
10
10
IV
4+
ꢀ4
A
H
U
G
R
N
U
G
2
A
H
N
T
E
U
G
2
A
H
N
T
E
N
N
(LOMe
)
2
]
IV
IV
IV
[
(OH)
A
H
U
G
R
N
N
(LOMe)Fe
A( m-O)Fe (O)
H
U
G
R
N
U
G
A( LOMe)]
H
U
G
R
N
N
0.027
2.7
0.090
IV
ACTHUNGERTNNUNG[ Fe (O) ACHTGNUTRENNUNG
ꢀ
IV
2+
A
H
U
G
R
N
U
G
A
H
U
G
R
N
G
3
tren)]
tion 2 acts as a 1e oxidant (Scheme 4), and that the yield of
the reaction with DHA is nearly quantitative.
2
[a]LOMe =tris[(4-methoxy-3,5-dimethylpyridin-2-yl)d -methyl]amine,
tmp=tetramesitylporphinate, TMG tren=1,1,1-tris{2- AHCTUTGNENRNGU[ N -(1,1,3,3-tetra-
3
2
methylguanidino)]ethyl}amine.
tions, reaction rates obtained with 2 are somewhat higher
Scheme 4.
IV
2+
than those obtained with [Fe (O)
ACHTUNGTRENNUNG( N4Py)] and nearly two
IV
orders of magnitude faster than with [Fe (O)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(tmc)-
2
+[24,34]
The kinetics of reactions of 2 with a series of substrates
were studied by UV/Vis spectroscopy following the decay of
the band at 750 nm characteristic of the iron(IV)-oxo spe-
cies 2. The decay of 2 followed good first-order kinetics
under conditions of excess substrate and kobs values were lin-
early dependent on substrate concentration in all cases, thus
allowing us to determine the corresponding second-order
rate constants, k (see the Supporting Information). These
values are collected in Table 3. Reaction of 2 with alkylaro-
matic substrates (toluene and ethylbenzene) caused rapid
ACHTUNGTNRENUNG( CH CN)]
3
but somewhat lower than that found for
IV
+2
[35]
[Fe (O)
A
H
U
G
R
N
N
(bpd)
A( CH CN)] (Scheme 2).
H
U
G
R
N
U
Furthermore, the
3
second-order rate constant for the oxidation of DHA with 2
appears to be several orders of magnitude higher than those
III
IV
IV
IV
reported for low-spin Fe Fe and Fe Fe oxo-dimers, and
ꢀ
1
ꢀ1
approaches the remarkably high rate (k =28m
s
at
2
193 K) recently measured for the high-spin [(HO)-
III
IV
2+
ACHTUNGRTNEG(NUN L )Fe -O-Fe (O) ACHTUNGTRENNNU(G LOMe)] , in which LOMe =tris[(4-me-
OMe
2
[65]
thoxy-3,5-dimethyl-pyridin-2-yl)methyl]amine (Table 4).
The high reactivity of the latter has been rationalized on the
Chem. Eur. J. 2011, 17, 1622 – 1634
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
1629

E. Mꢂnck, J. M. Luis, L. Que, Jr., M. Costas et al.
[
74,77,78]
basis of the high spin (S=2) state of the iron ions, and the
compound has limited stability even at ꢀ808C. On the other
hand, as already discussed, the metal ion found in 2 has an
S=1 spin state, and it is moderately stable at room tempera-
ture. As expected from its higher oxidation state, reaction
rates with 2 are 2–3 orders of magnitude faster than those
nism dominated by quantum mechanical tunneling.
For comparison, KIE values around 17 were obtained for
IV
n+
the oxidation of DHA by [Fe (O) ACTHUNGTERNN(UNG tmc)(X)] (X=CH CN,
3
CF COO , or N ) and even larger values were reported
in the oxidation of benzyl alcohol by [Fe (O) ACHUNTGRNENUG( N4Py)]
(KIE=48) and [Fe (O) ACHUTNGRNEGUN( tpa)(S)] (KIE=58). Moreover,
such large isotope effects have also been observed in hydro-
gen-atom abstraction reactions performed by the iron(IV)-
ꢀ
ꢀ
3
[34]
3
IV
+2
IV
+2
[33]
III
2+
obtained with the lipoxygenase model [Fe
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
[66]
(
Py5=2,6-bis[bis(2-pyridyl)methoxymethane]pyridine).
