Nutrient Requirements

Total Antioxidant Capacity of Plant Foods, Beverages and Oils Consumed

in Italy Assessed by Three Different In Vitro Assays1
Nicoletta Pellegrini,2 Mauro Serafini,* Barbara Colombi, Daniele Del Rio, Sara Salvatore,
Marta Bianchi and Furio Brighenti
Department of Public Health, University of Parma, Parma, Italy and *Antioxidant Research Laboratory at the
Unit of Human Nutrition, National Institute for Food and Nutrition Research, Rome, Italy

KEY WORDS:

antioxidant capacity

fruits

vegetables

beverages

oils

the synergic and redox interactions among the different molecules present in the food. Finally, geographical differences in
food composition data should be considered when applying
compositional databases to regional surveys.
Several methods were developed recently for measuring the
total antioxidant capacity of food and beverages (9 11); these
assays differ in their chemistry (generation of different radicals
and/or target molecules) and in the way end points are measured (12). Because different antioxidant compounds may act
in vivo through different mechanisms, no single method can
fully evaluate the TAC of foods. The objective of this study
was to assess the TAC of plant food, beverages and oils
commonly consumed in Italy using different methods to obtain
robust data useful for determining the potential intake of
antioxidants in Italian population studies. To this aim, three
methods, i.e., Trolox equivalent antioxidant capacity (TEAC)
(12), total radical-trapping antioxidant parameter (TRAP)
(13) and ferric reducing-antioxidant power (FRAP) (11), were

The consumption of fruits and vegetables has been inversely associated with morbidity and mortality from degenerative diseases (15). It is not known which dietary constituents are responsible for this association, but antioxidants
appear to play a major role in the protective effect of plant
foods (6 8). Epidemiologic studies that analyze the health
implications of dietary components rely on the intake estimates in sample populations found in databases that list the
components content in commonly consumed foods. Therefore, the availability of appropriate and complete food composition data is crucial. Due to the chemical diversity of
antioxidant compounds present in foods, complete databases
on food antioxidant content are not yet available. In addition,
levels of single antioxidants in food do not necessarily reflect
their total antioxidant capacity (TAC)3; this also depends on

1
Supported by the EC project Healthy Market (IST-200133204) and the
National Research Council of Italy (CNR 01.00923. CT26/115.25178).
2
To whom correspondence should be addressed.
E-mail: mailnico@nemo.unipr.it.
3
Abbreviations used: ABAP, 2,2-azobis(2-amidinopropane) dihydrochloride;
ABTS, 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid); ABTS, 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation; EVOO, extra virgin olive
oil; FRAP, ferric reducing-antioxidant power; OO, olive oil; R-PE, R-phycoerythrin;

RSD, relative standard deviation; TAC, total antioxidant capacity; TEAC, Trolox
equivalent antioxidant capacity; TPTZ, 2,4,6-tripyridyl-s-triazine; TRAP, total radical-trapping antioxidant parameter; Trolox, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid.

0022-3166/03 $3.00 2003 American Society for Nutritional Sciences.

Manuscript received 3 April 2003. Initial review completed 7 May 2003. Revision accepted 20 June 2003.
2812

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ABSTRACT Epidemiologic studies have demonstrated an inverse association between consumption of fruits and
vegetables and morbidity and mortality from degenerative diseases. The antioxidant content of fruits and vegetables may contribute to the protection they offer from disease. Because plant foods contain many different classes
and types of antioxidants, knowledge of their total antioxidant capacity (TAC), which is the cumulative capacity of
food components to scavenge free radicals, would be useful for epidemiologic purposes. To accomplish this, a
variety of foods commonly consumed in Italy, including 34 vegetables, 30 fruits, 34 beverages and 6 vegetable oils,
were analyzed using three different assays, i.e., Trolox equivalent antioxidant capacity (TEAC), total radicaltrapping antioxidant parameter (TRAP) and ferric reducing-antioxidant power (FRAP). These assays, based on
different chemical mechanisms, were selected to take into account the wide variety and range of action of
antioxidant compounds present in actual foods. Among vegetables, spinach had the highest antioxidant capacity
in the TEAC and FRAP assays followed by peppers, whereas asparagus had the greatest antioxidant capacity in
the TRAP assay. Among fruits, the highest antioxidant activities were found in berries (i.e., blackberry, redcurrant
and raspberry) regardless of the assay used. Among beverages, coffee had the greatest TAC, regardless of the
method of preparation or analysis, followed by citrus juices, which exhibited the highest value among soft
beverages. Finally, of the oils, soybean oil had the highest antioxidant capacity, followed by extra virgin olive oil,
whereas peanut oil was less effective. Such data, coupled with an appropriate questionnaire to estimate antioxidant intake, will allow the investigation of the relation between dietary antioxidants and oxidative stressinduced
diseases. J. Nutr. 133: 28122819, 2003.

TOTAL ANTIOXIDANT CAPACITY OF PLANT FOODS

selected. The TEAC assay measures the ability of antioxidants

to quench a radical cation (ABTS) in both lipophilic and
hydrophilic environments (12). The TRAP and FRAP assays
evaluate the chain-breaking antioxidant potential (13) and
the reducing power of the sample (11), respectively.
MATERIALS AND METHODS
Chemicals
The 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid
(Trolox), 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), and 2,4,6-tripyridyl-s-triazine (TPTZ) were
purchased from Sigma-Aldrich (St. Louis, MO). R-Phycoerythrin
(R-PE) was purchased from Prozyme (San Leandro, CA); 2,2-azobis(2-amidinopropane) dihydrochloride (ABAP) was purchased from
Waco Chemicals (Richmond, VA). All chemicals and solvents used
were HPLC-grade and purchased from Carlo Erba (Milan, Italy).
High-purity water was produced in the laboratory by using an AlphaQ system (Millipore, Marlborough, MA).

