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Redox properties of echinochrome A
JOURNAL OF
ELSEVIER
Journal of Electroanalytical Chemistry 384 (1995) 131-137
Redox properties of echinochrome A S.A. Petrova, O.S. Ksenzhek, M.V. Kolodyazhny Ukrainian State Unicersity of Chemical Technology, Dniepropetrocsk 32000.5, Ukraine
Received 3 June 1994; in revised form 19 August 1994
Abstract The electrochemical properties of polyhydroxynaphthoquinone from the shells of sea-urchins Strongylocentrotus intermedius (echinochrome A) and related compounds were investigated using thin layer voltammetry on pyrolytic graphite electrodes. On the basis of experimental data pH-potential diagrams for ecbinochrome A and its trimethyl ether are plotted. Five different forms of echinochrome A and six forms of the trimethyl ether of echinochrome A are shown to exist over the whole range of pH studied. Equilibrium constants and formal redox potentials for all conjugated forms are obtained. Electrochemical oxidation of echinochrome A and its trimethyl ether lead to the formation of different products. Tetraketotetralin is formed in the former case and naphthodiquinone in the latter. Keywords: Redox properties; Echinochrome A
1. Introduction Quinones are widespread in the cells of living organisms where they perform various biochemical functions. Several types of quinone act as components of electron transport chains in the respiratory and photosynthetic systems. Some quinones possess the properties of vitamins and drugs. Quinones are also found to play the role of "chemical weapon" for some insects and as toxins of plants. Naturally occurring quinones show a great diversity of structure, including various benzo-, naphtho- and anthraquinones. Among them is a group of polyhydroxynaphthoquinones which includes echinochrome A and a number of spinochromes. These quinones are incorporated in appreciable amounts (0.01% to 0.04% by weight) in the eggs, needles and shells of sea-urchins [1]. Their biological functions are not yet established and their chemical features, especially electrochemical features, have been scarcely studied. Nevertheless it is known that echinochrome A and some spinochromes possess pronounced activity as antioxidants and bactericides [2-4]. Recent investigations have shown that the compounds of this group may find useful application in medicine, food chemistry and other spheres [4,51. The aim of the present work was to study the redox properties of the red pigment echinochrome A and estimate the thermodynamic parameters of its possible 0022-0728/95/$09.50 © 1995 Elsevier Science S.A. All rights reserved SSDI 0022-0728(94)03737-X
transitions in aqueous solutions. Two related compounds, the trimethyl ether of echinochrome A and 5,7,8-trihydroxy-6-ethyl- 1,2,3,4-tetraketotetralin were also examined for comparison.
OH 0 HO~OH HsC~Y "~ OH OH 0 Compound I Echinochrome A (2,3,5,7,8-pentahydroxy-6-ethyl-1,4-naphthoquinone)
OH 0 OH 0 N3CO~ . ~ OCH, H O . . ~ O H,C~ y W OCH, HsC2"'~T>"~/I *0 OH 0 OH 0 CompoundII Trimethylether of echinochromeA (5,8-dihydroxy2,3,7-trimethoxy-6-ethyl-1,4-naphthoquinone)
CompoundIIl 5,7,8-trihydroxy-6-ethyl-1,2,3,4tetraketotetralin
2. Experimental The investigation was carried out using thin layer voltammetry. A cell with two plane-parallel pyrolytic graphite electrodes with a total area of 2.26 cm 2 was used. The effective thickness of the solution layer was 15 ~m. The reference and auxiliary electrodes were silver]silver chloride; potential values quoted are given vs. the SHE. Measurements were taken at T = 22 _+ 0.5°C.
S.A. Petro~'a et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
132
Preparations of echinochrome A from the shells of sea-urchins Strongylocentrotus intermedius and compounds II and III were supplied by Dr. N. Mistchenko (Pacific Institute of Bioorganic Chemistry, Russian Academy of Sciences, Vladivostok). Echinochrome A and its trimethyl ether are insoluble in water. These substances were deposited onto the surface of the pyrolytic carbon electrode from dimethylformamide solution in the form of a very thin film containing a few molecular layers. In most cases the amount of quinone deposited on the electrode was about 2.2 x 10 - 9 mol c m - 2 . Several experiments were run in which the quinone content ranged from 5 x 10 10 t o 5 X 10 - 9 mol cm 2. The water-soluble compound III was investigated at a concentration of about 0.6 to 0.7 mM. The experiments carried out with compounds of moderate solubility which could be studied both in the dissolved state and in solid film on the pyrolytic carbon electrode have shown quite coincident results. The potentials of redox conversion obtained this way may be considered as conforming with those obtained in solution phase. A solution of N a z S O 4 w a s used as supporting electrolyte. To maintain the desired value of pH, one of the following buffers was added to the solution: glycine + HC1 (pH 1.1 to 3.5), acetate (pH 3.6 to 4.9), phthalate + N a O H (pH 4.1 to 5.9), phosphate (pH 5.8 to 8.0), "tris" (pH 7.2 to 9.1), glycine + N a O H (pH 8.5 to 12.9). The ionic strength of the solution was about 1.0. The redox properties of echinochrome A and compound III were investigated in the pH range up to 8 because in more basic media they are unstable. Compound II was examined up to pH 13.
it
A potentiostat was used to control the linear potential sweep with a rate of 1 mV s-~ for cyclic voltammetric curves. The midpoint potentials at various values of p H were determined from the voltammetric curves. The results are represented in the form of pH-potential diagrams. The structures of conjugated forms of the compounds investigated as shown in the diagrams are regarded as the most probable in view of the values of the redox potentials and ionization constants found in the experiments.
3. Results and discussion
Echinochrome A and its trimethyl ether possess both the properties of electron donor and of electron acceptor. Thus these compounds may be either reduced cathodically or oxidized anodically.
3.1. Reduction of polyhydroxynaphthoquinones 3.1.1. Echinochrome A (compound I) In the pH range up to 5 the electrochemical reduction of echinochrome A occurs by binding of two hydrogen atoms to the carbonyl oxygens in positions 1 and 4 of the molecule with the formation of 1,2,3,4,5,7,8-heptahydroxy-6-ethylnaphthalene as product. The reverse reaction of anodic reoxidation may proceed by different routes with the generation of alternative quinones. This is due to the presence of many hydroxy groups in the molecule of quinone, up to seven in its reduced form. Several voltammetric curves for electrochemical
,2
2
o
04U
~ ~ f ~2 - - ~0
\1 t~-. I
?p2 o--.. I
(a)
(b)
(c)
Fig. 1. Cyclic voltammetric curves for echinochrome A (m = 2.2 x 10 9 rnol cm-2); E r resting potential: (a) pH 0.21; (b) pH 1.01; (c) pH 2.20; 1,2 sequence of cycling.
