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Antioxidant activity of extract from Polygonum aviculare L

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Free radicals induce numerous diseases by lipid peroxidation, protein peroxidation, and DNA damage. It has been reported that numerous plant extracts have antioxidant activities to scavenge free radicals. Whether Polygonum aviculare L. (Polygonaceae) has antioxidant activity is unknown. In this study, dried Polygonum aviculare L. was extracted by ethanol, and the extract was lyophilized. The antioxidant activities of extract powder were examined by free radical scavenging assays, superoxide radical scavenging assays, lipid peroxidation assays and hydroxyl radical-induced DNA strand scission assays. The results show that the IC50 value of Polygonum aviculare L. extract is 50 microg/ml in free radical scavenging assays, 0.8 microg/ml in superoxide radical scavenging assays, and 15 microg/ml in lipid peroxidation assays, respectively. Furthermore, Polygonum aviculare L. extract has DNA protective effect in hydroxyl radical-induced DNA strand scission assays. The total phenolics and flavonoid content of extract is 677.4 +/- 62.7 microg/g and 112.7 +/- 13 microg/g. The results indicate that Polygonum aviculare L. extract clearly has antioxidant effects.
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281
HSU Biol Res 39, 2006, 281-288
Biol Res 39: 281-288, 2006
BR
Antioxidant activity of extract from Polygonum
aviculare L.
CHIN-YUAN HSU
Department of Life Science, Chang Gung University, Tao-Yuan, Taiwan
ABSTRACT
Free radicals induce numerous diseases by lipid peroxidation, protein peroxidation, and DNA damage. It has
been reported that numerous plant extracts have antioxidant activities to scavenge free radicals. Whether
Polygonum aviculare L. (Polygonaceae) has antioxidant activity is unknown. In this study, dried Polygonum
aviculare L. was extracted by ethanol, and the extract was lyophilized. The antioxidant activities of extract
powder were examined by free radical scavenging assays, superoxide radical scavenging assays, lipid
peroxidation assays and hydroxyl radical-induced DNA strand scission assays. The results show that the IC50
value of Polygonum aviculare L. extract is 50 µg/ml in free radical scavenging assays, 0.8 µg/ml in
superoxide radical scavenging assays, and 15 µg/ml in lipid peroxidation assays, respectively. Furthermore,
Polygonum aviculare L. extract has DNA protective effect in hydroxyl radical-induced DNA strand scission
assays. The total phenolics and flavonoid content of extract is 677.4 ± 62.7 mg/g and 112.7 ± 13 mg/g. The
results indicate that Polygonum aviculare L. extract clearly has antioxidant effects.
Key terms: antioxidant activity, free radical, phenolics, lipid peroxidation, DNA damage Polygonum
aviculare L.
Corresponding author: Dr. Chin-Yuan Hsu, Department of Life Science, Chang Gung University, 259, Wen-Hwa 1st Road,
Kwei-Shan, Tao-Yuan 333, Taiwan, Tel: (886-3) 211-8800, ext 3402, E-mail: hsu@mail.cgu.edu.tw
Received: July 19, 2005. In revised form: September 26, 2005. Accepted: October 7, 2005
INTRODUCTION
Reactive oxygen species produced by
ultraviolet light, ionizing radiation,
chemical reactions, and metabolic processes
have numerous pathological effects, such as
causing lipid peroxidation, protein
peroxidation, DNA damage, and cellular
degeneration related to cardiovascular
disease, ageing, cancer, inflammatory
diseases, and a variety of other disorders (2,
6, 9, 16, 20, 28). They include superoxide
radical anion (
O
2-
), hydroxyl radicals
(
OH), singlet oxygen (
1
O
2
), and hydrogen
peroxide (H
2
O
2
). In cellular oxidation
reactions, superoxide radical normally is
formed first, and its effects can be
magnified because it produces other kinds
of cell-damaging free radicals and oxidizing
agents. The damaging action of the
hydroxyl radical is the strongest among free
radicals (18).
Phenolics have been reported to have a
capacity to scavenge free radicals. They are
commonly found in both edible and non-
edible plants and have multiple biological
effects, including antioxidant activity (13,
27). The antioxidant activity of phenolics is
mainly due to their redox properties, which
allow them to act as reducing agents,
hydrogen donators, and singlet oxygen
quenchers. In addition, they have a metal
chelation potential (22). Phenolics, such as
flavonoids, phenolic acids, stilbenes,
lignans, lignin, and tannins, are especially
common in leaves, flowering tissues, and
woody parts, such as stems and barks (17).
They have been suggested to play a
preventive role in the development of
cancer, heart disease, and ageing-related
diseases.
The importance of the antioxidant
constituents of plant materials in the
maintenance of health and protection from
ageing-related diseases has intrigued
scientist for a long time. I have screened the
HSU Biol Res 39, 2006, 281-288
282
antioxidant activity of a variety of wild
plants and other substances used in
traditional Oriental medicine by 1,1-
Diphenyl-2-picrylhydrazyl (DPPH) free
radical scavenging assays. The results show
that Polygonum aviculare L. (Polygonaceae)
exhibits a DPPH free radical scavenging
effect. Polygonum aviculare L. is used in
traditional Oriental medicine and belongs to
the “li-shui-shen-shih” category of drugs. It
is used traditionally to treat ailments caused
by high humidity, because of its diuretic
property. In the present study, I wish to
study the antioxidant effects of Polygonum
aviculare L. on superoxide radical
scavenging, lipid peroxidation, and DNA
damage.
METHODS
Chemicals
1,1-Diphenyl-2-picrylhydrazyl (DPPH),
nitroblue tetrazolium chloride (NBT), 2-
thiobarbituric acid (TBA), ΦX174 RF1
supercoiled DNA, Folin-Ciocalteu reagent,
L-ascorbic acid, and (+)-catechin were
purchased from Sigma Chemical Co. The
other chemicals and solvents used in this
experiment were of the highest quality
available. Dried Polygonum avicular was
purchased from a local drugstore.