ACHTUNGTRENNUNG
[
79]
Indeed, 2 proved to be competent for performing the oxida-
tion of strong CꢀH bonds such as those in THF (BDE=
oxo intermediate of TauD (KIEꢁ37),
the iron ACHTUNGTRENNUNG( III)-hy-
[
80]
droxo active species of lypoxygenase (KIEꢁ50),
and
ꢀ
1
IV
IV
9
1
3 kcalmol ) with
a
reaction rate k =(2.3ꢂ0.3)ꢈ
compound Q (Fe Fe ) of the diiron enzyme soluble meth-
2
ꢀ
3
ꢀ1 ꢀ1
[67]
[81]
0
m
s
at 258 K. Most remarkably, 2 is also competent
ane monooxygenase (sMMO) (KIE>50).
to perform the oxidation of cyclohexane (BDE=99.3 kcal
mol ) at room temperature with a second-order rate con-
A particular case that deserves some comment is the reac-
tion with the nicotinamide adenine dinucleotide (NADH)
ꢀ
1
stant, after correction for the number of CꢀH bonds, of
analogue 10-methyl-9,10-dihydroacridine (AcrH ), which is
2
ꢀ
5
ꢀ1 ꢀ1
[82]
k’ =3.6ꢈ10
2
m
s . GC analysis of the reactions revealed
considered a hydride donor. The reaction of AcrH with 2
2
ꢀ1
ꢀ1
the formation of 0.12 mol cyclohexanone per mol of 2 and
.28 mol cyclohexanol per mol of 2. Nevertheless, this reac-
results in rapid formation (k =(86ꢂ19)m s ) of the acri-
2
+
0
dinium cation AcrH , as evidenced by UV/Vis spectroscopy.
tion rate is relatively slow when compared with the fast oxi-
UV/Vis quantification based on the characteristic spectral
+
dation observed in catalytic 1/H O cyclohexane oxidations,
features of the cation accounts for 1 mmol AcrH per mmol
2
2
V
Me,H
2+
ꢀ
in which an [Fe (O)(OH)(
Pytacn)] species has been
2, which indicates that 2 acts as a 2e oxidant and that the
inferred from isotopic labeling experiments and DFT analy-
sis.
reaction is best described as a formal hydride transfer to the
iron(IV)-oxo moiety. Interestingly, the corresponding oxida-
[
45,50]
Comparative reactivity profiles of the reactions of 2
against different substrates show that the rate constants de-
creased with the increase of the CꢀH BDE and more inter-
tion rate constant (k’ ) appears to be only slightly faster
than expected from the log ACHTUNGTRENN(UNG k’ ) versus BDE graph correla-
2
tion (Figure 9), thus suggesting that a common rate-deter-
mining step is operative for all the series of substrates plot-
ted. In addition, the KIE value obtained from the reaction
2
estingly the log
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(k’ ) values correlated linearly with the CꢀH
2
BDE values of the substrates, giving a slope of approximate-
ly ꢀ0.2 (Table 3, Figure 9). The slope is comparable to those
of 2 with AcrD at 258 K is 4.7 (Figure S11 in the Support-
2
ing Information), which is intermediate between the values
IV
+2
[82]
observed for [Fe (O)
A
H
U
G
R
N
N
(N4Py)]
(KIE=13.5)
(bpd)(CH CN)]
.3; Scheme 2). On this basis, we propose that the reac-
and those
IV
2+
obtained recently with [Fe (O)
A
H
U
T
E
N
N
A
H
U
T
E
N
N
(KIE=
3
[
35]
2
tion takes place through a rate-determining hydrogen-atom
abstraction that can be regarded as a proton-coupled elec-
[68,76,83,84]
tron transfer, followed by a fast electron transfer.
Considering that CꢀH bond oxidation by 2 occurs through
[73,85,86]
a hydrogen-atom abstraction mechanism,
the balance
between the strength of the cleaved CꢀH bond and of the
III
formed Fe ꢀOH bond must determine the observed reac-
tion rates. Along this interpretation, we notice that reactions
mediated by 2 are comparable to those associated with the
Figure 9. Plot of log
of different substrates.
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(k’
2
) (determined at ꢀ158C) against the CꢀH BDE
IV
most reactive Fe (O) species containing nitrogen-based
IV
2+
pentadentate ligands such as [Fe (O) ACHUTNGRNENUG( N4Py)] , which con-
stitute one of the very rare examples of well-defined metal–
obtained for hydrogen abstraction reactions mediated by
oxo species that are capable of breaking the strong CꢀH
IV
2+ [16]
III
2+ [66]
[18,87–89]
[
Fe (O)
mediate between the ꢀ0.4 and ꢀ0.1 observed for [Ru (O)-
(bpy) (py)] and [Mn (OMe)(Py5)] respectively.