S.A. PetroL,a et aL /Journal of Electroanalytical Chemistry 384 (1995) 131-137
conversion of echinochrome A in the negative range against the resting potential E r a r e shown in Fig. 1. The primary reduction of echinochrome A showed a single cathodic peak (dashed line) but on anodic reoxidation and subsequent cycling two pairs of conjugated anodic-cathodic peaks appeared on the curve. This indicates that two quinones with different values of standard potential are formed in the acidic region. The quinone with less positive value of E °' is echinochrome A, and the other quinone is probably a derivative of 1,2-naphthoquinone. The reaction scheme may be as follows:
1~
Formed in the first reduction
process
",,,~ 558m'
o, o
',X,\
u~ -0.3l "-. Lu -04
" "tgov
HO
0
A~
3
AH4 "X'~\X 4 -58 mV
?-87dE
mV
t H 4-
05 ~ S ~ + e _
)2",,
-061
OH 0
®
2 xx~ - 87 my
pH i
I~
J
H0v'~.~.,.- OH
/
HBC2TOH "ii" OH 0
OH o
AH2
-°'1-
I
T 0H i~_---_~H
OH OH
/
-
(t)
HsC~ T
I",'~
OH 0 H0.,~0H
(AH4) OH OH H 0 ~ 0 H
[
133
0
L
i
2
4
6
i
8
Fig. 2. p H - p o t e n t i a l diagram for echinochrome A (compound I). T h e symbol AH 2 stands for the compound I shown in the figure. The dashed line (here and in Fig. 3) corresponds to the H 2 / H + equilibrium.
HsC~,T "T" "OH OH OH
The quantitative ratio between the two quinones varies. The fraction of echinochrome A rises as the p H is increased. Essentially in the p H range from 2.5 to 5, echinochrome A becomes the single product of the electrochemical conversion. With a further shift in p H to the neutral region, double or even triple pairs of peaks on the voltammetric curves appeared again, owing to the formation of other isomers of echinochrome A. The electrochemical conversion of echinochrome A proceeds rather reversibly. The difference between the potentials of anodic and cathodic peaks (AEp = a E p - c E p ) did not exceed 20 to 25 mV and the heights were equal within about _+10%. The voltammetric curves had a quite symmetric shape, but their width at mid-height (AEp/2) exceeded the theoretical value for a two-electron process (45 mV), the a v e r a g e AEp/2 value of the anodic peak being 57 _+ 3 m V in the p H range 0 to 5. The widening of the peaks may be caused by the formation of semiquinone as an intermediate product of the two-electron reaction. The equilibrium constant of the reaction of semiquinone disproportionation in this region was found from the values of AEp/2 [6] tO be about 4 to 5. Accordingly, the maximum concentration of semiquinone did not exceed 18 to 20% of the total amount of the compound involved in the process. The p H - p o t e n t i a l diagram of echinochrome A is shown in Fig. 2. In the lower left-hand corner of the figure the overall scheme of the redox processes in the system is depicted. On this scheme a shift to the left corresponds to the addition of a proton and a shift downward to the addition of an electron.
As one can see, the diagram consists of four intercepts with different slopes of the p H - p o t e n t i a l line: - 5 8 m V (pH < 4.9), - 8 7 mV (4.9 < pH < 5.8), - 5 8 mV (5.8 < p H < 6.6) and - 87 m V (pH > 6.6). Accordingly, five different forms of echinochrome A may exist in the p H range 0 to 8. Three of them are oxidized forms, while the other two are reduced. The uncharged forms ( A H z and A H 4) are stable in the acid and weak acid regions. Redox transitions between them are symbolized by scheme (1). In the p H range 5 to 8 the oxidized form may exist either as an anion A H - or (pH > 6.6) as a dianion A 2 . The anionic reduced form A H 3 is stable in solutions of p H > 5.8. The scheme of redox and a c i d - b a s e reactions in this region may be written as follows: ('2)
(AH-)
OH 0 HO~_..~OHsC2 "Y" "ii""OH OH 0
pKo~=6.6II-H+ OH 0
(A2-) H O ~ . ~ O H~C2 ~r" Y oOH 0
+-= 5H++2e~
~ "
OH OH HO~OH
(AH.)
H~C2 T ~" OH OH OH -H+IIpKred=5.8 OH OH
+SH++2e- H O ~ . O = ~
(AH;)
H~C2 T "T" OH OH OH
The midpoint potential of echinochrome A was not found to be dependent on the amount of quinone deposited onto the electrode surface within fairly wide limits. Thus, the average value of E m a t p H 3.45 was - 159 -+ 1.8 mV as the quinone content varied in the
S.A. Petroua et aL /Journal of Electroanalytical Chemistry 384 (1995) 131-137
134
Table 1 Redox equilibria in the echinochromeA system Transfer " Reaction E°'/V
]03~r/V
1-4 2-4 2-5 3-5
4.4 3.0 2.8 1.0
AH2 +2H+ +2e- ~ AH4 AH- +3H+ +2e ~ A H 4 AH +2H++2e ~AH~ A2 +3H+ +2e- ~ AH.~
0.044 0.188 0.018 0.211
Numbers correspond to Fig. 2; o- standard deviation.
Table 2 Acid-base equilibria in the echinochromeA system Transfer a Reaction pK
o-
1-2 2-3 4-5
0.17 0.09 0.11
AH 2 ~ AH- +H + AH- ~ A2 + H + AH 4 ~ AH 3 +H +
4.90 6.59 5.81
In the oxidized state the compound II shows less acidic features than does echinochrome A. The molecule of echinochrome A loses its first proton at pH > 4.9, whereas the formation of a singly charged anion of compound II occurs only in the weakly basic region (pH > 9.3). This difference arises because the protons are being removed in these two cases from different positions. In the molecule of echinochrome A the protons are derived from the strongly acidic quinoid hydroxylic groups, whereas in compound II the only possible sources of protons are the less acidic O H groups located in the aromatic moiety of the molecule. The acid-base and redox equilibria of compound II in solutions of pH > 9.3 may be depicted by the following scheme:
" Numbers correspond to Fig. 2.
0- O
H ~ C 0I~ range ( 0 . 2 5 - 1 ) x 10 8 mol cm -2. Beyond these limits the E m value was a few millivolts more negative. The values of E °' and p K quoted in Tables 1 and 2 were obtained from experiments carried out in the range indicated above.
