Bio-inspired amino acid oxidation by a non-heme iron catalyst

Article abstract of DOI:10.1016/j.jinorgbio.2013.02.007

This study reports the kinetics and mechanism of Fe(III)-catalyzed oxidative decarboxylation and deamination of a series of acyclic (α-aminoisobutyric acid, α-(methylamino)isobutyric acid, alanine, norvaline, and 2-aminobutyric acid) and cyclic (1-aminocyclopropane-1-carboxylic acid, 1-amino-1-cyclobutanecarboxylic acid, 1-aminocyclopentanecarboxylic acid, and 1-aminocyclohexanecarboxylicacid) amino acids using hydrogen peroxide, t-butyl hydroperoxide, iodosylbenzene, m-chloroperbenzoic acid, and peroxomonosulphate as oxidant in 75% DMF-25% water solvent mixture. Model complex [Fe^IVO(SALEN)]^?+ (SALENH₂: N,N′-bis(salicylidene)ethylenediamine) was generated by the reaction of Fe^III(SALEN)Cl and H₂O₂ in CH₃CN at 278 K as reported earlier. This method provided us high-valent oxoiron species, stable enough to ensure the direct observation of the reaction with amino acids.

Full text of DOI:10.1016/j.jinorgbio.2013.02.007

Journal of Inorganic Biochemistry 123 (2013) 46–52
Contents lists available at SciVerse ScienceDirect
Journal of Inorganic Biochemistry
journal homepage: www.elsevier.com/locate/jinorgbio
Bio-inspired amino acid oxidation by a non-heme iron catalyst
a
a
a
b
a
a,
Szabina Góger , Dóra Bogáth , Gábor Baráth , A. Jalila Simaan , Gábor Speier , József Kaizer ⁎
a
Department of Chemistry, University of Pannonia, 8201 Veszprém, Hungary
Aix-Marseille Université, FR1739, Spectropole, Campus St. Jérôme, Avenue Escadrille Normandie-Niemen, 13397 Marseille Cedex 20, France
b
a r t i c l e i n f o
a b s t r a c t
Article history:
This study reports the kinetics and mechanism of Fe(III)-catalyzed oxidative decarboxylation and deamina-
Received 4 December 2012
Received in revised form 11 February 2013
Accepted 11 February 2013
Available online 26 February 2013
tion of a series of acyclic (α-aminoisobutyric acid, α-(methylamino)isobutyric acid, alanine, norvaline, and
2-aminobutyric acid) and cyclic (1-aminocyclopropane-1-carboxylic acid, 1-amino-1-cyclobutanecarboxylic
acid, 1-aminocyclopentanecarboxylic acid, and 1-aminocyclohexanecarboxylicacid) amino acids using hydrogen
peroxide, t-butyl hydroperoxide, iodosylbenzene, m-chloroperbenzoic acid, and peroxomonosulphate as oxidant
IV
•+
in 75% DMF–25% water solvent mixture. Model complex [Fe O(SALEN)] (SALENH
2
: N,N′-bis(salicylidene)
Keywords:
Iron
III
ethylenediamine) was generated by the reaction of Fe (SALEN)Cl and H
2
O
2
3
in CH CN at 278 K as reported
Salen
earlier. This method provided us high-valent oxoiron species, stable enough to ensure the direct observation
of the reaction with amino acids.
Amino acids
Oxidation
ACC oxidase
Catalysis
© 2013 Elsevier Inc. All rights reserved.
1
. Introduction
oxoiron(IV) intermediate has been trapped in the reaction of
2-oxoglutarate-dependent dioxygenases (TauD and P4H) with dioxygen
Model chemistry in relevance to metalloenzymes has progressed
in the presence of substrate and cofactor [15,16]. In addition several syn-
thetic oxoiron(IV) species were successfully prepared from biomimetic
non-heme model complexes [17–26]. Pirrung et al. have determined
ACC amine radical cation to be a key intermediate in the ACCO active
site. A precedent for this kind of radical generation was established
in earlier studies on the inactivation of cytochrome P450 (oxoiron)
enzymes by cyclopropylamines [27]. They have proposed two types of
reactions from the key intermediate: (1) direct ring opening of ACC
amine radical cation followed by proton abstraction from the amino
group (Path A in Scheme 2) and (2) proton abstraction from the amino
group, followed by ring opening of ACC aminyl radical (Path B in
Scheme 2) [28,29]. The stability and reactivity of ACC amine radical
cation were also examined by using quantum chemical calculations. It
was quantitatively demonstrated, that reaction (1) was more favorable
than reaction (2). Therefore, ethylene synthesis should proceed predom-
inantly through reaction (1) [30].
