Hydroxyalkyloxy substituted tetraphenylporphyrins: Mechanism and superoxide scavenging activity

Article abstract of DOI:10.1142/S1088424616501212

The novel electrochemical approach based on coulometric response of electro-generated superoxide (O₂^?-) was used to determine scavenging properties of 2H-5,10,15,20-tetrakis (4-hydroxyphenyl)porphyrin (H₂T(4-OHPh)P); 2H-5,

Full text of DOI:10.1142/S1088424616501212

Journal of Porphyrins and Phthalocyanines
J. Porphyrins Phthalocyanines 2016; 20: 1–9
DOI: 10.1142/S1088424616501212
Published at http://www.worldscinet.com/jpp/
Hydroxyalkyloxy substituted tetraphenylporphyrins:
Mechanism and superoxide scavenging activity
a,b
a
a,c◊
Sergey M. Kuzmin* , Svetlana A. Chulovskaya and Vladimir I. Parfenyuk
a
G.A. Krestov Institute of Solution Chemistry of the Russian Academy of Sciences, Ivanovo 153045, Russia
Ivanovo State Power Engineering University, Ivanovo 153003, Russia
Nekrasov Kostroma State University, Kostroma 156961, Russia
b
c
Received 30 May 2016
Accepted 16 August 2016
ABSTRACT: The novel electrochemical approach based on coulometric response of electro-
•
–
generated superoxide (O ) was used to determine scavenging properties of 2H-5,10,15,20-tetrakis
2
(
4-hydroxyphenyl)porphyrin (H T(4-OHPh)P); 2H-5,10,15,20-tetrakis[4-(2-hydroxyethyloxy)phenyl]
2
porphyrin (H T(4-OH(CH ) OPh)P); 2H-5,10,15,20-tetrakis[4-(4-hydroxybutyloxy)phenyl]porphyrin
2
2 2
(
H T(4-OH(CH ) OPh)P); Zn-5,10,15,20-tetrakis[4-(4-hydroxyethyloxy)phenyl]porphyrin (ZnT(4-
2 2 4
OH(CH ) OPh)P) superoxide. It has been identified that the porphyrins under study possess good
2
2
antioxidant properties. The analysis of possible interactions between porphyrins and superoxide anion-
radical has shown that high values of superoxide scavenging activity of tetraphenylporphyrins with
•
–
alcohol chains can be explained by the nucleophilic attack of O2 on the C–C bonds of alcohol.
KEYWORDS: para-substituted tetraphenylporphyrins, superoxide scavenging activity, coulometric
approach, mechanisms.
superoxide dismutase (SOD) activity [15–22] which is
INTRODUCTION
explained by the redox activity of the metal ion and the
electron transfer mechanism. On the other hand, it is well-
known that there are nonenzymatic antioxidants with a
heterocyclic structure [23–25], as well as antioxidants
with phenol and polyphenol moieties [26–30]. Therefore,
it is resonable that some porphyrin ligands [31–36] display
a high antioxidant activity. Phenyl-substituted porphyrins
have been investigated by Milgrom et al. [37–41] who
have found that porphyrin phenyl groups contain a labile
hydrogen atom that is lost at the first oxidation stage. This
results in the formation of phenoxyl groups that promote
further macrocycle oxidation.
Antioxidant activity of enzymatic and nonenzymatic
types makes porphyrins a promising investigation object.
Earlier, we showed the antioxidant properties of some
amino- and hydroxyphenylporphyrins and their mech-
anism of superoxide scavenging activity [32–36]. This
paper is focused on determining superoxide scavenging
activity and its mechanisms in tetraphenylporphyrins with
hydrocarbon chains between the phenyl rings and the OH
groups. The high absorption coefficients of porphyrins
Oxygen cellular metabolism is realized via formation
of reactive oxygen species (ROS) including free radicals
•
−
•
such as superoxide (O ), hydroxyl radical (HO ), etc.
2
[
1]. As a rule, the radicals are aggressive oxidants, so
ROS overproduction induces damage at various sites in
the cell, especially cell membranes, and pathologies of
biochemical processes [2–4]. Different enzymatic and
nonenzymatic antioxidants prevent the damaging effects
of free radicals on living cells [4–6]. At the same time, an
urgent problem of therapeutic compounds development
is finding natural and synthetic compounds with a high
level of antioxidant activity.