Preparation of plant extract
Dried Polygonum aviculare L. was made
into powder form. 5 g of dried powder were
extracted in 50 ml 50% ethanol solution at
25
o
C for 30 min with shaking. The extract
was centrifuged at 15000 rpm for 3 min,
and the supernatant was collected. The
supernatant was concentrated in a rotary
evaporator and then lyophilized. The
resulting powder extract was used in this
study (4).
DPPH assay
1 mg extract powder was dissolved in 1 ml
of 50% ethanol solution to obtain 1000 µg/
ml sample solution. 1000 µg/ml solutions
were series diluted into 1 µg/ml, 5 µg/ml, 10
µg/ml, 20 µg/ml, 50 µg/ml, 100 µg/ml, 200
µg/ml, 500 µg/ml, and 1000 µg/ml with 50%
ethanol. In each reaction, the solutions were
mixed with 1 ml of 0.1 mM 1,1-Diphenyl-2-
picrylhydrazyl (DPPH), 0.45 ml of 50 mM
Tris-HCl buffer (pH 7.4), and 0.05 ml
samples at room temperature for 30 min.
50% ethanol solution was used as control.
The reduction of the DPPH free radical was
measured by reading the absorbance at 517
nm. DPPH, a purple-colored, stable free
radical is reduced to the yellow-colored
diphenylpicrylhydrazine when antioxidants
are added. L-ascorbic acid and (+)-catechin
were used as positive controls. The
inhibition ratio (percent) was calculated
from the following equation: % inhibition =
[(absorbance of control – absorbance of test
sample)/absorbance of control] x 100%. The
antioxidant activity of each sample was
expressed in terms of IC50 (micromolar
concentration required to inhibit DPPH
radical formation by 50%), calculated from
the inhibition curve (5, 8, 30).
NBT (superoxide scavenging) assay
The superoxide anion radical scavenging
activity was performed by using the
methods of Liu and Ng (18). Superoxide
radicals were generated in 3.0 ml of Tris-
HCl buffer (16 mM, pH 8.0), which
contained 78 µM β-nicotinamide adenine
dinucleotide (reduced form, NADH), 50
µM nitroblue tetrazolium (NBT), 10 µM
phenazin methosulfate (PMS), and test
samples in 50% ethanol solution (final
concentrations were 1, 5, 10, 20, 50, and
100 µg/ml, respectively). The color reaction
of superoxide radicals and NBT was
detected at OD 560 nm. (+)-catechin was
used as a positive control. The inhibition
ratio (%) was calculated from the following
equation: % inhibition = [(absorbance of
control – absorbance of test sample)/
absorbance of control] x 100%.
Lipid peroxidation assay
The brain of young adult male Balb/c mice
were dissected and homogenized with a
homogenizer in ice-cold Tris-HCl buffer (20
mM, pH 7.4) to produce a 1/10 homogenate.
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HSU Biol Res 39, 2006, 281-288
The homogenate was centrifuged at 12000g
for 15 min at 4
o
C, and the supernatant was
used for in vitro lipid peroxidation assay. A
1 ml aliquot of liposome was incubated with
the test samples (final concentrations were 1,
5, 10, 20, 50, and 100 µg/ml, respectively) in
the presence of 10 mM FeSO
4
and 0.1 mM
ascorbic acid at 37
o
C for 1 h. The reaction
was terminated by the addition of 1.0 ml of
trichloroacetic acid (TCA; 28%, w/v) and
1.5 ml of TBA (1%, w/v), followed by
heating at 100
o
C for 15 min. The absorbance
of the malondialdehyde (MDA)-TBA
complex was measured at 532 nm. (+)-
catechin was used as a positive control. The
inhibition ratio (%) was calculated from the
following equation: % inhibition =
[(absorbance of control – absorbance of test
sample)/absorbance of control] x 100% (4).
DNA strand scission assay
The assay was performed according to the
method of Keum et al., with minor
modifications (15). The reaction mixture
(30 µl) contained 10 mM Tris-HCl, 1 mM
EDTA buffer (pH 8.0), ΦX174 RF1
supercoiled DNA (0.6 µg), and H
2
O
2
(0.04M). Various amounts of the test
extract samples dissolved in 10 µl of
ethanol (final concentrations of the plant
extract in each assay were 1, 10, 100, 500,
and 1000 µg/ml, respectively) were added
prior to H
2
O
2
addition. Hydroxyl radicals
were generated by irradiation of the
reaction mixtures at a distance of 5 cm with
a 12 W UV lamp. After incubation at room
temperature for 20 min, the reaction was
terminated by the addition of a loading
buffer (0.25% bromophenol blue tracking
dye and 40% sucrose), and the mixtures
were then analyzed by 0.8% submarine
agarose gel electrophoresis (50eV, 1.5 h).
The gel was stained with ethidium bromide,
destained in water, and photographed on a
transilluminator (4).
Determination of total flavonoid
1 mg samples were added in 1ml of 80%
ethanol. A aliquot of 0.5 ml was added to
test tubes containing 0.1 ml of 10%
aluminum nitrate, 0.1 ml of 1 M potassium
acetate, and 4.3 ml of 80% ethanol. The
absorbance of the supernatant was measured
at 415 nm after 40 min at room temperature.
Total flavonoid concentration was calculated
using quercetin as standard (19).
Determination of Total Phenolics
Total phenolics content was determined
according to the Folin-Ciocalteu method
(23), using gallic acid as a standards 1 mg
extract powders were dissolved in 1 ml 50%
methanol solution. 0.5 ml extract solution
was mixed with 0.5 ml of 50% Folin-
Ciocalteu reagent. The mixture was let sit
for 2-5 min before the addition of 1.0 ml of
20% Na
2
CO
3
. The mixture was centrifuged
at 150 g for 8 min after 10 min of incubation
at room temperature. The absorbance of the
supernatant was measured at 730 nm. The
total phenolic content was expressed as
gallic acid equivalents (GAE) in milligrams
per gram sample (4).