remarkably in recent years and contributed greatly to clarification of
structure and mechanism of various enzymes. The other important
contribution of the model systems to progress in enzymatic studies
is to obtain information about structures and reactivities of substrate–
metal intermediates and investigate how the model chemistry is related
to the development of efficient catalysis by metal complexes. In nature
ethylene is an important phytohormone, which regulates many physio-
logical processes in plant growth and development such as fruit ripen-
ing [1,2]. Many efforts have been made to assay the ethylene forming
enzyme (EFE) and to determine the genes encoding it. EFE is now called
1
-aminocyclopropane-1-carboxylic acid oxidase (ACCO) [3–8]. ACCO
is a dioxygen-activating ascorbate-dependent nonheme iron enzyme,
which catalyzes the two electron oxidation of 1-aminocyclopropane-
1
2
-carboxylic acid (ACCH) to ethylene, CO , and HCN (Scheme 1). The
structure of ACCO from Petunia hybrida has recently been reported [9,10].
It confirms, that the active site contains a single Fe(II) ion coordinat-
ed to three residues (two histidines and one aspartate), referred to as a
Known alternate substrates for the ACCO include 2-alkyl ACCH ana-
logs and α-aminoisobutyric acid (AIBH) (Scheme 3). The former is
processed to the corresponding 1-alkenes, whereas the latter is converted
“
2-His-1-carboxylate facial triad” within the family of nonheme iron
oxidases/oxygenases [11–14]. The mechanism of α-ketoglutarate-
dependent enzymes has been studied extensively and it is accepted,
that dioxygen activation at the iron center is linked to oxidative
decarboxylation of the coligand, forming a high-valent oxoiron(IV)
species as the reactive intermediate. Recently, a mononuclear
2
to acetone imine and CO [28]. 1-amino-1-cyclobutanecarboxylic acid
(ACBH), which has slightly lower ring strain than ACCH, was also used
as a possible ACCH analog. At least three reaction pathways are available
to the radical cations derived from this amino acid, namely decarboxyl-
ation (Path A in Scheme 3), hydroxyl rebound (Path B in Scheme 3) and
ring opening path (Path C in Scheme 3). It was found, that the process-
ing of ACBH by the EFE proceeds via ring expansion to produce
dehydroproline. This result is consistent with a one-electron oxidation
mechanism. The absence of hydroxylated intermediates refutes the
⁎
Corresponding author. Tel.: +36 303098298; fax: +36 88624469.
E-mail address: kaizer@almos.vein.hu (J. Kaizer).
0162-0134/$ – see front matter © 2013 Elsevier Inc. All rights reserved.
http://dx.doi.org/10.1016/j.jinorgbio.2013.02.007

S. Góger et al. / Journal of Inorganic Biochemistry 123 (2013) 46–52
47
Scheme 1. Reaction catalyzed by ACC oxidase.
proposed N-hydroxylation mechanism previously suggested for ethyl-
ene biosynthesis [29].
There are only few reported functional models of ACCO, and even
less using iron [31]. Very recently we have demonstrated that the
III
complex Fe (SALEN)Cl highly selectively and efficiently catalyzes
the oxidation of ACCH and AIBH to ethylene and acetone, respectively
and thus serves as one of the first functional models for ACCO [32].
This study reports the first kinetic data obtained by direct observation
III
of the stoichiometric and catalytic reactions of Fe (SALEN)Cl with a
series of acyclic (α-aminoisobutyric acid (AIBH), α-(methylamino)
isobutyric acid (N-Me-AIBH), alanine (ALAH), norvaline (NORH), and
2
1
-aminobutyric acid (ABH)) and cyclic (1-aminocyclopropane-
-carboxylic acid (ACCH), 1-amino-1-cyclobutanecarboxylic acid
Scheme 3. Proposed mechanism for the oxidation of cyclic and acyclic substrates by
ACC oxidase.
