Reductive metalation of cyclic and acyclic pseudopeptidic bis-disulfides and back conversion of the resulting diamidato/dithiolato complexes to bis-disulfides

Article abstract of DOI:10.1021/ic101148c

Cyclic and acyclic pseudopeptidic bis-disulfides built on an o-phenylene diamine scaffold were prepared: (N₂H₂S₂) ₂, 1a, N₂H₂(S-SCH₃)₂, 1b, and N₂H₂(S-StBu)₂, 1c. Reductive metalation of these disulfides with (PF₆)[Cu(CH₃CN)₄] in the presence of Et₄NOH as a base, or with (Et₄N)[Fe(SEt) ₄] and Et₄NCl, yields the corresponding diamidato/dithiolato copper(III) or iron(III) complex, (Et₄N) [Cu(N₂S₂)], 2, or (Et₄N)₂[Fe(N ₂S₂)Cl], 5. These complexes display characteristics similar to those previously described in the literature. The mechanism of the metalation with copper has been investigated by X-band electron paramagnetic resonance (EPR) spectroscopy at 10 K. After metalation of the bis-disulfide 1c and deprotonation of the amide nitrogens, the reductive cleavage of the S-S bonds occurs by two one-electron transfers leading to the intermediate formation of a copper(II) complex and a thyil radical. Complexes 2 and 5 can be converted back to the cyclic bis-disulfide 1a with iodine in an 80% yield. Reaction of 5 with iodine in the presence of CH₃S-SCH₃ affords a 1/1 mixture of the acyclic N₂H₂(S-SCH₃)₂ disulfide 1b and cyclic bis-disulfide 1a. From 2, the reaction was monitored by ¹H NMR and gives 1b as major product. While there is no reaction of 2 or 5 with tBuS-StBu and iodine, reaction with an excess of tBuSI affords quantitatively the di-tert-butyl disulfide 1c. To assess the role of the Cu ^III oxidation state, control experiments were carried out under strictly anaerobic conditions with the copper(II) complex, (Et ₄N)₂[Cu(N₂S₂)], 6. Complex 6 is oxidized to 2 by iodine, and it reacts with an excess of tBuSI, yielding 1c as final product, through the intermediate formation of complex 2.

Full text of DOI:10.1021/ic101148c

Inorg. Chem. 2010, 49, 8637–8644 8637
DOI: 10.1021/ic101148c
Reductive Metalation of Cyclic and Acyclic Pseudopeptidic Bis-Disulfides and
Back Conversion of the Resulting Diamidato/Dithiolato Complexes
to Bis-Disulfides
Nicolas Desbenoıt,†,‡ Erwan Galardon,† Yves Frapart,† Alain Tomas,‡ and Isabelle Artaud*,†
ˆ
†
ꢀ
Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, Universite Paris Descartes,
UMR 8601, CNRS, 45 rue des Saints Peres, 75270 Paris Cedex 06, France, and ‡Laboratoire de
ꢁ
ꢀ
Crystallographie et RMN Biologiques, Universite Paris Descartes, UMR 8015, CNRS,
4 avenue de l’Observatoire, 75270 Paris Cedex 06, France
Received June 8, 2010
Cyclic and acyclic pseudopeptidic bis-disulfides built on an o-phenylene diamine scaffold were prepared: (N2H2S2)2,
1a, N2H2(S-SCH3)2, 1b, and N2H2(S-StBu)2, 1c. Reductive metalation of these disulfides with (PF6)[Cu(CH3CN)4]
in the presence of Et4NOH as a base, or with (Et4N)[Fe(SEt)4] and Et4NCl, yields the corresponding diamidato/
dithiolato copper(III) or iron(III) complex, (Et4N)[Cu(N2S2)], 2, or (Et4N)2[Fe(N2S2)Cl], 5. These complexes display
characteristics similar to those previously described in the literature. The mechanism of the metalation with copper has
been investigated by X-band electron paramagnetic resonance (EPR) spectroscopy at 10 K. After metalation of the
bis-disulfide 1c and deprotonation of the amide nitrogens, the reductive cleavage of the S-S bonds occurs by two one-
electron transfers leading to the intermediate formation of a copper(II) complex and a thyil radical. Complexes 2 and 5
can be converted back to the cyclic bis-disulfide 1a with iodine in an 80% yield. Reaction of 5 with iodine in the presence
of CH3S-SCH3 affords a 1/1 mixture of the acyclic N2H2(S-SCH3)2 disulfide 1b and cyclic bis-disulfide 1a. From 2, the
reaction was monitored by 1H NMR and gives 1b as major product. While there is no reaction of 2 or 5 with tBuS-StBu
and iodine, reaction with an excess of tBuSI affords quantitatively the di-tert-butyl disulfide 1c. To assess the role of the
CuIII oxidation state, control experiments were carried out under strictly anaerobic conditions with the copper(II)
complex, (Et4N)2[Cu(N2S2)], 6. Complex 6 is oxidized to 2 by iodine, and it reacts with an excess of tBuSI, yielding 1c
as final product, through the intermediate formation of complex 2.
Introduction
second nickel cation is required for activation of ACS/CODH.2
In this latter case, the two cysteine thiolates are bridging
ligands between the two Ni atoms. Very recently, hepcidin, a
25 amino acid, cysteine rich peptide was found to act as the
central regulator of body iron metabolism.3 Hepcidin binds
to the iron exporter ferroportin leading to its internalization
and degradation, thus regulating the export of iron from cells
to plasma.4 This peptide contains eight cysteines involved in
four disulfide bridges, the connectivities of which were assig-
ned by NMR experiments.5 With so many cysteines, hepcidin
is proposed to bind iron and to be an iron sensor. A first
hypothesis is the formation of an iron sulfur center. Indeed, a
proportion of urine purified hepcidin retains iron, and electron
paramagnetic resonance (EPR) spectroscopy analysis suggests
The Cys-X-Cys tripeptide motif provides, through the de-
protonated peptide nitrogens and the thiolate sulfurs, the
N2S2 donor set to the metal cation at the active site of nitrile
hydratase (NHase)1 and acetyl coenzyme A synthase/carbon
monoxide dehydrogenase (ACS/CODH).2 The reactivity of
these two metalloproteins are finely tuned by specific metal
coordination and post-translational modifications of the
cysteines thiolates. Oxygenation to sulfinate and sulfenate
is required for activity of iron and cobalt NHase1 and while a
first nickel cation is found at the N2S2 site, incorporation of a
*To whom correspondence should be addressed. E-mail: isabelle.artaud@
parisdescartes.fr. Phone: þ33-1-42 86 21 89. Fax: þ33-1-42 86 83 87.
