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Anaerobic oxidation of cysteine to cystine by manganese(III) in aqueous acetic acid
Inorganica Chimica Acta 357 (2004) 41–50 www.elsevier.com/locate/ica
Anaerobic oxidation of cysteine to cystine by manganese(III) in aqueous acetic acid Christian W. Salamon, Reginald F. Jameson, Wolfgang Linert
*
Institute of Applied Synthetic Chemistry, Technical University of Vienna, Getreidemarkt 9/163, A-1060 Vienna, Austria Received 11 March 2003; accepted 7 June 2003
Abstract The anaerobic oxidation of cysteine, Cys, by Mn(III) in acetic acid solutions has been followed by use of a stopped-flow spectrophotometric method at a temperature of 20 °C. The formation and disappearance of the [Mn(OAc)2 Cys] complex was monitored at 350 nm. The rate depends strongly on the acetic acid concentration (and hence also on pH) and led to the conclusion that more than one cysteine-containing species was involved. These mono-cysteinyl complexes are formed by the loss of two protons from the cysteine – one from the – SH and the other from either the –NHþ 3 or, more likely, the –COOH which is partially protonated at the low pH values involved (0.5–2.5). The rate-determining reprotonation of the bound –COO (or –NH2 ) is then accompanied by internal electron transfer yielding Mn(II) and the cysteinyl radical, Cys, which then dimerises to form (inactive) cystine. At high acetic acid concentrations (60–90% AcOH) the tris-acetato species, [Mn(OAc)3 ], predominates together with some of the biscomplex, [Mn(OAc)2 ]þ , and the active species is [Mn(OAc)2 Cys] which decomposes with a rate constant of k2 ¼ 16:8 0:9 M1 s1 . At low acetic acid concentrations (20–30% AcOH) the mono-acetato species predominates and the reactive species is [Mn(OH)Cys] for which the rate of decomposition ¼ k20 ¼ ð1:32 0:11Þ 104 M1 s1 . The relative values of the rate constants obtained are discussed, as is the bonding of cysteine to manganese(III). Ó 2003 Elsevier B.V. All rights reserved. Keywords: Kinetics; Oxidation; Manganese(III) in acetic acid; Cysteine
1. Introduction The reaction of cysteine, Cys (which exists as the zwitter-ion, HS CH2 CH(NHþ 3 ) COO , in neutral solutions), with metal ions in the absence of oxygen has been the subject of many investigations [1,2] but no study of oxidation with MnIII has so far been reported. The overall reaction is 2MnIII þ 2Cysteine ! 2MnII þ Cystine þ 2Hþ but the actual reaction takes place by internal one-electron transfer within a manganese(III)–cysteine complex,
Abbreviation: Cys; Cysteine. Corresponding author. Tel.: +43-1-58801-15350; fax: +4315880116299. E-mail address: [email protected] (W. Linert). *
0020-1693/$ - see front matter Ó 2003 Elsevier B.V. All rights reserved. doi:10.1016/S0020-1693(03)00379-7
releasing the free-radical S CH2 CH(NHþ 3 ) COO . This free-radical then dimerises, without further involvement of metal ions, to yield cystine, þ OOC CHðNHþ 3 Þ CH2 S–S CH2 CH(NH3 ) COO . A major problem encountered when working with manganese(III) solutions is their ready disproportionation to give MnII and MnIV . We managed to obtain some data by using strong perchloric acid solutions containing a large excess of MnII , but obtaining completely stable solutions and monitoring the [Hþ ] at such low pH values (pH < 1) proved too difficult and although a value for the decomposition of the [Mn(OH)Cys] was obtained, it could not be considered very accurate (but see Section 5). It was therefore decided to stabilise the MnIII by means of complex formation and, after consulting the literature, it was decided to use the acetate complexes in strong acetic acid, as they are not only relatively stable, but also fairly reactive. We were also influenced, of course, by the fact that acetic acid
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C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
solutions of manganese(III) have been long used as an oxidant in organic chemistry (e.g., to oxidise aromatic hydrocarbons [3]). The stopped-flow measurements were carried out over a pH range of 0.6–2.5. Higher pH values could be achieved with the use of acetate buffers, but such solutions always led to slow precipitation.
2. Experimental Time-dependant reaction spectra were recorded by use of a diode-array stopped-flow spectrophotometer SF-61 supplied by Hi-Tech Scientific Ltd., Salisbury. All kinetic measurements were made at a constant wavelength (350 nm) and were carried out on a photomultiplier stopped-flow spectrophotometer SX18.MV supplied by Applied Photophysics Ltd., London. Cysteine solutions were made up with L -cysteine from Sigma. manganese(III)-acetate was prepared by dissolution of Mn(NO3 )2 4H2 O (Merck) in acetic anhydride (Merck) by a method described previously [4]. The solutions were deoxygenated for 20 min (Ar) prior to every kinetic run and then transferred to the stopped-flow apparatus in sealed syringes. The pH was obtained by measuring the pH of 1:1 mixtures of the two reactant solutions. The instrument used was a PHM 84 Research pH meter made by Radiometer, Copenhagen. 3. Results The use of a diode-array stopped-flow spectrometer enabled changes in the absorption spectrum of a react-
ing mixture to be followed and an example of the timedependent spectra obtained is displayed in Fig. 1. Although the absorption band at 475 nm (due to the trisacetato complex) is clearly visible, and falls over time, there is only a slight indication of another band at about 350 nm, not present in cysteine-free solutions, that we attribute to the [Mn(OAc)2 Cys] complex (This band is a shoulder on an absorption band due to other Mn(III) species). This assignment is confirmed by Fig. 2 in which a typical kinetic run is displayed. This shows an initial rise in absorption attributable to the initial formation of the complex followed by a fall indicating its subsequent disappearance. The wavelength of 350 nm was chosen in order to maximise the initial rise in absorption. This enables one to formulate the overall reaction in general terms as kþ1
k2
MnIII þ Cys MnIII Cys!MnII þ Cys k1
ð1Þ
As is evident from the kinetic curve, it was not possible to gain any information regarding the formation of the complex and hence a value for kþ1 . In order to work with a reduced pseudo-order, one of the two reactants was always used in large excess and the results for excess cysteine and excess MnIII are presented separately below. 3.1. Cysteine in excess 3.1.1. Dependence on cysteine concentration The MnIII and acetic acid concentrations were kept constant in order to examine the effect of varying the cysteine concentration and Table 1 lists some typical results. Unfortunately the carboxyl group of cysteine is partially protonated over the pH range studied and so
Fig. 1. Typical time-dependant reaction spectrum. Spectral scan from 300 to 600 nm First spectrum taken after 1.6 s. ([Cys]T ¼ 5 102 M; [MnIII ]T ¼ 1.5 103 M; 60% AcOH (10.5 M); pH 1.63).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
43
1
Absorbance
0.8
0.6
0.4
0.2
0 0
0.002
0.004
0.006
0.008
0.01
0.012
0.014
0.016
0.018
0.02
Time/s Fig. 2. Typical kinetic run. (k ¼ 350 nm; [Cys]T ¼ 5 102 M; [MnIII ]T ¼ 1.5 103 M; 60% AcOH (10.5 M); pH 1.63).
