Oxidation of thiols to disulfides by dioxygen catalyzed by a bioinspired organocatalyst

Article abstract of DOI:10.1039/c5ra05529f

2,3-Dihydro-2,2,2-triphenylphenanthro[9,10-d]-1,3,2-λ⁵-oxazaphosphole serves as good catalyst for the oxidation of thiophenol, cysteine and glutathione to their disulfides by molecular oxygen. The kinetics of the reactions unveiled an overall second order rate equation for all reactions and pure dioxygen chemistry for all three substrates. The formation of an unstable hydroperoxide from the catalyst is assumed to be a key step during the reaction.

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Oxidation of thiols to disulfides by dioxygen catalyzed by a
bioinspired organocatalyst†
Nárcisz Bagi, József Kaizer, Gábor Speier*
5
Received (in XXX, XXX) Xth XXXXXXXXX 20XX, Accepted Xth XXXXXXXXX 20XX
DOI: 10.1039/b000000x
5
2,3-Dihydro-2,2,2-triphenylphenanthro[9,10-d]-1,3,2-
-
oxazaphosphole serves as good catalyst for the oxidation of
thiophenol, cysteine and glutathione to their disulfides by
10 molecular oxygen. The kinetics of the reactions unveiled
an overall second order rate equation for all reactions and
pure dioxygen chemistry for all three substrates. The
formation of an unstable hydroperoxide from the catalyst
is assumed as a key step during the reaction.
50 Scheme 1 The tautomeric forms of 1,3,2-oxazaphosphole (1).
15 Oxidative coupling of thiols to disulfides has attracted
interest in biological1 and synthetic contexts.2 Cysteine
residues are possible catalysts for protein modification3 and
quiescinsulfhydryl oxidases4 are responsible for oxidative
protein folding. Glutathione5 and disulfides6 prevent oxidative
20 stress and oxidative damage in biological systems.
in the range from -23 to -8 ppm. In more basic solvents the
oxazaphosphole form 1 is dominant. In solution they
decompose readily in air due to dioxygen or moisture.
1,3,2-Oxazaphospholes mimic flavoproteins,16 pterins17 and
55 deprotonated uric acid18 (e. g. Scheme 2). That means that
they react with triplet dioxygen to form unstable
Oxidative coupling of thiols is a very common reaction for
the synthesis of disulfides. Oxidative S-S coupling can be
achieved by permanganate,7 halogens,8 FeCl3,9 organometallic
complexes,10 peroxides11 and nanoparticles.12
60
25 In this paper we report that an 1,3,2-oxazaphosphole is a
good bioinspired catalyst for the oxidation of thiophenol,
cysteine and glutathione to their corresponding disulfides by
triplet dioxygen.
1,2-Quinones undergo with tertiary phosphines a [4+1]
30 electrocyclic reaction giving dioxaphosphole heterocyclic
rings.13 1,2-Azaquinones give 1,3,2-oxazaphospholes (1) by
the use of tertiary phosphines in good yields.14,15 Where the
monoimines are unstable they can also be prepared „in situ“
from the corresponding quinones and triphenylphosphine in a
35 sealed tube under ammonia. Electron-releasing groups on the
quinones make the nucleophilic replacement of the C=O bond
with NH3 to a C=NH functionality difficult or impossible (e.
g. NMe2, OR).
65
70
Scheme 2 Similarities of flavoprotein and 1,3,2-oxazaphosphole reactions
with 3O2.
