Phenolics, Aroma Profile, and In Vitro Antioxidant Activity of Italian Dessert Passito Wine from Saracena (Italy)

Phenolics, Aroma Profile, and In Vitro
Antioxidant Activity of Italian Dessert
Passito Wine from Saracena (Italy)
Monica R. Loizzo, Marco Bonesi, Giuseppe Di Lecce, Emanuele Boselli, Rosa Tundis, Alessandro Pugliese, Francesco Menichini,

                                                                                                                                                                   C: Food Chemistry
and Natale Giuseppe Frega

    Abstract: A traditional sweet dessert wine from Saracena (Italy), made with nonmacerated local white grapes (Guar-
    naccia, Malvasia and Moscato), was analyzed for phenolics and aroma profile and antioxidant activities. The most
    abundant classes of phenols identified by high-performance liquid chromatography were hydroxybenzoic acids and
    flavan-3-ols, where gallic acid showed the highest content (376.5 mg/L). The analysis by solid phase microextraction-gas
    chromatography-mass spectrometry revealed the presence of superior alcohols (from iso-butanol and iso-amyl alcohol up
    to 2-phenylethanol) and their ethyl esters, terpenes (such as linalool), furfuryl compounds, and free fatty acids (up to
    palmitic acid) as the key odorants of this wine. The antioxidant activity, evaluated by different in vitro assays 2,2-diphenyl-1-
    picrylhydrazyl (DPPH), 2,2 -azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS), and β-carotene bleaching test),
    showed that passito wine had a radical scavenging activity (IC50 value of 0.03 v/v against DPPH·) and inhibited linoleic
    acid oxidation with an IC50 value of 0.4 v/v after 30 min of incubation.

    Keywords: aroma, antioxidant activity, dessert wine, phenolics, HPLC tandem MS, SPME-GC-MS

    Practical Application:   It is well known that moderate consumption of wine is actually recommended since it appears
    associated with a decreased incidence of several diseases. Passito of Saracena, a well-appreciated Italian dessert wine,
    demonstrated an interesting antioxidant activity. Moreover, the aroma profile contributed and defined the chemical
    markers of the quality of this wine and their quantitative ranges, which are needed to assess the authenticity of local niche
    production claiming a quality designation.

Introduction                                                                             2000). In Italy, 3 different categories of dessert wine are pro-
   According to our new report entitled “US Wine Market Fore-                            duced: passito wines (Passito di Pantelleria and Vinsanto); Port-
cast to 2012” (2010), the U.S. wine market is one of the fastest                         like wine (Marsala), and sparkling sweet wines (Brachetto d’Acqui
growing markets in the world that at present accounts for around                         or Vernaccia di Serrapetrona). These wines are consumed also
12% of the global wine consumption. This market is expected                              as aperitif, to accompany aged cheese and blue cheese, and in
to grow about 3% during 2010 to 2013. The market is domi-                                cooking.
nated by table or still wine, which accounts for nearly 90% of                              The traditional way to produce passito in Calabria (Italy) in-
the market while dessert and sparkling wine together hold just                           volves picking grape bunches and place them on a cannizza and
over 10% market share. However, some experts believe that there                          allowing grapes to dry in the sun for 10 to 15 d but covered dur-
is potential for expanding this category market by re-educating                          ing the night. This process called appassimento (raisining) generally
and re-introducing consumers to high-quality products. Dessert                           increases the sugar content up to 30% to 40%. Some grape juice is
wines are distributed worldwide and some of the most impor-                              also left out in a jar for a few days. Then the dried grapes are placed
tant dessert wines are Beerenauslesen and Trockenbeerenauslesen                          in a jar for a few days and pressed. This sweet, slightly fermented
(Germany), Banyuls, Sauternes, Monbazillac, Jurancon, Maury,                             liquid is mixed with the juice that was also exposed to the sun.
and Rivesaltes (France), Sherry (Spain), Porto and Madeira (Por-                         The entire mixture is then closed in a jar and allowed to ferment
tugal). Leaving Europe for North America Canadian and Up-                                for 3 to 4 wk.
state New York ice wine were produced (Brooke 1987; Jackson                                 The effect of dehydration by sun exposure on Pedro Ximénez
                                                                                         grapes that are used to obtain a famous Andalusia sweet wine was
                                                                                         investigated by Peinado and others (2009) and Lopez de Lerma and
MS 20121784 Submitted 12/27/2012, Accepted 02/18/2013. Authors Loizzo,
Bonesi, Tundis, Pugliese, and Menichini are with Dept. of Pharmacy, Health Sciences      others (2012). Grapes subjected to this process undergo significant
and Nutrition, Univ. of Calabria, 87036 Rende (CS), Italy. Authors Lecce, Boselli,       modifications in terms of higher sugar concentration, change in
and Frega are with Dept. of Agricultural, Food and Environmental Sciences, Marche        volatile and phenolics profile, and enzymatic activity. In partic-
Polytechnic Univ., via Brecce Bianche, 60131 Ancona, Italy. Direct inquiries to author   ular, sun drying enriches musts in phenolics thus increasing the
Loizzo (E-mail:
                                                                                         antioxidant activity.
This work is dedicated to the memory of Professor Francesco Orlandi (1927                   In Europe, this production is enforced by the Commission
to 2012), who was an inspiration to scientists and played a key role in alcohol          Regulation (EC) No 606/2009 and its subsequent modifica-
research, not only in Italy, but also in the international arena.                        tions. Passito of Saracena (Italy) has often been awarded as a