Table 1 Typical results illustrating the dependence of the pseudo first-order rate constants, k2obs , on [Cys] ([MnIII ]T ¼ 1.5 103 M; in 60% AcOH) [Cys]T (mM)
pH
[Hþ ] (M)
102 [Cys]/(1+95.5[Hþ ]) (M)
K2obs (s1 )
5.00
1.28 1.06
0.0525 0.0871
0.0832 0.0537
0.145 0.111
12.5
1.14
0.0724
0.1579
0.231
25.0
1.72 1.76 1.38 1.51 1.27
0.0191 0.0174 0.0417 0.0309 0.0537
0.8866 0.9400 0.5019 0.6327 0.4079
0.636 0.490 0.383 0.318 0.411
37.5
1.81 1.81 1.48
0.0155 0.0155 0.0331
1.5126 1.5126 0.9009
0.809 0.688 0.621
50.0
1.92 1.87 1.55 1.76 1.38
0.0120 0.0135 0.0282 0.0174 0.0417
2.3276 2.1851 1.3544 1.8800 1.0038
1.05 0.925 0.786 0.891 0.696
62.5
1.93 1.61
0.0117 0.0245
2.9453 1.8689
1.07 0.873
75.0
1.95 1.67 1.81 1.48
0.0112 0.0214 0.0155 0.0331
3.6205 2.4657 3.0253 1.8019
1.32 1.04 1.25 0.947
87.5
1.97 1.71
0.0107 0.0195
4.3246 3.0572
1.53 1.18
100
1.73 1.55
0.0186 0.0282
3.5993 2.7089
1.35 1.18
the cysteine speciation is pH dependent. However, this variation could be allowed for because the (micro) protonation constants are known [1,4].
KN
Protonation of the –NH2 : Cys2 þ Hþ Cys log KN ¼ 10:32
ð2Þ
44
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50 KS
Protonation of the –S : Cys þ Hþ Cys log KS ¼ 8:23 þ KO
Protonation of the –COO : Cys þ H Cys log KO ¼ 1:96
3.2. Mn(III) in excess ð3Þ
ð4Þ
The protonation responsible for the variation in pH is that of the carboxyl group, and so the total amount of cysteine employed, [Cys]T , was replaced by [Cys] for the purposes of determining the dependence on cysteine concentration where, from above ½CysT ¼ ½Cysf1 þ KO ½Hþ g:
ð5Þ
The plot of k2obs versus [Cys]T /{1 + KO [Hþ ]} (Fig. 3) exhibits a clear curvature. This effect of a deviation from second-order kinetics has been known for many years [5] to be ascribable to the formation of an intermediate during the reaction, and so this curvature is further confirmation of the proposed reaction scheme (1). 3.1.2. Dependence on [Hþ ] Table 2 summarises the results for a series of kinetic runs in which the cysteine concentration was kept constant while varying the acetic acid concentration; several series are included for cysteine concentration of 2.5 102 , 5.0 102 , and 7.5 102 M. As usual, the pH values reported are for 1:1 mixtures of the reacting solutions. Fig. 4 shows a plot of k2obs versus 1/[Hþ ] (this form of plot was used because [Cys] is inversely proportional to [Hþ ]) and is seen to be independent of total cysteine concentration. It also shows a concave increase with increasing 1/[Hþ ] (which demonstrates that k2obs must involve an inverse power series in [Hþ ]).
All measurements were performed using a cysteine concentration of 5 105 M and [MnIII ]T varied over the range (0.5–1.5) 103 M. The solutions contained from 30 to 80% of acetic acid (3.5–14 M), and a typical set of results is given in Table 3. In order to allow for the variation in [Hþ ] and [AcOH], k2obs /[AcOH] was plotted against [MnIII ]/[Hþ ] (Fig. 5) yielding, after an initial steep rise, a linear correlation between the variables. This shows that the rate of reaction is strongly dependent on [OAc ], but no more conclusions can be drawn from this presentation of the results.
4. Interpretation of the results 4.1. Excess cysteine The observed rate equation is d½coloured complex=dt ¼ k2obs ½MnIII T
and the fact that the end-point of a run is at zero absorbance shows that the coloured complex is acting as an indicator for [MnIII ] during the decomposition reaction. To explain the dependencies reported above, we begin by assuming that the reaction proceeds via protonation of the intermediate complex, yielding the theoretical rate expression d½MnIII =dt ¼ k2 ½MnðOAcÞ2 Cys ½Hþ :
1.600 1.400
obs
k2 /s
-1
1.200 1.000 0.800 0.600 0.400 0.200 0.000 0.50
1.00
1.50
2.00
ð7Þ
To simplify (7) we need to define the following equilibria and mass-balance equations.
1.800
0.00
ð6Þ
2.50
3.00
3.50
4.00
4.50
5.00
+
100[Cys]T/(1+KO[H ]) (M) Fig. 3. Dependence of k2obs on [Cys] ¼ [Cys]T /(1+KO [Hþ ]). ([MnIII ]T ¼ 1.5 103 M; [Cys]T ¼ 2.5 103 – 0.2 M; 60% AcOH (10.5 M)).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
45
Table 2 Typical values of k2obs with varying [AcOH] and [Cys]T ([MnIII ]T ¼ 1.5 103 M) AcOH (%)
[AcOH]T (M)
pH
[Hþ ] (mM)
10k2obs (s1 )
[Cys]T ¼ 0.025 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.26 2.16 1.98 1.75 1.56 1.47 1.38 1.28 1.14 1.1 1
5.5 6.9 10.6 17.9 27.4 34 42.1 52.1 71.8 80 99.3
16.4 12.3 7.57 5.67 3.60 2.15 1.98 1.43 1.10 0.913 0.724
[Cys]T ¼ 0.050 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.49 2.36 2.24 1.99 1.86 1.75 1.66 1.57 1.45 1.4 1.31
3.3 4.4 5.8 10.3 13.7 17.9 21.8 26.7 35.6 39.8 48.6
47.4 32.9 19.1 11.2 6.48 5.79 3.99 3.36 2.67 2.23 1.56
[Cys]T ¼ 0.075 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.58 2.41 2.23 2.08 1.97 1.83 1.76 1.68 1.54 1.49 1.32
2.6 3.9 5.9 8.3 10.7 14.8 17.5 20.9 28.7 32.1 48.3
61.8 36.5 28.5 14.2 8.44 7.23 5.39 4.68 3.70 3.22 1.83
70 60
10k 2obs/s
50 40 30 20 10 0 0
50
100
150
200
250
300
350
400
10 /[H+] (M ) -2
-1
Fig. 4. Dependence of k2obs on 1/[Hþ ]. ([MnIII ]T ¼ 1.5 103 M; [Cys]T ¼ 2.5 102 , 5 102 , and 7.5 102 M).
