Mechanistic study of the oxidation of L-phenylalanine by hexacyanoferrate(III) catalyzed by iridium(III) in aqueous alkaline medium

Article abstract of DOI:10.1007/s11243-010-9362-1

The oxidation of L-phenylalanine by hexacyanoferrate(III) (abbreviated as HCF) catalyzed by Ir(III) has been studied spectrophotometrically at 35 °C and at a constant ionic strength of 0.50 mol dm^-3. The main oxidation product was identified as phenylpyruvic acid by physico-chemical and spectroscopic methods. The stoichiometry was found to be 2:1, i.e. 2 mol of hexacyanoferrate(III) reacted with 1 mol of phenylalanine. The reaction was first order with respect to both HCF and alkali concentration. The order with respect to [Phe] changed from first to zero as the concentration was increased. The effect of ionic strength was also investigated. Thermodynamic parameters were evaluated by studying the reaction at four different temperatures between 35 and 50 °C. Based on the experimental results, a suitable mechanism involving complex formation has been proposed. Springer Science+Business Media B.V. 2010.

Full text of DOI:10.1007/s11243-010-9362-1

Transition Met Chem (2010) 35:549–554
DOI 10.1007/s11243-010-9362-1
Mechanistic study of the oxidation of L-phenylalanine
by hexacyanoferrate(III) catalyzed by iridium(III)
in aqueous alkaline medium
Anjali Goel Shivani Sharma
•
Received: 29 January 2010 / Accepted: 19 April 2010 / Published online: 14 May 2010
Ó Springer Science+Business Media B.V. 2010
Abstract The oxidation of L-phenylalanine by hexacya-
noferrate(III) (abbreviated as HCF) catalyzed by Ir(III) has
been studied spectrophotometrically at 35 °C and at a
presence of other oxidants [5, 6], but little attention has so
far been given to iridium(III)-catalyzed oxidation of amino
acids by alkaline HCF. Other metal ions like Os(VIII),
Ru(III), and Ag(I) are known to catalyze the oxidation of
amino acids by HCF [7–9]. Oxidation of amino acids has
also been studied using other oxidants and different prod-
ucts have been reported [10, 11]. Thus, a precise under-
standing of the mechanism of such biological redox
reactions is important as it helps in the synthesis of specific
reaction products. The study of oxidation of Phe may help
in the understanding some aspects of enzyme kinetics.
Hence in the present study we have investigated the title
reaction.
-
3
constant ionic strength of 0.50 mol dm . The main oxi-
dation product was identified as phenylpyruvic acid by
physico-chemical and spectroscopic methods. The stoi-
chiometry was found to be 2:1, i.e. 2 mol of hexacyano-
ferrate(III) reacted with 1 mol of phenylalanine. The
reaction was first order with respect to both HCF and alkali
concentration. The order with respect to [Phe] changed
from first to zero as the concentration was increased. The
effect of ionic strength was also investigated. Thermody-
namic parameters were evaluated by studying the reaction
at four different temperatures between 35 and 50 °C. Based
on the experimental results, a suitable mechanism involv-
ing complex formation has been proposed.
Experimental
The solutions were prepared afresh in water which was
twice distilled in an all glass apparatus in the presence of
potassium permanganate. AR grade chemicals were used.
K [Fe(CN) ] was recrystallized before use. Its solutions
Introduction
In recent years, interest in the oxidation of inorganic [1]
and organic substrates [2] by hexacyanoferrate(III) (HCF)
has been growing. These studies revealed that oxidation by
the HCF ion generally proceeds through an outer sphere
electron transfer mechanism, which depends not only on
the nature of the substrate but also on the medium of the
reaction. The catalytic nature of iridium(III) chloride in the
oxidation of many substrates by HCF is also well docu-
mented in acidic and alkaline media [3, 4]. Homogeneous
catalysis by iridium(III) chloride has been studied in the
3
6
were kept in amber colored bottles to prevent photode-
composition. The stock solution of iridium trichloride
(SRL, AR) was prepared in dilute HCl to a final concen-
-
3
-3
tration of 3.35 9 10 mol dm . KCl (Merck) and NaOH
(Merck) were used to provide the required ionic strength
and alkalinity, respectively.
The kinetic measurements were performed at constant
-
3
ionic strength of 0.50 mol dm and at constant tempera-
ture (35.0 ± 0.1 °C). The requisite amount of each reactant
was thermostatted at 35 °C to attain thermal equilibrium.
The appropriate quantities of reactants were mixed in a
A. Goel (&) ꢀ S. Sharma
100-mL iodine flask. The reaction was initiated by inject-
Department of Chemistry, Kanya Gurukul Mahavidyalaya,
Gurukul Kangri University, Jwalapur, Hardwar 249407, UK
e-mail: anjaligoel10@gmail.com
ing a solution of Phe into the aforementioned reaction
mixture. The reaction mixture was properly shaken and
123