Synthetic metalloporphyrins are an appropriate model
of an enzyme catalytic center [7–10] where the metal ion
is able to form superoxo complexes in the superoxide
presence [11–14].Transitionmetalcomplexesdemonstrate

SPP full member in good standing
*
Correspondence to: Sergey M. Kuzmin, email: smk@isc-ras.
ru, tel: +7 915-824-9554
Copyright © 2016 World Scientific Publishing Company

2
S. M. KUZMIN ET AL.
[
42] make spectrometric measurements of antioxidant
2H-5,10,15,20-Tetrakis(4-hydroxyphenyl)
porphyrin. A solution of 0.5 mL (5.29 mmol) of boron
tribromide in 10 mL of methylene chloride was added
to the stirring and cooling solution of 1.0 g (1.36 mmol)
of 5,10,15,20-tetrakis(4′-methoxyphenyl) porphyrin in
200 mL of dried methylene chloride. The mixture was
stirred at room temperature for 2 h, and then 5 mL of
methanol was added. The mixture was neutralized by
ammonia until the color changed from green to dark
cherry, washed with water, dried over sodium sulfate and
evaporated to dryness. The deposit was dissolved in ethyl
acetate and purified on silica gel by eluting with ethyl
acetate. The eluate was evaporated and precipitated with
activity [43] more complicated; for that reason, the
electrochemical assay was found to be most suitable for
such purpose. The determination of radical scavenging
properties of the compounds is based on monitoring the
elecrochemical response of superoxide in the presence of
an antioxidant [32–36, 44–50].
Procedure of synthesis
H T(4-OHPh)P was synthesized by the two-step
2
method via demethylation of 2H-5,10,15,20-tetrakis(4-
methoxyphenyl)porphyrins [51] obtained in high yield
by condensation of benzaldehydes with pyrrole [52,
petroleum ether. The yield was 0.9 g (98%). R = 0.33
5
3]. Then the H T(4-OHPh)P was used to synthesize
f
2
1
(
-
(
CHCl ). H NMR (500 MHz, CDCl , Me Si): δ , ppm
porphyrins and complexes shown in Fig. 1.
3
3
4
H
2.92 (2H, s, pyrrole-NH), 7.70 (8H, d, m-H-Ph), 8.08
8H, d, o-H-Ph), 8.79 (8H, s pyrrole-H). UV-vis (CHCl ):
The purified products were studied by thin-layer
chromatography (silufol plates), UV-vis spectrometry
3
1
λmax, nm (log ε) 650 (3.72), 595 (3.71), 556 (3.90), 519
(
(
Varian Cary 50 spectrometer) and H NMR spectrometry
Bruker AVANCE-500 spectrometer) methods. The mass
(4.06), 423 (5.43).
The 2H-5,10,15,20-tetrakis(4-hydroxyphenyl)porphyrin
spectra were recorded on a Shimadzu Axima Confidence
MALDI-TOF) mass spectrometer.
H-5,10,15,20-Tetrakis(4-methoxyphenyl)
porphyrin {intermediate compound}. A solution of
.0 mL (72.2 mmol) of pyrrole and 8.8 mL (72.2 mmol)
of anisaldehyde was drop added to a boiling solution of
6 g of chloroacetic acid in 300 mL of an isomeric xylenes
characteristics agree quite well with the reported data
54, 55].