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C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
Table 3 Typical results for the pseudo first-order rate constant k2obs illustrating variation with [MnIII ]T ([Cys]T ¼ 5 105 M) [AcOH]T (M)
pH
[Hþ ] (mM)
10K2obs (s1 )
3.49 6.99 8.73 10.48 12.23 13.97
2.02 1.64 1.48 1.21 0.92 0.56
9.66 22.70 35.97 61.94 120.50 273.53
0.85 0.80 0.64 0.66 0.59 0.54
[MnIII ]T ¼ 0.75 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.65 1.45 1.21 0.92 0.06
9.40 22.65 35.81 61.80 119.95 877.20
1.01 1.00 0.97 0.83 0.76 0.79
[MnIII ]T ¼ 0.80 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.64 1.45 1.22 0.93 0.57
9.35 22.70 35.89 60.53 118.30 270.40
1.10 1.09 1.00 0.99 0.86 0.79
[MnIII ]T ¼ 1.00 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.66 1.45 1.22 0.93 0.58
9.27 22.13 35.48 60.12 117.76 265.46
1.24 1.19 1.17 1.10 1.06 1.05
[MnIII ]T ¼ 1.25 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.04 1.66 1.46 1.23 0.94 0.58
9.08 21.83 34.99 59.29 115.88 262.42
1.28 1.25 1.27 1.13 1.12 1.09
[MnIII ]T ¼ 1.50 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.04 1.66 1.46 1.23 0.94 0.59
9.08 21.68 34.75 58.88 115.08 255.27
1.59 1.45 1.39 1.35 1.32 1.25
AcOH (%) 3
III
[Mn ]T ¼ 0.50 10 20 40 50 60 70 80
M
Equilibria:
Mass balance equations: K
þ ðiÞ MnðOAcÞþ 2 þ Cys MnðOAcÞ2 Cys þ H
for which
¼ þ
K3
þ MnðOAcÞþ 2 þ AcOH MnðOAcÞ3 þ H
for which
½MnIII T ¼ ½MnðOAcÞ3 þ ½MnðOAcÞ2 Cys
K ¼ ½MnðOAcÞ2 Cys ½Hþ =½MnðOAcÞ2 ½Cys :
ðiiÞ
ð8Þ
K3
¼ ½MnðOAcÞ3 ½H ½AcOH:
þ
ð9Þ ð10Þ
þ K½Cys =½H g
ð11Þ
þ
þ
½CysT ¼ ½Cys KS ½H f1 þ KO ½H g:
ð12Þ ð13Þ ð14Þ
Substitution of these relationships into (7) yields
þ =½MnðOAcÞ2
þ ½MnðOAcÞ2 fK3 ½AcOH=½Hþ þ
d½MnIII k2 K½Hþ ½CysT ½MnIII T ¼ : þ dt K3 K½H ð1 þ KO ½Hþ Þ½AcOH þ K½CysT ð15Þ
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
47
0.6
-1 -1
10k2obs/[AcOH] T (M s )
0.5 0.4 0.3 0.2 0.1 0 0
0.0005
0.001 -
0.0015
0.002
+
10 ² [Mn]T/[H ] Fig. 5. Dependence of k2obs =½AcOHT on [MnIII ]T /[Hþ ]. ([Cys]T ¼ 5 105 M; [MnIII ]T ¼ 5 104 –1.5 103 M; [AcOH] ¼ 3.5–14 M ¼ 20–80% AcOH).
1012 M3 s, and hence, from slope/intercept, K3 /K ¼ (1.61 0.71) 1010 M2 . The data from solutions of low acetic acid concentrations (20–30%) are represented, mutates mutandis, in Fig. 7. If, under these conditions, it is assumed that the active species is [Mn(OH)Cys], accompanied by [MnOAc]2þ , then the same expression (16) is obtained with K3 being replaced by K1 and k2 by k20 . The intercept ¼ 1=k20 ¼ ð7:57 0:42Þ 105 Ms making k20 ¼ ð1:32 0:11Þ 104 M1 s1 . The slope¼ K1 =Kk20 ¼ ð1:00 0:06Þ 1011 s which makes K1 /K ¼ (1.32 0.11) 107 M1 .
From which it follows that ½Hþ =k2obs ¼ K3 KS ½Hþ ð1 þ KO ½Hþ Þ ½AcOH=k2 K½CysT þ 1=k2 :
ð16Þ
Since the protonation constants of cysteine and of acetic acid are known, it is possible to plot [Hþ ]/k2obs versus KS [Hþ ](1 + KO [Hþ ])[AcOH]/[Cys]T . Fig. 6 shows the result of this plot, and we draw attention to the results obtained for high acetic acid concentrations (>12 M) which can be seen to obey (16). From (16) the intercept ¼ 1=k2 ¼ ð5:94 0:35Þ 102 M, making k2 ¼ 16:8 0:9 M1 s1 . The slope¼K3 =Kk2 ¼ ð9:56 0:64Þ
1
+
10 [H ]/k 2
obs
(Ms)
1.5
0.5
0 0
2
4
6 10
8
10
12
14
+
10 KS(1+KO[H ])[AcOH]/[Cys]T
Fig. 6. Dependence of ½Hþ =k2obs on [Hþ ]KS (1+KO [Hþ ])[AcOH]/[Cys]T . ([MnIII ]T ¼ 1.5 103 M; [cys]T ¼ 5 102 M; (s) ¼ high [AcOH] ¼ 10.5–15.7 M ¼ 60–90% AcOH; (d) ¼ lower [AcOH] ¼ 3.5–15.7 M ¼ 20–60% AcOH).
48
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50 4 3.5
2.5 2
3
+
10 [H ]/k 2
obs
(Ms)
3
1.5 1 0.5 0 0
0.5
1 8
10 [H
1.5 +
2
2.5
3
+ H ]K1 (1+KO[H ])[AcOH]/[Cys]T
Fig. 7. Dependence of ½Hþ =k2obs on [Hþ ]KS (1+KO [Hþ ])[AcOH]/[Cys]T for low values of [AcOH] (20–30% AcOH ¼ 3.5–5.2 M). ([MnIII ]T ¼ 1.5 103 M; [cys]T ¼ 5 102 M).
