547

Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553

International Journal of Medical Research
&
Health Sciences
www.ijmrhs.com Volume 3 Issue 3 Coden: IJMRHS Copyright @2014 ISSN: 2319-5886
Received: 15
th
Mar 2014 Revised: 24
th
Apr 2014 Accepted: 29
th
Apr 2014
Research Article
EFFECTS OF SCIRPUSIN B, A POLYPHENOL IN PASSION FRUIT SEEDS, ON THE CORONARY
CIRCULATION OF THE ISOLATED PERFUSED RAT HEART
Yutaka Matsumoto
1
, Nozomi Gotoh
1
, Shoko Sano
2
, Kenkichi Sugiyama
2
, Tatsuhiko Ito
2
, Yohei Abe
3
, Yumi
Katano
1
, *Akira Ishihata
1
1
Division of Theoretical Nursing and Pathophysiology, Yamagata University School of Medicine, Japan
2
Research Institute, Morinaga & Co.,Ltd.
3
Department of Pharmacy, Yamagata University School of Medicine, Japan
*Corresponding author email:comedic2013@gmail.com
ABSTRACT
Objective:Piceatannol, a polyphenol which is contained in passion fruits seed, is a derivative of resveratrol and is
known to have antioxidant, anti-inflammatory and vasorelaxing activities. Passion fruits seed also contains a
dimer of Piceatannol, Scirpusin B. The aim of this study was to investigate the effect of Scirpusin B on the
coronary circulation of the isolated rat heart. Methods: Hearts were isolated from male Fischer 344 rats (5 - 6
months old), and perfused with modified Krebs-Henseleit solution aerated with 95% O
2
and 5% CO
2
(37 C) at
constant pressure (75 cmH
2
O) by Langendorff's method. Piceatannol or Scirpusin B (10, 30 and 100 M)was
injected as a bolus into the aortic cannula and coronary flow (CF) was continuously measured by the
electromagnetic flow meter. In some experiments, rat hearts were pretreated with L-NAME (an inhibitor of nitric
oxide synthase) or Diclofenac (an inhibitor of cyclooxygenase) to reveal the possible involvement of nitric oxide
(NO) and vasodilating prostanoids in the effect of Scirpusin B. Results:Scirpusin B increased CF up to 108.2 %
of the initial value, while Piceatannol did not increase CF. In addition;Scirpusin B increased CF concentration-
dependently. Pretreatment with L-NAME or Diclofenac significantly attenuated the Scirpusin B-induced coronary
vasodilatation. Scirpusin B did not change the heart rate either left ventricular pressure. Conclusion:This study
shows that Scirpusin B could increase CF via production of NO and vasodilating prostanoids.
Key words: Scirpusin B, Piceatannol, coronary flow, NO, Perfused heart
INTRODUCTION
Over the past few decades, patients suffering from
arteriosclerosis have been increasing in many
countries. Arteriosclerosis causes a wide variety of
complications such as coronary heart disease (CHD),
heart failure, cerebral infarction and arteriosclerosis.
Mortality in CHD in France is about 25% of that in
Britain, although the major risk factors are similar.
1
In
spite of a high intake of saturated fats commonly
found in the French diet, the low rate of CHD in
France compared with other developed countries with
comparable diets, has been called the French
Paradox.
2
In this epidemiological paradox, it has
been speculated that a higher intake of red wine with
Resveratrol(3,5,4-trihydroxy-trans-stilbene) may be
correlated with the incidence of myocardial infarction
in France, which is about 40 percent lower than that
in other European countries.
2
It has been reported that
Resveratrol is converted in vivo to Piceatannol
(3,4,3,5-tetrahydroxy-trans-stilbene) via the
cytochrome P450 enzyme CYP1B1.
3
The only
DOI: 10.5958/2319-5886.2014.00394.4
548
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
difference between Resveratrol and Piceatannol is the
presence of an extra hydroxyl group in one of the
aromatic rings of Piceatannol. Piceatannol and
Resveratrol are phenolic compounds produced
naturally in grapes and red wine.
4
In general,
phenolic compounds have been recognized as
important natural anti-oxidants found in many kinds
of foods and plants. Resveratrol has strong anti-
oxidative,
5,6
anti-inflammatory,
7
anti-cancer,
8,9
melanogenesis inhibitory,
5,10
and collagen synthesis
activating effects
10
. Passion fruits (Passiflora edulis),
especially the seeds of passion fruits contain
Piceatannol in a natural state, which is nearly 50
times larger than that found in grapes.