5
50
Transition Met Chem (2010) 35:549–554
-
-3
[
OH ] mol dm
Results
0.4
0.3
0.2
0.1
0.0
0
Stoichiometry and product analysis
5
4
3
2
1
0
1
2
3
4
5
Different sets of reaction mixture containing excess of
hexacyanoferrate(III) to L-phenylalanine at constant ionic
strength and alkali concentration were prepared. Estimation
of residual oxidant by Cerium(IV) using ferroin indicator
after definite intervals of time revealed a 2:1 stoichiometry
between HCF and Phe.
Plot 1(a)
The main oxidation products were identified as phen-
ylpyruvic acid by spot test [12] and ammonia by Nessler’s
reagent. The nature of the keto acid was confirmed by its
UV–Vis and IR spectra (Shimadzu-8400). Absorptions at
Plot 1(b)
242 and 365 nm were observed in the UV–Vis spectrum.
0
1
2
3
4
5
Peaks observed in the IR spectra were as follows: 3,119–
-1
4
-3
[
HCF] X 10 mol dm
-1
-1
3
,044 cm [m OH]; 1625 cm [m C=O]; 1698 cm [m
-
1
-4
-
Fig. 1 Plots of a rate versus [HCF] 9 10 and b rate versus [OH ]
C=O] and 1,493 cm [m C=C]. The melting point of the
product was found to be 157 °C (lit. 154 °C). The peaks
obtained in the UV–Vis and IR spectra were comparable to
those given in the literature [13].
(
conditions as in Table 1)
5
4
3
2
1
0
Effect of reactants and medium conditions
-
The [HCF] was varied in the range 1.0 9 10 –5 9
4
-
4
-3
mol dm keeping concentrations of all other reac-
1
0
-3
-3
tants fixed as [Phe] = 3 9 10 mol dm , [NaOH] =
-5
.4 mol dm , [Ir(III)] = 3.35 9 10 mol dm . The
-3
-3
0
reaction rate increased with increase in [HCF] showing first
order behavior with respect to [HCF] (Table 1). The [Phe]
-4
-4
-3
was varied from 0.2 9 10 –6 9 10 mol dm . The
reaction followed first order kinetics at lower concentration
of Phe tending to zero order at higher concentration
(Table 1).
-
The effect of [OH ] on the initial rate of reaction was
0
1
2
3
4
5
6
-
3
4
-3
studied in the concentration range of 0.05–0.4 mol dm at
constant concentrations of substrate and oxidant. The rate
increased with increase in alkali concentration, showing
unit order dependence of on alkali concentration (Table 1).
By varying the concentration of KCl at constant [HCF],
[
Phe] x 10 mol dm
4
Fig. 2 Plot of rate versus [Phe] 9 10 (conditions as in Table 1)
-
quickly transferred to a cuvette, which was previously
maintained at a constant temperature through circulatory
arrangement of water from a thermostatic bath. The pro-
gress of the reaction was monitored spectrophotometrically
using a Shimadzu double beam spectrophotometer, UV-
[Phe], and [OH ], it was found that increasing ionic
strength of the reaction mixture increased the rate. The
linear increase in log(-dA/dt) with Hl suggests a positive
salt effect, i.e. the involvement of two similarly charged
reacting species in the reaction (Fig. 3).
3
-
1
700 at 420 nm corresponding to the kmax of [Fe(CN)6]
.
From the aforementioned results, the experimental rate
laws are given as follows:-
It was verified that there is a negligible interference from
other species present in the reaction mixture at this wave-
length. Initial rates -dA/dt were calculated from absor-
bance versus time plots. Reaction orders were calculated
from -dA/dt versus concentration plots (Figs. 1, 2).
At low [Phe]
ꢁd½HCFꢂ
¼
kobs½HCFꢂ½Pheꢂ½OHꢁꢂ:
dt
1
23