3.1.2. Trimethyl ether of echinochrome A (compound II) Substitution of O H groups in positions 2,3 and 7 of the molecule of echinochrome A by methoxy groups affects the redox properties of the quinone appreciably. Thus, for compound I! only one pair of conjugated cathodic-anodic peaks was observed on the voltammetric curves in the whole range of pH. The redox transition occurs quite reversibly, but the voltammetric curves obtained in subsequent redox cycles show a gradual decrease in height of the peaks, especially in acid and weak acid regions. The p H - p o t e n t i a l diagram of compound II is shown in Fig. 3. The areas of stability of six different forms of the quinone are plotted on the diagram. The neutral oxidized form A H 2 is stable in the pH range up to 9.3. The stoichiometry of the reduction of A H 2 depends on the pH of the solution. In the solutions of pH < 6, A H z undergoes a two-proton and two-electron reduction, producing a neutral leuco-form, A H 4. At pH > 6 the reduced forms, namely A H y (pH < 8) and AH22 (pH > 8) are negatively charged:
O- OH
OH 0 H3CO~-~OCH3
H,C0~0CH, (AH;) ,~*.2~e- H,C2 T T 0CH, . ~ OH OH
HsC2 T Y 0CH3 " ~ OH 0 (AH2)
tl-H + o- OH HsC0~0CH.
H,C2 T T OCH, 0- OH
(3)
(AH-)
(4)
OCHa
HsC~ T "iF 0CH3 ~ OH 0
pKox2~t4 II-H + (A2-) H 3 C 0 ~
O- OH H~CO~OCH3
.
. ~
O- 0
H,C~ y y 0CH3 O- OH
0CH3
(AH~)
HBC ~ ~l~ "if" OCHs
O- 0
The values of the standard redox potentials and the ionization constants of the trimethyl ether of echinochrome A are summarized in Tables 3 and 4. Both the quinones studied show rather complicated redox behaviour. It may be noted that unsubstituted 1,4-naphthoquinone undergoes a simple two-proton, two-electron reduction in the entire range of pH ( E °'
[-I2
0.1 '\
\
0
~
\\\\\
-0.,
"£~
H3CO
OCH3
.502
OCH3
dE _ d-~-- 58 mV
@
\xx A H 4 X
AH-I 2-
> -0.2 -0.3
+H+ \ \
AH~ -29
_o,
-,,58 mV
""4, 7-
-0.5 ~
I \\ I
(AH~T)
II
0
,
I
2
,
I
4
,
6
,
|
8
pH ,'~
I
I0
,
I
12
i
Fig. 3. pH-potential diagram for trimethyl ether of echinochrome A (compound lI). The symbolAH 2 stands for the compound II shown in the figure.
S.A. Petroua et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137 Table 3 Redox equilibria in the system of the trimethyl ether of echinochrome A Transfer a
Reaction
1-4 1-5 1-6 2-6 3-6
AH2
a
+2H+
+2e
~4~-AH 4
A H 2 + H + 2 e - ~- A H 3 AH2 +2e ~AH 2 A H - + H + + 2 e - ~ A H 2A 2 + 2 H + + 2 e - ~ A H 2-
E°'/V
103~r/V
0.157 -0.017 -0.252 0.021 0.342
2,1 1,2 4.4 2,4 7.3
8
135
Ia
~o 4
~ .SHE I
I
i
N u m b e r s correspond to Fig. 3.
A_
= 0.470 V[7]). Introduction of one ethyl and five hydroxy groups not only decreases the value of the standard redox potential of 1,4-naphthoquinone by 426 mV ( E ° ' = 0.044 V for echinochrome A), but also affects the structure of the p H - p o t e n t i a l diagram noticeably. The topology of the p H - p o t e n t i a l diagram of compound II which contains three methoxy instead of hydroxy groups in positions 2,3 and 7 of the molecule, differs both from that of echinochrome A and from that of the unsubstituted 1,4-naphthoquinone. In addition, all redox transitions of compound II proceed at potentials more positive than that of compound I. The difference in the values of E °' attains 113 mV in the acid region. Both hydroxy and methoxy groups are electrondonating substitutents. Introduction of an O H group in position 2 of 1,4-naphthoquinone decreases the redox potential of the molecule by 128 mV [8], whereas the shift in potential induced by an O C H 3 group in the same position is 131 m V [9]. The effect is opposite for 2,3-substituted-l,4-naphthoquinones that contain either two O H or two O C H 3 groups. So the E °' of 2,3-dihydroxy-l,4-napthoquinone was found to be 200202 mV less positive than that of 1,4-naphthoquinone [9,10]. In contrast, the negative shift in redox potential for 2,3-dimethoxy-l,4-naphthoquinone is only 95 mV [10]. The effect we have observed in this study is quite similar. The available literature data relating to the present study are rather limited. Therefore the results obtained can be compared only with a few sources. So according to Fieser and Fieser [11], E °' of echinochrome A at 30°C equals 0.08 V. This is 0.036 V higher than our value. The considerably more positive value reported
Table 4 A c i d - b a s e equilibria echinochrome A
in the system of the trimethyl ether of
Transfer a
Reaction
pK
1-2 2-3 4-5 5-6
AH 2 ~ AH- +H + A H ~ A 2- + H ÷ AH 4 ~ AH 3 + H + A H 3 ~ AH22- + H +
9.31 10.95 5.96 8,01
a
Numbers correspond to Fig. 3.
0.15 0.23 0.08 0.14
~
-E lIIa
(o)
16
I1
I /]a
E/V vs. SHE ,
-04
~
t
I
/
12 <~
\
8
//
,
-0.2
0
C
0.2
[0.4- j . O . 6 ~
. V
°
Fig. 4. Cyclic voltammetric curves for echinochrome A (m = 2.2× 10 . 9 mol cm 2, pH 3.03): (a) initial scan to the negative side of Er; (b) initial scan to the positive side of Er; 1,2 sequence of cycling.
by Cannan [12], 0.200 V, may be assigned probably to the redox potential of an isomer of echinochrome A.