10,11
Actually,
88% of the total polyphenols are contained in the
seeds.
10
In addition, the extract of passion fruit seeds
contains another polyphenol named Scirpusin B,
which is a dimmer of Piceatannol (Fig. 1).Previous
studies reported that Scirpusin B also shows the
strong vasorelaxant effect in the rat thoracic aorta
probably due to nitric oxide generation,
12
superoxide
anion scavenging activity
13
as well as DPPH radical
scavenging activity.
12
It is very interesting that
scirpsin B was demonstrated to be more potent in the
aortic vasorelaxing effect and in radical scavenging
effect than Piceatannol.
12
In addition, it has been
reported that Scirpusin B could improve glucose
metabolism to prevent the postprandial elevation of
blood glucose.
14
These properties strongly suggest
that Scirpusin B could have vasodilatating effect and
anti-arteriosclerotic effect, those are clinically
important especially in maintaining blood supply to
myocardium. Therefore, the aim of this paper is to
investigate the effect of Scirpusin B isolated from
passion fruit seeds on the coronary circulation of the
perfused rat heart.
MATERIALS AND METHODS
Animals and Ethics; Experiments were performed in
accordance with the Guide for Care and Use of
Laboratory Animals by US National Institute of
Health and in accordance with the Regulation OF
Animal Experiment in Yamagata University
15
under
the regulation of the Animal Care Committee of
Yamagata University School of Medicine
(Identification number 25072). In this study, 5 - 6
months old (340 - 370 g) male Fischer rats (27
animals in total) obtained from Charles River Japan
(Atsugi, Japan) were used. Each rat was used for the
experiment of the isolated heart during the ages
between 20 to 26 weeks-old to avoid ageing effects.
Female rats were not used because of the possible
effects of changes in sex hormone levels (i.e. estrogen
and progesterone). Rats were bred in the Laboratory
Animal Center, Yamagata University School of
Medicine. They were maintained on standard rat
chow with water ad libitum. Scirpusin B was
extracted from passion fruit seeds and purified by
high-performance liquid chromatography (HPLC).
12
In brief, the extracts of passion fruit seeds were
fractionated by reverse-phase HPLC. Each fraction
was collected by an Inertsil ODS-3 column (GL
Sciences Inc., Tokyo, Japan) with conditions of a (A)
water and/or (B) acetonitrile mobile phase at a flow
rate of 5 mL/min. A gradient elution of 0 80% (B)
at 0 90 min was used for fractionation. The
fractionated samples were analyzed by using ODS-3
column. The analytical HPLC was carried out with
(A) water and/or (B) acetonitrile mobile phase at a
flow rate of 0.75 mL/min. A gradient elution of 0
45% (B) at 0 25 min was used for this analysis.
Fig 1: Chemical structures of Piceatannol and
Scirpusin B
Measurement of the coronary flow in isolated
perfused hearts
Fischer-344 rats were deeply anesthetized with
diethyl ether and sacrificed; then the hearts were
quickly excised by performing thoracotomies and
placed in an ice-cold solution of modified Krebs-
Henseleit (118 mM NaCl, 4.7 mM KCl, 24.9 mM
NaHCO
3
, 1.18 mM MgSO
4
, 1.18 mM KH
2
PO
4
, 1.8
mM CaCl
2
, 5.0 mM glucose, 2.0 mM pyruvic acid,
0.057 mM ascorbic acid). The isolated hearts were
immediately perfused by Langendorffs method
under constant pressure (75 cm H
2
O) with modified
Krebs-Henseleit solution. The buffer solution was
548
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
difference between Resveratrol and Piceatannol is the
presence of an extra hydroxyl group in one of the
aromatic rings of Piceatannol. Piceatannol and
Resveratrol are phenolic compounds produced
naturally in grapes and red wine.
4
In general,
phenolic compounds have been recognized as
important natural anti-oxidants found in many kinds
of foods and plants. Resveratrol has strong anti-
oxidative,
5,6
anti-inflammatory,
7
anti-cancer,
8,9
melanogenesis inhibitory,
5,10
and collagen synthesis
activating effects
10
. Passion fruits (Passiflora edulis),
especially the seeds of passion fruits contain
Piceatannol in a natural state, which is nearly 50
times larger than that found in grapes.
10,11
Actually,
88% of the total polyphenols are contained in the
seeds.
