Food Chemistry 129 (2011) 514519

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Anthocyanin composition in aged Chilean Cabernet Sauvignon red wines

Mara J. Aguirre a, Mauricio Isaacs b, Betty Matsuhiro a,, Leonora Mendoza a, Leonardo S. Santos c,
Simonet Torres a
a

Faculty of Chemistry and Biology, Universidad de Santiago de Chile, Av. B. OHiggins 3363, Santiago, Chile
Faculty of Sciences, Universidad de Chile, Las Palmeras 3425, Santiago, Chile
c
Laboratory of Asymmetric Synthesis, Chemistry Institute of Natural Resources, Universidad de Talca, P.O. Box 747, Talca, Chile
b

a r t i c l e

i n f o

Article history:
Received 20 October 2010
Received in revised form 21 March 2011
Accepted 3 May 2011
Available online 8 May 2011
Keywords:
Anthocyanins
Condensation
Differential pulse voltammetry
Aged red wine

a b s t r a c t
Anthocyanins in aged Cabernet Sauvignon red wines were studied by HPLCMS. The major anthocyanin in 6, 7, and 8 year old red wine extracts was the dimer vitisin A produced by condensation of
malvidin-3-O-glucoside mediated by pyruvic acid. In aged wine, the content of malvidin-3-O-glucoside
decreased with age with a concomitant increase of vitisin A. The latter is accompanied by several
condensation products with molecular weight up to 1500 m/z. Differential pulse voltammetry indicated
that aged wines have lower antioxidant capacity than young wine (400 mV), but higher than
malvidin-3-O-glucoside (483 mV). Tafels plots showed that the electrochemical process occurring in
aged wines is different from young wines. Six, seven and eight year old wines show similar behaviour
with plots of 234, 177 and 188 mV/dec, respectively. These values are higher than the expected
120 mV/dec corresponding to a rst electronic transfer but smaller compared to the 523 mV/dec
corresponding to young wine.
2011 Elsevier Ltd. All rights reserved.

1. Introduction
The main anthocyanin identied in Chilean Cabernet Sauvignon
red wine is malvidin-3-O-glucoside, besides minor amounts of
delphinidin-3-O-glucoside, peonidin-3-O-glucoside-4-viniylphenol
and condensation products (Aguirre et al., 2010). During red wine
ageing, anthocyanins interact with phenolics compounds, pyruvic
acid and acetaldehyde to give new pigments (Alcalde-Eon,
Escribano-Bailn, Santos-Buelga, & Rivas-Gonzalo, 2006, 2007;
Brouillard, Chassaing, & Fougerousse, 2003; Francia-Aricha, Guerra,
Rivas-Gonzalo, & Santos-Buelga, 1997; He, Santos-Buelga, Silva,
Mateus, & de Freitas, 2006; Mateus, Silva, Vercauteren, & De Feitas,
2001; Monagas, Gomez-Cordoves, & Bartolome, 2005). Atanasova,
Fulcrand, Le Guernev, Cheynier, and Moutounet (2002) reported
the formation of an ethyl-linked dimer by reaction between malvidin-3-O-glucoside and acetaldehyde in model wine solution.
Fulcrand, Bernabdeljalil, Rigaud, Cheynier, and Moutounet (1998)
obtained a very stable pigment by reaction of malvidin-3-Omonoglucoside with pyruvic acid in model wine solution, known
as vitisin A (Fig. 1) it was also found in red and Porto wines (Bakker
et al., 1997; Mateus & de Freitas, 2001; Vivar-Quintana, SantosBuelga, & Rivas-Gonzalo, 2002; Wang, Race, & Shrikhande, 2003).
Hayasaka and Asenstorfer (2002) identied various vinyl adducts

Corresponding author. Tel.: +56 2 7181159; fax: +56 2 6812108.

E-mail address: betty.matsuhiro@usach.cl (B. Matsuhiro).
0308-8146/$ - see front matter 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodchem.2011.05.009