hydroperoxide, as shown earlier.19 These hydroperoxides are
75 then able to oxidize/oxygenate various substrates, such as
PPh319 and CO2.20 We extended the oxidation by 3O2 catalyzed
by the 1,3,2-oxazaphosphole 1 to thiols, such as thiophenol,
cysteine and glutathione. Thiophenol represents thiols, while
cysteine oxidation to disulfide has some relevance to protein
80 folding3,4 and glutathione is very important against oxidative
stress in biological systems.5,6 Bulk oxidation of the substrates
in methanol or in methanol/water mixture at 298 K resulted in
yields of 93, 92 and 76% (GC and O2-uptake, isolated yields:
1,3,2-Oxazaphospholes exhibit tautomerism as shown in
40 Scheme 1 between the oxazaphosphole form (1) and the
iminophosphorane form (2) dependent on the consistency or
the solvent of solution (Scheme 1).14,15 1,3,2-Oxazaphospholes
(1) seem to be stable in the solid form. In solution, however
the tautomeric compounds of the iminophosphorane form (2)
45 are also present. 31P NMR spectra in the range of 39-51 ppm
are characteristic for 1 and those of the form 2 have a shift
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78, 70 and 65%) of the disulfide. In the first cases the yields
k2PhSH = 5.5 0.3 × 10-1 M-1s-1, k2cysteine = 12.5 1.8 × 10-1 M-
glutathione
were calculated from the GC and dioxygen-uptake (see ESI 45 1s-1 and k2
= 4.2 0.5 × 10-1 M-1s-1 with the activation
general part and Fig. S1). No other oxidized sulfur-containing
products were formed, though they have been detected in
numerous other oxidation formats (H2O2 and peroxy acids, for
example, may lead to sulfinic acid.21,22 We measured
parameters PhSH: ꢀEa = 24.5 0.4 kJmol-1, ꢀH‡ = 21.6 1.1
kJmol-1and ꢀS‡ = -177.0 0.4 Jmol-1K-1, cysteine: ꢀEa = 8.1
0.1 kJmol-1,ꢀH‡ = 5.6 0.1 kJmol-1, ꢀS‡ = -224.2 0.2 J mol-
5
1
K-1 and glutathione: ꢀEa = 9.9 0.1 kJmol-1, ꢀH‡ = 7.0
autoxidation of the substrates under identical condition and 50 0.1 kJ mol-1, ꢀS‡ = -228.7 0.2 Jmol-1K-1 (Fig. S22 - Fig. S27
deduced their rate from the catalytic reactions (Fig. S2 - Fig
and Tables S1 - S3).
S4). The disulfides were characterized by comparing them
This means that in these of reactions the catalyst reacts with
triplet dioxygen in a slow step and this is rate-determining.
The formation of the hydroperoxide 5 determines the
10 with authentic samples (mps, mixed mps, IR spectra, MS and
1
NMR). GC-MS (thiophenol), H- and 13C-NMR (cysteine and
glutathione)) (Fig S.5 - Fig. S12). Kinetic studies on the 55 scenario. All the consecutive reactions of the hydroperoxide 5
oxidation of the three substrates were carried out measuring
the disulfide amount by gas chromatograph (thiophenol) and
15 uptake of 3O2 in the case of cysteine and glutathione (Fig. S1).
Typical time courses can be seen in Fig. 1. The oxidation
with thiophenol, cysteine and glutathione and the formation of
the disulfides, are fast reactions, which do not influence the
reaction rates. The reactions can be considered as pure
dioxygen chemistry as depicted in Scheme 3. If this is true the
reactions have been carried out in a thermostated vessel 60 reaction rates must be close to each other. Of course we have
connected to a gas buret filled with O2 (constant pressure) and
a septum to take samples for GC. Duplicate runs were done in
20 each case.
to consider that in the case of cysteine and glutathione
MeOH:H2O = 2:1 mixture have to be used as solvent due to
solubility reason. This roughly holds if we look at the rate
constants. However, the O2-solubility in the solvents and the
65 various substrates probably have some effect on the reaction
rates too. There is also a difference in the product formation.
In the case of PhSH
Figure 1 Time course of the oxidation of thiols by 3O2 catalyzed by 1.
● [cysteine] = 5.50 × 10-2 M, [1,3,2-oxazaphosphole] = 6.00 × 10-4 M,
[O2] = 3.54 × 10-3 M, V(MeOH:H2O=2:1) = 30 mL, T = 25 °C.
25 ■ [glutathione] = 5.50 × 10-2 M, [1,3,2-oxazaphosphole] = 6.00 × 10-4 M,
[O2] = 3.54 × 10-3 M, V(MeOH:H2O=2:1) = 30 mL, T = 25 °C.