H-5,10,15,20-Tetrakis[4-(2-hydroxyethyloxy)
phenyl])porphyrin. A mixture of 0.3 g (0.44 mmol) of
,10,15,20-tetrakis(4-hydroxyphenyl)porphyrin, 1.0 mL
15.2 mmol) of 2-chloroethanol and 0.5 g (3.62 mmol)
(
[
2
2
5
5
(
1
potassium carbonate in 10 mL of dry DMF was refluxed
for 10 h. Then 1.0 mL (15.2 mmol) of 2-chloroethanol
was added and the mixture was refluxed again for
mixture for 20 min. The resulting mixture was refluxed
by air bubbling for 1 more hour. After xylene steaming,
the deposit was filtered off, washed with water and dried
in air at 80 °C. The deposit was dissolved in chloroform
and purified on aluminum oxide (Brockmann Activity
III) eluting with chloroform. The first red zone was
collected, the eluate was evaporated, and the porphyrin
was precipitated with methanol, filtered and dried at
1
0 h. The mixture was poured into water, the precipitate
was filtered off, washed with water and dried in air at
room temperature to constant weight. The precipitate
was Soxhlet extracted with methanol, the solution was
chromatographed on silica gel, eluting with methanol,
the eluate was evaporated, diluted with water and the
precipitate was filtered off and dried. The yield was
room temperature in air. The yield was 5.6 g (42%). R =
f
1
0
.33 (CHCl ). H NMR (500 MHz, CDCl , Me Si): δ ,
3
3
4
H
2
50 mg (66.5%). R = 0.61 (MeOH). UV-vis (MeOH):
ppm -2.79 (2H, s, pyrrole-NH), 4.03 (12H, s, -O-CH3),
.22 (8H, d, m-H-Ph), 8.06 (8H, d, o-H-Ph), 8.79 (8H,
f
λmax, nm (log ε) 650 (3.90), 593 (3.93), 554 (4.14), 517
7
-1
(4.24), 418 (5.57). IR (KBr tablet): λmax, cm 3423,
s, pyrrole-H 8H). UV-vis (CHCl ): λmax, nm (log ε) 652
3
2
923, 2850, 1607, 1509, 1471, 1245, 1172, 967, 802.
(3.87), 595 (3.78), 557 (4.07), 520 (4.25), 423 (5.69).
R
R1 =
O
O
O
H
N
N
N
N
R
M
R
R2 =
R3 =
CH2 CH OH
2
(CH2)3
CH OH
2
R
(
(
(
(
1) M = 2H R = R1 H2T(4-OHPh)P
2) M = 2H R = R2 H2T(4-OH(CH2)2OPh)P
3) M = 2H R = R3 H2T(4-OH(CH2)4OPh)P
4) M = Zn R = R2 ZnT(4-OH(CH2)2OPh)P
Fig. 1. Structural formulas of the studied porphyrins
Copyright © 2016 World Scientific Publishing Company
J. Porphyrins Phthalocyanines 2016; 20: 2–9

HYDROXYALKYLOXY SUBSTITUTED TETRAPHENYLPORPHYRINS
3
1
temperature to constant weight. The precipitate was
H NMR (500 MHz, CDCl , Me Si): δ , ppm -2.91 (2H,
3
4
H
Soxhlet extracted with methanol, then 0.5 g (2.28 mmol)
of zinc acetate dihydrate was added and the mixture
was boiled for 1 h. The solution was chromatographed
on silica gel, eluting with methanol, the eluate was
evaporated, diluted with water and the precipitate filtered
s, pyrrole-NH), 3.43 (8H, m, CH O(H)), 4.38 (8H, m,
CH O(Ph)), 5.27 (4H, s, OH), 7.55 (16H, m, C H ), 8.89
(
2
2
6
4
8H, s, β-pyrrole). MALDI-TOFF-MS: m/z [M + H –
+
H O] experimental 838.01, calcd. 837.95.
2
The 2H-5,10,15,20-tetrakis[4-(2-hydroxyethyloxy)
phenyl])porphyrin characteristics agreed well with the
reported data [56].
off. The yield was 250 mg (61.9%). R = 0.76 (MeOH).
f
UV-vis (DMSO): λmax, nm (log ε) 600 (4.12), 559 (4.30),
4
5
3
24 (5.74). UV-vis (MeOH): λmax, nm (log ε) 605 (4.14),
5
,10,15,20-Tetrakis[4-(4 acetoxybutyloxy)phenyl]
porphyrin (intermediate compound). A mixture of
.3 g (0.44 mmol) of 5,10,15,20-tetrakis(4-hydroxy-
-1
63 (4.14), 431 (5.49). IR (KBr tablet): λmax, cm
449, 1608, 1509, 1245, 1173, 997, 803, 722. MALDI-
0
+
TOFF-MS: m/z [M] experimental 918.28, calcd. 918.33.
phenyl)porphyrin, 1.0 mL (7.7 mmol) of 4-chlorobutyl
acetate and 0.5 g (3.62 mmol) potassium carbonate in
1
The low solubility of Zn-complex in deuterochloroform
1
did not allow us to obtain H NMR data.
