Industrial Crops and Products 62 (2014) 507–514

Contents lists available at ScienceDirect

Industrial Crops and Products

journal homepage: www.elsevier.com/locate/indcrop

Comprehensive two-dimensional gas chromatography with mass

spectrometry applied to the analysis of volatiles in artichoke
(Cynara scolymus L.) leaves
Caroline Saucier a , Allan dos S. Polidoro a , Anaí L. dos Santos a , Jaderson K. Schneider a ,
Elina B. Caramão a,b , Rosângela A. Jacques a,∗
a
Institute of Chemistry, Federal University of Rio Grande do Sul (UFRGS), 9500 Bento Gonçalves Av., ZIP 91501-970 Porto Alegre, RS, Brazil
b
INCT-E&A, Federal University of Rio Grande do Sul, RS, Brazil

a r t i c l e i n f o a b s t r a c t

Article history: Artichoke (Cynara scolymus L.) is well known due to its medicinal properties and, as a result, a large
Received 5 June 2014 number of studies have been conducted to determine the chemical constituents produced by the plant.
Received in revised form 20 August 2014 However, investigations were mainly focused on the non-volatile compounds, while the volatile con-
Accepted 10 September 2014
stituents remained largely neglected. This study was aimed at obtaining a deeper understanding of the
Available online 30 September 2014
volatile composition of artichoke. For this propose, comprehensive two-dimensional gas chromatography
coupled to a rapid scanning quadrupole mass spectrometer (GC × GC/qMS) and retention indices were
Keywords:
used to improve the chemical characterization of volatiles from leaves. A total of 130 compounds were
Volatile compounds
Hydrodistillation
found, 109 of which are reported for the first time in C. scolymus L., including oxygenated monoter-
GC × GC/qMS penes, sesquiterpenes, oxygenated sesquiterpenes, norisoprenoids, lactones, alcohols, ketones and
Terpenes aldehydes. The major compounds were 1-octen-3-one (3.85%), (E)-2-hexenal (3.75%), benzene acetalde-
Bioactive compounds hyde (2.90%), 2,2-dimethyl-4-pentenal (2.81%), ␤-ionone (1.94%), furfural (1.65%), (E)-␤-damescenone
Plant analysis (1.59%), ␣-methyl-␥-butirolactone (1.53%), benzaldehyde (1.47%) and dihydroactinidiolide (1.44%). The
comprehensive GC × GC/qMS approach enabled a greater number of analytes to be identified, approxi-
mately four times higher than that obtained for GC/qMS. Additionally, the results imply that artichoke
leaves are a potential source of volatile bioactive compounds.
© 2014 Elsevier B.V. All rights reserved.

1. Introduction of therapeutic activities was ascribed to several active compounds

Artichoke (Cynara scolymus L.) is a herbaceous perennial crop
that originates from the Mediterranean area and is widely culti-
vated around the world. Since ancient times, artichoke extracts
have been used in herbal medicine because of their recognized
therapeutic effects. Several studies have demonstrated hepatopro-
tective, anticarcinogenic, antioxidative, antibacterial, antifungal,
anti-HIV, urinative, anticholesterol and glycaemia reduction phar-
macological activities (Adzet et al., 1987; Brown and Rice-Evans,
1998; Cairella and Vecchi, 1969; Clifford, 2000; Clifford and Walker,
1987; Gebhardt, 1997; Preziosi, 1969; Rondanelli et al., 2011;
Rodriguez et al., 2002; Zhu et al., 2005). Water and polar organic
solvents were typically used in these studies. Such a broad range
∗ Corresponding author. Tel.: +55 51 3308 7213; fax: +55 51 3308 7213.
E-mail addresses: rosangela.j@gmail.com, rosangela.j@globo.com,
rosangela.j@iq.ufrgs.br (R.A. Jacques).
providing synergistic pharmacological effects (Romano et al., 2005;
Zhu et al., 2004).
Because of these diverse beneficial properties, the literature
contains numerous studies on non-volatile artichoke constituents,
particularly the phenolic compounds (as reviewed by Lattanzio
et al., 2009). However, there are few investigations on their volatile
constituents. Some researchers have reported that monoterpenes,
sesquiterpenes, alcohols, aldehydes and ketones are the main
volatile constituents of the leaves and heads of C. scolymus L.
(Buttery et al., 1978; Ghanem et al., 2009; Guillén-Ríos et al., 2006;
Hădărugă et al., 2009; MacLeod et al., 1982; Nassar et al., 2013).
Plant volatiles are usually complex samples that contain
hundreds of compounds, including alcohols, aldehydes, esters,
ketones, phenols, lactones, phenylpropanoids and terpenoids
(Brielmann et al., 2006; Rowshan et al., 2013). One-dimensional
gas chromatography (1D-GC) has been used for many years as
the standard tool for separating volatiles from plants. However,
obtaining the best component separation via 1D-GC is difficult due

