Evidence for the Michaelis-Menton type mechanism in the electrocatalytic oxidation of mercaptopropionic acid by an amavadine model

Article abstract of DOI:10.1021/ja9607042

The amavadine complex and its model [VL₂]^2- (L = ^-ON[CH(CH³)COO^-]₂ (HIDPA^3-) or ^-ON(CH₂COO^-)₂ (HIDA^3-), respectively) undergo, in aqueous medium and at a Pt electrode, a fully electrochemical and chemical reversible V(iv/v) oxidation and act as electron-transfer mediators in the electrocatalytic oxidation of some thiols (HSR) such as HS(CH₂)(n)COOH (n = 1 or 2, i.e, mercaptoacetic or mercaptopropionic acid, respectively) and HSCH₂CH(NH₂)COOH (cysteine) to the corresponding disulfides (RS-SR) which were isolated upon bulk preparative electrolyses. As shown by digital simulation of cyclic voltammetry, this redox catalysis process occurs through an unprecedented mechanism involving Michaelis-Menten type kinetics with formation (k₁ = 1.2 x 10³ M^-1 s^-1) of an intermediate species (with half-life time of ca. 0.3 s) derived from the interaction of the oxidized vanadium complex (the active form of the mediator) with the substrate. A possible biological role for amavadine is suggested by these results.

Full text of DOI:10.1021/ja9607042

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7568
J. Am. Chem. Soc. 1996, 118, 7568-7573
Evidence for a Michaelis-Menten Type Mechanism in the
Electrocatalytic Oxidation of Mercaptopropionic Acid by an
AmaVadine Model
M. Fa´tima C. Guedes da Silva,† J. Armando L. da Silva,†
Joa˜o J. R. Frau´sto da Silva,*,† Armando J. L. Pombeiro,*,†
Christian Amatore,*,‡ and Jean-Noe1l Verpeaux‡
Contribution from the Complexo I, Instituto Superior Te´cnico, AV. RoVisco Pais,
1096 Lisboa Codex, Portugal, and Ecole Normale Supe´rieure, De´partement de Chimie,
URA CNRS 1679, 24 rue Lhomond, F-75231 Paris Cedex 05, France
ReceiVed March 4, 1996. ReVised Manuscript ReceiVed June 4, 1996X
Abstract: The amaVadine complex and its model [VL2]2- (L ) -ON[CH(CH3)COO-]2 (HIDPA3-) or -ON(CH2COO-)2
(HIDA3-), respectively) undergo, in aqueous medium and at a Pt electrode, a fully electrochemical and chemical
reversible Viv/v oxidation and act as electron-transfer mediators in the electrocatalytic oxidation of some thiols (HSR)
such as HS(CH2)nCOOH (n ) 1 or 2, i.e, mercaptoacetic or mercaptopropionic acid, respectively) and HSCH2CH-
(NH2)COOH (cysteine) to the corresponding disulfides (RS-SR) which were isolated upon bulk preparative electrolyses.
As shown by digital simulation of cyclic voltammetry, this redox catalysis process occurs through an unprecedented
mechanism involving Michaelis-Menten type kinetics with formation (k1 ) 1.2 × 103 M-1 s-1) of an intermediate
species (with half-life time of ca. 0.3 s) derived from the interaction of the oxidized vanadium complex (the active
form of the mediator) with the substrate. A possible biological role for amaVadine is suggested by these results.
Introduction
where the disulfide bridges would bring about the cross-linking
of protein fibers necessary for the regeneration of damaged
tissues.