C 2013 Institute of Food Technologists

doi: 10.1111/1750-3841.12110                                                                           Vol. 78, Nr. 5, 2013 r Journal of Food Science C703
Further reproduction without permission is prohibited
Passito chemistry and bioactivity . . .

                    high-quality Italian dessert wine. The must is obtained by crush-       Wine samples
                    ing autochthonous Malvasia and Guarnaccia overripe grapes. The             The survey was carried out on Passito of Saracena wine (5
                    special aroma and taste also derive from Moscato grapes, which          samples) aged from 1 to 4 y. All samples were acquired directly
                    are collected and dried a few weeks before harvest. The Moscato         from the wineries. All bottles were stored in the dark (10 ◦ C) and
                    grape berries are dried, selected, crushed, and then manually           analyzed immediately after opening. A sample aliquot (50 mL)
                    added to the first-pressing must. After a long and slow fermenta-       was taken and mixed with equal aliquots of the others to obtain a
                    tion, a wine with sweet amber and golden reflections, intense           single sample for analysis.
C: Food Chemistry

                    aroma, and the taste of honey, dried figs, and exotic fruits is
                    obtained.                                                               Passito wine analyses
                       The aroma profile of wine, also called bouquet, is determined by        The chemical parameters (pH, total acidity, volatile acidity, re-
                    alcohols, aldehydes, esters, acids, monoterpenes, and other minor       ducing sugars, alcohol percentage, ash, dry extract, sulfates, and
                    components such as sulfur and nitrogen compounds which are              chloride) were determined according to the Commission Reg-
                    already present in the grapes or are formed during the fermen-          ulation (EEC) No 2676/90. Sugars (glucose and fructose) were
                    tation process and storage, and participate to the overall pleasant     quantified by HPLC (Thermo Electron Corp., Waltham, Mass.,
                    flavor perceived on drinking. The aroma profile is influenced by        U.S.A.) equipped with a UV100 set to 210 nm and a RI-150. The
                    environment, ripeness, and grape variety, as well as winemaking         analyses were performed isocratically at 0.8 mL/min and 65 ◦ C
                    procedures (Verzera and others 2008). Recently, solid phase mi-         with a 300 mm × 7.8 mm i.d. cation-exchange column (Aminex
                    croextraction (SPME), which allows to determine the low-level           HPX-87H) and a Cation H+ Microguard cartridge (Bio-Rad Lab-
                    components in complex mixtures, has been used to study the wine         oratories, Hercules, Calif., U.S.A.) using 0.003 N H2 SO4 as mo-
                    aroma compositions (Verzera and others 2008).                           bile phase (Gerbi and Tortia 1991). The total phenolic content was
                       Several phenolic compounds are present in wine and their con-        evaluated by using the Folin–Ciocalteu method (Menichini and
                    centrations depend on grape variety, climatic conditions, and,          others 2009) and expressed as mg/L (+)-catechin, while proan-
                    of course, viticultural and winemaking techniques. These com-           thocyanidins were spectrophotometrically determined according
                    pounds are responsible for wine color and also flavor and astrin-       to Di Stefano and others (1989). Moreover, color density, poly-
                    gent taste (Lesschaeve and Noble 2005). Also, they manifest a           meric color, and percent of polymeric color were determined
                    wide range of beneficial health effects including antioxidant activ-    according to the methodology previously reported by Giusti and
                    ity (Boselli and others 2009). Pellegrini and others (2003) stated      Wrolstad (2001).
                    that to correctly investigate antioxidant activity several methods
                    must be applied.                                                        HPLC-UV and mass spectrometry conditions
                       This study was designed to gain knowledge of an Italian Passito         The determination of low molecular weight phenolics in pas-
