Total phenols, ascorbic acid, b-carotene and lycopene in Portuguese wild edible mushrooms and their antioxidant activities

Food
Chemistry
Food Chemistry 103 (2007) 413–419
www.elsevier.com/locate/foodchem
Total phenols, ascorbic acid, b-carotene and lycopene in
Portuguese wild edible mushrooms and their antioxidant activities
Lillian Barros a, Maria-Joa˜o Ferreira a,b, Bruno Queiro´s a,b,
Isabel C.F.R. Ferreira a,*, Paula Baptista a
a CIMO- Escola Superior Agra´ria, Instituto Polite´cnico de Braganc¸a, Campus de Sta. Apolo´nia, Apartado 1172, 5301-855 Braganc¸a, Portugal
b Escola Superior de Sau´de, Instituto Polite´cnico de Braganc¸a, Av. D. Afonso V, 5300-121 Braganc¸a, Portugal
Received 2 February 2006; received in revised form 9 May 2006; accepted 17 July 2006
Abstract
The antioxidant activities of three Portuguese wild edible mushroom species, Leucopaxillus giganteus, Sarcodon imbricatus, andAgar-
icus arvensis, were evaluated. Methanolic extracts were screened for their reducing power, 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-
scavenging capacity, inhibition of erythrocytes hemolysis and antioxidant activity using the b-carotene linoleate model system. The
amounts of ascorbic acid, b-carotene and lycopene found in the mushroom extracts were very low. Otherwise, the high contents of phe-
nolic compounds might account for the good antioxidant properties found in all species. L. giganteus had the highest content of phenols
and proved to be the most active, presenting lower EC50 values in all the antioxidant activity assays.
Ó 2006 Elsevier Ltd. All rights reserved.
Keywords: Mushrooms; Antioxidant; Phenolics; Ascorbic acid; Beta-carotene; Lycopene
1. Introduction
ioli, Bellomo, & Galli, 1998a, Visioli, Bellosta, & Galli,
1998b). Free radicals and their uncontrolled production,
Free radical formation is associated with the normal
in fact, are responsible for several pathological processes,
natural metabolism of aerobic cells. The oxygen consump-
such as certain tumours (prostate and colon cancers)
tion inherent in cell growth leads to the generation of a ser-
(Keys, 1995) and coronary heart disease (Lipworth, Marti-
ies of oxygen free radicals. The interaction of these species
nez, Angell, Hsien, & Trichopoulos, 1997).
with molecules of a lipidic nature produces new radicals:
In the past few years, the suspected toxicity of some syn-
hydroperoxides and di?erent peroxides (Aust & Sringen,
thetic compounds used in food has raised interest in natu-
1982; Pryor, Lightsey, & Prier, 1982; Torel, Cillard, & Cil-
ral products (Fukushima & Tsuda, 1985; Stone, Leclair,
lard, 1986). This group of radicals (superoxide, hydroxyl
Ponder, Bagss, & Barret-Reis, 2003). Some industries, such
and lipoid peroxides) may interact with biological systems
as those related to food additive production, cosmetics,
in a clearly cytotoxic manner. In this respect, ?avonoids
and pharmaceuticals, have increased their e?orts in prepar-
and phenols have been shown to possess an important anti-
ing bioactive compounds from natural products by
oxidant activity toward these radicals, which is principally
extraction and puri?cation. Antioxidant compounds can
based on the redox properties of their phenolic hydroxyl
scavenge free radicals and increase shelf life by retarding
groups and the structural relationships between di?erent
the process of lipid peroxidation, which is one of the major
parts of their chemical structure (Bors & Saran, 1987; Vis-
reasons for deterioration of food products during process-
ing and storage (Halliwell, 1997; Halliwell & Gutteridge,
*
1999). Thus a need for identifying alternative natural and
Corresponding author. Tel.: +351 273303219; fax: +351 273325405.