4.2. Excess MnIII The first point to note is that in this series of experiments the k2obs values (Table 3) are considerably lower than those for the runs carried out with excess cysteine (Table 2), but this is not significant as the speciation is very different. Unfortunately, it has often been reported [6,7] that MnIII dissolved in acetic acid contains polymeric species, mainly dimers and trimers, but no quantitative data seem to have been published. It was decided, therefore, to proceed on the assumption that these were minority species. In fact, the interpretation of
the data on this basis seems to produce constants that agree with those obtained using excess cysteine. The observed rate expression is d½Coloured complex=dt ¼ k2obs ½CysT
ð17Þ
(It takes this form because now, with excess MnIII , the coloured complex is tracking the disappearance of the cysteine.) We now re-write the theoretical rate expression (7) as (18) d½Cys=dt ¼ k2 ½MnðOAcÞ2 Cys ½Hþ :
ð18Þ
1
+2
[H ] /k2
obs
2
(M s)
1.5
0.5
0 0
5
10 4
+
+
15
20
III
10 [H ](1+KO[H ])[AcOH]/[Mn ]T Fig. 8. Dependence of ½Hþ 2 =k2obs on [Hþ ](1+KO [Hþ ])[AcOH]/[MnIII ]T . ([Cys]T ¼ 5 105 M; [MnIII ]T ¼ 0.5 103 – 1.5 103 M; [AcOH] ¼ 3.5–14 M ¼ 30–80% AcOH).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
In order to simplify the resulting expression, we will re0 place the equilibrium constant K in Eq. (9) by K ¼ K/KS yielding þ
2
ð19Þ K 0 ¼ ½MnðOAcÞ2 Cys ½Hþ =½MnðOAcÞ2 ½Cys: Furthermore, the mass balance for MnIII is now ð20Þ ½MnIII T ¼ ½MnðOAcÞ2 Cys þ ½MnðOAcÞ3 : Making the required substitutions into (18) and rearranging enables the following expression for k2obs to be extracted 2
½Hþ =k2obs ¼ K3 ½Hþ ð1 þ KO ½Hþ Þ ½AcOH=k2 ½MnIII þ K 0 =k2 : þ 2
/k2obs
þ
ð21Þ þ
A plot of [H ] versus [H ](1 + KO [H ])[AcOH]/ [MnIII ] (Fig. 8) yields a straight line with slope ¼ K3 =k2 ¼ ð7:28 0:65Þ 106 M1 s1 and intercept ¼ K 0 =k2 ¼ ð2:02 0:83Þ 104 M2 s. Although a value of the rate constant cannot be extracted from these values, 0 the slope/intercept ¼ K3 /K ¼ (3.64 0.19) 102 M. 0 Since K ¼ K/KS and KS ¼ 108:32 , this makes K3 / K ¼ (1.74 0.88) 1010 M which is in excellent agreement with the value of (1.61 0.71) 1010 M obtained using the excess cysteine data. 5. Discussion 5.1. The bonding of cysteine to Mn(III) Spectroscopic evidence has been reported [8,9] that cysteine exhibits O,N chelation to MnIII (However, note that IR spectroscopy is unlikely to show any S-bonded species as these decompose rapidly.). In the case of iron(III) the bonding has been demonstrated [2] to be almost quantitatively via S,N chelation, but MnIII is a much ÔharderÕ species and so it could be that the active species in this case are S,O bonded. If, however, the same kinetics applied at higher pH, then since the second proton loss can then have been only from the amino group, this would confirm N,S bonding. Unfortunately the required kinetic studies for this to be examined were not possible due to precipitation from the solutions at higher pH. The spectroscopic evidence [8,9] does, however, imply that a considerable amount of the cysteine is O,N chelated to the MnIII and these species are expected to be kinetically inactive. (This is also supported by the fact that the kinetic evidence is that [Mn(OAc)2 Cys ] [Mn(OAc)2þ ].) Because this cannot be allowed for in the treatment of the kinetic data we must conclude that the rate constants here reported will have been underestimated (see also the discussion, below). 5.2. Evaluation of the rate constants It is interesting to note that the bonding of acetate ions to the MnCys entity reduces the rate of reaction
49
drastically (from k20 ¼ ð1:32 0:11Þ 104 M1 s1 for [Mn(OH)Cys] to k2 ¼ 16:8 0:9 M1 s1 for the decomposition of [Mn(OAc)2 Cys ]). This effect of the coordination of other species to the reactive centre has also been observed in the iron(III)–cysteine system [2] in that the [FeCys]þ species formed in acid solution reacts very much faster than the [Fe(OH)Cys] species that is formed at a higher pH. This is probably due to partial electron transfer to the MnIII from the negatively charged ligands. Unfortunately, this effect could not be extended by examining the complex [Mn(OAc)Cys] formed in the intermediate range of acetic acid concentrations because the formation constants of the acetato-complexes are not available. Work in these laboratories was also carried out using strong perchloric acid solutions stabilised by addition of a large excess of MnII . The result of these studies yielded value of ca. k2 ¼ 1:11 104 M1 s1 which is in good agreement with the present work, but because of the difficulty of obtaining reliable [Hþ ] values at pHÕs < 1 the agreement is perhaps rather more surprising than confirming! The accepted value of (1.32 0.11) 104 M1 s1 is interesting when compared with the value of k2 ¼ 1:08 105 M1 s1 obtained [2] for the decomposition of [Fe(Cys)]þ in which N,S bonding predominates [1,2]. Even allowing for the lowering of the rate because of the bonding of OH to the MnIII , it seems that the rate for electron transfer to MnIII in [Mn(OH)Cys] is much slower than expected. This underestimation is readily explicable, and indeed expected, if attention is once again drawn to the spectroscopic evidence [8,9] mentioned above that there is a considerable amount of O,N chelation involved. Since any such species are also formed by the loss of two protons they would not affect the form of the equilibria discussed in this work. It would, however, mean that the amount of reactive species present has been significantly overestimated leading to the calculation of low rate-constants from the experimental data.
Acknowledgements Thanks for financial support are due to the ‘‘Fonds zur F€ orderung der Wissenschaftlichen Forschung € sterreich’’ (Project 15874-NO3). Dr. E. Herlinger in O is thanked for proposing this study and for useful discussions.
References [1] R.F. Jameson, W. Linert, Monatsh. 122 (1991) 877. [2] R.F. Jameson, W. Linert, A. Tschinkowitz, V. Gutmann, J. Chem. Soc., Dalton Trans. (1988) 943.
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C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
[3] E.I. Heiba, R.M. Dessau, W.J. Koehl Jr., J. Am. Chem. Soc. 91 (1969) 138. [4] A. Albert, Biochem. J. 50 (1952) 690. [5] W.G. M€ ovius, R.G. Linck, J. Am. Chem. Soc. 91 (1969) 5394. [6] S. Chandraju, B.S. Sherigara, Int. J. Chem. Kinet. 26 (1994) 1105.
[7] B.S. Sherigara, K.I. Bhat, I. Pinto, Int. J. Chem. Kin. 27 (1995) 675. [8] H. Guntner, K.E. Schwarzhans, Z. Naturforsch. 31b (1964) 448. [9] M. Kaneko, N. Ishihara, A. Yamada, Makromol. Chem. 182 (1981) 89.