The importance of vanadium in Biology is a matter of current
and growing interest, and the natural occurrence of this metal
has already been recognized in a few cases, in particular in the
cofactor of an alternative nitrogenase in some azotobacteria (e.g.,
A. Vinelandii and A. chroococcum), in haloperoxidases present
in some algae, lichen, and terrestrial fungi (e.g., Filariopsis
breVipes, Ascophyllum nodosum, etc.), in some ascideans
(tunicates) (e.g., Ascidia nigra, Phallusia mammillata, etc.) and
fan-worms (Pseudopotamilla occelata), and in some Amanita
toadstools (A. muscaria, A. regalis, and A. Velatipes).1-4
A natural vanadium (IV) complex called amaVadine could
be isolated from Amanita mushrooms.7 Firstly described as an
oxovanadium (IV) species,7 this compound has then been shown
to display a remarkable octacoordinated V4+ center with two
tribasic HIDPA3- ligands (tribasic form of 2,2′-(hydroxiimino)-
dipropionic acid HON{CH(CH3)CO2H}).5,6 Even if the biologi-
cal function of amaVadine has not yet been fully elucidated, it
has been assumed to play a significant role in the specific
oxidation of some thiols into disulfides. This could namely
happen in the protective/defensive system of the mushrooms8
Organic thiols or thiolates can be oxidized to disulfides by
simple electron uptake, by means of outersphere oxidants or at
the electrode.9 In this respect, amaVadine with its stable
octacoordinated structure could very well belong to the class
of transition metal centered natural compounds acting as pure
electron donors or acceptors, in the same way as blue copper
proteins, cytochromes, or iron sulfur clusters. This hypothesis
is even reinforced by the fully reversible cyclic voltammogram
obtained upon oxidation of amaVadine10a and the spectroscopic
data of this compound and its oxidation product,6 all studies
leading to an identical geometrical structure for both vanadium
(IV) and (V) derivatives.
In the search of better analytical methods for thiols, Riechel
et al. showed that amaVadine can indeed act as a redox mediator
for the oxidation of natural thiols such as glutathione or
cysteine.10b Although not orientated toward mechanistical goals,
some of the data therein suggested that a nonclassical redox
catalysis mechanism was operative. This looked rather surpris-
ing with regards to the expected outersphere oxidation mech-
anism.6 We therefore decided to investigate thoroughly the
mechanism of amaVadine mediated oxidation of biological thiols
in aqueous medium in order to bring evidence for a specific
interaction between the substrate and the oxidized form of the
mediator rather than a simple outersphere electron transfer. The
† Instituto Superior Te´cnico.
‡ Ecole Normale Supe´rieure.
X Abstract published in AdVance ACS Abstracts, July 15, 1996.
(1) Frau´sto da Silva, J. J. R.; Williams, R. P. The Biological Chemistry
of the Elements. The Inorganic Chemistry of Life; Clarendon Press: Oxford,
1993.
(2) Metal ions in Biological systems; Siegel, H., Siegel, A., Eds.; Marcel
Dekker, Inc.: 1995; Vol. 31.
(3) Frau´sto da Silva, J. J. R. et. al., work in progress.
(4) Vollenbroek, E. G. M.; Simons, L. H.; van Schijndel, J. W. P. M.;
Barnett, P.; Bolzar, M.; Dekker, H.; van der Linden, C.; Weber, R. Plant
Peroxidases, Struct. Mol. Biol. 1995, 23, 267.
(7) Bayer, E.; Kneifel, H. Z. Naturforsch. 1972, 27B, 207. Kneifel, H.;
Bayer, E. Angew. Chem., Int. Ed. Engl. 1973, 12, 508.
(8) Frau´sto da Silva, J. J. R. Chem. Speciation BioaVailability 1989, 1,
139. A similar function for vanadium in tunicates has also been proposed,
although the metal is in a lower oxidation state, see: Smith, K. J. Experientia
1989, 45, 452, and references therein.
(5) Carrondo, M. A. A. F. de C. T.; Duarte, M. T. L. S.; Pessoa, J. C.;
Silva, J. A. L.; Frau´sto da Silva, J. J. R.; Vaz, M. C. T. A.; Vilas-Boas, L.
F. J. Chem. Soc., Chem Commun. 1988, 1158.
(6) Armstrong, E. M.; Beddoes, R. L.; Calviou, L. J.; Charnock, J. M.;
Collison, D.; Ertok, N.; Naismith, J. H.; Garner, C. D. J. Am. Chem. Soc.
1993, 115, 807.
(9) Svensmark, B.; Hammerich, O. In Organic electrochemistry. An
introduction and a guide, 3rd ed.; Lund, H., Baizer, M. M., Eds.; Chapter
17. Oxidation of sulfur containing compounds; Marcel Dekker: New York,
1991; p 659.