(1) (a) Nagashima, S.; Nakasako, M.; Dohmae, N.; Tsujimura, M.;
Takio, K.; Odaka, M.; Yohda, M.; Kamiya, N.; Endo, I. Nat. Struct. Biol.
1998, 5, 347–351. (b) Miyanago, A.; Fushinobu, S.; Ito, K.; Wakagi, T. Biochem.
Biophys. Res. Commun. 2001, 288, 1169–1174.
(2) (a) Darnault, C.; Volbeda, A.; Kim, E. J.; Legrand, P.; de Verne, X.;
Lindahl, P. A.; Fontecilla-Camps, J. C. Nat. Struct. Biol. 2003, 10, 271–279.
(b) Doukov, T. I.; Iverson, T. M.; Seravalli, J.; Ragsdale, S. W.; Drennan, C. L.
Science 2002, 298, 567–572.
(3) Ganz, T.; Nemeth, E. Biochim. Biophys. Acta 2006, 1663, 690–699.
(4) Nemeth, E.; Tuttle, M. S.; Powelson, J.; Vaughn, M. B.; Donovan, A.;
Ward, D. M.; Ganz, T.; Kaplan, J. Science 2004, 306, 2090–2093.
(5) (a) Jordan, J. B.; Poppe, L.; Haniu, M.; Arvedson, T.; Syed, R.; Li, V.;
Kohno, H.; Helen Kim, H.; Schnier, P. D.; Miranda, L. P.; Cheetham, J.;
Sasu, B. J. J. Biol. Chem. 2009, 284, 24155–24167. (b) Hunter, H. N.; Fulton,
D. B.; Ganz, T.; Vogel, H. J. J. Biol. Chem. 2002, 277, 37597–37603.
r
2010 American Chemical Society
Published on Web 08/18/2010
pubs.acs.org/IC

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Desbenoıt et al.
8638 Inorganic Chemistry, Vol. 49, No. 18, 2010
Scheme 1. Reductive Metalation of the Bis-Disulfides with (PF6)[CuI(CH3CN)4]
a rubredoxin-like iron sulfur site.6 An iron sulfur cluster has
Reductive Metalation of the Bis-Disulfides 1a-1c with Tetra-
also been previously postulated.7 Nevertheless, all the char-
acterizations are very difficult because purified hepcidin is
only available in a very low amount. Whatever the iron sulfur
center, presently its formation always involves metalation of
the reduced form of hepcidin. Moreover, when looking at the
sequence of hepcidins, we can notice another potential metal
binding site defined by a Cys-X-Cys tripeptide, in which the
two cysteinesare post-translationallyoxidized into disulfides.
Why not consider that this motif defines a cavity able to
trap a metal cation such as iron or copper? Actually, some
urine purified hepcidin was also found to retain copper ion.6b
We have studied this hypothesis using pseudopeptidic cyclic
(N2H2S2)2 and acyclic bis-disulfide N2H2(S-SR)2 synthe-
tic analogues, and we have shown that direct metalation of
such proligands under reductive conditions using Fe3CO12 or
(Et4N)[FeIII(SEt)4] and (PF6)[CuI(CH3CN)4] leads to the
formation of the well-known (Et4N)2[FeIII(N2S2)(Cl)] and
(Et4N)[CuIII(N2S2)] complexes with the typical [N2S2]4- donors.
Interestingly upon oxidation with iodine, the iron and copper
complexes alone or in the presence of an exogenous linear
disulfide RSSR, revert back to the cyclic and acyclic disul-
fide ligands respectively, while the iron and copper ions are
ejected.
kis(acetonitrile)copper Hexafluorophosphate, (PF6)[CuI-
(CH3CN)4]. Reaction of the cyclic bis-disulfide, 1a, inaceto-
nitrile, with 2 equiv of the copper(I) salt, (PF6)[CuI(CH3CN)4]
and 4 equiv of Et4NOH in methanol solution, as a base,
affords cleanly the diamidato/dithiolato copper(III) com-
plex (Et4N)[Cu(N2S2)] 2 (Scheme 1), previously prepared
by Hanss and Kruger10 upon electrochemical oxidation of
€
the corresponding diamidato/dithiolato copper(II) com-
plex. While there is no reaction without adding a base, upon
addition of Et4NOH to deprotonate the amide nitrogens,
the solution turns instantaneously greenish yellow, exhibit-
ing an absorption at 417 nm in electronic spectroscopy in
CH3CN. After removing the solvents, complex 2 was pre-
cipitated in cold diethyl ether and characterized by mass
spectrometry, UV-vis spectroscopy, electrochemistry, and
1H NMR. It displays the same characteristics as those
previously reported in the literature.10,11 Complex 2 was
also quantitatively obtained starting from the acyclic bis-
disulfides 1b and 1c, after addition of 1 equiv of the copper-
(I) salt and 2 equiv of Et4NOH. This reaction also leads to
the formation of 1,2-dimethyl disulfide from 1b and 1,2-di-
tert-butyl disulfide from 1c, respectively, that were identified
by GC and 1H NMR, respectively.
Copper(I) is well-known to coordinate to sulfur-con-
taining groups, including organic disulfides. Several di-
nuclear copper(I) complexes have been isolated and char-
acterized by X-ray crystallography, such as those derived
from bis[2,(N,N0-dimethylamino)ethyl]disulfide12 or bis-
{2-[(alkyl)(2-pyridylethyl)amino]ethyl} disulfide.13 In these
complexes the two CuI ions are bridged by the disulfide.