E-mail address: iferreira@ipb.pt (I.C.F.R. Ferreira).
safe sources of food antioxidants has been created, and
0308-8146/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodchem.2006.07.038

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414
L. Barros et al. / Food Chemistry 103 (2007) 413–419
the search for natural antioxidants, especially of plant ori-
gal), in autumn 2005. After collection, the mushrooms were
gin, has notably increased in recent years (Skerget et al.,
grouped by taxon and were air-dried in a liophylizator (Ly-
2005).
8-FM-ULE, Snijders, HOLLAND) before analysis. Taxo-
Vegetables and fruits are rich sources of antioxidants,
nomic identi?cation was done according to several authors
such as vitamin A, vitamin C, vitamin E, carotenoids, poly-
(Bon, 1988; Courtecuisse, 1999; Courtecuisse & Duhem,
phenolic compounds and ?avonoids (Diplock et al., 1998),
1995;
Marchand,
1971–1986;
Moser,
1983)
and
which prevent free radical damage, reducing risk of chronic
representative voucher specimens were deposited at the
diseases. Thus, the consumption of dietary antioxidants
herbarium of Escola Superior Agra´ria of Instituto Polite´c-
from these sources is bene?cial in preventing cardiovascu-
nico de Braganc¸a.
lar diseases, especially atherosclerosis (Hu, 2000).
Mushrooms have been used for traditional foods and
2.3. Sample preparation
medicines in Asia (Chang, 1996). Mushrooms contain var-
ious polyphenolic compounds recognized as an excellent
A ?ne dried mushroom powder (20 mesh) sample (5 g)
antioxidant due to their ability to scavenge free radicals
was continuously extracted with methanol in a Soxhlet
by single-electron transfer (Hirano et al., 2001). Some com-
apparatus for 24 h. The methanolic extract was evaporated
mon edible mushrooms, which are widely consumed in
to dryness at 40 °C and redissolved in methanol at a con-
Asian culture, have currently been found to possess antiox-
centration of 5 mg/ml, and stored at 4 °C prior to further
idant activity, which is well correlated with their total phe-
use.
nolic content (Cheung & Cheung, 2005; Cheung, Cheung,
& Ooi, 2003; Lo & Cheung, 2005; Mau, Chang, Huang,
2.4. Determination of antioxidant components
& Chen, 2004, 2002; Yang, Lin, & Mau, 2002; Yen &
Hung, 2000).
Phenolic compounds in the mushroom methanolic
Recently, we described the ?rst study on the antioxidant
extracts were estimated by a colorimetric assay, based on
activity of Portuguese wild edible mushrooms (Lactarius
procedures described by Singleton and Rossi with some
deliciosus and Tricholoma portentosum), comparing the
modi?cations (Singleton & Rossi, 1965). Brie?y, 1 ml of
entire mushroom, the cap and stipe DPPHÅ-scavenging
sample was mixed with 1 ml of Folin and Ciocalteu’s phe-
capacities and reducing powers (Ferreira, Baptista, Vilas-
nol reagent. After 3 min, 1 ml of saturated sodium carbon-
Boas, & Barros, 2007). Herein, we report the antioxidant
ate solution was added to the mixture and it was adjusted
activity of three new Portuguese wild edible mushroom
to 10 ml with distilled water. The reaction was kept in the
species (Leucopaxillus giganteus, Sarcodon imbricatus,
dark for 90 min, after which the absorbance was read at
Agaricus arvensis), and their correlation with phenol,
725 nm (Analytikijena 200-2004 spectrophotometer). Gal-
ascorbic acid, beta-carotene and lycopene contents. For
lic acid was used to calculate the standard curve (0.01–
the screening of mushroom antioxidant properties, we eval-
0.4 mM). Estimation of the phenolic compounds was
uated their reducing power, DPPH radical scavenging
carried out in triplicate. The results were mean val-
activity and inhibition of erythrocyte hemolysis, and we
ues ± standard deviations and expressed as mg of gallic
also used the b-carotene linoleate model system.
acid equivalents (GAEs) per g of extract.
