Comprehensive bidimensional gas chromatography (GCxGC-ToF ... Betnga... · Comprehensive bidimensional gas-chromatography (GCxGC or 2DGC) is at the forefront of the technical innovation

P.F. TCHOUAKEU BETNGA ET AL., COMPREHENSIVE BIDIMENSIONAL GAS CHROMATOGRAPHY AND SENSORY ANALYSIS TO INVESTIGATE THE AROMA PROFILE OF A COMMERCIAL GRAPPA, PAG. 1

WWW.INFOWINE.COM, INTERNET JOURNAL OF VITICULTURE AND ENOLOGY, 2019, N. 11/3

Comprehensive bidimensional gas chromatography (GCxGC-ToF-MS) and sensory analysis to investigate the aroma profile of a commercial grappa Prudence Fleur Tchouakeu Betnga1,2, Amanda Dupas de Matos1,2, Edoardo Longo1,2, Vakare Merkyte1,2, Sebastiano Pantò3, Emanuele Boselli1,2

1Free University of Bozen-Bolzano, Faculty of Science and Technology, Piazza Università 5, 39100 Bolzano-Bozen, Italy 2NOITechPark Alto Adige/Südtirol, Via A. Volta, 13B - 39100 Bolzano, Italy 3European and Application Technology Center, EATC, Berlin [email protected]

1- Introduction

Grappa is one of the most important Italian distillates and its production is allowed only from

grapes produced and processed in Italy. It is obtained from fermented or semi-fermented grape

marc (DM n. 747, 2016), which is a by-product of the winemaking process. Grappa is rich in

volatile compounds (about 1% v/v) [1] and each compound contributes to the flavour according

to its concentration and sensory threshold. This aspect plays a fundamental role in the

consumers’ preference of the different styles of grappa present on the market. The constant

application of new and improved reliable analytical tools is fundamental to achieve a detailed

description of a complex aroma profile in relation to its sensory descriptors.

Comprehensive bidimensional gas-chromatography (GCxGC or 2DGC) is at the forefront of

the technical innovation in volatile compounds analysis. With respect to mono-dimensional

GC-MS (1D), the bidimensional (2D) separation allows for higher sensitivities as well as better

peaks separation and identification of co-eluting compounds. Although GCxGC has been

already proposed for the characterization of distillates [2-8], the applications specifically on

grappa are rare. The aim of this study is to apply Head Space (HS) Solid Phase Microextraction

(SPME) coupled on-line with GC-MS analysis and SPME-GCxGC-ToF-MS to investigate the

volatile profile of a commercial young grappa in relation to its sensory profile.

2. Materials and methods

2.1 Grappa samples

Bottles of 500-mL were filled with a white (young) grappa (38% ABV) provided by Roner

distillery (Termeno, Italy) and sealed with Supercap (Mombaroccio, Italy) closures. The volatile

profile of the distillate was analysed with two different high resolution techniques: HS-SPME-

GC-MS analysis (section 2.2) and HS-SPME-GCxGC-ToF-MS analysis (section 2.3).



2.2 Mono-dimensional analysis (GC-MS)

The volatile profile of the grappa sample was analysed with gas chromatography - mass

spectrometry (GC-MS) after extraction with head-space solid phase microextraction (HS-

SPME). The procedure consisted in pipetting 4 mL of a prepared solution (0.5 L milliQ water +

62.5 g of NaCl + 100 µL of pure internal standard 4-methyl-2-pentanol) and transferring it into

a 20-mL vial. After that, 4 mL of grappa was added into the 20-mL vial and closed with a

perforable screw cap. Thus, the final volume of the solution was 8 mL. The vial was kept in a

continuous heating bath at 40 ºC for 15 min with a continuous stirring at 250 rpm. Afterwards,

a SPME fiber (DVB/CAR/PDMS, 50/30 µm, 1 cm) was exposed into the headspace of the 20-

mL vial for 30 min under continuous heating and stirring.