of anthocyanins in red wine extracts using nanoelectrospray tandem mass spectrometry. On the other hand, avanols condense
with anthocyanins in the presence or absence of acetaldehyde to
give polymeric compounds (Dallas, Ricardo-da-Silva, & Laureano,
1996; Escribano-Bailn, Alvarez-Garca, Rivas-Gonzalo, Heredia, &
Santos-Buelga, 2001; Nave, Teixeira, Mateus, & De Freitas, 2010;
Remy-Tanneau, Le Guernev, Meudec, & Cheynier, 2003; RivasGonzalo, Bravo-Haro, & Santos-Buelga, 1995; Salas, Fulcrand, Meudec, & Cheynier, 2003). Free anthocyanins and polymeric anthocyanins were detected and identied by mass spectrometric
methods. Fast atom bombardment mass spectrometry (FAB-MS),
matrix-assisted laser desorption/ionisation mass spectrometry
(MALDI-MS), and electrospray (ESI) mass spectrometry have been
used (Bakker & Timberlake, 1997; Giusti, Rodrguez-Sanoa, Grifn,
& Wrolstad, 1999; Pati et al., 2006; Piovan, Filippini, & Favretto,
1998; Wang & Sporns, 1999).
It is known that phenolics compounds in wine have antioxidant
capacity and free radical scavenging activity. There are many
chemical methods to analyse the antioxidant capacity of
foodstuffs, but no single assay can provide the information needed
to evaluate antioxidant capacity (Aguirre et al., 2010; Ghiselli,
Nardini, Baldi, & Scaccini, 1997; Huang, Ou, & Prior, 2005; Piljac,
Martnez, Valek, Stipcevic, & Kovacevic Ganic, 2005).
The aim of this work was to establish anthocyanins proles in
aged Chilean Cabernet Sauvignon red wines using LCMS/MS and
study the antioxidant capacity of aged wines by electrochemical
methods.

M.J. Aguirre et al. / Food Chemistry 129 (2011) 514519

OCH

volume of n-hexane, chloroform, and ethyl acetate. The remaining

aqueous phase was concentrated to 100 ml and poured over
500 ml of cold ethyl alcohol. The mixture was centrifuged in a Universal 32 centrifuge (Hettich, Tuttlingen, Germany) and the supernatant was concentrated to a nal volume of 50 ml.

OH
+

HO

OCH

515

Ogluc
O

COOH
Fig. 1. Structure of vitisin A.

2. Materials and methods

2.1. Materials
One, six, seven, and eight year old Cabernet Sauvignon red wine
bottles were supplied by a local producer. All the solvents were reagent grade and/or HPLC grade (Merck, Darmstadt, Germany), all
the chemicals were reagent grade (Sigma, St. Louis, MO, USA).
2.2. HPLC analysis
For the high performance liquid chromatography (HPLC) analysis a Waters 600 HPLC chromatograph (Waters, Mildford, MA, USA)
equipped with a Waters 2990 diode array detector, and a Symmetry C-18 (5 lm) (Waters, Milford, MA, USA) column (3.9 
150 mm) were used. The solvent system consisted of 1% aqueous
formic acid (A) and 1% formic acid in acetonitrile (B). The initial
composition of the mobile phase was 95% A and 5% B. With linear
gradients the composition changed to 75% A and 25% B within
45 min, and 50% A and 50% B within 60 min. The ow rate was
0.8 ml/min. Semipreparative HPLC was performed in the same
equipment using a Waters Spherisorb S10 ODS2 column
(10  250 mm).
2.3. HPLCMS/MS analysis
HPLC analysis was performed on an Agilent 1100 equipment
(Agilent Technologies, Santa Clara, CA, USA). Extracts (20 lL) were
injected into a 5 l C-18 column (250  4.6 mm). The mobile phase
consisted of 1% formic acid in water (A), and 1% formic acid in acetonitrile (B). The gradient was from 5% to 50% mobile phase B at a
ow rate of 0.8 ml/min over a 60 min period. For HPLC/MS analyses, the HPLC apparatus was interfaced to an ESI-IT Esquire 4000
(Bruker Daltonics, Billerica, MA, USA) mass spectrometer equipped
with an ionisation electrospray chamber. Conditions for the mass
spectra analysis in the positive ion mode included a voltage of
4000 V, a nebulising pressure of 35 psi, a drying nitrogen ow of
9.0 ml/min and a temperature of 325 C (Santos & Metzger,
2006). Data were collected on a full scan mode over a mass range
of m/z 202200.