                    wine produced in the area of Saracena (Calabria, Italy). For this       sito wine was carried out using RP-HPLC–diode array detector
                    purpose, the high-performance liquid chromatography (HPLC)              (DAD). The HPLC ternary pump was a mod. 9010 from Var-
                    phenolics profile, SPME-gas chromatography-mass spectrometry            ian (Middelburg, the Netherlands). A Chrompack (Middelburg)
                    (SPME-GC-MS) of the volatile fraction, and determination of             25 cm × 4.6 mm i.d. column packed with Chromosphere C18
                    antioxidant activity through different in vitro assays (2,2-diphenyl-   (5 μm particle size) was used. The separation of phenolic com-
                    1-picrylhydrazyl [DPPH], ABTS, and β-carotene bleaching test)           pounds was carried out in 45 min under the following conditions:
                    were performed. The experimental results derived from this inves-       at 0 min, mobile phase A (formic acid 4.5% v/v) was pumped
                    tigation are the premise to assess the commercial value and health      isocratically for 5 min; then B (acetonitrile at a flow rate of 0.7
                    quality of this rather neglected dessert wine. The data will con-       mL/min) was increased from 0% to 15% in 35 min; successively,
                    tribute to the preservation of traditional and local agro-food pro-     the mobile phase B reached 40% in 25 min and was held for 20
                    duction method and protection against the cheaper mass-produced         min. The HPLC system was coupled to a Varian Prostar 330 DAD
                    industrial sweet wines.                                                 and a LCQ-DUO ion-trap mass spectrometer (Thermoquest, San
                                                                                            José, Calif., U.S.A.). The chromatograms were monitored at 3
                    Materials and Methods                                                   wavelengths (280, 320, and 365 nm) to analyze each group of
                                                                                            compounds. Each wavelength was suitable for a peculiar class of
                    Chemicals                                                               compounds: 280 nm was used for hydroxybenzoic acids, flavan-
                       ABTS solution, acetate buffer, β-carotene, caffeic acid,             3-ols and procyanidin, 320 nm for hydroxycinnamic acids and
                    (+)-catechin, chlorogenic acid, p-coumaric acid, DPPH,                  their tartaric esters, and 365 nm for flavonols. Mass spectra were
                    EDTA, (–)-epicatechin, ferulic acid, ferrozine, gallic acid,            obtained using electro-spray negative and positive ionization in a
                    homovanillic acid, kaempferol-3-O-glucoside, L-ascorbic acid,           range of m/z 100 to 800 and the tandem mass experiments were
                    linoleic acid, 2-nonanone, propyl gallate, protocatechuic acid,         carried out with relative collision energy of 30% to 40%. All the
                    quercetin, quercetin-3-O-glucoside, tripyridyltriazine, trans-          data were acquired with the Excalibur software ver. 1.2 by Finni-
                    resveratrol, Trolox, butylated hydroxytoluene, Tween 20 and             gan. Individual compounds were quantified using a calibration
                    vanillic acid, were purchased from Sigma-Aldrich (Milan, Italy).        curve of the corresponding standard compound. When reference
                    cis-Resveratrol was obtained after exposure of the trans-isomer         compounds were not available, the calibration of structurally re-
                    standard to UV light (Cvejic and others 2010). SPME poly-               lated substances was used. The data are reported as the averages
                    dimethylsiloxane (PDMS) 100 μm fiber (model 57300-U) and                of 3 determinations (Santos-Buelga and Williamson 2003; Boselli
                    sealed 15 mL vials were purchased by SUPELCO (Bellefonte, Pa.,          and others 2006, 2008).
                    U.S.A.). Spectrophotometric determinations were carried out us-
                    ing a UV-vis Jenway 6003 spectrophotometer (Carlo Erba, Milan, Solid phase microextraction
                    Italy) with cuvettes of 1 cm length. Solvents were HPLC-grade      A SPME PDMS 100 μm fiber (model 57300-U, Supelco) was
                    and were purchased from VWR Intl. (Milan, Italy).                employed for the extraction of volatiles from the passito wine

