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ANTHOCYANIN PROFILE AND ANTIOXIDANT ACTIVITY

FROM 24 GRAPE VARIETIES CULTIVATED
IN TWO PORTUGUESE WINE REGIONS

Elisa CoSTA1, Fernanda CoSME2, António M. JorDÃo1* and Arlete MEnDES-FAIA2

1: Polytechnic Institute of Viseu (Centre for the Study of Education, Technologies and Health),
Agrarian Higher School, Estrada de nelas, Quinta da Alagoa, ranhados, 3500-606 Viseu, Portugal
2: Institute for Biotechnology and Bioengineering – Centre of Genomics and Biotechnology (IBB-CGB-UTAD),
University of Trás-os-Montes and Alto Douro, School of Life Science and Environment,
5001-801 Vila real, Portugal

Abstract Résumé
Aims: The purpose of this work was to evaluate the general Objectifs : Le but de ce travail était d’évaluer la
phenolic composition and the anthocyanin profile of composition phénolique générale et le profil d’anthocyanes
24 grape varieties from two Portuguese wine regions as well de 24 cépages de deux régions viticoles portugaises ainsi
as their antioxidant activity in the different grape berry que leur activité antioxydante dans les différentes fractions
fractions (skins, pulps and seeds). de baie de raisin (pellicules, pulpes et pépins).

Methods and results: Individual anthocyanin composition Méthodes et résultats : Les anthocyanes obtenues à partir
of grape skin extracts was analyzed by HPLC, whereas total d’extraits de pellicules de raisin ont été analysées par HPLC
antioxidant activity was evaluated by two methods: DPPH tandis que l’activité antioxydante totale a été évaluée par
and ABTS. In general, a high variability was found among deux méthodes : DPPH et ABTS. En général, une grande
the different autochthonous and non-autochthonous grape variabilité a été observée parmi les différentes variétés de
varieties in relation to the polyphenolic compounds raisins autochtones et non autochtones en ce qui concerne
analyzed, especially the individual anthocyanins. The les composés polyphénoliques analysés comme les
individual anthocyanins in grape skin extracts were mainly anthocyanes. Dans les extraits de pellicules de raisin, les
malvidin (1.40-7.09 mg/g of skin), in particular malvidin-3- anthocyanes étaient principalement la malvidine (1.40-
glucoside (0.62-6.09 mg/g of skin). The highest antioxidant 7.09 mg/g de pellicule), et en particulier la malvidine-3-
activity was consistently detected in the seed extracts; glucoside (0.62-6.09 mg/g de pellicule). Dans tous les
however, it was not possible to establish a clear difference cépages étudiés, l’activité antioxydante la plus élevée a été
among the grape varieties analyzed. mesurée dans les pépins. Par contre, il n’a pas été possible
d’établir une différence claire entre les cépages.
Conclusion: High variability in polyphenolic content,
individual anthocyanin composition and antioxidant activity Conclusion : Une grande variabilité a été trouvée parmi les
was found among the diverse autochthonous and non- divers cépages autochtones et non-autochtones étudiés par
autochthonous grape varieties studied. Seeds showed the rapport à la teneur en polyphénols, ainsi que la composition
highest antioxidant activity, followed by skin and pulp, en anthocyanes et l’activité antioxydante. Les pépins étaient
irrespective of the grape variety. la fraction de la baie de raisin à l’activité antioxydante la
plus élevée suivie par les peaux et les pulpes, quel que soit
Significance and impact of the study: Most vineyards in le cépage.
Portugal grow Portuguese cultivars of Vitis Vinifera L. and
other cultivars grown worldwide. The phenolic compounds Signification et impact de l’étude : Au Portugal, la plupart
and antioxidant activity of these grape cultivars have never des vignobles possèdent des variétés portugaises de Vitis
been characterized under the environmental conditions of vinifera L. et d’autres espèces largement cultivées dans le
the Douro and Dão regions. The variability in phenolic monde. L’évaluation et la comparaison des composés
content among the grape varieties studied confirms the phénoliques et de l’activité antioxydante de ces cépages
hypothesis that genetic factors have an important role in the n’ont pas été exploitées, à savoir sous les conditions
biosynthesis of these compounds and, consequently, in the environnementales des régions du Douro et du Dão. La
antioxidant activity of grapes. variabilité observée dans le contenu phénolique des cépages
étudiés confirme l’hypothèse que des facteurs génétiques
Key words: antioxidant activity, individual anthocyanins, jouent un rôle important dans la biosynthèse de ces
red grapes, pulp, seed, skin composés et, par conséquent, sur l’activité antioxydante du
raisin.
Mots clés : activité antioxydante, anthocyanes, cépage
rouge, pulpe, pépin, pellicule

manuscript received 4th January 2013 - revised manuscript received 8th August 2013

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INTRODUCTION been conflicting evidence regarding the contribution

of anthocyanins to red grape and wine antioxidant
Grapes are the chief dietary sources of anthocyanins properties.
(rechkemmer and Pool-Zobel, 1996). Anthocyanins
are mainly localized in berry skin (Jordão et al., Dão and Douro are two important wine-producing
1998a), where they are found at concentrations of 200 regions in northeastern Portugal. The Dão region is
to 5,000 mg/Kg of fresh grapes (ribéreau-Gayon, sheltered on nearly all sides by high granite mountains
1982). Anthocyanin pigments are of noticeable and is situated in the transition zone between maritime
importance in grapes and wines because of their dual and continental climate, with abundant rainfall in the
role: first, they constitute an essential part of the winter months and long warm dry summers leading up
sensory attributes, as their concentration, different to harvest. The Douro region, which is situated around
forms and derivatives directly affect the color of the the Douro river basin, is sheltered from Atlantic
final wine and second, they are considered to have winds by several mountains and has a continental
diverse biological properties and therefore are climate, with hot and dry summers and cold winters.
regarded as secondary metabolites with potential Although Portuguese Vitis vinifera L. autochthonous
nutritional value. Grape anthocyanins are the 3-O- cultivars are the most frequently used cultivars for the
monoglucosides of delphinidin, cyanidin, petunidin, production of high quality red wines in both regions,
peonidin, and malvidin. Glucosylated derivatives of other varieties from other countries (especially from
these anthocyanins, esterified at the C6 position of France) have been introduced in the past few years.
glucose with acetyl or coumaroyl groups, have also
been detected, yet generally at low concentrations. A few studies have been performed on the anthocyanin
Malvidin is the predominant monomeric anthocyanin profile of Portuguese grape varieties. However, these
in grapes and is the reddest of all anthocyanins, studies were restricted to a small number of varieties
providing the characteristic color of young red wines. from southern (Dallas and Laureano, 1994a; Jordão et
al., 1998a) and northern Portugal (Dallas and
Anthocyanin concentration and profile is influenced Laureano, 1994a; Dopico-García et al., 2008; Mateus
by several factors such as varietal diversity et al., 2002) and did not address the antioxidant
(Kallithraka et al., 2005), soil type (Yokotsuka et al., activity of the different grape berry fractions.
1999), environmental conditions (Jackson and
Lombard, 1993; Mateus et al., 2002), vineyard Thus, the purpose of this work was to evaluate the
management (Jordão et al., 1998b), and grape anthocyanin profile in grape skin as well the
maturity (Jordão et al., 1998a). Additionally, some antioxidant activity in different grape berry fractions
grape varieties can have a high anthocyanin (skins, pulps and seeds) of 24 grape varieties from
concentration but a low extractability index (romero- both Portuguese wine regions. The results obtained in
Cascales et al., 2005). this work could be useful for selecting winemaking
techniques in order to improve anthocyanin extraction
According to several studies, flavan-3-ols, flavonols and hence increase the antioxidant capacity of the
and anthocyanins are the most important compounds wines produced in these two regions. Finally, we
that contribute to red grape and wine antioxidant should note that this is the first report of the
proprieties (Beecher, 2003; Simonetti et al., 1997). anthocyanin content and antioxidant activity of grape
For rivero-Pérez et al. (2008), the free anthocyanin cultivars from the Dão and Douro wine regions.
fraction is mainly responsible for the total antioxidant
capacity and scavenger activity in red wines. MATERIALS AND METHODS
However, other authors have reported that there is no 1. Samples
correlation between antioxidant activity and spectral
anthocyanin content in grapes and finished wines In 2010, 24 grape varieties (Vitis vinifera L.) were
(Arnous et al., 2002; Kallithraka et al., 2005). For harvested at technological maturity in good sanitary
example, considering some individual anthocyanins conditions from 6-year-old grapevines in two
(cyanidin-3-glucoside, peonidin-3-glucoside and experimental vineyards of northeastern Portugal (one
malvidin-3-glucoside) from Italian wines, Di Majo et in the Douro region and the other in the Dão region).
al. (2008) reported low correlations between these Grape berry samples (samples of 200 berries in
individual compounds and their antioxidant capacity duplicate) were picked randomly from 20 different
(ranging from 0.29 to 0.54). In addition, Jordão et al. vine plants in all possible conditions (e.g., different
(2012) reported differences in the correlation values height and exposure to sunlight). All grape samples
between individual anthocyanins and antioxidant were kept frozen at -20 ºC until processing. The grape
activity during the maceration process. Thus, there has varieties studied are listed in Table 1.