2.4.1. Spectrophotometric analysis of anthocyanins

The free and oligomeric anthocyanins in extracts were determined by the differential pH method, according to Wang and Lin
(2000). The absorbance was measured at 700 and 250 nm in a
Genesys 5 (ThermoSpectronic, Waltham, MA, USA) spectrophotometer, at pH 1.0 and 3.5. The anthocyanins were quantied as
malvidin-3-O-glucoside
using
the
extinction
coefcient
( = 28,000). Determination was conducted in triplicate. The results
are presented as mean standard error followed by Students ttest. Differences were considered to be statistically signicant if
P < 0.05.
2.4.2. Isolation of malvidin-3-O-glucoside and peonidin-3-O-glucoside
The major components of Cabernet Sauvignon 2008 vintage red
wine were identied by semi-preparative HPLC as previously described (Aguirre et al., 2010).
2.4.3. ESI-MS Monitoring of reactions of anthocyanins
ESI-MS and ESI-MS/MS analyses were conducted in a high resolution hybrid quadrupole (Q) and orthogonal time-of-ight
(TOF) mass spectrometer (Q-TOF Micro, Micromass, UK) with a
constant nebulizer temperature of 100 C. The ESI source and the
mass spectrometer were operated in the positive-ion mode, and
the cone and extractor potentials were set to 10 and 4.5 V, respectively, with a scan range of m/z 803000. The samples were directly
infused into the ESI source at ow rates of 510 ll min 1 by means
of a microsyringe pump. The MS/MS experiments were carried out
by mass selection of a specic ion in Q1 which was then submitted
to collision-induced dissociation (CID) with argon in the collision
chamber. The product-ion MS analysis was accomplished with
the high-resolution orthogonal TOF analyser (Santos & Metzger,
2006).
2.5. Reaction between peonidin-3-O-glucoside and pyruvic acid
To a solution of peonidin-3-O-glucoside (2 mg) in 12% aqueous
ethyl alcohol, pyruvic acid (40 mg) was added (Fulcrand et al.,
1998). The reaction mixture was adjusted to pH 2 in 12% ethyl
alcohol in water, and stirred at 40 C for 35 h. During this period
of time, ESI-MS monitoring of the reaction mixture was carried
out by taking aliquots every hour during the rst 12 h period,
and then at 35 h, and by direct analysis of the species that have
been produced and consumed along the reaction (Santos & Metzger, 2006).
2.6. Reaction between malvidin-3-O-glucoside and acetaldehyde
To a solution of malvidin-3-O-glucoside (2 mg) in 12% ethyl
alcohol in water, acetaldehyde (0.5 ml) was added, and the pH
was adjusted to 2. The reaction mixture was stirred at 40 C during
48 h (Atanasova et al., 2002). During this period of time, ESI-MS
monitoring of the reaction mixture was carried out by taking 12
aliquots and by direct analysis of species that have been produced
and consumed along the reaction (Santos & Metzger, 2006).
2.7. Voltammetric experiments

2.4. Extraction of anthocyanins and reaction monitoring by ESI-MS

A sample of Cabernet Sauvignon red wine (700 ml) was concentrated in vacuo to 200 ml, and extracted successively with the same

Electrochemical measurements were performed on a CHI 104

Scanning Electrochemical microscope, CHI 900B (CH Instruments,
Austin, TX, USA) at room temperature. The working electrode

516

M.J. Aguirre et al. / Food Chemistry 129 (2011) 514519

was glassy carbon, the counter electrode was a platinum wire, and
the reference electrode was Ag/AgCl. Prior to each measurement
the working electrode was polished with 0.3 lm alumina powder.
A wine model solution consisting of 0.1 M NaCl in 12% aqueous
ethanol was used. Malvidin-3-O-glucoside and the wine extracts
were dissolved in the model wine solution at concentrations ranging from 0.2 to 1.0 mg/ml. The pH was adjusted to 3.6 with 1 M
NaOH and the solution was purged with nitrogen for 20 min. The
electrochemical measurements were conducted also in pH 3.6 acetateacetic acid buffer prepared by mixing 10.9 ml of 0.1 M sodium
acetate and 89.1 ml 0.1 M acetic acid, and in pH 3.6 acetate-acetic
acid buffer solution containing 12% ethyl alcohol (10.9 ml of 0.1 M
sodium acetate and 89.1 ml 0.1 M acetic acid containing 13.5%
ethyl alcohol). Linear voltammetry measurements were performed
at a scan rate of 5 mV/s in the range of 0.21.0 V (vs Ag/AgCl). The
cyclic voltammograms were acquired in the range of 0+0.8 V at a
scan rate of 0.1 V/s at 0.001 V intervals. Ascorbic acid (0.050.1 mg/
ml) was used as reference substance. Differential pulse voltammetry measurements were conducted with a pulse amplitude of
50 mV and a pulse width of 50 ms (Aguirre et al., 2010). Experiments were run in triplicate.

