Fe-amino acid complexes immobilized on silica gel as active and highly selective catalysts in cyclohexene epoxidation

Article abstract of DOI:10.1007/s11164-015-1920-x

In this work, the syntheses, structure, superoxide dismutase (SOD) activity, and the catalytic use in the oxidative transformations of cyclohexene of covalently grafted Fe(III)-complexes formed with various or various combinations of C-protected amino acid (L-histidine, L-tyrosine, L-cysteine and L-cystine) ligands are presented. The structural features of the surface complexes were studied by XANES/EXAFS and mid/far-IR spectroscopies. The compositions of the complexes were determined by ICP-MS and the Kjeldahl method. The SOD activities of the materials were evaluated in a biochemical test reaction. The obtained materials were used as catalysts for the oxidation of cyclohexene with peracetic acid in acetone. Both covalent grafting and building the complex onto the surface of the chloropropylated silica gel were successful in most cases. In many instances, the obtained structures and the coordinating groups were found to substantially vary upon changing the conditions of the syntheses. All the covalently immobilized Fe(III)-complexes displayed SOD activities, and most of them were found to be capable of catalyzing the oxidation of cyclohexene with appreciably high activities and outstanding epoxide selectivities.

Full text of DOI:10.1007/s11164-015-1920-x

Res Chem Intermed
DOI 10.1007/s11164-015-1920-x
Fe-amino acid complexes immobilized on silica gel
as active and highly selective catalysts in cyclohexene
epoxidation
´
Gabor Varga Zita Csendes Eva G. Bajnoczi
•
•
•
´
Stefan Carlson Pal Sipos Istvan Palinko
´
•
•
´
´
´
´
Received: 1 October 2014 / Accepted: 2 January 2015
Ó Springer Science+Business Media Dordrecht 2015
Abstract In this work, the syntheses, structure, superoxide dismutase (SOD)
activity, and the catalytic use in the oxidative transformations of cyclohexene of
covalently grafted Fe(III)-complexes formed with various or various combinations
of C-protected amino acid (L-histidine, L-tyrosine, L-cysteine and L-cystine) ligands
are presented. The structural features of the surface complexes were studied by
XANES/EXAFS and mid/far-IR spectroscopies. The compositions of the complexes
were determined by ICP-MS and the Kjeldahl method. The SOD activities of the
materials were evaluated in a biochemical test reaction. The obtained materials were
used as catalysts for the oxidation of cyclohexene with peracetic acid in acetone.
Both covalent grafting and building the complex onto the surface of the chloro-
propylated silica gel were successful in most cases. In many instances, the obtained
structures and the coordinating groups were found to substantially vary upon
changing the conditions of the syntheses. All the covalently immobilized Fe(III)-
complexes displayed SOD activities, and most of them were found to be capable of
catalyzing the oxidation of cyclohexene with appreciably high activities and out-
standing epoxide selectivities.
´
´
G. Varga ꢀ Z. Csendes ꢀ I. Palinko (&)
´ ´
Department of Organic Chemistry, University of Szeged, Dom ter 8, Szeged 6720, Hungary
e-mail: palinko@chem.u-szeged.hu
´
´
´
´
G. Varga ꢀ Z. Csendes ꢀ E. G. Bajnoczi ꢀ P. Sipos ꢀ I. Palinko
´
Material and Solution Structure Research Group, Institute of Chemistry, University of Szeged, Dom
´
ter 7-8, Szeged 6720, Hungary
´
´
E. G. Bajnoczi ꢀ P. Sipos
´ ´
Department of Inorganic and Analytical Chemistry, University of Szeged, Dom ter 7, Szeged 6720,
Hungary
S. Carlson
MaxIV-Lab, Lund University, 223 63 Lund, Sweden
123

G. Varga et al.
Keywords Silica-anchored ꢀ Fe(III)-C-protected amino acids ꢀ Covalent
anchoring ꢀ Structural characterization ꢀ Catalytic activity and selectivity
Introduction
Enzymes catalyze a wide range of chemical transformations. They often provide
high regio- and stereoselectivity and operate under physiological conditions.
Enzyme-catalyzed reactions can be alternatives to traditional organic syntheses
under environmentally benign conditions [1]. Outside the usual physiological
environment, such as temperatures higher than the physiological, pH, or presence of
non-aqueous solvents, enzymes are unstable and are easily inactivated. Therefore,
they have several limitations for broader applications like catalysts in the synthesis
of fine chemicals. Moreover, the recovery and the reuse of enzymes are also
cumbersome. These drawbacks can be eliminated and more stable and reusable
catalysts may be produced by immobilizing the enzyme over various supports by
employing methods that preserve the catalytic activity and selectivity of the
support-free enzyme [2, 3].
Active and selective solid catalysts can also be fabricated by immobilizing either
the active site itself, or its structural or functional model [4]. Both approaches are
applied in various laboratories, and promising results emerge. These biomimetic
catalysts comprise redox-active transition metal ions [5] complexed by amino acids
[6], or other molecules that are capable of coordination [7]. The immobilized
complexes are often called bioinspired catalysts, and their activities and selectivities
may resemble those of the enzymes. These substances are capable of operating
under more rigorous conditions, and they can easily be recovered and recycled [8].
In this contribution, the active center of the Fe(III) superoxide dismutase (SOD)
enzyme was used for inspiration [9]. It appears in the form of dimers or tetramers
with one Fe ion per monomer unit and with a molecular weight of 22 kDa. The
geometry around the metal ion is trigonal bipyramidal, coordinated by two histidine
and one aspartate ligands in the equatorial plane and a histidine and a solvent
molecule in the axial plane. The solvent molecule, which is OH- in the oxidized
and H2O in the reduced form of the enzyme, is supported by an extensive network
of H-bonds [10, 11].
There is hope to obtain efficient, durable, and recoverable electron transfer
catalysts that can be reused, if one can immobilize the functional mimics of this
enzyme in one way or another (e.g., hydrogen bonding, ion exchange, covalent
anchoring) on solid supports of various kinds (e.g., zeolites, layered materials,
resins, unmodified or surface-modified silica gel). This hope is supported by a range
of literature results [12–17].
Catalytic epoxidation of alkenes by various oxidants is of interest, since epoxides
are intermediates and precursors to many useful chemical products [18], such as
food additives, agrochemicals, drugs, [19] perfumes, and sweeteners [20].
Cyclohexene oxide is used in the synthesis of many products, e.g., chiral
pharmaceuticals, epoxy paints, pesticides, dyestuffs, and rubber promoters [21, 22].
123