0 mL of dry DMF was boiled for 4 h, then 1.0 mL
(
7.7 mmol) of 4-chlorobutyl acetate was added and the
mixture was boiled again for 10 h. Then the mixture was
pouredintowater,theprecipitatewasfiltered,washedwith
water and dried in air at room temperature. The precipitate
was dissolved in dichloromethane and chromatographed
on silica gel eluting with the dichloromethane-methanol,
then the eluate was evaporated, precipitated with
methanol and dried. The yield was 230 mg (45.9%).
Electrochemical procedure
Dimethylsulfoxide (DMSO ≥ 99.5, ALDRICH) was
purified by zone melting and then stored over molecular
sieves in a dry box before use. Tetrabutylammonium
perchlorate (TBAP ≥ 98.0, ALDRICH) was purified by
recrystallization from ethanol. Concentrated solutions of
porphyrins containing 0.02 M TBAP as the supporting
electrolyte were prepared by the gravimetric method
using the electronic analytical balance «Sartorius»
ME215S (the mass determination error did not exceed
3%). The solutions of smaller concentrations were
prepared by the method of serial dilution.
A potentiostat PI-50PRO3 (Elins, Russia) was used for
electrochemical measurements. The experiments were
carried out in a three-electrode temperature-controlled
(25 ± 0.5°C) electrochemical cell in freshly prepared
solutions. The saturated calomel electrode (SCE) inserted
into the electrochemical cell through the Luggin capillary
was used as the reference electrode. The Pt wire was used
as an auxiliary electrode.
R = 0.85 (ethyl acetate), 0.77 (benzene-methanol, 20:1).
f
1
H NMR (500 MHz, CDCl , Me Si): δ , ppm -2.73 (2H,
3
4
H
bs, pyrrole-NH), 2.00–2.13 (16H, m, CH ), 2.16 (12H,
2
s, CH CO), 4.28–4.34 (16H, m, -O-CH ), 7.29 (8H, d,
3
2
J = 8.2 Hz, m-H-Ph), 8.14 (8H, d, J = 8.2 Hz, o-H-Ph),
8
6
.89 (8H, s, pyrrole-H). UV-vis (CHCl ): λmax, nm (log ε)
3
51 (3.89), 594 (3.89), 557 (4.14), 519 (4.29), 423 (5.69).
+
MALDI-TOFF-MS: m/z [M – H] experimental 1134.61,
calcd. 1134.31.
2
H-5,10,15,20-Tetrakis[4-(4-hydroxybutyloxy)
phenyl]porphyrin. A solution of 220 mg (0.19 mmol)
of 5,10,15,20-tetrakis[4-(4 acetoxybutyloxy)phenyl]por-
phyrin in 20 mL THF was mixed with a solution of 0.5 g
(
8.91 mmol) of potassium hydroxide in 1.0 mL of water,
then the mixture was refluxed for 20 h. The mixture
was poured into water and the precipitate was filtered
off, washed with water and dried. The yield was 183
mg (98.0%). R = 0.13 (ethyl acetate). UV-vis (CHCl ):
As the working electrode, we used a polishing Pt strip
(the working surface equaled 1.2 cm ) rigidly fixed in the
2
fluoroplastic lid. Before every measurement, the active
surface of the working electrode was mechanically mirror-
polished, degreased with ethanol, etched with a chromic
mixture for 20 min, carefully cleaned in distilled water and
then in the solution under study. The working electrode
was immersed in the cell with the test solution where the
potential of the working electrode reached a steady value
in 10 min. In order to degas or oxygenate solutions before
the electrochemical measurements, argon or oxygen was
bubbled through the capillary tube for 30 min.
f
3
λmax, nm (log ε) 651 (3.89), 594 (3.85), 557 (4.10), 520
1
(
4.24), 423 (5.63). H NMR (500 MHz, CDCl , Me Si):
3 4
δH, ppm -2.76 (2H, bs, pyrrole-NH), 1.95 (8H, qv, J = 6.8
Hz, -CH ), 2.10 (8H, qv, J = 6.8 Hz, -CH ), 3.88 (8H, t, J
2
2
=
6.1 Hz, -CH O), 4.31 (8H, t, J = 6.1 Hz, -CH O), 7.29
2
2
(
8H, d, J = 8.4 Hz, m-H-Ph), 8.13 (8H, d, J = 8.4 Hz,
o-H-Ph), 8.87 (8H, s, pyrrole-H). MALDI-TOFF-MS:
m/z [M – H] experimental 966.33, calcd. 966.16.