The GC-MS analysis was carried out on an Agilent 7890A GC coupled to an Agilent 5975 mass

detector. The thermal desorption took place at 240 °C for 3.5 min. The separation was

performed on a MEGA-WAX Spirit column 0.30 µm/0.18 mm/40 m (MEGA srl, Legnano, Italy),

with He carrier gas (GC-grade) at 0.7 mL/min, in split mode (1:10 split ratio). The oven

temperature was programmed at 40 °C for 0.2 min, then raised to 180 °C at 3 °C/min and to

230 °C at 10 °C/min. The mass spectrometer was operated in electron ionization mode at 70

eV. The mass range was 34-360 m/z and the ion source temperature at 230 °C; the quadrupole

temperature was set at 150 °C and the acquisition rate was 1 spectrum/sec.

2.3 Bidimensional analysis (GCxGC-ToF-MS)

Volatile compounds of grappa were also separated with a bidimensional gas chromatographic

system using an Agilent 7890 GC equipped with a Pegasus BT-4D system (LECO, Berlin,

Germany) after HS-SPME. For the first dimension, the column used was a Rxi-5 MS (30 m x

0.25 mm i.d. x 0.25 µm coating) and for the second dimension the column used was a Rxi-

17Sil MS (0.9 m x 0.25 mm i.d. x 0.25 µm coating) (Restek, Bellefonte, USA). The second

column was operated in a secondary oven. The first oven temperature was programmed as

follow: 40 ºC (hold 3 min), ramp 4 ºC/min to 220 ºC, ramp 15 ºC/min to 280 ºC hold 1 min; the

secondary oven temperature was kept at +5 ºC than the first dimension. The MS transfer line

was set at 280 ºC, the ion source temperature was set at 250ºC, the mass range was operated

between 34-600 m/z, the acquisition rate was 200 spectra/s and the extraction frequency was

at 30 Hz.



2.4 Sensory analysis

The samples were evaluated using QDA (Quantitative Descriptive Analysis), which is one of

the main descriptive analysis techniques in sensory evaluation [9]. The grappa samples were

given randomly to a trained panel (19 subjects, 58% females and 42% males, 23±4 years old)

in ISO glasses codified with 3-digit number containing 10 mL of grappa/glass at around 16-18

°C, in two different sessions according to the UNI 10957:2003 guideline.

3. Results and discussion

3.1 Volatile profile of grappa by GC-MS

The volatile profile of grappa was analysed in two replicates (bottle 1 and bottle 2); no

significant difference was observed. The major volatile compounds observed are

shown in Figure 1.

Figure 1. Volatile profile of grappa by SPME/GC-MS analysis.

The list of the main volatile compounds present in grappa is reported in Table 1. The main

chemical families are related to ethanol, higher alcohols and esters.



Table 1. Assignment of the most abundant peaks present in the chromatograms with the respective retention times.

Peak number * Retention time (min) Assignement 1 5.12 ethyl acetate 2 6.19 ethanol 3 10.85 1-propanol, 2-methyl 4 11.97 1-butanol-3-methyl acetate

5** 13.78 4-methyl, 2-pentanol 6 15.52 3-methyl-1-butanol 7 16.64 ethyl hexanoate 8 21.85 1-hexanol 9 25.59 ethyl octanoate 10 26.46 isopenthyl hexanoate 11 32.06 methyl decanoate 12 34.08 ethyl decanoate 13 41.48 ethyl dodecanoate

*peak numbers used in Figure 1; **Internal standard.

3.2 Volatile profile of grappa by GCxGC-ToF-MS

With respect to GC-MS analysis, in GCxGC many classes of volatile compounds were eluted

in separate patterns according to their main chemical classes (regions circled in red Figure 2).

As an example, the identification of the peaks in the regions of volatile aldehydes (Figure 2)

and fatty acid ethyl esters (Figure 3) is reported. The three volatile aldehydes shown are

located in the same region of the bidimensional trace (high retention times of the second

dimension). This result is very helpful for the correct identification of species belonging to the

same chemical class, compared to mono-dimensional GC, where the volatile classes of

compounds are not resolved (for instance the area delimited by the white frame in Figure 3).

Figure 2. Volatile profile of grappa by GCxGC analysis and zoom for three volatile

aldehydes.



Figure 3. Volatile profile of grappa by GCxGC analysis and zoom for fatty acid ethyl esters.

4. Sensory evaluation

4.1 Definition of the sensory descriptors for grappa

The aroma descriptors defined by the panel were grouped in classes [10] and are reported in

Figure 4.