http://dx.doi.org/10.1016/j.indcrop.2014.09.023
0926-6690/© 2014 Elsevier B.V. All rights reserved.
508 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514
to the insufficient resolution power of a single column (Purcaro
et al., 2009). This problem persists despite the continuous develop-
ment of chromatographs, techniques and analytical methodologies
(Mateus et al., 2010). Overlapping peaks usually significantly com-
plicate compound identification and accurate quantification (Zhu
et al., 2007). In addition, plant volatiles are present in a wide range
of concentrations, and the trace analytes that are occasionally the
biologically active matrix components may not be detected, par-
ticularly if they are co-eluted with high concentration compounds
(Mateus et al., 2010).
Comprehensive two-dimensional gas chromatography
(GC × GC) has emerged as a powerful separation technique
for overcoming these limitations and is widely used to character-
ize complex samples. GC × GC uses two orthogonal mechanisms to
separate the sample constituents through two columns connected
in series with different stationary phases and a transfer device,
defined as the modulator. The modulator continuously isolates, re-
concentrates, and introduces small portions of the primary column
effluent to a secondary column. The time required to complete this
process is defined as the modulation period (Marriott et al., 2012;
Mondello et al., 2008). The chromatographic resolution is greatly
enhanced by this technology (Marriott et al., 2004).
The detectors used in GC × GC represent an additional challenge
because high spectral acquisition rates are required for correct
peak assignment and quantification (Adahchour et al., 2005). The
time-of-flight mass spectrometer (TOFMS) can be used to obtain
such data; however, its high cost limits its laboratory utilization
(Mondello et al., 2005). In contrast, a quadrupole mass spectrom-
eter (qMS) is much less expensive and more user-friendly, and
several authors have reported the use of qMS hyphenated to
GC × GC (Adahchour et al., 2008; Mondello et al., 2008). Recent
studies have shown the effectiveness of GC × GC in combination
with qMS operating in the rapid scanning mode to achieve satis-
factory data acquisition rates (Cordero et al., 2007; Purcaro et al.,
2010; Tranchida et al., 2013).
The aim of this study was to investigate the volatile con-
stituents of artichoke leaves via comprehensive two-dimensional
gas chromatography coupled to a rapid scanning quadrupole
mass spectrometer. The linear temperature programmed reten-
tion indices (LTPRI) were used to confirm the peak assignments. To
the best of the authors’ knowledge, this is the first report detailing
volatile compounds in artichoke leaves using GC × GC.
2. Materials and methods
2.1. Plant material
Artichoke (C. scolymus L.) was collected in the municipal dis-
trict of Riozinho (29◦ 38 27 S and 50◦ 27 10 W), Rio Grande do
Sul State, Brazil, in February 2011 and identified by the specialist
botanist Dr. Eduardo Pasini (Bio-sciences Institute, Department of
Botany, UFRGS, Rio Grande do Sul, Brazil). A voucher specimen (ICN:
166985) was deposited in the Herbarium of UFRGS, Rio Grande do
Sul, Brazil. The leaves were dried at 35 ◦ C until their weight was
constant, and they were then stored in dark bags to protect them
from humidity and light.
2.2. Chemicals and solvents
All of the solvents and standards (linear alkanes) used were
purchased from Sigma–Aldrich (St. Louis, MO, USA). All water was
purified using a Milli-Q system (Millipore, Bedford, MA, USA). Other
unmarked reagents were of analytical grade.
2.3. Volatile compounds extraction
Dried artichoke leaves (100 g) were hydrodistilled for 4 h using a
Clevenger-type apparatus following the method recommended by
the Brazilian Official Pharmacopoeia V (2010). The obtained distil-
late was extracted with dichloromethane and dried over anhydrous
sodium sulfate. The organic layer was transferred into dark vials
and stored at 4 ◦ C until its analysis.
2.4. GC/qMS analysis
The GC/qMS measurements were performed on a Shimadzu
GC/qMS system, consisting of a GC2010 Gas Chromatograph and
a QP2010 Plus Mass Spectrometer (qMS) (Kyoto, Japan). The
GC was equipped with an AOC-20i auto-injector (split/splitless).
The separation was performed on a ZB-5MS (5% phenyl,
95% dimethylpolysiloxane) column with 60 m length × 0.25 mm
I.D. × 0.25 ␮m film thickness (Phenomenex, Torrance, CA, USA). The
GC oven started at 40 ◦ C, was heated at 2 ◦ C/min to 300 ◦ C, and then
maintained for 20 min. The injector temperature was 300 ◦ C, and
the injection was performed in the splitless mode using 0.5 ␮L. High
purity helium (99.99%, Linde Gases, Canoas, Porto Alegre, RS, Brazil)
at a flow rate of 0.91 mL/min was used as the carrier gas. The inter-
face and ion source temperature was 300 ◦ C. The mass spectrometer
operated in the electron impact mode (EI) at 70 eV and scanned
from 40 to 500 m/z in a full scan acquisition mode. The data were
acquired using GCMS-solution software version 2.6 (Shimadzu,
Kyoto, Japan). The mass spectrum of each detected compound was
compared with those in the NIST-05 mass spectral library, using
similarity matches of at least 80% for identification. This identi-
fication was supported by the experimentally determined linear
temperature programmed retention index (LTPRI) values and com-
pared with the values reported in the bibliography when available
(Adams, 2007; NIST 11, 2013). The LTPRI values were determined
using a C6 –C30 n-alkane series and calculated from the van den
Dool and Kratz equation (van den Dool and Kratz, 1963). The rel-
ative amounts (%) for each individual component in the sample
were expressed as percent peak areas relative to the total peak
area.
2.5. GC × GC/qMS analysis
The GC × GC/qMS was performed using a Shimadzu GC × GC sys-
tem consisting of a GC2010 gas chromatograph and a QP2010 Ultra
mass spectrometer (qMS) (Kyoto, Japan). The GC was equipped
with an AOC-20i auto-injector (split/splitless) and dual-stage loop-
type modulator ZX1-GC × GC (Zoex Corporation, Houston, TX,
USA). Cryogenic modulation occurred every 5 s, with a hot jet
duration of 0.5 s. The first-dimension chromatographic separation
was performed on a OV-5 (5% phenyl, 95% dimethylpolysiloxane)
column with 60 m length × 0.25 mm I.D. × 0.10 ␮m film thick-
ness (Ohio Valley Specialty Company, Marietta, OH, USA). A
DB-17 (50% phenyl and 50% dimethylpolysiloxane) column with
2.15 m × 0.18 mm × 0.18 ␮m (J&W Scientific, Agilent Technologies,
Palo Alto, CA, USA) was used as the second dimension. The GC
conditions are the same as for GC/qMS. The data were acquired
using the GC Image software version 2.2 (ZOEX Corporation, Hous-
ton, TX, USA). The identification methods were the same as those
used previously. The relative amounts (%) for each individual com-
ponent in sample were expressed as the percent of the peak
volume relative to the total peak volume. The sum of the retention
time from the first and second dimensions were used to iden-
tify each peak. This procedure was validated by von Mühlen et al.
(2008).
 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514 509