3. Results and discussion

3.1. Anthocyanins characterisation
The anthocyanin contents in 6, 7 and 8 year old Cabernet Sauvignon red wine from the Central Valley of Chile were studied. It is
shown in Table 1 that the content of free anthocyanins decreases
signicantly with age, being almost half of the amount in young
wine at seven years of storage in bottles. Fig. 2 shows the HPLC
chromatograms of some wine extracts. The elution prole corresponding to one year old sample is very similar to that previously
reported for Cabernet Sauvignon red wine (Aguirre et al., 2010);
the main peak is assigned to malvidin-3-O-glucoside. At highest
retention time very small amounts of complex anthocyanins were
detected, indicating that in the young wine, the reaction of anthocyanins with other compounds has already taken place. In order to
analyse the formation of new anthocyanins-derived compounds in
red wine, we tried the formation of condensation products through
the reaction between peonidin-3-O-glucoside and pyruvic acid in a
model wine solution. In this work, we applied off-line monitoring
ESI-MS(/MS) of anthocyanins reactions with pyruvic acid and acetaldehyde. Since ESI normally releases ions preformed in solution,
we expected that species from reactions should be detectable by
ESI-MS in the reacting solution. ESI is also known for the mild conditions used to form the gaseous ions; even very labile molecules
can be transferred to the gas phase (Santos, 2008, 2010). Analysis
of the reaction of peonidin-3-O-glucoside and pyruvic acid by
ESI/MS indicated that the original anthocyanin (at m/z 463) after
35 h of reaction disappeared, with the formation of a new
compound at m/z 531. This molecular ion corresponds to the
compound obtained in the cycloaddition of pyruvic acid to peoni-

Table 1
Anthocyanins composition in red wine extracts.
Red wine extracts

Eight year old

Seven year old
Six year old
One year old

Anthocyaninsa
Total

Polymeric

Free

% Free anthocyanins

73.21 2.0
69.21 5.3
70.21 4.5
75.55 3.50

51.25 1.9
44.29 1.5
42.13 2.5
22.67 3.9

21.96
24.92
28.08
52.88

30
36
40
70

a
Anthocyanins content expressed as milligrams of malvidin-3-O-glucoside per
100 ml of wine.

Fig. 2. HPLCMS chromatograms of Cabernet Sauvignon red wine extracts. (A) eight
year old, (B) one year old.

din-3-O-glucoside known as vitisin A of peonidin-3-O-glucoside

(Fulcrand et al., 1998). It has been previously reported in red wines
(Alcalde-Eon et al., 2007; Atanasova et al., 2002; Pati et al., 2006).
On the other hand, the reaction of malvidin-3-O-glucoside and
acetaldehyde in wine model solution showed, by ESI/MS analysis
after 48 h, the formation of a malvidin-3-O-glucoside dimer mediated by acetaldehyde, with a molecular ion at m/z 1012. In this
work, both condensation products were detected in aged Cabernet
Sauvignon red wines.
The anthocyanins content in aged Cabernet Sauvignon and
young red wines was analysed by HPLCMS. In the chromatograms
of aged wines (Fig. 2) the presence of vitisin A of peonidin-3-O-glucoside (m/z 531), was easily recognised, although the most abundant compound is vitisin A of malvidin-3-O-glucoside (peak 15)
(Fig. 1), with a molecular ion at m/z 561 and a fragment at m/z
399 (M+ glucose) (Alcalde-Eon et al., 2006; He, Santos-Buelga,
Silva, Mateus, & de Freitas, 2006). Peak 4 (Fig. 2) in the elution
prole of young Cabernet Sauvignon red wine corresponds to
malvidin-3-O-glucoside, with a molecular ion at m/z 493 and a
fragment at m/z 331 due to the aglycone. It is seen in Fig. 2 that
the peak due to malvidin-3-O-glucoside gradually decreases with
age while peak 15 increases with age from 10% in young wine to
39% in eight year old wine. Similar results were found by Mateus
and de Freitas (2001) on the evolution of free anthocyanins in Port
wines; however, they detected also degradation of anthocyanin
pyruvic acid adducts during storage in oak barrels. Table 2 presents
assignments of the most important peaks in HPLCMS of aged
wines through interpretation of MS/MS spectra and literature data
(Alcalde-Eon, Escribano-Bailn, Santos-Buelga, & Rivas-Gonzalo,
2007; Pati, Liberatore, Gambacorta, Antonacci, & La Notte, 2009).
The glucosides of peonidin, delphinidin and petudinin are present,
they showed similar fragmentation patterns, the most important
one being the loss of glucose moiety. Also acetyl, coumaryl and
caffeyl-glucosides were detected. Pati et al. (2006) reported the
formation in young red wine of condensation products of malvidin-3-O-glucoside with catechin and epicatechin mediated by
ethyl or vinyl bridges. The resolution of the chromatograms over
50 min is low, which indicates that compounds with molecular
weights between m/z 900 and 1500 are co-eluting (Fig. 2). Among
them various dimers formed by condensation between two different anthocyanins, such as dimers of malvidin-3-O-glucoside and
cyanidin-3-O-glucoside (at m/z 941), malvidin-3-O-glucoside
and peonidin-3-O-glucoside (at m/z 955), malvidin-3-O-glucoside
and petunidin-3-O-glucoside (at m/z 971) were identied. The

