Antioxidant action of the antihypertensive drug, carvedilol, against lipid peroxidation

Article abstract of DOI:10.1016/S0006-2952(99)00417-7

The action of carvedilol, a vasodilating, β-adrenoceptor blocking agent, against lipid peroxidation has been the subject of many studies, but the results reported thus far are contradictory. In an attempt to define the antioxidant mechanism of carvedilol

Full text of DOI:10.1016/S0006-2952(99)00417-7

Biochemical Pharmacology, Vol. 59, pp. 1069–1076, 2000.
© 2000 Elsevier Science Inc. All rights reserved.
ISSN 0006-2952/00/$–see front matter
PII S0006-2952(99)00417-7
Antioxidant Action of the Antihypertensive Drug,
Carvedilol, against Lipid Peroxidation
Noriko Noguchi,* Kimihiro Nishino and Etsuo Niki
RESEARCH CENTER FOR ADVANCED SCIENCE AND TECHNOLOGY, UNIVERSITY OF TOKYO,
MEGURO, TOKYO 153-8904, JAPAN
ABSTRACT. The action of carvedilol, a vasodilating, ␤-adrenoceptor blocking agent, against lipid peroxida-
tion has been the subject of many studies, but the results reported thus far are contradictory. In an attempt to
define the antioxidant mechanism of carvedilol against lipid peroxidation, the dynamics of the action of
carvedilol were studied in several oxidation systems. We investigated the reactivity of carvedilol toward radicals
and its inhibitory effect on lipid peroxidation induced by several kinds of initiating species such as azo
compounds and metal ions in solution, micelles, membranes, and low-density lipoprotein. Carvedilol exerted
poor reactivity toward phenoxyl, alkoxyl, and peroxyl radicals in acetonitrile solution nor did it show an
appreciable antioxidant effect against either the peroxyl radical-induced oxidation of methyl linoleate in
acetonitrile or against phosphatidylcholine liposomal membranes in aqueous suspension. Carvedilol completely
inhibited the ferric ion-induced oxidation of methyl linoleate micelles by sequestering ferric ions, but not by
reducing hydroperoxide. It was shown that carvedilol enhanced the oxidation of micelles induced by either
methemoglobin or peroxyl radical. Carvedilol, which was added exogenously, did not suppress the oxidation of
isolated low-density lipoprotein induced by peroxyl radical or cupric ion. These results show that carvedilol does
not act as a radical-scavenging antioxidant, but that it does act most efficiently as an antioxidant against ferric
ion-induced oxidation by sequestering ferric ion. BIOCHEM PHARMACOL 59;9:1069–1076, 2000. © 2000 Elsevier
Science Inc.
KEY WORDS. carvedilol; antioxidant; lipid peroxidation; radical; ferric ion; methemoglobin
Carvedilol, 1-[carbazole-(4)-oxy]-3-[(2-methoxy-phenoxy-
ethyl)-amino]-2-propanol, is a vasodilator/␤-adrenoceptor
antagonist used for the treatment of hypertension [1, 2].
Carvedilol has been shown to have multiple functions such
as neuroprotection [3–5] and myocardial [6, 7], and endo-
thelial protection [8, 9]. Preventive effects of carvedilol
against smooth muscle cell proliferation have been shown
[10, 11]. It has also been reported that treatment with
carvedilol both in vitro and in vivo increased the resistance
of isolated LDL† against oxidation induced by macrophage
or copper [12, 13]. In many of these studies, antioxidant
properties of carvedilol are thought to account for the
protective effects. The mechanisms of lipid peroxidation
have been well documented [14, 15], and the action of
antioxidants against this peroxidation has also been studied
extensively [15]. The antioxidant properties of carvedilol
have received much attention [4–9, 12, 13, 16–24]. How-
ever, the dynamics of the antioxidant action of carvedilol
are not consistently understood yet. For example, it has
been reported that carvedilol inhibits lipid peroxidation by
scavenging free radicals [18], while another report con-
cludes that carvedilol is not a radical scavenger but rather
sequesters ferric ion [25]. In the present study, the dynamics
of the action of carvedilol as an antioxidant against lipid
peroxidation were studied in several oxidation systems to
elucidate the molecular mechanisms of its antioxidant
action.
MATERIALS AND METHODS
Materials
Carvedilol and probucol were supplied from Daiichi Phar-
maceutical Co.. Troglitazone and 2R,4ЈR,8ЈR-␣-tocopherol
were supplied by Sankyo Co. and Eisai Co., respectively.