Such a correlation strongly suggests that reactions take
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(N4Py)]
and [Fe
A
H
U
G
R
N
N
(OMe)
A
H
U
G
E
N
N
(Py5)]
,
and inter-
bond of cyclohexane at room temperature.
The simi-
IV
IV
2+
lar reaction rates observed for 2 and [Fe (O) ACHUNTGRNENUG( N4Py)]
2
+
III
2+ [69,70]
III
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
A
H
U
G
R
N
U
G
A
H
U
G
R
N
U
G
,
should be interpreted as both of them forming a Fe OꢀH
2
bond of approximately the same energy. Previous literature
[69,71–73]
III
IV
place through a H-atom abstraction mechanism,
earlier established for related Fe compounds.
as
reports on the estimation of the Fe OꢀH bond in [Fe (O)-
IV
[18,34,74]
2+
ACHTUNGTRENNUN(G N4Py)] have provided somewhat different values. Two in-
III
Parallel reactions with deuterated DHA (d -DHA) yield-
dependent theoretical studies yielded Fe OꢀH BDEs of
4
[90,91]
ꢀ1 [92]
ed a kinetic isotope effect (KIE) of 27, a value which is con-
84
and 97 kcalmol , and some of us have experimen-
ꢀ
1
sistent with CꢀH bond cleavage being the rate-determining
tally estimated a more modest BDE=78 kcalmol from its
experimentally observed redox potential in water. We can
[68]
[75]
step. This large KIE value is well above the semiclassical
limit of 7, thus suggesting a hydrogen-atom transfer mecha-
take the latter value as a lower limit estimate of the corre-
1630
www.chemeurj.org
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2011, 17, 1622 – 1634

Modeling the Binding Site Motif of Non-Heme Iron Oxygenases
FULL PAPER
III
sponding Fe OꢀH bond formed by H abstraction in 2.
Conclusion
IV
2+ [90]
Thus, as previously noted for [Fe (O)
A
H
U
G
R
N
U
G
,
despite
its rather surprising thermal stability, 2 is a very powerful
oxidant. Besides the high oxidation state, one of the reasons
at the origin of such a high reactivity is likely to be the posi-
We have reported the preparation of a new S=1 iron(IV)-
oxo species (2) with a tripodal tetradentate ligand that has
remarkable thermal stability, especially when compared with
IV
2+
IV
tive charge of the [LFe (O)] unit when L is a neutral
structurally related complexes such as [Fe (O)-
Me,H
2+
IV
2+
IV
ligand (N4Py or
philic oxidant.
Pytacn), which results in a very electro-
A
H
U
G
E
N
N
(bpmcn)(S)]
,
[Fe (O)
A
H
U
G
R
N
N
(tpa)(S)]
,
and [Fe (O) ACHTUNGTRENNUNG( bpd)-
+
2
AHCTUNGTREGUN(NN CH CN)] . Complex 2 has a labile site cis to the oxo
3
The remarkably high oxidative reactivity exhibited by 2
both in O-atom transfer and in H abstraction reactions is
unprecedented and deserves some comment. The factors
that determine the reactivity of oxo-iron(IV) species have
been the focus of intense debate. The accessibility of multi-
ple spin states modulated by the nature of the ligand trans
to the oxo group has been invoked to explain inverted ten-
dencies in oxygen-atom transfer versus H abstraction reac-
group, which constitutes a very common structural feature
of non-heme iron oxygenases. Complex 2 rapidly exchanges
1
8
its oxygen atom with H2 O following a mechanism in which
an exogenous water molecule assists the hydrogen transfer
from the coordinated water molecule to the oxo group. De-
spite its remarkable thermal stability, 2 is a very good oxi-
dant both with respect to oxygen-atom transfer to sulfides
and hydrogen-atom abstraction of alkane CꢀH bonds, thus
IV
[34]
tion by a series of Fe compounds.
Basic anionic li-
demonstrating a highly electrophilic character arising from
IV
+
IV
gands X in a series of [Fe (O)(X)
A
H
U
G
E
N
N
(tmc)] complexes de-
the Fe oxidation state in a neutral N-based ligand environ-
crease their electrophilicity and O-atom transfer reactivity,
but enhance their H abstraction reactivity by populating a
reactive quintet state. Population of an excited, more highly
reactive S=2 state has also been proposed to account for
ment. The higher oxidative reactivity of 2 when compared
with complexes containing pentadentate ligands is attributed
Me,H
to the tetradentate nature of the
Pytacn ligand, which
does not provide steric encumbrance of the oxo ligand in 2,
thus providing the basis of its remarkable oxidative reactivi-
ty.