10
In addition, the extract of passion fruit seeds
contains another polyphenol named Scirpusin B,
which is a dimmer of Piceatannol (Fig. 1).Previous
studies reported that Scirpusin B also shows the
strong vasorelaxant effect in the rat thoracic aorta
probably due to nitric oxide generation,
12
superoxide
anion scavenging activity
13
as well as DPPH radical
scavenging activity.
12
It is very interesting that
scirpsin B was demonstrated to be more potent in the
aortic vasorelaxing effect and in radical scavenging
effect than Piceatannol.
12
In addition, it has been
reported that Scirpusin B could improve glucose
metabolism to prevent the postprandial elevation of
blood glucose.
14
These properties strongly suggest
that Scirpusin B could have vasodilatating effect and
anti-arteriosclerotic effect, those are clinically
important especially in maintaining blood supply to
myocardium. Therefore, the aim of this paper is to
investigate the effect of Scirpusin B isolated from
passion fruit seeds on the coronary circulation of the
perfused rat heart.
MATERIALS AND METHODS
Animals and Ethics; Experiments were performed in
accordance with the Guide for Care and Use of
Laboratory Animals by US National Institute of
Health and in accordance with the Regulation OF
Animal Experiment in Yamagata University
15
under
the regulation of the Animal Care Committee of
Yamagata University School of Medicine
(Identification number 25072). In this study, 5 - 6
months old (340 - 370 g) male Fischer rats (27
animals in total) obtained from Charles River Japan
(Atsugi, Japan) were used. Each rat was used for the
experiment of the isolated heart during the ages
between 20 to 26 weeks-old to avoid ageing effects.
Female rats were not used because of the possible
effects of changes in sex hormone levels (i.e. estrogen
and progesterone). Rats were bred in the Laboratory
Animal Center, Yamagata University School of
Medicine. They were maintained on standard rat
chow with water ad libitum. Scirpusin B was
extracted from passion fruit seeds and purified by
high-performance liquid chromatography (HPLC).
12
In brief, the extracts of passion fruit seeds were
fractionated by reverse-phase HPLC. Each fraction
was collected by an Inertsil ODS-3 column (GL
Sciences Inc., Tokyo, Japan) with conditions of a (A)
water and/or (B) acetonitrile mobile phase at a flow
rate of 5 mL/min. A gradient elution of 0 80% (B)
at 0 90 min was used for fractionation. The
fractionated samples were analyzed by using ODS-3
column. The analytical HPLC was carried out with
(A) water and/or (B) acetonitrile mobile phase at a
flow rate of 0.75 mL/min. A gradient elution of 0
45% (B) at 0 25 min was used for this analysis.
Fig 1: Chemical structures of Piceatannol and
Scirpusin B
Measurement of the coronary flow in isolated
perfused hearts
Fischer-344 rats were deeply anesthetized with
diethyl ether and sacrificed; then the hearts were
quickly excised by performing thoracotomies and
placed in an ice-cold solution of modified Krebs-
Henseleit (118 mM NaCl, 4.7 mM KCl, 24.9 mM
NaHCO
3
, 1.18 mM MgSO
4
, 1.18 mM KH
2
PO
4
, 1.8
mM CaCl
2
, 5.0 mM glucose, 2.0 mM pyruvic acid,
0.057 mM ascorbic acid). The isolated hearts were
immediately perfused by Langendorffs method
under constant pressure (75 cm H
2
O) with modified
Krebs-Henseleit solution. The buffer solution was
548
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
difference between Resveratrol and Piceatannol is the
presence of an extra hydroxyl group in one of the
aromatic rings of Piceatannol. Piceatannol and
Resveratrol are phenolic compounds produced
naturally in grapes and red wine.
4
In general,
phenolic compounds have been recognized as
important natural anti-oxidants found in many kinds
of foods and plants. Resveratrol has strong anti-
oxidative,
5,6
anti-inflammatory,
7
anti-cancer,
8,9
melanogenesis inhibitory,
5,10
and collagen synthesis
activating effects
10
. Passion fruits (Passiflora edulis),
especially the seeds of passion fruits contain
Piceatannol in a natural state, which is nearly 50
times larger than that found in grapes.
10,11
Actually,
88% of the total polyphenols are contained in the
seeds.