(
ACBH), 1-aminocyclopentanecarboxylic acid (ACPH), and 1-
aminocyclohexanecarboxylicacid (ACHH)) amino acids using hydro-
gen peroxide (H ), t-butyl hydroperoxide (TBHP), iodosylbenzene
PhIO), m-chloroperbenzoic acid (MCPBA), and peroxomonosulphate
2.2. Analytical and physical measurements
2 2
O
(
(
Infrared spectra were recorded on an Avatar 330 FT-IR Thermo Nicolet
instrument. UV–vis spectra were recorded on an Agilent 8453 diode-
array spectrophotometer using quartz cells. GC analyses were performed
on a Hewlett Packard 5890 gas chromatograph equipped with a flame
ionization detector and a 30 m Supelcowax column. Microanalyses
were done by the Microanalytical Service of the University of Pannonia.
IV
•+
PMS) as oxidants. A synthetic model complex, [Fe O(SALEN)] was
III
generated by the reaction of Fe (SALEN)Cl and H
2 2 3
O in CH CN at
2
78 K, as reported earlier [33,34]. This preparation provided us
high-valent oxoiron species stable enough to ensure the direct
observation of the reaction with amino acids. In contrast to the for-
mation of high-valent oxo species with H
III
2
O
2
, [Fe (SALEN)(OIPh)]
and iron(III)-phenoxyl radical are formed with PhIO and MCPBA,
2.3. Determination of products
respectively [35–37].
Reactivity assays were performed as follows: a respective amino acid
(ABH, ACCH, ACBH, ACHH, ACPH, AIBH, N-Me-AIBH, ALAH, NORH) was
2
. Experimental
dissolved in 10 mL of DMF/H
(3/1) in a sealable tube of 20 mL. With MeCN (10 μL) as inner standard,
NH OH and the catalyst were then added to the mixture. Hydrogen
2
O (DMF: N,N-dimethylformamide) mixture
2
.1. Materials
4
All manipulations were performed under pure dinitrogen or argon
atmosphere unless otherwise stated, using standard Schlenk-type
peroxide was added through the septum with a syringe and the
evolved acetone, acetaldehyde, methyl-ethylketone cyclobutanone,
cyclopentanone, cyclohexanone or ethylene was measured by removing
0.25 mL of the headspace with a gastight syringe and the sample
was injected into a gas chromatograph. The concentration of the corre-
sponding product in the headspace is linearly proportional to the con-
centration of the product in the reaction mixture (Fig. S1–S9). In case
inert-gas techniques. Solvents used for the reactions were purified
III
by literature methods and stored under argon. Fe (SALEN)Cl was
prepared according to the literature [38]. All other chemicals were
commercial products and were used as received without further
purification.
Scheme 2. Possible mechanisms for oxidation of ACCH by ACC oxidase.

48
S. Góger et al. / Journal of Inorganic Biochemistry 123 (2013) 46–52
Scheme 4. Reactions catalyzed by FeIII(SALEN)Cl.
of ACBH two other peaks were observed in the chromatogram in addi-
tion to the peak of cyclobutanone. In order to investigate the nature of
these species, ACBH was oxidized also with KMnO
determined [28]. Concentrated NH OH (0.540 mL; 25%) and 89.7 mg
of KMnO were added to an aqueous solution (10 mL) of ACBH
41.5 mg) in a sealable tube of 20 mL. The reaction was monitored
4
and products were
4
4
(
Fig. 1. Conversion and TOF values for amino acid oxidations (substrate (reaction time in
first with a gas chromatograph by removing 0.25 mL of the headspace
minutes): ACCH (100), ACBH (110), ACPH (35), ACHH (25), AIBH (25), ABH (60), NORH
−
2
with a gastight syringe. The observed peak was identical with one of
(60), N-Me-AIB (100), ALAH (120)) in DMF/water (3:1) at 35 °C. [S]
0
= 3.6 × 10 M,
III
III
−6
−2
−2
[
Fe (SALEN)Cl]
0
= 7.2 × 10 M, [H
2
O
2
]
0
= 3.6 × 10 M, [NH
4
OH]
0
= 3.6 × 10 M.
the products in the oxidation reaction of ACBH using Fe (SALEN)Cl as
TOF: the molecules of substrate converted per unit of catalyst per unit of time.
2 2
catalyst and H O as oxidant in the presence of base (RT = 0.935, RT:
retention time; the elapsed time from injection until the
highest-concentration part of the peak has eluted from the column).