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Table 1. Grape variety and origin 3. Sample preparation for antioxidant activity and
of the grape samples studied. individual anthocyanin composition analysis
Skins, pulps and seeds were removed manually from
the berries, washed separately several times with
Autochthonous/

distilled water, and dried on blotting paper. Before the

Grape variety Non-autochthonous Origin
!
extraction process, seeds were crushed. The extraction
grape variety

process followed the procedure described by Sun et

Camarate! AG! Dão!

al. (1996): 50 mL of 80 % methanol followed by

Gewürztraminer*! NAG! Dão!

50 mL of 75 % acetone, each for 3 h with agitation.

Monvedro! AG! Dão!

After clarification of the suspension by centrifugation

Moreto Boal! AG! Dão!

(10 min at 3500 rpm), all the extracts were combined.

Negro Mole! AG! Dão!

Aliquots of each extract were filtered (Whatman

Negro Mouro! AG! Dão!
Alfrocheiro! AG! Douro!
0.45 µm) and frozen at -20 ºC until processing for
Alvarilhão! AG! Douro!

antioxidant activity (skin, pulp and seed extracts) and

Aramon! NAG! Douro!

individual anthocyanin composition (skin extracts)

Bastardo! AG! Douro!

analysis. Extractions were done in duplicate for each

Cabernet Franc! NAG! Douro!

grape berry fraction.

Carignan Noir! NAG! Douro!
Cornifesto! AG! Douro!

4. Antioxidant activity determination

Gamay! NAG! Douro!
Grenache! NAG! Douro!

The total antioxidant activity of each grape berry

Jean! AG! Douro!
Malvasia Preta! AG! Douro!
Rufete! AG! Douro!
fraction was determined from the extracts according
to two previously described methods: ABTS and
Sousão! AG! Douro!

DPPH. The ABTS method is based on the

Tinta Amarela! AG! Douro!

discoloration that occurs when the radical cation

Tinta Barca! AG! Douro!

ABTS .+ is reduced to ABTS (2,2’-azinobis-3-

Tinta Barroca! AG! Douro!

ethylbenzothiazoline-6-sulfonic acid) (re et al.,

Tinta Miúda! AG! Douro!
Tinto Cão! AG! Douro!
AG - autochthonous grape variety; nAG - non- 1999). The radical was generated by reaction of a
autochthonous grape variety; * white grape variety with
colored skin 7 mM solution of ABTS in water with 2.45 mM
potassium persulphate (1:1). The assay was made with
The evolutions of the climatic characteristics (rainfall, 980 μL of ABTS .+ solution and 20 μL of sample
temperature and relative humidity) for the 2010 (diluted 1:50 in water). The reaction was incubated for
vintage in the two regions considered are summarized 15 min in darkness at room temperature, and then
in Figure 1. In general, it was clear that during the absorbance at 734 nm was measured.
three months (June, July and August) prior to
sampling (at the end of September) the average The procedure using the DPPH method is described
temperature and average relative humidity was higher by Brand-Williams et al. (1995). Briefly, 0.1 mL of
in the Dão region than in the Douro region. In different sample concentrations was added to 3.9 mL
addition, average rainfall for the months of July, of 2,2-diphenyl-1-picrylhydrazyl (DPPH) methanolic
August and September was lower in the Dão region. solution (25 mg/L). The DPPH solution was prepared
daily and protected from the light. The reaction was
2. General physico-chemical and phenolic carried out for 30 min at 20 ºC in closed Eppendorf
parameters tubes under shaking, and then absorbance at 515 nm
was measured. Methanol was used as a blank
Estimated alcohol degree, pH, titratable acidity and
reference. The antioxidant activity results were
tartaric acid were measured directly from the must
expressed as Trolox equivalents (TEAC mM), using
obtained after grape pressing, using the analytical
the relevant calibration curve. All measurements were
methods recommended by the oIV (2006). Total
phenolic compounds were determined by measuring performed in duplicate.
absorbance at 280 nm (ribéreau-Gayon et al., 1982). 5. Chromatographic analysis of individual
non-flavonoids and flavonoids were determined using anthocyanins
the method described by Kramling and Singleton
(1969). Color intensity (A420 + A520 + A620) and hue The analysis of individual anthocyanins from skin
(A420/A520) were estimated using the method described extracts was performed on a HPLC Dionex Ultimate
by the oIV (2006). All analyses were done in 3000 Chromatographic System (Sunnyvale,
duplicate. California, USA) equipped with a quaternary pump

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Elisa CoSTA et al.

Figure 1. Evolution of climatic characteristics (temperature, relative humidity and rainfall)

for the 2010 vintage in the two regions considered.