In the pyridyl series, changing the spacer from ethyl to
methyl, as in bis{2-[(benzyl)(2-pyridylmethyl)amino]ethyl}
disulfide, promotes the reductive cleavage of the S-S
bond and the conversion of the disulfide-dicopper(I) com-
plex into a bis(μ-thiolato)dicopper(II) complex.13 This effect
Results and Discussion
The cyclic bis-disulfide, 1a (Scheme 1), was selectively pre-
pared, as previously described,8 by iodine oxidation of the
Ni complex derived from the dithiol, N,N0-(1,2-phenylene)bis-
(2-mercapto-2-methylpropanamide), N2S2H4. The acyclic bis-
disulfides, 1b and 1c, (Scheme 1) were synthesized by reaction
of the dithiol, N2S2H4, with S-methyl methane thiosulfonate
and S-tert-butyl methane thiosulfonate, respectively, follow-
ing a procedure described by Grayson et al.9
(6) (a) Farnaud, S.; Rapisarda, C.; Bui, T.; Drake, A.; Cammak, R.;
Evans, R. W. Biochem. J. 2008, 413, 553–557. (b) Farnaud, S.; Patel, A.; Evans,
R. W. Biometals 2006, 19, 527–533.
€
(10) Hanss, J.; Kruger, H.-J. Angew. Chem., Int. Ed. Engl. 1996, 35, 2827–
2830.
(7) Gerardi, G.; Biasiotto, G.; Santambrogio, P.; Zanella, I.; Ingrassia,
R.; Corrado, M.; Cavadini, P.; Derosas, M.; Levi, S.; Aorosio, P. Blood Cells
Mol. Dis. 2005, 35, 177–191.
(11) Desbenoit, N.; Galardon, E.; Roussel, P.; Artaud, I.; Tomas, A.
J. Coord. Chem. 2009, 62, 2472–2479.
(12) Ottersen, T.; Warner, L. G.; Seff, K. Inorg. Chem. 1974, 13, 1904–
1911.
ꢁ
(8) Bourles, E.; Alves de Sousa, R.; Selkti, M.; Tomas, A.; Artaud, I.
(13) (a) Itoh, S.; Nagagawa, M.; Fukuzumi, S. J. Am. Chem. Soc. 2001,
123, 4087–4088. (b) Osako, T.; Ueno, Y.; Itoh, S. Inorg. Chem. 2003, 42, 8087–
8097.
Tetrahedron 2007, 63, 2466–2471.
(9) Grayson, E. J.; Ward, S. J.; Hall, A. L.; Rendle, P. M.; Gamblin, D. P.;
Andrei, S.; Batsanov, A. S.; Davis, B. G. J. Org. Chem. 2005, 70, 9740–9754.

Article
Inorganic Chemistry, Vol. 49, No. 18, 2010 8639
between 4 and 30 K, at g = 2.07 for the copper(II) com-
plex, and g=2.01 for the organic radical species. As expected,
and as shown in Figure 1b from the plot of I versus P1/2 in
mW at 30 K, the intensity of the radical signal is easily
saturated and further decreases, while the intensity of the
copper(II) signal slowly increases and reaches a plateau.
Such an experiment enables to separate radical and metal
spectra.14 The variation of the intensity versus the power
can be modeled from eq 1 derived from that of Innes and
Brudvig15 that has been modified to take in account the
presence of two different magnetic species.16
pffiffiffi
P
pffiffiffi
P
Ccopper
P
P1=2copper
ꢀ
Cradical
P
P1=2radical
ꢀ
þ
ð1Þ
!
!
b
copper=2
b
radical=2
1 þ
1 þ
In this equation, Ci is a proportionality factor, P1/2i the
microwave power at half saturation, and b the inhomo-
geneity factor taking in account the different type of spin
packets. Using this equation we fitted the saturation curves
obtained at g=2.07 (Ccopper =391, P1/2copper =11.9897,
bcopper=1 and Cradical=72, P1/2radical=1.4483, bradical
=
3) and 2.01 (Ccopper =156, P1/2copper = 11,99, bcopper = 1
and Cradical = 185,6, P1/2radical = 1.45, bradical = 3). This
result is coherent with a S=1/2 copper(II) and a radical.
This result supports a reductive cleavage of the S-S
bond of the cyclic disulfide 1c by two successive one-
electron transfers, as proposed in Scheme 2. In a first step,
incorporation of copper(I) into 1c should lead to a bis-
imine/bis-disulfide copper(I) complex, in which the CuI
ion is coordinated to the amide nitrogens under their
iminol forms and to the internal sulfur atom of each
disulfide unit. Deprotonation of the amides promotes the
first electron-transfer, yielding species 4, which can be
described as a bis-amidato/thiolato/disulfide copper(II)
complex, and a thyil radical. The second electron transfer
from the copper(II) results in the formation of the isolated
reaction products, the copper(III) complex 2 and the di-
tert-butyl disulfide.
Similarly incorporation of two Cu(I) ions in the cyclic
bis-disulfide 1a in CH3CN allows to detect by EPR at liquid
nitrogen temperature, only after addition of base (4 equiv
of Et4NOH/1a), a Cu(II) signal (Figure S1 in Supporting
Information). Using Cu(ClO4)2 as an internal standard,
we evaluated the Cu(II)/1a ratio to be 2. The EPR pattern
is very similar to that of the copper(II) complex in species
4 (Figure 1a). Such observations are consistent with a
binuclear Cu(II) complex 1a0 (Scheme 3) in which the two
copper cations are in a similar N2SS* tetragonal environ-
ment and bridged by one disulfide. The cleavage of one
disulfide bond of 1a should result from a one-electron
transfer from each Cu(I) cation. Allowing the tempera-
ture to reach room temperature, leads to the formation of
the copper(III) complex 2, after reductive cleavage of the
second S-S bridge in 1a0. Such a binuclear Cu(II) com-
plex, in which the two copper(II) sites are connected
Figure 1. (a) EPR spectrum of complex 6 in dashed line and of species 4
in black solid line; (b) Intensity of the EPR signal versus the square root of
the microwave power for the CuII complex of 4 at g = 2.07 (dotted gray
line) and tBuS• at g = 2.01 (solid black line), the g value of the power
saturation study are indicated by arrows.