Table 1 impure mass spectra. With this in mind, artichoke leaf volatiles
Volatile compounds identified in Cynara scolymus L. leaves using GC/qMS.
were analyzed via GC × GC/qMS. Table 2 shows the identified com-
No. Compound tR (min) A (%) LTPRIexp LTPRIlit pounds, both their first and second dimension retention times, their
1 Furfural 14.47 1.84 834 828a volume percentages and their LTPRI values. These values are listed
2 (E)-2-Hexenal 15.37 1.42 853 846a in order of their elution through the OV-5 capillary column.
3 Benzaldehyde 21.48 1.44 961 952a Figs. 1 and 2 present the 1D-GC/qMS(A) and GC × GC/qMS(B)
4 1-Octen-3-one 22.55 11.30 978 980b total ion current chromatogram expansions of C. scolymus L.
5 6-Methyl-5-hepten-2-one 23.19 0.37 988 981a
leaf volatiles. The 1D chromatogram contains several overlap-
6 Octanal 24.21 0.40 1004 998a
7 Benzene acetaldehyde 27.01 8.86 1044 1036a ping peaks. However, adding a second dimension to separate the
8 (E)-2-Octenal 28.05 0.62 1058 1060b compounds according to their polarity increased the GC chro-
9 Acetophenone 28.62 0.11 1067 1059a matographic space and enhanced the separation potential. Many
10 (E,E)-3,5-Octadien-2-one 28.97 0.25 1072 1072b
components that were obscured in the 1D-GC analysis are imme-
11 Nonanal 31.20 1.68 1103 1100a
12 (E)-6-Methyl-3,5-heptadien-2-one 31.35 0.62 1106 1106b diately revealed using GC × GC.
13 Phenethyl alcohol 31.93 0.46 1114 1107a Fig. 3 shows a section of the contour plot as an example (the
14 Isophorone 32.47 1.18 1122 1118a TIC is shown in Figs. 1 and 2) of the second-dimension separa-
15 3-Nonen-2-one 33.74 0.26 1140 1141b tion (upon polarity) for the four compounds that overlapped in
16 (E,Z)-2,6-Nonadienal 34.68 0.30 1153 1150a
the first dimension (separation upon volatility). They were sep-
17 4-Methyl-acetophenone 36.93 0.36 1185 1179a
18 Safranal 37.95 1.15 1200 1196a arated by GC × GC due to their different polarities with a higher
19 Decanal 38.29 0.28 1205 1201a chromatographic resolution than for 1D-GC. These compounds
20 ␤-Cyclocitral 39.39 1.35 1221 1220b were not identified by the 1D-GC system because of the low
21 Neral 40.77 0.22 1241 1235a
spectral match given by the library database search. Another
22 ␤-Homocyclocitral 41.87 0.24 1257 1254b
23 Geranial 42.76 0.80 1270 1264a
consequence of GC × GC is the increased signal to noise ratio (S/N)
24 p-Vinylguaiacol 45.66 0.68 1314 1314b for all of the analyte peaks. The increased S/N combines with the
25 Eugenol 48.51 0.69 1358 1356a enhanced resolution of GC × GC to maximize the purity of the
26 ␥-Nonalactone 48.86 0.85 1363 1358a obtained mass spectra, which allows for a more accurate identi-
27 (E)-␤-Damascenone 50.24 5.02 1385 1386a
fication of the sample compounds. Peaks B–D (Fig. 3) are present in
28 Geranyl acetone 54.40 2.23 1452 1455a
29 ␤-Ionone 56.51 6.05 1487 1487a low concentrations; however, their improved S/N ratio due to the
30 Dicyclohexyl-methanone 58.50 5.18 1520 – modulator provides a better separation from the system noise than
31 Dihydroactinidiolide 59.25 3.43 1533 1532b that obtained for the 1D-GC. Consequently, better mass spectra
32 Phytone 75.90 0.19 1844 1844b
are recorded, which allowed the compounds tentative identifica-
tR (min): retention times (in minutes), A (%): peak area percentages, LTPRIexp : exper- tion. A survey of the Adams database allowed the compounds to be
imental linear temperature programmed retention indices values, LTPRIlit : linear identified by their retention index.
temperature programmed retention indices values for the correspondent compound
reported in the literature.
A detailed GC × GC/qMS analysis of the sample tentatively iden-
a
Adams. tified 130 compounds, 109 of which were reported for the first
b
NIST 11 library. time for artichoke (C. scolymus L.). The tentatively identified com-
Compounds in bold are previously reported for Cynara scolymus L. (Buttery et al., pounds included 24 aldehydes, 24 ketones, 14 norisoprenoids, 19
1978; Ghanem et al., 2009; Guillén-Ríos et al., 2006; Hădărugă et al., 2009; Nassar oxygenated monoterpenes, 9 oxygenated sesquiterpenes, 9 alco-
et al., 2013). Data are the mean of three replicates. The retention times showed
variation coefficient less than 2%.
hols, 9 lactones, 5 phenylpropanoids, 4 unsaturated hydrocarbons,
3 sesquiterpenes, 3 fatty acids, 2 furans, 2 nitrogenated compounds,
1 flavonoid, 1 ether and 1 tiazol. The major components identi-
3. Results and discussion fied in the artichoke leaves via GC × GC/qMS were 1-octen-3-one
(3.85%), (E)-2-hexenal (3.75%), benzene acetaldehyde (2.90%), 2,2-
3.1. GC/qMS analysis dimethyl-4-pentenal (2.81%), ␤-ionone (1.94%), furfural (1.65%),
(E)-␤-damescenone (1.59%), ␣-methyl-␥-butirolactone (1.53%),
The yield (w/w) of essential oil obtained from artichoke leaves benzaldehyde (1.47%) and dihydroactinidiolide (1.44%).
was 0.1%. As a preliminary application, the volatile composition of The first authors to study the volatile fraction of C. scolymus L.
artichoke leaves was investigated via GC/qMS. Table 1 shows the were Buttery et al. (1978). They analyzed steam distillates from arti-
identified compounds, their retention times and area percentages, choke heads using gas–liquid chromatography/mass spectrometry.
and their LTPRI values. These values are listed in the order of their A total of 32 compounds were characterized, including alcohols,
elution through the ZB-5MS capillary column. aldehydes, ketones, oxygenated terpenes and sesquiterpenes. The
For GC/qMS, 32 compounds were tentatively identified using major components were ␤-selinene and caryophyllene. Similarly,
the NIST mass spectra library and retention index criteria, includ- MacLeod et al. (1982) used steam distillation and GC/MS to analyze
ing 7 aldehydes, 7 norisoprenoids, 6 phenylpropanoids, 6 ketones, C. scolymus L. volatiles. They reported the tentative identification
4 oxygenated monoterpenes, 1 lactone and 1 alcohol. According of 28 compounds, and sesquiterpenes composed the major group
to Table 1, the major components present in artichoke leaves components for C. scolymus L. with ␤-selinene as the major con-
were identified as 1-octen-3-one (11.30%), benzene acetaldehyde stituent.
(8.86%), ␤-ionone (6.05%), dicyclohexyl-methanone (5.18%), (E)- The prominent volatile compounds found in dichloromethane
␤-damescenone (5.02%), dihydroactinidiolide (3.43%) and geranyl extracts from artichoke hearts by Guillén-Ríos et al. (2006) using
acetone (2.23%). GC/MS were ␤-selinene, isoamyl acetate, trans-caryophyllene,
limonene, 2-hexanol, benzaldehyde and aromadrendene.
3.2. GC × GC/qMS analysis Ghanem et al. (2009) investigated the potential protective effect
of C. scolymus L. leaves versus the hepatic and renal toxicity of
Artichoke leaf essential oil can be considered a complex sample, lead in male rats and explored the volatile constituents using a
and thus, 1D-GC could fail to satisfactorily elucidate its composi- dynamic headspace system and GC/MS analysis. A total of 23 com-
tion. The main consequence of an insufficient resolving power is pounds were identified. This analysis indicated that benzaldehyde,
overlapping analytes, which should be avoided because it can yield selinene, phenethyl alcohol and caryophyllene oxide were the main
510 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514