Model LPG-3400 A, an auto sampler Model ACC- treatment means were carried out using Statistica 7
3000, a thermostated column compartment (adjusted software (StatSoft, Tulsa, USA). Scheffle’s test was
to 25 ºC) and a multiple Wavelength Detector MWD- applied to the data as comparison test to determine
300. The column (250 x 4.6 mm, particle size 5 μm) when samples are significantly different after AnoVA
was a C 18 Acclaim ® 120 (Dionex, Sunnyvale, (p < 0.05). Principal component analysis (PCA) was
California, USA) protected by a guard column of the used to analyze the data and study the relations
same material. The solvents were (A) 40 % formic between the grape varieties and their anthocyanin
acid, (B) pure acetonitrile and (C) bidistilled water. profile or antioxidant activity.
The anthocyanin composition was analyzed by HPLC
using the method described by Dallas and Laureano RESULTS AND DISCUSSION
(1994b). Thus, initial conditions were 25 % A, 10 % 1. General physico-chemical composition
B and 65 % C, followed by a linear gradient from 10
to 30 % B and 65 to 45 % C for 40 min, with a flow The physico-chemical parameters of the grape
rate of 0.7 mL/min. The injection volume was 40 µL. varieties at technological maturity are summarized in
Detection was made at 520 nm using Chromeleon Table 2. The estimated alcohol degree ranged from
6.8 software (Sunnyvale, California, USA). Individual 9.53 % (v/v) (Grenache) to 14.84 % (v/v) (Malvasia
anthocyanins were quantified by means of calibration Preta) with an average of 12.3 % (v/v). Titratable
curve obtained with standard solutions of malvidin-3- acidity, expressed as equivalent of tartaric acid, varied
glucoside chloride (>95 % purity) from Extrasynthese from 3.9 (Moreto Boal) to 13.5 g/L (Jean and Tinta
(Genay, France). The chromatographic peaks of Miúda) with an average of 7.3 g/L. The high titratable
anthocyanins were identified according to reference acidity and low pH values found in the majority of the
data previously described by Dallas and Laureano grape varieties are probably a consequence of two
(1994b). All analyses were done in duplicate. main factors: their natural acidity and the ripening
process (associated to low degradation rates of organic
6. Statistical analysis acids). In addition, these results are in agreement with
The data are presented as mean ± standard deviation. the tartaric acid content of all grape varieties.
To determine whether there is a statistically significant 2. General phenolic composition and anthocyanin
difference between the data obtained for antioxidant profile
activity and for the diverse phenolic compounds
quantified in the different grape varieties, an analysis It is well known that the genetic potential for
of variance (AnoVA, one-way) and comparison of polyphenol biosynthesis and the degree of ripeness of

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Table 2. General physico-chemical composition of grape varieties at technological maturity.

Grape berry Must volume Estimated alcohol degree pH Titratable acidity Tartaric acid
Grape variety
weight (g)a (mL)a (%, v:v) (g/L tartaric acid) (g/L)

Camarate 241 ± 10a 128 ± 5c 11.00 ± 0.02e 2.93 ± 0.04ef 5.1 ± 0.1abc 3.19 ± 0.23abc
Gewürztraminer 444 ± 1l 177 ± 1f 12.76 ± 0.01m 2.41± 0.00a 8.8 ± 0.2fgh 6.99 ± 0.02ijkl
Monvedro 273 ± 8bcd 130 ± 4cd 12.35 ± 0.01k 3.11± 0.00hij 4.7 ± 0.3ab 2.56 ± 0.02a
Moreto Boal 325 ± 2fg 152 ± 1e 13.59 ± 0.02p 3.17 ± 0.00ijk 3.9 ± 0.3a 3.35 ± 0.02bc
Negro Mole 318 ± 8efg 159 ± 4e 11.47 ± 0.01f 2.75 ± 0.00cd 6.2 ± 0.1cd 5.97 ± 0.08gh
Negro Mouro 319 ± 10efg 152 ± 5e 12.15 ± 0.02j 3.22 ± 0.00jk 4.1 ± 0.1a 3.38 ± 0.01bc
Alfrocheiro 260 ± 8ab 131± 6cd 11.96 ± 0.02h 2.91 ± 0.00ef 6.0 ± 0.4bcd 3.19 ± 0.23abc
Alvarilhão 297 ± 9def 144 ± 5de 13.45 ± 0.02o 2.83 ± 0.00de 8.0 ± 0.1ef 6.95 ± 0.06ijkl
Aramon 727 ± 6o 322 ± 3k 10.59 ± 0.02c 2.72 ± 0.01bc 9.5 ± 0.3gh 7.70 ± 0.12k
Bastardo 408 ± 2k 181 ± 1f 12.08 ± 0.02i 3.49 ± 0.00m 4.1 ± 0.2a 2.49 ± 0.68a
Cabernet Franc 268 ± 4abc 66 ± 0a 13.73 ± 0.01q 2.94 ± 0.01ef 6.0 ± 0.4bcd 5.56 ± 0.04fg
Carignan Noir 450 ± 3l 179 ± 1f 12.49 ± 0.02l 3.14 ± 0.01hij 8.5 ± 0.3fg 6.01 ± 0.11gh
Cornifesto 303 ± 6ef 108 ± 2b 14.01 ± 0.02r 3.26 ± 0.00k 6.0 ± 0.5bcd 3.35 ± 0.02bc
Gamay 487 ± 5m 228 ± 2h 11.67 ± 0.01g 3.09 ± 0.09hi 8.0 ± 0.1ef 6.38 ± 0.01hij
Grenache 378 ± 6ij 203 ± 4g 9.53 ± 0.01a 3.22 ± 0.07jk 10.0 ± 0.4h 6.22 ± 0.00ghi
Jean 387 ± 3jk 179 ± 1f 13.45 ± 0.02o 2.92 ± 0.01ef 13.5 ± 0.5i 7.04 ± 0.05jkl
Malvasia Preta 295 ± 8cde 129 ± 4cd 14.84 ±0.02s 3.38 ± 0.01l 8.5 ± 0.4fg 3.88 ± 0.17cd
Rufete 484 ± 8m 269 ± 5j 12.08 ± 0.01i 3.05 ± 0.01gh 5.5 ± 0.3bcd 2.87 ± 0.02ab
Sousão 336 ± 7gh 178 ± 4f 9.66 ± 0.02b 2.62 ± 0.01b 8.5 ± 0.1fg 7.24 ± 0.02kl
Tinta Amarela 530 ± 9n 270 ± 9j 10.93 ± 0.01d 2.91 ± 0.02ef 6.3 ± 0.1cd 3.50 ± 0.16bc
Tinta Barca 459 ± 9lm 230 ± 5hi 11.67 ± 0.02g 2.88 ± 0.01ef 9.0 ± 0.2fgh 6.53 ± 0.03ijk
Tinta Barroca 527 ± 7n 246 ± 3i 12.76 ± 0.01m 3.15 ± 0.01hij 5.5 ± 0.4bc 4.57 ± 0.37de
Tinta Miúda 358 ± 7hi 197 ± 4g 13.45 ± 0.02o 2.88 ± 0.00ef 13.5 ± 0.3i 7.25 ± 0.24kl
Tinto Cão 272 ± 6bcd 111 ± 2b 13.04 ± 0.01n 2.94 ± 0.00fg 6.8 ± 0.5de 4.96 ± 0.16ef
aGrape berry weight and must volume of 200 berries; data are the average of two replicates ± standard deviation; different letters above means
indicate statistically significant differences between grape varieties (p < 0.05).