(10) (a) Nawi, M. A.; Riechel, T. L. Inorg. Chim Acta 1987, 136, 33.
(b) Thackerey, R. D.; Riechel, T. L. J. Electroanal. Chem. 1988, 245, 131.
S0002-7863(96)00704-4 CCC: $12 00 © 1996 American Chemical Society

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Electrocatalytic Oxidation with an AmaVadine Model
J. Am. Chem. Soc., Vol. 118, No. 32, 1996 7569
results described in this work show that a Michaelis-Menten
type mechanism involving the interaction of the substrate with
the oxidized form of the mediator actually takes place. It is
established that such a mechanism, frequently invoked to
interprete the enzymatic activity,11 can also operate in electro-
catalytic processes, a result with evident significance in biology.
Results
Electrochemical Behavior of the Amavadine Complex and
Its Model. The electrochemical behavior of the complexes
[VL2]2- (L ) hidpa3- or HIDA3-) was studied by cyclic
voltammetry and controlled potential electrolysis, in 0.1 M KCl
aqueous solutions and at a platinum-disc (φ ) 0.5 mm) or -gauze
working electrode, respectively. The complexes undergo a
single-electron reversible oxidation (at E° ) 0.49 or 0.52 V Vs
SCE, respectively) corresponding to the Viv h Vv interconver-
sion (Figure 1). In agreement, the heterogeneous electron
transfers are fast, as indicated by peak-to-peak separation (∆Ep
) Epa - Epc) and half-width (∆Ep/2 ) Epa - Ep/2a) values (70
( 5 mV and 55 ( 5 mV, respectively, determined at a scan
rate of 0.2 V s-1) which are identical to those obtained for K4-
[Fe(CN)6] under the same experimental conditions. Moreover,
controlled potential electrolysis at a potential slightly higher than
Epa consumes 1 Faraday/mol to afford the Vv species, which is
unstable and consequently difficult to isolate since it slowly
reverts to the parent Viv presumably by oxidation of water. In
the scan rate range under study, 0.05-20 V s-1, the current
function is constant.
Electrochemical Behavior of [V(HIDA)2]2- in the Presence
of Mercaptopropionic Acid. The [VL2]2-/- systems of this
study are involved in the electrocatalytic oxidation of thiols such
as mercaptoacetic and mercaptopropionic acids [HSCH2COOH
and HS(CH2)2COOH, respectively] as well as, for example,
cysteine [HSCH2CH(NH2)COOH], in unbuffered solutions.12 It
was also observed that the electrocatalytic effect is completely
lost by replacement of the thiol group by an hydroxyl or an
alkylthioether moiety or of the apparently necessary activating
carboxylic group by an amine, a methyl, a methylenethiol, a
sulfonate, or an hydroxide. In fact, no catalytic effect was
detected for serine [HOCH2CH(NH2)COOH], methionine
[CH3S(CH2)2CH(NH2)COOH], 2-aminoethanethiol (HSCH2-
CH2NH2), propanethiol (HSCH2CH2CH3), 1,3-propanedithiol
(HSCH2CH2CH2SH), 2-mercaptoethanesulfonic acid, sodium
salt (HSCH2CH2SO3Na) (coenzyme M), or 2-mercaptoethanol
(HSCH2CH2OH).14
Figure 1. Cyclic voltammograms for the [V(HIDA)2]2- complex (solid
line) and for the vanadium complex/HS(CH2)2COOH system (symbols)
in 0.2 M KCl/H2O, at a platinum disc working electrode (φ ) 0.5 mm)
and at a scan rate of ν ) 0.2 V s-1. The γ values indicate the ratio
between the thiol and the vanadium complex concentrations.
the addition of increasing amounts of mercaptopropionic acid
(as indicated by the different values of the excess factor γ,
defined as the ratio between the substrate and the electron
mediator concentrations [S]/[M]) for which, in the absence of
the vanadium system, no direct anodic oxidation has been
detected. However, a simple EC catalytic mechanism15,16 was
in contradiction with the dramatic suppression effect on the
catalytic activity of the mediator (Figure 1) by a relatively high
thiol concentration. This is evident from the plot of (ic - io)/io
Vs [S] at different scan rates (Figure 2). It is also noteworthy
that the different plots (Figure 3) of ic/io Vs -log(ν) establish
that there is no saturation effect with the scan rate since (ic -
io)/io increases upon decreasing the scan rate even in the range
where the saturation effect with the substrate concentration is
observed.