Methyl linoleate obtained from Sigma was purified before
use as described previously [26]. The hydrophilic azo com-
pound AAPH and the lipophilic azo compounds AMVN
[27] and MeO-AMVN [28], with the latter used as a peroxyl
radical generator, were purchased from Wako Pure Chem-
ical Industries and used as received. BHN, used as an
alkoxyl radical generator, was prepared as previously re-
* Corresponding author: Dr. Noriko Noguchi, Research Center for Ad-
vanced Science and Technology, The University of Tokyo, 4-6-1 Komaba,
Meguro, Tokyo 153-8904, Japan. Tel. 81-35452-5204; FAX 81-35452-
5201; E-mail: nonoriko@oxygen.rcast.u-tokyo.ac.jp
† Abbreviations: LDL, low-density lipoprotein; AAPH, 2,2Ј-azobis(2-
amidinopropane) dihydrochloride; AMVN, 2,2Ј-azobis(2,4-dimethyl-
valeronitrile);
MeO-AMVN,
2,2Ј-azobis(4-methoxy-2,4-dimethyl-
valeronitrile); BHN, di-tert-butylhyponitrite; DPPD, N,NЈ-diphenyl-1,4-
phenylenediamine; DPBQ, N,NЈ-diphenyl-1,4-benzoquinone diimine;
BOOH, tert-butyl hydroperoxide; MeLOOH, methyl linoleate hydroper-
oxide; and PC, phosphatidylcholine.
Received 13 April 1999; accepted 14 October 1999.

1070
N. Noguchi et al.
ported [29]. LDL was separated from human plasma of
healthy donors and by ultracentrifugation within a density
cut-off of 1.019 to 1.063 g/mL, followed by dialysis as
described previously [30]. Galvinoxyl, DPPD, and other
chemicals were of the highest grade available commercially.
Interaction of Antioxidant with Galvinoxyl
The interactions of carvedilol, probucol, troglitazone, or
␣-tocopherol with galvinoxyl were measured in acetonitrile
with a spectrophotometer equipped with a rapid-mixing
stopped-flow apparatus (RX-1000, Applied Photophysics)
by following the decrease in maximum absorption of galvi-
noxyl at 429 nm [31].
FIG. 1. Reaction of carvedilol with galvinoxyl. Antioxidant (10
␮M) and galvinoxyl (5 ␮M) in acetonitrile were reacted, and the
consumption of galvinoxyl was followed by its absorption at 429
nm. The data for ␣-tocopherol, troglitazone, and probucol are
also included for comparison.
Estimation of Reactivity of Antioxidant toward Alkoxyl
and Peroxyl Radicals
The reactivities of carvedilol toward alkoxyl and peroxyl
radicals were estimated from the competition with DPPD as
follows. One molecule of DPPD reacts rapidly with two
molecules of alkoxyl or peroxyl radical to give DPBQ,
which has a strong absorption at 440 nm [28, 32]. Appro-
priate amounts of DPPD and either BHN or AMVN were
incubated in acetonitrile at 37° under air in the absence or
presence of antioxidant, and the formation of DPBQ was
followed spectrophotometrically at 440 nm. The reactivity
of the antioxidant toward alkoxyl and peroxyl radicals was
assessed from the extent of reduction of formation of DPBQ
by the test antioxidant.
induced by adding BHN or AMVN to aqueous dispersion as
a methanol solution. AAPH, CuCl2, FeCl3, or methemo-
globin was added as an aqueous solution. Hydrogen perox-
ide, BOOH, or MeLOOH was added as a methanol solu-
tion. The rate of oxidation was followed by measuring
oxygen uptake with an oxygen monitor equipped with a
Clark-type oxygen electrode (YSI Model 5300).
Oxidation of LDL
The oxidation of LDL (0.1 mg protein/mL PBS) was
induced by either CuCl2 or MeO-AMVN. Carvedilol was
added as a methanol solution prior to the induction of the
oxidation. The oxidation was followed by measuring the
increase in conjugated diene formation with a spectropho-
tometer at 234 nm [36].