3.2. Oxidation of polyhydroxynaphthoquinones 3.2.1. Echinochrome A When the redox cycling of echinochrome A was carried out in the potential region more negative than E r, the usual voltammetric curve was obtained (a pair of peaks I c - I . in Fig. 4(a)). However, when the potential was scanned in the positive direction from the resting potential, more complete oxidation of the substance occurred. Thereby the anodic p e a k II a appeared on the curve. The peak potential was 0.63 V more positive than the midpoint potential of the reductionreoxidation of echinochrome A. Peak II a had no cathodic counter peak when the scanning was reversed. Only when the potential attained the value of redox conversion of echinochrome A in the negative region was the strongly oxidized form reduced. Accordingly
136
S.A. Petroua et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
the cathodic peak II c was almost twice the size of the initial cathodic peak I x. Subsequent cycling in the negative region showed symmetric anodic and cathodic peaks that practically coincided with the initial peaks (I a and lc). Probably peak IIa corresponds to the oxidation of echinochrome A according to the reaction: OH 0 HO
--
"
~
OH
_2H+_2e -
HO...~.O
OH OH 0
echinochrome A
L
II
J
I
I]a
2
// # :-#-o'.2 o
OH O ---
HsC
ia
I E"VvsSHE
/d6-
(5)
HsC~'~'~O OH 0
compoundl~
This substance was found to be the product of the reaction of chemical oxidation of echinochrome A [13]. The polyketone formed in reaction (5) may be hydrated easily. This is the probable reason for the high irreversibility of the subsequent process of electrochemical reduction of the compound (peak IIc). The product of reaction (5) may also undergo further oxidation, as indicated by the anodic peak IIIa (Fig. 4(b)). This reaction is likely to proceed with the destruction of the tetralin ring system. Most of the oxidation products are electrochemically inactive. The peaks corresponding to the conversion of echinochrome A disappeared in subsequent cycles. Some minor peaks observed on the curve may be attributed to derivatives of 1,4-benzoquinone formed on oxidative decomposition of compound III [5]. In the acid region the values of potentials of anodic peaks depend on the p H according to the expressions aEp = (0.671 - 0.058 p H )
( p e a k II~)
~Ep = (0.895 - 0.058 p H )
(peak IIIa)
3.2.2. C o m p o u n d I I I
The electrochemical properties of this compound have been examined with a view to verifying its role in the process of anodic oxidation of echinochrome A. The reduction of compound III is displayed in Fig. 5(a) as a large cathodic peak II c at the potential near that of the reduction of echinochrome A. Further cycling in this potential range resulted in the appearance of conjugated anodic-cathodic peaks Ia--I c of an area half that of the peak II c. It stands to reason that in the initial process four electrons were added to c o m p o u n d I I I to produce the dihydroform of echinochrome A. Further reconversion of echinochrome A and its leuco-compound proceeded with participation of two electrons. Compound III can be regenerated by oxidation of echinochrome A in the region more positive than the resting potential (peak IIa). The behaviour of the system when polarized positively in relation to the resting potential (Fig. 5(b)) did
L -4
(b
\ \ II
2o
- 4 "Y
Fig. 5. Cyclic voltammetric curves for compound III (c = 0.62 mM, pH 3.34): (a) initial scan to the negative side of Er; (b) initial scan to the positive side of Er; 1,2 sequence of cycling. not differ noticeably from that of echinochrome A (Fig. 4(b)). The only distinction was that in this case peak II a practically disappeared. This confirms our suggestion
< 4
4
~
2 E / V v s SHE
/ -2 .x
~2
-4
Fig. 6. Cyclicvoltammetriccurves for trimethyl ether of echinochrome A (m = 2.2x 10 9 mol per cm -2, pH 2.26); 1,2 sequence of cycling.
S.A. Petrova et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
that peak IIa corresponds to the reaction of electrochemical oxidation of echinochrome A into compound III.
3.2.3. Trimethyl ether of echinochrome A The processes of electrochemical oxidation of echinochrome A and compound II are found to differ. Fig. 6 shows a voltammetric curve obtained under redox cycling of compound II over a wide potential range. Two pairs of conjugated cathodic-anodic peaks can be observed on the curve. The left pair corresponds to the reduction and reoxidation of the compound II ( E ° ' = 0.157 V), whereas the right pair may be assigned to the formation of the derivative of 1,4,5,8-naphthodiquinone: OH
0
H~CO
OCH3
HsC& "y "y OH
0
-2H+-2e -
0CH~ E°'=t.02V
H ~PO ~
0 II
0 II OC",,3
(6)
H5 C2~ 0IIC H II 0
0
Reaction (6) proceeds rather reversibly, but the product is chemically unstable, especially outside the acid region. In solutions of p H > 6 the naphthodiquinone is completely decomposed by water. When comparing reactions (5) and (6) one should note that the oxidation sites involved in these two processes are different. There are the O H groups located in positions 2 and 3 of the quinoid ring (reaction (5)) and in positions 5 and 8 of the aromatic moiety of the molecule. Electrochemical oxidation by the first path demands less energy to proceed than oxidation by the second path. This may be proved by comparing the potential shifts needed for the reactions
137
to occur. The difference between the values of a E p of peak II a (0.671 V at pH 0) and E °' of the echinochrome-dihydroechinochrome couple (0.044 V) is 0.627 V, whereas the corresponding value for reaction (6) is considerably greater (0.863 V). Exactly this energy relation may be the cause of the formation of compound iII and not the naphthodiquinone during electrochemical oxidation of echinochrome A. References [1] V.L. Novikov and V.F. Anufriev, Proc. 3rd All-Union Conf. on Bioantioxidants, Moscow, 1989, Vol.1, p. 233 (in Russian). [2] L.V. Boguslavskaya, N.G. Khrapova and O.B. Maksimov, Izv. Akad. Nauk SSSR, Ser. Khim., 7 (1985) 1471 (in Russian). [3] L.V. Boguslavskaya and N.P. Mistchenko, Izv. Akad. Nauk SSSR, Set. Khim., 2 (1991) 329 (in Russian). [4] V.L. Novikov and V.F. Anufriev, Proc. All-Union Seminar on Chemistry of Physiologically Active Compounds, Chernogotovka, 1989, p. 182 (in Russian). [5] E.A. Kol'tsova and O.B. Maksimov, Proc. of All-Union Conf. on Chemistry of Quinones and Quinoid Compounds, Novosibirsk, 1991, p. 165 (in Russian). [6] S.A. Petrova and O.S. Ksenzhek, Elektrokhimiya, 22 (1986) 137 (in Russian). [7] O.S. Ksenzhek and S.A. Petrova, Electrochemical Properties of Reversible Biological Redox Systems, Nauka, Moscow, 1986, p. 94 (in Russian). [8] J.B. Conant and L.F. Fieser, J. Am. Chem. Soc., 46 (1924) 1858. [9] L.F. Fieser and M. Fieser, J. Am. Chem. Soc., 57 (1935) 491. [10] K. Wallenfels and W. Mohle, Bet. Dtsch. Chem. Ges., 76 (1943) 924. [11] L.F. Fieser and M. Fieser, Advanced Organic Chemistry, Vol. 2, Russian translation, Khimiya, Moscow, 1970, p.436 (in Russian). [12] R.K. Cannan, Biochcm. J., 21 (1927) 184. [13] K. Wallenfels and A. Gauhe, Ber. Dtsch. Chem. Ges., 76 (1942) 413.