RESULTS
The free radical scavenging activity of
Polygonum aviculare L. extract was
assessed by 1,1-Diphenyl-2-picrylhydrazyl
(DPPH) assay. (+)-catechin and L-ascorbic
acid were used as controls. Both are well-
known antioxidant compounds. The result
is shown in Figure 1. The IC
50
values (the
concentration required to inhibit radical
formation by 50%) of Polygonum aviculare
L. extract are 50 µg/ml. The IC
50
values of
(+)catechin and ascorbic acid are 35 µg/ml
and 50 µg/ml, respectively; they exhibit a
similar curve of antioxidant activity
compared to (+)-catechin and L-ascorbic
acid. The IC
50
value of Polygonum
aviculare L. extract is lower than that of
(+)-catechin and is similar to that of L-
ascorbic acid. This result demonstrates that
Polygonum aviculare L. extract has an
inhibitory effect on the DPPH radical.
The superoxide scavenging activity of
Polygonum aviculare L. extract was
evaluated by NBT (Superoxide Scavenging)
assay. (+)-catechin served as a control. The
result is shown in Figure 2. The IC
50
value
of Polygonum aviculare L. extract is 0.8 µg/
HSU Biol Res 39, 2006, 281-288
284
ml. The IC
50
value of (+)-catechin is 40 µg/
ml. Almost all superoxide radicals were
inhibited by 10 µg/ml Polygonum aviculare
L. extract. The superoxide scavenging
activity of Polygonum aviculare L. extract is
apparently higher than that of (+)-catechin.
Therefore, Polygonum aviculare L. seems to
be a potential source of superoxide radicals
scavenging. This result shows that
Polygonum aviculare L. extract has a
significant superoxide scavenging activity.
The lipid peroxidation suppressing
activity of Polygonum aviculare L. extract
was estimated by TBA assay. (+)-catechin
was employed as control. The result is
shown in Figure 3. The IC
50
values of
Polygonum aviculare L. extract is 16 µg/
ml. The IC
50
value of (+)-catechin is 17 µg/
ml. Compared to (+)-catechin, they exhibit
the similar curve of antioxidant activity.
The lipid peroxidation suppressing activity
of Polygonum aviculare L. extract is
identical to that of (+)-catechin. This result
indicates that Polygonum aviculare L.
extract has suppressing activity on lipid
peroxidation.
Figure 1: Free-radical scavenging activity of
Polygonum aviculare L. extract are measured
by using the DPPH assay: () (+)catechin; (O)
ascorbic acid; (L) Polygonum aviculare L.
extract. Results are mean ± SD (N=5).
Figure 2: Superoxide scavenging activity of
Polygonum aviculare L. extract are measured by
using the NBT assay: () (+)catechin; (L)
Polygonum aviculare L. extract. Results are
mean ± SD (N =5).
Figure 3: Effects of Polygonum aviculare L.
extract on both ferric ion and ascorbic acid
induced lipid peroxidation on mouse brain
homogenates: () (+)catechin; (L )
Polygonum aviculare L. extract. Results are
mean ± SD (N =5).
ΦX174 RF1 DNA strand scission
induced by UV photolysis of H
2
O
2
elevated the protective effect of DNA of
Polygonum aviculare L. extract. The result
is shown in Figure 4. ΦX174 RF1
supercoiled DNA was utilized as control
(lane 1). UV illumination alone did not
cause DNA strand cleavage (lane 2). The
treatment of supercoiled DNA with UV
plus H
2
O
2
led to the conversion of the
DNA to open circular form (lane 3). The
treatment of supercoiled DNA with UV,
H
2
O
2
plus the different concentration of
285
HSU Biol Res 39, 2006, 281-288
Polygonum aviculare L. extract led to the
maintenance of the DNA in the
supercoiled form (lanes 4-8). Almost
complete protection was expressed at a
dose of 1000 µg/ml. This protective effect
of DNA exhibits dose-dependency. This
result shows that Polygonum aviculare L.
extract has DNA protective activity under
oxidative stress.
TABLE 1
Total phenolic content and total flavonoid
content of ethanolic extract from
Polygonum aviculare
Extract Total phenolic Total flavonoid
(mg of GAE/g) (mg/g of samples)
Polygonum
aviculare 677.4 ± 62.7 (N=8) 112.7 ± 13 (N=10)
Total phenolics are expresed as gallic acid equivalent
(GAE).
Total flavonoid are expresed as mg of total flavonoid
content / g of samples based on quercetin as standard.
Values represent mean ± S.D.
DISCUSSION
Antioxidant activity of Polygonum
aviculare L. extract has been found by
means of free radical scavenging assays,
superoxide radical scavenging assays, lipid
peroxidation assays, and hydroxyl radical-
induced DNA strand scission assays. In
addition, Polygonum aviculare L. extract
has high phenolics and flavonoid contents.
This study indicates that Polygonum
aviculare L. extract obviously has
antioxidant effects.
DPPH is a stable radical that has been
used widely to evaluate the antioxidant
activity of various natural products (12). In
this study, DPPH scavenging activity has
been found in Polygonum aviculare L.
extract. The maximum inhibition of
Polygonum aviculare, (+)catechin and
ascorbic acid is about 80% in this study.
The maximum inhibition concentration of
Polygonum aviculare L. and (+)-catechin is
approximately 100 µg/ml. The maximum
inhibition concentration of ascorbic acid is
approximately 200 µg/ml. The inhibitory
curve of DPPH scavenging activity of
Polygonum aviculare L. is similar to that of
Acacia confusa (4), Cat’s claw (Uncaria
tomentosa) (24), and Anthriscus cerefolium
(5). However, the IC
50
value of Polygonum
aviculare L. (50 µg/ml) is less than that of
Acacia confusa (5 µg/ml), Cat’s claw
(Uncaria tomentosa) (18 µg/ml), and
Anthriscus cerefolium (45 µg/ml) (4, 24, 5).