Fe-amino acid complexes
Mainly hydrogen peroxide, organic peroxides (tert-butyl hydroperoxide),
peracids (peracetic acid, m-chloroperbenzoic acid), and molecular oxygen are used
as oxidants for oxidation of alkenes [23–25].
Peracids can epoxidize alkenes without adding catalyst; however, for the
uncatalyzed reaction, relatively high reaction temperatures and long reaction times
are needed [26]. These drawbacks can be eliminated by adding transition metal
catalyst to the reaction [27].
In the following section, a summary is given based on previously published
results [28, 29], on the construction, some aspects of structural characterization and
SOD-like activity testing of Fe(III)-C-protected uniform or mixed amino acid
complexes covalently anchored onto chloropropylated silica gel, inspired by the
active centre of the FeSOD enzyme, and an account is provided on these features of
newly constructed silica-anchored complexes. Furthermore, the hitherto unpub-
lished results concerning the catalytic activities and selectivities of all these
anchored complexes are also communicated here.
Experimental
Materials and methods of syntheses
For the syntheses, C-protected (in the form of methyl ester) L-histidine, L-tyrosine,
L-cysteine, and L-cystine were used as ligands. The metal ion source was
FeCl3ꢀ6H2O. Chloropropylated silica gel (SG particle size, 230–400 mesh, BET
surface area, 500 m2/g, functionalization, 8 %)] was used as support. These
materials as well as the 2-propanol solvent were products of Aldrich Chemical Co.
All chemicals were of analytical grade and were used without further purification.
The general features of the syntheses are as follows. The first step of
immobilization was the reaction of the appropriately protected amino acid
(1.75 mmol) and the support (0.5 g, containing 0.35 mmol of chlorine atoms).
The C-protected amino acids were covalently grafted onto the support through
N-alkylation like reaction by refluxing the mixture in 2-propanol (60 cm3) under
alkaline conditions. After 24 h, the solid substance was filtered, washed several
times in order to remove the uncoupled amino acid excess, and then dried.
Complexation followed the anchoring: the grafted support was soaked in the
2-propanolic (60 cm3) solution of the metal salt (1.75 mmol) under continuous
stirring at room temperature for 24 h. After filtering and washing, the obtained
material was divided into two parts, and then half of it was set aside. This is what we
call covalent grafting under ligand-poor conditions, i.e., only the immobilized
protected amino acids were available for coordination. To the other half,
2-propanolic (60 cm3) solution of the appropriately protected amino acid derivative
was added in excess (0.875 mmol), and the suspension was continuously stirred at
room temperature for 24 h. Then, the solid material was filtered, rinsed with
2-propanol several times, and dried. The latter was named covalent grafting under
ligand-excess conditions, i.e., the surface–grafted complex might have rearranged in
the presence of excess amino acid mixture.
123