Table 2
Volatile compounds identified in Cynara scolymus L. leaves using GC × GC/qMS.
1 2
No. Compound tR (min) tR (s) V (%) LTPRIexp LTPRIlit

1 2,3-Dihydro-3-methyl-furan 11.00 0.58 0.38 766 –

2 4-pentenal 11.33 0.58 0.51 773 –
3 2,2-dimethyl-4-pentenal 12.67 0.55 2.81 802 –
4 Furfural 14.08 1.54 1.65 833 828a
5 3-Furfural 14.17 1.27 0.36 835 831b
6 (E)-2-Hexenal 15.00 0.82 3.75 853 846a
7 2-Cyclopentene-1,4-dione 16.75 1.75 0.57 891 884b
8 2-Heptanone 16.92 0.82 0.35 895 889a
9 Heptanal 17.42 0.82 0.96 904 901a
10 (E,E)-2,4-Hexadienal 18.08 1.33 0.45 915 907a
11 Dihydro-3-(2H)-thiophenone 20.42 2.20 0.34 953 952b
12 6-Methyl-2-heptanone 20.67 0.91 0.17 957 956b
13 (Z)-2-Heptenal 20.75 1.12 0.39 958 964b
14 Benzaldehyde 21.00 1.72 1.47 962 952a
15 ␣-Methyl-␥-butyrolactone 21.08 2.35 1.53 964 957b
16 7-Oxabicyclo[4.1.0]heptan-2-one 21.67 1.66 0.23 973 –
17 1-Octen-3-one 22.08 1.06 3.85 980 980b
18 6-Methyl-5-hepten-2-one 22.67 1.12 0.74 989 981a
19 2-Pentyl-furan 22.83 0.85 0.23 992 993b
20 (E,E)-2,4-Heptadienal 23.50 1.39 0.30 1003 1003b
21 Octanal 23.67 1.00 0.72 1005 998a
22 2-Etil-1-hexanol 25.50 0.94 0.36 1031 1030b
23 2,2,6-Trimethyl-cyclo-hexanone 25.75 1.42 0.50 1035 1035b
24 Benzyl alcohol 25.92 1.99 0.45 1038 1026a
25 Benzene acetaldehyde 26.50 2.08 2.90 1046 1036a
26 (E)-2-Octenal 27.50 1.48 0.69 1060 1060b
27 ␥-Caprolactone 27.50 2.53 0.25 1061 1063b
28 Acetophenone 28.08 2.11 0.40 1069 1059a
29 4-Methyl-benzaldehyde 28.25 1.93 0.17 1071 1076b
30 Octanol 28.42 1.06 0.34 1074 1063a
31 (E,E)-3,5-Octadien-2-one 28.50 1.51 0.32 1075 1072b
32 (Z)-5-Undecene 28.92 1.18 0.15 1081 –
33 1-Nonen-4-ol 30.17 1.33 0.13 1099 1109b
34 Linalool 30.33 1.03 0.26 1101 1095a
35 Nonanal 30.67 1.12 0.75 1106 1100a
36 (E)-6-Methyl-3,5-heptadien-2-one 30.83 1.63 0.51 1109 1107b
37 2,5-Dimethyl-cyclohexanol 31.00 1.48 0.68 1111 1099b
38 Phenethyl alcohol 31.42 2.11 0.60 1117 1107a
39 2,4-Dimethyl-2,4-heptadienal 31.58 1.51 0.12 1119 1129b
40 (Z)-2-Undecene 31.75 1.06 0.17 1122 1114b
41 Isophorone 31.92 1.75 0.66 1124 1118a
42 3-Nonen-2-one 33.17 1.72 0.30 1142 1141b
43 trans-3-Nonen-2-one 33.25 1.33 0.09 1143 1144b
44 Oxophorone 33.50 2.02 0.46 1147 1147b
45 Camphor 33.58 1.63 0.24 1148 1141a
46 cis-Verbenol 33.58 1.39 0.14 1148 1147b
47 5-Ethyl-6-methyl-(E)-3-hepten-2-one 33.67 1.15 0.16 1149 1144b
48 4-(5-Methyl-2-furanyl)-2-butanone 34.00 1.78 0.20 1154 –
49 (E,Z)-2,6-Nonadienal 34.25 1.45 0.50 1157 1150a
50 Phenyl-2-propenal 34.42 2.38 0.26 1160 1161b
51 (E)-2-Nonenal 34.58 1.60 0.37 1162 1162a
52 Propiophenone 35.00 2.05 0.34 1168 1164b
53 p-mentha-1,5-dien-8-ol 35.17 1.54 0.41 1170 1166a
54 1-Nonanol 35.50 1.36 0.35 1175 1165a
55 Caprylic acid 35.58 1.21 0.15 1176 1177b
56 Terpinen-4-ol 35.83 1.33 0.11 1180 1174a
57 4-Methyl-acetophenone 36.42 2.14 0.27 1188 1179a
58 p-Cymen-8-ol 36.42 1.78 0.18 1188 1179a
59 Safranal 37.42 1.75 0.64 1203 1196a
60 Decanal 37.75 1.18 0.19 1207 1201a
61 3-Phenyl-butanal 38.58 1.78 0.14 1220 –
62 Coumaran 38.83 2.20 0.53 1223 1223b
63 ␤-Cyclocitral 38.92 1.66 0.70 1224 1224b
64 Benzothiazol 39.17 2.98 0.23 1228 1227b
65 Nerol 39.42 1.36 0.19 1231 1227a
66 2-Pentyl-cyclopentanone 39.67 1.87 0.09 1235 –
67 Neral 40.33 1.54 0.44 1245 1235a
68 Carvone 40.50 1.84 0.08 1247 1239a
69 Geraniol 41.17 1.42 0.11 1257 1249a
70 Piperitone 41.25 1.84 0.09 1258 1249a
71 ␤-Homocyclocitral 41.42 1.51 0.19 1260 1261b
72 ␥-Octanolactone 41.50 2.38 0.12 1262 1262b
73 (E)-2-Decenal 41.58 1.66 0.16 1263 1264a
74 1,12-Tridecadiene 42.17 2.02 0.17 1271 –
75 Pelargonic acid 42.17 1.30 0.27 1271 1267a
76 Geranial 42.33 1.60 0.25 1274 1264a
 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514 511