Samples

local supermarkets. After washing and cutting, equal amounts of each

food were pooled, mixed and homogenized under nitrogen in a high
speed blender. A precisely weighed amount of the homogenized
sample (1 g) was extracted with 4 mL of water under agitation for
15 min at room temperature, centrifuged at 1000 g for 10 min and
the supernatant collected. The extraction was repeated with 2 mL of
water and the two supernatants were combined. The pulp residue was
reextracted by the addition of 4 mL of acetone under agitation for 15
min at room temperature, centrifuged at 1000 g for 10 min and the
supernatant collected. The extraction was repeated with 2 mL of
acetone and the two supernatants were combined. In the case of
lipid-rich foods (e.g., avocados and olives), the pulp residue was
reextracted twice by the addition of 2 mL of chloroform under
agitation for 15 min at room temperature, centrifuged at 1000 g for
10 min and the supernatant collected. All food extracts were adequately diluted in the appropriate solvent (depending on their activity) and immediately analyzed in duplicate for their antioxidant
capacity. The variation in the TEAC, TRAP and FRAP values for
replicates was always between 3 and 10% relative standard deviation
(RSD). When the RSD was higher than 10%, the analyses were
repeated to confirm the value.
TEAC (Trolox equivalent antioxidant capacity) assay. The
method is based on the ability of antioxidant molecules to quench the
long-lived ABTS, a blue-green chromophore with characteristic
absorption at 734 nm, compared with that of Trolox, a water-soluble
vitamin E analog. The addition of antioxidants to the preformed
radical cation reduces it to ABTS, determining a decolorization. A
stable stock solution of ABTS was produced by reacting a 7 mmol/L
aqueous solution of ABTS with 2.45 mmol/L potassium persulfate
(final concentration) and allowing the mixture to stand in the dark at
room temperature for 1216 h before use (12). At the beginning of
the analysis day, an ABTS working solution was obtained by the
dilution in ethanol of the stock solution to an absorbance of 0.70
0.02 AU at 734 nm, verified by a Hewlett-Packard 8453 Diode
Array spectrophotometer (HP, Waldbronn, Germany), and used as
mobile phase in a flow-injection system, according to Pellegrini et al.
(15). Results were expressed as TEAC in mmol of Trolox per kg (solid
foods and oils) or per L (beverages) of sample. All solid food extracts,
obtained by water, acetone and chloroform, were analyzed by this
assay.
TRAP (total radical-trapping antioxidant parameter) assay.
The TRAP was determined according to the method of Ghiselli et al.
(13) based on the protection provided by antioxidants on the fluorescence decay of R-phycoerythrin (lag-phase) during a controlled
peroxidation reaction. Briefly, 120 L of diluted sample were added
to 2.4 mL of phosphate buffer (pH 7.4), 375 L of bidistilled water,
30 L of diluted R-PE and 75 L of ABAP; the reaction kinetics at
38C were recorded for 45 min (or more, if necessary) by a LS-55
luminescence spectrometer (Perkin Elmer, Wellesley, MA). TRAP
values were calculated from the length of the lag-phase due to the
sample compared with that of Trolox and expressed as mmol of
Trolox per kg (solid foods) or per L (beverages) of sample. All solid
food extracts, with the exception of those in chloroform, were analyzed by this procedure.
FRAP (ferric reducing-antioxidant power) assay. The FRAP
was assessed according to Benzie and Strain (11) using a HewlettPackard 8453 diode array spectrophotometer. The method is based on
the reduction of the Fe3-TPTZ complex to the ferrous form at low
pH. This reduction is monitored by measuring the absorption change
at 593 nm. Briefly, 3 mL of working FRAP reagent prepared daily was
mixed with 100 L of diluted sample; the absorbance at 593 nm was
recorded after a 30-min incubation at 37C. FRAP values were
obtained by comparing the absorption change in the test mixture
with those obtained from increasing concentrations of Fe3 and
expressed as mmol of Fe2 equivalents per kg (solid food) or per L
(beverages) of sample. All solid food extracts, with the exception of
those in chloroform, were analyzed by this procedure.
Statistical analysis. To verify the association among methods,
Pearson correlation analysis was performed using a statistical package
running on a PC (Statistical Statsoft, Tulsa, OK); P-values 0.05
were considered significant.

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The selection of the samples was based on food consumption data

of the EPIC cohort of Varese province (Italy) kindly provided by the
Epidemiology Unit of the National Cancer Institute based in Milan
and developed on the basis of 24-h recalls recorded in the North Italy
area (Dr V. Krogh, Department of Epidemiology, National Cancer
Institute, Milan, Italy, personal communication). Once the main
putative contributors of antioxidants in the Italian diet were identified, three food samples for each food item were purchased, selecting
the three cultivars and/or brands with the highest sales in the Italian
market. Samples were then prepared, mixed in equal proportions and
analyzed for TAC, as reported below. Like other nutrients, the
estimation of the overall dietary intake of TAC does not require an
estimate of the variance for any single food item if the value of the
given food as consumed by the responder is sufficiently close to the
average value. The same approach was used previously to generate
food TAC data (14).
Beverages. The following beverages were purchased in local
supermarkets: soft drinks (cola and lemon iced tea), fruit juices
(orange, lemon, apple, pineapple, pear, grapefruit, peach, apricot,
mixed fruits and tropical fruits), alcoholic drinks (beer, rum, whiskey,
grappa, cognac, and red, rose and white wines of different geographical origins) and vinegar from red wine. Teas and chamomile were
prepared by brewing an infusion (2 g) in 250 mL boiling water for
5 min. Soluble coffee was prepared by solubilizing a coffee serving
(2 g) in 40 mL of boiling water. Extracted coffee was prepared using
an Italian Moka coffee machine. Espresso coffees were purchased in
local coffee shops.
Before analysis, beverages were adequately diluted in high purity
water, depending on their presumed activity. Carbon dioxide from
cola and beer was removed completely by magnetic stirring under
nitrogen. Diluted beverages were centrifuged for 5 min at 1000 g,
and the supernatant was collected and analyzed without further
preparation.
Oils. Oils (olive, extra virgin olive, sunflower, corn, soybean and
peanut oils) were purchased in local supermarkets. Before analysis,
oils were diluted in n-hexane, depending on their presumed activity.
Fruits and vegetables. Fruits (n 23; red Delicious and yellow
Golden apples, apricot, banana, clementine, yellow grapefruit, orange, yellow peach, pear, honeydew and cantaloupe melons, pineapple, red plum, tangerine, water melon, kiwi fruit, black and white
grapes, loquat, fig, prickly pear, black and green olives), berries (n
7; cultivated and wild strawberries, blackberry, blueberry, raspberry, cherry, and redcurrant) and vegetables (n 34; artichoke,
arugula, asparagus, avocado, beet, red cooked beetroot, broccoli,
green and Savoy cabbages, carrot, cauliflower, celery, chicory, cucumber, eggplant, endive, fennel, green bean, leek, green lettuce, mushroom, yellow onion, chili and red bell peppers, potato, pumpkin,
radicchio, Swiss chard stalk, red radish, spinach, salad and sauce
tomatoes, turnip tops and zucchini) were collected at three different

2813

PELLEGRINI ET AL.