G. Varga et al.
Surface–grafted complexes were prepared having both uniform and mixed amino
acid derivatives (two protected amino acids were used) as ligands. Two methods
were applied for the syntheses when mixed ligands were used. In method ‘A’, one of
the protected amino acid esters was covalently anchored to the surface of the
support; then, it was soaked in the metal salt solution, and after filtering and
thorough washing, the final substance was made by allowing complexation with
excess amounts of the other amino acid ester. In method ‘B’ a 1:1 molar mixture of
the protected amino acids was grafted onto the surface of the support; then, the
metal complex was formed. Parts of the materials thus formed were further treated
in excess 1:1 protected amino acid mixtures resulting in the formation of surface-
anchored complexes under ligand-excess conditions. The other experimental
parameters were the same as described earlier.
The materials used in structural characterization and for catalytic testing together
and their codes used in the followings are listed as follows:
SG–Tyr-OMe–Fe(III)a
SG–Cys-OMe–Fe(III), complex made under ligand-poor conditions
SG–Cys-OMe–Fe(III)–H-Cys-OMe, complex made under ligand-excess conditions
SG–(Cys-OMe)2–Fe(III),
SG–(Cys-OMe)2–Fe(III)M–H-(Cys-OMe)2,
SG–His-OMe–Fe(III)–H-Cys-OMeb, complex made with method ‘A’
SG–Cys-OMe–Fe(III)–H-His-OMeb,
SG–His-OMe;Cys-OMe–Fe(III)b, complex made with method ‘B’, under ligand-
poor conditions
SG–His-OMe;Cys-OMe–Fe(III)–H-His-OMe;H-Cys-OMeb, complex made with
method ‘B’ under ligand-excess conditions
SG–His-OMe–Fe(III)–(H-Cys-OMe)2,
SG–(Cys-OMe)2–Fe(III)–H-His-OMe,
SG–His-OMe;(Cys-OMe)2–Fe(III),
SG–His-OMe;(Cys-OMe)2–Fe(III)–H-His-OMe;(H-Cys-OMe)2.
a, b—Synthesis, structural characterization and SOD-like activity is described in
Refs. [28, 29], respectively.
Analytical measurements
The amount of Fe(III) ions on the surface-modified silica gel was measured by an
Agilent 7,7009 ICP-MS. Before measurements, a few milligrams of the anchored
complexes measured by analytical accuracy were digested in 1 cm3 cc. H2SO4;
then, they were diluted with distilled water to 50 cm3 and filtered. The possibility of
metal ion leaching was also studied by the ICP-MS method with the same
instrument.
The nitrogen content of the samples was determined by the Kjeldahl method (the
composition of the reaction mixture: 5 cm3 cc. H2SO4, 1 cm3 30 % solution of
H2O2, 100 mg of the grafted complexes).
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Fe-amino acid complexes
X-ray absorption spectroscopy (XAS) measurements
The measurements were carried out on the K-edge of the metals at MAX-lab at
beamline I811. This is a superconducting multipole wiggler beamline equipped with
a water-cooled channel cut Si(111) double crystal monochromator delivering at
10 keV, approximately 2 9 1015 photons/s/0.1 % bandwidth with horizontal and
vertical FWHM of
7 and 0.3 mrad, respectively [30]. A beamsize of
0.5 mm 9 1.0 mm (width 9 height) was used. The incident beam intensity (I0)
was measured with an ionization chamber filled with a mixture of He/N2. Higher-
order harmonics were reduced by detuning the second monochromator to 50–70 %
of the maximum intensity, depending on the metal. Data collection was performed
in transmission mode. Samples (*300 mg) were measured in Teflon spacers with
Kapton tape windows.
XAS spectra were normalized to an edge jump of unity and the background
absorption was removed.
The EXAFS data were k3-weighted and Fourier transformed in the range of
-1
˚
˚
k = 2–12 A . The ranges for the backtransform were 1–3 A for all complexes.
The fitted parameters included the amplitude reduction factor (S20), interatomic
distances (R), Debye–Waller factors (r2), and energy shift (DE0). The coordination
numbers (N) were kept constant during each optimization, but a range of
coordination numbers were used to find the best fit.
The main objectives of these measurements were to determine the coordination
numbers, geometries around the Fe(III) ion, and to find out whether the sulphur
atom was coordinated to the central ion in the anchored Fe(III) complexes.
Mid/far range FT-IR spectroscopy
Structural information on each step of the synthesis procedure was obtained by far- and
mid-range infrared spectroscopy. Mid-range spectra were recorded with a BIO-RAD
Digilab Division FTS-65 A/896 FT-IR spectrophotometer with 4 cm-1 resolution,
measuring diffuse reflectance. A total of 256 Scans were collected for each spectrum;
300 mg KBr and 10 mg sample were combined and finely ground. Spectra were
evaluated by the Win-IR package. They were baseline-corrected, smoothed (if it was
necessary), and the spectra of the supports were subtracted. The 3,800–600 cm-1
wavenumber range was investigated. The comparison of the difference mid-IR spectra of
the anchored amino acid derivatives with and without metal ion and the spectra of the
pristine amino acid derivatives gives indirect information on the coordinating groups. The
difference D [D = masym(COO-) - msym(COO-)] between the asymmetric and symmetric
carboxylate vibrations gives information about the coordination mode of the carboxylate
group. The coordination can be either bidentate chelating (Dcomplex \Dligand) or bidentate
bridging (Dcomplex * Dligand) or monodentate (Dcomplex [Dligand) [31]. A shift in the
position of the carbonyl group and the phenolic C–O or the absence of the S–H stretching
vibration indicates the participation of these groups in complexation [32, 33].
Far-range spectra were recorded with a BIO-RAD Digilab Division FTS-40
vacuum FT-IR spectrophotometer with 4 cm-1 resolution. A total of 256 scans were
collected for each spectrum. The Nujol mull technique was used between two
123