Nevertheless, Polygonum aviculare L.
Figure 4: Protection effect of Polygonum
aviculare L. extract on DNA strand scission
induced by H
2
O
2
and UV. ΦX174 RF1
supercoiled DNA as control (lane 1), ΦX174
RF1 supercoiled DNA was exposed to UV
alone (lane 2), UV plus H
2
O
2
(lane 3), or plus
H
2
O
2
in the presence of final concentration of
1000 µg/ml (lane 4), 500 µg/ml (lane 5), 100
µg/ml (lane 6), 10 µg/ml (lane 7), 1 µg/ml
(lane 8) of Polygonum aviculare L. extract.
Lane 1 represents native ΦX174 RF1
supercoiled DNA without any treatment. OC:
Open circular; SC: Super coiled.
Plant phenolics are widely distributed in
plants. They are highly effective free
radical scavengers and exhibit strong
antioxidant activity. The content of total
phenolics in the Polygonum aviculare L.
was determined spectrometrically according
to the Folin-Ciocalteu procedure and
calculated as gallic acid equivalent
contents. The result is shown in Table 1.
The total phenolic content of Polygonum
aviculare L. extract is 677.4 ± 62.7 mg/g.
The content of total flavonoid in the
Polygonum aviculare L. also was
determined spectrometrically and calculated
as quercetin equivalents content. The result
is shown in Table 1. The total flavonoid
content of Polygonum aviculare L. extract
is 112.7 ± 13 mg/g. These results imply that
Polygonum aviculare L. extract contains a
high quantity of phenolics and flavonoids.
HSU Biol Res 39, 2006, 281-288
286
extract is a potential source of natural
antioxidants.
In cellular oxidation reactions,
superoxide radicals normally are formed
first, and their effects can be magnified
because they produce other kinds of free
radicals and oxidizing agents (18).
Additionally, xanthine oxidase is one of the
main enzymatic sources of those reactive
oxygen species in vivo. In this study,
superoxide radicals scavenging property has
been found in Polygonum aviculare L.
extract. The IC
50
value of Polygonum
aviculare L. extract is 0.8 µg/ml, whereas
the IC
50
value of (+)-catechin is 40 µg/ml.
The maximum inhibitory effect of
Polygonum aviculare L. is about 100%. The
maximum inhibition concentration of
Polygonum aviculare L. is approximately 10
µg/ml. However, the maximum inhibition
concentration of (+)-catechin is higher than
100 µg/ml. It is noteworthy that the
superoxide scavenging activity of
Polygonum aviculare L. extract is superior
to that of (+)-catechin. Moreover, the IC
50
value of Polygonum aviculare L. (0.8 µg/ml)
is larger than that of Paeonia suffruticosa
(50 µg/ml) (18). In other words, Polygonum
aviculare L. has better superoxide radicals
scavenging activity than Paeonia
suffruticosa. These results show that
Polygonum aviculare L. is an important
source for superoxide radical scavenging.
In the current study, lipid peroxidation
of mouse brain homogenates was induced
by ferric ion plus ascorbic acid. Lipid
peroxidation scavenging activity has been
found in Polygonum aviculare L. extract.
The IC
50
value of Polygonum aviculare L.
extract is about 16 µg/ml. The IC
50
value of
(+)-catechin is about 17 µg/ml. The
maximum inhibitory effect of Polygonum
aviculare L. is about 75%. The maximum
inhibition concentration of Polygonum
aviculare L. is approximately 20 µg/ml.
The inhibitory effect of Polygonum
aviculare L. is higher than that of
(+)catechin when the concentration is
higher than IC
50
values. In other words,
Polygonum aviculare L. extract has better
scavenging effect than (+)-catechin when
the concentration is higher than 17 µg/ml.
This result indicates that Polygonum
aviculare L. extract is a good source of
lipid peroxidation scavenging.
The cellular damage resulting from
hydroxyl radicals is strongest among free
radicals. Hydroxyl radicals can be
generated by biochemical reactions.
Superoxide radical is converted by
superoxide dismutase to hydrogen peroxide,
which subsequently can produce extremely
reactive hydroxyl radicals in the presence
of transition metal ions, such as iron and
copper or by UV photolysis. Hydroxyl
radicals can attack DNA to cause strand
scission. That is, incubation of ΦX174 RF1
supercoiled DNA with H
2
O
2
and then UV
radiation resulted in complete conversion of
supercoiled DNA to the open circular form.
In this study, the administration of
Polygonum aviculare L. extract to the
reaction mixture substantially decreased the
DNA strand scission induced by both H
2
O
2
and UV radiation. It shows a dose-
dependent protection of DNA under
oxidative stress. The higher the
concentration of Polygonum aviculare L.
extracts, the better the DNA protection.
There is almost complete protection at a
dose of 1000 µg/ml. The effect of DNA
protection of Polygonum aviculare L. is
similar to that of Acacia confusa (4). These
results reveal that Polygonum aviculare L.
extract is an excellent DNA protector.
Phenolics are found in large quantities in
the plant kingdom, and they have been
shown to have multiple biological functions,
including antioxidant activity (21, 25, 14). In
this study, we examined the content of
phenolics from the extract of Polygonum
aviculare. The result showed that
Polygonum aviculare L. extract contains
677.4 ± 62.7 mg/g phenolics. It indicated
that the Polygonum aviculare extract
contained a higher amount of phenolics than
the bark and heartwood extracts of Acacia
confusa based on Folin-Ciocalteu procedures
(4). Therefore, Polygonum aviculare L. is a
significant source of phenolics. The results
in this study suggest that the effectiveness of
the antioxidant activity of Polygonum
aviculare extract is probably related to the
high contents of phenolics, and the observed
antioxidant activities of the extract may be
due to the hydroxyl groups in phenolics (10).
287
HSU Biol Res 39, 2006, 281-288
A similar finding has been demonstrated in
the plant extracts of Eucommia ulmoides
(Du-zhong) and Acacia confusa in which
enriched phenolics correlated well with their
antioxidant activities (4, 29).