individual grape varieties may affect the polyphenolic aging processes, but also on the potential
content in grape berries at harvest. In addition, the concentration of anthocyanins in grape skin as well as
concentration of phenolic compounds is highly the easiness of anthocyanin extraction from skin into
influenced by viticulture and environmental factors must (ortega-regules et al., 2006).
such as sunlight, temperature, altitude, soil type, and
water and nutritional status (Jackson and Lombard, The individual anthocyanin composition of the skin
1993; Yokotsuka et al., 1999). As expected, due to the extracts of the grape varieties at technological
reasons mentioned above, differences in phenolic maturity is presented in Table 4. Malvidin-3-glucoside
composition were observed among grape varieties was the major individual anthocyanin (concentrations
(Table 3). ranging from 0.62 to 6.09 mg/g of skin) in all varieties
except Alvarilhão and Rufete, where the major
The total phenolic compounds, expressed as individual anthocyanins were peonidin-3-glucoside
equivalent of gallic acid, ranged from 989 to (1.04 mg/g of skin) and malvidin-3-p-coumaroyl
3033 mg/L with an average of 1608.5 mg/L. The glucoside (1.48 mg/g of skin), respectively. Malvidin-
highest concentration of total phenols (ranging from 3-p-coumaroyl glucoside was the second main
2119 to 3033 mg/L) was detected in Tinta Barca, Tinta individual anthocyanin for the majority of the
Barroca, Sousão and Tinta Miúda, while Camarate, varieties and the values ranged from 0.12 to 3.44 mg/g
Moreto Boal, Cornifesto and Grenache showed the of skin. These results are in agreement with previous
lowest values (ranging from 989 to 1088 mg/L). findings in other grape varieties (Dallas and Laureano,
1994a; Jordão et al., 1998a; Kallithraka et al., 2005;
The interest of winemakers for grape polyphenolic ortega-regules et al., 2006). According to ribéreau-
content is increasing, as it offers tools to influence the Gayon et al. (2006), malvidin derivative forms are
color, bitterness, astringency, ‘mouth-feel’ and ‘age- stable molecules and their presence give stability to
ability’ of wines. Therefore, the color of red wines and the wine during the winemaking process because of
its evolution depend not only on the winemaking and their relative resistance to oxidation. Mateus et al.

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Elisa CoSTA et al.

(2002) found that malvidin-3-glucoside and its important group, ranging from 0.23 (Bastardo) to 2.87
acylated esters were the major anthocyanin (Tinto Cão) mg/g of skin, except in four varieties
monoglucosides in Touriga Nacional and Touriga (Camarate, Gewürztraminer, Monvedro and Jean)
Francesa at harvest date. where the acetyl glucoside group was the second main
group. A high concentration of p-coumaroyl-
In general, cyanidin derivatives (cyanidin-3-acetyl anthocyanin derivatives, mainly from malvidin and
glucoside and petunidin-3-acetyl glucoside) were the petunidin, has been associated with warm climates
less abundant individual anthocyanins (ranging from (Downey et al., 2006). However, according to
0.01 to 0.05 and 0.01 to 0.39 mg/g of skin, Férnandez-Lopez et al. (1998) it is commonly
respectively), which is in line with the results accepted that the anthocyanin composition of each
published by several authors (Dimitrovska et al., cultivar is closely linked to its genetic inheritance and,
2011; ortega-Meder et al., 1994; roggero et al., from a qualitative point of view, is quite independent
1986). In addition, these two anthocyanins were not of seasonal conditions or production area.
detected in a great number of the grape varieties
studied. According to Di Stefano and Flamini (2008), The acetyl glucoside group, which participates in an
cyanidin is the precursor of peonidin-3-glucoside by intra-molecular copigmentation process leading to an
the action of the UFGT (which transform cyanidin increase in wine color intensity (ortega-regules et
into cyanidin-3-glucoside) and MT enzymes (which al., 2006), was the minor group, ranging from 0.01
transform cyanidin-3-glucoside into peonidin-3- (Grenache) to 2.03 (Gewürztraminer) mg/g of skin.
glucoside). Although the anthocyanin profile may be complex
and quite different for each variety studied, in general
The glucoside group was the main anthocyanin Carignan Noir, Monvedro and Negro Mole showed
chemical group in all grape varieties, ranging from the highest individual anthocyanin concentration
1.40 (Rufete) to 7.09 (Carignan Noir) mg/g of skin while Bastardo, Aramon, Grenache and Alvarilhão
(sum of all five 3-glucoside anthocyanins). The showed the lowest concentration. It is important to
coumaroyl glucoside group was the second most note that grape anthocyanin concentration is largely

Table 3. General phenolic composition of grape varieties at technological maturity.

Total phenolic Flavonoid Non-flavonoid

compounds Color intensity Color hue
Grape variety compounds compounds
(mg/L gallic acid) (abs.u.)a (abs.u.)a
(mg/L gallic acid) (mg/L gallic acid)
Camarate 989 ± 13a 946 ± 11abc 43 ± 2a 13.57 ± 0.83h 6.31 ± 0.35cde
Gewürztraminer 1124 ± 17ab 1031 ± 16bcd 93 ± 1abc 9.34 ± 0.24cde 7.96 ± 0.25ghi
Monvedro 1240 ± 14abc 1118 ± 13bcde 122 ± 1bcd 7.62 ± 0.16b 7.46 ± 0.15efgh
Moreto Boal 1032 ± 39a 961 ± 9abc 71 ± 3ab 5.21 ± 0.13a 11.33 ± 0.26j
Negro Mole 1375 ± 17abcd 1278 ± 16defg 97 ± 1abc 10.15 ± 0.38ef 5.54 ± 0.08abc
Negro Mouro 1593 ± 26def 1408 ± 23efgh 185 ± 3def 12.17 ± 0.11gh 6.46 ± 0.12cdef
Alfrocheiro 1492 ± 13bcd 1262 ± 11defg 229 ± 2efgh 9.94 ± 0.20def 7.79 ± 0.11fghi
Alvarilhão 1820 ± 39fgh 1582 ± 34ghi 238 ± 5efgh 9.17 ± 0.16cde 7.28 ± 0.22defgh
Aramon 1590 ± 13def 1341 ± 11fgh 248 ± 2fghi 8.56 ± 0.14bcd 8.74 ± 0.10i
Bastardo 2024 ± 37gh 1662 ± 30hi 362 ± 7k 9.93 ± 0.23def 8.59 ± 0.25hi
Cabernet Franc 1900 ± 17fgh 1535 ± 14fghi 365 ± 6k 13.26 ± 0.15h 8.26 ± 0.07hi
Carignan Noir 1476 ± 22bcd 1268 ± 19defg 208 ± 5efg 10.28 ± 0.87ef 8.35 ± 0.51hi
Cornifesto 1088 ± 18ab 874 ± 13a 214 ± 3efg 8.25 ± 0.04bc 6.82 ± 0.10cdefg
Gamay 1529 ± 18cde 1309 ± 12efgh 220 ± 2efgh 12.86 ± 0.43h 8.08 ± 0.48ghi
Grenache 1038 ± 52a 907 ± 9ab 131 ± 7bcd 5.41 ± 0.60a 7.43 ± 0.35efgh
Jean 1530 ± 24cde 1334 ± 10efgh 196 ± 5defg 10.99 ± 0.30fg 5.94 ± 0.29bcd
Malvasia Preta 1352 ± 28abcd 1083 ± 23bcde 269 ± 6ghij 10.05 ± 0.14ef 6.18 ± 0.27bcde
Rufete 1297 ± 33abc 1133 ± 28bcde 164 ± 4cde 11.16 ± 0.33fg 5.86 ± 0.00bc
Sousão 2130 ± 74h 1898 ± 12i 232 ± 8efgh 18.68 ± 0.21jk 5.78 ± 0.22bc
Tinta Amarela 1490 ± 28bcd 1197 ± 23cdef 294 ± 6hijk 16.22 ± 0.37i 4.86 ± 0.03ab
Tinta Barca 3033 ± 19i 2700 ± 16j 333 ± 13jk 19.15 ± 0.12k 5.99 ± 0.02bcd
Tinta Barroca 2783 ± 13i 2418 ± 18j 365 ± 15k 17.68 ± 0.10f 6.14 ± 0.16bcde
Tinta Miúda 2119 ± 37h 1857 ± 32i 263 ± 5ghij 21.68 ± 0.16l 4.19 ± 0.12a
Tinto Cão 1562 ± 33def 1254 ± 26defg 308 ± 11ijk 13.35 ± 0.42h 5.78 ± 0.18bc
a abs.u. – absorbance units referred to a 1 mm path length cell used; data are the average of two replicates ± standard deviation;