Inhibition of Oxidation of Lipids by Carvedilol
The oxidations of methyl linoleate and soybean PC lipo-
somes were carried out at 37° in air. Methyl linoleate was
incubated in acetonitrile in the presence of AMVN and an
appropriate amount of antioxidant, and the formation of
methyl linoleate hydroperoxides was measured by HPLC
equipped with a reversed-phase LC-18 column (particle size
5 ␮m; 4.6 mm ϫ 25 cm; Supelco). Methanol: H2O (95:5,
v/v) was used as an eluent at a flow rate of 1.0 mL/min, and
the hydroperoxides were detected by UV absorption at 234
nm [33]. The multilamellar PC liposomal membranes were
prepared as reported previously [34]. PC, AMVN, and
antioxidants, when used, were incorporated into the mem-
branes simultaneously by dissolving them into the solvent
before preparation of the membranes. The PC hydroperox-
ides were analyzed by absorption at 234 nm by HPLC using
an LC-Si column (particle size 5 ␮m; 4.6 mm ϫ 25 cm;
Supelco), and methanol: 40 mM phosphate buffer (90:10
v/v) was used as an eluent at a flow rate of 1.0 mL/min.
The oxidation of methyl linoleate was performed in
aqueous dispersions at 37° in air. The methyl linoleate
micelles were prepared as reported previously [35] by using
SDS as a surfactant. Carvedilol was added as a methanol
solution at the time of preparation of micelles or at an
appropriate time during oxidation. The oxidation was
RESULTS
Reactivity of Carvedilol toward Radical
As mentioned above, some reports indicate radical-scav-
enging activity of carvedilol, whereas other reports deny it.
Hence, the reactivities of carvedilol toward radicals were
first measured. Galvinoxyl radical has often been used in
the estimation of reactivity of antioxidant toward radicals.
Potent radical-scavenging antioxidants having reactive hy-
drogens react rapidly with galvinoxyl. It was found that
while ␣-tocopherol and troglitazone very rapidly reduced
the absorption of galvinoxyl radical at 429 nm, carvedilol
did not reduce it at an appreciable rate (Fig. 1). The
capability of carvedilol to act as a hydrogen donor is even
smaller than that of probucol [33]. Secondly, the reactivi-
ties toward alkoxyl and peroxyl radicals were measured. The
actual rate of free radical flux was measured by using DPPD
as a radical scavenger. One molecule of DPPD scavenges
two radicals rapidly to give DPBQ [28, 32], which has a

Antioxidant Action of Carvedilol
1071
FIG. 2. Reactivity of carvedilol toward alkoxyl radical and peroxyl radical. DPPD (100 ␮M) was incubated with either (A) BHN (20
mM) or (B) AMVN (40 mM) at 37° in the absence (no antioxidant [IH]) and presence of antioxidant in acetonitrile, and the formation
of DPBQ was followed at 440 nm. While a radical-scavenging antioxidant such as ␣-tocopherol (100 ␮M) suppressed the formation
of DPBQ, carvedilol showed little effect.
strong absorption at 440 nm. This absorption increased
with time due to the formation of DPBQ from DPPD via
the reaction with the radicals generated from BHN or
AMVN. In Fig. 2 is shown the increase in absorbance at
440 nm observed when DPPD and BHN or AMVN were
incubated at 37° in air. These radical initiators induced the
increase in the absorption at 440 nm linearly with time in
the absence of antioxidant, the absorbance became con-
stant when all the DPPD had been converted to DPBQ.
The plateau absorbance was directly proportional to the
initial DPPD concentration. The addition of ␣-tocopherol
to this mixture suppressed the formation of DPBQ, appar-
ently because ␣-tocopherol competed with DPPD in scav-
enging alkoxyl or peroxyl radicals. On the other hand,
carvedilol exerted little effect on the formation of DPBQ,
suggesting that carvedilol was not capable of competing with
DPPD in scavenging these radicals.
branes. Antioxidant and AMVN were incorporated into
multilamellar liposomal membranes simultaneously. The re-
sults summarized in Fig. 3B show that ␣-tocopherol efficiently
suppressed the oxidation of PC membranes and produced a
clear induction period. The rates of oxidation after the
induction period were the same as that in the absence of
antioxidant. Carvedilol acted as a weak antioxidant and
reduced the rate of oxidation at the high concentration.
Antioxidant Activity of Carvedilol in Oxidation of
Methyl Linoleate Micelles
The antioxidant action of carvedilol against the oxidations
of methyl linoleate micelles in aqueous dispersions, which
were induced by several initiating species, was studied. The
extent of oxidation was followed by measuring oxygen
consumption. It was found that carvedilol did not inhibit
the oxidation induced by alkoxyl radical (data not shown)
and even enhanced that induced by peroxyl radical (Fig. 4).
It was found that an equimolar amount of carvedilol
completely inhibited the oxidation of micelles induced by
ferric chloride. The complete inhibition of ferric ion-
induced oxidation by carvedilol was observed under the
condition in which the ratio of carvedilol to ferric ion was
higher than 1 (Fig. 5A). Addition of an excess amount of
either MeLOOH or BOOH did not resume the oxidation
(Fig. 5B).