+
Zn-5,10,15,20-Tetrakis[4-(2-hydroxyethyloxy)
phenyl]porphyrin. A mixture of 0.3 g (0.44 mmol) of
,10,15,20-tetrakis(4-hydroxyphenyl)porphyrin, 1.0 mL
15.2 mmol) of 2-chloroethanol and 0.5 g (3.62 mmol)
potassium carbonate in 10 mL of dry DMF was refluxed
for 10 h. Then 1.0 mL (15.2 mmol) of 2-chloroethanol
was added and the mixture was refluxed again for 10 h.
The mixture was poured into water, the precipitate was
filtered off, washed with water and dried in air at room
The oxygen saturation condition was verified by the
cyclic voltammetry (CV) method. In saturated conditions
the concentration of the dissolved oxygen in DMSO at
25°C was 2.1 mM [57, 58]. The free convection mode
was reached 3 min. after the capillary removal from
the solution. After that, the CV response was recorded
at scan rates from 0.01 to 1.00 V/s. The CV data were
corrected for Ohmic (iR) losses using the current
interruption technique [59]. Only the first cycle was used
5
(
Copyright © 2016 World Scientific Publishing Company
J. Porphyrins Phthalocyanines 2016; 20: 3–9

4
S. M. KUZMIN ET AL.
•-
2
Fig. 2. Electrochemical response of the O /O redox process at H T(4-OH(CH ) OPh)P (a) and ZnT(4-OH(CH ) OPh)P
2
2
2
2
2 2
(b) concentration of 0.00, 0.10, 0.25, 0.50, 1.00, 1.50 mM. The scan rate is 0.02 V/s
to study the interaction of the porphyrins and superoxide
anion-radical.
presence, we found value Q –Q (the gray colored area
3ox 1ox
between lines 2 and 4 for positive currents in Fig. 3).
The radical scavenging activity of the para-
substituted tetraphenylporphyrins was determined using
dimensionless parameters w and k:
RESULTS AND DISCUSSION
Figure 2 shows a typical influence of porphyrins
admixture on the electrochemical response of the O /
w = (Q3ox–Q1ox)/(Q3red–Q1red
)
(2)
2
•
-
O2 redox couple. With small porphyrin additions (up to
.1 mM), the superoxide anion-radical electrochemical
•–
2
0
where (Q3ox–Q1ox) is the quantity of electricity of O
oxidation in porphyrin presence; (Q3red-Q1red) is the quantity
of electricity of O reduction in the same experiment.
oxidation peak increases and shifts slightly, which
indicates the increasing of heterogeneous electron
transfer rate [60, 61]. A further increase in the con-
centration of H T(4-OHPh)P, H T(4-OH(CH ) OPh)P,
2
k = (w –w)/w
(3)
0
0
2
2
2 2
H T(4-OH(CH ) OPh)P and ZnT(4-OH(CH ) OPh)P
2
2
4
2 4
where w is the value obtained by extrapolating the w
•
-
0
leads to a decay of the O oxidation currents. The
2
dependence to the zero porphyrin concentration.
The k value vs. antioxidant concentration plots are
shown in Fig. 4. The antioxidant activity of the compound
•-
losses of O2 current values allow us to determine the
superoxide scavenging activity using the jC50 and Kb
parameters [32–34, 48, 50]. But it has been shown [35,
under study is characterized by the slope (A ) (the bigger
k
36] that the coulometric response of electrogenerated
is the slope, the higher is the activity).
superoxide allows us to obtain more suitable parameters
for superoxide scavenging activity determination.