The reaction was monitored then by TLC (TLC: thin layer chromatogra-
phy) using n-butanol:glacial acetic acid:water (4:1:1) as a solvent sys-
tem and ninhydrin as a staining reagent. TLC displayed a brown spot
below the starting material ACBH, which was a pink spot. The brown
II
III
metal ions, e.g. Mn, Co, Fe /Fe , Pd, Cu and Ni. These metal complexes
are active and selective in the oxidation of alkanes, alkenes, phenols, al-
cohols, organic sulfides and sulfoxides [33,34,38]. As a functional model
III
of the ACCO we have found, that Fe (SALEN)Cl (1) catalyzes the oxida-
tion of ACCH and AIBH with various oxidants such as H
MCPBA, and PMS in DMF/H O (3:1) solution at 35 °C (Scheme 4).
2 2
O , TBHP, PhIO,
1
spot corresponded to Δ -pyrroline-2-carboxylic acid when compared
2
with authentic TLC data. The liberated CO
tional limewater test. Ammonia – formed in the reaction – was deter-
2
was identified by the conven-
Amino acids are inert against oxidants – either in catalytic and autoxida-
tion reactions – hence reactions were carried out in the presence of a cer-
mined according to the micro Kjeldahl procedure.
4 4
tain base (NH OH or NBu OH). This allows substrates to react in their
more active anionic form. The solution was placed in a hermetically
sealed tube at 35 °C, and then the appropriate oxidant was injected
through the septum. The evolved ethylene and acetone were measured
by removing 0.25 mL of the headspace with a gastight syringe and
injecting the sample into a gas chromatograph. Reactions were selective.
Ethylene and acetone were formed as main reaction products. The
results obtained are given in Table 1.
2
.4. Kinetic measurements
Kinetic study on the oxidation of amino acids by Fe (SALEN)Cl in
III
acetonitrile is performed under pseudo first order conditions. The
progress of reaction is measured following the decay of absorbance
of [Fe O(SALEN)] intermediate with time at appropriate wavelength
IV
•+
III
[34]. The concentrations of [Fe (SALEN)Cl], amino acid tetrabutyl-
In general, the highest catalytic efficiency was obtained with H
2
O
2
,
,
−
4
−3
IV
•+
ammonium salt and hydrogen peroxide are 2 × 10
and 5 × 10
wise stated. The proposed active intermediate was generated first
by addition of hydrogen peroxide. Substrate was then added to the
mixture and spectral changes were recorded.
M, 2 × 10
M
where it is believed that the active oxidant is [Fe O(SALEN)]
−
3
III
M, respectively, temperature is 278.15 K, unless other-
which can be drawn by oxygen transfer between the Fe (SALEN)Cl
and H [33]. Since ACCO catalysis involves several substrates, we
2 2
O
have extended our study to various acyclic and cyclic amino acids.
The optimum conditions for amino acid oxidation by this catalytic
system were catalyst, oxidant, base and substrate in a molar ratio of
1:5000:5000:5000, respectively (Fig. 1). Where in the case of cyclic
amino acids base is needed, in the case of acyclic amino acids the cor-
3
. Results and discussion
3
.1. Catalytic oxidations
responding product was formed without the addition of NH
induction period was observed in the latter case [32], which can be
explained by the in situ formation of NH OH. In the presence of equi-
4
OH. An
The SALENH
2
is a tetradentate ligand with two oxygen and two nitro-
4
gen atoms and can form stable complexes with a variety of transition
molar amount of base (S/base ratio is 1 to 1) there was no induction
Table 1
= 3.6 × 10− M, [FeIII(SALEN)Cl]
2
= 7.2 × 10−6 M, [H
O
2
]
=
Comparison of product formation in the oxidation of amino acids (ACCH, AIBH) in DMF/water (3:1) at 35 °C. [S]
0
0
2
0
−
2
−2
3.6 × 10
M, [NH
4
OH]
0
= 3.6 × 10
M.
2
H O
2
PhIO
TBHP
MCPBA
PMS
a
ACCH
AIBH
TOF (catalyzed) [1/h]
565 ± 56
1 ± 0.1
9273 ± 349
460 ± 59
310 ± 22
190 ± 14
3 ± 0.2
–
734 ± 65
522 ± 41
1328 ± 36
288 ± 19
578 ±
353 ±
1508 ± 48
378 ± 23
6
3
a
TOF (uncatalyzed) [1/h]
a
TOF (catalyzed) [1/h]
27 ±
7 ±
3
1
61 ±
33 ±
5
2
a
TOF (uncatalyzed) [1/h]
a
Turnover frequency: molecules of substrate converted per unit of catalyst per unit of time.