10
In addition, the extract of passion fruit seeds
contains another polyphenol named Scirpusin B,
which is a dimmer of Piceatannol (Fig. 1).Previous
studies reported that Scirpusin B also shows the
strong vasorelaxant effect in the rat thoracic aorta
probably due to nitric oxide generation,
12
superoxide
anion scavenging activity
13
as well as DPPH radical
scavenging activity.
12
It is very interesting that
scirpsin B was demonstrated to be more potent in the
aortic vasorelaxing effect and in radical scavenging
effect than Piceatannol.
12
In addition, it has been
reported that Scirpusin B could improve glucose
metabolism to prevent the postprandial elevation of
blood glucose.
14
These properties strongly suggest
that Scirpusin B could have vasodilatating effect and
anti-arteriosclerotic effect, those are clinically
important especially in maintaining blood supply to
myocardium. Therefore, the aim of this paper is to
investigate the effect of Scirpusin B isolated from
passion fruit seeds on the coronary circulation of the
perfused rat heart.
MATERIALS AND METHODS
Animals and Ethics; Experiments were performed in
accordance with the Guide for Care and Use of
Laboratory Animals by US National Institute of
Health and in accordance with the Regulation OF
Animal Experiment in Yamagata University
15
under
the regulation of the Animal Care Committee of
Yamagata University School of Medicine
(Identification number 25072). In this study, 5 - 6
months old (340 - 370 g) male Fischer rats (27
animals in total) obtained from Charles River Japan
(Atsugi, Japan) were used. Each rat was used for the
experiment of the isolated heart during the ages
between 20 to 26 weeks-old to avoid ageing effects.
Female rats were not used because of the possible
effects of changes in sex hormone levels (i.e. estrogen
and progesterone). Rats were bred in the Laboratory
Animal Center, Yamagata University School of
Medicine. They were maintained on standard rat
chow with water ad libitum. Scirpusin B was
extracted from passion fruit seeds and purified by
high-performance liquid chromatography (HPLC).
12
In brief, the extracts of passion fruit seeds were
fractionated by reverse-phase HPLC. Each fraction
was collected by an Inertsil ODS-3 column (GL
Sciences Inc., Tokyo, Japan) with conditions of a (A)
water and/or (B) acetonitrile mobile phase at a flow
rate of 5 mL/min. A gradient elution of 0 80% (B)
at 0 90 min was used for fractionation. The
fractionated samples were analyzed by using ODS-3
column. The analytical HPLC was carried out with
(A) water and/or (B) acetonitrile mobile phase at a
flow rate of 0.75 mL/min. A gradient elution of 0
45% (B) at 0 25 min was used for this analysis.
Fig 1: Chemical structures of Piceatannol and
Scirpusin B
Measurement of the coronary flow in isolated
perfused hearts
Fischer-344 rats were deeply anesthetized with
diethyl ether and sacrificed; then the hearts were
quickly excised by performing thoracotomies and
placed in an ice-cold solution of modified Krebs-
Henseleit (118 mM NaCl, 4.7 mM KCl, 24.9 mM
NaHCO
3
, 1.18 mM MgSO
4
, 1.18 mM KH
2
PO
4
, 1.8
mM CaCl
2
, 5.0 mM glucose, 2.0 mM pyruvic acid,
0.057 mM ascorbic acid). The isolated hearts were
immediately perfused by Langendorffs method
under constant pressure (75 cm H
2
O) with modified
Krebs-Henseleit solution. The buffer solution was
549
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
continuously aerated with 95% O
2
- 5% CO
2
mixture
(pH 7.4), and its temperature was kept constant at 37
0.1C with a water-jacketed column.
Piceatannol (10, 30 and 100 M; n = 7) and the
Scirpusin B (10, 30 and 100 M; n = 10) were
injected into the coronary artery as a bolus for 10
seconds. The changes in the coronary flow were
continuously recorded and expressed as a percentage
of the basal flow just before the injection of the
Scirpusin B and Peceatannol. The coronary flow
(ml/min) was measured with an electromagnetic flow
meter (MFV 1100, Nihon Kohden, Tokyo, Japan).
The left ventricular pressure was recorded with a
pressure transducer (Statham P-50, Gould). The heart
rate was detected with a heart rate counter (AT-601G,
Nihon Kohden, Tokyo, Japan).
Effects of L-NAME and Diclofenac on the Scirpusin
B-induced coronary vasodilatation N
G
-nitro-L-
arginine methyl ester hydrochloride (L-NAME) or
Diclofenac were applied by continuous infusion via
the rubber tubing connected to the aortic cannula. The
concentrations of L-NAME (final concentration: 100
mM) and Diclofenac (final concentration: 10 mM)
were sufficient to inhibit the synthesis of nitric oxide
(NO) and prostaglandins, respectively.