The possibility of coordination of the thiol to any of the redox
forms of the vanadium complex was also taken into account.
However, the cyclic voltammetric investigation at high scan rates
(ν > 5 V s-1, i.e., when a completely chemically reversible
wave is observed for the Viv/Vv couple) indicated that the E°
of the Viv/Vv redox pair was not affected by the presence of
the thiol (γ e 8) as apparent by an identical value for the E°
and for the peak-to-peak separation.
As shown in Figure 1, the catalytic nature of the anodic waves
is evident from the enhancement of the anodic peak current upon
(11) Bailey, J. E.; Ollis, D. F. Biochemical Enginnering Fundamentals,
2nd ed.; MacGraw-Hill Book Company: New York, 1986.
(12) No pH effect on the electrochemical behavior of the vanadium
system was detected under our experimental conditions, either by using
the previously prepared complexes [VL2](H3O)2 (L ) HIDA3- or HIDPA3-),
see Experimental Section, pH ca. 5, or when generating in situ the [VL2]2-
species (pH ca. 2). This can be accounted for in terms of the following
arguments: (i) the stability constants of the amaVadine complex
[V(HIDPA)2]2- and its model [V(HIDA)2]2- are so high (greater than 1021,
see ref 13a) that they remain practically undissociated at quite low pH values
(even lower than those observed in our systems); (ii) for pH values (as
those of our experiments and in biological medium) well below the pKa
value of the functional thiol group of our substrates (e.g., 10.8 for
mercaptoacetic acid, see ref 13b), these are practically all in the acid (-SH)
form; (iii) the form of the carboxylic functional groups of our substrates
should not play a relevant role in view of the recognized (see also ref 10b)
activity for thiols with either a carboxylic or an ester group.
(13) (a) Anderegg, G.; Koch, E.; Bayer, E. Inorg. Chim. Acta 1987, 127,
183. (b) Smith, R. M.; Martell, A. E. Critical Stability Constants; Plenum
Press: New York: 1977; Vol. 3.
(14) Frau´sto da Silva, J. J. R.; Guedes da Silva, M. F. C.; da Silva, J. A.
L.; Pombeiro, A. J. L. Molecular Electrochemistry of Inorganic, Bioinorganic
and Organometallic Compounds; Kluwer Academic Publishers: The
Netherlands, 1993; p 411.
Preparative scale electrolysis was performed in the presence
of 8 equiv of the thiol (HSR) leading to a consumption of 9
(15) Andrieux, C. P.; Dumas-Bouchiat, J. M.; Save´ant, J. M. J.
Electroanal. Chem. 1980, 113, 1.
(16) Bard, A. J.; Faulkner, L. R. Electrochemical Methods. Fundamentals
and Applications; John Wiley & Sons: New York, 1980; p 438, Table II.2.1,
case 6.

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Guedes da SilVa et al.
process.15,16 Indeed, a saturation of the redox catalyst upon
increasing the substrate concentration is observed. Moreover,
this saturation effect with the thiol concentration is not associated
to a similar saturation upon decreasing the scan rate. This
indicates that the process at hand cannot be described in terms
of a classical redox catalysis mechanism17 which implies no
saturation effect with the scan rate or substrate concentration
provided that the substrate is in excess. It does not fit either to
a complicated redox catalysis involving a progressive quenching
of the mediator by formation of a stable product with the
activated substrate.18 This would imply a saturation of the redox
catalytic effect with both the scan rate and the concentration.
Moreover, and in contrast with the latter mechanism, product
analysis of the electrolysed solution of the thiol in the presence
of the vanadium complex yielded no indication of the formation
of a stable adduct between the mediator and the substrate.