It also has been reported that Polygonum
aviculare L. can be employed supportively
in the therapy of gingivitis by oral rinse (7).
It was suggested that this phenomenon was
attributed to the flavonoid components that
decrease capillary fragility and exert a
cortisone-like effect on gingival tissues (7).
In this study, we examined the content of
flavonoids from the extract of Polygonum
aviculare. The result showed that
Polygonum aviculare L. extract contains
high flavonoids, 112.7 ± 13 mg/g. It has
been suggested that the therapeutic effect of
Polygonum aviculare L. on gingivitis is
derived from its high flavonoid.
Additionally, the flavonoid content of
Polygonum aviculare L. also is higher than
that of propolis (29).
It is well known that free radicals are the
principal cause of several diseases, including
Parkinson’s disease, coronary heart disease,
cancer, and Alzheimer’s disease (3, 1, 11,
26). This study demonstrated that
Polygonum aviculare L. has high phenolics
contents and excellent antioxidant activity. It
would be interesting to investigate further
the potential effectiveness of Polygonum
aviculare L. for treating diseases caused by
the overproduction of free radicals. Also, the
antimicrobial effect, bioavailability and
potential toxicity of Polygonum aviculare L
need to be studied in vivo.
ACKNOWLEDGEMENTS
This work was supported by NSC 94-2311-
B-182-008 grants from the National
Science Council, R. O. C.
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... More specifically, the inhibition concentration (IC 50 ) values of plants during the sampling times are showed. A low value of IC 50 indicates a strong antioxidant activity, meaning that low concentrations of antioxidant substances in the extract can inhibit 50% of the radical reaction [32]. scientific evidence. ...
... More specifically, the inhibition concentration (IC50) values of plants during the sampling times are showed. A low value of IC50 indicates a strong antioxidant activity, meaning that low concentrations of antioxidant substances in the extract can inhibit 50% of the radical reaction [32]. Our findings (Figure 1) proved that, in general, the highest antioxidant activity was shown by plant samples collected during the third sampling period (November/December) whereas the lowest values were detected in plants collected during the first sampling in June. ...
... F. circummediterranea and D. glomerata were reported twice because they were collected from two different areas (Guardata and Guado Cannavina, respectively). A low value of IC50 indicates a strong antioxidant activity since low concentrations of extract are able to inhibit 50% of the radical reaction[32].Our findings(Figure 1) proved that, in general, the highest antioxidant activity was shown by plant samples collected during the third sampling period (November/December) whereas the lowest values were detected in plants collected during the first sampling ...
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Polyphenols are secondary metabolites of interest due to their potential application in various fields. This study is supposed to analyse the content of total polyphenols, total tannins, condensed tannins and antioxidant activity of ten wild plant species of nutritive interest to better understand their potential applications. Furthermore, the effect of heavy metals on the production of the investigated secondary metabolites was analysed. The different phenolic compounds were determined in methanol extracts obtained from edible plants collected during three sampling periods (June, September and November–December) in four areas of the Central Italy. Analyses were carried out by applying standard methodologies. In particular, total polyphenols were determined by the Folin–Ciocalteu method, total tannins by the polyvinylpolypyrrolidone (PVPP) reagent and condensed tannins by the 4-(dimethylamino)cinnamaldehyde (DMCA). Antioxidant activity was determined by assessing the scavenging capacity of 2,2-diphenyl-1-picrylhydrazyl (DPPH). The results showed a satisfactory content of phenolic compounds and antioxidant activity for all species analysed. Furthermore, the existence of a negative correlation between the presence of heavy metals and phenolic compounds was found. Results proved the potential use of these plants for balanced feeding of ruminants.
... This is in agreement with the yield of the extraction where the ethanolic extract of P. istanbulicum (14.5%) produced the highest yield in comparison with methanolic (11.6%) and chloroform (10.6%) extracts ( Table 2). The total flavonoid content of the other Polygonum species have previously been reported, for example the methanolic leaf extract of P. minus (53.19 ± 0.71 mg QE/g extract) (Abdullah et al., 2017), the ethanolic (70%) leaf extract of P. aviculare (19.78 ± 0.67 mg RE/g DW) (Cai et al., 2020), and the ethanolic (50%) extract of P. aviculare (112.7 ± 13 mg QE/g sample) (Hsu, 2006). With regard to the ethanolic extract of P. aviculare, the total phenolic and flavonoid contents obtained in the present work were lower as compared to the values reported by Hsu (2006), although they were higher than the values reported by Cai et al. (2020). ...
... The total flavonoid content of the other Polygonum species have previously been reported, for example the methanolic leaf extract of P. minus (53.19 ± 0.71 mg QE/g extract) (Abdullah et al., 2017), the ethanolic (70%) leaf extract of P. aviculare (19.78 ± 0.67 mg RE/g DW) (Cai et al., 2020), and the ethanolic (50%) extract of P. aviculare (112.7 ± 13 mg QE/g sample) (Hsu, 2006). With regard to the ethanolic extract of P. aviculare, the total phenolic and flavonoid contents obtained in the present work were lower as compared to the values reported by Hsu (2006), although they were higher than the values reported by Cai et al. (2020). This variation could be due to the application of different extraction methods and assay procedures or variations in the plants' origin. ...
... This variation could be due to the application of different extraction methods and assay procedures or variations in the plants' origin. In the study of Hsu (2006), dried P. aviculare was purchased from a local drugstore in Taiwan; while in the study of Cai et al. (2020), P. aviculare was collected from Shandong, China. Nevertheless, the results of the present work suggest that the tested Polygonum species, especially P. istanbulicum, could be a good source of phenolics and flavonoids. ...