16,17
In the
groups receiving pretreatment with either L-NAME
or Diclofenac, L-NAME or Diclofenac were
continuously infused through a micro syringe pump
(IC3200, KD Scientific Inc., Holliston, MS, USA) for
10 minutes prior to and during the application of the
Scirpusin B. The response to the Scirpusin B in the
presence and absence of L-NAME or Diclofenac was
recorded in each experiment by using different hearts.
Statistical analysis : All data were expressed as means
standard error of the mean (SEM). Differences
between two groups were compared for statistical
significance using unpaired Students t-test.
Differences between the three groups were compared
using ANOVA (analysis of variance), followed by
Tukeys post-hoc tests for multiple comparisons.
Differences were considered significant at p < 0.05.
Materials used: Chemicals used in these experiments
were 3,4,3,5-tetrahydroxy-trans-stilbene
(Piceatannol, Cayman Chemical Co., Ann Arbor, MI,
USA),N
G
-Nitro-L-arginine methyl ester
hydrochloride (L-NAME, Sigma-Aldrich Co., St.
Louis, MO, USA) and sodium Diclofenac (Sigma-
Aldrich). The Scirpusin B (purity > 96%) derived
from passion fruit (Passiflora edulis) seeds was
purified and provided by Morinaga & Co., Ltd.
(Kanagawa, Japan). Piceatannol and the Scirpusin B
were dissolved in DMSO. The vehicle used for
dissolving L-NAME and Diclofenac was
physiological saline. Each solution was prepared
freshly on the day of experiment.
RESULTS
Effects of Piceatannol and Scirpusin B on the
coronary flow; Fig. 2 shows the effect of the
Scirpusin B (100 M) and Piceatannol (100 M) on
the coronary flow in the perfused rat heart. Each drug
was infused into the rubber tubing connected to the
aortic cannula. The Scirpusin B increased coronary
flow up to 108.2% of the initial value within 15
seconds, then it gradually decreased and returned to
the basal level after 2 minutes. Scirpusin B did not
change heart rate and cardiac contractility. In
contrast, Piceatannol did not significantly increase the
coronary flow. The Scirpusin B-increased the
increase in coronary flow was concentration-
dependent (Fig. 3).
Effects of L-NAME and Diclofenac on the Scirpusin
B-induced coronary vasodilatation in an isolated
perfused heart.
For revealing the role of NO and vasorelaxing
prostanoids on coronary vasodilatation induced by
Scirpusin B, rat hearts were pretreated with their
inhibitors. In order to determine whether NO was
involved in the Scirpusin B-induced vasodilatation,
N
G
-Nitro-L-arginine methyl ester hydrochloride (L-
NAME) was used as an inhibitor of NO synthase. In
the rats pretreated with L-NAME (100 M), the
Scirpusin B-induced coronary vasodilatation was
significantly attenuated compared with administration
of the Scirpusin B alone (Fig. 4). In order to elucidate
whether vasorelaxing prostanoids (for example,
prostacyclin and PGE
2
) were involved in the
Scirpusin B-induced coronary vasodilatation,
Diclofenac was used as an inhibitor of
cyclooxygenase. In the rats pretreated with
Diclofenac (10 M), the Scirpusin B-induced
coronary vasodilatation was also significantly
attenuated (Fig. 4). Although coronary flow was
slightly diminished by treatment with L-NAME
alone, treatment with Diclofenac alone did not change
coronary flow at all.
550
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
Fig 2: Comparison of the effects of Scirpusin B and
Piceatannol on the coronary flow in perfused rat
hearts.
Each drug was infused into the aortic cannula.
Results are expressed as mean SEM. *P< 0.05 vs.
Piceatannol, **P< 0.01 vs. Piceatannol.
Fig 3: Effect of scirpusin B on the coronary flow of the
Langendorff-perfused rat hearts.
Scirpusin B was infused into the aortic cannula.
Scirpusin B increased coronary flow in a
concentration-dependent manner. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
basal value.
Fig4: Effects of nitric oxide synthase inhibitor L-
NAME and of cyclooxygenase inhibitor diclofenac on
the scirpusin B-induced coronary vasodilatation in
Langendorff-perfused rat hearts
Continuous infusion of each inhibitor through a
syringe pump began 10 minutes before and continued
during the application of the scirpusin B. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
scirpusin B alone.