The antioxidant activity of carvedilol was assessed in the
oxidation of micelles induced by methemoglobin. Al-
though methemoglobin contains ferric ion in the molecule,
carvedilol did not inhibit methemoglobin-induced oxida-
tion of micelles, but rather enhanced it significantly (Fig.
6). The enhancement of the reaction of methemoglobin-
induced oxidation by carvedilol in micelles was confirmed
by following a change in the spectrum of methemoglobin in
the absence and presence of carvedilol. Methemoglobin
exhibited absorption at 400 nm in methyl linoleate micelle
Antioxidant Action of Carvedilol against Lipid
Peroxidation in Solution and Liposomal Membranes
The antioxidant activity of carvedilol was assessed in the
oxidation of methyl linoleate induced by AMVN in aceto-
nitrile solution. The rate of spontaneous oxidation of
methyl linoleate in acetonitrile at 37° under air was quite
small (data not shown), but the addition of AMVN
induced the oxidation and the accumulation of methyl
linoleate hydroperoxide was observed at a constant rate
without any appreciable induction period (Fig. 3A). A
small amount of troglitazone suppressed the oxidation and
produced a clear induction period, after which oxidation
proceeded at a similar rate as oxidation without antioxidant.
Similar results have been observed for ␣-tocopherol. On the
other hand, the antioxidant activity of carvedilol was ex-
tremely small. Liposomal membranes are often used as a model
of biological membranes. The antioxidant action of carvedilol
was studied in the oxidations of soybean PC liposomal mem-

1072
N. Noguchi et al.
FIG. 3. Effect of carvedilol on lipid peroxidation in solution and membranes induced by peroxyl radical. (A) Methyl linoleate (453 mM)
was oxidized in acetonitrile in the presence of AMVN (0.20 mM) without antioxidant (E), with 20 (‚) or 200 ␮M (ᮀ) carvedilol,
or with 5 ␮M troglitazone (F) at 37° in air, and MeLOOHs were measured as described in Materials and Methods. (B) Soybean PC
(5.15 mM) multilamellar liposomal membranes containing AMVN (0.50 mM) without antioxidant (E), with 20 (‚) or 200 ␮M (ᮀ)
carvedilol, or with 2 ␮M ␣-tocopherol (Œ) were incubated at 37° in air, and the formation of PC hydroperoxides (PCOOH) was
followed by HPLC as described in Materials and Methods.
dispersion, with the absorbance decreasing with time (Fig.
7A). Carvedilol showed no absorption at about 400 nm and
enhanced the change in the absorbance of methemoglobin
(Fig. 7B).
234 nm. As expected, carvedilol did not show an inhibitory
effect on the peroxyl radical-induced oxidation of LDL
(data not shown). It was also shown that carvedilol did not
reduce the rate of formation of conjugated diene in the
copper-induced oxidation of LDL to an appreciable degree
(Fig. 8).
Inhibition of Oxidation of LDL by Carvedilol
The antioxidant activity of carvedilol in the oxidation of
LDL is of great interest. The oxidation of human LDL was
carried out by using either cupric ion or a lipophilic radical
initiator, MeO-AMVN, and the extent of the oxidation
was measured by following the increase in absorbance at
DISCUSSION
The antioxidant exerts its function by different mecha-
nisms such as radical scavenging, peroxide reduction, and
metal ion sequestration [37]. Peroxyl radical is one of the
FIG. 4. Effect of carvedilol on peroxyl radical-induced oxidation of methyl linoleate micelles. Oxidation of methyl linoleate micelles (75
mM/0.5 M SDS in H2O) was induced by either (A) a hydrophilic radical initiator, AAPH (2 mM), or (B) a lipophilic radical initiator,
AMVN (2 mM), at 37°. The oxidation was followed by oxygen consumption, and carvedilol (100 ␮M) was added as a methanol
solution at the point indicated in the figure.