Article
Polygonum species are used in traditional medicine in many countries; some are also consumed as vegetables in Turkey. The ethanolic, methanolic, and chloroform extracts of four Polygonum species growing in Istanbul, namely P. aviculare, P. patulum subsp. pulchellum, P. lapathifolium, and the only endemic species P. istanbulicum were evaluated for their antioxidant, anti-acetylcholinesterase (AChE), and anticancer potentials. The total phenolic and flavonoid contents of the extracts were determined by Folin-Ciocalteu and aluminium chloride methods, respectively. The antioxidant capacities of the extracts were determined using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and ferric-reducing antioxidant power (FRAP) assays. The AChE inhibitory activities of the extracts were determined using the Ellman method. Each extract was screened for cytotoxic activity against NRK-52E and HeLa cervical cancer cell lines using the MTT assay. Among the extracts screened, ethanolic extract of P. istanbulicum showed the highest total phenolic (207.03 ± 14.12 mg GAE/g extract) and total flavonoid (124.95 ± 7.84 mg CE/g extract) contents, and antioxidant activity (DPPH EC50, 8.09 ± 0.50 mg/mL). The chloroform extract of P. lapathifolium exhibited the lowest total phenolic (22.33 ± 3.05 mg GAE/g extract) and total flavonoid (11.66 ± 0.36 mg CE/g extract) contents, and antioxidant activity (DPPH EC50, 218.44 ± 24.46 mg/mL). The extracts exhibited AChE inhibitory activity in a dose-dependent manner, particularly the ethanolic extract of P. istanbulicum which displayed strongest inhibition against AChE (88.2 ± 3.44%). AChE inhibition was minimal (32.19 ± 2.09 to 48.34 ± 3.41%) in the chloroform extracts. All ethanolic extracts revealed cytotoxic activity toward HeLa cells, while they were not cytotoxic toward NRK-52E cells. The ethanolic extract of P. lapathifolium showed the most potent cytotoxicity against HeLa cells (IC50, 8.70 ± 1.35 µg/mL). Results suggested that ethanol was the best solvent for extracting the phenolic, antioxidant, and anti-AChE compounds, and P. istanbulicum may be a potential source of these compounds. Further investigations are nevertheless required to identify the bioactive compounds present in Polygonum species.
... In addition, the antioxidant activity of the sample can be expressed in various ways. The first is to express the ability of a food substance to remove ROS from a sample as a percentage of ROS present after the reactive oxygen species scavenging reaction (Akinmoladun et al., 2007;Hsu, 2006). The antioxidant activity expressed as a percentage represents only the free radical removal rate at the specific concentration used for the analysis. ...
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Abstract The quantitative antioxidant activity of foods is expressed as the equivalent weight of the authentic standard compounds with antioxidant function. Even in the same food sample, antioxidant activity is defined differently depending on the standard antioxidant compounds, making it difficult to compare them. Antioxidant activity was measured by DPPH radical and ABTS radical scavenging assay using Trolox, ascorbic acid, catechin, and gallic acid as standard antioxidant compounds. The relationship between the antioxidant activity and the standard was used to convert the antioxidant activity expressed as the equivalent of a specific antioxidant standard to the equivalent of another antioxidant standard. To verify the converted antioxidant activity, the antioxidant activity of apples was analyzed with several antioxidant standards and compared with the antioxidant activity converted with other standard compounds. As a result of comparing the antioxidant activity measured using various antioxidant standards and the converted antioxidant activity of apples, it was confirmed that there was no difference. The results of this study show that antioxidant activity expressed as a specific antioxidant standard compound can be accurately converted into the equivalent of another standard antioxidant compound.
... The TPC of crude CNSL was determined by Folin-Ciocalteau's method, as described by [42], with modifications. Briefly, a set of working standards of concentrations ranging from (0 to 5 mg/L) were prepared from a stock solution of standard gallic acid (10 mg/L). ...
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Cashew nutshell liquid (CNSL) is a cheap source of natural phenolic compounds that have numerous applications. These phenolic compounds have chemical structures with chromophores similar to those found in synthetic chemical UV-filters, which are present in commercial sunscreen products (SSPs). Thus, this study investigated the impact of solvents on the yield, total phenol content (TPC), total flavonoid content (TFC), and the sun protection factor (SPF) of crude CNSL. The percent yield ranged from (30.4 ± 0.7% to 49.3 ± 3.2%); hexane recorded the lowest yield, while ethanol recorded the highest. Acetone (101.2 ± 2.5 mg GA/g), methanol (99.5 ± 0.10), and chloroform (95.4 ± 3.7 mg GAE/g), recorded the highest TPC respectively, while hexane (33.3 ± 0.7 mg QE/g) recorded the highest TFC. The SPFs ranged from (22.1 ± 1.1 to 16.4 ± 0.8), chloroform (22.1 ± 1.1), acetone (21.5 ± 1.1), and methanol (19.3 ± 1.0) again recorded the highest values respectively, while hexane (16.4 ± 0.8) recorded the lowest. Our results revealed that extracting solvents has a significant impact on the yield and SPF of CNSL. Therefore, we propose that acetone, chloroform, and methanol, either alone or as mixtures, could be the best solvents for extracting CNSL with a good TPC and SPF.
... In addition, the study of the phenolic content corroborated these results obtained (18.00 ± 0.25 mg GAE/g) [72]. Other studies showed that the ethanolic extract in the determination of the free radical DPPH and scavenging of superoxide (NBT) has an antioxidant activity similar to that of ascorbic acid (IC 50 = 50 µg/ml) and inhibition of superoxide radicals (IC 50 = 0.8 µl/ml) [73]. On the other hand, used a model of the evaluation of ROS production by human neutrophils, the flavonol glucuronides present in the P. aviculare plant indicated that all compounds isolated at concentrations of 1 and 10 µM inhibited significantly the production of reactive oxygen species by neutrophils stimulated by f-MLP [74]. ...