IV
2+
the exceptional ability of [Fe (O)
A
H
U
G
R
N
U
G
to cleave
strong CꢀH bonds through a H-atom abstraction mecha-
[90]
nism.
The nature of the ligands cis to the oxo group is
considered to have a more modest effect on the oxidative
[93]
reactivity. Computational studies have thus suggested that
IV
high-spin S=2 Fe (O) species could be much more reactive
than their corresponding low-spin S=1 analogues. However,
Experimental Section
the recent preparation and characterization of the first syn-
thetic S=2 Fe (O) species raises some questions about this
Materials and methods: Reagents and solvents used were of commercial-
ly available reagent quality unless otherwise stated. Preparation and han-
dling of air-sensitive materials were performed under an inert atmos-
phere either on a Schlenk line or in a glove box. Acetonitrile was pur-
IV
prediction, because it exhibits reactivity only comparable to
IV
2+ [24,25]
that of [Fe (O)
A
C
H
T
U
N
G
T
R
E
N
N
U
N
G
(N4Py)]
.
Steric effects were invoked
1
8
18
chased from Scharlau. H
ICON Isotopes.
2
O (95% O-enriched) was received from
by the authors to account for these observations. Complex 2
contains an S=1 spin state, and DFT computations show
that it is well separated in energy from the S=2 spin state
Me,H
Synthesis of complexes: The starting complex [Fe
A
T
N
T
E
N
N
3
SO
3
)
2
(
Pytacn)]
(
1) was prepared by following the previously reported experimental pro-
[45]
(
see Table S7 in the Supporting Information). Therefore, a
cedure. Preparation of iron(IV)-oxo complex 2 was performed as fol-
lows. Peracetic acid solution (2 equiv, 100 mL of a 40 mm solution in
two-state reactivity (TSR) scenario appears highly unlikely.
In addition, high reaction rates are measured for both O-
atom transfer and H-atom abstraction reactions. Its high oxi-
dative reactivity may then add to the rationale of substrate
accessibility to the reactive oxo ligand as a major factor dic-
tating reactivity in these species. The high reactivity of 2 in
C–H oxidation reactions finds a nice precedent in that re-
3
CH CN obtained by dilution of commercially available 32% w/w perace-
tic acid solution in acetic acid) was added at 288 K to a solution of 1 in
acetonitrile (1 mm, 2 mL total volume). The formation of 2 was followed
by UV/Vis spectroscopy, which showed the appearance of a band at
ꢀ
1
ꢀ1
750 nm (lmax =200m cm ) with a shoulder at 900 nm that developed in
about 10 min. The resulting solution of 2 (1 mm) was directly used for
subsequent reactivity studies.
III
Me,H
IV
+2
Direct
(ClO
solved in MeCN (2 mL) under an inert atmosphere. An acetonitrile solu-
tion (200 mL) 0.30m in CH COOH and Et N (60 mmol CH COOH and
Et N) were added at once, which caused an immediate color change
from dark pink to bright yellow. Immediately a balloon filled with O
preparation
of
[Fe
A
H
U
G
R
N
N
(m-O)
A
H
U
G
R
N
U
G
2
Pytacn) ]-
2
3
cently
(
described
for
[Fe (O) AHCTNUGTRENNUNG( bpd) ACHTUNGTNERNUNG( CH CN)]
3
A
H
U
G
R
N
U
G
4
)
3
·CH CN (3·CH
3
3
CN): Compound 1 (90 mmol, 54.6 mg) was dis-
[
35]
Scheme 2). Both compounds have a N -based ligand that
4
enforces a cis-oxo-labile site. However, the bispidine is a
3
3
3
[32,62]
Me,H
[50]
weaker-field ligand
than
Pytacn, and the TSR sce-
3
nario is more likely. In this scenario, we propose that 2 is
more reactive than most low-spin iron(IV)-oxo species bear-
ing pentadentate and/or planar tetradentate ligands, because
the oxo ligand in 2 is more exposed and susceptible to inter-
2
was connected to the reaction vessel and the solution became red-brown
in a few seconds. After stirring for 3 h, the solvent from the resulting so-
lution was removed under reduced pressure which afforded a brown oil.