G. Varga et al.
polyethylene windows (the suspension of 10 mg sample and a drop of Nujol mull).
Spectra were evaluated by the Win-IR package. They were baseline-corrected and
smoothed (if it was necessary). Unfortunately, in several cases the spectra could not be
used for evaluation. The spectra in the far-IR region provide direct information on
metal ion–functional group coordination, although assignation of the vibrations is not
a trivial exercise. For making it easier, probe complexes having uniform, thus easily
identifiable, coordinating groups were prepared and their far-IR spectra were
registered. Fe(III)-complexes of imidazole, isopropylamine, and monosodium
malonate were prepared. Each probe complex was synthesized via using 2-propanol
(10 cm3) as solvent. The metal salt (4 9 10-4 mol) and the ligand (2.4 9 10-3 mol)
were stirred for 24 h at room temperature to get a solid precipitate. The obtained
materials were filtered and washed with 2-propanol.
Testing the superoxide dismutase activity
The SOD activity was tested by the Beauchamp–Fridovich reaction [34]. A brief
description of this biochemical test reaction is as follows. For this reaction,
riboflavin, L-methionine, and nitro blue tetrazolium were used. Under aerobic
conditions, reaction takes place on illumination between riboflavin and L-
methionine. It is a reduction, and the reduced form of riboflavin reacts with
oxygen forming a peroxide derivative. This derivative decomposes giving the
superoxide radical anion. This radical ion is captured by the nitro blue tetrazolium
(NBT) and its original yellow color turns blue.
The transformation can be followed by Vis spectrophotometry, measuring the
absorbance at 560 nm. If our enzyme-mimicking material works well, it success-
fully competes with NBT in capturing the superoxide radical ion. Thus, the
photoreduction of NBT is inhibited. The SOD probe reaction was carried out at
room temperature in a suspension of the immobilized complex at pH = 7 ensured
with a phosphate or for the Mn(II) complexes, 4-(2-hydroxyethyl)-1-piperazi-
neethanesulfonic acid (HEPES) buffer. The reaction mixture contained 0.1 cm3 of
0.2 mM riboflavin, 0.1 cm3 of 5 mM NBT, 2.8 cm3 of 50 mM buffer, containing
EDTA (0.1 mM), L-methionine (13 mM), and the catalyst. Riboflavin was added
last, and the reaction was initiated by illuminating the tubes with two 15-W
fluorescent lamps. Equilibrium could be reached in 10 min. EDTA masks the
interfering metal ion traces, since the metal ion–EDTA complexes have no SOD
activity. From the resulting graph, the volume of enzyme-mimicking complex
corresponding to 50 % inhibition (IC50) was registered to allow a comparison with
the efficiency of the real enzyme and other SOD mimics. The enzyme mimic works
the better when the IC50 is the smaller. There was no reaction without illumination,
and the support did not display SOD activity either.
Catalytic oxidation of cyclohexene
In the reaction, a vial with septum was loaded with the catalyst (25 mg), acetone
(10 ml), cyclohexene (5 mmol), and 2.5 mmol peracetic acid (*39 % in acetic
123

Fe-amino acid complexes
acid). After 3 h of continuous stirring at room temperature, the mixture was
analyzed quantitatively with a Hewlett-Packard 5,890 Series II gas chromatograph
(GC) using an Agilent HP-1 column and the internal standard technique. The
temperature was increased in stages from 50 to 250 °C. The products were identified
via the use of authentic samples.
Results and discussion
Analytical measurements
The results of the Kjeldahl method and the ICP-MS measurements for the grafted
Fe(III)-complexes containing uniform ligands are displayed in Table 1.
In all cases, the metal ion-to-amino acid ratios indicate (they are never integers)
that mixtures of various kinds of complexes were formed on the surface. The amino
acid content of SG–Tyr-OMe–Fe(III) and SG–Tyr-OMe–Fe(III)–H-Tyr-OMe is
almost the same, suggesting (just as the mid-IR spectra published in Ref. [28]) that
the preparation of the complex under ligand-excess conditions was not successful.
X-ray absorption measurements
Fe K-edge XAS spectra were recorded for some complexes formed with sulphur-
containing ligands. The XANES spectra of the materials are depicted in Fig. 1.
Since the local symmetry around the Fe is higher in octahedral than in tetrahedral
complexes, the intensity of the characteristic pre-edge peak around 7,113 eV
decreases in the following order: Itetrahedral [ Isquare pyramidal [ Ioctahedral [35].
Comparison of the normalized intensity of the pre-edge peak in the anchored
Fe(III)-complexes (I * 0.05) with that of some reference compounds indicates that
the complexes are 5-coordinate; they are square pyramidal [35].
The Fourier-transformed EXAFS data (without phase correction) are displayed in
Fig. 2, and the results of the fitting are shown in Table 2.
Table 1 Amino acid and Fe3? content of the samples obtained from Kjeldahl type N-determination and
ICP-MS measurements
Sample
Amino acid
content (mmol/g)
Fe3? content
(mmol/g)
SG–Tyr-OMe–Fe(III)a
SG–Tyr-OMe–Fe(III)–H-Tyr-OMea
0.572
0.581
0.460
0.654
0.445
0.924
0.380
0.382
0.333
0.333
0.370
0.371
SG–Cys-OMe–Fe(III)
SG–Cys-OMe–Fe(III)–H-Cys-OMe
SG–(Cys-OMe)2–Fe(III)
SG–(Cys-OMe)2–Fe(III)–H-(Cys-OMe)2
a
Their syntheses and mid-IR characterization was published in Ref. [28], but Kjeldahl measurements
were not performed then
123