S. Góger et al. / Journal of Inorganic Biochemistry 123 (2013) 46–52
49
Fig. 2. Correlation between substrate concentration and the rate of the reaction of ACCH in
Fig. 3. Hydrogen peroxide dependence of amino acid oxidation reactions in DMF/water (3:1)
III
−6
−2
DMF/water (3:1) at 35 °C. [Fe (SALEN)Cl]
0
= 7.2 × 10 M, [H
O ]
2 2 0
= 3.6 × 10 M,
−2
III
−6
at 35 °C for cyclic substrates. [S]
0
0
= 3.6 × 10 M, [Fe (SALEN)Cl] = 7.2 × 10 M,
= 3.6 × 10− M.
2
−2
[
NH
4
OH]
0
[
NH OH] = 3.6 × 10 M. ■ ACCH, × ACBH, ● ACPH, ▼ ACHH.
4 0
period visible. The oxidation of ACCH occurs with moderate conversion
through Michaelis–Menten kinetics, i.e. saturation kinetics is observed
with respect to the substrate. Similar saturation kinetics was found with
other acyclic and cyclic amino acids too, establishing, that the reaction fol-
lows Michaelis–Menten type kinetics involving an intermediate complex
formation between the oxidant and the substrate (Supplementary data,
Table S2–S9, Figs. S11–S18). The rate and the Michaelis–Menten con-
stants, kcat and K , obtained from the 1/V vs. 1/[substrate] plots are
M 0
shown in Table 2. Michaelis–Menten constant values ensure the stronger
binding of acyclic amino acids to the oxidant compared to cyclic amino
−
1
(
20%; TOF = 565 h ). The highest attained TOF values and con-
−
1
−1
versions are 8340 h
(71%) with ACHH and 9273 h
(92%) with
AIBH. The relative reactivity of substrates is in the following order:
ACHH > ACPH > ACBH > ACCH in the case of cyclic amino acids,
and AIBH > ABH > NORH > N-Me-AIBH > ALAH for acyclic amino
acids. All reactions are selective – based on the results – except for the
oxidation of ACBH. The corresponding carbonyl compound such as ace-
tone, acetaldehyde, methyl-ethylketone cyclobutanone, cyclopentanone,
cyclohexanone or ethylene was formed as main product.
acids because of low K
The kinetic parameters for ACCH measured in both DMF/H
and DMF/D O (3:1) show the presence of SIEs. Similar to the ACCO
M
value observed with acyclic derivatives.
The kinetic runs of the oxidation of amino acids with hydrogen
2
O (3:1)
III
peroxide in the presence of Fe (SALEN)Cl were conducted in 75%
2
DMF–25% water at 35 °C under pseudo-first-order conditions. The
progress of the reaction was followed gas chromatographically by
measuring the evolved product from the headspace gas. The depen-
dence of the initial rate (see Supplementary data, Fig. S10) of the
reaction on substrate concentration is studied by measuring the rate
of the reaction at different concentrations. It follows, in Fig. 2 and
the data collected in Table S1, that the oxidation of ACCH proceeds
containing system investigated earlier, the large number of protons in
the active site can exchange with solvent. On the basis of the results
(SIE values of b5) the amino acid oxidation reactions by Fe (SALEN)Cl
complex is proposed to occur via an electron transfer–proton transfer
(ET–PT) mechanism. Studies of oxidations of O\H and N\H bonds usu-
ally invoke stepwise mechanisms (ET/PT or PT/ET) rather than concerted
PCET [39,40]. The observed isotope effects are not large, but SIEs in ET–PT
III
Table 2
Steady state kinetic parameters for [FeIII(SALEN)Cl] catalyzed amino acid oxidation.