has been attributed to the stronger donor ability of the
pyridine nitrogen in the latter ligand that enhances the
electron transfer from copper(I) to the disulfide leading to
the S-S bond scission.13b In our case, deprotonation of
the amide nitrogens is likely to promote the reductive
cleavage of the S-S bonds by enhancing the electron
density on the copper center, and to stabilize the copper-
(III) dithiolato complex. This reduction can occur by a
two-electron transfer from CuI, or by two one-electron
transfers leading to the intermediate formation of a
copper(II) complex. To discriminate between these two
processes, a mixture of 1c and (PF6)[CuI(CH3CN)4], in
acetonitrile was trapped at liquid nitrogen temperature
after the addition of 2 equiv of Et4NOH, and was ana-
lyzed by X-band EPR spectroscopy at 10 K. The EPR
spectrum, shown in Figure 1a, exhibits the typical pattern
of a copper(II) complex in a tetragonal geometry with
bands at g^=2.03 and g|| =2.12 and a hyperfine coupling
constant A|| =181 G as the best fit of the g-values, and an
additional signal at g=2.01 attributed to the thyil radical
tBuS•. The superimposition of the two paramagnetic
entities has been confirmed by recording the variation
of the intensities of the signals related to the two species
with the microwave power (P) between 0 and 200 mW and
(14) Frapart, Y.-M.; Boussac, A.; Anxolabehere-Mallard, E.; Delroissse,
M.; Verlhac, J.-B.; Blondin, G.; Girerd, J.-J.; Guilhem, J.; cesario, M.;
Rutherford, A. W.; Lexa, D. J. Am. Chem. Soc. 1996, 118, 2669–2678.
(15) Innes, J. B.; G. W. Brudvig, G. W. Biochemistry 1989, 28, 1116–1125.
(16) Merdy, P.; Guillon, E.; Frapart, Y.-M.; Aplincourt, M. New J.
Chem. 2003, 27, 577–582.

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Desbenoıt et al.
8640 Inorganic Chemistry, Vol. 49, No. 18, 2010
Scheme 2. Proposed Mechanism for the Reductive Metalation of the tert-Butyl Bis-Disulfide 1c with (PF6)[CuI(CH3CN)4]
Scheme 3. Proposed Structure of the Intermediate Trapped by EPR
during Copper Insertion in 1a
sample prepared as previously detailed by iron(III) insertion
in the dithiol N2S2H4.18
To incorporate iron in 1a, we turned to the tetraethyl-
ammonium iron(III) tetraethylthiolate (Et4N)[Fe(SEt)4]19
(Scheme 4). The bis-disulfide 1a was readily and quantita-
tively converted into 5 upon adding 2 equiv of (Et4N)[Fe-
(SEt)4] in DMF and 2 equiv of Et4NCl, as indicated by the
X-band EPR spectrum recorded at 5 K, which shows the
disappearance of the signals attributed to the HS ferric
state of (Et4N)[Fe(SEt)4] at g=4.2520 and the appearance
of the signals characteristic of the S=3/2 state of complex 5
(Figure S2, Supporting Information). Similarly the reac-
tion was monitored by electronic spectroscopy (Figure S3,
Supporting Information). For a set point of comparison, we
checked that reacting 1b with (Et4N)[Fe(SEt)4] (1 equiv)
in DMF also yields complex 5 (data not shown).
With Fe3CO12, the reductive metalation of 1b is likely
to involve an electron transfer from the iron(0) to the
disulfide, promoting the scission of the S-S bonds when
deprotonating the amides with NaOMe. This probably
leads to the formation of an iron(II) complex equivalent
to 5 (that has not been isolated), along with dimethyl
disulfide. Complex 5 is then formed upon air oxidation.
As previously detailed, the five-coordinate chloro species
is air stable.18,21 It is reduced at a very low potential ex-
plaining why the iron(II) form is so unstable and is readily
oxidized to the iron(III) state.18 The steric hindrance of
the bis-disulfide 1a probably disfavors the reaction with
Fe3CO12.
When using either 1a or 1b and (Et4N)[Fe(SEt)4], addi-
tion of base is not necessary. In this reaction the ethyl
thiolates ligands are both the reducing agent able to trigger
the S-S bond cleavage through several successive thio-
late/disulfide exchanges, and the base able to deprotonate
the amides. By forcing the cleavage of the S-S bonds,
they drive the reaction toward the formation of the stable
[(Et4N)][FeIII(N2S2)Cl], complex 5. Such thiolate/disulfide
exchange has been previously reported to induce the
interconversion of iron dinitrosyl complexes by reacting
through a disulfide bridge, has been previously prepared
from oxidized glutathione and crystallized. The magnetic
susceptibility and the EPR showed that the Cu(II);Cu(II)
interaction was negligibly small.17
Reductive Metalation of the Bis-Disulfides 1a-1c with
Triiron Dodecacarbonyl, Fe3CO12, or Iron(tetraethylthiolate)
Tetraethylammonium, (Et4N)[Fe(SEt)4]. Since simple iron(I)
complexes are not easily accessible, we first tried the meta-
lation experiments of the bis-disulfides with an iron(0) species,
Fe3CO12. These reactions were performed at 60 ꢀC in DMF
under argon, with 1a and 1b, and Fe3CO12 (2/3 equiv versus
1a and 1/3 equiv versus 1b) in the presence of sodium metha-
nolate as a base to deprotonate the amides (4 equiv relative to
1a and 2 equiv relative to 1b). After adding Et4NCl (4 equiv
relative to 1a and 2 equiv relative to 1b), the solution was
opened to the air. While there was no reaction from 1a, the
diamidato/dithiolato complex 5, [(Et4N)2][Fe(N2S2)Cl], was
isolated from 1b in a 33% yield after purification over
Sephadex LH20 (Scheme 4). The iron complex 5 was char-
acterized by mass spectrometry, UV-vis spectroscopy, and
EPR. Typically, it exhibits an absorption at 475 nm in DMF
solution, and its X-band EPR spectrum in frozen DMF at
5 K displays the two typical bands at g=4.52 and g=2.05
corresponding to an iron(III) center with a S = 3/2 ground
state. These characteristics are similar to those of an authentic
(18) Chatel, S.; Chauvin, A.-S.; Tuchagues, J.-P.; Leduc, P.; Bill, E.;
Chottard, J.-C; Mansuy, D.; Artaud, I. Inorg. Chim. Acta 2002, 336, 19–28.