G. Varga et al.
C
B
A
Main edge position eV
A
B
C
7133.6
7133.2
7133.5
7100
7120
7140
7160
7180
7200
Energy (eV)
Fig. 1 The Fe K-edge XANES spectra of A—SG–(Cys-OMe)2–Fe(III), B—SG–Cys-OMe–Fe(III) and
C—SG–His-OMe–Fe(III)–H-Cys-OMe
C
B
A
0
1
2
3
4
5
Radial distance (Å)
Fig. 2 The Fourier-transformed EXAFS data (without phase correction) of A—SG–(Cys-OMe)2–Fe(III),
B—SG–Cys-OMe–Fe(III) and C—SG–His-OMe–Fe(III)–H-Cys-OMe, red line—fit, black line—
experimental. (Color figure online)
In the first coordination shell of SG–(Cys-OMe)2–Fe(III), there are five oxygen/
˚
nitrogen atoms, with a Fe(III)–O/N bond length of 1.98 A. For SG–Cys-OMe–
Fe(III), the first coordination sphere contains four O/N atoms and one sulphur atom,
˚
where the bond distances were fitted to be 1.98 and 2.36 A, respectively. In SG–
123

Fe-amino acid complexes
Table 2 Parameters deduced from the fitted EXAFS spectra
2
(Fe3?–)X
N
r2 (A )
DE0 (eV)
R factor
˚
R (A)
˚
Sample
SG–(Cys-OMe)2–Fe(III)
SG–Cys-OMe–Fe(III)
O/N
O/N
S
5
1.98
1.98
2.36
1.98
2.36
0.0080
0.0120
0.0327
0.0114
0.0232
-1.44
-5.37
0.0179
0.0135
4
1
SG–His-OMe–Fe(III)–H-Cys-OMe
O/N
S
3.6
1.4
-5.69
0.0120
N—coordination number, R—bond length, r2—Debye–Waller factor, DE0—energy shift, R factor—
goodness of fit
˚
His-OMe–Fe(III)–H-Cys-OMe, the Fe(III)–O/N distance is 1.98 A and the average
˚
coordination number is 3.6, while for Fe(III)–S, these date are 2.36 A and 1.4,
respectively. The thiolate sulphur donor atom is coordinated to the Fe(III)
irrespective to whether the cysteine is anchored or non-anchored on the surface
of the support.
Mid/far-range FT-IR spectroscopy
As far as SG–Cys-OMe–Fe(III) is concerned (Fig. 3, trace A), the carboxylate group
is most probably coordinated as monodentate ligand, since D increased from 190 to
216 cm-1 (1,602–1,386 cm-1). The XAS measurement revealed the coordination of
1734
1745
D
1625
1390
C
1745
B
A
1602
1600
1386
1400
1800
1200
1000
800
600
-1
Wavenumber (cm )
Fig. 3 The difference IR spectra of A—SG–Cys-OMe–Fe(III), B—SG–Cys-OMe–Fe(III)–H-Cys-OMe,
C—SG–(Cys-OMe)2–Fe(III) and D—SG–(Cys-OMe)2–Fe(III)–(H-Cys-OMe)2 (the spectrum of the
support was subtracted)
123

G. Varga et al.
the thiolate sulphur, and the lack of the S–H vibration further confirms that. Under
ligand-excess conditions (Fig. 3, trace B), the carbonyl oxygen is not involved in the
coordination, since the stretching vibration of the carbonyl band did not shift to
lower wavenumbers (1,745 cm-1). There is no S–H vibration at around
2,500 cm-1; therefore, the thiolate sulphur is coordinated.
For the SG–(Cys-OMe)2–Fe(III), D = 1,625 - 1,390 cm-1 = 235 cm-1 sug-
gesting the monodentate ligation of the carboxylate groups (Fig. 3, trace C). Under
ligand-excess conditions (Fig. 3, trace D), the carbonyl oxygens do not take part in
the complexation, since the positions of the bands (1,745, 1,734 cm-1) do not move
relative to the pristine amino acid.
In the far-IR range (Fig. 4, trace A), the bands at 383 and 294 cm-1 correspond
to the Fe(III)–Namino vibrations.
Using all these pieces of information structural proposals for the surface–bound
complexes are offered as follows:
SG–Cys-OMe–Fe(III)
SthiolateCyssurfOcarboxylateCysssurfH2O; H2O; H2O;
SG–Cys-OMe–Fe(III)–H-Cys-OMe
SthiolateCyssurfOcarboxylateCysssurfSthiolateCysSthiolateCysH2O;
SG–(Cys-OMe)2–Fe(III)
OcarboxylateðCysÞ2surfOcarboxylateðCysÞ2surfH2O; H2O; H2O;
SG–(Cys-OMe)2–Fe(III)–(H-Cys-OMe)2
294
312
A
383
366
350
500
450
400
300
250
200
-1
Wavenumber (cm )
Fig. 4 The difference far-IR spectra of A—SG–(Cys-OMe)2–Fe(III)–(H-Cys-OMe)2 (the spectrum of the
support was subtracted)
123