DISCUSSION
It is widely known that moderate red wine
consumption is associated with reducing the risk of
coronary heart diseases (CHDs). Polyphenolic
antioxidants found in red wine, includingResveratrol
and piceatannol, are thought to be responsible for the
cardiovascular benefits associated with moderate red
wine consumption.
18
Oxidative stress can cause
endothelial dysfunction and is associated with the
development of cardiovascular diseases such as
hypertension and atherosclerosis.
19
In general,
polyphenol exhibit various biological activities such
as the decrease of LDL oxidation, inhibition of
platelet aggregation
20
and improvement of endothelial
function.
21
These biological activities indicate that
polyphenol possess cardioprotective properties.
The phytoalexin Resveratrol is produced naturally by
some spermatophytes in response to fungal attack or
injury. Resveratrol is commonly found in food and
drinks, including red wine, grapes, mulberries,
passion fruit
10
and peanuts.
22
Many studies have
revealed that Resveratrol has anti-inflammatory
properties, suppression of ICAM-1 gene expression
23
and health benefits to prevent CHD.
On the other hand, piceatannol is an analogue of
Resveratrol, which also has a wide variety of
bioactivities, such as anti-oxidative effects,
5,6
anti-
inflammatory effects,
7,24,25
inhibition of vascular
smooth muscle cell proliferation,
26
anti-arrhythmic
activity.
27
Taken together, both Resveratrol and
piceatannol could have preventative properties for
atherosclerosis, which in turn can help prevent CHD.
The genus Passiflora, comprising about 500 species,
is the largest in the family Passifloraceae.
Passifloraceae were introduced into medicine in
1840, and have been widely used as medical herbs in
many countries. For instance, Passifloraceae have
been used as hypnotic,
25
anxiolytic,
29,30
sedative, anti-
convulsant,
31
anti-tussive, analgesic, wormicidal, and
against inflammatory skin diseases. In addition, anti-
inflammatory effects
7
and the acceleration of the
healing of incisions
32
have been reported on the
550
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
Fig 2: Comparison of the effects of Scirpusin B and
Piceatannol on the coronary flow in perfused rat
hearts.
Each drug was infused into the aortic cannula.
Results are expressed as mean SEM. *P< 0.05 vs.
Piceatannol, **P< 0.01 vs. Piceatannol.
Fig 3: Effect of scirpusin B on the coronary flow of the
Langendorff-perfused rat hearts.
Scirpusin B was infused into the aortic cannula.
Scirpusin B increased coronary flow in a
concentration-dependent manner. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
basal value.
Fig4: Effects of nitric oxide synthase inhibitor L-
NAME and of cyclooxygenase inhibitor diclofenac on
the scirpusin B-induced coronary vasodilatation in
Langendorff-perfused rat hearts
Continuous infusion of each inhibitor through a
syringe pump began 10 minutes before and continued
during the application of the scirpusin B. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
scirpusin B alone.
DISCUSSION
It is widely known that moderate red wine
consumption is associated with reducing the risk of
coronary heart diseases (CHDs). Polyphenolic
antioxidants found in red wine, includingResveratrol
and piceatannol, are thought to be responsible for the
cardiovascular benefits associated with moderate red
wine consumption.
18
Oxidative stress can cause
endothelial dysfunction and is associated with the
development of cardiovascular diseases such as
hypertension and atherosclerosis.
19
In general,
polyphenol exhibit various biological activities such
as the decrease of LDL oxidation, inhibition of
platelet aggregation
20
and improvement of endothelial
function.
21
These biological activities indicate that
polyphenol possess cardioprotective properties.
The phytoalexin Resveratrol is produced naturally by
some spermatophytes in response to fungal attack or
injury. Resveratrol is commonly found in food and
drinks, including red wine, grapes, mulberries,
passion fruit
10
and peanuts.
22
Many studies have
revealed that Resveratrol has anti-inflammatory
properties, suppression of ICAM-1 gene expression
23
and health benefits to prevent CHD.
On the other hand, piceatannol is an analogue of
Resveratrol, which also has a wide variety of
bioactivities, such as anti-oxidative effects,
5,6
anti-
inflammatory effects,
7,24,25
inhibition of vascular
smooth muscle cell proliferation,
26
anti-arrhythmic
activity.