(19) Maelia, L.; Millar, M.; Koch, S. A. Inorg. Chem. 1992, 31, 4594–
4600.
(20) Millar, M.; Lee, J. F.; O’Sullivan, T.; Koch, S. A.; Fikar, R. Inorg.
Chim. Acta 1996, 243, 333–343.
(21) Galardon, E.; Giorgi, M.; Artaud, I. Chem; Commun. 2004, 286–287.
(17) Miyoshi, K.; Sugiura, Y.; Ishizu, K.; Iitaka, Y.; Nakamura, H.
J. Am. Chem. Soc. 1980, 102, 6130–6136.

Article
Inorganic Chemistry, Vol. 49, No. 18, 2010 8641
Scheme 4. Reductive Metalation of Bis-Disulfides with Iron Species
R0S-SR0 with [Fe(SR)2(NO)2]- to yield [Fe(SR0)2(NO)2]-
and RS-SR.22
More interestingly, complexes 2 and 5 can also be con-
verted back to the acyclic bis-disulfides N2H2(S-S-CH3)2
1b and N2H2(S-S-tBu)2 1c. For this, 2 and 5 were first
treated with iodine in the presence of the dimethyl disul-
fide, 3b (Scheme 5). In both cases, no reaction takes place
in the absence of iodine. Oxidation of 5 with 1 equiv of I2
and 1 equiv of 3b affords a 1/1 mixture of 1a and 1b as
determined by 1H NMR analysis of the material isolated
after precipitation in cold diethyl ether (see Figure S4,
Supporting Information). Products 1a and 1b were iso-
lated in a 50 and 45% yield, respectively.
The reaction of 2 with 3b and iodine was monitored by
1H NMR in CD3CN. Figure 2 displays the aromatic
region of a typical spectrum obtained after addition of
iodine (2 equiv) and 3b (1 equiv relative to 2). It clearly
shows a mixture of three main products: the starting
complex 2, along with a mixture of the two disulfides 1a
and 1b in a 20/80 ratio. For an equimolar ratio of 3b and 2,
when increasing the I2/2 ratio from 1 to 3 the conversion
yield (see the Experimental Section) determined after a
reaction time of 22 h at room temperature, increases from
45 to 100% (I2/2: 1, 1.5, 2, and 3, conversion yield: 45, 60,
80, and 100%). In a preparative experiment with 1 equiv
of 3b and 3 equiv of iodine, 1b and 1a were isolated in a
50% and 8% yield after purification over silica gel,
respectively. No other product has been identified by
1H NMR. Using a 3b/2 ratio of 2 neither significantly in-
creases the conversion yield of 2, nor changes the selec-
tivity of the reaction.
Reoxidation of the Copper and Iron Complexes 2 and 5
into Cyclic and Acyclic Bis-Disulfides. Some years ago,
Fox et al.23 described the synthesis of macrocyclic bis-
(disulfide)tetramine ligands by oxidative coupling of the
corresponding bis-imine/bis-thiolate Ni complex [NiII(N2S2)]
or copper complex [CuII(N2S2)] with iodine. We used a
similar procedure to prepare 1a from the diamidato/
dithiolato nickel complex, [(Et4N)2][Ni(N2S2)]8, and here
we similarly submitted complexes 2 and 5 to iodine
oxidation in CH3CN at room temperature. Oxidation
of the iron complex 5 with 1 equiv of iodine affords, after
purification over silica gel, the cyclic bis-disulfide 1a in an
80% yield. Since iodine24 and bromine25 adducts of nickel
thiolate as well as nickel sulfenyl bromide25 were reported
in the literature, we suspected the intermediate formation
of an iron(III) sulfenyl iodide during the course of iodine
oxidation of 5, but unfortunately we did not succeed in
isolating any intermediate.
With 2, for unclear reasons which are very likely due to
a change in the redox state of the copper ion, the reaction
requires an excess of iodine for completion (3 equiv).
After purification, 1a was isolated with a yield similar to
that obtained from the iron complex 5. In contrast to Fox
et al.,23 we did not isolate any dinuclear complexes
derived from the cyclic bis-disulfide ligand but directly
the free ligand, indicating that this iodine oxidation of
diamidato/dithiolato complexes is associated to proton-
ation of the amidates, probably with the solvent, and to
release of iron or copper ions in solution. We have already
noticed that N2S2 complexes with carboxamido nitrogen
donors tend to lose their metal cation when the donating
properties of the sulfur atoms are too decreased upon
oxidation.26
Despite several attempts, we were unable to define the
final oxidation state of the copper at the end of the
reaction or to trap and identify at low temperature any
intermediate complexes. Nevertheless, it is reasonable to
postulate the involvement of a dimethyl disulfide iodo
sulfonium species,27 resulting from the reaction of 3b with
iodine. On the basis of the known reactivity of disulfide
halo sulfonium,28 it could react with a thiolate of 2 to
give the methyl sulfenyl iodide (CH3SI) and of a mono
S-methyldisulfidespecies. Then, thesecondthiolateshouldbe
(22) Tsai, F.-T.; Chiou, S.-J.; Tsai, M.-C.; Huang, H.-W.; Chang, M.-H.;
Liaw, W.-F. Inorg. Chem. 2005, 44, 5872–5881.
(23) Fox, S.; Stibrany, R. T.; Potenza, J. A.; Knapp, S.; Schugar, H. J.
Inorg. Chem. 2000, 39, 4950–4961.
(24) Lyon, E. J.; Musie, G.; Reibenspies, J. H.; Darensbourg, M. Y. Inorg.
Chem. 1998, 37, 6942–6946.
(27) Gooda, M.; Major, A.; Nag-Chaughuri, J.; McGlynn, S. P. J. Am.
Chem. Soc. 1961, 83, 4329–4333.
€
(25) Steinfeld, G.; Lozan, V.; Kruger, H.-J.; Kersting, B. Angew. Chem.,
(28) (a) Billard, T.; Langlois, B. R.; Large, S.; Anker, D.; Roidot, N.;
Roure, P. J. Org. Chem. 1996, 61, 7545–7550. (b) Doi, J. T.; Luehr, G. W.;
Musker, W. K. J. Org. Chem. 1985, 50, 5716–5719.
Int. Ed. 2009, 48, 1954–1957.