Transition Met Chem (2010) 35:549–554
551
-
Table 1 Effect of [HCF], [Phe], [OH ], and [Ir(III)] on pseudo
reaction rates at 35.0 ± 0.1 °C and l = 0.5 mol dm
At intermediate [Phe]
-3
ꢁ
d½HCFꢂ
0
0:67
½OHꢁꢂ:
4
HCF] 9 10 [Phe] 9 10 [OH ]
3
-
5
[Ir(III)] 9 10 [-dA/
¼ kobs½HCFꢂ½Pheꢂ
dt
[
(
-3
-3
-3
-3
3
mol dm
)
(mol dm
)
(mol dm
)
(mol dm
)
dt] 9 10
(
-
1
)
abs min
At high [Phe]
1
2
3
4
5
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3
3.00
3.00
3.00
3.00
3.00
0.02
0.06
0.1
0.4
0.4
0.4
0.4
0.4
0.4
0.4
0.4
0.4
0.4
0.4
0.05
0.1
0.2
0.3
0.4
0.4
0.4
0.4
0.4
0.4
0.4
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
3.35
0.67
0.80
1.00
1.21
1.34
1.67
1.2
2.1
3.0
3.9
4.8
0.3
0.7
1.4
2.4
4.0
4.6
0.6
1.2
2.4
3.6
4.8
1.0
1.1
1.4
1.7
1.8
2.4
ꢁd½HCFðIIIÞꢂ
dt
0
ꢁ
¼
kobs½HCFðIIIÞꢂ½Pheꢂ ½OH ꢂ:
The kinetics was studied at four different temperatures
(35, 40, 45 and 50 °C) under varying concentrations of
phenylalanine and hexacyanoferrate(III), keeping other
conditions constant. The rate increased with increase in
temperature. Activation parameters were evaluated from
the linear Arrhenius plots (Table 2).
0.2
0.4
Test for free radicals
0.6
3.0
The possibility of free radical formation was examined as
follows. The reaction mixture, to which a known quantity
of acrylonitrile had been added initially, was kept in an
inert atmosphere for 24 h at room temperature. When the
reaction mixture was diluted with methanol, no precipita-
tion resulted, suggesting there were no free radicals
present.
3.0
3.0
3.0
3.0
3.0
3.0
3.0
3.0
3.0
3.0
Table 2 Thermodynamic activation parameters for the oxidation of
Phe by HCF catalyzed by Ir(III) in aqueous alkaline medium with
respect to the slow step of Scheme 2
Parameter
Value
(
a) Activation parameters
-1
1
1
1
0
.4
.2
.0
.8
Ea (kcal mol
)
5.34
-31.81
4.71
14.69
6.32
-3
-
1
-1
DS (cal K mol
)
-
1
DH (kcal mol
DG (kcal mol
log A
)
)
-
1
-3
Temperature(K)
K
1
9 10
2
K 9 10
(
b) Effect of temperature on equilibrium constants
3
3
3
3
08
13
18
23
2.18
3.06
4.38
5.78
3.51
3.98
4.72
5.94
Parameters
Value from K
1
Value from K
2
0
.3
0.4
0.5
0.6
0.7
(
c) Thermodynamic parameters
1
/2
µ
-1
DE (kcal mol
)
15.25
14.62
3.98
9.15
-
1
DH (kcal mol
)
8.53
Fig. 3 Plot for the oxidation of Phe by HCF catalyzed by Ir(III) in
-3
-1
-1
)
-
aqueous alkaline medium at 308 K; [Phe] = 3 9 10 mol dm
3
DS (cal K mol
-1
DG (kcal mol
-15.67
13.47
,
-3,
-4
-3
mol dm ,[Ir(III)] = 3.35 9 10 mol dm
-5
[
[
HCF] = 3 9 10
-
OH ] = 0.1 mol dm
)
13.36
-3
123