The resulting product was redissolved in acetonitrile and NaClO ·H O
4
2
action with small substrate molecules. However, compared
(135 mmol, 19 mg) was added. The solution was stirred for about 2 h, fil-
tered through Celite, and diethyl ether was slowly diffused. Brown crys-
Me,H
with previously reported tetradentate ligands,
Pytacn af-
3
tals of 3·CH CN (37 mg) suitable for X-ray diffraction were obtained
fords a very significant degree of steric protection against bi-
metallic dimerization reactions.
1
(
1
C
38 mmol, 84%). H NMR (200 MHz, CDCl
3
, 300 K): d=26.43, 18.27,
COO), 1447 (C=
4 3
ar), 1073, 621 cm (ClO ); UV/Vis (CH CN): lmax (e/Fe)=427 (660),
7.41, 14.47, 6.57 ppm; FTIR (ATR) n=1611, 1523 (CH
3
ꢀ
1
Chem. Eur. J. 2011, 17, 1622 – 1634
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
1631

E. Mꢂnck, J. M. Luis, L. Que, Jr., M. Costas et al.
ꢀ
1
ꢀ1
4
62 (690), 513 (500), 692 nm (58 mol cm ); ESI-MS: m/z: 227.6
grants. We thank RahuCat for a generous gift of tritosyl tacn. We thank
W. Han and L. Noodleman for providing the utility programs to compute
the Mçssbauer parameters of the iron complexes and M. Swart for his
useful comments about the Mçssbauer calculations.
+
3
+
[
Mꢀ3ClO
4
]
, 880.9 [MꢀClO
4
] .
Oxygen-atom exchange with water: Kinetic studies on oxygen-atom ex-
1
8
change of 2 with H
2
O were performed by quenching aliquots of the re-
action mixture with thioanisole at different times, and analyzing by GC-
1
8
MS the percentage of O-labeled methylphenyl sulfoxide generated. The
pH dependence of the reaction rates was not studied. In a typical experi-
1
8
2
ment, an appropriate amount of H O (6–60 mL) was added to a stirred
solution of 2 in acetonitrile (2.5 mL, 1 mm) at once. At different reaction
times, an aliquot of the reaction mixture (350 mL) was directly poured
[
[
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1] M. M. Abu-Omar, A. Loaiza, N. Hontzeas, Chem. Rev. 2005, 105,
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2
3
into a solution containing thioanisole (7 mL) in CH CN (300 mL). The re-
1
sulting yellow solution was stirred for 30 min at room temperature, fil-
tered through a short path of basic alumina, and washed with ethyl ace-
1
8
tate (2 mL). The sample was analyzed by GC-MS. The percentage of O-
labeled sulfoxide was determined by the isotopic pattern shown by the
peaks at m/z 125 and 140. In a typical experiment, a total of seven ali-
quots were removed from the reaction mixture. The temperature of the
reaction mixture (0–308C) was controlled by means of a cryostat or a
2
000, 100, 235.
[
[
[
[
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1
8
thermostatized water bath. The percentage of O-labeled sulfoxide is di-
rectly related to the percentage of iron(IV)-oxo species 2 that has ex-
1
8
18
2
changed its oxygen atom with H O. The percentage of O-labeled sulf-
oxide over time could be fitted to a pseudo-first-order kinetics that al-
lowed us to measure the rate constants (kobs) corresponding to oxygen-
1
8
atom exchange with O-labeled water in 2.
Oxidation of activated CꢀH bonds and sulfides: Appropriate amounts of
substrates (diluted in acetonitrile) were added to a solution of 2 (1 mm)
and the subsequent decay of the spectral changes corresponding to the
iron(IV)-oxo species was directly monitored by UV/Vis spectroscopy.
The kinetic studies were performed at specific temperatures: 273 K for
the oxidation of sulfides and 258 K for the oxidation of activated CꢀH
bonds. Unless specifically stated, reactions were carried out under air.
Rate constants, kobs, were determined by pseudo-first-order fitting of the
decrease of the absorption band at 750 nm. Product analyses for the oxi-
dation of thioanisole and DHA were performed by gas chromatography.
Prior to injection, an internal standard (biphenyl) was added to the solu-
tion, which was further filtered through basic alumina and washed with
ethyl acetate. Calibration curves with authentic products were generated
to allow quantitative determination of the oxidized products.
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Financial support for this work was provided by MEC-Spain (Projects
CTQ2009–08464/BQU and CTQ2008–06696/BQU), and the US National
Institutes of Health (GM-33162 to L.Q. and EB-001475 to E.M.). M.C.
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www.chemeurj.org
ꢅ 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2011, 17, 1622 – 1634