Fe-amino acid complexes
OcarboxylateðCysÞ2surfOcarboxylateðCysÞ2surfNaminoðCysÞ2NaminoðCysÞ2H2O:
SG–His-OMe–Fe(III) was rearranged by adding C-protected cystine in excess
(Fig. 5, trace A). The stretching vibration of the carbonyl band (1,739 cm-1) did not
move relative to the free C-protected cysteine; therefore, it can only coordinate with
the amino nitrogens.
In Fig. 5, trace B depicts the spectrum of SG–(Cys-OMe)2–Fe(III)–H-His-OMe.
The spectrum is very similar to that of the C-protected histidine—the carbonyl
oxygen does not participate in the complexation.
In SG–His-OMe;(Cys-OMe)2–Fe(III), the anchored amino acids are again
assumed to coordinate as they did alone (Fig. 5, trace C).
Under ligand-excess conditions (Fig. 5, trace D), three unshifted carbonyl
vibrations can be seen at 1,760 (C-protected histidine), 1,746 and 1,734 cm-1 (C-
protected cystine).
Unfortunately, their far-IR spectra are unusable, but on the basis of these
observations and the accumulated knowledge described at the complexes formed
with uniform ligands, the following structures may be proposed:
SG–His-OMe–Fe(III)–(H-Cys-OMe)2
NimidazoleHissurfOcarboxylateHissurf
N
aminoðCysÞ2NaminoðCysÞ2H2O;
SG–(Cys-OMe)2–Fe(III)–H-His-OMe
OcarboxylateðCysÞ2surfOcarboxylateðCysÞ2surfNimidazoleHisNimidazoleHisH2O;
SG–His-OMe;(Cys-OMe)2–Fe(III)
1746
1760
1734
D
1632
1392
C
1760
B
1739
A
1800
1600
1400
1200
1000
800
600
-1
Wavenumber (cm )
Fig. 5 The difference IR spectra of A—SG–His-OMe–Fe(III)–(H-Cys-OMe)2, B—SG–(Cys-OMe)2–
Fe(III)–H-His-OMe, C—SG–His-OMe;(Cys-OMe)2–Fe(III), D—SG–His-OMe;(Cys-OMe)2–Fe(III)–H-
His-OMe;(H-Cys-OMe)2 (the spectrum of the support was subtracted)
123

G. Varga et al.
NimidazoleHissurfOcarboxylateHissurf
O
carboxylateðCysÞ2surfOcarboxylateðCysÞ2surfH2O;
SG–His-OMe;(Cys-OMe)2–Fe(III)–H-His-OMe;(H-Cys-OMe)2
NimidazoleHissurfOcarboxylateHissurfOcarboxylateðCysÞ2surfNimidazoleHisNaminoðCysÞ2
:
Superoxide dismutase activity of the complexes
All materials were active in catalyzing the dismutation reaction of the superoxide
radical anion. Catalytic activities differed widely though (Table 3).
Data reveal that there were catalysts with activity close to that of the Cu,ZnSOD
enzyme. The most active substances contain cysteine, indicating the key role of
cysteine-like structures in determining catalytic activity. They seem to have the
optimum structures for promoting this reaction. SG-Tyr-OMe-Fe(III) was the
second most active material, in which tyrosine coordinates as tridentate ligand,
making the complex more strained.
Catalytic oxidation of cyclohexene
The catalytic oxidation of cyclohexene takes place according to Scheme 1 [36].
After performing numerous optimization experiments, we found that for our
supported Fe(III) complexes, 3 h reaction time and 2.5 mmol peracetic acid were
needed to obtain the highest epoxide selectivity, and the reactions must have been
performed in acetone to avoid the decomposition of the oxidant.
Table 3 The SOD activities of
the surface-grafted complexes
Materials
IC50
(lM)
Cu,ZnSOD enzyme
SG–Tyr-OMe–Fe(III)a
0.4
5
SG–Cys-OMe–Fe(III)
6
SG–Cys-OMe–Fe(III)–H-Cys-OMe
SG–(Cys-OMe)2–Fe(III)
31
36
33
28
25
21
4
SG–(Cys-OMe)2–Fe(III)–H-(Cys-OMe)2
SG–His-OMe–Fe(III)–H-Cys-OMeb
SG–Cys-OMe–Fe(III)–H-His-OMeb
SG–His-OMe;Cys-OMe–Fe(III)b
SG–His-OMe;Cys-OMe–Fe(III)–H-His-OMe;H-Cys-
OMeb
SG–His-OMe–Fe(III)–(H-Cys-OMe)2
SG–(Cys-OMe)2–Fe(III)–H-His-OMe
SG–His-OMe;(Cys-OMe)2–Fe(III)
24
32
31
a,b
Data published in Refs. [28,
29], respectively, are shown for
comparison
SG–His-OMe;(Cys-OMe)2–Fe(III)–H-His-OMe;(H-Cys- 28
OMe)2
123