Ascorbic acid was determined according to the method
2. Materials and methods
of Klein and Perry (1982). The dried methanolic extract
(100 mg) was extracted with 10 ml of 1% metaphosphoric
2.1. Standards and reagents
acid for 45 min at room temperature and ?ltered through
Whatman No. 4 ?lter paper. The ?ltrate (1 ml) was mixed
Standards BHA (2-tert-butyl-4-methoxyphenol), TBHQ
with 9 ml of 2,6-dichlorophenolindophenol and the absor-
(tert-butylhydroquinone), L-ascorbic acid, a-tocopherol
bance was measured within 30 min at 515 nm against a
and gallic acid were purchased from Sigma (St. Louis,
blank. Content of ascorbic acid was calculated on the basis
MO, USA). 2,2-Diphenyl-1-picrylhydrazyl (DPPH) was
of the calibration curve of authentic L-ascorbic acid (0.020–
obtained from Alfa Aesar (Ward Hill, MA, USA). All
0.12 mg/ml). The assays were carried out in triplicate; the
other chemicals were obtained from Sigma Chemical Co.
results were mean values ± standard deviations and
(St. Louis, MO, USA). Methanol was obtained from
expressed as mg of ascorbic acid/g of extract.
Pronalab (Lisbon, Portugal). Water was treated in a Mili-
b-Carotene and lycopene were determined according to
Q water puri?cation system (TGI Pure Water Systems,
the method of Nagata and Yamashita (1992). The dried
USA).
methanolic extract (100 mg) was vigorously shaken with
10 ml of acetone–hexane mixture (4:6) for 1 min and ?l-
2.2. Samples
tered through Whatman No. 4 ?lter paper. The absorbance
of the ?ltrate was measured at 453, 505, 645 and 663 nm.
Samples of L. giganteus and A. arvensis were colleted
Contents of b-carotene and lycopene were calculated
under grassland whereas S. imbricatus was collected under
according to the following equations: lycopene (mg/
live pine trees (Pinus sp.), in Braganc¸a (northeast of Portu-
100 ml) = À0.0458 A663 + 0.372 A505 À 0.0806 A453; b-car-

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L. Barros et al. / Food Chemistry 103 (2007) 413–419
415
otene (mg/100 ml) = 0.216 A663 À 0.304 A505 + 0.452 A453.
rated from the plasma and the bu?y coat, were washed
The assays were carried out in triplicate; the results were
three times with 10 ml of 10 mM phosphate bu?er saline
mean values ± standard deviations and expressed as mg
(PBS) at pH 7.4 (prepared by mixing 10 mM NaH2PO4
of carotenoid/g of extract.
and Na2HPO4, and 125 mM NaCl in 1 l of distilled water)
and centrifuged at 1500 g for 5 min. During the last wash-
2.5. Reducing power assay
ing, the erythrocytes were obtained by centrifugation at
1500 g for 10 min. 0.1 ml of a 20% suspension of erythro-
The reducing power was determined according to the
cytes in PBS was added to 0.2 ml of 200 mM 2,20-azo-
method of Oyaizu (1986). Various concentrations of mush-
bis(2-amidinopropane)dihydrochloride (AAPH) solution
room methanolic extracts (2.5 ml) were mixed with 2.5 ml
(in PBS) and 0.1 ml of mushroom methanolic extracts of
of 200 mM sodium phosphate bu?er (pH 6.6) and 2.5 ml
di?erent concentrations. The reaction mixture was shaken
of 1% potassium ferricyanide. The mixture was incubated
gently (30 rpm) while being incubated at 37 °C for 3 h.
at 50 °C for 20 min. After 2.5 ml of 10% trichloroacetic
The reaction mixture was diluted with 8 ml of PBS and cen-
acid (w/v) were added, the mixture was centrifuged at
trifuged at 3000 g for 10 min; the absorbance of its super-
1000 rpm for 8 min (Centorion K24OR-2003 refrigerated
natant was then read at 540 nm by a spectrophotometer,
centrifuge). The upper layer (5 ml) was mixed with 5 ml
after ?ltration with a syringe ?lter (cellulose membrane
of deionised water and 1 ml of 0.1% of ferric chloride,
30 mm, 0.20 lm, Titan). The percentage hemolysis inhibi-
and the absorbance was measured spectrophotometrically
tion was calculated by the equation: % hemolysis inhibi-
at 700 nm. The assays were carried out in triplicate and
tion = [(AAAPH–AS)/AAAPH] · 100,
where
AS
is
the
the results expressed as mean values ± standard deviations.