2814

RESULTS AND DISCUSSION

The total antioxidant capacity of a variety of foods used in
the Italian diet (i.e., vegetables, fruits, beverages and oils) was
evaluated using three different assays. The food extracts had
different antioxidant capacities in relation to the method
applied; thus, the same item often ranked differently depending on the assay. Moreover, even for values obtained from the
same assay, a comparison with the data in the literature was
problematic due to the large variability within the food item
and to the lack of standardization of the assays. For this
discussion, the ranking order of the TAC values will be used.
In the case of solid foods (i.e., vegetables and fruits), the waterand lipid-soluble extracts were analyzed separately and the
overall TAC values were obtained from the sum of the two
extract values. The approach of using only one solvent for
fruits and vegetables extraction, generally used in TAC literature, may underestimate TAC values because antioxidant
compounds at the extremities of the lipophilic or hydrophilic
scale are incompletely extracted. In our case, food extracts

obtained with two different solvents were analyzed separately

and their sum reported in the tables. The approach of summing values of lipophilic and hydrophilic extracts permits the
inclusion of the different contributors to the TAC of the food.
However, it cannot be excluded that there may be a synergistic
interaction between water and lipid-soluble antioxidants that
is not evaluated by simply summing the components.
Total antioxidant capacity of vegetables. The values of
TAC of vegetables and the ranking order for each assay are
shown in Table 1. In examining results of the FRAP and the
TEAC assays, spinach was the vegetable with the greatest
antioxidant capacity, followed by peppers (red bell for TEAC
and chili pepper for FRAP assay), whereas cucumber and
endive exhibited the lowest TAC values for the FRAP and
TEAC assays, respectively. In the case of the TRAP assay, the
highest TAC value was found for asparagus, whereas the TAC
values of zucchini and cucumber were not detectable. The
high TAC value assigned to spinach, when analyzed by the
FRAP and TEAC assays, is in agreement with Proteggente et

Ferric reducing-antioxidant power (FRAP), total radical-trapping antioxidant parameter (TRAP) and Trolox equivalent antioxidant
capacity (TEAC) of vegetable extracts1,2
FRAP
Vegetable

Value

TRAP
Rank

Value

(mmol Fe2/kg FW3)

Artichoke
Arugula
Asparagus
Avocado
Beet
Beetroot (red cooked)
Broccoli
Cabbage (green)
Cabbage (Savoy)
Carrot
Cauliflower
Celery
Chicory
Cucumber
Eggplant
Endive
Fennel
Green bean
Leek
Lettuce (green)
Mushroom
Onion (yellow)
Pepper (chili)
Pepper (red bell)
Potato
Pumpkin
Radicchio
Radish (red)
Spinach
Swiss chard (stalk)
Tomato (salad)
Tomato (sauce)
Turnip tops
Zucchini
1
2
3
4

11.09
14.30
10.60
4.90
13.13
15.31
11.67
5.79
8.17
1.06
4.27
1.16
6.72
0.71
3.77
3.24
2.33
2.35
2.15
4.94
16.39
5.28
23.54
20.98
3.67
4.00
11.39
3.77
26.94
11.60
5.12
6.15
17.77
3.33

12
7
13
21
8
6
9
17
14
32
22
31
15
33
24
27
29
28
30
20
5
18
2
3
25
23
11
24
1
10
19
16
4
26

Values are means, n 2.

Values represent the sum of the water- and lipid-soluble extracts.
Fresh weight.
ND, not detectable; NF, not found.

TEAC
Rank

Value

Rank

(mmol Trolox/kg FW)

6.85
4.22
9.71
1.87
2.70
7.67
3.07
2.83
2.35
0.70
1.61
0.47
1.88
ND4
2.82
0.91
0.78
0.65
1.02
2.31
6.26
2.43
6.42
5.47
0.85
NF
6.27
3.62
5.79
2.91
1.31
1.69
6.62
ND

3
10
1
21
16
2
12
14
18
29
23
31
20
32
15
26
28
30
25
19
7
17
5
9
27
33
6
11
8
13
24
22
4
32

1.55
3.55
3.92
2.22
5.21
2.94
3.04
1.15
2.08
0.44
1.10
0.49
1.86
0.43
1.10
0.30
0.43
1.27
0.72
1.33
4.93
1.82
7.62
8.40
0.80
3.71
3.24
2.22
8.49
3.53
1.65
1.47
5.52
2.86

20
9
7
15
5
13
12
24
16
29
25
28
17
30
25
31
30
23
27
22
6
18
3
2
26
8
11
15
1
10
19
21
4
14