Anaerobic oxidation of cysteine to cystine by manganese(III) in aqueous acetic acid Christian W. Salamon, Reginald F. Jameson, Wolfgang Linert
*
Institute of Applied Synthetic Chemistry, Technical University of Vienna, Getreidemarkt 9/163, A-1060 Vienna, Austria Received 11 March 2003; accepted 7 June 2003
Abstract The anaerobic oxidation of cysteine, Cys, by Mn(III) in acetic acid solutions has been followed by use of a stopped-flow spectrophotometric method at a temperature of 20 °C. The formation and disappearance of the [Mn(OAc)2 Cys] complex was monitored at 350 nm. The rate depends strongly on the acetic acid concentration (and hence also on pH) and led to the conclusion that more than one cysteine-containing species was involved. These mono-cysteinyl complexes are formed by the loss of two protons from the cysteine – one from the – SH and the other from either the –NHþ 3 or, more likely, the –COOH which is partially protonated at the low pH values involved (0.5–2.5). The rate-determining reprotonation of the bound –COO (or –NH2 ) is then accompanied by internal electron transfer yielding Mn(II) and the cysteinyl radical, Cys, which then dimerises to form (inactive) cystine. At high acetic acid concentrations (60–90% AcOH) the tris-acetato species, [Mn(OAc)3 ], predominates together with some of the biscomplex, [Mn(OAc)2 ]þ , and the active species is [Mn(OAc)2 Cys] which decomposes with a rate constant of k2 ¼ 16:8 0:9 M1 s1 . At low acetic acid concentrations (20–30% AcOH) the mono-acetato species predominates and the reactive species is [Mn(OH)Cys] for which the rate of decomposition ¼ k20 ¼ ð1:32 0:11Þ 104 M1 s1 . The relative values of the rate constants obtained are discussed, as is the bonding of cysteine to manganese(III). Ó 2003 Elsevier B.V. All rights reserved. Keywords: Kinetics; Oxidation; Manganese(III) in acetic acid; Cysteine
1. Introduction The reaction of cysteine, Cys (which exists as the zwitter-ion, HS CH2 CH(NHþ 3 ) COO , in neutral solutions), with metal ions in the absence of oxygen has been the subject of many investigations [1,2] but no study of oxidation with MnIII has so far been reported. The overall reaction is 2MnIII þ 2Cysteine ! 2MnII þ Cystine þ 2Hþ but the actual reaction takes place by internal one-electron transfer within a manganese(III)–cysteine complex,
Abbreviation: Cys; Cysteine. Corresponding author. Tel.: +43-1-58801-15350; fax: +4315880116299. E-mail address: [email protected] (W. Linert). *
0020-1693/$ - see front matter Ó 2003 Elsevier B.V. All rights reserved. doi:10.1016/S0020-1693(03)00379-7
releasing the free-radical S CH2 CH(NHþ 3 ) COO . This free-radical then dimerises, without further involvement of metal ions, to yield cystine, þ OOC CHðNHþ 3 Þ CH2 S–S CH2 CH(NH3 ) COO . A major problem encountered when working with manganese(III) solutions is their ready disproportionation to give MnII and MnIV . We managed to obtain some data by using strong perchloric acid solutions containing a large excess of MnII , but obtaining completely stable solutions and monitoring the [Hþ ] at such low pH values (pH < 1) proved too difficult and although a value for the decomposition of the [Mn(OH)Cys] was obtained, it could not be considered very accurate (but see Section 5). It was therefore decided to stabilise the MnIII by means of complex formation and, after consulting the literature, it was decided to use the acetate complexes in strong acetic acid, as they are not only relatively stable, but also fairly reactive. We were also influenced, of course, by the fact that acetic acid
42
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
solutions of manganese(III) have been long used as an oxidant in organic chemistry (e.g., to oxidise aromatic hydrocarbons [3]). The stopped-flow measurements were carried out over a pH range of 0.6–2.5. Higher pH values could be achieved with the use of acetate buffers, but such solutions always led to slow precipitation.
2. Experimental Time-dependant reaction spectra were recorded by use of a diode-array stopped-flow spectrophotometer SF-61 supplied by Hi-Tech Scientific Ltd., Salisbury. All kinetic measurements were made at a constant wavelength (350 nm) and were carried out on a photomultiplier stopped-flow spectrophotometer SX18.MV supplied by Applied Photophysics Ltd., London. Cysteine solutions were made up with L -cysteine from Sigma. manganese(III)-acetate was prepared by dissolution of Mn(NO3 )2 4H2 O (Merck) in acetic anhydride (Merck) by a method described previously [4]. The solutions were deoxygenated for 20 min (Ar) prior to every kinetic run and then transferred to the stopped-flow apparatus in sealed syringes. The pH was obtained by measuring the pH of 1:1 mixtures of the two reactant solutions. The instrument used was a PHM 84 Research pH meter made by Radiometer, Copenhagen. 3. Results The use of a diode-array stopped-flow spectrometer enabled changes in the absorption spectrum of a react-
ing mixture to be followed and an example of the timedependent spectra obtained is displayed in Fig. 1. Although the absorption band at 475 nm (due to the trisacetato complex) is clearly visible, and falls over time, there is only a slight indication of another band at about 350 nm, not present in cysteine-free solutions, that we attribute to the [Mn(OAc)2 Cys] complex (This band is a shoulder on an absorption band due to other Mn(III) species). This assignment is confirmed by Fig. 2 in which a typical kinetic run is displayed. This shows an initial rise in absorption attributable to the initial formation of the complex followed by a fall indicating its subsequent disappearance. The wavelength of 350 nm was chosen in order to maximise the initial rise in absorption. This enables one to formulate the overall reaction in general terms as kþ1
k2
MnIII þ Cys MnIII Cys!MnII þ Cys k1
ð1Þ
As is evident from the kinetic curve, it was not possible to gain any information regarding the formation of the complex and hence a value for kþ1 . In order to work with a reduced pseudo-order, one of the two reactants was always used in large excess and the results for excess cysteine and excess MnIII are presented separately below. 3.1. Cysteine in excess 3.1.1. Dependence on cysteine concentration The MnIII and acetic acid concentrations were kept constant in order to examine the effect of varying the cysteine concentration and Table 1 lists some typical results. Unfortunately the carboxyl group of cysteine is partially protonated over the pH range studied and so
Fig. 1. Typical time-dependant reaction spectrum. Spectral scan from 300 to 600 nm First spectrum taken after 1.6 s. ([Cys]T ¼ 5 102 M; [MnIII ]T ¼ 1.5 103 M; 60% AcOH (10.5 M); pH 1.63).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
43
1
Absorbance
0.8
0.6
0.4
0.2
0 0
0.002
0.004
0.006
0.008
0.01
0.012
0.014
0.016
0.018
0.02
Time/s Fig. 2. Typical kinetic run. (k ¼ 350 nm; [Cys]T ¼ 5 102 M; [MnIII ]T ¼ 1.5 103 M; 60% AcOH (10.5 M); pH 1.63).