absorbance of the sample containing the mushroom
The extract concentration providing 0.5 of absorbance
extract, and AAAPH is the absorbance of the control sample
(EC50) was calculated from the graph of absorbance at
containing no mushroom extract. The assays were carried
700 nm against extract concentration. BHA and a-tocoph-
out in triplicate and the results expressed as mean val-
erol were used as standards.
ues ± standard deviations. The extract concentration pro-
viding 50% inhibition (EC50) was calculated from the
2.6. DPPH radical-scavenging assay
graph of hemolysis inhibition percentage against extract
concentration. L-ascorbic acid was used as standard.
The capacity to scavenge the ‘‘stable’’ free radical
DPPHÅ was monitored according to the method of Hatano,
2.8. Antioxidant assay using the b-carotene linoleate model
Kagawa, Yasuhara, and Okuda (1988). Various concentra-
system
tions of methanolic extracts from mushrooms (0.3 ml) were
mixed with 2.7 ml of methanolic solution containing
The antioxidant activity of mushroom extracts was eval-
DPPH radicals (6 · 10À5 mol/l). The mixture was shaken
uated by the b-carotene linoleate model system (Mi-Yae,
vigorously and left to stand for 60 min in the dark (until
Tae-Hun, & Nak-Ju, 2003). A solution of b-carotene was
stable absorption values were obtained). The reduction of
prepared by dissolving 2 mg of b-carotene in 10 ml of chlo-
the DPPH radical was determined by measuring the
roform. Two millilitres of this solution were pipetted into a
absorption at 517 nm. The radical-scavenging activity
100 ml round-bottom ?ask. After the chloroform was
(RSA) was calculated as a percentage of DPPH discolor-
removed at 40 °C under vacuum, 40 mg of linoleic acid,
ation
using
the
equation:
%RSA = [(ADPPH-AS)/
400 mg of Tween 80 emulsi?er and 100 ml of distilled water
ADPPH] · 100, where AS is the absorbance of the solution
were added to the ?ask with vigorous shaking. Aliquots
when the sample extract has been added at a particular
(4.8 ml) of this emulsion were transferred into di?erent test
level, and ADPPH is the absorbance of the DPPH solution.
tubes containing 0.2 ml of di?erent concentrations of the
The assays were carried out in triplicate and the results
mushroom extracts. The tubes were shaken and incubated
expressed as mean values ± standard deviations. The
at 50 °C in a water bath. As soon as the emulsion was
extract concentration providing 50% inhibition (EC50)
added to each tube, the zero time absorbance was mea-
was calculated from the graph of RSA percentage against
sured at 470 nm using a spectrophotometer. Absorbance
extract concentration. BHA and a-tocopherol were used
readings were then recorded at 20 min intervals until the
as standards.
control sample had changed colour. A blank, devoid of
b-carotene, was prepared for background subtraction.
2.7. Assay for erythrocyte hemolysis mediated by peroxyl
Antioxidant activity was calculated using the following
free radicals
equation: antioxidant activity = (b-carotene content after
2 h of assay/initial b-carotene content) · 100. The assays
The antioxidant activity of the mushroom methanolic
were carried out in triplicate and the results expressed as
extracts was measured as the inhibition of erythrocyte
mean values ± standard deviations. The extract concentra-
hemolysis (Miki, Tamia, Mino, Yamamoto, & Niki,
tion providing 50% antioxidant activity (EC50) was calcu-
1987). Blood was obtained from a male ram (churra galega
lated from the graph of antioxidant activity percentage
transmontana) of body weight $67 kg. Erythrocytes, sepa-
against extract concentration. TBHQ was used as standard.