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TABLE 1

TOTAL ANTIOXIDANT CAPACITY OF PLANT FOODS

and TEAC the ability to scavenge the ABTS radical cation)

could be the reason for the different results.
Total antioxidant capacity of fruits. The TAC values of
fruits and the ranking order for each assay are shown in Table
2. In general, berries had the greatest antioxidant capacity,
with blackberry being the most effective. Its high antioxidant
capacity, in agreement with the literature (18,25,26), is likely
due to the high content of phenolic acids and flavonoids such
as anthocyanins (27,28), which have demonstrated strong
antioxidant activities in different model systems (9,29,30). In
ranking TAC values of fruits, olives were second in antioxidant capacity, likely due to their high levels of hydroxytyrosol
and tyrosol (31). Citrus fruits exhibited intermediate antioxidant capacity, with oranges as the most effective followed by
grapefruit. This result is in agreement with the higher concentrations of phenolic compounds and vitamin C present in
orange with respect to grapefruit (32). Among the fruits belonging to the Rosaceae family (i.e., plum, apricot, apple, pear
and peach), plums had the highest antioxidant capacity for all
methods. This observation is consistent with the higher phenolic content of plums with respect to other stone fruits (e.g.,
nectarine and peach) described by Gil et al. (33). Moreover,
even if different rankings of antioxidant capacity were reported among these fruits, there is agreement in the literature
that plums are the most effective (14,18,21,34). Finally, as
already reported by other authors (17,18,35), fruits from the
Cucurbitaceae family (i.e., honeydew and cantaloupe melons,
and watermelon) had low TAC values.
Regardless of the assay applied, the data were well correlated. Correlations were better than those described for vegetables, especially for the TRAP assay with the other two assays
(r 0.922, P 0.0001 and r 0.924, P 0.0001 for FRAP
and TEAC assays, respectively). Moreover, as already reported
for vegetables, the best correlation was observed between the
TEAC and FRAP values (r 0.977, P 0.0001).
Total antioxidant capacity of soft beverages. The TAC
values of soft beverages and the ranking order for each assay
are shown in Table 3. Among these, citrus juices had the
highest amount of antioxidants, even though different ranking
orders were obtained from the three assays. The higher antioxidant capacity of citrus juices with respect to other soft
drinks was observed by van der Berg et al. (36). Other fruit
juices had intermediate TAC values; the values varied from
5.01 to 8.76 mmol Fe2/L for the FRAP assay, and from 1.58
to 2.19 mmol Trolox/L and 1.50 to 2.56 mmol Trolox/L for the
TRAP and TEAC assays, respectively. Cola drinks had the
lowest TAC values, which were not detectable with the
TRAP. This result is consistent with the publication of Karakaya et al. (37), who could not detect TAC activity for this
beverage using the spectrophotometric TEAC assay. Finally,
in contrast to the results for vegetables and fruits, the TAC
values of soft beverages from the different assays were not
correlated, with the exception of those obtained by the FRAP
and TEAC assays (r 0.941, P 0.0001). The absence or
presence of correlations among assays when applied to different food items highlights once again the importance of assessing TAC using a battery of different assays so as to obtain a
detailed picture of the phenomenon.
Total antioxidant capacity of alcoholic beverages, teas and
coffees. The TAC of alcoholic and caffeine-rich beverages
and the ranking order for each assay are shown in Table 4.
Among the beverages analyzed, coffee drinks were the most
effective, regardless of the assay applied, with espresso having
the greatest antioxidant capacity. The removal of caffeine
from the espresso coffee led to a decrease in TAC values of
2530%, likely due to the antioxidant capacity of caffeine

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al. (14), whose ranking of TAC in 18 vegetables were similar

to our results. The high antioxidant capacity of spinach is due
to both the water- and lipid-soluble fractions; the former
contains glucuronic acid derivates of flavonoids and derivates
and isomers of p-coumaric acid (16), and the latter is rich in
lutein and chlorophylls (17).
Considering the plant families, the vegetables analyzed in
our study that belonged to the Cucurbitaceae family (i.e.,
pumpkin, zucchini and cucumber) exhibited low TAC values,
in agreement with the literature data (18), with the exception
of pumpkin when analyzed using the TEAC assay. This is not
surprising because this vegetable contains high level of carotenoids (19), especially less lipid-soluble carotenoids such as
lutein, which had high antioxidant efficiency in the TEAC
assay (20). The vegetables analyzed from the Liliaceae family
(i.e., yellow onion and leek) had different TAC values: onion,
in particular, exhibited an intermediate TAC value, whereas
leek had a low antioxidant capacity, regardless of the method
applied. The difference observed between these two vegetables
is consistent with the data of Proteggente et al. (14), whereas
Szeto et al. (21) using the FRAP assay found that onion was
quite high in antioxidants. The antioxidant capacity varied
considerably for the Brassicaceae family of vegetables; turnip
tops had the highest TAC values for all methods, broccoli and
cabbages exhibited intermediate capacity, whereas the TAC
value of cauliflower was low. This large variability of TAC
values observed in this plant family is in agreement with the
literature (14,18,22). Vegetables from the Solanaceae family
(i.e., eggplant, tomato and potato) had low antioxidant values,
as described in the literature (14,17,18,2123). This is surprising for tomato, which is a rich source of lycopene, a carotene
that can to quench different reactive species, including singlet
oxygen. A possible reason might be that all the three assays
employed (TEAC, TRAP and FRAP) as well as the majority
of the assays present in the literature, do not utilize singlet
oxygen as the oxidizing system. However, because the TEAC
assay is able to recognize at least in part the antioxidant
properties of all carotenoids, which indeed demonstrate high
antioxidant activities with this assay (20), another possible
explanation for the low results obtained in tomatoes is incomplete extraction. Indeed, in the case of tomato, carotenoids are
present in the chromoplasts in crystalline form, and this might
reduce the extraction capacity of acetone, the only liposoluble
solvent compatible with the assays.
Finally, among salads, endive had the lowest antioxidant
capacity, whereas radicchio exhibited a relatively high antioxidant capacity regardless of the applied method. This high
antioxidant capacity of radicchio is consistent with the publication of Papetti et al. (24) and is possibly due to the
presence of anthocyanins in the leaf.
In contrast to Ou et al. (22), who analyzing different
varieties of vegetables using the FRAP and the oxygen radical
absorption capacity (ORAC) assay, and did not find agreement among methods for the vegetables analyzed, with the
exception of a few, our data were well correlated among assays.
The data obtained using the FRAP and TEAC assays had the
best correlation coefficients (r 0.917, P 0.0001), whereas
the TEAC and TRAP values (r 0.656, P 0.0001) and
TRAP and FRAP values (r 0.744, P 0.0001) were less
well correlated. The differences observed in the ranking order
between the TRAP and TEAC values of some vegetables
could be the results of different variables, such as the sensitivity of the assays and the ability of TEAC assay, with respect to
TRAP, to measure the antioxidant capacity of lipid-soluble
antioxidants. Moreover, the different information furnished by
the two assays (TRAP measures the chain breaking potential

2815

PELLEGRINI ET AL.