Antioxidant Action of Carvedilol
1073
FIG. 5. Inhibition of ferric ion-induced oxidation of methyl linoleate micelles by carvedilol. (A) Oxidation of methyl linoleate micelles
(75 mM/0.5 M SDS in H2O) was induced by 50 ␮M FeCl3 and inhibited by equimolar carvedilol. (B) Oxidation of methyl linoleate
micelles (75 mM/0.5 M SDS in H2O) was induced by 100 ␮M FeCl3 and inhibited by equimolar carvedilol. The oxidation was not
restarted by the addition of MeLOOH (100 ␮M) or BOOH (300 ␮M).
most important radicals to act as a chain-carrying species in
lipid peroxidation. Antioxidants such as ␣-tocopherol and
ubiquinol, which scavenge peroxyl radical very rapidly, act
as chain-breaking antioxidants. We first investigated the
reactivity of carvedilol toward radicals. It was shown clearly
in Figs. 1 and 2 that carvedilol does not act as a potent
radical scavenger. A carbazole group of carvedilol has been
attributed to its antioxidant properties. It has previously
been shown [38, 39] that carbazoles with a free hydroxyl
group suppress oxidation effectively but those without do
not. Since the carbazole group of carvedilol does not have
a free hydroxyl group, poor activity of carvedilol as a radical
scavenger seems reasonable. On the other hand, Yue et al.
[18] reported that carvedilol acted as a radical-scavenging
antioxidant, which is not consistent with our results.
However, they used iron as an initiating species, and their
results can also be explained by sequestration of iron by
carvedilol.
Secondly, the hydroperoxide-reducing activity of carve-
dilol was studied. The oxidation of methyl linoleate mi-
celles induced by ferric ion is dependent on MeLOOH,
which is contained in the micelles [35, 40]. It was previ-
ously shown that ebselen, which mimics glutathione per-
oxidase, completely inhibited metal-induced lipid peroxi-
dation by reducing seeding hydroperoxide [40–43]. If
carvedilol inhibits ferric ion-induced oxidation of micelles
by reducing hydroperoxide, the addition of an excess
amount of hydroperoxide should resume oxidation. Since,
as shown in Fig. 5B, the addition of a high concentration of
MeLOOH and BOOH showed no effect, the reducing
activity of hydroperoxide by carvedilol is ruled out. Indeed,
we confirmed that cholesteryl ester hydroperoxide was not
FIG. 6. Effect of carvedilol on oxidation of methyl linoleate micelles induced by methemoglobin. (A) Oxidation of methyl linoleate
micelles (15 mM/0.5 M SDS in H2O) was induced by 1 ␮M methemoglobin (mtHb). The addition of carvedilol (100 ␮M) after onset
of the oxidation showed no effect. (B) When methyl linoleate micelles (15 mM/0.5 M SDS in H2O) containing 100 ␮M carvedilol were
oxidized by 1 ␮M mtHb, the rate of oxidation was greater than that without carvedilol.

1074
N. Noguchi et al.
FIG. 7. Enhancement by carvedilol of the change in spectrum of methemoglobin in the oxidation of micelles. Methemoglobin (1 ␮M)
was incubated with methyl linoleate micelles (15 mM/0.5 M SDS in H2O) in the absence (A) and presence of 100 ␮M carvedilol (B)
at 37°. The spectrum was measured every 3 min.
reduced by carvedilol. On the other hand, carvedilol
inhibited the ferric ion-induced oxidation of micelles in a
concentration-dependent manner. The equimolar carve-
dilol to ferric ion showed a complete inhibitory effect. The
addition of an excess amount of ferric ion to the oxidation
system in which carvedilol stopped oxidation restarted this
oxidation. These results are in agreement with those of
Tadolini and Franconi [25], who showed that carvedilol
could make a complex with ferric ion in a 1:1 ratio in
methanol/H2O solution. The present study showed that
carvedilol, a lipophilic compound which is located in the
lipid phase, could react with ferric ion added to the aqueous
phase. The rate of oxidation of micelles induced by a
certain concentration of ferric ion was the same as that
induced by the same concentration of ferric ion which
remained after sequestration by carvedilol in a 1:1 ratio
(data not shown).
peroxidation induced by methemoglobin, which may be
important in vivo. Although methemoglobin contains ferric
ion, contrary to our expectations, carvedilol did not inhibit
the methemoglobin-induced oxidation of methyl linoleate
micelles but even enhanced it. The reason for this is not
clear at present. Further investigation of the several differ-
ent oxidation systems is required.