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L. Barros et al. / Food Chemistry 103 (2007) 413–419
3. Results and discussion
1.5
3.1. Determination of antioxidant components
1
Table 1 shows the phenol, ascorbic acid, b-carotene and
lycopene concentration in the mushroom extracts. Whereas
0.5
total phenols were the major antioxidant components
Abs at 700 nm
found in the mushroom extracts, ascorbic acid was found
in small amounts (0.13–0.35 mg/g), and b-carotene and
0
lycopene were only found in vestigial amounts (<3
0
1
2
3
4
5
6
lg/g),
Concentration (mg/ml)
which is in agreement with other authors (Mau, Lin, &
Song, 2002). These antioxidants were determined in di?er-
L. giganteus
S. imbricatus
A. arvensis
ent mushrooms but ascorbic acid and b-carotene were not
Fig. 1. Reducing power of mushroom methanolic extracts (higher
detected by spectrophotometry and HPLC, respectively.
absorbance indicates higher reducing power). Each value is expressed as
L. giganteus extracts showed the highest phenolic content
mean ± standard deviation (n = 3).
(6.29 ± 0.20 mg/g); the amount found in A. arvensis
extracts (2.83 ± 0.09 mg/g) was slightly lower than the con-
ing power values than did those from S. imbricatus (Fig. 1).
tent found in S. imbricatus (3.76 ± 0.11 mg/g). The highest
It was reported that the reducing properties are generally
content of total phenols in the L. giganteus extracts might
associated with the presence of reductones, which have
account for the better results found for their antioxidant
been shown to exert antioxidant action by breaking the free
activity. In fact, it had been reported that the antioxidant
radical chain by donating a hydrogen atom (Shimada,
activity of plant materials is well correlated with the content
Fujikawa, Yahara, & Nakamura, 1992). Accordingly, L.
of phenolic compounds. Polyphenols, such as BHT (butyl-
giganteus might contain higher amounts of reductone,
ated hydroxytoluene) and gallate, are known to be e?ective
which could react with free radicals to stabilise and block
antioxidants (Velioglu, Mazza, Gao, & Oomah, 1998).
radical chain reactions.
3.2. Reducing power assay
3.3. Radical-scavenging activity (RSA) assay
Fig. 1 shows the reducing power of mushroom methan-
The free radical DPPHÅ possesses a characteristic
olic extracts as a function of their concentration. In this
absorption at 517 nm (purple in colour), which decreases
assay, the yellow colour of the test solution changes to var-
signi?cantly on exposure to radical-scavengers (by provid-
ious shades of green and blue, depending on the reducing
ing hydrogen atoms or by electron donation). A lower
power of each compound. The presence of reducers (i.e.
absorbance at 517 nm indicates a higher radical-scavenging
antioxidants) causes the conversion of the Fe3+/ferricyanide
activity of the extract. Free radical-scavenging is one of the
complex used in this method to the ferrous form. Therefore,
known mechanisms by which antioxidants inhibit lipid oxi-
by measuring the formation of Perl’s Prussian blue at
dation. This test is a standard assay in antioxidant activity
700 nm, we can monitor the Fe2+ concentration; a higher
studies and o?ers a rapid technique for screening the RSA
absorbance at 700 nm indicates a higher reducing power.
of speci?c compounds or extracts (Amarowicz, Pegg, Rah-
The reducing power of the mushroom methanolic
imi-Moghaddam, Barl, & Weil, 2004).
extracts increased with concentration. Reducing powers
The RSA values of mushroom methanolic extracts are
obtained for all the mushrooms were excellent (Fig. 1); at
presented in Fig. 2; results are expressed as the ratio per-
5 mg/ml they were above 0.67 and in the order: L. gigan-
centage of sample absorbance decrease and the absorbance
teus > S. imbricatus $ A. arvensis. At 5 mg/ml, reducing
of DPPH solution in the absence of extract at 517 nm.
powers of methanolic extracts from wild edible mushrooms
From the analysis of Fig. 2, we can conclude that the scav-
were 0.67–1.47, and at 1 mg/ml were 0.072–0.26. Reducing
enging e?ects of mushrooms methanolic extracts on DPPH
powers of BHA at 3.6 mg/ml and a-tocopherol at 8.6 mg/
radicals increased with the concentration increase and were
ml were only 0.12 and 0.13, respectively. Methanolic
excellent for L. giganteus (100% at 5 mg/ml), even higher
extracts from A. arvensis showed only slightly lower reduc-
than the scavenging e?ects of BHA (96% at 3.6 mg/ml)
and a-tocopherol (95% at 8.6 mg/ml). The RSA values
were good for S. imbricatus (80% at 5 mg/ml) and moder-
Table 1
Contents of total phenols, ascorbic acid, b-carotene and lycopene in the
ate for A. arvensis (68.3% at 5 mg/ml).