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Polygonum aviculare is a plant widely distributed in all continents and very common in temperate regions of the world. The different parts of this plant are used as an anthelmintic, antidiabetic and to treat kidney disorders and skin problems. In order to highlight the importance of P. aviculare L., This review is undertaken with the aim to highlight some aspects of this plant, specifically the taxonomy, ethnomedicinal, morphological, phytochemical and biological studies of the plant. The taxonomic analysis indicated that the plant is descended from an inbred line and the cytogenetic results showed that P. aviculare L. is dominated by the number of chromosomes x = 10, therefore it was found that this herb has a rich phytochemical composition and many biological activities have been discussed in this review, mainly antioxidant, anti-inflammatory, anti-diabetic, anti-cancer, and dermato-protective activities. Graphical abstract
... Due to their redox characteristics, which enable them to function as reducing agents, hydrogen donors, and singlet oxygen quenchers, phenolics have a significant antioxidant activity. (Chin-Yuan Hsu, 2006). Recent years have seen a rise in interest in the hunt for natural antioxidants since research has revealed that oxidative stress and the formation of reactive oxygen species are important contributors to a variety of illnesses. ...
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Screening of antioxidant potential in different extracts of Adenocalymma alliaceum leaves were analysed by free radical scavenging activity. Aqueous, hexane and methanolic extracts of Adenocalymma alliaceum leaves (AAAE, AAHE and AAME,) were determined using various in vitro radical scavenging activities viz., DPPH, superoxide anion, nitricoxide, hydroxyl, iron chelating and reducing power assays at different concentrations. Phytochemical constituents, total polyphenolic and total flavonoid content of each of the extracts were determined. AAME showed high free radical scavenging activity as evidenced by the low IC50 values in DPPH(142.05µg/mL), superoxide (133.14µg/mL), nitricoxide(161.05µg/mL), hydroxyl (145.47 µg/mL) and in ferrous ion chelating (152.94) assays than AAAE and AAHE. In addition, the high reducing ability was reported in AAME followed by AAAE and AAHE. Furthermore, the total polyphenolic and total flavonoid content of the extracts were found to be high in AAME of 310.88±0.217µg gallic acid equivalent per mg of extract and 86.98±0.012µg quercetin equivalent per mg of extract respectively followed by AAAE and AAHE. Adenocalymma alliaceum could be used as available source of natural antioxidants, which might be helpful in preventing or reducing the progress of oxidative stress and the disorders associated with free radicals induced tissue damage.
... Interestingly, the publications' impact was directly related to the number of indigenous species in the authors' countries. Currently, the trend of research has shifted from identifying and recording the medicinal plant species used in traditional medicine [30,[43][44][45][46][47][48][49][50][51][52][53][54] to the evaluation of specific properties or treatment effects of crude plant extracts [55][56][57][58][59][60][61][62][63], or particular naturally derived products, such as flavonoids [64], alkaloids [65], tannins [66], saponins [67], phenols [68], and terpenoids [69] (an analysis of the provided bibliography together with applied methodology is presented in Table 1). Table 1. ...
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Ethnopharmacology, through the description of the beneficial effects of plants, has provided an early framework for the therapeutic use of natural compounds. Natural products, either in their native form or after crude extraction of their active ingredients, have long been used by different populations and explored as invaluable sources for drug design. The transition from traditional ethnopharmacology to drug discovery has followed a straightforward path, assisted by the evolution of isolation and characterization methods, the increase in computational power, and the development of specific chemoinformatic methods. The deriving extensive exploitation of the natural product chemical space has led to the discovery of novel compounds with pharmaceutical properties, although this was not followed by an analogous increase in novel drugs. In this work, we discuss the evolution of ideas and methods, from traditional ethnopharmacology to in silico drug discovery, applied to natural products. We point out that, in the past, the starting point was the plant itself, identified by sustained ethnopharmacological research, with the active compound deriving after extensive analysis and testing. In contrast, in recent years, the active substance has been pinpointed by computational methods (in silico docking and molecular dynamics, network pharmacology), followed by the identification of the plant(s) containing the active ingredient, identified by existing or putative ethnopharmacological information. We further stress the potential pitfalls of recent in silico methods and discuss the absolute need for in vitro and in vivo validation as an absolute requirement. Finally, we present our contribution to natural products’ drug discovery by discussing specific examples, applying the whole continuum of this rapidly evolving field. In detail, we report the isolation of novel antiviral compounds, based on natural products active against influenza and SARS-CoV-2 and novel substances active on a specific GPCR, OXER1.
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Hard Keratin (HK) from goat horn discards was dissolved using Ionic Liquid (IL) called Tetrabutylammonium hydroxide (TBAH, 40% aqueous solution), particles of HK were extracted, evaluated for antioxidant potential and in vitro cell viability against A549 and HEK-293 cell lines. Different material characterization techniques viz. PFM, XRD, FESEM/EDAS, FTIR, and TGA were employed to characterize the extracted hard keratin. PFM characterization revealed a piezoelectric coefficient (d33) of about 22.6 pm/V. FTIR analysis confirmed the presence of reactive Amides I, II & III at 1681, 1527, and 1249 cm-1 respectively. XRD spectrum demonstrated the amorphous phase of the material, and the Crystalline Index (C.I.) is 0.38. Characterization by TGA represents high thermal stability of extracted HK with a mass loss of 64.6% at 974.5°C temperature, reflective of the desired thermal stability for different applications. HK particles showed considerable antioxidant activity and inhibition of Human cell lines to an extent of 40.4% at 10 μg concentration for A549 (Lung Carcinoma cells) and 8.38% at 100 μg for HEK-293 (Human embryonic kidney cells).