2816

TABLE 2
Ferric reducing-antioxidant power (FRAP), total radical-trapping antioxidant parameter (TRAP) and Trolox equivalent antioxidant
capacity (TEAC) of fruit extracts1,2
FRAP
Fruit

Value

TRAP
Rank

Value

(mmol Fe2/kg FW3)

3.84
3.23
4.02
2.28
51.53
18.61
8.10
8.88
5.82
11.09
3.25
10.20
7.41
2.70
5.73
2.27
39.99
24.59
20.50
6.57
5.00
15.73
12.79
6.97
43.03
44.86
22.74
28.00
9.60
1.13

24
26
23
28
1
9
16
15
20
12
25
13
17
27
21
29
4
6
8
19
22
10
11
18
3
2
7
5
14
30

Rank

Value

Rank

(mmol Trolox/kg FW)

2.23
1.54
2.29
1.05
21.01
9.30
4.17
2.74
2.06
2.50
1.59
4.04
2.30
1.73
0.95
1.12
18.08
14.64
5.65
1.49
3.87
5.92
8.09
2.06
10.48
12.14
8.56
10.34
2.76
0.46

20
24
19
27
1
7
12
16
21
17
23
13
18
22
28
26
2
3
11
25
14
10
9
21
5
4
8
6
15
29

1.59
1.31
1.44
0.64
20.24
7.43
2.69
3.10
2.47
3.85
2.48
3.05
2.28
0.75
1.20
0.65
14.73
10.43
8.74
1.67
2.19
9.91
5.11
1.46
16.79
14.05
10.94
11.34
4.16
0.69

22
25
24
30
1
10
16
14
18
13
17
15
19
27
26
29
3
7
9
21
20
8
11
23
2
4
6
5
12
28

1 Values are means, n 2.

2 Values represent the sum of the water- and lipid-soluble extracts.
3 Fresh weight.

(38). During coffee making, the roasting process leads to

profound changes in the chemical composition and biological
activities of the coffee bean, resulting in the generation of
compounds derived from the Maillard reaction, carbohydrate
caramelization and pyrolysis of organic compounds (39). In
roasted coffee most polyphenolic compounds are destroyed,
but Maillard reaction products with antioxidant properties are
generated (40), resulting in an increased antioxidant activity
in the -carotene-linoleic acid model system (39).
In teas, the antioxidant capacity of green tea is considerably
higher than that of black tea, according to the literature
(41,42). This different behavior of teas is due to the changes
occurring during the process of fermentation; the flavanols in
green tea leaves (mainly catechins and their gallic esters)
undergo an oxidative polymerization by polyphenol oxidase,
which turns the leaves black. During oxidation, much of the
catechin content of green tea is converted to oxyproducts,
such as thearubingens and theaflavins, with a loss of antioxidant capacity (42).
Among alcoholic beverages, red wines had the most antioxidant capacity followed by rose and white wines, in agreement with the literature (43 46). This is not surprising because phenolic compounds in wine derive mainly from the
skin, seeds and stems of grapes, making them important
sources of the polyphenols that are transferred to the juice at

the first stage of wine fermentation. Thus, the content of

polyphenols is high in red wine in which the contact between
juice and pomace is prolonged; it is intermediate in rose wine
in which the contact is reduced compared with red wine and
relatively low in white wine, which is usually made from the
free-running juice without contact with the grape skins. The
TEAC values of red and white wines were in the same range
of those described by Simonetti et al. (43), whereas, in the
case of red wines, other authors reported TEAC values slightly
lower than ours (45). These differences could be due to the
winemaking procedure as well as to the grape variety and
growing conditions.
The TAC values of distilled spirits (i.e., cognac, grappa,
rum and whiskey) were lower than those of wines, with the
exception of whiskey; for the TRAP and TEAC assays, whiskey outranked some white and rose wines. This higher antioxidant capacity of whiskey with respect to other distilled
spirits could be due to its higher content of phenols and furans,
as described by Goldberg et al. (47). Among distilled spirits,
cognac followed whiskey in the ranking order obtained for all
of the assays applied. This is likely due to polyphenols extracted from wood during the aging process (47). Finally,
grappa and rum, distilled from grape pomace and molasses,
respectively, but not aged in wood, had very low TEAC values
with undetectable FRAP and TRAP values.

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Apple (red Delicious)

Apple (yellow Golden)
Apricot
Banana
Blackberry
Blueberry
Cherry
Clementine
Fig
Grape (black)
Grape (white)
Grapefruit (yellow)
Kiwi fruit
Loquat
Melon (cantaloupe)
Melon (honeydew)
Olive (black)
Olive (green)
Orange
Peach (yellow)
Pear
Pineapple
Plum (red)
Prickly pear
Raspberry
Redcurrant
Strawberry (cultivated)
Strawberry (wild)
Tangerine
Watermelon

TEAC

TOTAL ANTIOXIDANT CAPACITY OF PLANT FOODS

2817

TABLE 3
Ferric reducing antioxidant power (FRAP), total radical-trapping antioxidant parameter (TRAP) and Trolox equivalent antioxidant
capacity (TEAC) of soft beverages1
FRAP
Beverage

TRAP

Value

Rank

Value

TEAC
Rank

(mmol Fe2/L)
Apple juice
Apricot juice
Cola
Grapefruit juice
Lemon iced tea
Lemon juice
Mixed fruit juice
Orange juice
Peach juice
Pear juice
Pineapple juice
Tropical juice

5.01
7.15
0.92
8.22
7.43
8.37
8.76
9.44
7.79
7.43
5.16
6.18

Value

Rank

1.83
2.48
0.09
3.30
2.28
2.21
2.65
3.02
2.51
2.56
1.50
1.97

10
6
12
1
7
8
3
2
5
4
11
9

(mmol Trolox/L)
10
7
11
4
6
3
2
1
5
6
9
8

1.97
2.19
ND2
2.65
1.92
3.67
1.58
2.27
1.81
2.15
1.79
2.06

7
4
12
2
8
1
11
3
9
5
10
6

The TAC values of alcoholic beverages, teas and coffees

obtained by different assays were well correlated (TRAP vs
FRAP: r 0.993, P 0.0001; TEAC vs FRAP: r 0.997, P
0.0001; TEAC vs TRAP: r 0.993, P 0.0001). This is in
keeping with Schlesier et al. (48) who analyzed some beverages, including different teas, and observed strong correlations
between assays based on different chemistry (TEAC and
DPPH).