ELSEVIER
Journal of Electroanalytical Chemistry 384 (1995) 131-137
Redox properties of echinochrome A S.A. Petrova, O.S. Ksenzhek, M.V. Kolodyazhny Ukrainian State Unicersity of Chemical Technology, Dniepropetrocsk 32000.5, Ukraine
Received 3 June 1994; in revised form 19 August 1994
Abstract The electrochemical properties of polyhydroxynaphthoquinone from the shells of sea-urchins Strongylocentrotus intermedius (echinochrome A) and related compounds were investigated using thin layer voltammetry on pyrolytic graphite electrodes. On the basis of experimental data pH-potential diagrams for ecbinochrome A and its trimethyl ether are plotted. Five different forms of echinochrome A and six forms of the trimethyl ether of echinochrome A are shown to exist over the whole range of pH studied. Equilibrium constants and formal redox potentials for all conjugated forms are obtained. Electrochemical oxidation of echinochrome A and its trimethyl ether lead to the formation of different products. Tetraketotetralin is formed in the former case and naphthodiquinone in the latter. Keywords: Redox properties; Echinochrome A
1. Introduction Quinones are widespread in the cells of living organisms where they perform various biochemical functions. Several types of quinone act as components of electron transport chains in the respiratory and photosynthetic systems. Some quinones possess the properties of vitamins and drugs. Quinones are also found to play the role of "chemical weapon" for some insects and as toxins of plants. Naturally occurring quinones show a great diversity of structure, including various benzo-, naphtho- and anthraquinones. Among them is a group of polyhydroxynaphthoquinones which includes echinochrome A and a number of spinochromes. These quinones are incorporated in appreciable amounts (0.01% to 0.04% by weight) in the eggs, needles and shells of sea-urchins [1]. Their biological functions are not yet established and their chemical features, especially electrochemical features, have been scarcely studied. Nevertheless it is known that echinochrome A and some spinochromes possess pronounced activity as antioxidants and bactericides [2-4]. Recent investigations have shown that the compounds of this group may find useful application in medicine, food chemistry and other spheres [4,51. The aim of the present work was to study the redox properties of the red pigment echinochrome A and estimate the thermodynamic parameters of its possible 0022-0728/95/$09.50 © 1995 Elsevier Science S.A. All rights reserved SSDI 0022-0728(94)03737-X
transitions in aqueous solutions. Two related compounds, the trimethyl ether of echinochrome A and 5,7,8-trihydroxy-6-ethyl- 1,2,3,4-tetraketotetralin were also examined for comparison.
OH 0 HO~OH HsC~Y "~ OH OH 0 Compound I Echinochrome A (2,3,5,7,8-pentahydroxy-6-ethyl-1,4-naphthoquinone)
OH 0 OH 0 N3CO~ . ~ OCH, H O . . ~ O H,C~ y W OCH, HsC2"'~T>"~/I *0 OH 0 OH 0 CompoundII Trimethylether of echinochromeA (5,8-dihydroxy2,3,7-trimethoxy-6-ethyl-1,4-naphthoquinone)
CompoundIIl 5,7,8-trihydroxy-6-ethyl-1,2,3,4tetraketotetralin
2. Experimental The investigation was carried out using thin layer voltammetry. A cell with two plane-parallel pyrolytic graphite electrodes with a total area of 2.26 cm 2 was used. The effective thickness of the solution layer was 15 ~m. The reference and auxiliary electrodes were silver]silver chloride; potential values quoted are given vs. the SHE. Measurements were taken at T = 22 _+ 0.5°C.
S.A. Petro~'a et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
132
Preparations of echinochrome A from the shells of sea-urchins Strongylocentrotus intermedius and compounds II and III were supplied by Dr. N. Mistchenko (Pacific Institute of Bioorganic Chemistry, Russian Academy of Sciences, Vladivostok). Echinochrome A and its trimethyl ether are insoluble in water. These substances were deposited onto the surface of the pyrolytic carbon electrode from dimethylformamide solution in the form of a very thin film containing a few molecular layers. In most cases the amount of quinone deposited on the electrode was about 2.2 x 10 - 9 mol c m - 2 . Several experiments were run in which the quinone content ranged from 5 x 10 10 t o 5 X 10 - 9 mol cm 2. The water-soluble compound III was investigated at a concentration of about 0.6 to 0.7 mM. The experiments carried out with compounds of moderate solubility which could be studied both in the dissolved state and in solid film on the pyrolytic carbon electrode have shown quite coincident results. The potentials of redox conversion obtained this way may be considered as conforming with those obtained in solution phase. A solution of N a z S O 4 w a s used as supporting electrolyte. To maintain the desired value of pH, one of the following buffers was added to the solution: glycine + HC1 (pH 1.1 to 3.5), acetate (pH 3.6 to 4.9), phthalate + N a O H (pH 4.1 to 5.9), phosphate (pH 5.8 to 8.0), "tris" (pH 7.2 to 9.1), glycine + N a O H (pH 8.5 to 12.9). The ionic strength of the solution was about 1.0. The redox properties of echinochrome A and compound III were investigated in the pH range up to 8 because in more basic media they are unstable. Compound II was examined up to pH 13.
it
A potentiostat was used to control the linear potential sweep with a rate of 1 mV s-~ for cyclic voltammetric curves. The midpoint potentials at various values of p H were determined from the voltammetric curves. The results are represented in the form of pH-potential diagrams. The structures of conjugated forms of the compounds investigated as shown in the diagrams are regarded as the most probable in view of the values of the redox potentials and ionization constants found in the experiments.
3. Results and discussion
Echinochrome A and its trimethyl ether possess both the properties of electron donor and of electron acceptor. Thus these compounds may be either reduced cathodically or oxidized anodically.
3.1. Reduction of polyhydroxynaphthoquinones 3.1.1. Echinochrome A (compound I) In the pH range up to 5 the electrochemical reduction of echinochrome A occurs by binding of two hydrogen atoms to the carbonyl oxygens in positions 1 and 4 of the molecule with the formation of 1,2,3,4,5,7,8-heptahydroxy-6-ethylnaphthalene as product. The reverse reaction of anodic reoxidation may proceed by different routes with the generation of alternative quinones. This is due to the presence of many hydroxy groups in the molecule of quinone, up to seven in its reduced form. Several voltammetric curves for electrochemical
,2
2
o
04U
~ ~ f ~2 - - ~0
\1 t~-. I
?p2 o--.. I
(a)
(b)
(c)
Fig. 1. Cyclic voltammetric curves for echinochrome A (m = 2.2 x 10 9 rnol cm-2); E r resting potential: (a) pH 0.21; (b) pH 1.01; (c) pH 2.20; 1,2 sequence of cycling.