                    C704 Journal of Food Science r Vol. 78, Nr. 5, 2013
Passito chemistry and bioactivity . . .

headspace according to the modified methodology previously de-         Table 1–Chemical and chromatic parameters of Passito wine.
scribed by Rodrı´guez-Bencomo and others (2003). In brief, each        Parameters                                Mean ± standard deviation
wine sample was sealed into a 15 mL vial (Supelco) and heated for
10 min at 60 ◦ C. Then the SPME fiber was inserted into the vial  Alcoholic percentage (ethanol, % vol)                  14.2   ±   0.6
                                                                  Total acidity (g/L as tartaric acid)                    6.9   ±   0.3
for 60 min at 60 ◦ C. Afterwards the fiber was inserted into the gas
                                                                  Volatile acidity (g/L as acetic acid)                   0.9   ±   0.07
chromatograph injector (15 min at 250 ◦ C).                       Reducing sugars (g/L)                                   116   ±   4.5
                                                                  Ashes (g/L)                                             2.1   ±   0.8

                                                                                                                                                 C: Food Chemistry
                                                                  Total dry extract                                       151   ±   2.8
GC-MS analysis                                                    Dry extract without sugar                              35.3   ±   0.8
   A Hewlett-Packard 6890 gas chromatograph equipped with a Sulfates (as K2 SO4 )                                         0.2   ±   0.05
DB-WAX capillary column (30 m length, 0.25 mm i.d., 0.25 μm Chloride (as NaCl)                                           0.06       0.008
                                                                  pH                                                      3.8   ±   0.03
film thickness) (Agilent Technologies, Palo Alto, Calif., U.S.A.) Total polyphenols (mg/L as (+)-catechin )               249   ±   5.8
and interfaced with a Hewlett Packard 5973 was used for analysis. Proanthocyanidins (mg/L as cyaniding chloride)         76.8   ±   1.9
Mass selective ionization of the sample components was performed Color density                                           3.06   ±   0.09
in electron impact mode (EI, 70 eV) using a m/z range of 30 to Polymeric color                                           2.63   ±   0.07
                                                                  % Polymeric color                                      86.1   ±   2.4
400 and a scan time of 3.89 s, with a flux of 0.8 mL/min of helium.
The sample was injected by placing the SPME fiber into the GC
inlet for 15 min in the splitless mode. The oven temperature
program started with an isothermal step of 5 min at 45 ◦ C. Then,
the temperature was raised to 220 ◦ C at a rate of 4 ◦ C/min and       Results and Discussion
kept at 220 ◦ C for 20 min. The injector temperature was set at        Physico chemical analyses
250 ◦ C and the detector at 280 ◦ C. Analyses were carried out            Mean values and SDs for each chemical parameter determined
in triplicate. GC retention times were used for identification of      on passito wine are reported in Table 1. Passito wine showed a total
the compounds using the Wiley 138, Wiley 275, or NIST 98               acidity of 6.9 g/L as tartaric acid and a volatile acidity of 0.9 g/L;
libraries. 2-Nonanone was used as internal standard to quantify        these results are quite similar to other Passito wine values reported
the individual compounds.                                              in the literature. In fact, Giordano and others (2009) investigated
                                                                       3 different protected designation of origin (PDO) passito wines
Antioxidant activity                                                   namely Caluso Passito, Cinque Terre Sciacchetrà, and Passito from
   DPPH radical-scavenging activity assay. Radical-                    Pantelleria and found a total acidity ranging from 5.4 to 7.3 g/L
scavenging activity was investigated using the technique reported      as tartaric acid and a volatile acidity of 1.0 to 1.2 g/L. Our sam-
by Blois (1958) with some modifications. An aliquot of 1.2 mL          ples are characterized by a lower content of reducing sugar (116