Figure 4. Young grappa aroma wheel.

The trained panel was asked to rate the intensity of sensory attributes for: i) visual; ii) olfactory;

and iii) gustatory perceptions. The sensory attributes and respective meanings are shown in

Table 2.

Table 2. Sensory attributes listed in the score sheet and their definition.



Sensory attribute Definition

Visual

Clarity Absence of faults or taints

Color tonality Tonality or shade of color

Fluidity Ability of a substance to flow easily

Olfactory

Overall intensity Total smell intensity perceived through the nose

Pungent (alcohol) Pungency due to the alcohol perceived through the nose

Floral Rose, orange blossom

Dried fruit Raisin, plum, fig, date

Tree fruit Apple, pear, peach

Spicy Clove, licorice, anise, black pepper

Fresh vegetative Sage, mint, rosemary

Dried vegetative Hay, straw, fennel, tea

Caramelized Honey

Cleanness Absence of faults/taints/unpleasant odors

Unpleasant odors Presence of faults/taints/unpleasant odors

Gustatory

Overall intensity Total gustatory intensity by retronasal perception

Sweetness Taste of a sucrose solution

Bitterness Taste of a caffeine solution

Pungent (alcohol) Burning sensation due to alcohol

Floral Rose, orange blossom

Dried fruit Raisin, plum, fig, date

Tree fruit Apple, pear, peach

Spicy Clove, licorice, anise, black pepper

Fresh vegetative Sage, mint, rosemary

Dried vegetative Hay, straw, fennel, tea

Caramelized Honey

Cleanness Absence of faults/taints/unpleasant flavors

Unpleasant flavors Presence of faults/taints/unpleasant flavors

4.2 Sensory characterization of grappa samples

The sensory profile of grappa is shown in Figure 5. For visual evaluation, the grappa was

highly rated for clarity and fluidity, meaning that the sample did not present suspensions into

the distillate neither had a high viscosity (Figure 5A). For color tonality, the grappa was

characterized by weak intensity, meaning that it was colorless. For olfactory evaluation, the

grappa was described by weak intensity of floral, dried fruit, tree fruit, spicy, fresh vegetative,

dried vegetative, and caramelized aromas. However, the overall intensity perceived after

sniffing the sample was rated high, which could be explained by the high pungency due to the

alcohol content (38%, ABV). The sample was described also for its high cleanness (absence

of off-odors) and weak unpleasant odors (presence of off-odors), which confirm its good



sensory quality by olfactory evaluation (Figure 5B). For gustatory evaluation, sweetness,

bitterness and pungency were perceived at moderate-strong intensities by the panel. The

aromas perceived by retro-nasal evaluation (flavors) were also evaluated with a weak intensity

(Figure 5C). Indeed, the cleanness and absence of unpleasant flavors were confirmed.

Therefore, grappa was evaluated similarly for both olfactory and gustatory descriptors.

Figure 5. Intensity of sensory attributes of grappa by 9-point intensity scale. A: visual, B:

olfactory, C: gustatory evaluation.

5. Conclusion

The use of GCxGC allowed a better understanding of the complexity of grappa volatile profile,

since some resolved peaks in the second dimension of bidimensional GC were overlapped in

a monodimensional system (GC-MS). Moreover, comprehensive bidimensional GC provides

a peak list of about two hundred identified volatile compounds, which can be useful to identify

the molecules responsible for the main sensory descriptors. The most abundant classes of

compounds were ethyl esters, alcohol and aromatic aldehydes. Regarding the sensory

analysis, the main aroma classes reported by the trained panel were floral, fruity, spicy,

vegetative and caramelized. Ethyl esters of monocarboxylic acids (the series from butanoic to

dodecanoic acid) are related to the pleasant fruity aroma. Benzaldehyde is usually associated

with a dry fruit. Furfural has a bitter and spicy flavor, whereas benzenacetaldehyde is perceived

as a floral and fresh aroma. This research is an ongoing project in which the evaluation of

grappa samples will be carried out over a storage at three months, six months, one year and

two years.



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Comprehensive bidimensional gas chromatography (GCxGC-ToF ... Betnga... · Comprehensive bidimensional gas-chromatography (GCxGC or 2DGC) is at the forefront of the technical innovation