different letters above means indicate statistically significant differences between grape varieties (p < 0.05).

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due, among other factors, to the berry size of each of skin) and malvidin-3-p-coumaroyl glucoside
grape variety. on the other hand, Gewürztraminer, (ranging from 0.88 to 3.44 mg/g of skin). The
Monvedro, Moreto Boal, Negro Mole, Alvarilhão, varieties Moreto Boal, Monvedro, Gewürztraminer
Cabernet Franc, Jean and Tinta Amarela presented and Camarate appeared in the positive part of PC2,
the widest variety of individual anthocyanins while due to their high content in malvidin-3-acetyl
Rufete presented the narrowest variety of individual glucoside (ranging from 1.01 to 1.65 mg/g of skin).
anthocyanins quantified.
From the description presented above, anthocyanins
To better understand the relationship between grape could be considered useful markers to distinguish
variety and anthocyanin concentration, a principal grape varieties; however, this characteristic should be
component analysis (PCA) was performed used with caution since anthocyanin concentration is
(Figure 2A). The first two principal components (PCs) influenced not only by genetic background but also by
explained 92.22 % of the total variance and showed agroecological factors, such as maturation (Conde et
that grape varieties can be distinguished according to al., 2007; Jordão et al., 1998a), ripening stage (Segade
their individual anthocyanin concentration. Figure 2A et al., 2008), climate (Gil and Yuste, 2004), stress
shows the corresponding loading plots that established levels (Gatto et al., 2008) and cultural practices
the relative importance of each variable. The first (Jordão et al., 1998b).
principal component (PC1), which explained 81.89 %
of the variance, was negatively correlated with the 3. Antioxidant activity from different grape berry
variables malvidin-3-glucoside and malvidin-3-p- fractions
coumaroyl glucoside. The second PC (PC2, 10.33 %
of the variance) was positively correlated with the The data in Table 5 show the antioxidant activity
variable peonidin-3-glucoside and negatively quantified in the different grape berry fractions (pulps,
correlated with malvidin-3-acetyl glucoside. Five skins and seeds) from the grape varieties studied. The
groups could be distinguished. The varieties highest antioxidant activity was found in seeds
Camarate, Gewürztraminer, Monvedro, Negro Mole, (ranging from 77.59 to 867.81 and from 75.52 to
Negro Mouro, Cabernet Franc, Carignan Noir, 363.47 µmol/g of seed, for ABTS and DPPH method,
Cornifesto and Tinta Barroca were rather grouped on respectively), followed by skins (ranging from 1.13 to
the negative side of PC1, due to their high content in
Figure
Figure 22..
292.05 and from 1.78 to 299.99 µmol/g of skin, for
malvidin-3-glucoside (ranging from 4.10 to 6.09 mg/g ABTS and DPPH method, respectively) and pulps
A B

Figure 2. Principal component analysis score plot (PC1 and PC2) of grape varieties:
individual anthocyanins (A) and antioxidant activity (B).
CA, Camarate; GW, Gewürztraminer; Mn, Monvedro; MB, Moreto Boal; nM, negro Mole; nMr, negro Mouro; AF,
Alfrocheiro; AV, Alvarilhão; Ar, Aramon; BT, Bastardo; CF, Cabernet Franc; Cn, Carignan noir; Cr, Cornifesto; GY, Gamay;
Gr, Grenache; J, Jean; MP, Malvasia Preta; r, rufete; S, Sousão; TA, Tinta Amarela; TB, Tinta Barca; TBr, Tinta Barroca;
TM, Tinta Miúda; TC, Tinto Cão.
Delp gluc, delphinidin-3-glucoside; Cyan gluc, cyanidin-3-glucoside; Petun gluc, petunidin-3-glucoside; Peon gluc, peonidin-3-
glucoside; Malv gluc, malvidin-3-glucoside; Cyan acet-gluc, cyanidin-3 acetyl glucoside; Petun acet-gluc, petunidin-3-acetyl
glucoside; Peon acet-gluc, peonidin-3-acetyl glucoside; Malv acet-gluc, malvidin-3-acetyl glucoside; Peon coum-gluc, peonidin-

J. Int. Sci. Vigne Vin, 2014, 48, n°1, 51-62 31

- 57 - ©Vigne et Vin Publications Internationales (Bordeaux, France)
 Table
Table4.4.Individual
Individualanthocyanins quantified in
anthocyanins quantified ingrape
grapeskin
skinatattechnological
technologicalmaturity.
maturity.

Cyan Petun Peon Malv Petun Peon Malv

Grape variety Delp gluc Cyan gluc Petun gluc Peon gluc Malv gluc
acet-gluc acet-gluc acet-gluc acet-gluc coum-gluc coum-gluc coum-gluc
Elisa CoSTA et al.