We are then forced to conclude that the vanadium mediated
oxidation of thiols proceeds along a mechanism that differs
significantly from the redox catalysis mechanisms that have been
considered and investigated up to now.15-18
The detection of a saturation of the catalytic efficiency with
the substrate concentration associated with no saturation upon
increasing the time scale of the experiment (i.e., decreasing the
scan rate) is very reminiscent of the classical Michaelis-Menten
enzymatic kinetics.11 These observations suggest that the
oxidation of the thiol by the oxidized vanadium species proceeds
Via the transient formation of an adduct between the Vv species
and the thiol (see Scheme 1 where V stands for [VL2]2- with L
) HIDA3- or HIDPA3-).
Figure 2. Plot of the normalized peak current (ic - io)/io vs the substrate
concentration [S], for the [V(HIDA)2]2-/HS(CH2)2COOH system. Scan
rate of 0.05 (b), 0.1 (2), 0.2 (9), and 0.4 (() V s-1. [S] in M.
Concentration of the HIDA complex, c ) 8.4 mM.
Faraday/mol of vanadium complex. The final product, with low
solubility in the aqueous electrolyte solution, was isolated by
filtration and characterized by the usual techniques of elemental
microanalysis and IR spectroscopy. A comparison of the
obtained results with those of an authentic sample identified
the product as the dimer RS-SR (ca. 85% isolated average yield).
The electrochemistry of the amaVadine and its model in the
presence of other thiols (cysteine and mercaptoacetic acid)
essentially showed identical results, with the Vv species mediat-
ing their oxidation in a process involving a saturation with the
substrate concentration and not with the lowering of the potential
scan rate. Moreover, preparative scale electrolysis of each thiol
in the presence of the vanadium complex (γ ) 12 for cysteine,
γ ) 8 for mercaptoacetic acid) yielded the corresponding
dissulfides with the consumption of 1 e-/molecule of thiol in
addition to 1 e-/molecule of the vanadium complex.
Scheme 1
Viv - e- h Vv
Vv + HSR yk1z {V‚HSR}
k-1
Discussion
k2
iv
+
1
{V‚HSR}
98 V + RS-SR + H
2
The complexes [V(HIDA)2]2- and [V(HIDPA)2]2- undergo
a fully electrochemical and chemical reversible Viv/v oxidation
process (eq 1 for the former species) in the range of scan rates
under study (0.05-20 V s-1).
Indeed, the mechanism in Scheme 1 is qualitatively expected
to give rise to all effects that have been observed experimentally.
Thus, when the thiol is not in large excess, the regeneration of
the Viv species is kinetically controlled by the slow reaction of
the Vv complex with the thiol leading to the observation of a
classical kinetic behavior for redox catalysis.16 Increasing the
thiol concentration will result in a progressive shift of the kinetic
control of the regeneration process to the decomposition of the
transient adduct. Then the redox catalysis overall rate becomes
independent of the substrate concentration but remains depend-
ent on the scan rate. Note that this occurs only when k-1 , k2
and k1[HSR], since otherwise the formation of {V‚HSR} would
be equilibrated and no saturation with the substrate would be
observed (Vide infra).
-
[V(HIDA)2]2- y-e z [V(HIDA)2]-
(1)
In the presence of the investigated thiols HSR (mercaptopro-
pionic acid, mercaptoacetic acid and cysteine), the oxidized
vanadium species is able to promote their oxidation converting
them into the corresponding disulfide derivatives (eq 2).
1
2
HSR + [V(HIDA)2]- f RSSR + H+ + [V(HIDA)2]2-
(2)
Under preparative and transient electrochemical experiments,
combination of the processes in eqs 1 and 2 yields to mediated
oxidation of the thiols at the oxidation potential of the Viv
complex. This agrees with observations reported by others10
for the behavior of some biological thiols (glutathione, cysteine,
cysteine methyl ester, or penicillamine) in the presence of the
amaVadine complex, at a glassy carbon electrode, although the
mechanism was then not investigated. We now report the results
of the detailed mechanistic study we have performed on the
abovementioned electrocatalytic processes.