of 1 × 10−4 M DPPH solution in ethanol and 300 μL of wine at           g/L) and ash (2.1 g/L). These parameters may vary according to
different dilutions in water (not diluted, 1/2, 1/3, 1/4, 1/5) were    the origin and processing technology of wines (Frias and others
mixed. The bleaching of DPPH was determined by measuring               2003). Other passito wines, such as Greco di Bianco DOC and
the absorbance at λ = 517 nm.                                          Mantonico DOC, both produced with Calabrian autochthonous
                                                                       Greco di Bianco and Mantonico grapes, exhibited a total acidity
   Antioxidant capacity determined by radical cation
                                                                       of 6.49 and 9.90 g/L as tartaric acid, respectively. The content of
(ABTS+ ). ABTS assay was based on the method of Re and others
                                                                       reducing sugar was 98 and 82 g/L for Greco di Bianco and Man-
(1996). After addition of 25 μL of samples at different dilutions
                                                                       tonico wines, respectively, and was therefore lower with respect to
in water (not diluted, 1/2, 1/3, 1/4, 1/5) or Trolox standard to
                                                                       our passito wine. This fact could be explained considering that the
diluted ABTS+ solution, absorbance at λ = 734 nm was measured.
                                                                       wine samples of passito had lower acidity and higher sugar content
   Antioxidant activity by β-carotene bleaching test. An-              due to the different state of ripeness of the grapes. Growing area,
tioxidant activity was determined using the β-carotene bleach-         climate conditions (temperature and humidity can vary very sig-
ing test previously described by Conforti and others (2007). The       nificantly according to the light exposure and height above sea and
bleaching of β-carotene was measured and expressed as antioxidant      date of harvest are crucial factors determining the sugar content
activity (AA):                                                         and total acidity of the grapes. Usually, a higher sugar content is
                                                                       always associated with a lower total acidity, as it was the case of the
            AA = [1 − (A0 − At )/(A◦0 − A◦t )] × 100                   passito wine of Saracena.
                                                                          Winemaking techniques including maceration, modalities of
where A0 and A◦ 0 are the absorbance values measured at the            fermentation, and aging can influence the content of total
incubation t = 0 min for samples and control, respectively, while      polyphenols significantly (Zeppa and others 2001). Saracena pas-
At and A◦ t are the absorbance values measured in the samples and      sito wine had a total phenolics content of 249 mg/L as (+)-
control, respectively, at t = 30 min and t = 60 min.                   catechin. This value is similar to the total polyphenols content
                                                                       of Calluso Passito wine (251.3 mg/L as (+)-catechin) produced
                                                                       in the north of Italy from Erbaluce grapes (Giordano and others
Statistical analysis                                                   2009) and is similar to the total phenolics content of white wines of
  All experiments were carried out in triplicate. Data were ex-        good quality. Fuda and others (2007) had analyzed several passito
pressed as means ± standard deviation (SD). Prism GraphPad             wines produced in Calabria (Italy) and reported a high content
Prism version 4.0 for Windows (GraphPad Software, San Diego,           of (+)-catechin (7.5 to 33.1 ppm) for Mantonico DOC sweet
Calif., U.S.A.) was used for plotting the concentration-response       wine, whereas a lower content of apigenin was found in Greco
curve and one-way ANOVA test followed by multicomparison               di Bianco 2004 (4.2 to 17.3 mg/L). A total phenolics content of
Dunnet’s test.                                                         450 mg/L as gallic acid equivalent was found in Muscat of Samos,