According to the A value, the antioxidant activity
k
series can be arranged as follows:
The quantity of electricity (Q) of reduction processes
was determined by integrating dependences I(t) for
the cathode branch of the CV curve. The quantity of
the electricity spent on oxidation was calculated by
integrating dependences I(t) for the anode branch of the
CV curve. The region between curves 1 and 2 corresponds
to the quantity of electricity Q1red required for porphyrin
electroreduction in a degassed porphyrin solution. The
region between curves 1 and 3 determines the quantity
H T(4-OHPh)P ≥ H T(4-OH(CH ) OPh)P
2
2
2 2
≈
H T(4-OH(CH ) OPh)P
2 2 4
> ZnT(4-OH(CH ) OPh)P
(4)
2
2
Comparing the binding constants of superoxide
anion radical of the studied porphyrins and those of
flavanoids [48, 50], we can describe H T(4-OHPh)P as
2
of electricity Q2red, of dissolved O electroreduction
molecular systems with a high antioxidant activity. Our
2
in oxygenated DMSO region between curves 1 and 3
Q3red — the quantity of electricity corresponding to the
simultaneous porphyrin and oxygen reduction in an
oxygenated solution. The region between curves 2 and 4
results also indicate the considerable activity of H T(4-
2
OH(CH ) OPh)P, H T(4-OH(CH ) OPh)P and ZnT(4-
2 2
2
2 4
OH(CH ) OPh)P.
2
2
Porphyrins antioxidant activity has not been studied
well enough yet, only a few works deal with this problem
[15–22, 31–36]. It is assumed that the antioxidant action
of the most common antioxidants (aromatic amines,
(
gray colored) evaluates the quantity of electricity Q3red
–
Q1red of O reduction in porphyrin presence. To calculate
2
•
-
the quantity of electricity of O2 oxidation in porphyrin
Copyright © 2016 World Scientific Publishing Company
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HYDROXYALKYLOXY SUBSTITUTED TETRAPHENYLPORPHYRINS
5
Fig. 3. Redox currents vs. time plots: (1) background; (2) porphyrins H T(4-OH(CH ) OPh)P (a) and ZnT(4-OH(CH ) OPh)P (b) in
2
2
2
2 2
degassing solution; (3) oxygen in oxygenated solution; (4) respective porphyrins and oxygen in oxygenated solution. The scan rate
is 0.02 V/s. The porphyrins concentrations are 1 mM
•
-
+
2-
2
O2 +AO → AO + O
Scheme 1. Electron donating mechanism of superoxide
scavenging
Fig. 4. Coulometric parameter k of H T(4-OHPh)P (1); H T(4-
2
2
OH(CH ) OPh)P (2); H T(4-OH(CH ) OPh)P (3); ZnT(4-
2
2
2
2 4
OH(CH ) OPh)P (4)
2
2
Fig. 5. CV of redox processes in DMSO: (1) H T(4-OHPh)P,
2
phenols, naphthols, etc.) consists in breaking the reacting
(
2) H T(4-OH(CH ) OPh)P, (3) H T(4-OH(CH ) OPh)P, (4) ZnT
2 2 2 2 2 4
chains. Antioxidants neutralize free radicals donating
(4-OH(CH ) OPh)P. The scan rate is 0.02 V/s. The porphyrins
2 4
(
accepting) an electron or a hydrogen atom. Antioxidant
concentration is 1 mM
interaction with active radicals results in formation
of low activity radicals, which reduces the oxidation
rate. A number of works have reported a correlation
between oxidation potential and antioxidant activity of
compounds. This correlation is thought to be connected
with electron transfer, namely: easier antioxidant
oxidation leads to a more efficient process of superoxide
scavenging according to Scheme 1, where AO is the
antioxidant molecule.