27
Taken together, both Resveratrol and
piceatannol could have preventative properties for
atherosclerosis, which in turn can help prevent CHD.
The genus Passiflora, comprising about 500 species,
is the largest in the family Passifloraceae.
Passifloraceae were introduced into medicine in
1840, and have been widely used as medical herbs in
many countries. For instance, Passifloraceae have
been used as hypnotic,
25
anxiolytic,
29,30
sedative, anti-
convulsant,
31
anti-tussive, analgesic, wormicidal, and
against inflammatory skin diseases. In addition, anti-
inflammatory effects
7
and the acceleration of the
healing of incisions
32
have been reported on the
550
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
Fig 2: Comparison of the effects of Scirpusin B and
Piceatannol on the coronary flow in perfused rat
hearts.
Each drug was infused into the aortic cannula.
Results are expressed as mean SEM. *P< 0.05 vs.
Piceatannol, **P< 0.01 vs. Piceatannol.
Fig 3: Effect of scirpusin B on the coronary flow of the
Langendorff-perfused rat hearts.
Scirpusin B was infused into the aortic cannula.
Scirpusin B increased coronary flow in a
concentration-dependent manner. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
basal value.
Fig4: Effects of nitric oxide synthase inhibitor L-
NAME and of cyclooxygenase inhibitor diclofenac on
the scirpusin B-induced coronary vasodilatation in
Langendorff-perfused rat hearts
Continuous infusion of each inhibitor through a
syringe pump began 10 minutes before and continued
during the application of the scirpusin B. Results are
expressed as mean SEM. *P< 0.05, **P< 0.01 vs.
scirpusin B alone.
DISCUSSION
It is widely known that moderate red wine
consumption is associated with reducing the risk of
coronary heart diseases (CHDs). Polyphenolic
antioxidants found in red wine, includingResveratrol
and piceatannol, are thought to be responsible for the
cardiovascular benefits associated with moderate red
wine consumption.
18
Oxidative stress can cause
endothelial dysfunction and is associated with the
development of cardiovascular diseases such as
hypertension and atherosclerosis.
19
In general,
polyphenol exhibit various biological activities such
as the decrease of LDL oxidation, inhibition of
platelet aggregation
20
and improvement of endothelial
function.
21
These biological activities indicate that
polyphenol possess cardioprotective properties.
The phytoalexin Resveratrol is produced naturally by
some spermatophytes in response to fungal attack or
injury. Resveratrol is commonly found in food and
drinks, including red wine, grapes, mulberries,
passion fruit
10
and peanuts.
22
Many studies have
revealed that Resveratrol has anti-inflammatory
properties, suppression of ICAM-1 gene expression
23
and health benefits to prevent CHD.
On the other hand, piceatannol is an analogue of
Resveratrol, which also has a wide variety of
bioactivities, such as anti-oxidative effects,
5,6
anti-
inflammatory effects,
7,24,25
inhibition of vascular
smooth muscle cell proliferation,
26
anti-arrhythmic
activity.
27
Taken together, both Resveratrol and
piceatannol could have preventative properties for
atherosclerosis, which in turn can help prevent CHD.
The genus Passiflora, comprising about 500 species,
is the largest in the family Passifloraceae.
Passifloraceae were introduced into medicine in
1840, and have been widely used as medical herbs in
many countries. For instance, Passifloraceae have
been used as hypnotic,
25
anxiolytic,
29,30
sedative, anti-
convulsant,
31
anti-tussive, analgesic, wormicidal, and
against inflammatory skin diseases. In addition, anti-
inflammatory effects
7
and the acceleration of the
healing of incisions
32
have been reported on the
551
Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
experimental basis. Passion fruit (Passiflora edulis) is
rich in piceatannol.
10, 11
Besides, in passion fruit seeds
contain a much larger amount of polyphenol
compared to its rind and pulp.
10
Thus, we took notice
of passion fruit seed extract. Passion fruit (Passiflora
edulis) seed polyphenol is mainly comprised of
piceatannol and another polyphenol, scirpusin B. In
the present study, we have evaluated the effects of
scirpusin B and piceatannol in the coronary
vasodilatation effect observed for passion fruit seed
extract.