(26) Alves de Sousa, R.; Galardon, E.; Rat, M.; Giorgi, M.; Artaud, I.
J. Inorg. Biochem. 2005, 99, 690–697.

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8642 Inorganic Chemistry, Vol. 49, No. 18, 2010
Scheme 5. Conversion of the Iron and Copper Diamidato/Dithiolato Complexes Back to the Bis-Disulfide Ligands
Scheme 6. Proposed Mechanism for the Reaction of the Copper(II)
Complex 6 with tert-Butyl Sulfenyl Iodide
Figure 2. Typical 1H NMR spectra, recorded at 500 MHz, obtained
after addition of 1 equiv of dimethyl disulfide and 2 equiv of iodine to a
0.04 M solution of complex 2 in CD3CN (bottom), along with the spectra
of the authentic compounds 1a, 1b and 2 for comparison (top).
its reaction with tBuSI (10 equiv) selectively yields the bis-
disulfide 1c (Scheme 6). It is noteworthy that lowering the
amount of tBuSI allows the detection in the reaction mix-
ture of the copper(III) derivative 2 along with 1c (Figure S5,
Supporting Information). This suggests that thionylation
of 6 with tBuSI probably promotes, as shown in Scheme 6,
the intermediate formation of a CuII diamidato/thiolato/
S-disulfide and further the reductive cleavage of the S-S
bond and the formation of the CuIII complex 2 along with
a thyil radical that dimerizes into disulfide. Further reac-
tion of 2 with tBuSI yields as previously detailed the acyclic
bis-disulfide 1c.
thionylated by reaction with CH3SI, leading to the for-
mation of the observed bis-S-methyl disulfide 1b (see
Scheme S1 in the Supporting Information).
All attempts to convert back the copper or iron com-
plexes 2 and 5 into 1c by an analogous reaction with tBuS-
StBu 3c and iodine have failed, and we only observed the
cyclic bis-disulfide 1a. The di-tert-butyl disulfide iodo
sulfonium is probably too sterically hindered, and we
turned to the reaction with tert-butylsulfenyl iodide.
Hence, reaction of an excess of tBuSI (10 equiv) with 2
or 5 in CH3CN affords quantitatively and selectively the
di-tert-butyl disulfide 1c.
Conclusion
Finally, to assess the role of the CuIII oxidation state,
we carried out some control experiments with the copper-
(II) complex 6, [(Et4N)2][Cu(N2S2)], prepared as pre-
viously described.10 All the experiments were monitored
by 1H NMR in CD3CN. The NMR tubes were prepared
in a glovebox under strictly anaerobic conditions to avoid
any air oxidation of 6 into 2. The copper(II) complex 6 is
oxidized with 1 equiv of iodine into the copper(III) species 2,
as expected from the low CuII/CuIII redox potential.10
Like 2, the Cu(II) derivative 6 does not react with dimethyl
or di-tert-butyl disulfide in the absence of iodine. However,
In this paper, we have shown that pseudopeptidic bis-
disulfides built on an o-phenylene diamide scaffold, mimick-
ing a Cys*-X-Cys* tripeptide motif containing two cysteines
modified into disulfides, can be easily and directly metalated
under reductive conditions to produce the iron or copper
diamidato/dithiolato species (Et4N)2[FeIII(N2S2)Cl] or (Et4N)-
[CuIII(N2S2)]. Interestingly, these complexes revert back to
the starting cyclic bis-disulfide or acyclic bis-disulfide N2H2-
(SSR)2 when exposed to iodine or to a mixture of an exo-
genous disulfide RSSR and iodine, when this disulfide is not
too sterically hindered. With bulky R groups such as tBu, the

Article
Inorganic Chemistry, Vol. 49, No. 18, 2010 8643
corresponding sulfenyl iodide, tBuSI, appears as an alternative
to turn back to the bis-disulfide N2H2(S-S-tBu)2. Even though
iodine is not a biologically relevant oxidant, the reversibility
of the overall process, metalation/reoxidation, might be of
interest for the investigation of the reactivity of disulfides on
a peptide backbone with metal cations. Moreover, it is an
additional example of the reactivity of sulfur oxidized species
toward metal cations that we study from a general point of
view and that includes persulfides,29 pseudopeptidic sulfi-
nates,30 and pseudopeptidic cyclic thiosulfinates.31
chloride, CH3SO2Cl, (1.1 equiv, 1.74 mmol, 134 μL) in 3 mL
of CH2Cl2 was added over 20 min. After stirring for 16 h, the
solution was successively washed with water, saturated aqueous
NaHCO3 solution, then water, and dried over MgSO4. After eva-
porating the solvents, CH3SO2StBu was isolated as a white oil
(186 mg, 70% yield) and characterized by its 1H NMR as previ-
ously described: 1H NMR (δ, CDCl3): 3.37 (s, 3H), 1.63 (s, 18H).33
General Procedure for the Synthesis of the Acyclic Bis-Disulfide
Ligands. One equivalent of triethylamine (99 μL, 0.71 mmol)
was added to a CH2Cl2 solution (5 mL) of the alkyl methane
thiosulfonate (0.71 mmol, 72 μL of S-methyl methanethiosulfo-
nate or 119 mg of S-tert-butyl methanethiosulfonate). After
cooling the solution to 0 ꢀC, a solution of 0.45 equiv of N,N0-1,2-
phenylenebis(2-mercapto-2-methylpropanamide) (100 mg, 0.32
mmol) in CH2Cl2 (8 mL) was added dropwise over 30 min, and
the reaction mixture was allowed to warm to room temperature
and was stirred for 2 h. Volatiles were then removed under
vacuo. The residue was purified by column chromatography
(silica gel, 90/10 (v:v) CH2Cl2/EtOAc mixture) and was precipi-
tated in diethyl ether yielding the pure product as a white powder.
2-Methyl-N-(2-(3-methyl-3-(methyldisulfanyl)butan-2-ylamino)-
phenyl)-2-(methyldisulfanyl)propanamide (1b). Yield: 103 mg (80%).