517

M.J. Aguirre et al. / Food Chemistry 129 (2011) 514519

Table 2
Anthocyanins identied in young and aged red wine extracts by HPLCMS/MS. Asterik symbol () denotes presence of anthocyanin in wine sample.
Peak

Retention time (min)

[M]+ (m/z)

Product ion 1

Product ion 2

Anthocyanin

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

17.517.8
22.323.3
25.526.2
26.427.7
29.930.8
32.132.4
32.532.7
32.832.9
33.133.3
33.333.4
33.433.5
33.633.8
339
33.934.0
34.134.8
34.935.0
35.135.6
35.835.9
36.036.3

465
479
463
493
507
535
611
595
625
609
639
627
547
531
561
707
581
579
609
651
727
741
625
551
565
753
783
767
797
943
751
781

303
317
301
331
303
331
303
287
317
301
331
303
385
369
399
399
419
417
447
447
419
433
301
317
331
591
621
605
635

257

Delphinidin-3-O-glucoside
Petunidin-3-O-glucoside
Peonidin-3-O-glucoside
Malvidin-3-O-glucoside
Delphinidin-3-O-acetylglucoside
Malvidin-3-O-acetylglucoside
Delphinidin-3-O-coumaryl-glucoside
Cyanidin-3-O-coumaryl-glucoside
Petudidin-3-O-coumarylglucoside o
Peonidin-3-O-coumarylglucoside
Malvidin-3-O-coumarylglucoside
Delphinidin-3-O-caffeylglucoside
Visitin A of petunidin-3-O-glucoside
Visitin A of peonidin-3-O-glucoside
Visitin A
Visitin A of malvidin-3-O-coumarylglucoside
Adduct 4-vinylphenol-delphinidin-3-O-glucoside
Adduct 4-vinylphenol-peonidin-3-O-glucoside
Adduct 4-vinylphenol-malvidin-3-O-glucoside
Adduct 4-vinylphenol-malvidin-3-O-acetylglucoside
Adduct 4-vinylphenol-delphinidin-3-O-acetyl-glucoside
Adduct 4-vinylphenol-petunidin-3-O-coumarylglucoside
Peonidin-3,7-O-diglucoside
petunidin-3-O-glucoside + lactic acid
Malvidin-3-O-glucoside + lactic acid
Dimer delphinidin-3-O-glucoside
Adduct gallocatechin-petunidin-3-O-glucoside
Adduct gallocatechin-peonidin-3-O-glucoside
Adduct epigallocatechin-malvidin-3-O-glucoside
Adduct gallocatechin-malvidin-3-O-glucoside
Adduct epicatechin-peonidin-3-O-glucoside
Adduct catechin-malvidin-3-O-glucoside

Age, year old

1

40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0
40.055.0

286
299

589
619

presence of malvidin-3-O-glucoside dimer mediated by acetaldehyde (at m/z 1012) was probed to be formed by performing reactions of pure malvidin-3-O-glucoside with acetaldehyde, in the
condensed phase, followed by direct ESI-MS monitoring of the
mixture with time. The structure of the dimer corresponds to a
condensation compound in which two molecules of malvidin-3O-glucoside (MW 493) are linked through positions C-8 of B rings
by an ethyl bridge. Furthermore, two trimers, epicatechincyanidin-3-O-glucoside-malvidin-3-O-glucoside (at m/z 1229) and
epicatechin-delphinidin-3-O-glucosidemalvidin-3-O-glucoside (at
m/z 1245) were also detected. Oligomeric anthocyanins in red wine
are formed by condensation of a avanol, e.g. epicatechin, through
position C4 with a dimeric anthocyanin. One of the subunits is
always malvidin-3-O-glucoside, whereas the other anthocyanin
could be any of the monomeric anthocyanins (Alcalde-Eon,
Escribano-Bailn, Santos-Buelga, & Rivas-Gonzalo, 2007).