S.A. PetroL,a et aL /Journal of Electroanalytical Chemistry 384 (1995) 131-137
conversion of echinochrome A in the negative range against the resting potential E r a r e shown in Fig. 1. The primary reduction of echinochrome A showed a single cathodic peak (dashed line) but on anodic reoxidation and subsequent cycling two pairs of conjugated anodic-cathodic peaks appeared on the curve. This indicates that two quinones with different values of standard potential are formed in the acidic region. The quinone with less positive value of E °' is echinochrome A, and the other quinone is probably a derivative of 1,2-naphthoquinone. The reaction scheme may be as follows:
1~
Formed in the first reduction
process
",,,~ 558m'
o, o
',X,\
u~ -0.3l "-. Lu -04
" "tgov
HO
0
A~
3
AH4 "X'~\X 4 -58 mV
?-87dE
mV
t H 4-
05 ~ S ~ + e _
)2",,
-061
OH 0
®
2 xx~ - 87 my
pH i
I~
J
H0v'~.~.,.- OH
/
HBC2TOH "ii" OH 0
OH o
AH2
-°'1-
I
T 0H i~_---_~H
OH OH
/
-
(t)
HsC~ T
I",'~
OH 0 H0.,~0H
(AH4) OH OH H 0 ~ 0 H
[
133
0
L
i
2
4
6
i
8
Fig. 2. p H - p o t e n t i a l diagram for echinochrome A (compound I). T h e symbol AH 2 stands for the compound I shown in the figure. The dashed line (here and in Fig. 3) corresponds to the H 2 / H + equilibrium.
HsC~,T "T" "OH OH OH
The quantitative ratio between the two quinones varies. The fraction of echinochrome A rises as the p H is increased. Essentially in the p H range from 2.5 to 5, echinochrome A becomes the single product of the electrochemical conversion. With a further shift in p H to the neutral region, double or even triple pairs of peaks on the voltammetric curves appeared again, owing to the formation of other isomers of echinochrome A. The electrochemical conversion of echinochrome A proceeds rather reversibly. The difference between the potentials of anodic and cathodic peaks (AEp = a E p - c E p ) did not exceed 20 to 25 mV and the heights were equal within about _+10%. The voltammetric curves had a quite symmetric shape, but their width at mid-height (AEp/2) exceeded the theoretical value for a two-electron process (45 mV), the a v e r a g e AEp/2 value of the anodic peak being 57 _+ 3 m V in the p H range 0 to 5. The widening of the peaks may be caused by the formation of semiquinone as an intermediate product of the two-electron reaction. The equilibrium constant of the reaction of semiquinone disproportionation in this region was found from the values of AEp/2 [6] tO be about 4 to 5. Accordingly, the maximum concentration of semiquinone did not exceed 18 to 20% of the total amount of the compound involved in the process. The p H - p o t e n t i a l diagram of echinochrome A is shown in Fig. 2. In the lower left-hand corner of the figure the overall scheme of the redox processes in the system is depicted. On this scheme a shift to the left corresponds to the addition of a proton and a shift downward to the addition of an electron.
As one can see, the diagram consists of four intercepts with different slopes of the p H - p o t e n t i a l line: - 5 8 m V (pH < 4.9), - 8 7 mV (4.9 < pH < 5.8), - 5 8 mV (5.8 < p H < 6.6) and - 87 m V (pH > 6.6). Accordingly, five different forms of echinochrome A may exist in the p H range 0 to 8. Three of them are oxidized forms, while the other two are reduced. The uncharged forms ( A H z and A H 4) are stable in the acid and weak acid regions. Redox transitions between them are symbolized by scheme (1). In the p H range 5 to 8 the oxidized form may exist either as an anion A H - or (pH > 6.6) as a dianion A 2 . The anionic reduced form A H 3 is stable in solutions of p H > 5.8. The scheme of redox and a c i d - b a s e reactions in this region may be written as follows: ('2)
(AH-)
OH 0 HO~_..~OHsC2 "Y" "ii""OH OH 0
pKo~=6.6II-H+ OH 0
(A2-) H O ~ . ~ O H~C2 ~r" Y oOH 0
+-= 5H++2e~
~ "
OH OH HO~OH
(AH.)
H~C2 T ~" OH OH OH -H+IIpKred=5.8 OH OH
+SH++2e- H O ~ . O = ~
(AH;)
H~C2 T "T" OH OH OH
The midpoint potential of echinochrome A was not found to be dependent on the amount of quinone deposited onto the electrode surface within fairly wide limits. Thus, the average value of E m a t p H 3.45 was - 159 -+ 1.8 mV as the quinone content varied in the
S.A. Petroua et aL /Journal of Electroanalytical Chemistry 384 (1995) 131-137
134
Table 1 Redox equilibria in the echinochromeA system Transfer " Reaction E°'/V
]03~r/V
1-4 2-4 2-5 3-5
4.4 3.0 2.8 1.0
AH2 +2H+ +2e- ~ AH4 AH- +3H+ +2e ~ A H 4 AH +2H++2e ~AH~ A2 +3H+ +2e- ~ AH.~
0.044 0.188 0.018 0.211
Numbers correspond to Fig. 2; o- standard deviation.
Table 2 Acid-base equilibria in the echinochromeA system Transfer a Reaction pK
o-
1-2 2-3 4-5
0.17 0.09 0.11
AH 2 ~ AH- +H + AH- ~ A2 + H + AH 4 ~ AH 3 +H +
4.90 6.59 5.81
In the oxidized state the compound II shows less acidic features than does echinochrome A. The molecule of echinochrome A loses its first proton at pH > 4.9, whereas the formation of a singly charged anion of compound II occurs only in the weakly basic region (pH > 9.3). This difference arises because the protons are being removed in these two cases from different positions. In the molecule of echinochrome A the protons are derived from the strongly acidic quinoid hydroxylic groups, whereas in compound II the only possible sources of protons are the less acidic O H groups located in the aromatic moiety of the molecule. The acid-base and redox equilibria of compound II in solutions of pH > 9.3 may be depicted by the following scheme:
" Numbers correspond to Fig. 2.
0- O
H ~ C 0I~ range ( 0 . 2 5 - 1 ) x 10 8 mol cm -2. Beyond these limits the E m value was a few millivolts more negative. The values of E °' and p K quoted in Tables 1 and 2 were obtained from experiments carried out in the range indicated above.