Total antioxidant capacity of oils. To assess the total

antioxidant capacity of oils, samples were directly diluted in
n-hexane; this procedure allows the contribution of all antioxidant compounds present, such as phenolic compounds,
tocopherols, and carotenoids, to be considered (49). However,
the use of this solvent limits the assessment of TAC of oils to
the TEAC assay, because it is not compatible with the TRAP
and FRAP assays. In Table 5, the TEAC values of the analyzed

TABLE 4
Ferric reducing antioxidant power (FRAP), total radical-trapping antioxidant parameter (TRAP) and Trolox equivalent antioxidant
capacity (TEAC) of alcoholic beverages, teas and coffees1
FRAP
Beverage

Value

TRAP
Rank

Value

TEAC
Rank

(mmol Fe2/L)
Beer (lager)
Chamomile
Coffee (espresso)
Coffee (espresso, decaffeinated)
Coffee (extracted)
Coffee (soluble)
Cognac
Grappa
Rum
Tea (black)
Tea (green)
Vinegar (red)
Whiskey
Wine (Aglianico, red)
Wine (Chianti, red)
Wine (Sauvignon, red)
Wine (Villa Torre, rose )
Wine (Tamerici, rose )
Wine (Bardolino, rose )
Wine (Vernaccia, white)
Wine (Pinot, white)
Wine (Greco di Tufo, white)
1 Values are means, n 2.
2 NF, not found; ND, not detectable.

2.78
0.65
129.38
93.01
96.40
108.56
2.25
ND
ND
10.09
18.00
9.50
3.45
30.53
31.53
23.90
8.33
7.22
4.66
5.04
3.72
3.83

Value

Rank

(mmol Trolox/L)
18
20
1
4
3
2
19
21
21
9
8
10
17
6
5
7
11
12
14
13
16
15

NF2
1.26
66.00
45.82
59.57
52.37
1.46
ND
ND
4.87
7.63
4.80
2.31
16.09
14.84
11.73
2.24
3.20
1.98
2.32
2.10
1.86

22
19
1
4
2
3
18
21
21
9
8
10
13
5
6
7
14
11
16
12
15
17

1.04
0.61
36.54
26.96
30.29
32.48
1.29
0.18
0.04
3.60
6.01
3.12
1.68
12.14
11.43
8.95
2.42
2.18
1.52
1.94
1.68
1.61

18
19
1
4
3
2
17
20
21
9
8
10
14
5
6
7
11
12
16
13
14
15

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1 Values are means, n 2.

2 ND, not detectable.

PELLEGRINI ET AL.