K
M
k
cat
k
cat/K
M
V
(10
max
−5
SIEa
−
3
(s−1)
(M s−1
−1
Ms−1
)
(
10
M)
)
1
7.03 ± 2.50 0.10 ± 0.01
5.45
0.07 ± 0.01
0.29 ± 0.01
1.70 ± 0.12
1.38
2.50
1.33
1
2
3.59 ± 0.05 0.41 ± 0.01
30.27
1.29 ± 2.03 2.35 ± 0.16 110.51
3.78 ± 3.13 3.04 ± 0.28 127.85
2
2.19 ± 0.20
1.80
1
2
1
1
5.99 ± 3.46 0.06 ± 0.01
0.52 ± 4.24 1.57 ± 0.24
3.69
0.05 ± 0.01
1.13 ± 0.18
1.49 ± 0.12
2.98 ± 0.20
2.10
2.13
2.47
1.24
76.41
4.86 ± 1.74 2.07 ± 0.17 139.36
2.92 ± 1.09 4.13 ± 0.28 319.68
Fig. 4. Hydrogen peroxide dependence of amino acid oxidation reactions in DMF/water (3:1)
= 3.6 × 10− M, [Fe (SALEN)Cl]
2
III
= 7.2 × 10 M,
−6
at 35 °C for acyclic substrates. [S]
0
0
a
−2
SIE = V
i
(H)/V
i
(D).
[NH OH] = 3.6 × 10 M. ■ ALAH, 0 N-Me-AIB, ▲ NORH, ● ABH, × AIBH.
4 0

50
S. Góger et al. / Journal of Inorganic Biochemistry 123 (2013) 46–52
Fig. 7. Correlation between catalyst concentration and the rate of the reaction of amino acid
−
2
−2
Fig. 5. (a) Hydrogen peroxide dependence of amino acid oxidation reactions in DMF/
oxidation in DMF/water (3:1) at 35 °C. [S] = 3.6 × 10 M, [H
2 2
O ] = 3.6 × 10 M,
= 3.6 × 10− M, [Fe (SALEN)Cl]
= 3.6 × 10− M. ● n-butyronitrile, ■ cyclobutanone, ▲ dehydroproline.
b) Proportion of products formed.
2
III
−6
[NH OH] = 3.6 × 10−2 M. × ACBH, ▼ACHH, ● ACPH, ■ ACCH.
water (3:1) at 35 °C. [ACBH]
0
0
= 7.2 × 10
M,
4
2
4 0
[NH OH]
(
3.2. Stoichiometric oxidations
reactions are not necessarily large, as demonstrated both experimentally
and theoretically [17]. Thus, for example, the observed SIE in the range
Model complex [FeIVO(SALEN)]•+, was generated by the reaction
III
1.3–2.1 are not uncommon for an ET–PT reaction.
of Fe (SALEN)Cl and H
2
O
2
3
in CH CN at 278 K, as reported earlier
Figs. 3 and 4 present the dependence of the cyclic and acyclic
amino acid oxidations as a function of the quantity of hydrogen perox-
ide. The catalyst remains comparably active even after addition of large
[33,34]. This preparation provided us with high-valent oxoiron species
stable enough to ensure the direct observation of the reactions with
amino acids. The decrease in absorbance with time at 470 nm upon
addition of substrate (in its anionic form) to the in situ generated
2 2
excess of H O . There is a linear relationship between the amount of
IV
•+
H
2
O
2
added and the rate of the formation of ethylene or the correspond-
[Fe O(SALEN)] species is followed spectrophotometrically (Fig. 8).
The new peak at ca. 420 nm – compared to the literature data
(370 nm) – can be assigned to the coordination of the anionic substrate
to the oxoiron moiety. Parallel with the above described procedure the
formation of the product (ethylene) was followed by GC analysis. Data
2 2
ing carbonyls, suggesting a first-order dependence on H O concentra-
tion for all cases studied.
In the case of ACBH – which has slightly lower ring strain than
ACCH – three kinds of main products were obtained: cyclobutanone,
n-butyronitrile and dehydroproline. This suggests a possible interpre-
tation of conversion in two different reaction pathways, decarboxyl-
ation and ring opening path (Fig. 5). The absence of an hydroxylated
intermediate refutes the proposed N-hydroxylation mechanism previ-
ously suggested for ethylene biosynthesis.
IV
•+
correspond well with each other. The [Fe O(SALEN)] species was
thermally stable even at room temperature, but capable of the activa-
tion of the N\H bond. Detailed mechanistic studies were carried out
IV
•+
to understand whether the [Fe O(SALEN)] mediated amino acid
oxidation – as a possible elemental step of the catalytic system – occurs
via a proton coupled electron transfer mechanism or a hydrogen atom
A straight line was obtained in a plot of the reaction rate versus
III
the initial concentration of [Fe (SALEN)Cl] suggesting a first-order
transfer (Supplementary data, Table S10). The calculated k
H
/k
O/D
two times larger than that of the Fe (SALEN)Cl catalyzed ACCH
D
value
dependence on catalyst concentration for all cases studied (Figs. 6, 7).