5
52
Transition Met Chem (2010) 35:549–554
Discussion
(Fig. 4c). The carboxyl oxygen of Phe is most likely
involved in the formation of complex C . In the next step,
1
It is well known that amino acids exist as zwitterions in
aqueous solution. The pKa of the ammonium group of
amino acids is generally *9.6 at 25 °C. Since the pH of
the reaction mixture in the present study was greater than
HCF combines with C through electron abstraction to
1
form complex C2.
The complex C then slowly disproportionates to give
2
the final product along with iridium(I) and hexacyanofer-
3
?
1
0, the anion of Phe was assumed to be the reactive species
14]. It is reported that Ir(I) and Ir(II) are the most stable
species of iridium in acidic medium [15, 16], but in alka-
rate(II). The complex formation between Ir and oxidant
was also reported in the literature [19]. The probable
structures of C and C are shown in Scheme 1.
[
1
2
3
-
1?
3?
line medium [IrCl6] is the only reacting species of irid-
ium [17]. Srivastava et al. [18] have reported that the
oxidation of amino acids involves the cleavage of N–H and
C–H bonds in the rate-determining step. Based on the
aforementioned results, a mechanism can be proposed for
the present case. In the first equilibrium step, the amino
Ir is reoxidized to Ir by two molecules of hexa-
cyanoferrate(III) via one electron transfer processes.
Metal ion complexes with organic substrates make the
electron transfer easier [20, 21]. A similar type of
mechanism for the oxidation of amino acids has been
reported in the literature [22]. Repeated spectral scans
were recorded as a function of time at different concen-
trations of Phe. At low concentration of Phe, three distinct
peaks due to HCF [23] at 280, 310 and 420 nm and an
additional peak at 224 nm due to Ir(III) were recorded.
The peak heights decreased with time, due to progress of
the reaction. No peaks due to intermediates were recorded
due to their transient existence. But at higher concentra-
tions of Phe, sharp peaks were not observed due to
accumulation of adduct (Fig. 5).
acid anion forms a loosely bound complex C with
1
3
IrCl6] . Spectroscopic evidence for this catalyst–sub-
-
[
strate complex was obtained from the UV–Vis spectra of
iridium(III) and Phe and iridium(III)-Phe mixtures. Peaks
were observed for pure iridium(III) and Phe at 224 and
2
06 nm, respectively. A bathochromic shift of 5 nm in
Ir(III)-Phe mixture from 224 to 229 nm was observed in
the spectrum of catalyst. The kinetic evidence for complex
formation was obtained from Michaelis–Menton plots
(a)
(b) 0.9
3.5
3.0
2.5
2.0
1.5
1.0
0.5
0.0
0
.8
.7
0
0.6
0.5
0.4
0.3
0.2
0.1
0.0
0
1
2
3
4
5
0.0
0.2
0.4
0.6
0.8
1.0
-
1
-4
-3
-1
-1
-4
-1
3
[
Phe] x 10 dm mol
[HCF] X 10 mol dm
(c)
5
4
3
2
1
0
0
1
2
3
4
5
6
7
8
-1
-5
-1
3
[Ir(III)] X 10 mol dm
-
1
-1
-4
-1
-4
-1
-1
-5
Fig. 4 Plots of (a) rate versus [Phe] 9 10 (b) rate versus [HCF] 9 10 ; (c) rate versus [Ir(III)] 9 10 (conditions as in Table 1)
1
23