Finally, we studied the antioxidant potency of carvedilol
against LDL oxidation induced by either a radical initiator
or copper ion. As observed in other oxidation systems,
carvedilol did not inhibit the peroxyl radical-induced
oxidation of LDL. The antioxidant effect of carvedilol on
the copper-induced oxidation of LDL has been studied both
in vivo and in vitro [12, 13]. In the present study, it was
shown that carvedilol did not suppress the oxidation of
LDL induced by copper to an appreciable degree. We also
confirmed that carvedilol did not inhibit the copper-
induced oxidation of methyl linoleate micelles (data not
shown). These results are not in agreement with those of
Yue et al. [12], who observed antioxidant activity of
carvedilol against LDL oxidation induced by macrophage
and copper. It is known that metal is required for oxidation
by macrophage, and it may be possible that carvedilol
suppressed LDL oxidation induced by macrophage by se-
questering iron. The effect of carvedilol on copper-induced
LDL oxidation was reported to be less significant than that
for macrophage-induced oxidation [12]. Furthermore, the
thiobarbituric acid assay used in their work is affected by
metal ion and chelating agents. It was reported that
treatment with carvedilol in vivo increased the lag time in
copper-induced LDL oxidation [13]. Considering the con-
centration of carvedilol in plasma of patients treated with
carvedilol [44], it is difficult to explain the antioxidant
effect in LDL particle by carvedilol alone, but its metabo-
lite, having a hydroxyl group on the phenyl ring, might act
as an antioxidant against LDL oxidation [45].
We investigated the action of carvedilol against lipid
FIG. 8. Effect of carvedilol on oxidation of LDL induced by
copper. LDL (0.1 mg protein/mL PBS) was oxidized by 2 ␮M
CuCl2 in the absence (E) or presence of 5 (F) and 10 ␮M (Œ)
carvedilol at 37° in air. The oxidation was followed by measur-
ing the increase in absorbance at 234 nm.
It may be concluded that carvedilol does not act as a

Antioxidant Action of Carvedilol
1075
dilating ␤-blocker with the potential for cardiovascular organ
protection. Eur Heart J 17 (Suppl B): 24–29, 1996.
17. Yue TL, McKenna PJ, Ruffolo RR Jr and Feuerstein GZ,
Carvedilol, a new ␤-adrenoceptor antagonist and vasodilator
antihypertensive drug, inhibits superoxide release from hu-
man neutrophils. Eur J Pharmacol 214: 277–280, 1992.
18. Yue TL, Cheng HY, Lysko PG, McKenna PJ, Feuerstein R,
Gu JL, Lysko KA, Davis LL and Feuerstein GZ, Carvedilol, a
new vasodilator and ␤-adrenoceptor antagonist, is an antiox-
idant and free radical scavenger. J Pharmacol Exp Ther 263:
92–98, 1992.
radical-scavenging antioxidant and does not reduce hy-
droperoxides, but that it does act as an antioxidant against
iron-induced lipid peroxidation by sequestering ferric ion.
References
1. Feuerstein GZ and Ruffolo RR Jr, Carvedilol, a novel multiple
action antihypertensive agent with antioxidant activity and
the potential for myocardial and vascular protection. Eur
Heart J 16 (Suppl F): 38–42, 1995.
19. Christopher TA, Lopez BL, Yue TL, Feuerstein GZ, Ruffolo
RR Jr and Ma X, Carvedilol, a new ␤-adrenoceptor blocker,
vasodilator and free-radical scavenger, exerts an anti-shock
and endothelial protective effect in rat splanchnic ischemia
and reperfusion. J Pharmacol Exp Ther 273: 64–71, 1995.
20. Lopez BL, Christopher TA, Yue TL, Ruffolo RR Jr, Feuerstein
GZ and Ma XL, Carvedilol, a new ␤-adrenoceptor blocker
antihypertensive drug, protects against free-radical-induced
endothelial dysfunction. Pharmacology 51: 165–173, 1995.
21. Kramer JH and Weglicki WB, A hydroxylated analog of the
␤-adrenoceptor antagonist, carvedilol, affords exceptional
antioxidant protection to postischemic rat hearts. Free Radic
Biol Med 21: 813–825, 1996.
22. Ma XL, Yue TL, Lopez BL, Barone FC, Christopher TA,
Ruffolo RR Jr and Feuerstein GZ, Carvedilol, a new ␤-adre-
noceptor blocker and free radical scavenger, attenuates myo-
cardial ischemia–reperfusion injury in hypercholesterolemic
rabbits. J Pharmacol Exp Ther 277: 128–136, 1996.
23. Ruffolo RR Jr and Feuerstein GZ, Pharmacology of carvedilol:
Rationale for use in hypertension, coronary artery disease, and
congestive heart failure. Cardiovasc Drugs Ther 11 (Suppl 1):
247–256, 1997.
24. Rabasseda X, Carvedilol: An effective antihypertensive drug
with antiischemic/antioxidant cardioprotective properties.
Drugs Today 34: 905–926, 1998.