Camarate n.d. 0.05 ± 0.01e 0.22 ± 0.01defg 0.49 ± 0.02e 5.08 ± 0.02g 0.02 ± 0.0bc 0.02 ± 0.00a 0.12 ± 0.01g 1.29 ± 0.07i 0.12 ± 0.00g 0.11 ± 0.00def 0.90 ± 0.17def
Gewürztraminer 0.21 ± 0.05cd 0.01 ± 0.00a 0.44 ± 0.11j 0.26 ± 0.02cd 4.92 ± 0.04g 0.03 ± 0.00d 0.39 ± 0.00d 0.07 ± 0.00ef 1.54 ± 0.03jk 0.39 ± 0.00k 0.07 ± 0.00bcde 0.88 ± 0.13def
Monvedro 0.60 ± 0.00f 0.01 ± 0.01a 0.94 ± 0.02m 0.30 ± 0.05d 6.09 ± 0.04i 0.05 ± 0.00e 0.14 ± 0.00bc 0.01 ± 0.00a 1.65 ± 0.01k 0.11 ± 0.01g 0.09 ± 0.00cdef 1.48 ± 0.03efg
Moreto Boal 0.08 ± 0.02ab 0.01 ± 0.00a 0.06 ± 0.00ab 0.49 ± 0.03e 3.96 ± 0.08f 0.01 ± 0.00ab 0.06 ± 0.00a 0.11 ± 0.00g 1.01 ± 0.00h 0.10 ± 0.00fg 0.12 ± 0.01ef 1.27 ± 0.03efg
08-jordao_05b-tomazic 01/04/14 20:26 Page58

Negro Mole 0.04 ± 0.01ab 0.01 ± 0.00a 0.24 ± 0.05efgh 0.51 ± 0.10e 5.92 ± 0.00hi 0.02 ± 0.00bc 0.21 ± 0.00c 0.11 ± 0.00g 1.43 ± 0.07j 0.08 ± 0.00ef 0.11 ± 0.01def 2.21 ± 0.07gh
Negro Mouro 0.03 ± 0.01ab n.d. 0.21 ± 0.00cdefg 0.54 ± 0.05ef 5.88 ± 0.04hi 0.02 ± 0.00bc 0.08 ± 0.01ab 0.10 ± 0.00fg 1.45 ± 0.04j 0.03 ± 0.01a 0.09 ± 0.03cdef 2.23 ± 0.05gh
ab bcdef b e abcd bcde cde ab
Alfrocheiro 0.03 ± 0.01 n.d. 0.16 ± 0.01 0.14 ± 0.00 2.90 ± 0.15 n.d. n.d. 0.04 ± 0.01 0.18 ± 0.01 0.06 ± 0.00 0.04 ± 0.01 1.46 ± 0.15efg

J. Int. Sci. Vigne Vin, 2014, 48, n°1, 51-62

Alvarilhão 0.08 ± 0.01ab 0.24 ± 0.01h 0.12 ± 0.00bcd 1.04 ± 0.04h 0.99 ± 0.05ab 0.01 ± 0.00a 0.01 ± 0.00a 0.06 ± 0.00cde 0.05 ± 0.00ab 0.01 ± 0.00a 0.12 ± 0.01ef 0.12 ± 0.03a
ab bc ab bc b a a bcde a a
Aramon 0.05 ± 0.00 0.03 ± 0.00 0.08 ± 0.00 0.17 ± 0.01 1.24 ± 0.01 n.d. 0.01 ± 0.00 0.01 ± 0.00 0.15 ± 0.01 0.03 ± 0.00 0.01 ± 0.00 0.32 ± 0.08bc
Bastardo 0.02 ± 0.00a 0.03 ± 0.00bc 0.01 ± 0.00a 0.13 ± 0.00ab 0.62 ± 0.00a n.d. n.d. 0.02 ± 0.00abc 0.05 ± 0.00ab 0.02 ± 0.00a 0.05 ± 0.00abc 0.16 ± 0.01a
g de m bc f a a i ef i a
Cabernet Franc 0.95 ± 0.00 0.04 ± 0.00 1.03 ± 0.01 0.17 ± 0.02 4.10 ± 0.13 0.01 ± 0.00 0.02 ± 0.00 0.20 ± 0.01 0.27 ± 0.00 0.22 ± 0.01 0.02 ± 0.00 1.52 ± 0.10fgh
Carignan Noir 0.34 ± 0.05e 0.02 ± 0.01ab 0.57 ± 0.01k 0.56 ± 0.03ef 5.60 ± 0.37h n.d. n.d. 0.04 ± 0.01abcd 0.53 ± 0.01g 0.16 ± 0.01h 0.35 ± 0.03g 3.44 ± 0.43i
Cornifesto 0.03 ± 0.01a n.d. 0.10 ± 0.00abc 0.56 ± 0.03ef 5.60 ± 0.37h n.d. n.d. 0.02 ± 0.00abc 0.17 ± 0.00bcde 0.06 ± 0.00cde 0.03 ± 0.00a 1.38 ± 0.13efg
Gamay 0.02 ± 0.01a n.d. 0.08 ± 0.02ab 0.12 ± 0.00ab 1.77 ± 0.03c n.d. n.d. 0.04 ± 0.00abcd 0.23 ± 0.00cde 0.02 ± 0.00a 0.11 ± 0.02def 1.61 ± 0.01fgh

©Vigne et Vin Publications Internationales (Bordeaux, France)

Grenache 0.25 ± 0.00cd 0.04 ± 0.00de 0.14 ± 0.01bcde 0.18 ± 0.00bcd 1.44 ± 0.01bc n.d. n.d. n.d. 0.01 ± 0.00a 0.05 ± 0.00bcd 0.05 ± 0.00abc 0.17 ± 0.01a
b ab cdef cd de bc a abc h abc a
Jean 0.09 ± 0.01 0.02 ± 0.00 0.20 ± 0.01 0.27 ± 0.01 2.75 ± 0.05 0.02 ± 0.00 0.04 ± 0.00 0.03 ± 0.03 0.89 ± 0.07 0.04 ± 0.01 0.02 ± 0.00 0.77 ± 0.02cde
Malvasia Preta 0.21 ± 0.00cd 0.06 ± 0.01ef 0.31 ± 0.00ghi 0.55 ± 0.02ef 2.32 ± 0.06d n.d. 0.01 ± 0.00a 0.01 ± 0.00a 0.10 ± 0.00abc 0.02 ± 0.00a 0.04 ± 0.02abc 0.28 ± 0.05ab
a bc ab bcde a abc
Rufete n.d. n.d. n.d. 0.01 ± 0.00 1.39 ± 0.01 n.d. n.d. 0.02 ± 0.00 0.14 ± 0.01 0.03 ± 0.00 0.05 ± 0.00 1.48 ± 0.03efg

- 58 -
Sousão 0.21 ± 0.01cd 0.09 ± 0.01g 0.33 ± 0.01hi 1.34 ± 0.01i 2.76 ± 0.05de n.d. n.d. n.d. 0.06 ± 0.00ab 0.02 ± 0.00a 0.04 ± 0.00abc 0.18 ± 0.08a
f fg l g e bc a abcd ab bcd abcd
Tinta Amarela 0.58 ± 0.02 0.07 ± 0.01 0.74 ± 0.01 0.69 ± 0.02 2.91 ± 0.07 0.02 ± 0.00 0.04 ± 0.01 0.03 ± 0.01 0.10 ± 0.01 0.05 ± 0.00 0.06 ± 0.01 0.65 ± 0.01cde
Tinta Barca 0.34 ± 0.00e 0.04 ± 0.00de 0.46 ± 0.01jk 0.23 ± 0.00bcd 2.41 ± 0.01d n.d. n.d. 0.05 ± 0.00bcde 0.13 ± 0.01bcd 0.04 ± 0.01abc n.d. 0.98 ± 0.01def
Tinta Barroca 0.21± 0.01cd 0.06 ± 0.00ef 0.41 ± 0.01ij 0.57 ± 0.00efg 4.93 ± 0.05g n.d. n.d. 0.07 ± 0.00de 0.26 ± 0.00def 0.11 ± 0.01g 0.13 ± 0.01f 1.94 ± 0.03fgh
Tinta Miúda 0.20 ± 0.01c 0.01 ± 0.00a 0.25 ± 0.01fgh 0.64 ± 0.03fg 2.31 ± 0.06d n.d. 0.02 ± 0.00a 0.04 ± 0.00bcde 0.26 ± 0.00def 0.02 ± 0.01a 0.11 ± 0.02def 0.41 ± 0.04bcd
Tinto Cão 0.27 ± 0.01d 0.01 ± 0.00a 0.39 ± 0.01ij 0.13 ± 0.02ab 2.65 ± 0.01de n.d. 0.01 ± 0.00a 0.16 ± 0.01h 0.37 ± 0.00f 0.25 ± 0.01j 0.05 ± 0.03abc 2.57 ± 0.02hi