Anal. Calcd (found) for C16H24N2O2S4: C, 47.37 (47.51); H, 5.98
(5.84); N, 6.92 (7.01). HRMS (CIþ, CH4, m/z): Calcd (found) for
[C16H24N2O2S4 þ Naþ]: 427.0618 (427.0621). 1H NMR (δ,
CD3CN): 8.67 (s, 2H), 7.53 (m, 2H), 7.27 (m, 2H), 2.43 (s, 6H),
1.66 (s, 12). 13C NMR (CD3CN): 173.2, 131.3, 126.4, 125.7, 54.0,
25.3, 25.1. FT-IR (ATR, cm-1): 1666.
Experimental Section
Physical Measurements. IR spectra were obtained with a
Perkin-Elmer Spectrum One FT-IR spectrometer equipped with
a MIRacleTM single reflection horizontal ATR unit (zirconium-
selenium crystal). ESI-MS mass spectra were recorded on a Thermo
Finnigan LCD Advantage spectrometer. UV spectra were recor-
ded on a SAFAS mc2 spectrometer. Elemental analyses and high
resolution ESI-MS mass analyses were carried out in CNRS at
Gif-sur-Yvette. 1H NMR and 13C NMR spectra were recorded
on a Bruker ARX-250 spectrometer or AVANCEII-500, with
chemical shifts reported in parts per million (ppm) relative to the
residual solvent (CH3CN, δH = 1.96 ppm). Cyclic voltammo-
grams were obtained in deaerated CH3CN solution at room
temperature using an EGGR-PAR model 173 potentiostat and
model interface instruments with a three electrode system which
consists of a NaCl saturated calomel electrode (SCE), a plati-
num auxiliary electrode, and a glassy carbon working electrode.
Tetrabutylammonium hexafluorophosphate (NBu4PF6, 0.1 M
in CH3CN) was used as the supporting electrolyte. The potential
sweep rate was 50 mV.s-1, and under our conditions the poten-
tial of the ferrocene/ferrocenium ion couple was 0.38 V. EPR
spectra were recorded on a Bruker Elexsys 500 EPR spectro-
meter operating at X-band frequency (9.44 GHz) equipped with
a shq0011 cavity fitted with an Oxford Instrument liquid helium
probe. The following instrument settings were used: field modu-
lation amplitude frequency of 100 kHz, field modulation ampli-
tude of 0.1 mT, time constant of 0.04 s, sampling time 41 ms,
8192 sampling points, field sweep 1 T, microwave power of 10 mW.
Xsophe has been used for EPR spectra simulation of the non
saturated experimental spectrum after subtraction of a solvent
tube spectrum. The g, D (E/D), and A values are those found to
reasonably fit the experimental spectrum with the calculated one
using a g and A strain line width mode.
2-(tert-Butyldisulfanyl)-N-(2-(3-(tert-butyldisulfanyl)-3-methyl-
butan-2-ylamino)phenyl)-2-methylpropanamide (1c). 125 mg, (80%).
HRMS (CIþ, CH4, m/z): Calcd (found) for [C22H36N2O2S4
þ
1
Naþ]: 511.1557 (511.1571). H NMR (δ, CD3CN): 8.80 (s, 2H),
7.54 (m, 2H), 7.25 (m, 2H), 1.65 (s, 12), 1.32 (s, 18H). 13C NMR
(CD3CN): 173.4, 131.3, 126.2, 125.6, 56.1, 47.7, 30.2, 25.8. FT-IR
(ATR, cm-1): 1663.
Reductive Metalation of the Bis-disulfide 1a-1c with (PF6)[Cu-
(CH3CN)4]. From 1a. In an acetonitrile solution (5 mL) of bis-
disulfide (310 mg, 0.5 mmol) were added successively under argon
tetrakis(acetonitrile)copper(I)hexafluorophosphate (PF6)[Cu(CH3-
CN)4] (372 mg, 1 mmol) and tetraethylammonium hydroxide in
methanol (2.0 mmol, 1.43 mL of a 1.4 M solution in methanol).
After stirring for 2 h at room temperature, the solvent was
removed. The crude product was dissolved in acetone (2 mL),
filtered over Celite, and the solvent evaporated to dryness. After
dissolving the product in the minimum amount of acetonitrile
(1 mL), the complex was precipitated several times in cold
diethyl ether, and pure compound 2 was isolated as a greenish
yellow powder in a quantitative yield. Analyses were identical to
those reported in the literature:10 UV (CH3CN) λmax: 417 nm
Materials. CH3CN, DMF, and MeOH were dried and dis-
tilled before use, following standard procedures. Anhydrous
€
diethyl ether was purchased from Riedel-de-Haen.
Synthesis. The bis-disulfide 1a was synthesized according to
1
(εM = 6130 M-1 cm-1); H NMR(δ, CD3CN): 8.48 (dd, 2H,
8
Bourles et al. N,N00-1,2-phenylenebis(2-mercapto-2-methylpro-
ꢁ
3JH-H=6.3 Hz and 4JH-H=3.5 Hz), 6.84 (dd, 2H, 3JH-H=6.3
panamide) was prepared according to Chatel et al.18 and com-
4
3
Hz and JH-H = 3.5 Hz), 3.18 (q, 8H, JH-H = 7.3 Hz), 1.51
(s, 12H), 1.23 (t, 12H, 3JH-H =7.3 Hz); MS(ESI-): 371 (100%,
[2-Et4N]-); E1/2 (CH3CN): -1.16 V versus the ferrocene/ferro-
cenium (Fc/Fcþ) redox couple.
plexes 2, 6, and 5 according to Hanss and Kruger,10 and Chatel
€
et al.,18 respectively. tert-Butyl sulfenyl iodide in solution in
CCl4 was prepared as described by Field et al.32 The concentra-
tion of the solution was estimated by H NMR using dioxane
1
From 1b or 1c. Using a similar procedure, 250 mg of 2 (100%)
were obtained starting from 1b (202 mg, 0.5 mmol) or 1c (244 mg,
0.5mmol) uponreactionwith(PF6)[Cu(CH3CN)4] (1 equiv, 184 mg,
0.5 mmol) and Et4NOH (1.4 M in methanol, 2 equiv, 1.0 mmol,
707 μL). From 1b the dimethyl disulfide 3b also formed and was
identified by GC, and from 1c, the di-tert-butyl disulfide 3c
also formed and was identified by 1H NMR (tBu: 1.34 ppm in
CD3CN).