of 2.45, determined kinetically (in the presence of 5 v/v% H
2
2
O) is
III
Fig. 6. Correlation between catalyst concentration and the rate of the reaction of amino acid
Fig. 8. (a) Oxidation of ACC in CH
3
CN at 5 °C in the presence of FeIVO species. [ACC]
=
0
−
2
−2
−3
IV
−4
IV
oxidation in DMF/water (3:1) at 35 °C. [S] = 3.6 × 10 M, [H
2 2
O ] = 3.6 × 10 M,
2 × 10
M, [Fe O(SALEN)Cl]
0
= 2 × 10
M. (b) Time course of decay of Fe O
OH] = 3.6 × 10− M. ▲ NORH, × AIBH, ● ABH, ■ ALAH.
2
species under the conditions described above, monitored at 470 nm.
[
NH
4

S. Góger et al. / Journal of Inorganic Biochemistry 123 (2013) 46–52
51
oxidation, where the lower SIEs represent the result of much more mul-
tiple effects compared to the elemental step discussed above. The result
above indicates an ET–PT mechanism.
To determine the rate dependence on the two reactants, oxidation
IV
•+
runs were performed at various initial [Fe O(SALEN)] and ACC
concentrations under pseudo-first-order conditions (Fig. 9). Kinetic mea-
surements revealed first-order rate dependence on both the oxidant and
the substrate concentrations with kinetic parameters: k = (0.36 ± 0.02)
−
1
−1
‡
−1
‡
−1 −1
M
2
s
, ΔH = 38 ± 2 kJ mol , ΔS = −116 ± 7 J mol
K
at
78.16 K. Though activation parameters are often not discriminating
factors in recognizing the reaction pathway, the large negative entropy
‡
of activation (ΔS ) clearly indicates an associative mode of activation in
the rate-determining step (Fig. 10).
The effect of substituents on SALEN ligand in the 5 position was
investigated. While electron withdrawing groups enhance the rate
of the reaction – by increasing the electrophilic character of the active
−
1
3
−1
intermediate – k(NO
2
-SALEN) = 21.81 mol
dm
s
, electron donat-
−
4
−1
3
−1
ing groups decrease it k(MeO-SALEN) = 5.22 × 10
mol
dm
s
.
Fig. 9. (a) Catalyst concentration dependence of the oxidation of ACC in CH
3
CN at 5 °C.
= 1 × 10− M. (b) Substrate concentration dependence of the oxidation of
2
[
ACC]
0
3.3. Mechanism
−4
ACC in CH
3
CN at 5 °C. [FeIVO(SALEN)Cl]
0
= 2 × 10
M.
The mechanism for the Fe(III)-catalyzed model system, that is
suggested on the bases of the kinetics of stoichiometric and catalytic
systems, linear free energy correlation, intramolecular kinetic isotope
effects, and product analysis, is shown below (Scheme 5). We assume
the formation of the ternary high-valent oxoiron complex between
catalyst, substrate (see Michaelis–Menten correlation and Fig. 8) and
2 2
H O in a fast pre-equilibrium step, followed by ET–PT between the co-
ordinated substrate and the oxoiron moieties forming a substrate
radical in the rate-determining step. After that – depending on the
substrates – two main pathways can be written down. In case of sub-
strates with high ring strain the rapid radical rearrangement results
in cleavage of the cyclopropane and cyclobutane ring and formation
of ethylene and dehydroproline (n-butyronitrile), respectively (Path A
in Scheme 5). In case of other acyclic and cyclic amino acids the corre-
sponding ketimines were formed as a result of decarboxylation path-
ways, which can easily hydrolyze to carbonyls (Path B in Scheme 5).
4
. Conclusions
In this study we have demonstrated, that Fe (SALEN)Cl is an active
III
CN. [ACC] = 2 × 10− M,
3
Fig. 10. Eyring plot for the oxidation reaction of ACC in CH
[
3
and selective catalyst in the oxidation of amino acids. The reaction was
investigated first for ACCH and AIBH – possible alternate substrates –
Fe O(SALEN)Cl] = 2 × 10−4 M.
IV
Scheme 5. Possible mechanism and proposed intermediates for the amino acid oxidations with hydrogen peroxide by FeIII(SALEN)Cl.

52
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Appendix A. Supplementary data
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Kinetic details: Table S1–S10, and Figures S1–S18. This material is
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