Table 2 (Continued)
1 2
No. Compound tR (min) tR (s) V (%) LTPRIexp LTPRIlit

77 2-Phenyl-2-butenal 42.42 2.47 0.14 1275 1273b

78 Isopiperitenone 42.42 2.14 0.07 1275 1272b
79 (E)-2-Decen-1-ol 42.67 1.21 0.18 1278 1271a
80 3-Ethoxy-benzaldehyde 43.25 2.23 0.10 1287 –
81 Anethole 43.33 1.96 0.13 1288 1282a
82 p-mentha-1(7),8(10)-dien-9-ol 43.58 1.78 0.23 1292 –
83 2-Undecanone 43.75 1.21 0.07 1294 1293a
84 Indole 43.92 3.19 1.41 1297 1290a
85 p-Vinylguaiacol 45.17 2.26 0.71 1316 1316b
86 (E,E)-2,4-Decadienal 45.33 1.63 0.35 1318 1315a
87 Megastigma-4,6(Z),8(Z)-triene 46.42 1.63 0.31 1335 1324b
88 Piperitenone 47.00 2.26 0.19 1345 1340a
89 1-Methoxy-4-methyl-bicyclo[2.2.2]octane 47.08 2.35 0.46 1346 1341b
90 Ionene 47.58 1.45 0.16 1353 1349b
91 1,2-Dihydro-1,1,6-trimethyl-naphthalene 47.75 1.72 0.16 1356 1355b
92 Eugenol 48.00 2.11 0.70 1360 1356a
93 ␥-Nonalactone 48.33 2.35 0.47 1365 1358a
94 (Z)-␤-Damascenone 48.33 1.66 0.23 1365 1361a
95 Capric acid 48.50 1.33 0.24 1368 1373b
96 Oxide piperitenone 48.58 2.29 0.68 1369 1366a
97 Biphenyl 49.25 2.41 0.13 1380 1375a
98 Isolongifolene 49.58 1.60 0.11 1385 1386b
99 (E)-␤-Damescenone 49.75 1.75 1.59 1387 1386a
100 5-Methyl-indole 49.92 3.07 0.15 1390 1381b
101 6,10-Dimethyl-2-undecanone 50.92 1.15 0.12 1405 1407b
102 Methyleugenol 51.00 2.14 0.07 1407 1403a
103 7,8-Dihydro-3,4-dehydro-␤-ionone 51.67 1.51 0.97 1418 1424b
104 Geranyl acetone 53.92 1.48 0.86 1454 1455a
105 ␥-Decalactone 54.92 2.20 0.11 1471 1465a
106 Massoia lactone 55.58 2.47 0.10 1482 –
107 Dihydro-␤-ionone 55.83 1.84 0.33 1485 1485b
108 ␤-Ionone 56.08 1.72 1.94 1489 1487a
109 ␦-Decalactone 56.67 2.41 0.11 1499 1493a
110 ␣-Farnesene 56.75 1.84 0.20 1500 1506a
111 Dicyclohexyl-methanone 58.00 2.11 1.32 1522 –
112 Pseudoionone 58.67 1.72 0.11 1533 1527b
113 Dihydroactinidiolide 58.75 2.95 1.44 1535 1537b
114 (E)-Nerolidol 60.50 1.48 0.12 1564 1561a
115 Ledol 60.92 1.51 0.07 1572 1569a
116 Spathulenol 61.50 1.72 0.28 1582 1577a
117 Caryophyllene oxide 61.83 1.78 0.56 1587 1582a
118 Isoaromadendrene epoxide 62.75 1.75 0.15 1603 1594b
119 Humulene epoxide II 63.25 1.81 0.08 1613 1608a
120 Benzophenone 64.25 3.13 0.21 1631 1626a
121 Cubenol 64.33 1.66 0.13 1633 1627a
122 ␶-Cadinol 65.00 1.75 0.13 1645 1638a
123 Hedione 65.67 2.05 0.14 1657 –
124 cis-␥-6-Dodecenolactone 65.75 1.66 0.32 1659 1657b
125 Bisabolol oxide II 65.75 2.35 0.39 1659 1656a
126 Ageratochromene 65.92 2.68 0.08 1662 1658a
127 ␥-Dodecalactone 67.00 2.20 0.17 1682 1676a
128 ␣-Bisabolol 67.25 1.69 0.11 1686 1685a
129 Phytone 75.42 1.36 0.23 1846 1846b
130 Farnesyl acetone 79.08 1.84 0.10 1921 1920b
1
tR (min): first dimension retention times (in minutes), 2 tR (s): second dimension retention times (in seconds), V (%): peak volume percentages, LTPRIexp : experimental
linear temperature programmed retention indices values, LTPRIlit : linear temperature programmed retention indices values for the correspondent compound reported in
the literature.
a
Adams.
b
NIST 11 library.
Compounds in bold are previously reported for Cynara scolymus L. (Buttery et al., 1978; Ghanem et al., 2009; Guillén-Ríos et al., 2006; Hădărugă et al., 2009; Nassar et al.,
2013).