Table 1 Typical results illustrating the dependence of the pseudo first-order rate constants, k2obs , on [Cys] ([MnIII ]T ¼ 1.5 103 M; in 60% AcOH) [Cys]T (mM)
pH
[Hþ ] (M)
102 [Cys]/(1+95.5[Hþ ]) (M)
K2obs (s1 )
5.00
1.28 1.06
0.0525 0.0871
0.0832 0.0537
0.145 0.111
12.5
1.14
0.0724
0.1579
0.231
25.0
1.72 1.76 1.38 1.51 1.27
0.0191 0.0174 0.0417 0.0309 0.0537
0.8866 0.9400 0.5019 0.6327 0.4079
0.636 0.490 0.383 0.318 0.411
37.5
1.81 1.81 1.48
0.0155 0.0155 0.0331
1.5126 1.5126 0.9009
0.809 0.688 0.621
50.0
1.92 1.87 1.55 1.76 1.38
0.0120 0.0135 0.0282 0.0174 0.0417
2.3276 2.1851 1.3544 1.8800 1.0038
1.05 0.925 0.786 0.891 0.696
62.5
1.93 1.61
0.0117 0.0245
2.9453 1.8689
1.07 0.873
75.0
1.95 1.67 1.81 1.48
0.0112 0.0214 0.0155 0.0331
3.6205 2.4657 3.0253 1.8019
1.32 1.04 1.25 0.947
87.5
1.97 1.71
0.0107 0.0195
4.3246 3.0572
1.53 1.18
100
1.73 1.55
0.0186 0.0282
3.5993 2.7089
1.35 1.18
the cysteine speciation is pH dependent. However, this variation could be allowed for because the (micro) protonation constants are known [1,4].
KN
Protonation of the –NH2 : Cys2 þ Hþ Cys log KN ¼ 10:32
ð2Þ
44
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50 KS
Protonation of the –S : Cys þ Hþ Cys log KS ¼ 8:23 þ KO
Protonation of the –COO : Cys þ H Cys log KO ¼ 1:96
3.2. Mn(III) in excess ð3Þ
ð4Þ
The protonation responsible for the variation in pH is that of the carboxyl group, and so the total amount of cysteine employed, [Cys]T , was replaced by [Cys] for the purposes of determining the dependence on cysteine concentration where, from above ½CysT ¼ ½Cysf1 þ KO ½Hþ g:
ð5Þ
The plot of k2obs versus [Cys]T /{1 + KO [Hþ ]} (Fig. 3) exhibits a clear curvature. This effect of a deviation from second-order kinetics has been known for many years [5] to be ascribable to the formation of an intermediate during the reaction, and so this curvature is further confirmation of the proposed reaction scheme (1). 3.1.2. Dependence on [Hþ ] Table 2 summarises the results for a series of kinetic runs in which the cysteine concentration was kept constant while varying the acetic acid concentration; several series are included for cysteine concentration of 2.5 102 , 5.0 102 , and 7.5 102 M. As usual, the pH values reported are for 1:1 mixtures of the reacting solutions. Fig. 4 shows a plot of k2obs versus 1/[Hþ ] (this form of plot was used because [Cys] is inversely proportional to [Hþ ]) and is seen to be independent of total cysteine concentration. It also shows a concave increase with increasing 1/[Hþ ] (which demonstrates that k2obs must involve an inverse power series in [Hþ ]).
All measurements were performed using a cysteine concentration of 5 105 M and [MnIII ]T varied over the range (0.5–1.5) 103 M. The solutions contained from 30 to 80% of acetic acid (3.5–14 M), and a typical set of results is given in Table 3. In order to allow for the variation in [Hþ ] and [AcOH], k2obs /[AcOH] was plotted against [MnIII ]/[Hþ ] (Fig. 5) yielding, after an initial steep rise, a linear correlation between the variables. This shows that the rate of reaction is strongly dependent on [OAc ], but no more conclusions can be drawn from this presentation of the results.
4. Interpretation of the results 4.1. Excess cysteine The observed rate equation is d½coloured complex=dt ¼ k2obs ½MnIII T
and the fact that the end-point of a run is at zero absorbance shows that the coloured complex is acting as an indicator for [MnIII ] during the decomposition reaction. To explain the dependencies reported above, we begin by assuming that the reaction proceeds via protonation of the intermediate complex, yielding the theoretical rate expression d½MnIII =dt ¼ k2 ½MnðOAcÞ2 Cys ½Hþ :
1.600 1.400
obs
k2 /s
-1
1.200 1.000 0.800 0.600 0.400 0.200 0.000 0.50
1.00
1.50
2.00
ð7Þ
To simplify (7) we need to define the following equilibria and mass-balance equations.
1.800
0.00
ð6Þ
2.50
3.00
3.50
4.00
4.50
5.00
+
100[Cys]T/(1+KO[H ]) (M) Fig. 3. Dependence of k2obs on [Cys] ¼ [Cys]T /(1+KO [Hþ ]). ([MnIII ]T ¼ 1.5 103 M; [Cys]T ¼ 2.5 103 – 0.2 M; 60% AcOH (10.5 M)).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
45
Table 2 Typical values of k2obs with varying [AcOH] and [Cys]T ([MnIII ]T ¼ 1.5 103 M) AcOH (%)
[AcOH]T (M)
pH
[Hþ ] (mM)
10k2obs (s1 )
[Cys]T ¼ 0.025 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.26 2.16 1.98 1.75 1.56 1.47 1.38 1.28 1.14 1.1 1
5.5 6.9 10.6 17.9 27.4 34 42.1 52.1 71.8 80 99.3
16.4 12.3 7.57 5.67 3.60 2.15 1.98 1.43 1.10 0.913 0.724
[Cys]T ¼ 0.050 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.49 2.36 2.24 1.99 1.86 1.75 1.66 1.57 1.45 1.4 1.31
3.3 4.4 5.8 10.3 13.7 17.9 21.8 26.7 35.6 39.8 48.6
47.4 32.9 19.1 11.2 6.48 5.79 3.99 3.36 2.67 2.23 1.56
[Cys]T ¼ 0.075 M 20 30 40 50 60 65 70 75 80 85 90
3.49 5.24 6.99 8.73 10.48 11.35 12.23 13.1 13.97 14.85 15.72
2.58 2.41 2.23 2.08 1.97 1.83 1.76 1.68 1.54 1.49 1.32
2.6 3.9 5.9 8.3 10.7 14.8 17.5 20.9 28.7 32.1 48.3
61.8 36.5 28.5 14.2 8.44 7.23 5.39 4.68 3.70 3.22 1.83
70 60
10k 2obs/s
50 40 30 20 10 0 0
50
100
150
200
250
300
350
400
10 /[H+] (M ) -2
-1
Fig. 4. Dependence of k2obs on 1/[Hþ ]. ([MnIII ]T ¼ 1.5 103 M; [Cys]T ¼ 2.5 102 , 5 102 , and 7.5 102 M).