Fe-amino acid complexes
Scheme 1 The oxidative transformations of cyclohexene
Table 4 The conversion and selectivity results of the oxidation of cyclohexene after 3 h; the best
catalysts are highlighted in italics
Catalyst
Conversion Epoxide Alcohol Ketone Diol
(%)
(%)
(%)
(%)
(%)
–
21
45
59
56
25
10
44
65
49
32
64
94
87
99
97
95
88
85
98
97
4
2
3
1
1
2
2
2
1
1
2
1
8
0
1
1
0
9
0
1
30
3
SG–Tyr-OMe–Fe(III)
SG–Cys-OMe–Fe(III)
2
SG–Cys-OMe–Fe(III)–H-Cys-OMe
SG–(Cys-OMe)2–Fe(III)
0
1
SG–(Cys-OMe)2–Fe(III)–H-(Cys-OMe)2
SG–His-OMe–Fe(III)–H-Cys-OMe
SG–Cys-OMe–Fe(III)–H-His-OMe
SG–His-OMe;Cys-OMe–Fe(III)
2
10
4
1
SG–His-OMe;Cys-OMe–Fe(III)–H-His-OMe;H-
Cys-OMe
1
Initially, for activity (and selectivity) testing, three selection criteria were
applied. First, those complexes were chosen, which displayed high activities in the
biochemical reaction; second, their ‘‘pairs’’ (syntheses under ligand-poor—ligand
excess conditions) were included (if they existed); and third, complexes constructed
by all varieties must be represented.
The activity and selectivity data in Table 4 attest that all the chosen immobilized
complexes were catalytically active, and that the major product of the reactions was
the epoxide. It is to be seen that in many cases, the conversion of catalyzed reaction
was significantly higher than that of the stoichiometric. The cystine-containing
complexes only had lower conversions than the stoichiometric reaction; therefore,
all the other cystine-containing complexes were excluded from the tested catalyst
group. The selectivities, which were very significantly higher for the catalyzed
reaction than for the stoichiometric one, virtually did not depend on the identity of
the amino acid ligand. The presence of the ligands was necessary though, since
silica gel impregnated with the Fe(III) ions only decomposed the peracetic acid.
Concerning epoxide selectivity and catalytic activity, the best catalyst is SG–
Cys-OMe–Fe(III)–H-Cys-OMe with 99 and 56 % selectivity and conversion,
respectively. In general, cystine-containing materials did not catalyze this reaction
well (but the epoxide selectivities are still high), moreover, for SG–(Cys-OMe)2–
123

G. Varga et al.
Fe(III)–H-(Cys-OMe)2 the observed conversion was lower than that of the
homogeneous, uncatalyzed reaction.
Leaching of the metal ion was not observed, and the catalysts, highlighted in
italics in Table 4, could be reused twice just after rinsing it with the solvent.
The mechanism of the reaction is suggested as follows: one of the coordinated
water molecules is replaced by the peroxidic oxygen donor oxidant, forming
(hydro)peroxo-metal species, then, the O–O bond is cleaved heterolytically to form
high-valent metal-oxo species as an active intermediate, which is responsible for the
epoxidation of the uncoordinated cyclohexene. If both reactants were coordinated,
there would be plenty of time for further reactions. If cyclohexene was coordinated
alone, the situation would not be much different from the stoichiometric reaction.
Thus, probably, the role of the ligands is to exert steric influence on the accessibility
of the central ion by the reactants.
Conclusions
All the SOD-mimicking surface-anchored Fe(III)–amino acid complexes were
successfully constructed. It was possible to prepare the covalently anchored
complexes with uniform and mixed ligands as well. Covalent anchoring gave good
control over the mode of immobilization.
Analytical measurements revealed that mixtures of complexes with 1:1, 1:2, and
1:3 metal ion-to-ligand ratios were formed on the surface of the support. The
coordination numbers and coordinating sites could be identified with the combi-
nation of XAS measurements, mid- and far-IR spectroscopies and chemical
considerations. It was proven that the Fe(III) complexes were square pyramidal. The
major coordinating sites were proposed to be the carboxylate oxygen, the imidazole
nitrogen, the phenolate oxygen and sulphur atom of the thiolate group. The other
coordination sites depended on the conditions of the synthesis and the structures of
the molecules. In all cases, water molecules saturated the coordination sphere. In
most cases under ligand–excess conditions, the surface–anchored ligand–poor
complexes were rearranged.
All the covalently anchored complexes were active in a SOD test reaction: they
could catalyze the dismutation reaction of the superoxide radical anion. The
activity, in some cases, was only one magnitude lower that of the native Cu,ZnSOD
enzyme.
The complexes displayed catalytic activity in the oxidation of cyclohexene, and
all of them were extremely selective to cyclohexene oxide formation. Some of the
best catalysts were reused twice without significant loss in the catalytic activity and
selectivity. The activities were, but the selectivities were basically not dependent on
the coordinating groups.
Immobilization of transition metal–amino acid complexes turned out to be a
viable route for preparing efficient electron transfer catalysts, since they are very
active and selective catalysts that can be easily recovered and recycled. They show
the promise of becoming efficient catalysts in the synthesis of fine chemicals.
123