mushroom extract
L. giganteus
S. imbricatus
A. arvensis
3.4. Assay for erythrocyte hemolysis mediated by peroxyl
free radicals
Total phenols (mg/g)
6.29 ± 0.20
3.76 ± 0.11
2.83 ± 0.09
Ascorbic acid (mg/g)
0.13 ± 0.0069
0.16 ± 0.0072
0.35 ± 0.0015
b-Carotene (lg/g)
1.88 ± 0.090
2.53 ± 0.11
2.97 ± 0.12
The oxidative hemolysis in erythrocytes induced by
Lycopene (lg/g)
0.69 ± 0.034
1.3 ± 0.070
1.0 ± 0.049
AAPH has been extensively studied as a model for perox-

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L. Barros et al. / Food Chemistry 103 (2007) 413–419
417
100
3.5. Antioxidant assay using the b-carotene linoleate model
system
75
Fig. 4 shows the antioxidant activity of the mushroom
50
extracts as measured by the bleaching of b-carotene. The
antioxidant activity of carotenoids is based on the radical
25
adducts of carotenoids with free radicals from linoleic acid.
Scavenging Effect (%)
The linoleic acid free radical attacks the highly unsaturated
0
0
1
2
3
4
5
6
b-carotene models. The presence of di?erent antioxidants
Concentration (mg/ml)
can hinder the extent of b-carotene-bleaching by neutraliz-
ing the linoleate-free radical and other free radicals formed
L. giganteus
S. imbricatus
A. arvensis
in the system (Jayaprakasha, Singh, & Sakariah, 2001).
Fig. 2. Scavenging activity (%) on DPPH radicals of mushroom methan-
Accordingly, the absorbance decreased rapidly in samples
olic extracts. Each value is expressed as mean ± standard deviation
without antioxidant whereas, in the presence of an antiox-
(n = 3).
idant, they retained their colour, and thus absorbance, for
a longer time. Antioxidant activities of L. giganteus, S.
idative damage in biomembranes (Zhang et al., 1997).
imbricatus and A. arvensis extracts increased with their
AAPH is a peroxyl radical initiator that generates free rad-
increasing concentration. Their antioxidant activities were
icals by its thermal decomposition and will attack the
61.4%, 54.3% and 46.7% at 5 mg/ml, but antioxidant activ-
erythrocytes to induce the chain oxidation of lipid and pro-
ity of TBHQ standard reached 82.2% at 2 mg/ml. It is
tein, disturbing the membrane organization and eventually
probable that the antioxidative components in the mush-
leading to hemolysis. In this study, the protective e?ect of
room extracts can reduce the extent of b-carotene destruc-
the mushroom extracts on hemolysis by peroxyl radical-
tion by neutralizing the linoleate free radical and other free
scavenging activity was investigated. Fig. 3 shows inhibi-
radicals formed in the system. Again, L. giganteus was the
tion percentage of hemolysis, as a result of protection
most e?ective, with an EC50 value of 2 mg/ml.
against the oxidative damage of cell membranes of erythro-
Table 2 shows the EC50 values for the antioxidant activ-
cytes from ram, induced by AAPH. The mushroom
ity assays obtained from each mushroom methanolic
extracts inhibited hemolysis of erythrocytes in a concentra-
extract.
tion-dependent manner. Once more, L. giganteus showed
Overall, L. giganteus revealed better antioxidant proper-
higher protective e?ect against erythrocytes hemolysis
ties (lower EC50 values) than did either S. imbricatus or A.