constituents in the sample. The study suggested that artichoke may
be useful in combating the damaging effect of lead toxicity due to
its volatile constituents with antioxidative properties.
Hădărugă et al. (2009) reported that the main compounds
of artichoke (C. scolymus L.) flowers, steam and root extracts
isolated via steam distillation and GC/MS were sesquiterpenes,
specifically ␤-cubebene and caryophyllene oxide. Nevertheless,
the major compound in the leaf extracts was (E)-2-hexenal,
followed by ␤-cubebene and caryophyllene oxide. Bisabolol
oxide II and ␣-bisabolol were also found in the artichoke
leaves.
Another approach to identifying the volatile composition of
C. scolymus L. was used by Nassar et al. (2013). Artichoke head
scales were hydrodistilled and analyzed using GC/MS. A total of
37 compounds were identified, with the majority including mono
and sesquiterpenes. The main constituents were cyclosativene and
myrtenal.
Comparing the cited literature data with the results presented
in this study verifies the different morphological parts of C. scoly-
mus L. yielded similar volatile compositions during the qualitative
analyses. However, the abundance of certain compounds differed
significantly between the different plant parts. Analogous behavior
512 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514

Fig. 1. 1D-GC/qMS (A) and GC × GC/qMS (B) total ion current chromatograms expansions of Cynara scolymus L. leaf volatiles.

was observed by Ramos et al. (2013), who compared the lipophilic
composition of stalks, capitula and leaves from Cynara cardunculus
L. var. altilis, another artichoke variety.
Several compounds from C. scolymus L. reported here (Table 2)
have not yet been studied for their biological activities, though they
are known to occur in other natural products. The activity of the
monoterpenes nerol and geraniol against Mycobacterium tubercu-
losis was studied by Rajab et al. (1998). Carvone, an oxygenated
monoterpene, was analyzed as a pulmonary adenoma reduc-
ing agent (Raphael and Kuttan, 2003). Linalool is an oxygenated
monoterpene with known anti-inflammatory activity that is used
as flavor and fragrance in foods and cosmetics (Ajikumar et al.,
2008; Dewhirst, 1980). The oxygenated sesquiterpene ␣-bisabolol
can be isolated from Matricaria chamolilla and is commonly used
in herbal medicine to treat skin inflammation and as an antibac-
terial and antifungal agent (Dewick, 2002). Caryophyllene oxide,
an oxygenated sesquiterpene, is used as a preservative in food,
drugs and cosmetics and has been tested in vitro for antibacterial
and antifungal activity (Rajeswari et al., 2011). Some research has
shown that phytone is a significant constituent of medicinal plants
(Leonurus japonicus Houtt. and Centaurium erythraea Rafn.) that
has antimicrobial potential for Escherichia coli, Salmonella enter-
itidis, and Staphylococcus aureus, among others (Jerković et al.,
2012; Xiong et al., 2013). ␤-Ionone, a norisoprenoid, has shown
cholesterol-suppressive action (Yu et al., 1994). Another noriso-
prenoid, ␤-damascenone, has been identified in many fruits and