Figure 5 presents redox behavior of the studied por-
phyrins in degassed solutions DMSO. As the represented
CV curves show, the electrochemical response of oxida-
tion of para-substituted tetraphenylporphins is within
the electrochemical window (up to +0.8 V vs. SCE). The
oxidation of H T(4-OHPh)P, H T(4-OH(CH ) OPh)P,
2
2
2 2
H T(4-OH(CH ) OPh)P and ZnT(4-OH(CH ) OPh)P is
2
2 4
2 2
irreversible due to the fast intermolecular electron transfer
and π-cation radical formation [62, 63]. H T(4-OHPh)P
2
oxidation includes two peaks with the maximum
potentials of +0.20 and +0.48 V described elsewhere
[35].Incaseofoxidation,thefirstoxidationwaveofH T(4-
2
OH(CH ) OPh)P, H T(4-OH(CH ) OPh)P and ZnT(4-
2
2
2
2 4
OH(CH ) OPh)P occurs, with the maximums around
2
2
+0.30, +0.24, and +0.20 V, respectively. The oxidation
Copyright © 2016 World Scientific Publishing Company
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6
S. M. KUZMIN ET AL.
current passes through a plateau and then transforms
into the second oxidation wave at the borders of the
electrochemical window. Thus, the oxidation potential
decreases as follows:
Additionally, the electrochemical reduction of such
porphyrins takes place at more negative potential values
(-1.0 ÷ -1.5 V, Fig. 5) than that of oxygen; therefore, the
mechanism shown in Scheme 2 cannot be used to explain
the series of antioxidant activity as the electron transfer
is an energetically unfavorable process.
H T(4-OHPh)P ≈ ZnT(4-OH(CH ) OPh)P
2
2 2
Figure 6 represents the influence of potential scan
rate on the electrochemical response of the porphyrins
oxidation-reduction processes.
<
<
H T(4-OH(CH ) OPh)P
2 2 4
H T(4-OH(CH ) OPh)P
(5)
2
2 2
In order to determine the nature of the anode peak
observed at potential values of about -0.5 V, the potential
was cycled within the range from 0 to -1.6 V at the
scan rate of 0.01 V/s (20 cycles). As a result, the same
electrochemical responses (similar to those in Figs 5 and
6 at low scan rates) were observed, without increasing
of oxidation peaks with the potential values of around
-0.5 V. If the scan rate is high, the potential cycling in the
range -1.70 ÷ -0.80 V leads to a CV response (Fig. 6b,
curve 2), which coincides with those obtained within a
wider range (Fig. 6b, curve 1). If the potential is cycled
in the range -0.90 ÷ +0.10 V (Fig. 6b, curve 3), no anode
peak of about -0.4 V is observed either. Based on the
facts given above, the occurrence of the anode peak II’
should be interpreted as an intermediate oxidation.
Porphyrin reduction accompanied by intermediates
formation was described elsewhere [35, 36]. The
intermediates redox potentials allow us to assume
intermediates involvement in the superoxide scavenging
according to both Schemes 1 and 2. But in the case of
H T(4-OH(CH ) OPh)P with high superoxide scavenging
The series of porphyrins oxidation potential differs from
that of antioxidant activity, which means that the process
described in Scheme 1 cannot explain the differences in
antioxidant properties of the studied porphyrins.
The deactivation mechanism of superoxide scavenging
in case of electron transfer from the superoxide anion-
radical to the antioxidant is represented in Scheme 2.
The antioxidant efficiency in such a mechanism must
correlate with the electron affinity or reduction potential:
the lower the antioxidant reduction potential is, the
more preferable Scheme 2 is. As the CV curve shows
(
Fig. 5), the first reduction peaks of H T(4-OHPh)P,
2
H T(4-OH(CH ) OPh)P, H T(4-OH(CH ) OPh)P and
2
2 2
2
2 4
ZnT(4-OH(CH ) OPh)P reaches the maximum at -1.08,
2
2
-1.17, -1.14 and -1.40 V potential values, respectively.
Therefore, the reduction stability increases as follows:
H T(4-OHPh)P < H T(4-OH(CH ) OPh)P
2
2
2 4
<
<
H T(4-OH(CH ) OPh)P
2
2 2
2
2 4
ZnT(4-OH(CH ) OPh)P
(6)
activity there are no observed intermediates. Therefore,
the assumption of a primary role of intermediates in the
superoxide scavenging activity is unfounded.
Thus, the mechanisms of direct electron transfer cannot
explain the porphyrin antioxidant activity series. At the
same time, the hydrogen transfer mechanism cannot
explain the porphyrin antioxidant activity series in the case
2
2
The reduction stability series differs from that of
antioxidant activity of the porphyrins under study.