46
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
Table 3 Typical results for the pseudo first-order rate constant k2obs illustrating variation with [MnIII ]T ([Cys]T ¼ 5 105 M) [AcOH]T (M)
pH
[Hþ ] (mM)
10K2obs (s1 )
3.49 6.99 8.73 10.48 12.23 13.97
2.02 1.64 1.48 1.21 0.92 0.56
9.66 22.70 35.97 61.94 120.50 273.53
0.85 0.80 0.64 0.66 0.59 0.54
[MnIII ]T ¼ 0.75 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.65 1.45 1.21 0.92 0.06
9.40 22.65 35.81 61.80 119.95 877.20
1.01 1.00 0.97 0.83 0.76 0.79
[MnIII ]T ¼ 0.80 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.64 1.45 1.22 0.93 0.57
9.35 22.70 35.89 60.53 118.30 270.40
1.10 1.09 1.00 0.99 0.86 0.79
[MnIII ]T ¼ 1.00 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.03 1.66 1.45 1.22 0.93 0.58
9.27 22.13 35.48 60.12 117.76 265.46
1.24 1.19 1.17 1.10 1.06 1.05
[MnIII ]T ¼ 1.25 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.04 1.66 1.46 1.23 0.94 0.58
9.08 21.83 34.99 59.29 115.88 262.42
1.28 1.25 1.27 1.13 1.12 1.09
[MnIII ]T ¼ 1.50 103 M 20 40 50 60 70 80
3.49 6.99 8.73 10.48 12.23 13.97
2.04 1.66 1.46 1.23 0.94 0.59
9.08 21.68 34.75 58.88 115.08 255.27
1.59 1.45 1.39 1.35 1.32 1.25
AcOH (%) 3
III
[Mn ]T ¼ 0.50 10 20 40 50 60 70 80
M
Equilibria:
Mass balance equations: K
þ ðiÞ MnðOAcÞþ 2 þ Cys MnðOAcÞ2 Cys þ H
for which
¼ þ
K3
þ MnðOAcÞþ 2 þ AcOH MnðOAcÞ3 þ H
for which
½MnIII T ¼ ½MnðOAcÞ3 þ ½MnðOAcÞ2 Cys
K ¼ ½MnðOAcÞ2 Cys ½Hþ =½MnðOAcÞ2 ½Cys :
ðiiÞ
ð8Þ
K3
¼ ½MnðOAcÞ3 ½H ½AcOH:
þ
ð9Þ ð10Þ
þ K½Cys =½H g
ð11Þ
þ
þ
½CysT ¼ ½Cys KS ½H f1 þ KO ½H g:
ð12Þ ð13Þ ð14Þ
Substitution of these relationships into (7) yields
þ =½MnðOAcÞ2
þ ½MnðOAcÞ2 fK3 ½AcOH=½Hþ þ
d½MnIII k2 K½Hþ ½CysT ½MnIII T ¼ : þ dt K3 K½H ð1 þ KO ½Hþ Þ½AcOH þ K½CysT ð15Þ
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
47
0.6
-1 -1
10k2obs/[AcOH] T (M s )
0.5 0.4 0.3 0.2 0.1 0 0
0.0005
0.001 -
0.0015
0.002
+
10 ² [Mn]T/[H ] Fig. 5. Dependence of k2obs =½AcOHT on [MnIII ]T /[Hþ ]. ([Cys]T ¼ 5 105 M; [MnIII ]T ¼ 5 104 –1.5 103 M; [AcOH] ¼ 3.5–14 M ¼ 20–80% AcOH).
1012 M3 s, and hence, from slope/intercept, K3 /K ¼ (1.61 0.71) 1010 M2 . The data from solutions of low acetic acid concentrations (20–30%) are represented, mutates mutandis, in Fig. 7. If, under these conditions, it is assumed that the active species is [Mn(OH)Cys], accompanied by [MnOAc]2þ , then the same expression (16) is obtained with K3 being replaced by K1 and k2 by k20 . The intercept ¼ 1=k20 ¼ ð7:57 0:42Þ 105 Ms making k20 ¼ ð1:32 0:11Þ 104 M1 s1 . The slope¼ K1 =Kk20 ¼ ð1:00 0:06Þ 1011 s which makes K1 /K ¼ (1.32 0.11) 107 M1 .
From which it follows that ½Hþ =k2obs ¼ K3 KS ½Hþ ð1 þ KO ½Hþ Þ ½AcOH=k2 K½CysT þ 1=k2 :
ð16Þ
Since the protonation constants of cysteine and of acetic acid are known, it is possible to plot [Hþ ]/k2obs versus KS [Hþ ](1 + KO [Hþ ])[AcOH]/[Cys]T . Fig. 6 shows the result of this plot, and we draw attention to the results obtained for high acetic acid concentrations (>12 M) which can be seen to obey (16). From (16) the intercept ¼ 1=k2 ¼ ð5:94 0:35Þ 102 M, making k2 ¼ 16:8 0:9 M1 s1 . The slope¼K3 =Kk2 ¼ ð9:56 0:64Þ
1
+
10 [H ]/k 2
obs
(Ms)
1.5
0.5
0 0
2
4
6 10
8
10
12
14
+
10 KS(1+KO[H ])[AcOH]/[Cys]T
Fig. 6. Dependence of ½Hþ =k2obs on [Hþ ]KS (1+KO [Hþ ])[AcOH]/[Cys]T . ([MnIII ]T ¼ 1.5 103 M; [cys]T ¼ 5 102 M; (s) ¼ high [AcOH] ¼ 10.5–15.7 M ¼ 60–90% AcOH; (d) ¼ lower [AcOH] ¼ 3.5–15.7 M ¼ 20–60% AcOH).
48
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50 4 3.5
2.5 2
3
+
10 [H ]/k 2
obs
(Ms)
3
1.5 1 0.5 0 0
0.5
1 8
10 [H
1.5 +
2
2.5
3
+ H ]K1 (1+KO[H ])[AcOH]/[Cys]T
Fig. 7. Dependence of ½Hþ =k2obs on [Hþ ]KS (1+KO [Hþ ])[AcOH]/[Cys]T for low values of [AcOH] (20–30% AcOH ¼ 3.5–5.2 M). ([MnIII ]T ¼ 1.5 103 M; [cys]T ¼ 5 102 M).