Fig. 2. 1D-GC/qMS (A) and GC × GC/qMS (B) total ion current chromatograms expansions of Cynara scolymus L. leaf volatiles.
 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514
Fig. 3. Example of section of the contour plot (TIC already shown in Figs. 1 and 2), showing the second dimension separation (upon polarity) of four compounds that were
overlapping in the first dimension (separation upon volatility).
is important to the perfume and flavoring industries (Carneiro
et al., 2006). This discussion could include many of the compounds
reported in Table 2; however, that is outside the scope of this
study, which aimed to unveil the complex nature of artichoke leaf
volatiles.
4. Conclusions
GC × GC coupled to a rapid scanning quadrupole mass spec-
trometer proved adequate for analyzing the volatile components
of artichoke leaves. The use of GC × GC/qMS enhanced the separa-
tion efficiency and increased the signal to noise ratio (S/N) for the
analyte peaks, which maximized the obtained mass spectra purity
and improved the accuracy tentative identification of the artichoke
leaf components. Analyzing the mass spectra data and retention
index of the analytes allowed a total of 130 compounds to be tenta-
tively identified, 109 of were reported for the first time for artichoke
(C. scolymus L). The GC × GC mass spectra were of better quality
than those obtained by 1D-GC, which increased the match factors
for the NIST library and was thus more effective at identifying the
compounds in the samples compared with the one-dimensional GC
analysis.
Finally, the obtained results represent an initial contribution
toward a detailed knowledge of the chemical composition of
artichoke leaves. GC × GC/qMS confirmed the identity of many pre-
viously detected and new compounds relative to the 1D-GC/MS
systems in the existing literature. The presence of biologically
active molecules in artichoke leaves indicates that they can serve
as an alternative commercial source for these compounds.
Acknowledgements
The authors are grateful to The National Council for Scientific
and Technological Development (CNPq, Brazil), to The Coordination
of Improvement of Higher Education Personnel (CAPES, Brazil), and
to the Foundation for Research Support in the State of Rio Grande
do Sul (FAPERGS, Brazil) for financial support and fellowships.
513
References
Adahchour, M., Brandt, M., Baier, H., Vreuls, R.J.J., Batenburg, A.M., Brinkman,
U.A.T., 2005. Comprehensive two-dimensional gas chromatography coupled to
a rapid-scanning quadrupole mass spectrometer: principles and applications. J.
Chromatogr. A 1067, 245–254.
Adahchour, M., Beens, J., Brinkman, U.A.T., 2008. Recent developments in the appli-
cation of comprehensive two-dimensional gas chromatography. J. Chromatogr.
A 1186, 67–108.
Adams, R.P., 2007. Identification of Essential Oil Components by Gas Chromatogra-
phy/Mass Spectrometry, 4th ed. Allured Publishing Corporation, Illinois.
Adzet, T., Camarassa, J., Laguna, C.J., 1987. Hepatoprotective activity of polyphenolic
compounds from Cynara scolymus against CCl4 toxicity in isolated rat hepato-
cytes. J. Nat. Prod. 50, 612–617.
Ajikumar, P.K., Tyo, K., Carlsen, S., Mucha, O., Phon, T.H., Stephanopoulos, G., 2008.
Terpenoids: opportunities for biosynthesis of natural product drugs using engi-
neered microorganisms. Mol. Pharm. 5, 167–190.
Brazilian Health Surveillance Agency, 2010. Brazilian Official Pharmacopoeia
(FBRAS), 5th ed. Brasília, DF.
Brielmann, H.L., Setzer, W.N., Kaufman, P.B., Kirakosyan, A., Cseke, L.J., 2006. Phy-
tochemicals: the chemical components of plants. In: Cseke, L.J., Kirakosyan, A.,
Kaufman, P.B., Warber, S.L., Duke, J.A., Brielmann, H.L. (Eds.), Natural Products
From Plants. CRC Press, Boca Ratón, pp. 1–42.
Brown, J.E., Rice-Evans, C.A., 1998. Luteolin-rich artichoke extracts protects low
density lipoproteins from oxidation in vitro. Free Radic. Res. 29, 247–255.
Buttery, R.G., Guadagni, D.G., Ling, L.C., 1978. Volatile aroma components of cooked
artichoke. J. Agric. Food Chem. 26, 791–793.
Cairella, M., Vecchi, L., 1969. I principi attivi del carciofo in terapia. In: Atti Del I
Congresso Internazionale Di Studi Sul Carciofo. Edizioni Minerva Medica, Torino,
pp. 283–293.
Carneiro, J.R., Ferreira, J.A., Guido, L.F., Almeida, P.J., Rodrigues, J.A., Barros, A.A., 2006.
Determination of ␤-damascenone in alcoholic beverages by reversed-phase liq-
uid chromatography with ultraviolet detection. Food Chem. 99, 51–56.
Clifford, M., 2000. Chlorogenic acids and other cinnamates nature, occur-
rence, dietary burden, absorption and metabolism. J. Sci. Food Agric. 80,
1033–1043.
Clifford, M., Walker, R., 1987. Chlorogenic acids confounders of coffee–serum choles-
terol relationships. Food Chem. 24, 77–80.
Cordero, C., Bicchi, C., Joulain, D., Rubiolo, P., 2007. Identification, quantitation and
method validation for the analysis of suspected allergens in fragrances by com-
prehensive two-dimensional gas chromatography coupled with quadrupole
mass spectrometry and with flame ionization detection. J. Chromatogr. A 1150,
37–49.
Dewhirst, F.E., 1980. Structure–activity relationships for inhibition of prostaglandin
cyclooxygenase by phenolic compounds. Prostaglandins 20, 209–222.
Dewick, P.M., 2002. Medicinal Natural Products: A Biosynthetic Approach, 2nd ed.
John Wiley & Sons, New York.
Gebhardt, R., 1997. Antioxidative and protective properties of extracts from leaves
of the artichoke against hydroperoxide-induced oxidative stress in cultured rat
hepatocytes. Toxicol. Appl. Pharmacol. 144, 279–286.
514 C. Saucier et al. / Industrial Crops and Products 62 (2014) 507–514
Ghanem, K.Z., Ramadan, M.M., Farrag, A.R.H., Ghanem, H.Z., Farouk, A., 2009. Egyp-
tian artichoke volatile compounds protect against lead-induced hepatic and
renal toxicity in male rats. Polish J. Food Nutr. Sci. 59, 175–181.
Guillén-Ríos, P., Burló, F., Martínez-Sánchez, F., Carbonell-Barrachina, A.A., 2006.
Effects of processing on the quality of preserved quartered artichokes hearts. J.
Food Sci. 71, 176–180.
Hădărugă, N.G., Hădărugă, D.I., Tatu, C., Gruia, A., Costescu, C., Lupea, A.X., 2009.