(δ=3.57 ppm) as an internal standard (δ=1.45 ppm for tBuSI).
Synthesis of S-tert-butyl methane Thiosulfonate. To a solution
of tBuSH (100 μL, 1.58 mmol) in dry CH2Cl2 was added
triethylamine (1.1 equiv, 244 μL, 1.74 mmol); then the reaction
mixture was cooled to 0 ꢀC, and a solution of methane sulfonyl
(29) Galardon, E.; Tomas, A.; Selkti, M.; Roussel, P.; Artaud, I. Inorg.
Chem. 2009, 48, 5921–5927.
Reductive Metalation of 1b with Triiron Dodecacarbonyl. To a
solution of 1b (100 mg, 0.25 mmol) in 2 mL of DMF were
successively added under argon Fe3CO12 (43 mg, 0.08 mmol),
(30) Alves de Sousa, R.; Artaud, I. Tetrahedron 2008, 64, 2198–2206.
ꢁ
(31) (a) Bourles, E.; Alves de Sousa, R.; Galardon, E.; Artaud, I. Angew.
ꢁ
Chem., Int. Ed. 2005, 44, 6162–6165. (b) Galardon, E.; Bourles, E.; Artaud, I.;
Daran, J.-C.; Roussel, P.; Tomas, A. Inorg. Chem. 2007, 46, 4515–4522.
(32) Field, L.; Vanhorne, J. L.; Cunningham, L. W. J. Org. Chem. 1970,
35, 3267–3273.
(33) Block, E.; Bentley, M. D.; Davis, F. A.; Douglass, I. B.; Lacadielb,
J. A. J. Org. Chem. 1975, 40, 2770–2773.

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Desbenoıt et al.
8644 Inorganic Chemistry, Vol. 49, No. 18, 2010
sodium methoxide (1.0 M in methanol, 0.50 mmol, 500 μL), and
Et4NCl (82 mg, 0.50 mmol). After stirring for 4 h at 60 ꢀC, the
solvents were removed, and the crude product purified by
exclusion chromatography (Sephadex LH-20 eluted with
MeOH). The brown fraction was collected and found to have
the same spectroscopic properties as 518 (55 mg, 33% yield).
Reductive Metalation of 1a with (Et4N)[Fe(SEt)4]. To a solu-
tion of 1a (20 mg, 0.032 mmol) in DMF (1 mL) was added under
argon [(Et4N][Fe(SEt)4] (27.5 mg, 0.064 mmol) followed by
tetraethylammonium chloride (10.6 mg, 0.064 mmol). After
stirring for 5 h, the solvent was removed, and the resulting
orange powder analyzed as 5 (49 mg, quantitative yield).
Synthesis of the Bis-disulfides 1a and 1b by Reaction of Com-
plex 5 with Iodine or Iodine and Dimethyl Disulfide. To a solution
of complex 5 (100 mg, 0.15 mmol) in CH3CN (2 mL) were
successively added under argon iodine alone (38 mg, 0.15 mmol)
or dimethyl disulfide (266 μL, 3 mmol) and iodine (38 mg, 0.15
mmol). The mixture was stirred for 16 h at room temperature,
and the solvent removed. The crude product was dissolved in
dichloromethane and washed abundantly with water, aqueous
sodium thiosulfate (0.05 M), and water, and then was dried over
MgSO4. After removing the solvent to dryness, the products
were purified over column chromatography and identified by
comparing their 1H NMR data with those of authentic samples
prepared in this work (1b) or previously published (1a).8 Treat-
ment of 5 with I2 afforded only 1a, that was purified over
chromatography (silica gel, 80/20 (v:v) CH2Cl2/EtOAc mixture)
and isolated as a white powder (38 mg, 80% yield). Treatment of
5 with dimethyl disulfide 3b and iodine afforded a mixture of 1b
and 1a that was separated by elution over silica gel with a 90/10
(v:v) mixture of CH2Cl2/EtOAc for 1b and a 80/20 mixture for
1a. After removal of the solvents, 1b (30 mg, 50% yield) and 1a
(22 mg, 45% yield) were isolated as white powders.
Synthesis of the Bis-disulfide 1c by Reaction of Complexes 2 or
5, with tert-Butyl Sulfenyl Iodide. To an acetonitrile solution
(3 mL) of complexes 2 (100 mg, 0.2 mmol) or 5 (100 mg, 0.15
mmol) was added under argon an excess (10 equiv) of a 0.13 M
solution of tert-butyl sulfenyl iodide in CCl4 (2 mmol, 15 mL for
2 and 1,5 mmol, 11 mL for 5). The resulting mixture was stirred
for 16 h at room temperature in a dark room. The solvent was
removed, and after dissolving the crude product in CH2Cl2, the
solution was washed abundantly with water, sodium thiosulfate
(0.05 M), and water, and was then dried over MgSO4. Evapora-
tion of the solvent to dryness gave 1c as a white powder in a
quantitative yield (98 mg). 1H NMR and mass spectra of 1c were
identical to those of an authentic sample prepared in this work.
NMR Studies. Reactions from Complex 2. In a deuterated
acetonitrile solution (0.5 mL) of complex 2 (10 mg, 0.02 mmol)
were added 1 or 2 equiv of methyl disulfide as well as 1, 1.5, 2, or
3 equiv of iodine. The conversion yield of complex 2 was
determined after 22 h by 1H NMR by comparing the integration
ratio of the CH2 signal at 3.24 ppm of the countercation of 2,
Et4Nþ, used as an internal standard, and the aromatic signals of
complex 2 at 6.82 ppm.
Reaction from Complex 6. All these experiments were per-
formed in NMR tubes prepared in a glovebox under strictly
anaerobic conditions. Ten milligrams of the paramagnetic
complex 6 (0.016 mmol) were dissolved in 0.5 mL of CD3CN,
and 1 equiv of iodine or 1 equiv of dimethyl disulfide 3b or 4
equiv of tBuSI in solution in CCl4 were added.
Supporting Information Available: Synthesis of (Et4N)[Fe-
(SEt)4], Figures S1-S5 and Scheme S1 as a PDF file. This
material is available free of charge via the Internet at http://
pubs.acs.org.