Cyclic voltammetric simulation, by explicit finite differences,
was used to validate quantitatively the process given in Scheme
1. The corresponding results are presented for mercaptopro-
pionic acid as solid curves in Figure 3. The best fit was obtained
for k1 ) 1.2 × 103 M-1 s-1, k2 ) 2.5 s-1, and k-1 , k2 , and
this yielded to an excellent agreement with the experimental
data at several concentrations of substrate, all over the range of
scan rates investigated. These results are also presented in
Figure 4 under a form reminiscent of a classical Lineweaver-
Burk plot11 used in the treatment of homogeneous Michaelis-
Our cyclic voltammetric investigation showed that despite
the facile mediated electro-oxidation of the thiols, the overall
process did not behave as expected for a classical redox catalytic
(17) Andrieux, C. P.; Blocman, C.; Dumas-Bouchiat, J. M.; Save´ant, J.
M. J. Am. Chem. Soc. 1979, 101, 3431.
(18) Pedersen, S. U. Acta Chim. Scand. 1987, A 41, 391.

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Electrocatalytic Oxidation with an AmaVadine Model
J. Am. Chem. Soc., Vol. 118, No. 32, 1996 7571
sequence in Scheme 1 is the complexation of the oxidized
vanadium species by the thiol substrate, since k1[HSR] , k2.
Then, the concentration of the intermediate complex {V‚HSR}
obeys the steady-state approximation and the sequence in
Scheme 1 becomes kinetically equivalent to the simplified one
in Scheme 3.
Scheme 3
Viv - e- h Vv
k1
9
Vv + HSR
8 Viv (+ products)
This latter sequence is also equivalent to a classical redox
catalysis mechanism and the kinetics depend only on k1[HSR]
(RT/Fν). The dependence of ic/io on the scan rate and the thiol
concentration could then be determined from previously reported
works.20,21 However, based on the values of k1 (1.2 × 103 M-1
s-1) and k2 (2.5 s-1) this would require (see above) a concentra-
tion of the thiol much less than the Michaelis constant Km
)
k2/k1 ) 2.1 mM, that is, outside the range of concentration that
could be investigated by our method. Therefore, our data in
Figure 4 correspond to an intermediate kinetic behavior between
the two limiting cases usually considered in Michaelis-Menten
kinetics. The slope could, however, be evaluated by means of
a simulation procedure, and its variation with the scan rate is
shown in Figure 5b. Again, a reasonable fit is obtained between
the experimental data and the theoretical prediction.
Figure 3. Plots of experimental (symbols) and simulated (lines) ic/io
vs -log(ν), for excess factor of 1 (a), 2 (b), 4 (c), 6 (d), 8 (e), (ν in V
s-1), and (f) superimposition of the simulated curves in (a-e).
In Scheme 1, a disulfide is produced. Although the mech-
anism of the S-S bond formation could not be investigated in
the present study, our results show that the formation of the
disulfide from the {V‚HSR} species obeys a rate determining
step that is first order in {V‚HSR} and zero order in thiol, and
in the vanadium Vv or Viv species. Thus the disulfide formation
Menten enzymatic kinetics. Despite the fact that, in electro-
chemical experiments, reaction kinetics occur in nonhomogeneous
conditions (because of the development of concentration profiles
as a function of the distance from the electrode surface), it is
observed that almost linear plots are obtained when 1/[(ic/io) -
1] is plotted as a function of 1/[S]. As shown in Figure 4, the
experimental values fit reasonably well (within the experimental
error)19 with the theoretical plot (solid line) obtained Via
simulation.
Of interest in the classical Lineweaver-Burk plot is the
intercept and the slope of the regression line that give access,
respectively, to k2 and k1.11 In the present case, due to the
electrochemical character of the method used here, the relation-
ship between the intercept and slope of the regression lines
(shown in Figure 4) and k1 or k2 is less straightforward. The
intercept corresponds to a kinetic situation in which all the Vv
mediator is trapped instantaneously to afford the complex {V‚-
HSR} because k1[HSR](RT/Fν) f ∞. In this case, the kinetic
sequence in Scheme 1 is kinetically equivalent to the simplified
sequence in Scheme 2 where V* represents the sum of the Vv
and {V‚HSR} species.