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Kochiae Fructus (KF) was listed as ‘top grade’ medicinal material by the ‘Shennong's Herbal Classic of Materia Medica’ and has been used in traditional Chinese medicine to delay aging and treat inflammation, such as rubella, eczema, cutaneous pruritus, etc. Our research focused on the antioxidant capability of water decoction and fractions from KF based on 2,2-iphenyl-1-picrylhydrazyl (DPPH) free radical and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) cation radical scavenging assay, the ferric reducing antioxidant power assay, and inhibitory effects on DNA and protein oxidative damage. The results of total phenolics and flavonoids contents showed that ethyl acetate fraction (EAF) possessed the highest phenolics and flavonoids with values of 112.90 ± 9.58 mg gallic acid equivalents/g and 329.60 ± 20.93 mg rutin equivalents/g, respectively. At the same time, the results of antioxidant capacities showed that EAF possessed best antioxidant abilities. In addition, in this work, we evaluated the oral safety of the water decoction of KF (KFWD) via the 14-day acute and 28-day subacute toxicity tests. The results of in vivo toxicity assessment showed that KFWD did not cause significant changes in the general clinical symptoms, hematology and biochemical parameters, organ weights, or histopathological appearances in mice or rats. In summary, the reason why KF has the traditional effect on delaying aging may be related to the fact that its rich in flavonoids and phenolics. Simultaneously, no toxicity was detected after acute or subacute treatment of KFWD, providing valuable evidence for the traditional safe use of KF.
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The present study reports, the nutrient composition, total phenolic content and utilization potential of Ficus racemosa stem bark and Terminalia arjuna bark as an ingredient in tea, a popular non alcoholic beverage. The bark was found to be a good source of dietary fibre, minerals, and phenolic compounds. Further, the bark powder was used as an ingredient in the preparation of tea and the bark incorporated black tea (nutra tea) was found to contain significantly higher amounts of phenolic compounds compared to control tea. Sensory analysis of the nutra tea indicated no perceptible off-taste or off-aroma and the overall quality was similar to that of control and was acceptable in terms of all sensory attributes. The results suggest that the bark could be effectively used in the preparation of tea to derive its beneficial effects particularly attributable to those of phenolics. Keywords: Ficus racemosa, Terminalia arjuna, black tea, Nutra tea
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The antioxidant action of medicinal herbs used in Ghana for treating various ailments was evaluated in vitro and in vivo. Five plants, Desmodium adscendens, Indigofera arrecta, Trema occidentalis, Caparis erythrocarpus, and Thonningia sanguinea were tested for their free radical scavenging action by their interaction with 1,1-diphenyl-2-picrylhydrazyl (DPPH). Of these five plants, only Thonningia sanguinea was found to scavenge the DPPH radical. Lipid peroxidation in liver microsomes induced by H2O2 was also inhibited by T. sanguinea. The hepatoprotective effect of T. sanguinea was studied on acute hepatitis induced in rats by a single dose of galactosamine (GalN, 400 mg/kg, IP) and in mice by carbon tetrachloride (CCl4, 25 μl/kg, IP). GalN induced hepatotoxicity in rats as evidenced by an increase in alanine aminotransferase (ALT) and glutathione (GSH) S-transferase activities in serum was significantly inhibited when T. sanguinea extract (5 ml/kg, IP) was given to rats 12 hr and 1 hr before GalN treatment. The activity of liver microsomal GSH S-transferase, which is known to be activated by oxidative stress, was increased by the GalN treatment and this increase was blocked by T. sanguinea pretreatment. Similarly, T. sanguinea pretreatment also inhibited CCl4-induced hepatotoxicity in mice. These data indicate that T. sanguinea is a potent antioxidant and can offer protection against GalN- or CCl4-induced hepatotoxicity.
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Fifty-one tannins and forty-one flavonoids isolated from Oriental medicinal herbs were evaluated for their antioxidant ability with a 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-generating system. The results showed that tannins and certain flavonoids are potential free-radical scavengers, and that their activity against the DPPH radical is closely associated with their chemical structure. A comparison of the two classes of compounds showed that tannins have more potential than flavonoids because almost all the tannins demonstrated significant scavenging action within a low concentration range, whereas the activity of flavonoids varied distinctively among the different compounds. An increase of galloyl groups, molecular weight, and ortho-hydroxyl structure enhanced the activity of tannins, whereas the number and position of hydroxyl groups were important features for the scavenging of free radicals by flavonoids. Moreover, it appeared that when the free hydroxyl group was methoxylated or glycosylated, the inhibitory activity was obviously decreased or even abolished.
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Thirty-one wine samples differing in their origin of production and vintages were analyzed for total phenolic content, total and free sulfur dioxide contents, and superoxide radical scavenging potentials. The polyphenol content of red wine ranged from 735.9 to 2858 ppm, and that of white wine was in the range 259.4−720.5 ppm. Total sufur dioxide content ranged from 21.9 to 270.7 ppm, and had no correlation to the color of the wine. Superoxide radical scavenging activity values ranged from 39.3 to 215.9 units/mL for the white wine, and those of red varieties were 5−10 times higher. No correlation was observed between the free and total sulfur dioxide contents in the different wine samples tested and their superoxide radical scavenging activity values. A direct correlation between the color of the wine (r = 0.7517), its phenolic content (r = 0.9908), and the ability of the wine constituents to scavenge superoxide radical was, however, established by a simple regression analysis. Keywords: Wine; phenolics; radical scavenger; varietal difference
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The antioxidant activity of extracts from bark and heartwood of Acacia confusa was evaluated by various antioxidant assays, including free radical and superoxide radical scavenging assays and lipid peroxidation assay as well as hydroxyl radical-induced DNA strand scission assay. In addition, an ex vivo antioxidant assay using a flow cytometric technique was also employed in this study. The results indicate that both bark and heartwood extracts clearly have strong antioxidant effects. Similar inhibitory activities for each test sample were found for both 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical generation and lipid peroxidation. As for the superoxide radical scavenging activity, the heartwood extract was more effective than the bark extract. Furthermore, the heartwood extract protected ΦX174 supercoiled DNA against strand scission induced by ultraviolet photolysis of H2O2, and it reduced the amounts of intracellular hydrogen peroxide, a reactive oxygen species, when it was co-incubated with human promyelocytic leukemia (HL-60) cells under oxidative stress. Keywords: Acacia confusa; antioxidant activity; lipid peroxidation; reactive oxygen species; flow cytometry