The effects of scirpusin B and piceatannol on the
coronary flow of the perfused rat heart was compared
(Fig. 2). Although coronary flow was not
significantly increased by piceatannol, scirpusin B
(purity > 96%) increased coronary flow up to 108.2%
of the initial value within 15 seconds, then it
gradually decreased and returned to the basal level
after 2 minutes. These results would indicate that
scirpusin B itself has the vasodilating effect on
coronary arteries of the rat. In addition, scirpusin B
increased coronary flow in a concentration-dependent
manner (Fig. 3). Scirpusin B (10, 30 and 100 M)
increased coronary flow up to 101.4%, 102.8%,
108.2% on percentage of the initial value within 15
seconds, respectively.
In order to elucidate the coronary vasorelaxant
pathway, the perfused rat hearts were pretreated with
two different inhibitors. For the following reason, we
used L-NAME as an inhibitor of NO synthase and
diclofenac as an inhibitor of cyclooxygenase. The
endothelium is the monolayer of endothelial cells and
plays a critical role in regulating vascular tone and in
maintaining the cardiovascular function. Two of the
well-known factors involved in vasodilatation are
endothelium derived NO and vasorelaxing
prostanoids (i.e., prostacyclin and prostaglandin E
2
).
They may be released not only in the basal condition
but also in response to various vasodilatating
substances such as bradykinin, and to the
intravascular shear stress.
33,34
Atrial natriuretic
peptide (ANP) also plays an important role in
regulating coronary circulation in vivo.
35
In the rat
heart pretreated with L-NAME, the scirpusin B-
induced coronary vasodilatation was significantly
attenuated compared with that of scirpusin B alone
(Fig. 4). Also, in the rat heart pretreated with
diclofenac, the scirpusin B-induced coronary
vasodilatation was significantly attenuated (Fig. 4).
These results suggest that the vasodilating effect of
scirpusin B depends, at least in part, on the release of
NO and vasodilating prostanoids.
PGI
2
is the major prostaglandin released under basal
conditions of perfused heart. However, the basal
release of prostaglandins are known to be less than
100-300 pg/ml.
33,36
The functional significance of the
small amount of basal PGI
2
varies among species. For
example, coronary vascular tone was increased by
inhibition of PG synthesis in guinea-pig heart,
37
while
not affected in rat and rabbit heart
38,39
probably
depending on the level of basal PG release. In our
present study, inhibition of the basal release of PGs
by diclofenac could not significantly affect the basal
coronary flow probably because the level of basal
PGI
2
was low. On the other hand, the PGI
2
release
could be stimulated to be functionally sufficient
amounts by scirpusin B, so the CF was inhibited by
diclofenac.
Although scirpusin B significantly increased coronary
flow, it did not influence HR, LVP. These results
suggest that scirpusin B may have a protective effect
on an ischemic heart by increasing coronary flow
without affecting the cardiac function. Chronic
ischemia is caused by a mismatch of the oxygen
supply and demand, where significant fixed coronary
stenosis and/or excess myocardial oxygen demand
could result in ischemia. As to oxygen demand, the
coronary blood flow increases as the metabolic
activity of the heart. Although normal PaO
2
levels
range from 80 to 100 mmHg, the coronary sinus
blood has a PO
2
of about 20 mmHg. Therefore
oxygen extraction is very high in coronary
circulation. An increase in oxygen demand elicits an
increase in coronary blood flow as a result of
vasodilatation of the coronary vessels. Thus, we
speculated that scirpusin B might have a protective
effect on an ischemic heart.
In the present study, we found that scirpusin B has a
coronary vasodilating effect via production of both
NO and some prostanoids. It is known that these
vasodilator substances (NO and prostacyclin) have
anti-aggregatory effects on platelets. An intact
endothelium shows an anti-thrombotic, anti-
coagulatory and fibrinolytic properties.
40
In contrast,
endothelial dysfunction is associated with
cardiovascular events.
41
Therefore, scirpusin B would
exert the beneficial effect on coronary circulation of
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Matsumoto et al., Int J Med Res Health Sci. 2014;3(3):547-553
the intact endothelium and on preventing
arteriosclerosis.
CONCLUSION
This study shows that scirpusin B increases rat
coronary flow via production of NO and vasodilating
prostanoids. It is implicated that scirpusin B may
have beneficial effects on preventing cardiac events
and atherosclerosis by increasing these vasodilating
substances.
ACKNOWLEDGEMENTS
This study was supported partly by the Grants-in-Aid
for Scientific Research (C) No.24500846 (A.I.). We
wish to thank Erin MacNamara and Robert Jones for
correcting English editing of the manuscript.
Conflicts of interest: Declare no conflict of interest.
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