Multivariate analysis (PCA) in compositae biocompounds class. J. Agroaliment.
Process. Technol. 15, 201–210.
Jerković, I., Gašo-Sokač, D., Pavlović, H., Marijanović, Z., Gugić, M., Petrović, I.,
Kovač, S., 2012. Volatile organic compounds from Centaurium erythraea Rafn
(Croatia) and the antimicrobial potential of its essential oil. Molecules 17,
2058–2072.
Lattanzio, V.L., Kroon, P.A., Linsalata, V., Cardinali, A., 2009. Globe artichoke:
a functional food and source of nutraceutical ingredients. J. Funct. Foods,
131–144.
MacLeod, A.J., Pieris, N.M., de Tronconis, N.L., 1982. Aromavolatiles of Cynara scoly-
mus and Helianthus tuberosus. Phytochemistry 21, 1647–1651.
Marriott, P.J., Massil, T., Hügel, H., 2004. Molecular structure retention relation-
ships in comprehensive two-dimensional gas chromatography. J. Sep. Sci. 27,
1273–1284.
Marriott, P.J., Chin, S., Maikhunthod, B., Schmarr, H.G., Bieri, S., 2012. Multidimen-
sional gas chromatography. Trends Anal. Chem. 34, 1–21.
Mateus, E., Barata, R.C., Zrostlíková, J., Silva, M.D.R.G., Paiva, M.R., 2010. Characteri-
zation of the volatile fraction emitted by Pinus spp. by one- and two-dimensional
chromatographic techniques with mass spectrometric detection. J. Chromatogr.
A 1217, 1845–1855.
Mondello, L., Casilli, A., Tranchida, P.Q., Dugo, G., Dugo, P., 2005. Comprehen-
sive two-dimensional gas chromatography in combination with rapid scanning
quadrupole mass spectrometry in perfume analysis. J. Chromatogr. A 1067,
235–243.
Mondello, L., Tranchida, P.Q., Dugo, P., Dugo, G., 2008. Comprehensive two-
dimensional gas chromatography–mass spectrometry: a review. Mass Spec-
trom. Rev. 27, 101–124.
Nassar, M.I., Mohamed, T., Elshamy, A.I., El-Toumy, S.A., Lateef, A., Farrag, A.H., 2013.
Chemical constituents and anti-ulcerogenic potential of the scales of Cynara
scolymus (artichoke) heads. J. Sci. Food Agric. 93, 2494–2501.
NIST 11, 2013. Mass Spectral Library. http://chemdata.nist.gov (accessed 28.02.13).
Preziosi, P., 1969. Valutazione farmacologica dei principi attivi del carciofo. In: Atti
Del I Congresso Internazionale Di Studi Sul Carciofo. Edizioni Minerva Medica,
Torino, pp. 237–281.
Purcaro, G., Tranchida, P.Q., Jacques, R.A., Caramão, E.B., Moret, S., Conte, L., Dugo, P.,
Dugo, G., Mondello, L., 2009. Characterization of the yerba mate (Ilex paraguar-
iensis) volatile fraction using solidphase microextraction-comprehensive 2-D
GC–MS. J. Sep. Sci. 32, 3755–3763.
Purcaro, G., Tranchida, P.Q., Ragonese, C., Conte, L., Dugo, P., Dugo, G., Mondello, L.,
2010. Evaluation of a rapid-scanning quadrupole mass spectrometer in an apolar
× ionic-liquid comprehensive two-dimensional gas chromatography system.
Anal. Chem. 82, 8583–8590.
Rajab, M.S., Cantrell, C.L., Franzblau, S.G., Fischer, N.H., 1998. Antimycobacterial
activity of (e)-phytol and derivatives—a preliminary structure-activity study.
Planta Med. 64, 2–4.
Rajeswari, R., RamaLakshmi, N., Muthuchelian, K., 2011. GC–MS analysis of bioactive
components from the ethanolic leaf extract of Canthium dicoccum (Gaertn.). J.
Chem. Pharm. Res. 3, 792–798.
Ramos, P.A.B., Guerra, A.R., Guerreiro, O., Freire, C.S.R., Silva, A.M.S., Duarte, M.F.,
Silvestre, A.J.D., 2013. Lipophilic extracts of Cynara cardunculus L. var. altilis
(DC): a source of valuable bioactive terpenic compounds. J. Agric. Food Chem.
61, 8420–8429.
Raphael, T.J., Kuttan, G., 2003. Immunoregulatory activity of naturally occur-
ring monoterpenes carvone, limonene and perillic acid. Immunopharmacol.
Immunotoxicol. 25, 285–294.
Rodriguez, T.S., Giménez, D.G., Vázquez, R.P., 2002. Choleretic activity and biliary
elimination of lipids and bile acids induced by an artichoke leaf extract in rats.
Phytomedicine 9, 687–693.
Rondanelli, M., Giacosa, A., Orsini, F., Opizzi, A., Villani, S., 2011. Appetite control
and glycaemia reduction in overweight subjects treated with a combination of
two highly standardized extracts from Phaseolus vulgaris and Cynara scolymus.
Phytother. Res. 25, 1275–1282.
Romano, A., Pinelli, P., Cantini, C., Cimato, A., Heimler, D., 2005. Characterization of
Violetto di Toscana, a typical Italian variety of artichoke (Cynara scolymus L.).
Food Chem. 95, 221–225.
Rowshan, V., Bahmanzadegan, A., Saharkhiz, M.J., 2013. Influence of storage condi-
tions on the essential oil composition of Thymus daenensis Celak. Ind. Crop. Prod.
49, 97–101.
Tranchida, P.Q., Franchina, F.A., Zoccali, M., Pantò, S., Sciarrone, D., Dugo, P.,
Mondello, L., 2013. Untargeted and targeted comprehensive two-dimensional
GC analysis using a novel unified high-speed triple quadrupole mass spectrom-
eter. J. Chromatogr. A 1278, 153–159.
van den Dool, H., Kratz, P.D., 1963. Generalization of the retention index system
including linear temperature programmed gas–liquid partition chromatogra-
phy. J. Chromatogr. 11, 463–471.
von Mühlen, C., Zini, C.A., Caramão, E.B., Marriott, P.J., 2008. Comparative study
of Eucalyptus dunnii volatile oil composition using retention indices and com-
prehensive two-dimensional gas chromatography coupled to time-of-flight and
quadrupole mass spectrometry. J. Chromatogr. A 1200, 34–42.
Yu, S.G., Abuirmeileh, N.M., Qureshi, A.A., Elson, C.E., 1994. Dietary ␤-ionone sup-
presses hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase activity. J.
Agric. Food Chem. 42, 1493–1496.
Xiong, L., Peng, C., Zhou, Q., Wan, F., Xie, X., Guo, L., Li, X., He, C., Dai, O., 2013.
Chemical composition and antibacterial activity of essential oils from different
parts of Leonurus japonicus Houtt. Molecules 18, 963–973.
Zhu, X.F., Zhang, H.X., Lo, R., 2004. Phenolic compounds from the leaf extract of
artichoke (Cynara scolymus L.) and their antimicrobial activities. J. Agric. Food
Chem. 52, 7272–7278.
Zhu, X.F., Zhang, H.X., Lo, R., 2005. Antifungal activity of Cynara scolymus L. extracts.
Fitoterapia 76, 108–111.
Zhu, S., Lu, X., Ji, K., Guo, K., Li, Y., Wu, C., Xu, G., 2007. Characterization of fla-
vor compounds in Chinese liquor Moutai by comprehensive two-dimensional
gas chromatography/time-of-flight mass spectrometry. Anal. Chim. Acta 597,
340–348.