Delp gluc, delphinidin-3-glucoside; Cyan gluc, cyanidin-3-glucoside; Petun gluc, petunidin-3-glucoside; Peon gluc, peonidin-3-glucoside; Malv gluc, malvidin-3-glucoside; Cyan acet-gluc, cyanidin-3- acetyl
glucoside;
Delp gluc, Petun acet-gluc, petunidin-3-acetyl
delphinidin-3-glucoside; Cyan glucoside; Peon acet-gluc, peonidin-3-acetyl
gluc, cyanidin-3-glucoside; Petun gluc,glucoside; Malv acet-gluc, malvidin-3-acetyl
petunidin-3-glucoside; glucoside; Peon coum-gluc,
Peon gluc, peonidin-3-glucoside; Malvpeonidin-3-p-coumaroyl
gluc, malvidin-3-glucoside; Malv
glucoside; Cyan
coum-gluc,
acet-gluc,malvidin-3-p-coumaroyl glucoside; individual
cyanidin-3- acetyl glucoside; anthocyanins
Petun acet-gluc, are expressed as malvidin-3-glucoside
petunidin-3-acetyl glucoside; Peon equivalents
acet-gluc, (mg/g of skin); n.d., not
peonidin-3-acetyl detected; data
glucoside; Malvare acet-gluc,
means of twomalvidin-3-acetyl deviation;
replicates ± standardglucoside;
different letters above means
Peon coum-gluc, indicate statistically significant
peonidin-3-p-coumaroyl differences
glucoside; Malvbetween grape varieties
coum-gluc, (p < 0.05).
malvidin-3-p-coumaroyl glucoside; individual anthocyanins are expressed as malvidin-3-glucoside
equivalents (mg/g of skin); n.d., not detected; data are the average of two replicates ± standard deviation; different letters above means indicate statistically significant differences
between grape varieties (p < 0.05).

26
 Table 5. Antioxidant activities of grape skin, pulp and seed extracts as measured by the ABTS
Table 5. Antioxidant activities of grape skin, pulpand seed extracts
andDPPH methodsasinmeasured
grape varieties
by theatABTS and DPPH
technological methods in grape varieties at technological maturity.
maturity.

Skins Pulps Seeds

Grape variety
ABTS DPPH ABTS DPPH ABTS DPPH
abc efgh kl j
Camarate 47.64 ± 0.62 58.14 ± 2.02 2.57 ± 0.00 3.13 ± 0.27 161.13 ± 7.23bcde 129.00 ± 18.91abc
Gewürztraminer 78.78 ± 7.45gh 45.30 ± 0.00defg 2.14 ± 0.07ijk 2.02 ± 0.05ef 276.21 ± 6.03fghi 330.89 ± 1.89ij
Monvedro 34.48 ± 0.62b 66.71 ± 0.00ghi 1.58 ± 0.02fghi 1.84 ± 0.07de 77.59 ± 0.00a 96.91 ± 11.35ab
Moreto Boal 93.28 ± 1.86h 9.63 ± 1.09ab 3.65 ± 0.02n 3.12 ± 0.03j 310.31 ± 1.21hi 210.56 ± 9.45def
Negro Mole 71.78 ± 4.96fg 88.11 ± 2.02hi 0.95 ± 0.04bcde 1.05 ± 0.09b 185.15 ± 15.13cde
08-jordao_05b-tomazic 01/04/14 20:26 Page59

137.26 ± 4.82bcd
Negro Mouro 57.74 ± 4.96cdef 48.60 ± 3.78defg 1.95 ± 0.07ijk 2.48 ± 0.01ghi 145.79 ± 14.47bcd 239.97 ± 39.71efg
Alfrocheiro 47.20 ± 3.72abc 15.34 ± 4.04bc 0.04 ± 0.03a 0.18 ± 0.06a 105.72 ± 1.21ab 229.27 ± 54.83def
Alvarilhão 9.03 ± 3.10a 38.79 ± 20.18defg 0.05 ± 0.03a 2.01 ± 0.06ef 174.77 ± 4.82bcde 300.14 ± 3.78ghij
Aramon 133.21 ± 4.96i 203.27 ± 16.40k 4.80 ± 0.22o 2.59 ± 0.07hi 505.52 ± 16.88l 314.84 ± 1.89hij
Bastardo 1.13 ± 0.62a 1.78 ± 0.01a 0.82 ± 0.07bcd 2.02 ± 0.05ef 410.05 ± 19.29jk 363.47 ± 11.82j
Cabernet Franc 52.91 ± 1.86bcdef 30.32 ± 1.01cde 1.71 ± 0.12fghi 1.48 ± 0.02cd 506.37 ± 10.85l 264.87 ± 4.73fghi
Carignan Noir 192.01 ± 9.93j 182.37 ± 1.26k 3.42 ± 0.13nm 2.25 ± 0.01fgh 867.81 ± 18.08n 130.34 ± 9.45abc
Cornifesto 52.91 ± 0.62bcdef 23.40 ± 3.47bcd 1.16 ± 0.07cdef 1.25 ± 0.08bc 306.89 ± 44.61hi 106.27 ± 5.67ab
Gamay 42.38 ± 3.10bc 68.22 ± 11.35ghi 1.92 ± 0.07hij 2.24 ± 0.02fgh 224.21 ± 19.29efgh 253.57 ± 19.24efgh
Grenache 33.16 ± 6.21b 299.99 ± 5.04l 2.95 ± 0.06lm 2.71 ± 0.05i 348.67 ± 14.47ij 75.52 ± 11.35a
Jean 63.88 ± 13.65defg 94.08 ± 12.61i 3.30 ± 0.56nm 1.12 ± 0.06bc 453.52 ± 20.49kl 122.31 ± 1.89abc