Transition Met Chem (2010) 35:549–554
553
O
O
C
3
+
R
C H
N H
C
O
I r C l
6
R
C H
N H
O
I r C l
F e
6
2
2
C
1
C
2
1 2
Scheme 1 Probable structures for complex C and C
According to Eq. 7, plots of 1/rate versus 1/[Phe], 1/rate
versus 1/[HCF(III)] and 1/rate versus 1/[Ir(III)] were linear
(
Fig. 4). From the intercepts and slopes of these plots, the
2
reactionconstants k, K and K werecalculatedas4.86 9 10 ,
1
2
3
.18 9 10 and 3.51 9 10 dm mol min , respectively,
3
3
-1
-1
2
at 35 °C. The effect of ionic strength suggests a positive salt
effect, i.e. the involvement of two similarly charged species in
the reaction as seen in Scheme 2. Thermodynamic quantities
for the equilibrium steps of Scheme 2 were calculated as
follows.
The [Phe] and [HCF] were varied at different tem-
peratures. The plots of 1/rate versus 1/[Phe] and 1/rate
versus 1/[HCF(III)] were linear as in Fig. 4. From the
0
Fig. 5 Spectra indicating accumulation of the adduct at high concen-
-
3
-3
trations of amino acids [Phe] = 5 9 10 mol dm , [HCF] = 3 9
slopes and intercepts, the values of k, K1 and K were
2
-
0
4
-3
mol dm , [OH ] = 0.4 mol dm , [Ir(III)] = 3.35 9 10
-
-3
-5
1
calculated at different temperatures. Van’t hoff plots were
-
3
-3
mol dm l = 0.5 mol dm , Temp. = 35 °C
made for the variation of K and K with temperature [i.e.
1
2
0
log K1 versus 1/T and log K versus 1/T]. The values of
2
Scheme 2 leads to the following rate equation
DH, DS and DG were calculated for the equilibrium steps
of Scheme 2. These values are summarized in Table 2. A
comparison of these values with those obtained for the
slow step shows that they mainly refer to the rate-limiting
step, supporting the idea that the reaction before the rate-
limiting step is fairly slow and with a high activation
energy [24]. The moderate values of DH and DS were
both favorable for the electron transfer process. The
negative values of S suggest that the intermediate com-
plex is more ordered than the reactants [25]. The observed
activation energy and sizeable entropy of activation sup-
port a complex transition state in the reaction. The values
of activation parameters for the oxidation of some amino
acids by HCF catalyzed by iridium(III) in aqueous alka-
line medium are summarized in Table 3. The entropy of
activation for the title reaction falls within the observed
ranges.
ꢁ
d½HCFðIIIÞꢂ
dt
3
þ
ꢁ
kK1K2½HCFðIIIÞꢂ½Ir ꢂ½Pheꢂ½OH ꢂ
¼ 1
:
ꢁ
ꢁ
þ K1½Pheꢂ½OH ꢂ þ K1K2[Pheꢂ½OH ꢂ½HCF(IIIÞꢂ
ð6Þ
Equation 6 can also be written as
1
1
3
¼
Â
Ã
k Ir
þ
ꢁ
rate kK K ½HCFꢂ Ir ½Pheꢂ½OH ꢂ
1
2
ð7Þ
1
1
þ
Â
à þ  3þÃ
3þ
kK2½HCFꢂ Ir
b, the isokinetic temperature has been evaluated from
Exner’s equation [26, 27];
b ¼ T T ðb ꢁ 1Þ=T b ꢁ T
1
2
2
1
log k at 313 K was plotted against log k at 308 K (Fig. 6).
2
1
The slope ‘b’ of the plot gave the value of b as 339 K. The
higher value of b than the experimental temperature
(308 K) indicates that the rate is governed by the enthalpy
Scheme 2
of activation [28].
123

5
54
Transition Met Chem (2010) 35:549–554
Table 3 Activation parameters for some amino acids (for isokinetic temperature)
2
9 10 (at 308 K)
2
10 (at 313 K)
Amino acids
k
(
1
k
2
DS
(cal mol
DH
(kcal mol )
References
3
dm mol
-1
3
( dm mol
-1
-1 -1
K
-1
)
)
)
L-alanine
3.8
7.7
8.9
5.1
5.7
10.8
12.8
7.2
-20.0
-21.3
-18.4
-28.2
13.9
13.1
13.8
11.2
[20]
L-leucine
[20]
L-valine
[20]
L-phenylalanine
Present work
1.0
0.9
0.8
0.7
0.6
References
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The present study shows that Ir(III) chloride acts as an
effective catalyst in the oxidation of Phe by HCF. It was
observed previously that in the oxidation of some cyclic
alcohols [29], Ir(III) also acts as an effective catalyst and
the reaction proceeds through an electron transfer process.
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acids by acidic quinolinium fluorochromate catalyzed by
Ir(III) involves a transient complex between Ir(III), oxidant
and substrate. The oxidation of Phe occurs through inter-
mediate complex formation. The large negative value of
DS suggests that the intermediate complex is more ordered
than the reactants. The isokinetic temperature was found to
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enthalpy of activation.
1
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