(72.8% at 5 mg/ml) than did the other mushrooms (34.2%
arvensis, which is in agreement with the higher content of
for S. imbricatus and 31.8% for A. arvensis). However,
phenols found in the ?rst species. The EC50 values obtained
the inhibition percentage of the standard L-ascorbic acid
for reducing power and scavenging e?ects on DPPH radi-
on hemolysis of red blood cells was much higher (94.6%
cals were better than those for hemolysis inhibition medi-
at 1 mg/ml) than those of mushroom extracts.
ated by peroxyl free radicals and for the antioxidant
activity using the linoleate-bcarotene system. A relation-
ship between the reducing power, DPPHÅ-scavenging activ-
ity, hemolysis inhibition and b-carotene-bleaching extent
was found, indicating that the mechanisms of action of
the extracts for the antioxidant activity may be identical,
100
100
75
75
50
50
25
25
Antioxidant activity (%)
0
Hemolysis inhibition (%)
0
0
1
2
3
4
5
6
Concentration (mg/ml)
0
1
2
3
4
5
6
Concentration (mg/ml)
L. giganteus
S. imbricatus
A. arvensis
L. giganteus
S. imbricatus
A. arvensis
Fig. 4. Antioxidant activity (%) of the mushroom methanolic extracts by
Fig. 3. Hemolysis inhibition (%) of the mushroom methanolic extracts.
b-carotene bleaching method. Each value is expressed as mean ± standard
Each value is expressed as mean ± standard deviation (n = 3).
deviation (n = 3).

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418
L. Barros et al. / Food Chemistry 103 (2007) 413–419
Table 2
EC50 values (mg/ml) of mushroom extracts in the antioxidant activity evaluation assays
Samples
Reducing power (EC a
b
c
d
50 )
DPPH (EC50 )
Hemolysis inhibition (EC50 )
b-Carotene bleaching (EC50 )
L. giganteus
1.71
1.44
1.80
2.00
S. imbricatus
2.79
1.67
>5
3.97
A. arvensis
2.86
3.50
>5
>5
a EC50 (mg/ml): e?ective concentration at which the absorbance is 0.5.
b EC50 (mg/ml): e?ective concentration at which 50% of DPPH radicals are scavenged.
c EC50 (mg/ml): e?ective concentration at which 50% of the erythrocytes hemolysis are inhibited.
d EC50 (mg/ml): e?ective concentration at which the antioxidant activity is 50%.
being related to the content of total phenols. Though other
mushrooms from northeast Portugal: individual cap and stipe activity.
antioxidants were probably present in these mushroom
Food Chemistry, 100, 1511–1516.
Fukushima, N. S., & Tsuda, H. (1985). Carcinogenity and modi?cation of
extracts, the amounts of ascorbic acid, b-carotene and lyco-
the carcinogenic response by BHA, BHT and other antioxidants.
pene found in the three Portuguese mushroom extracts
Critical Reviews in Toxicology, 15, 109–150.
were very low, which emphasises the idea that phenolic
Halliwell, B. (1997). Antioxidants in human health and disease. Annual
compounds could make a signi?cant contribution to the
Review of Nutrition, 16, 33–50.
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Hatano, T., Kagawa, H., Yasuhara, T., & Okuda, T. (1988). Two new
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?avonoids and other constituents in licorice root: their relative
the membranes of erythrocytes incubated with AAPH, and
astringency and radical scavenging e?ects. Chemical and Pharmaceu-
can reduce the extent of b-carotene destruction by neutral-
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Document Outline

• Total phenols, ascorbic acid, beta -carotene and lycopene in Portuguese wild edible mushrooms and their antioxidant activities
  • Introduction
  • Materials and methods
    • Standards and reagents
    • Samples
    • Sample preparation
    • Determination of antioxidant components
    • Reducing power assay
    • DPPH radical-scavenging assay
    • Assay for erythrocyte hemolysis mediated by peroxyl free radicals
    • Antioxidant assay using the beta -carotene linoleate model system
  • Results and discussion
    • Determination of antioxidant components
    • Reducing power assay
    • Radical-scavenging activity (RSA) assay
    • Assay for erythrocyte hemolysis mediated by peroxyl free radicals
    • Antioxidant assay using the beta -carotene linoleate model system
  • Acknowledgements
  • References

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