4.2. Excess MnIII The first point to note is that in this series of experiments the k2obs values (Table 3) are considerably lower than those for the runs carried out with excess cysteine (Table 2), but this is not significant as the speciation is very different. Unfortunately, it has often been reported [6,7] that MnIII dissolved in acetic acid contains polymeric species, mainly dimers and trimers, but no quantitative data seem to have been published. It was decided, therefore, to proceed on the assumption that these were minority species. In fact, the interpretation of
the data on this basis seems to produce constants that agree with those obtained using excess cysteine. The observed rate expression is d½Coloured complex=dt ¼ k2obs ½CysT
ð17Þ
(It takes this form because now, with excess MnIII , the coloured complex is tracking the disappearance of the cysteine.) We now re-write the theoretical rate expression (7) as (18) d½Cys=dt ¼ k2 ½MnðOAcÞ2 Cys ½Hþ :
ð18Þ
1
+2
[H ] /k2
obs
2
(M s)
1.5
0.5
0 0
5
10 4
+
+
15
20
III
10 [H ](1+KO[H ])[AcOH]/[Mn ]T Fig. 8. Dependence of ½Hþ 2 =k2obs on [Hþ ](1+KO [Hþ ])[AcOH]/[MnIII ]T . ([Cys]T ¼ 5 105 M; [MnIII ]T ¼ 0.5 103 – 1.5 103 M; [AcOH] ¼ 3.5–14 M ¼ 30–80% AcOH).
C.W. Salamon et al. / Inorganica Chimica Acta 357 (2004) 41–50
In order to simplify the resulting expression, we will re0 place the equilibrium constant K in Eq. (9) by K ¼ K/KS yielding þ
2
ð19Þ K 0 ¼ ½MnðOAcÞ2 Cys ½Hþ =½MnðOAcÞ2 ½Cys: Furthermore, the mass balance for MnIII is now ð20Þ ½MnIII T ¼ ½MnðOAcÞ2 Cys þ ½MnðOAcÞ3 : Making the required substitutions into (18) and rearranging enables the following expression for k2obs to be extracted 2
½Hþ =k2obs ¼ K3 ½Hþ ð1 þ KO ½Hþ Þ ½AcOH=k2 ½MnIII þ K 0 =k2 : þ 2
/k2obs
þ
ð21Þ þ
A plot of [H ] versus [H ](1 + KO [H ])[AcOH]/ [MnIII ] (Fig. 8) yields a straight line with slope ¼ K3 =k2 ¼ ð7:28 0:65Þ 106 M1 s1 and intercept ¼ K 0 =k2 ¼ ð2:02 0:83Þ 104 M2 s. Although a value of the rate constant cannot be extracted from these values, 0 the slope/intercept ¼ K3 /K ¼ (3.64 0.19) 102 M. 0 Since K ¼ K/KS and KS ¼ 108:32 , this makes K3 / K ¼ (1.74 0.88) 1010 M which is in excellent agreement with the value of (1.61 0.71) 1010 M obtained using the excess cysteine data. 5. Discussion 5.1. The bonding of cysteine to Mn(III) Spectroscopic evidence has been reported [8,9] that cysteine exhibits O,N chelation to MnIII (However, note that IR spectroscopy is unlikely to show any S-bonded species as these decompose rapidly.). In the case of iron(III) the bonding has been demonstrated [2] to be almost quantitatively via S,N chelation, but MnIII is a much ÔharderÕ species and so it could be that the active species in this case are S,O bonded. If, however, the same kinetics applied at higher pH, then since the second proton loss can then have been only from the amino group, this would confirm N,S bonding. Unfortunately the required kinetic studies for this to be examined were not possible due to precipitation from the solutions at higher pH. The spectroscopic evidence [8,9] does, however, imply that a considerable amount of the cysteine is O,N chelated to the MnIII and these species are expected to be kinetically inactive. (This is also supported by the fact that the kinetic evidence is that [Mn(OAc)2 Cys ] [Mn(OAc)2þ ].) Because this cannot be allowed for in the treatment of the kinetic data we must conclude that the rate constants here reported will have been underestimated (see also the discussion, below). 5.2. Evaluation of the rate constants It is interesting to note that the bonding of acetate ions to the MnCys entity reduces the rate of reaction
49
drastically (from k20 ¼ ð1:32 0:11Þ 104 M1 s1 for [Mn(OH)Cys] to k2 ¼ 16:8 0:9 M1 s1 for the decomposition of [Mn(OAc)2 Cys ]). This effect of the coordination of other species to the reactive centre has also been observed in the iron(III)–cysteine system [2] in that the [FeCys]þ species formed in acid solution reacts very much faster than the [Fe(OH)Cys] species that is formed at a higher pH. This is probably due to partial electron transfer to the MnIII from the negatively charged ligands. Unfortunately, this effect could not be extended by examining the complex [Mn(OAc)Cys] formed in the intermediate range of acetic acid concentrations because the formation constants of the acetato-complexes are not available. Work in these laboratories was also carried out using strong perchloric acid solutions stabilised by addition of a large excess of MnII . The result of these studies yielded value of ca. k2 ¼ 1:11 104 M1 s1 which is in good agreement with the present work, but because of the difficulty of obtaining reliable [Hþ ] values at pHÕs < 1 the agreement is perhaps rather more surprising than confirming! The accepted value of (1.32 0.11) 104 M1 s1 is interesting when compared with the value of k2 ¼ 1:08 105 M1 s1 obtained [2] for the decomposition of [Fe(Cys)]þ in which N,S bonding predominates [1,2]. Even allowing for the lowering of the rate because of the bonding of OH to the MnIII , it seems that the rate for electron transfer to MnIII in [Mn(OH)Cys] is much slower than expected. This underestimation is readily explicable, and indeed expected, if attention is once again drawn to the spectroscopic evidence [8,9] mentioned above that there is a considerable amount of O,N chelation involved. Since any such species are also formed by the loss of two protons they would not affect the form of the equilibria discussed in this work. It would, however, mean that the amount of reactive species present has been significantly overestimated leading to the calculation of low rate-constants from the experimental data.
Acknowledgements Thanks for financial support are due to the ‘‘Fonds zur F€ orderung der Wissenschaftlichen Forschung € sterreich’’ (Project 15874-NO3). Dr. E. Herlinger in O is thanked for proposing this study and for useful discussions.
References [1] R.F. Jameson, W. Linert, Monatsh. 122 (1991) 877. [2] R.F. Jameson, W. Linert, A. Tschinkowitz, V. Gutmann, J. Chem. Soc., Dalton Trans. (1988) 943.
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[3] E.I. Heiba, R.M. Dessau, W.J. Koehl Jr., J. Am. Chem. Soc. 91 (1969) 138. [4] A. Albert, Biochem. J. 50 (1952) 690. [5] W.G. M€ ovius, R.G. Linck, J. Am. Chem. Soc. 91 (1969) 5394. [6] S. Chandraju, B.S. Sherigara, Int. J. Chem. Kinet. 26 (1994) 1105.
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