25. Tadolini B and Franconi F, Carvedilol inhibition of lipid
peroxidation. A new antioxidative mechanism. Free Radic Res
29: 377–387, 1998.
26. Komuro E, Takahashi M, Morita T, Tsuchiya J, Arakawa Y,
Yamamoto Y, Niki E, Sugioka K and Nakano M, Inhibition of
peroxidations of lipids and membranes by estrogens. J Phys
Org Chem 3: 309–315, 1990.
27. Niki E, Free radical initiators as source of water- or lipid-
soluble peroxyl radicals. In: Methods in Enzymology (Eds.
Packer L and Glazer AN), Vol. 186, pp. 100–108. Academic
Press, New York, 1990.
2. Dunn CJ, Lea AP and Wagstaff AJ, Carvedilol. A reappraisal
of its pharmacological properties and therapeutic use in
cardiovascular disorders. Drugs 54: 161–185, 1997.
3. Lysko PG, Lysko KA, Webb CL and Feuerstein GZ, Neuro-
protective effects of carvedilol, a new antihypertensive, at the
N-methyl-D-aspartate receptor. Neurosci Lett 148: 34–38,
1992.
4. Lysko PG, Lysko KA, Yue TL, Webb CL, Gu JL and
Feuerstein GZ, Neuroprotective effects of carvedilol, a new
antihypertensive agent, in cultured rat cerebellar neurons and
in gerbil global brain ischemia. Stroke 23: 1630–1635, 1992.
5. Yue TL, Lysko PG, Brone FC, Gu JL, Ruffolo RR Jr and
Feuerstein GZ, Carvedilol, a new antihypertensive drug with
unique antioxidant activity: Potential role in cerebroprotec-
tion. Ann N Y Acad Sci 738: 230–242, 1994.
6. Feuerstein GZ, Yue TL, Cheng HY and Ruffolo RR Jr,
Myocardial protection by the novel vasodilating beta-blocker,
carvedilol: Potential relevance of anti-oxidant activity. J Hy-
pertens 11 (Suppl 1): S41–S48, 1993.
7. Sugawara H, Tobise K and Kikuchi K, Antioxidant effects of
calcium antagonists on rat myocardial membrane lipid per-
oxidation. Hypertens Res 19: 223–228, 1996.
8. Yue TL, McKenna PJ, Gu JL, Cheng HY, Ruffolo RR Jr and
Feuerstein GZ, Carvedilol, a new antihypertensive agent,
prevents lipid peroxidation and oxidative injury to endothe-
lial cells. Hypertension 22: 922–928, 1993.
9. Yue TL, McKenna PJ, Gu JL, Cheng HY, Ruffolo RR Jr and
Feuerstein GZ, Carvedilol, a new vasodilating ␤ adrenoceptor
blocker antihypertensive drug, protects endothelial cells from
damage initiated by xanthine–xanthine oxidase and neutro-
phils. Cardiovasc Res 28: 400–406, 1994.
10. Ohlstein EH, Douglas SA, Sung CP, Yue TL, Louden C,
Arleth A, Poste G, Ruffolo RR Jr and Feuerstein GZ,
Carvedilol, a cardiovascular drug, prevents vascular smooth
muscle cell proliferation, migration, and neointimal forma-
tion following vascular injury. Proc Natl Acad Sci USA 90:
6189–6193, 1993.
28. Noguchi N, Yamashita H, Gotoh N, Yamamoto Y, Numano
R and Niki E, 2,2Ј-Azobis (4-methoxy-2,4-dimethylvaleroni-
trile), a new lipid-soluble azo initiator: Application to oxida-
tions of lipids and low-density lipoprotein in solution and in
aqueous dispersions. Free Radic Biol Med 24: 259–268, 1998.
29. Kigoshi M, Sato K and Niki E, Oxidations of lipids induced by
dioctadecyl hyponitrite and di-t-butyl hyponitrite in organic
solution and aqueous dispersions. Effect of reaction medium
and size of radicals on efficiency of chain initiation. Bull Chem
Soc Jpn 66: 2954–2959, 1993.
30. Ramos P, Gieseg SP, Schuster B and Esterbauer H, Effect of
temperature and phase transition on oxidation resistance of
low density lipoprotein. J Lipid Res 36: 2113–2128, 1995.
31. Iwatsuki M, Komuro E and Niki E, Antioxidant activities of
aminophenols against oxidation of methyl linoleate in solu-
tion. Bull Chem Soc Jpn 68: 620–624, 1995.