Fe-amino acid complexes
´
Acknowledgments This research was financed by the TAMOP 4.2.2.A-11/1/KONV-2012-0047 and the
OTKA 83889 Grants. The support is highly appreciated.
References
1. K.M. Koeller, C.H. Wong, Nature 409, 232 (2001)
2. U. Hanefeld, L. Gardossi, E. Magner, Chem. Soc. Rev. 38, 453 (2009)
3. U.T. Bornscheuer, Angew. Chem. Int. Ed. 42, 3336 (2003)
4. D.J. Xuereb, R. Raja, Catal. Sci. Technol. 1, 517 (2011)
5. J.A. Labinger, J. Mol. Catal. A Chem. 220, 27 (2004)
¨
6. M. Luechinger, A. Kienhofer, G.D. Pirngruber, Chem. Mater. 18, 1330 (2006)
7. K. Suzuki, P.D. Oldenburg, L. Que Jr, Angew. Chem. Int. Ed. 47, 1887 (2008)
8. B.M. Weckhuysen, J. Am. Chem. Soc. 128, 3208 (2006)
9. A.-F. Miller, in Handbook of Metalloproteins, ed. by A. Messerschmidt, R. Huber, K. Wieghardt, T.
Poulos (Wiley, Chichester, 2001), pp. 668–682
10. M.L. Ludwig, A.L. Metzger, K.A. Pattridge, W.C. Stallings, J. Mol. Biol. 219, 335 (1991)
11. M.S. Lath, M.M. Dixon, K.A. Pattridge, W.C. Stallings, J.A. Fee, M.L. Ludwig, Biochem. 34, 1646
(1995)
12. F. Farzaneh, S. Sohrabi, M. Ghiasi, M. Ghandi, V. Mehdi Ghandi, J. Porous Mater. 20, 267 (2013)
´
13. M. Halma, K.A.D. de Freitas Castro, C. Taviot-Gueho, V. Prevot, C. Forano, F. Wypych, S.
Nakagaki, J. Catal. 257, 233 (2008)
14. A. Baso, L.D. Martin, C. Ebert, L. Gardossi, P. Linda, F. Sibilla, Tetrahedron Lett. 44, 5889 (2003)
15. H.H.P. Yiu, P.A. Wright, J. Mater. Chem. 15, 3690 (2005)
16. C. Ispas, I. Sokolov, S. Andreescu, Anal. Bioanal. Chem. 393, 543 (2009)
17. G.D. Pirngruber, L. Frunz, M. Lu¨chinger, Phys. Chem. Chem. Phys. 11, 2928 (2009)
18. Y. Zhang, J. Zhao, L. He, D. Zhao, S. Zhang, Mic. Mes. Mater. 94, 159 (2006)
19. J. Gao, Y. Chen, B. Han, Z. Feng, C. Li, N. Zhou, Z. Gao, J. Mol. Catal., A 210, 197 (2004)
20. S. Samantaray, K. Parida, Catal. Commun. 6, 578 (2005)
21. S. Bhattacharjee, J.A. Anderson, J. Mol. Catal. A 249, 103 (2006)
´
22. C. Marchi-Delapierre, A. Jorge-Robin, A. Thibon, S. Menage, Chem. Commun. 1166 (2007)
23. J. Jiang, K. Ma, Y. Zheng, S. Cai, R. Li, J. Ma, Appl. Clay Sci. 45, 117 (2009)
24. R. Noyori, M. Aoki, K. Sato, Chem. Commun. 1977 (2003)
25. T. Punniyamurthy, L. Rout, Coord. Chem. Rev. 252, 134 (2008)
26. H. Shi, Z. Zhang, Y. Wang, J. Mol. Catal. A 238, 13 (2005)
27. K.-P. Ho, W.-L. Wong, K.-M. Lam, C.-P. Lai, T.H. Chan, K.-Y. Wong, Chem. Eur. J. 14, 7988
(2008)
´
28. Z. Csendes, Cs. Dudas, G. Varga, E.G. Bajnoczi, S.E. Canton, P. Sipos, I. Palinko, J. Mol. Struct.
´
´
´
´
1044, 39 (2013)
´
´
¨
29. Z. Csendes, N. Foldi, J.T. Kiss, P. Sipos, I. Palinko, J. Mol. Struct. 993, 203 (2011)
30. S. Carlson, M. Clausen, L. Gridneva, B. Sommarin, C.J. Svensson, J. Synchrotron Radiat. 13, 359
(2006)
31. S.K. Papageorgiou, E.P. Kouvelos, E.P. Favvas, A.A. Sapadilis, G.E. Romanos, F.K. Katsaros,
Carbohydr. Res. 345, 469 (2010)
32. S.A. Abdel-Latif, H.B. Hassib, Y.M. Issa, Spectrochim Acta Part A 67, 950 (2007)
33. T. Miura, T. Satoh, H. Takeuchi, Biochim. Biophys. Acta 1384, 171 (1998)
34. C. Beauchamp, I. Fridovich, Anal. Biochem. 44, 276 (1971)
35. P. Chutia, S. Kato, T. Kojima, S. Satokawa, Polyhedron 28, 370 (2009)
36. G. Bilis, K.C. Christoforidis, Y. Deligiannakis, M. Louloudi, Catal. Today 157, 101 (2010)
123