                                                                                       Vol. 78, Nr. 5, 2013 r Journal of Food Science C705
Passito chemistry and bioactivity . . .

                    Table 2–Principal phenolic components of Passito wine.                                     Table 3–The major volatile compounds identified in Passito wine
                                                                                                               after SPME extraction.
                    Phenolic compounds                                   Mean ± standard deviation
                                                                                                               Compound                                              RTa              mg/L
                    Benzoate   Ia                                                  84.9 ±   2.5
                    Benzoate IIa                                                   88.6 ±   6.4                iso-Butanol                                           8.50          22.1   ±   0.3
                    Gallic acida                                                    376 ±   24.5               iso-Amyl acetate                                      8.80           6.7   ±   0.2
                    Hydroxytyrosolb                                                78.3 ±   3.5                Limonene                                             10.72           1.6   ±   0.08
                    Protocatechuic acidc                                           37.3 ±   1.4                iso-Amyl alcohol                                     12.72           161   ±   3.6
C: Food Chemistry

                    Caftaric acidd                                                 68.2 ±   5.9                1-Hydroxy-2-propanone                                15.68           9.1   ±   0.2
                    Tyrosolb                                                        148 ±   12.9               1-Hexanol                                            19.60          23.2   ±   0.7
                    (+)-Catechine                                                  71.7 ±   4.9                1-Heptanol                                           19.86          18.9   ±   2.3
                    cis-Coutaric acidf                                              7.7 ±   0.4                Ethyl hexanoate                                      20.01           156   ±   9.7
                    trans-Coutaric acidf                                           15.0 ±   0.9                Acetic acid                                          20.57            75   ±   2.4
                                                                                                               2-Furancarboxaldehyde                                20.85           6.0   ±   0.3
                    Caffeic acidd                                                  27.9 ±   1.9
                                                                                                               Ethyl octanoate                                      21.98           6.4   ±   0.1
                    Fertaric acidg                                                 10.2 ±   0.4
                    Procyanidin dimere                                              195 ±   9.5                2,3-Butanediol                                       23.54          60.6   ±   4
                                                                                                               Linalool                                             23.68           0.6   ±   0.04
                    p-Coumaric acidf                                               13.0 ±   0.8
                                                                                                               5-Methyl-2-furancarboxaldehyde                       24.26           4.1   ±   0.9
                    Ferulic acidg                                                  11.5 ±   0.9
                                                                                                               Ethyl decanoate                                      26.26           135   ±   12.3
                    trans-Resveratrolh                                              4.9 ±   0.1                2-Furanmethanol                                      27.08           3.7   ±   1.0
                    Procyanidind                                                   18.8 ±   1.1                Butanedioic acid, diethyl ester                      27.41           9.9   ±   0.2
                    Oligomeric procyanidinse                                        203 ±   17.5               Ethyl dec-9-enoate                                   27.78           4.7   ±   0.3
                    Total phenolic                                                 1460                        α-Terpineol                                          28.04           1.1   ±   0.09
                    Phenolic compounds quantified as: a mg gallic acid/L; b mg tyrosol/L; c mg
                                                                                                               α-Muurolene                                          28.58           0.5   ±   0.04
                    protocatechuic acid/L; d mg caffeic acid/L; e mg (+)-catechin/L; f mg p-coumaric acid/L;   Isopropyl dodecanoate                                31.73           8.5   ±   0.6
                      mg ferulic acid/L; h mg trans-resveratrol/L.                                             Ethyl dodecanoate                                    31.98          22.9   ±   1.3
                                                                                                               Linalyl propinate                                    33.33           1.3   ±   0.1
                                                                                                               2-Phenylethanol                                      33.77          34.9   ±   1.7
                                                                                                               Nerolidol                                            37.02           0.4   ±   0.03
                    a dessert wine produced in Greece using Samos Muscat Blanc à                              Ethyl tetradecanoate                                 37.21          13.8   ±   1.4
                    Petits Grains grapes (Roussis and others 2005).                                            Octanoic acid                                        37.52           5.7   ±   0.7
                                                                                                               Ethyl hexadecanoate                                  42.00           187   ±   7.7