3.1.2. Trimethyl ether of echinochrome A (compound II) Substitution of O H groups in positions 2,3 and 7 of the molecule of echinochrome A by methoxy groups affects the redox properties of the quinone appreciably. Thus, for compound I! only one pair of conjugated cathodic-anodic peaks was observed on the voltammetric curves in the whole range of pH. The redox transition occurs quite reversibly, but the voltammetric curves obtained in subsequent redox cycles show a gradual decrease in height of the peaks, especially in acid and weak acid regions. The p H - p o t e n t i a l diagram of compound II is shown in Fig. 3. The areas of stability of six different forms of the quinone are plotted on the diagram. The neutral oxidized form A H 2 is stable in the pH range up to 9.3. The stoichiometry of the reduction of A H 2 depends on the pH of the solution. In the solutions of pH < 6, A H z undergoes a two-proton and two-electron reduction, producing a neutral leuco-form, A H 4. At pH > 6 the reduced forms, namely A H y (pH < 8) and AH22 (pH > 8) are negatively charged:
O- OH
OH 0 H3CO~-~OCH3
H,C0~0CH, (AH;) ,~*.2~e- H,C2 T T 0CH, . ~ OH OH
HsC2 T Y 0CH3 " ~ OH 0 (AH2)
tl-H + o- OH HsC0~0CH.
H,C2 T T OCH, 0- OH
(3)
(AH-)
(4)
OCHa
HsC~ T "iF 0CH3 ~ OH 0
pKox2~t4 II-H + (A2-) H 3 C 0 ~
O- OH H~CO~OCH3
.
. ~
O- 0
H,C~ y y 0CH3 O- OH
0CH3
(AH~)
HBC ~ ~l~ "if" OCHs
O- 0
The values of the standard redox potentials and the ionization constants of the trimethyl ether of echinochrome A are summarized in Tables 3 and 4. Both the quinones studied show rather complicated redox behaviour. It may be noted that unsubstituted 1,4-naphthoquinone undergoes a simple two-proton, two-electron reduction in the entire range of pH ( E °'
[-I2
0.1 '\
\
0
~
\\\\\
-0.,
"£~
H3CO
OCH3
.502
OCH3
dE _ d-~-- 58 mV
@
\xx A H 4 X
AH-I 2-
> -0.2 -0.3
+H+ \ \
AH~ -29
_o,
-,,58 mV
""4, 7-
-0.5 ~
I \\ I
(AH~T)
II
0
,
I
2
,
I
4
,
6
,
|
8
pH ,'~
I
I0
,
I
12
i
Fig. 3. pH-potential diagram for trimethyl ether of echinochrome A (compound lI). The symbolAH 2 stands for the compound II shown in the figure.
S.A. Petroua et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137 Table 3 Redox equilibria in the system of the trimethyl ether of echinochrome A Transfer a
Reaction
1-4 1-5 1-6 2-6 3-6
AH2
a
+2H+
+2e
~4~-AH 4
A H 2 + H + 2 e - ~- A H 3 AH2 +2e ~AH 2 A H - + H + + 2 e - ~ A H 2A 2 + 2 H + + 2 e - ~ A H 2-
E°'/V
103~r/V
0.157 -0.017 -0.252 0.021 0.342
2,1 1,2 4.4 2,4 7.3
8
135
Ia
~o 4
~ .SHE I
I
i
N u m b e r s correspond to Fig. 3.
A_
= 0.470 V[7]). Introduction of one ethyl and five hydroxy groups not only decreases the value of the standard redox potential of 1,4-naphthoquinone by 426 mV ( E ° ' = 0.044 V for echinochrome A), but also affects the structure of the p H - p o t e n t i a l diagram noticeably. The topology of the p H - p o t e n t i a l diagram of compound II which contains three methoxy instead of hydroxy groups in positions 2,3 and 7 of the molecule, differs both from that of echinochrome A and from that of the unsubstituted 1,4-naphthoquinone. In addition, all redox transitions of compound II proceed at potentials more positive than that of compound I. The difference in the values of E °' attains 113 mV in the acid region. Both hydroxy and methoxy groups are electrondonating substitutents. Introduction of an O H group in position 2 of 1,4-naphthoquinone decreases the redox potential of the molecule by 128 mV [8], whereas the shift in potential induced by an O C H 3 group in the same position is 131 m V [9]. The effect is opposite for 2,3-substituted-l,4-naphthoquinones that contain either two O H or two O C H 3 groups. So the E °' of 2,3-dihydroxy-l,4-napthoquinone was found to be 200202 mV less positive than that of 1,4-naphthoquinone [9,10]. In contrast, the negative shift in redox potential for 2,3-dimethoxy-l,4-naphthoquinone is only 95 mV [10]. The effect we have observed in this study is quite similar. The available literature data relating to the present study are rather limited. Therefore the results obtained can be compared only with a few sources. So according to Fieser and Fieser [11], E °' of echinochrome A at 30°C equals 0.08 V. This is 0.036 V higher than our value. The considerably more positive value reported
Table 4 A c i d - b a s e equilibria echinochrome A
in the system of the trimethyl ether of
Transfer a
Reaction
pK
1-2 2-3 4-5 5-6
AH 2 ~ AH- +H + A H ~ A 2- + H ÷ AH 4 ~ AH 3 + H + A H 3 ~ AH22- + H +
9.31 10.95 5.96 8,01
a
Numbers correspond to Fig. 3.
0.15 0.23 0.08 0.14
~
-E lIIa
(o)
16
I1
I /]a
E/V vs. SHE ,
-04
~
t
I
/
12 <~
\
8
//
,
-0.2
0
C
0.2
[0.4- j . O . 6 ~
. V
°
Fig. 4. Cyclic voltammetric curves for echinochrome A (m = 2.2× 10 . 9 mol cm 2, pH 3.03): (a) initial scan to the negative side of Er; (b) initial scan to the positive side of Er; 1,2 sequence of cycling.
by Cannan [12], 0.200 V, may be assigned probably to the redox potential of an isomer of echinochrome A.