2818

LITERATURE CITED

Trolox equivalent antioxidant capacity (TEAC) of oils1

1. Rimm, E. B., Aschiero, A., Giovannucci, E., Spiegelman, D., Stampfer,

M. J. & Willett, W. C. (1996) Vegetables, fruit, and cereal fiber intake and risk
of coronary heart disease among men. J. Am. Med. Assoc. 275: 447 451.
2. Gillman, M. W., Cupples, L. A., Gagnon, D., Posner, B. M., Ellison, R. C.,
Castelli, W. P. & Wolf, P. A. (1995) Protective effect of fruits and vegetables on
development of stroke in men. J. Am. Med. Assoc. 273: 11131117.
3. La Vecchia, C., Altieri, A. & Tavani, A. (2001) Vegetables, fruit, antioxidants and cancer: a review of Italian studies. Eur. J. Nutr. 40: 261267.
4. Terry, P., Terry, J. B. & Wolk, A. (2001) Fruit and vegetable consumption in the prevention of cancer: an update. J. Intern. Med. 250: 280 290.
5. Cohen, J. H., Kristal, A. R. & Stanford, J. L. (2000) Fruit and vegetable
intakes and prostate cancer risk. J. Natl. Cancer Inst. 92: 61 68.
6. Gey, K. F. (1990) The antioxidant hypothesis of cardiovascular disease: epidemiology and mechanisms. Biochem. Soc. Trans. 18: 10411045.
7. Gey, K. F., Puska, P., Jordan, P. & Moser, U. K. (1991) Total antioxidant capacity of plant foods. Inverse correlation between plasma vitamin E and
mortality from ischemic heart disease in cross-cultural epidemiology. Am. J. Clin.
Nutr. 53: 326S334S.
8. Willett, W. C. (1991) Micronutrients and cancer risk. Am. J. Nutr. Clin.
53: 265S269S.
9. Wang, H., Cao, G. & Prior, R. L. (1997) Oxygen radical absorbing
capacity of anthocyanins. J. Agric. Food Chem. 45: 304 309.
10. Pellegrini, N., Simonetti, P., Gardana, C., Brenna, O., Brighenti, F. &
Pietta, P. G. (2000) Polyphenol content and total antioxidant activity of vini
novelli (young red wines). J. Agric. Food Chem. 48: 732735.
11. Benzie, I.F.F. & Strain, J. J. (1999) Ferric reducing antioxidant power
assay: direct measure of total antioxidant activity of biological fluids and modified
version for simultaneous measurement of total antioxidant power and ascorbic
acid concentration. Methods Enzymol. 299: 1527.
12. Pellegrini, N., Re, R., Yang, M. & Rice-Evans, C. A. (1999) Screening
of dietary carotenoids and carotenoid-rich fruit extracts for antioxidant activities
applying the 2, 2-azobis(3-ethylenebenzothiazoline-6-sulfonic) acid radical cation decolorization assay. Methods Enzymol. 299: 379 389.
13. Ghiselli, A., Serafini, M., Maiani, G., Azzini, E. & Ferro-Luzzi, A. (1995)
A fluorescence-based method for measuring total plasma antioxidant capability.
Free Radic. Biol. Med. 18: 29 36.
14. Proteggente, A. R., Pannala, A. S., Paganga, G., van Buren, L., Wagner,
E., Wiseman, S., van de Put, F., Dacombe, C. & Rice-Evans, C. A. (2002) The
antioxidant activity of regularly consumed fruit and vegetables reflects their
phenolic and vitamin C composition. Free Radic. Res. 36: 217233.
15. Pellegrini, N., Del Rio, D., Colombi, B., Bianchi, M. & Brighenti, F. (2003)
Application of the 2, 2-azobis(3-ethylenebenzothiazoline-6-sulfonic acid) radical
cation assay to a flow injection system for the evaluation of antioxidant activity of
some pure compounds and beverages. J. Agric. Food Chem. 51: 260 264.
16. Bergman, M., Varshavsky, L., Gottlieb, H. E. & Grossman, S. (2001)
The antioxidant activity of aqueous spinach extract: chemical identification of
active fractions. Phytochemistry 58: 143152.
17. Buratti, S., Pellegrini, N., Brenna, O. V. & Mannino, S. (2001) Rapid
electrochemical method for the evaluation of the antioxidant power of some
lipophilic food extracts. J. Agric. Food Chem. 49: 5136 5141.
18. Halvorsen, B. L., Holte, K., Myhrstad, M.C.W., Barikmo, I., Hvattum, E.,
Fagertun Remberg, S., Wold, A. B., Haffner, K., Baugerod, H., Frost Andersen, L.,
Moskaug, J. O., Jacobs, D. R. & Blomhoff, R. (2002) A systematic screening of
total antioxidants in dietary plants. J. Nutr. 132: 461 471.
19. Muller, H. (1997) Determination of the carotenoid content in selected
vegetables and fruit by HPLC and photodiode array detection. Z. Lebensm.
Unters Forsch A 204: 88 94.
20. Miller, N. J., Sampson, J., Candeias, L. P., Bramley, P. M. & Rice-Evans,
C. A. (1996) Antioxidant activities of carotenes and xanthophylls. FEBS Lett.
384: 240 242.
21. Szeto, Y. T., Tomlinson, B. & Benzie, I.F.F. (2002) Total antioxidant
and ascorbic acid content of fresh fruits and vegetables: implication for dietary
planning and food preservation. Br. J. Nutr. 87: 5559.
22. Ou, B., Haung, D., Hampsch-Woodill, M., Flanagan, J. A. & Deemer, E. K.
(2002) Analysis of antioxidant activities of common vegetables employing oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power
(FRAP) assays: a comparative study. J. Agric. Food Chem. 50: 31223128.
23. Vinson, J. A., Hao, Y., Su, X. & Zubik, L. (1998) Phenol antioxidant
quantity and quality in foods: vegetables. J. Agric. Food Chem. 46: 3630 3634.
24. Papetti, A., Daglia, M. & Gazzani, G. (2002) Anti- and pro-oxidant
water soluble activity of Cichorium genus vegetables and effect of thermal treatment. J. Agric. Food Chem. 50: 4696 4704.
25. Ka hko nen, M. P., Hopia, A. I., Vuorela, H. J., Rauha, J. P., Pihlaja, K.,
Kujala, T. S. & Heinonen, M. (1999) Antioxidant activity of plant extracts
containing phenolic compounds. J. Agric. Food Chem. 47: 3954 3962.
26. Kalt, W., Forney, C. F., Martin, A. & Prior, R. L. (1999) Antioxidant
capacity, vitamin C, phenolics, and anthocyanins after fresh storage of small
fruits. J. Agric. Food Chem. 47: 4638 4644.
27. Macheix, J. J., Fleuriet, A. & Billot, J. (1990) Fruit Phenolics. CRC
Press, Boca Raton, FL.
28. Ha kkinen, S., Heinonen, M., Karenlampi, S., Mykkanen, H., Ruuskanen, J.
& Torronen, R. (1999) Screening of selected flavonoids and phenolic acids in
19 berries. Food Res. Int. 32: 345353.
29. Rice-Evans, C. A., Miller, N. J., Bolwell, P. G., Bramley, P. M. & Pridham,

TEAC
Oil

Value

Rank

(mmol Trolox/kg)
Corn
Extra virgin olive
Olive
Peanut
Soybean
Sunflower

1.29
1.79
0.63
0.61
2.20
1.17

3
2
5
6
1
4

1 Values are means, n 2.

oils are shown. Soybean oil had the greatest antioxidant capacity, likely due to its high tocopherol content (50), whereas
peanut oil was less effective. As already observed (51,52), the
TEAC value of extra virgin olive oil (EVOO) was higher than
that of olive oil (OO). The difference between OO and
EVOO arises from the different manufacturing processes (49),
leading to differences in the antioxidant composition. In particular, EVOO (obtained by cold pressure of the olive paste) is
much richer in phenolic compounds than refined oils (obtained by solvent extraction), which are virtually devoid of
phenols. OO is a vaguely defined mixture of refined olive oil
and EVOO in which the amount of EVOO may vary from 33
to 95% (53), thus affecting the amount of antioxidants
present.
The validity of the TAC approach for investigating the role
of antioxidants in the protective effect of plant food is growing. Our study did not pretend to explain components of
variance (such as cultivars, sun exposure, water availability,
organic agricultural practices, seasonality, food processing, domestic cooking or food combination) which, although important for characterization of a specific food, are descriptors that
are normally ignored in dietary surveys in which the consumption of a large number of food items is recorded.
The relevance of TAC as a new tool for investigating the
relationship between dietary antioxidants and oxidative
stressinduced pathologies seems confirmed by the data from a
recent population-based case-control study, which showed an
inverse correlation between the diet TAC and the risk of both
cardia and distal gastric cancer (54). These relationships
emerged in spite of the use of a very incomplete database of
total antioxidant potential (12 items only among vegetables
and fruits), highlighting the potentiality of the TAC as a
descriptor of the diet.
In summary, the total antioxidant activities of 64 foods, 34
beverages and 6 oils were measured by three different methods.
Such data, integrated with further food items of different
classes (i.e., cereals, pulses and nuts), will result in a complete
and versatile database of total antioxidant capacity. Coupled
with an appropriate questionnaire, this will allow the evaluation of the overall intake of antioxidant-equivalents in selected groups of the Italian population in relation to the
incidence of oxidative stressinduced diseases.
ACKNOWLEDGMENT
We thank Valeria Pala from the Epidemiology Unit of the National Cancer Institute (Milan, Italy) for the elaboration of food
consumption data.