eight year old

seven year old
six year old
one year old

3.2. Electrochemical behaviour of wine extracts

In a previous study it was suggested that pulse differential voltammetry was a good electrochemical technique for the study of
the antioxidant capacity of red wines (Aguirre et al., 2010).
Ragubeer, Beukes, and Limson (2010) found that voltammetric
methods represent suitable assays for rapid screening of antioxidant capacity in plant extracts. Here, we compare the antioxidant
capacity of aged Cabernet Sauvignon by electrochemical methods.
Fig. 3 shows the differential pulse voltammograms of aged wine
extracts. Six, seven and eight year red wines present only one
oxidation peak at approximately 400, 436 and 436 mV, respectively. The curves are not well resolved, probably due to the large
number of condensation products present in aged wine. It was previously found that malvidin-3-O-glucoside presented two oxidation
peaks at 483, and 698 mV (Aguirre et al., 2010). Fig. 4 presents the

0.0

0.2

0.4

0.6

0.8

1.0

E/V vs Ag/AgCl
Fig. 3. Differential pulse voltammograms of Cabernet Sauvignon red wine extracts
in model wine solution (12% ethanol, 0.1 M NaCl, and added NaOH to pH 3.6) of red
wine extracts. Arrows indicate the oxidation peaks.

linear voltammograms of young and aged wines. For young wine,

the rst oxidation peak appears at 401 mV, this value is more negative than the value previously found for Cabernet Sauvignon red
wine (435 mV). This may be explained by the fact that the latter
contained 16% less free anthocyanins. For 6, 7 and 8 year old wines,
values are 436, 443, and 451 mV, respectively. According to
Simic, Manojlovic, egan, and Todorovic (2007) and Zienlinska,
Wiczkowski, and Konrad (2008), as the oxidation potential

518

M.J. Aguirre et al. / Food Chemistry 129 (2011) 514519

wine. Free anthocyanins decreases signicantly with age, with the

concomitant increase in condensed products.
The condensation products of anthocyanins in aged Cabernet
Sauvignon red wines are responsible of their lower antioxidant
capacity in relation to young wines.
Voltammetric methods represent suitable assays for the analysis of the antioxidant capacity of red wine samples in ethyl alcoholwater solutions. Also, they may be use to differentiate
between young and aged red wine samples.

-5

1.6x10

eight year old

seven year old
six year old
one year old
d l wine
i solution
l i
model

-5

I//A

1 2x10
1.

-6

8.0x10

-6

Acknowledgements

4.0x10

The nancial support of Programa Bicentenario en Ciencia y

Tecnologa, CONICYT-Banco Mundial, Proyecto Anillo ACT-24 is
gratefully acknowledged. S. Torres thanks CONICYT for a doctoral
fellowship. L.S.S thanks FONDECYT (Project 1085308) for support
research activity.

0.0
0
0.0

0.2

0.4

0.6

0.8

E/V vs Ag/AgCl
Fig. 4. Linear voltammograms of Cabernet Sauvignon red wine extracts in model
wine solution (12% ethanol, 0.1 M NaCl, and added NaOH to pH 3.6) of red wine
extracts. Arrows indicate the oxidation peaks.

eight year old

seven year old
ld
six
i year old
ld
one year old

460

450

E// mV
V

440

430

420

410
-2.9
29

-2.8
28

-2.7
27

-2.6
26

25
-2.5

-2.4
24

-2.3
23

log I/A
Fig. 5. Tafels plots for the electrooxidation of Cabernet Sauvignon red wine
extracts in 12% ethyl alcohol solutions.

becomes more positive the antioxidant capacity decreases. In order

to know if the oxidation processes in aged wines are similar to those
of young wines, polarisation curves were registered at low speed
and the Tafels curves were calculated (Brett & Oliveira-Brett,
1993). Fig. 5 shows the calculated Tafels curves of young and aged
wines. It can be seen that 7 and 8 year wines show similar slopes.
Six, seven and eight year old extracts show Tafels plots of 234,
177 and 188 mV/dec, respectively. The values of Tafels plots for
eight and seven year old samples could be considered close to
120 mV/dec value corresponding to one electron transfer as the rate
determining step and formation of a radical. The value for the young
wine (523 mV/dec) is indicative that the electrochemical process is
not the same, probably due to the high content of free anthocyanins, which polymerised on the glassy carbon electrode. The value
for 6 year old wine is higher than the expected value which may
be due to the presence of higher amount of free anthocyanins in
relation to 7 and 8 year samples, that also, polymerised on the
electrode.
4. Conclusions
The anthocyanins content in 6, 7 and 8 year old Cabernet Sauvignon
red wine stored in bottles are very different from the content in young