3.2. Oxidation of polyhydroxynaphthoquinones 3.2.1. Echinochrome A When the redox cycling of echinochrome A was carried out in the potential region more negative than E r, the usual voltammetric curve was obtained (a pair of peaks I c - I . in Fig. 4(a)). However, when the potential was scanned in the positive direction from the resting potential, more complete oxidation of the substance occurred. Thereby the anodic p e a k II a appeared on the curve. The peak potential was 0.63 V more positive than the midpoint potential of the reductionreoxidation of echinochrome A. Peak II a had no cathodic counter peak when the scanning was reversed. Only when the potential attained the value of redox conversion of echinochrome A in the negative region was the strongly oxidized form reduced. Accordingly
136
S.A. Petroua et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
the cathodic peak II c was almost twice the size of the initial cathodic peak I x. Subsequent cycling in the negative region showed symmetric anodic and cathodic peaks that practically coincided with the initial peaks (I a and lc). Probably peak IIa corresponds to the oxidation of echinochrome A according to the reaction: OH 0 HO
--
"
~
OH
_2H+_2e -
HO...~.O
OH OH 0
echinochrome A
L
II
J
I
I]a
2
// # :-#-o'.2 o
OH O ---
HsC
ia
I E"VvsSHE
/d6-
(5)
HsC~'~'~O OH 0
compoundl~
This substance was found to be the product of the reaction of chemical oxidation of echinochrome A [13]. The polyketone formed in reaction (5) may be hydrated easily. This is the probable reason for the high irreversibility of the subsequent process of electrochemical reduction of the compound (peak IIc). The product of reaction (5) may also undergo further oxidation, as indicated by the anodic peak IIIa (Fig. 4(b)). This reaction is likely to proceed with the destruction of the tetralin ring system. Most of the oxidation products are electrochemically inactive. The peaks corresponding to the conversion of echinochrome A disappeared in subsequent cycles. Some minor peaks observed on the curve may be attributed to derivatives of 1,4-benzoquinone formed on oxidative decomposition of compound III [5]. In the acid region the values of potentials of anodic peaks depend on the p H according to the expressions aEp = (0.671 - 0.058 p H )
( p e a k II~)
~Ep = (0.895 - 0.058 p H )
(peak IIIa)
3.2.2. C o m p o u n d I I I
The electrochemical properties of this compound have been examined with a view to verifying its role in the process of anodic oxidation of echinochrome A. The reduction of compound III is displayed in Fig. 5(a) as a large cathodic peak II c at the potential near that of the reduction of echinochrome A. Further cycling in this potential range resulted in the appearance of conjugated anodic-cathodic peaks Ia--I c of an area half that of the peak II c. It stands to reason that in the initial process four electrons were added to c o m p o u n d I I I to produce the dihydroform of echinochrome A. Further reconversion of echinochrome A and its leuco-compound proceeded with participation of two electrons. Compound III can be regenerated by oxidation of echinochrome A in the region more positive than the resting potential (peak IIa). The behaviour of the system when polarized positively in relation to the resting potential (Fig. 5(b)) did
L -4
(b
\ \ II
2o
- 4 "Y
Fig. 5. Cyclic voltammetric curves for compound III (c = 0.62 mM, pH 3.34): (a) initial scan to the negative side of Er; (b) initial scan to the positive side of Er; 1,2 sequence of cycling. not differ noticeably from that of echinochrome A (Fig. 4(b)). The only distinction was that in this case peak II a practically disappeared. This confirms our suggestion
< 4
4
~
2 E / V v s SHE
/ -2 .x
~2
-4
Fig. 6. Cyclicvoltammetriccurves for trimethyl ether of echinochrome A (m = 2.2x 10 9 mol per cm -2, pH 2.26); 1,2 sequence of cycling.
S.A. Petrova et al. /Journal of Electroanalytical Chemistry 384 (1995) 131-137
that peak IIa corresponds to the reaction of electrochemical oxidation of echinochrome A into compound III.
3.2.3. Trimethyl ether of echinochrome A The processes of electrochemical oxidation of echinochrome A and compound II are found to differ. Fig. 6 shows a voltammetric curve obtained under redox cycling of compound II over a wide potential range. Two pairs of conjugated cathodic-anodic peaks can be observed on the curve. The left pair corresponds to the reduction and reoxidation of the compound II ( E ° ' = 0.157 V), whereas the right pair may be assigned to the formation of the derivative of 1,4,5,8-naphthodiquinone: OH
0
H~CO
OCH3
HsC& "y "y OH
0
-2H+-2e -
0CH~ E°'=t.02V
H ~PO ~
0 II
0 II OC",,3
(6)
H5 C2~ 0IIC H II 0
0
Reaction (6) proceeds rather reversibly, but the product is chemically unstable, especially outside the acid region. In solutions of p H > 6 the naphthodiquinone is completely decomposed by water. When comparing reactions (5) and (6) one should note that the oxidation sites involved in these two processes are different. There are the O H groups located in positions 2 and 3 of the quinoid ring (reaction (5)) and in positions 5 and 8 of the aromatic moiety of the molecule. Electrochemical oxidation by the first path demands less energy to proceed than oxidation by the second path. This may be proved by comparing the potential shifts needed for the reactions
137
to occur. The difference between the values of a E p of peak II a (0.671 V at pH 0) and E °' of the echinochrome-dihydroechinochrome couple (0.044 V) is 0.627 V, whereas the corresponding value for reaction (6) is considerably greater (0.863 V). Exactly this energy relation may be the cause of the formation of compound iII and not the naphthodiquinone during electrochemical oxidation of echinochrome A. References [1] V.L. Novikov and V.F. Anufriev, Proc. 3rd All-Union Conf. on Bioantioxidants, Moscow, 1989, Vol.1, p. 233 (in Russian). [2] L.V. Boguslavskaya, N.G. Khrapova and O.B. Maksimov, Izv. Akad. Nauk SSSR, Ser. Khim., 7 (1985) 1471 (in Russian). [3] L.V. Boguslavskaya and N.P. Mistchenko, Izv. Akad. Nauk SSSR, Set. Khim., 2 (1991) 329 (in Russian). [4] V.L. Novikov and V.F. Anufriev, Proc. All-Union Seminar on Chemistry of Physiologically Active Compounds, Chernogotovka, 1989, p. 182 (in Russian). [5] E.A. Kol'tsova and O.B. Maksimov, Proc. of All-Union Conf. on Chemistry of Quinones and Quinoid Compounds, Novosibirsk, 1991, p. 165 (in Russian). [6] S.A. Petrova and O.S. Ksenzhek, Elektrokhimiya, 22 (1986) 137 (in Russian). [7] O.S. Ksenzhek and S.A. Petrova, Electrochemical Properties of Reversible Biological Redox Systems, Nauka, Moscow, 1986, p. 94 (in Russian). [8] J.B. Conant and L.F. Fieser, J. Am. Chem. Soc., 46 (1924) 1858. [9] L.F. Fieser and M. Fieser, J. Am. Chem. Soc., 57 (1935) 491. [10] K. Wallenfels and W. Mohle, Bet. Dtsch. Chem. Ges., 76 (1943) 924. [11] L.F. Fieser and M. Fieser, Advanced Organic Chemistry, Vol. 2, Russian translation, Khimiya, Moscow, 1970, p.436 (in Russian). [12] R.K. Cannan, Biochcm. J., 21 (1927) 184. [13] K. Wallenfels and A. Gauhe, Ber. Dtsch. Chem. Ges., 76 (1942) 413.