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TABLE 5

TOTAL ANTIOXIDANT CAPACITY OF PLANT FOODS

42. Benzie, I.F.F. & Szeto, Y. T. (1999) Total antioxidant capacity of teas
by ferric reducing/antioxidant power assay. J. Agric. Food Chem. 47: 633 636.
43. Simonetti, P., Pietta, P. G. & Testolin, G. (1997) Polyphenol content
and total antioxidant activity potential of selected Italian wines. J. Agric. Food
Chem. 45: 11521155.
44. Mannino, S., Brenna, O., Buratti, S. & Cosio, M. S. (1998) A new
method for the evaluation of the antioxidant power of wines. Electroanalysis 10:
908 912.
45. Fogliano, V., Verde, V., Randazzo, G. & Ritieni, A. (1999) Method for
measuring antioxidant activity and its application to monitoring the antioxidant
capacity of wines. J. Agric. Food Chem. 47: 10351040.
46. Campos, A. M. & Lissi, E. A. (1996) Total antioxidant potential of
Chilean wines. Nutr. Res. 16: 385389.
47. Goldberg, D. M., Hoffman, B., Yang, J. & Soleas, G. J. (1999) Phenolic
constituents, furans, and total antioxidant status of distilled spirits. J. Agric. Food
Chem. 47: 3978 3985.
48. Schlesier, K., Harwat, M., Bohm, V. & Bitsch, R. (2002) Assessment of
antioxidant activity by using different in vitro methods. Free Radic. Res. 36:
177187.
49. Boskou, D. (1996) Olive oil composition. In: Olive Oil: Chemistry and
Technology (Boskou, D., ed.), pp. 52 83. AOCS Press, Champaign, IL.
50. Rovellini, P., Azzolini, M. & Cortesi, N. (1997) Tocoferoli e tocotrienoli in
oli e grassi vegetali mediante HPLC. Riv. Ital. Sostanze Grasse LXXIV: 15.
51. Pellegrini, N., Visioli, F., Buratti, S. & Brighenti, F. (2001) Direct analysis of total antioxidant activity of olive oil and studies on the influence of heating.
J. Agric. Food Chem. 49: 25322538.
52. Mannino, S., Buratti, S., Cosio, M. S. & Pellegrini, N. (1999) Evaluation
of the antioxidant power of olive oils based on a FIA system with amperometric
detection. Analyst 124: 11151118.
53. Andrikopoulos, N. K., Hassapidou, M. N. & Manoukas, A. G. (1989)
The tocopherol content of Greek olive oils. J. Sci. Food Agric. 46: 503509.
54. Serafini, M., Bellocco, R., Wolk, A. & Ekstrom, A. M. (2002) Total
antioxidant potential of fruit and vegetables and risk of gastric cancer. Gastroenterology 123: 98599.

Downloaded from jn.nutrition.org by guest on April 27, 2015

J. B. (1995) The relative antioxidant activities of plant-derived polyphenolic

flavonoids. Free Radic. Res. 22: 375383.
30. Satue-Gracia, M. T., Heinonen, M. & Frankel, E. N. (1997) Antioxidant
activity of anthocyanins in LDL and lecithin liposome system. J. Agric. Food
Chem. 45: 33623367.
31. Ryan, D. & Robards, K. (1998) Phenolic compounds in olives. Analyst
123: 31R 44R.
32. Gorinstein, S., Martin-Belloso, O., Park, Y. S., Haruenkit, R., Lojek, A.,
Ciz, M., Caspi, A., Libman, I. & Trakhtenberg, S. (2001) Comparison of some
biochemical characteristics of different citrus fruits. Food Chem. 74: 309 315.
33. Gil, M. I., Tomas-Barberan, F. A., Hess-Pierce, B. & Kader, A. A. (2002)
Antioxidant capacities, phenolic compounds, carotenoids, and vitamin C contents of nectarine, peach, and plum cultivars from California. J. Agric. Food Chem.
50: 4976 4982.
34. Wang, H., Cao, G. & Prior, R. L. (1996) Total antioxidant capacity of
fruits. J. Agric. Food Chem. 44: 701705.
35. Leong, L. P. & Shui, G. (2002) An investigation of antioxidant capacity
of fruits in Singapore markets. Food Chem. 76: 69 75.
36. van den Berg, R., Haenen, G. R. M. M., van den Berg, H. & Bast, A.
(1999) Applicability of an improved Trolox equivalent antioxidant capacity
(TEAC) assay for evaluation of antioxidant capacity measurements of mixtures.
Food Chem. 66: 511517.
37. Karakaya, S., El, S. N. & Tas, A. A. (2001) Antioxidant activity of some
foods containing phenolic compounds. Int. J. Food Sci. Nutr. 52: 501508.
38. Shi, X., Dalal, N. S. & Jain, A. C. (1991) Antioxidant behaviour of
caffeine: efficient scavenging of hydroxyl radicals. Food Chem. Toxicol. 29: 1 6.
39. Daglia, M., Papetti, A., Gregotti, C., Berte , F. & Gazzani, G. (2000) In
vitro antioxidant and ex vivo protective activities of green and roasted coffee. J.
Agric. Food Chem. 48: 1449 1454.
40. Tubaro, F., Micossi, E. & Ursini, F. (1996) The antioxidant capacity of
complex mixture by kinetic analysis of crocin bleaching inhibition. J. Am. Oil
Chem. Soc. 73: 173179.
41. Richelle, M., Tavazzi, I. & Offord, E. (2001) Comparison of the antioxidant activity of commonly consumed polyphenolic beverages (coffee, cocoa, and
tea) prepared per cup serving. J. Agric. Food Chem. 49: 3438 3442.

2819