•
-
–
O2 +AO → AO + O
2
Scheme 2. Electron accepting mechanism of superoxide
scavenging
Fig. 6. CV response of ZnT(4-OH(CH ) OPh)P in oxygen-free DMSO solutions: (a) at the scan rates: 0.02, 0.05, 0.10, 0.20, 0.50 V/s;
2
2
(
b) at the scan rate 0.50 V/s within potential ranges: -1.70 ÷ +0.10 (1), -1.70 ÷ -0.80 (2); -0.90 ÷ +0.10 (3) V. The porphyrin concentration
is equal to 1 mM
Copyright © 2016 World Scientific Publishing Company
J. Porphyrins Phthalocyanines 2016; 20: 6–9

HYDROXYALKYLOXY SUBSTITUTED TETRAPHENYLPORPHYRINS
7
R1
R1
N
N
N
N
N
N
N
HO2- + R1
O•
R1
M
R1
R2
O
H
M
N
attack
R1
O2•-
attack
N
N
N
R2
M
O
(CH2)n CH OH
products
2
N
n = 1 or 3
R2
R1
O
H
R2
O
(CH2)n CH OH
2
n = 1 or 3
Scheme 3. Hydrogen transfer mechanism of superoxide scavenging in case of hydroxy- and hydroxyalkyloxy substituted
tetraphenylporphyrins.
of H T(4-OH(CH ) OPh)P, H T(4-OH(CH ) OPh)P and
2
2 2
2
2 4
CONCLUSION
ZnT(4-OH(CH ) OPh)P porphyrins either. For antioxidants
2
2
A new approach to superoxide scavenging activity
determination of hydroxyalkyloxy substituted tetra-
phenylporphyrins has been applied. It has been found
that the tested porphyrins have a high activity despite
the absence of the labile hydrogen atom in the porphyrin
structure. The high experimental values of superoxide
scavenging activity of H T(4-OH(CH ) OPh)P, H T(4-
containing phenyl moieties, the hydrogen transfer
mechanism is provided by OH bond dissociation, which
leads to a phenoxyl radical formation. The dissociation
energy of OH bonds is in the range of 75–90 kcal/mol
(depending on the substituents and functional environment)
[27–30]. The dissociation energy of alcohol OH groups is
2
2 2
2
around 105 kcal/mol [64]. This dissociation energy value
does not allow the hydrogen atom to get involved in effective
superoxide scavenging. As a result, a strong decrease in the
superoxide scavenging activity should be observed.
OH(CH ) OPh)P and ZnT(4-OH(CH ) OPh)P can be
2
4
2 4
•-
explained by the effective nucleophilic attack of O2
on the C–C bonds of hydroxyalkyloxy substitutes. It is
supposed that the attack leads to the formation of CH OH
2
The high experimental values of superoxide scav-
enging activity of H T(4-OH(CH ) OPh)P, H T(4-OH
radical with a labile hydrogen atom and the hydrogen
transfer mechanism in what follows.
2
2 2
2
(
CH ) OPh)P and ZnT(4-OH(CH ) OPh)P can be
2
4
2
4
•
-
explained by the effective nucleophilic attack of O2 on
the C–C bonds of alcohol. It is a more favorable process
than OH dissociation (Scheme 3) because C–C bond
Acknowledgements
The authors thank Professor A. S. Semeikin (Ivanovo
State University of Chemistry and Technology) for
developing porphyrins synthesis methods and assistance,
the Upper Volga Region Centre of Physicochemical
Research (G.A. Krestov Institute of Solution Chemistry
of the Russian Academy of Sciences) for technical
support, the Russian Foundation for Basic Research
(grant no. 15-43-03006) for partial financial support.
dissociation leading to CH OH radical formation requires
2
about 85 kcal/mol only [64]. OH bond dissociation energy
in CH OH radical is equal to about 30 kcal/mol [64], so
2
hydrogen atom transfer mechanism is easily realized.
The obtained higher level of antioxidant activity
of the porphyrin-ligand than that of the Zn-porphyrin
is similar to the effects observed for hydroxyphenyl
porphyrins [36] previously. The phenomena, on
the one hand, can be caused by metal influence on
intramolecular transfer of unpaired electrons [37],
which leads to a decrease in the porphyrin oxidation
rate. On the other hand, the metal influences on the
structure of the molecular orbitals and, as a result, on
the bond dissociation energy of moieties involved in
the superoxide scavenging processes.
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