♦ [PhSH] = 5.50 × 10-2 M, [1,3,2-oxazaphosphole] = 6.00 × 10-4 M, [O2]
= 9.50 × 10-3 M, V(MeOH) = 30 mL, T = 25 °C.
Scheme
3 The proposed mechanism of thiophenol, cysteine and
70 glutathione oxidation by 3O2 catalyzed by 2
.
30 The partial rate orders of the reaction were secured also by
plotting the initial concentrations of PhSH, cysteine and
glutathione against the reaction rates (Fig. S13 – Fig. S15),
which gave also straight lines. These Figures, where the
reaction rates remained constant, reinforced the zero order
35 dependence on the thiol concentrations. Plotting the reaction
rates against the various starting concentrations of 1 and
dioxygen resulted also in straight lines (Fig. S16 - Fig. S21)
indicating first order dependence on 1 and dioxygen
concentration. The kinetic data suggested an overall second
40 order rate equation (1) and the second order rate constants are
hydrogen peroxide is formed (that was determined from
dioxygen-uptake and also iodometric titration to be 87%),23
while in the case of cysteine and glutathione water. Since all
75 these steps are after the rate-determining step the kinetic data
do not help us, we can only speculate or reasonable arguing
what can be assumed.
Based on that we believe that the hydroperoxide 5 reacts in a
fast reaction with the thiophenol, which is rather acidic, to an
80 adduct (6) and H2O2, and a further reaction of thiophenol with
6 to the disulfide and the catalyst (Scheme 4). It is interesting
to note that in the case of thiophenol oxidation beside of the
disulfide hydrogen peroxide is formed, while in the case of
cysteine and glutathione water. These reactions, after the rate-
Reaction rate = k2 [catalyst] [3O2]
(1)
2
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determining step, are fast ones and to our believe the unstable 35 resembles the classical hydroperoxide oxidation of the
hydroperoxide (5) reacts with PhSH deliberating H2O2 and the
compound 6 is formed. Compound like 6 was assumed in the
reaction of an electron-deficient isoalloxazine, 3,10-dimethyl-
8-cyano-isoalloxazine with thiophenol.24 Compound 6 reacts
then with a further thiophenol molecule to give the disulfide
and the catalyst 2. This reacts then again with O2 resulting in 5
just to close the catalytic cycle.
substrates.
Conclusions
5
In summary, we have disclosed that a 1,3,2-oxazaphosphole
(1) catalyses the oxidation of thiophenol, cysteine and
40 glutathione by molecular oxygen to their corresponding
disulfides. The oxidation of thiols to disulfides has some
importance on the structure of proteins, and in the last case
glutathiones role as antioxidant in the cell is well recognized.
Kinetics of the reactions resulted in an overall second order
45 rate equation. That means that all the reactions can be called
as pure dioxygen chemistry, since the rate-determining step is
the reaction of the catalyst with molecular oxygen forming the
hydroperoxide (5). After that 5 reacts with thiophenol to H2O2
and in the case of cysteine and glutathione to water and the
50 corresponding disulfides. 1,3,2-Oxazaphospholes in these
cases mimic flavoproteins exhibiting similar mechanistic
features.
If we look at the substrates cysteine or glutathione rather
10 different pictures were found. In the case of cysteine and
glutathione again a dioxygen chemistry could be observed,
what means that the rate-determining step of the reaction is
the reaction of the catalyst (2) with triplet molecular oxygen,
which reacts in a fast consecutive reaction with the cysteine or
15 glutathione and water and the catalyst is reformed.
55 This work was supported by a grant from The Hungarian Research Fund
(OTKA) # K108489 and COST Actions CM1205, CM1201 and CM1003.
Notes and references
Department of Chemistry, University of Pannonia, Egyetem u. 10,
Veszprém, Hungary. Fax:3688 624469; Tel: 36 88 624720; E-mail:
60 speier@almos.uni-pannon.hu
†Electronic Supplementary Information (ESI) available: Experimental
details of synthesis and structural characterization, kinetics and
spectroscopic data. For ESI and electronic format see
DOI: 10.1039/b000000x/
Scheme 4 The proposed fast reactions of the hydroperoxide 5 with
20 thiophenol.
65
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