•
could occur by the intermediate production of SR radicals
followed by fast coupling.9 Alternatively, one could also
conceive that the intermediate species {V‚HSR} undergoes some
kind of reorganization (with the rate constant k2) followed by
rapid reaction with another thiol and elimination of the disulfide.
It must be stressed that only thiols with carboxylate (or ester)
groups exhibit the behavior described in the present work. The
presence of “activating” carbonyl seems therefore to be a
prerequisite for the electrochemical oxidation of thiols mediated
by amaVadine.
Conclusions
The results obtained in this work indicate that the electro-
catalytic oxidation of some thiols in the presence of amaVadine
(or a related Viv complex) occurs through a novel type of redox
catalysis mechanism characterized by a saturation effect with
(19) It is noteworthy that, due to its reciprocal nature, the Lineweaver-
Burk kind of plot presents an experimental validity only when ic/io differs
significantly from unity as evidenced by the large error bars in Figure 4
for 1/[(ic - io) - 1] > 5, i.e., 1 e ic/io e 1.2, since our estimated accuracy
is (3% on ic/io. For this reason Figure 4 is presented as two different
panels: (a) presents the raw results, and (b) only these which bear kinetic
sense based on the experimental error.
Scheme 2
Viv - e- f V*
(20) Andrieux, C. P.; Blocman, C.; Dumas-Bouchiat, J. M.; M’Halla,
F.; Save´ant, J. M. J. Electroanal. Chem. 1980, 113, 19.
k2
9
V*
8 Viv (+ products)
(21) Note that the general sequence in Scheme 1 leads to another limiting
case that we do not consider here because its predictions disagree with the
experimental observations. This third limiting case corresponds to a situation
where k2 , k-1. Then, when k-1RT/Fν . 1 and k1 RT/Fν . 1 , {V‚HSR}
is in rapid equilibrium with VV and HSR, this equilibrium being pulled
continuously to its right-hand side (Viz., to the formation of VIV and 1/2-
RSSR) by the continuous evolution of {V‚HSR}. Thus the system is
controlled by the parameter (k1[HSR]/k-1)k2RT/Fν. This predicts therefore
a variation of the catalytic activity similar to that in Scheme 3, Viz., that
could be determined from literature.20
Scheme 2 is equivalent to the classical catalytic ErC’i process.16
The current ratio ic/io is then only a function of k2RT/Fν. This
is what is observed in Figure 5a, where the solid line represents
the theoretical variation for k2 ) 2.5 s-1. The relationship
between the slope and k1 or k2 is less obvious. Indeed, at low
concentrations of the substrate, the rate determining step of the

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7572 J. Am. Chem. Soc., Vol. 118, No. 32, 1996
Guedes da SilVa et al.
Figure 4. Plot of experimental (symbols) and simulated (lines) 1/[(ic/io) - 1] vs 1/[S] for the [V(HIDA)2]2-/HS(CH2)2COOH system. Scan rate of
0.05 (b), 0.1 (2), 0.2 (9), and 0.4 (() V s-1. [S} in M.
a coordination to the metal due to the geometrical analogy with
the initial HIDPA ligand.
Experimental Section
General Considerations. The electrochemical experiments were
carried out on an EG&G PAR 173 potentiostat/galvanostat and an
EG&G PARC 175 Universal programmer or on an EG&G PAR 273
potentiostat/galvanostat connected to a 386-SX personal computer
through a GPIB interface. Cyclic voltammetry was undertaken in a
two-compartment three electrode cell, at a Pt disc working electrode
(φ ) 0.5 mm), probed by a Luggin capillary connected to a SCE
reference electrode, on deionized water containing 0.1 M potassium
chloride as supporting electrolyte. Controlled-potential electrolysis was
carried out in a three electrode H-type cell with platinum gauze working
and counter electrodes in compartments separated by a glass frit; a
Luggin capillary, probing the working electrode, was connected to a
silver-wire pseudoreference electrode. The electrochemical studies were
performed in an inert atmosphere (N2).
The mechanism of the electrocatalytic process was investigated by
simulation (program CVSIM22 ) of the voltammograms at different scan
rates and various concentrations of reagents.