11. Sung CP, Arleth AJ and Ohlstein EH, Carvedilol inhibits
vascular smooth muscle cell proliferation. J Cardiovasc Phar-
macol 21: 221–227, 1993.
12. Yue TL, McKenna PJ, Lysko PG, Ruffolo RR Jr and Feuer-
stein GZ, Carvedilol, a new antihypertensive, prevents oxi-
dation of human low density lipoprotein by macrophages and
copper. Atherosclerosis 97: 209–216, 1992.
13. Maggi E, Marchesi E, Covini D, Negro C, Perani G and
Bellomi G, Protective effects of carvedilol, a vasodilating
␤-adrenoceptor blocker, against in vivo low density lipopro-
tein oxidation in essential hypertension. J Cardiovasc Pharma-
col 27: 532–538, 1996.
14. Porter NA, Caldwell SE and Mills KA, Mechanisms of free
radical oxidation of unsaturated lipids. Lipids 30: 277–290,
1995.
15. Noguchi N and Niki E, Chemistry of active oxygen species
and antioxidants, In: Antioxidant Status, Diet, Nutrition, and
Health (Ed. Papas AM), pp. 3–20. CRC Press, Boca Raton,
1999.
32. Takahashi M, Tsuchiya J and Niki E, Oxidation of lipids.
XVI. Inhibition of autoxidation of methyl linoleate by dia-
rylamines. Bull Chem Soc Jpn 62: 1880–1884, 1989.
33. Gotoh N, Shimizu K, Komuro E, Tsuchiya J, Noguchi N and
16. Feuerstein GZ and Ruffolo RR Jr, Carvedilol, a novel vaso-

1076
N. Noguchi et al.
Niki E, Antioxidant activities of probucol against lipid
Action of ebselen as an antioxidant against lipid peroxida-
tion. Biochem Pharmacol 44: 39–44, 1992.
peroxidations. Biochim Biophys Acta 1128: 147–154, 1992.
34. Yamamoto Y, Niki E, Kamiya Y and Shimasaki H, Oxidation
of lipids. VII. Oxidation of phosphatidylcholines in homoge-
neous solution and in water dispersion. Biochim Biophys Acta
795: 332–340, 1984.
41. Maiorino M, Roveri A and Ursini F, Antioxidant effect of
Ebselen (PZ 51): Peroxidase mimetic activity on phospholipid
and cholesterol hydroperoxides vs free radical scavenger
activity. Arch Biochem Biophys 295: 404–409, 1992.
42. O’Leary VJ, Darley-Usmar VM, Russell LJ and Stone D,
Pro-oxidant effects of lipoxygenase-derived peroxides on cop-
per-initiated oxidation of low-density lipoprotein. Biochem J
282: 631–634, 1992.
43. Noguchi N, Gotoh N and Niki E, Effects of ebselen and
probucol on oxidative modifications of lipid and protein of
low density lipoprotein induced by free radicals. Biochim
Biophys Acta 1213: 176–182, 1994.
44. Saijonmaa O, Metsarinne K and Fyhrquist F, Carvedilol and
its metabolites suppress endothelin-1 production in human
endothelial cell culture. Blood Press 6: 24–28, 1997.
45. Yue TL, McKenna PJ, Lysko PG, Gu JL, Lysko KA, Ruffolo
RR Jr and Feuerstein GZ, SB 211475, a metabolite of
carvedilol, a novel antihypertensive agent, is a potent anti-
oxidant. Eur J Pharmacol 251: 237–243, 1994.
35. Yoshida Y and Niki E, Oxidation of methyl linoleate in
aqueous dispersions induced by copper and iron. Arch Biochem
Biophys 295: 107–114, 1992.
36. Esterbauer H, Striegel G, Puhl H and Rotheneder M, Con-
tinuous monitoring of in vitro oxidation of human low density
lipoprotein. Free Radic Res Commun 6: 67–75, 1989.
37. Niki E, Noguchi N, Tsuchihashi H and Gotoh N, Interaction
among vitamin C, vitamin E, and ␤-carotene. Am J Clin Nutr
62 (Suppl 6): 1322S–1326S, 1995.
38. Yamamoto Y, Yamamoto Y, Niki E, Gee CA and Willson RL,
Antioxidant activity of 6-hydroxy-1,4-dimethylcarbazole. J
Jpn Oil Chem Soc 41: 622–628, 1992.
39. Iwatsuki M, Niki E and Kato S, Antioxidant activities of
natural and synthetic carbazoles. Biofactors 4: 123–128, 1993.
40. Noguchi N, Yoshida Y, Kaneda H, Yamamoto Y and Niki E,