References
Aguirre, M. J., Chen, Y. Y., Isaacs, M., Matsuhiro, B., Mendoza, L., & Torres, S. (2010).
Electrochemical behaviour and antioxidant capacity of anthocyanins from
Chilean red wine, grape and raspberry. Food Chemistry, 121, 4448.
Alcalde-Eon, C., Escribano-Bailn, M. T., Santos-Buelga, C., & Rivas-Gonzalo, J. C.
(2006). Changes in the detailed pigment composition of red wine during
maturity and ageing: A comprehensive study. Analytica Chimica Acta, 563,
238254.
Alcalde-Eon, C., Escribano-Bailn, M. T., Santos-Buelga, C., & Rivas-Gonzalo, J. C.
(2007). Identication of dimeric anthocyanins and new oligomeric pigments in
red wine by means of HPLCDADESI/Msn. Journal of Mass Spectrometry, 42,
735748.
Atanasova, V., Fulcrand, H., Le Guernev, C., Cheynier, V., & Moutounet, M. (2002).
Structure of a new dimeric acetaldehyde malvidin 3-glucoside condensation
product. Tetrahedron Letters, 43, 61516153.
Bakker, J., Bridle, P., Honda, T., Kuwano, H., Saito, N., Terahara, N., et al. (1997).
Identication of an anthocyanin occurring in some red wines. Phytochemistry,
44, 13751382.
Bakker, J., & Timberlake, C. F. (1997). Isolation, identication, and characterization
of new color-stable anthocyanins occurring in some red wines. Journal of
Agricultural and Food Chemistry, 45, 3543.
Brett, C. A., & Oliveira-Brett, A. M. (1993). Electrochemistry principles, methods and
applications. New York: Oxford University Press (pp. 102126).
Brouillard, R., Chassaing, S., & Fougerousse, A. (2003). Why are grape/fresh wine
anthocyanins so simple and why is it that red wine color lasts so long?
Phytochemistry, 64, 11791186.
Dallas, C., Ricardo-da-Silva, J. M., & Laureano, O. (1996). Products formed in model
wine solutions involving anthocyanins, procyanidin B2, and acetaldehyde.
Journal of Agricultural and Food Chemistry, 44, 24022407.
Escribano-Bailn, T., Alvarez-Garca, M., Rivas-Gonzalo, J. C., Heredia, F. J., & SantosBuelga, C. (2001). Color and stability of pigments derive from the acetaldehydemediated condensation between malvidin 3-O-glucoside and (+)-catechin.
Journal Agricultural and Food Chemistry, 49, 12131217.
Francia-Aricha, E. M., Guerra, M. T., Rivas-Gonzalo, J. C., & Santos-Buelga, C. (1997).
New anthocyanins pigments formed after condensation with avanols. Journal
of Agricultural and Food Chemistry, 45, 22622266.
Fulcrand, H., Bernabdeljalil, C., Rigaud, J., Cheynier, V., & Moutounet, M. (1998). A
new class of wine pigments generated by reaction between pyruvic acid and
grape anthocyanins. Phytochemistry, 47, 14011407.
Ghiselli, A., Nardini, M., Baldi, A., & Scaccini, C. (1997). Antioxidant activity of
different phenolic fractions separated from an Italian red wine. Journal of
Agricultural and Food Chemistry, 46, 361367.
Giusti, M. M., Rodrguez-Sanoa, L. E., Grifn, D., & Wrolstad, R. E. (1999).
Electrospray and tandem mass spectroscopy as tools for anthocyanin
characterization. Journal of Agricultural and Food Chemistry, 47, 46574664.
Hayasaka, Y., & Asenstorfer, R. E. (2002). Screening for potential pigments derived
from anthocyanins in red wine using nanoelectrospray tandem mass
spectrometry. Journal of Agricultural and Food Chemistry, 50, 756761.
He, J., Santos-Buelga, C., Silva, A. M. S., Mateus, N., & de Freitas, V. (2006). Isolation
and structural characterization of new anthocyanin-derived yellow pigments in
aged red wines. Journal of Agricultural and Food Chemistry, 54, 95989603.
Huang, D., Ou, B., & Prior, R. (2005). Chemistry behind antioxidant capacity assay.
Journal of Agricultural and Food Chemistry, 53, 18411856.
Mateus, N., Silva, A. M. S., Vercauteren, J., & De Feitas, V. (2001). Occurrence of
anthocyanins derived pigments in red wines. Journal of Agricultural and Food
Chemistry, 49, 48364840.
Mateus, N., & de Freitas, V. (2001). Evolution and stability of anthocyanins-derived
pigments during Port wine aging. Journal of Agricultural and Food Chemistry, 49,
52175222.