- 59 -
Malvasia Preta 1.57 ± 1.24a 3.21 ± 1.01a 0.55 ± 0.04ab 1.12 ± 0.06bc 237.85 ± 16.88efgh 95.57 ± 13.24ab
Rufete 37.55 ± 2.48bc 63.76 ± 7.57fghi 1.86 ± 0.03ghij 1.09 ± 0.05b 539.62 ± 69.92l 189.49 ± 13.24cde
Sousão 68.27 ± 0.00efg 43.88 ± 4.04defg 1.30 ± 0.16fgh 1.09 ± 0.02b 189.26 ± 1.21cdef 249.65 ± 5.67efg
Tinta Amarela 45.89 ± 1.86bcd 7.75 ± 0.96ab 0.63 ± 0.28abc 0.49 ± 0.00a 711.81 ± 14.47m 219.92 ± 0.00def
Tinta Barca 50.28 ± 0.62bcde 137.33 ± 7.06j 0.16 ± 0.06a 0.49 ± 0.01a 122.77 ± 3.62abc 161.09 ± 11.35bcd
Tinta Barroca 7.20 ± 1.96a 87.39 ± 1.01hi 1.24 ± 0.13defg 1.12 ± 0.00bc 206.31 ± 13.26defg 237.30 ± 1.89efg
Tinta Miúda 292.05 ± 3.72k 32.24 ± 6.73cdef 2.55 ± 0.06kl 2.14 ± 0.11efg 236.99 ± 22.91efgh 239.97 ± 1.89efg
Tinto Cão 123.55 ± 7.45i 25.33 ± 4.04bcd 2.40 ± 0.15jkl 2.48 ± 0.01ghi 283.88 ± 31.34ghi 318.35 ± 14.18hij
Data are the average of two replicates ± standard deviation; values are expressed as !mol/g of skin, pulp or seed; different letters above means indicate
Data are the average of two replicates ± standard deviation; values are expressed as µmol/g of skin, pulp or seed; different letters above means indicate statistically
statistically
significant differences differences
significantbetween grapebetween
varieties grape varieties (p < 0.05).
(p < 0.05).

27

©Vigne et Vin Publications Internationales (Bordeaux, France)

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Elisa CoSTA et al.

(ranging from 0.04 to 4.80 and from 0.18 to and non-autochthonous grape varieties and the grape
3.13 µmol/g of pulp, for ABTS and DPPH method, samples from the two wine regions considered. From
respectively). In addition, the average values were these results, it is clear that the antioxidant activity
314.17, 68.24 and 1.82 µmol/g, respectively for seeds, values depend on the method used. According to
skins and pulps considering the ABTS method and Villaño et al. (2006), this divergence is due to the
211.01, 89.78 and 1.72 µmol/g, respectively for seeds, different reagents of the polyphenols with each
skins and pulps considering the DPPH method. one method applied. For Wang et al. (2004), ABTS˙+ and
possible explanation for this distribution in the DPPH radicals have a different stereochemical
different grape berry fractions could be the higher structure and a different method of genesis and thus
amount of polyphenols such as monomeric flavanols they lend, after the reaction with the antioxidants, a
(catechin and epicatechin), dimeric, trimeric and qualitatively different response to the inactivation of
polymeric proanthocyanidins, and phenolic acids in their radical. Some authors reported that no single
seeds compared to skins (Di Majo et al., 2008; Yilmaz assay can provide all the information needed to
and Toledo, 2004). These findings are in agreement evaluate antioxidant capacity, and multiple assays are
with previous studies conducted with other red Vitis therefore required to build up an antioxidant profile of
vinifera grape varieties (Poudel et al., 2008; Xu et al., particular foodstuffs (rivero-Pérez et al., 2007).
2010).
To highlight the relation between grape varieties and
Taking into account the antioxidant activity of the red their antioxidant activity in the different grape berry
grape varieties analyzed here, a large variation was fractions (pulps, skins and seeds), a PCA was applied
found in the different grape berry fractions (Table 5). (Figure 2B). The PCA of the antioxidant activity
For seeds and considering the ABTS method, values obtained by two methods showed that the first
Carignan Noir, Tinta Amarela, Rufete, Cabernet two PCs explained 84.47 % of the total variance. The
Franc, Aramon, Jean and Bastardo showed the first PC (70.21 % of the variance) was positively
highest antioxidant activity values (ranging from correlated with seeds ABTS and the second PC
410.05 to 867.81 µmol/g of seed; p < 0.05) while
(14.26 % of the variance) was positively correlated
Monvedro and Alfrocheiro showed the lowest
with skins DPPH and negatively correlated with seeds
antioxidant values (77.59 and 105.72 µmol/g of seed,
DPPH. Grape varieties appeared grouped due to their
respectively; p < 0.05). However, considering the
different antioxidant activity of skins, seeds and pulps.
DPPH method, Bastardo, Gewürztraminer, Tinto Cão,
The grape varieties Carignan Noir and Tinta Amarela
Aramon and Alvarilhão showed the highest
antioxidant activity values (ranging from 300.14 to were on the right side of PC1, disclosing a good
363.47 µmol/g of seed; p < 0.05) while Monvedro, correlation with seeds ABTS. However, the grape
Grenache and Malvasia Preta showed the lowest varieties Bastardo, Gewürztraminer, Alvarilhão and
antioxidant values (ranging from 75.52 to Tinto Cão were on the negative side of PC2,
96.91 µmol/g of seed; p < 0.05). disclosing a good correlation with seeds DPPH.

For skins and considering the ABTS method, Tinta A linear regression analysis was performed to
Miúda, Carignan Noir, Aramon and Tinto Cão determine the correlation between polyphenol
showed the highest antioxidant activity values composition and respective antioxidant activities in
(ranging from 123.55 to 292.05 µmol/g of skin; p < the different grape berry fractions. This analysis found
0.05) while Bastardo, Malvasia Preta and Tinta no significant correlation (p > 0.05) between general
Barroca showed the lowest antioxidant values phenolic parameters and antioxidant activity in the
(ranging from 1.13 to 7.20 µmol/g of skin, grape berry fractions. on the one hand, correlations
respectively; p < 0.05). The results obtained by the between the antioxidant activity and the total
DPPH method showed similar distribution of high polyphenolic content of a great number of grape seed
(Carignan Noir and Aramon) and low (Malvasia and skin extracts from different grape varieties have
Preta and Bastardo) antioxidant activity values. been reported (Monagas et al., 2005; Xu et al., 2010).
Finally, for pulps, high antioxidant activity values on the other hand, other authors (Bozan et al., 2008)
were found for Aramon, Moreto Boal and Carignan reported no significant correlations between
Noir (4.80, 3.65 and 3.42 µmol/g of pulp for ABTS individual flavanols analyzed by HPLC or total
method, respectively) while low antioxidant values polyphenols and antioxidant activity in seed extracts
were found for Alfrocheiro, Alvarilhão, Tinta Barca from several grape varieties. Thus, there is conflicting
and Malvasia Preta (0.04, 0.05, 0.16 and 0.55 µmol/g evidence in the literature about the correlation
of pulp for ABTS method, respectively). overall, between polyphenol content and the antioxidant
there is no clear difference between the autochthonous activity of grapes.

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CONCLUSIONS Biochemical changes throughout grape berry

development and fruit and wine quality. Food 1, 1-22.
In this work, the influence of grape variety on the Dallas C. and Laureano o., 1994a. Effect of So2 on the
phenolic content, anthocyanin profile and antioxidant extraction of individual anthocyanins and colored
activity of the different grape berry fractions (skins, matter of three Portuguese grape varieties during
pulps and seeds) has been analyzed in twenty-four winemaking. Vitis 33, 41-47.
different grape varieties cultivated in two Portuguese Dallas C. and Laureano o., 1994b. Effects of pH, sulphur
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