                                                                                                               Decanoic acid                                        42.43           6.3   ±   0.4
                    Profiling of phenolics in passito wine                                                     1-Hexadecene                                         44.69          22.8   ±   0.8
                       The total content of phenolic compounds quantified with                                 Ethyl octadecanoate                                  46.35          57.8   ±   3.0
                    HPLC was 1460 mg/L (Table 2). The most abundant classes of                                 Ethyl 9-octadecanoate                                46.74          13.2   ±   1.3
                    phenols were hydroxybenzoic acids and flavan-3-ols, where gal-                             Dodecanoic acid                                      46.93           2.1   ±   0.1
                                                                                                               5-(Hydroxymethyl)-2-furancarboxaldehyde              47.07           8.2   ±   0.1
                    lic acid showed the highest content (376.5 mg/L). Four flavanols                           1-Octadecene                                         48.88           5.1   ±   0.2
                    were tentatively identified; they were monomeric (catechin) and                            Tetradecanoic acid                                   51.42          21.4   ±   0.2
                    dimeric forms (procyanidins). The high content of oligomeric                               Pentadecanoic acid                                   54.39          13.5   ±   0.6
                    procyanidins in a white wine can be due to polymerization of sim-                          Hexadecanoic acid                                    58.22         118.1   ±   7.5
                                                                                                               9-Hexadecenoic acid                                  59.73          22.6   ±   1.3
                    ple flavonoids, such as catechins, during aging. Procyanidins with
                    a high degree of polymerization were eluted as one peak at the                             a
                                                                                                                 Retention time (min) on DB-WAX capillary column. Results are given as means ±
                                                                                                               standard deviations of 3 replications in mg/L.
                    end of the chromatogram as it usually happens with a C18 station-
                    ary phase (Boselli and others 2006). The content of catechin and
                    procyanidin dimer was 72 and 195 mg/L. The phenyl ethyl alco-
                                                                                                               vineyard climate and soil parameters, cultivation practices, and
                    hols detected were: 2-phenylethanol, hydroxytyrosol, and tyrosol
                                                                                                               the aging time and conditions (Rodriguez-Delgado and others
                    at concentration of 35, 78, and 148 mg/L. 2-Phenylethanol is well
                                                                                                               2002). Based on this consideration, the differences in the chemi-
                    known for its sensory floral aroma and is derived from the yeast
                                                                                                               cal profile among Passito wines of different origin should not be
                    metabolism of the nonvolatile phenylalanine (Poláŝkova and others
                    2008). In addition, several different forms of hydroxycinnamic acid
                    and their tartaric esters were detected. These compounds were reg-
                    istered in considerable amounts; the content of hydroxycinnamic                            SPME-GC of aroma compounds
                    acid and derivatives ranged from 10.2 to 68.2 mg/L for fertaric                               Various compounds are responsible for wine aroma. They are
                    and caftaric acid, respectively. trans-Resveratrol was found at a                          classified as follows: primary aroma compounds (originate from
                    level of 4.9 mg/L. Roussis and others (2005) analyzed the HPLC                             the grape), secondary aroma compounds (are formed during fer-
                    phenolic profile of Muscat of Samos and found mainly benzoic                               mentation), and tertiary aroma compounds (are produced during
                    acid and cinnamic acid together with unclassified compounds and                            aging in wood barrels). In recent years, SPME has become the
                    flavonoids. The phenolic profile of Greco di Bianco and Man-                               most widely applied sampling procedure for the study of wine
                    tonico passito wines were reported by Fuda and others (2007)                               flavor composition (Rocha and others 2001; Torrens and others
                    who identified apigenin-7-glucoside, caffeic acid, (+)-catechin,                           2004; Riu-Aumatell and others 2006). A total of 42 compounds
                    epicatechin, ferulic acid, gallic acid, p-coumaric acid, rutin, sy-                        were identified in Saracena passito wine, including alcohols, es-
                    ringic acid, and vanillic acid as constituents. Among them a high                          ters, acids, terpenes, aldehydes, and ketones. Many of these com-
                    content of caffeic acid (38.9 mg/mL for Greco di Bianco and                                pounds are commonly found in wines (Table 3). Ethyl hexanoate
                    59.8 mg/L for Mantonico) and gallic acid (30.2 mg/L for Greco di                           (156 mg/L), ethyl decanoate (135 mg/L), and isoamyl alcohol (161
                    Bianco and 48.7 mg/L for Mantonico) were found in both wines.                              mg/L) were determined as the major constituents of passito wine
                    As previously underlined, the phenolic composition of wine de-                             and showed concentrations higher than threshold values of 0.005,
                    pends not only on grape varieties but also on the location of the                          0.2, and 30 mg/L, respectively previously reported by Jiang and