Infrared spectra were run with a Perkin Elmer 683 spectrophotometer.
The vanadyl sulfate, potassium chloride, and the organic substrates
were commercially available; HIDPA (L-N-hydroxyimino-2,2′-dipro-
pionic acid) and HIDA (N-hydroxyiminodiacetic acid) were prepared
according to a published method.23
Sample Preparation. The compounds [VL2](H3O)2 [L ) -ON(CH-
Figure 5. (a) Plot of the experimental (bars) and simulated (line)
(CH3)COO-)2 or -ON(CH2COO-)2] were prepared by a modification
intercept of 1/[(ic/io) -1] vs 1/[S] (see Figure 4), as a function of log
of the procedure followed5 for the synthesis of the corresponding
(ν). [S] in M, ν in V s-1. (b) Plot of the experimental (bars) and
simulated (line) slope of 1/(ic/io) - 1] vs 1/[S] (see Figure 4), as a
ammonium
tetramethylammonium
salt,
[V(HIDA)2][NH4]-
function of log (ν). [S] in M, ν in V s-1
.
[NMe4]: vanadyl sulfate VOSO4 was used instead of the chloride
VOCl2, and treatment with [NMe4]OH and NH4OH was avoided.
Alternatively, they were generated in situ by adding (in a twofold
excess) the appropriate acid, HIDPA or HIDA, respectively, to a VOSO4
aqueous solution. The electrochemical behavior of the complexes was
found to be independent of their generation process.
the concentration of the substrate. The kinetic analysis estab-
lished that the electrogenerated active form of amaVadine acts
as an innersphere oxidant forming a complex with the thiol,
the half-life time of this intermediate being in the 0.3 s range.
The occurrence of this Michaelis-Menten process is of biologi-
cal significance: the limitation of the efficiency of the redox
catalysis due to the saturation effect is compensated by the
specificity of the process associated with the recognition of the
substrate. Indeed, special conditions have to be fulfilled for
the oxidation of a given thiol to proceed: the necessary presence
of a second functional group such as a carboxylic derivative
two or three bonds away from the sulfhydryl group may suggest
Preparative Scale Electrolysis of RS-SR (R ) CH2CH2COOH
or CH2COOH). A mixture of 0.020 g (0.050 mmol) of [V(HIDPA)2]-
(H3O)2 and 30.6 µL (0.35 mmol) of mercaptopropionic acid was
electrochemically oxidized in 15 cm3 of a 0.1 M KCl/H2O electrolyte
solution, at 0.51 V Vs SCE, until the complete conversion of the Viv
complex into the Vv analogue (consumption of 8 Faraday/mole), as
(22) Gosser, D. K., Jr.; Zhang, F. J. Electroanal. Chem. 1991, 38, 715.
(23) Kneifel, H.; Bayer, E. J. Am. Chem. Soc. 1986, 108, 3075.

+
+
Electrocatalytic Oxidation with an AmaVadine Model
J. Am. Chem. Soc., Vol. 118, No. 32, 1996 7573
shown by the cyclic voltammogram of the electrolyzed solution. The
electrolyzed colorless solution was then transferred to a Schlenck tube
and the solvent was slowly evaporated under vacuum until the formation
of a white solid which was isolated by filtration and dried. Yield 85%.
Anal. Calcd (found) for S2C6O4H10‚1/2H2O: C, 32.87 (32.72); H, 4.24
(4.79); S, 28.48 (29.22). IR (KBr pellet) ν(OH) and ν(CH2-S) 3100-
2550 cm-1; ν(CdO) 1685 cm-1 (strong); ν(C-S) 650-635 cm-1
(medium); ν(S-S) 535-510 cm-1 (weak).
from the electrolyzed solution. Its IR spectrum was identical to that
of a genuine sample of cystine.
Acknowledgment. This work has been partially supported
by CNRS and MESR (France), the PRAXIS XXI (Portugal)
program, and by the bilateral JNICT/CNRS Portugal-France
protocol of collaboration.
A similar procedure was followed for the electrochemical synthesis
of HOOCCH2SSCH2COOH (cystine) which precipitated spontaneously
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