M.J. Aguirre et al. / Food Chemistry 129 (2011) 514519

Monagas, M., Gomez-Cordoves, C., & Bartolome, B. (2005). Evolution of polyphenols
in red wines from Vitis vinifera L during aging in the bottle. I. Anthocyanins and
pyranoanthocyanins. European Food Research and Technology, 220, 607614.
Nave, F., Teixeira, N., Mateus, N., & De Freitas, V. (2010). The fate of avanolanthocyanin adducts in wines: Study of their putative reaction patterns in the
presence of acetaldehyde. Food Chemistry, 121, 11291138.
Pati, S., Liberatore, M. T., Gambacorta, G., Antonacci, D., & La Notte, E. (2009). Rapid
screening for anthocyanins and anthocyanin dimers in crude grape extracts by
high performance liquid chromatography coupled with diode array detection
and tandem mass spectrometry. Journal of Chromatography A, 1216, 38643868.
Pati, S., Losito, I., Gambacorta, G., La Notte, E., Palmisano, F., & Zambonin, P. G.
(2006). Simultaneous separation and identication of oligomeric procyanidins
and anthocyanin-derived pigments in raw red wine by HPLC-UVESI-MS.
Journal of Mass Spectrometry, 41, 861867.
Piljac, J., Martnez, S., Valek, L., Stipcevic, T., & Kovacevic Ganic, K. (2005). Phenolic
content and antioxidant activity of Croatian wines. Food Technologies and
Biotechnology, 43, 271276.
Piovan, A., Filippini, R., & Favretto, D. (1998). Characterization of the anthocyanins
of Catharantus roseus (L.) G. Don in vivo and in vitro by electrosspray ionization
ion trap mass spectrometry. Rapid Communications Mass Spectrometry, 12,
361367.
Ragubeer, N., Beukes, D. R., & Limson, J. L. (2010). Critical assessment of
voltammetry for rapid screening of antioxidants in marine algae. Food
Chemistry, 121, 227232.
Remy-Tanneau, S., Le Guernev, C., Meudec, E., & Cheynier, V. (2003).
Characterization of a colorless anthocyanin-avan-3-ol dimer containing both
carbon-carbon and ether interavanoid linkages by NMR and mass
spectrometry. Journal of Agricultural and Food Chemistry, 57, 35923597.
Rivas-Gonzalo, J. C., Bravo-Haro, S., & Santos-Buelga, C. (1995). Detection of
compounds formed through the reaction of malvidin 3-monoglucoside and

519

catechin in the presence of acetaldehyde. Journal of Agricultural and Food

Chemistry, 43, 14441449.
Salas, E., Fulcrand, H., Meudec, E., & Cheynier, V. (2003). Reactions of anthocyanins
and tannins in model solutions. Journal of Agricultural and Food Chemistry, 51,
79517961.
Santos, L. S. (2008). Online mechanistic investigations of catalyzed reactions by
electrospray ionization mass spectrometry: A tool to intercept transient species
in solution. European Journal of Organic Chemistry, 235, 253.
Santos, L. S. (2010). Reaction intermediates: MS investigations in solution. Weinheim:
Wiley-VCH.
Santos, L. S., & Metzger, J. O. (2006). Study of homogeneously catalyzed Ziegler
Natta polymerization of ethene by ESI-MS. Angewandte Chemie International
Edition, 45, 977981.
Simic, A., Manojlovic, D., egan, D., & Todorovic, M. (2007). Electrochemical
behavior and antioxidant and prooxidant activity of natural phenolics.
Molecules, 12, 23272340.
Vivar-Quintana, A. M., Santos-Buelga, C., & Rivas-Gonzalo, J. C. (2002). Anthocyaninderived pigments and colour of red wines. Analytica Chimica Acta, 458, 147155.
Wang, S. Y., & Lin, H.-S. (2000). Antioxidant activity in fruits and leaves of
blackberry, raspberry, and strawberry varies with cultivar and development
stage. Journal of Agricultural and Food Chemistry, 48, 140146.
Wang, J., & Sporns, P. (1999). Analysisi of Anthocyanins in red wine and fruit juice
using MALDI-MS. Journal of Agricultural and Food Chemistry, 47, 20092015.
Wang, H., Race, E. J., & Shrikhande, A. J. (2003). Anthocyanin transformation in
Cabernet Sauvignon wine during aging. Journal of Agricultural and Food
Chemistry, 51, 79897994.
Zienlinska, D., Wiczkowski, W., & Konrad, M. (2008). Determination of the relative
contribution of quercetin and its glucoside to the antioxidant capacity of onion
by cyclic voltammetry and spectrophotometric methods. Journal of Agricultural
and Food Chemistry, 56, 35243531.