                    C706 Journal of Food Science r Vol. 78, Nr. 5, 2013
Passito chemistry and bioactivity . . .

Zhang (2010). Ethyl hexanoate is responsible for the apple peel,        Table 4–Antioxidant activities of Passito dessert wine.
and ethyl decanoate for the fruit, oily, and floral odors (Scacco and                                                                        β-Carotene
others 2010). Other abundant compounds were hexadecanoic acid                                   DPPH                 ABTS                   bleaching test
(118 mg/L), acetic acid (75 mg/L), 2,3-butanediol (61 mg/L),                                    assay                assay                    (IC50 v/v)
and ethyl octadecanoate (58 mg/L). The concentrations of ethyl                                (IC50 v/v)          (IC50 v/v)           30 min             60 min
esters of fatty acids, representing the most important group of
                                                                        Wine sample          0.03 ± 0.005          0.6 ± 0.08         0.4 ± 0.02        0.5 ± 0.04
aroma compounds, depend on several factors including type of

                                                                                                                                                                          C: Food Chemistry
yeast, fermentation temperature, aeration degree, and sugar con-        Data are expressed as mean ± SD (n = 3); DPPH radical scavenging activity assay;
                                                                        antioxidant capacity determined by radical cation (ABTS+ ); β-carotene bleaching test.
tent (Schreirer 1979). Moreover, ethyl esters are known to have         Ascorbic acid was used as positive control in DPPH (IC50 = 5.0 μg/mL) and ABTS
very pleasant fruits, honey, and sweet scents which contribute          (IC50 = 1.7 μg/mL); Propyl gallate was used as positive control in β-carotene bleaching
                                                                        test (IC50 = 1.0 μg/mL at 30 and 60 min incubation).
to the aromatic finesse of wines (Ugliano and Henschke 2009).
An amount of 6.7 mg/L was found for iso-amyl acetate, a com-
pound responsible for distinctly banana-like fragrance (Van Wyck           Data of antioxidant activity are reported in Table 4. Passito wine
and others 1979). This compound derives from the amino acid             scavenged the DPPH with an IC50 of 0.03 v/v, while a value of
metabolism of yeast and its concentration in our sample is in line      0.60 v/v was found using ABTS. Moreover, passito wine showed
with those reported for Caluso passito, Cinque Terre Sciacchetrà,      promising inhibition of linoleic acid oxidation (IC50 values of 0.4
and passito from Pantelleria (Giordano and others 2009). The con-       and 0.5 v/v at 30 and 60 min of incubation, respectively) that
centration of this ester in Saracena passito wine is higher than its    decreased linearly after dilution. Roussis and others (2005) had
flavor threshold (0.03 mg/L) (Jiang and Zhang 2010). Higher alco-       reported the scavenging of DPPH radical by Muscat wine from
hols represent the other important group of volatiles identified in     Greece founding EC50 values of 314 and 134 γ /(mg/L) at 5 and
wine. The group is composed of aliphatic and aromatic alcohols,         60 min of incubation and correlated the bioactivity to the high
most of which are products of yeast fermentation (Kotseridis and        phenolic content. The DPPH and ABTS tests of wines obtained by
others 2000). In Saracena passito wine, iso-butanol, 1-hexanol,         Muscat Hamburg grapes revealed that samples ranged from 1402
1-heptanol, 2,3-butanediol, and 2-furanmethanol were identi-            to 3410 and from 8841 to 14309 Trolox equivalents, respectively
fied. Among aldehydes, 2-furancarboxaldehyde, and 5-methyl-             (Li and others 2009).
2-furancarboxaldehyde were identified. In this passito wine, how-
ever, the bitter almond odor that is characteristic of the presence     Conclusion
of benzaldehyde was completely absent, while it is a significantly         In conclusion, this work analyzed Saracena passito wine for its
dominant bouquet component of Caluso Passito and Cinque Terre           phenolic and aroma profile and potential antioxidant properties.
Sciacchetrà Passito wines (Giordano and others 2009). Numerous         This wine is obtained using autochthonous Malvasia and Guar-
studies have shown that the terpenes are critical in determining the    naccia grapes with further addition of Muscat grapes. Chemical
bouquet of a wine. Seven terpenes were identified in passito wine.      analyses have provided a contribution to define the chemical mark-
They were α-terpinolene (0.5 mg/L), α-terpineol (1.1 mg/L),             ers of the quality of this wine, and their quantitative ranges, which
limonene (1.6 mg/L), linalool (0.6 mg/L), nerolidol (0.4 mg/L),         are needed to assess the authenticity of local niche production
linalyl propionate (1.3 mg/L), and α-muurolene (0.5 mg/L). Jiang        claiming a quality designation (such as PDO). Moreover, passito
and Zhang (2010) reported for linalool and limonene threshold           wine demonstrated good antioxidant properties. This bioactiv-
values of 0.0252 and 0.2 mg/L, respectively. In comparison with         ity could be related to the presence of flavonoids, anthocyans, and
the other constituents, their concentrations were low according         many other polyphenolic compounds, that showed highly positive
to previous studies on other Passito wines. Moreover, these com-        effects on human health.
pounds are not influenced by the fermentation process so could
be used as indicators of the grape variety and quality (Begala          Acknowledgment
and others 2002). Linalool is the floral representative monoter-          This work was supported by European Community POR Cal-
pene alcohol of sweet wine, followed by α-terpineol and nerodiol        abria FSE 2007/2013.
(Ribéreau-Gayon and others 2000). In particular, this monoter-
pene compound is related to acacia and honey odor. It has been          Conflict of Interest
found also in Greco di Bianco passito wine, but not in Manton-             The authors have declared no conflict of interest.
ico passito wine (Fuda and others 2007). The olfactory profile
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