Ultrasonic assist with headspace solid phase microextraction (UA-HS-SPME) have been applied, for the first time, for the extraction of volatile organic compounds of Carum carvi L. from Iran. The oils obtained were analyzed by GC-MS. Different experimental parameters such as fiber&apos;s coating type, sonication time, extraction time and temperature, and desorption time were investigated. Results showed that in the presence of UA-HS-SPME the highest extraction efficiency was achieved with a 100 μm polydimethylsiloxane (PDMS) fiber. Essential oil analysis showed that 10 compounds were identified through UA-HS-SPME-GC/MS method .The major component were carvone (57.7 %) and limonone (35.5 %)

Ali Abdollahi

Mohammad Hadi Meshkatalsadat

Reza Aminiradpoor

Sabah Salahvarzi

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Digest Journal of Nanomaterials and Biostructures                     Vol. 7, No. 2, April - June 2012, p. 637 - 640 
 
 
 
 
IDENTIFICATION OF ESSENTIAL OIL CONSTITUENTS OF CARAWAY 
(CARUM CARVI) USING ULTRASONIC ASSIST WITH HEADSPACE SOLID 
PHASE MICROEXTRACTION (UA-HS-SPME) 
 
 
MOHAMMAD HADI MESHKATALSADATa*,  SABAH  SALAHVARZIb , 
REZA AMINIRADPOORa, ALI ABDOLLAHIc 
aFaculty of Basic Engineering sciences, Qom University of technology ,Qom, Iran 
bDepartement of Chemistry, Azad University of khoramabad, khoramabad.Iran 
cDepartment of Medicinal Plants, Faculty of Agriculture and Natural Resources, 
Sistan & Baluchestan University, Saravan, Iran 
 
 
Ultrasonic assist with headspace solid phase microextraction (UA-HS-SPME) have been 
applied, for the first time, for the extraction of volatile organic compounds of Carum carvi 
L. from Iran. The oils obtained were analyzed by GC–MS. Different experimental 
parameters such as fiber's coating type, sonication time, extraction time and temperature, 
and desorption time were investigated. Results showed that in the presence of UA-HS-
SPME the highest extraction efficiency was achieved with a 100 μm polydimethylsiloxane 
(PDMS) fiber. Essential oil analysis showed that 10 compounds were identified through 
UA-HS-SPME-GC/MS method .The major component were carvone (57.7 %) and 
limonone (35.5 %). 
 
(Received February 6, 2012; Accepted May 7, 2012) 
 
Keywords: Carum carvi L., essential oil, solid phase microextraction, gas 
chromatography–mass spectrometry, carvone, limonene. 
 
 
1. Introduction 
 
The genus Carum belong to the Apiaceae or Umbelifereae family includes 25 spices, only 
Caraway (Carum carvi L.) has an economical importance, being used and cultivated in several 
regions (from Northern Europe to the Mediterranean regions, Russia, Iran, Indonesia and North 
America). In numerous countries it is a very common species and, as a result, an integral part of 
their folk medicines. C. carvi essential oil has been used as a flavouring for liquors and toothpaste, 
while the seeds have been used as a spice and flavouring. C. carvi essential oils or their pure 
constituents (such as carvone) showed several biological activities including insecticidal or insect 
repellent effects, antibacterial and antifungal effects, inhibition of seed germination and inhibition 
of sprouting of potatoes [1-2]. The percentages of essential oil available in C. carvi seeds were 
reported in different studies between 1% (v/w) and 6% (v/w). Caraway seeds contain a several 
components. Carvone and limonene are the main components available in their oil. Also, their 
seeds contain trace amounts of other compounds including acetaldehyde, furfural, carveole, 
pinene, thujone, camphene, phellandrene, etc [3].  
Hydrodistillation (HD), steam distillation, steam and water distillation are the 
conventional methods to obtain essential oils from the essential oil-bearing plants. Among these 
methods, HD has been the most common approach to extract the essential oils from the medicinal 
herbs/plants. All conventional methods to isolate valuable compounds from aromatic plants have 
important drawbacks, such as low yields, formation of by-products – owing to degradation of 
thermally unstable and unsaturated compounds by temperature or hydrolytic effects, respectively – 
                                                 
*Corresponding author: mhmeshkatalsadat@yahoo.com, meshkatalsadat.m@qut.ac.ir 
638 
 
large extraction times and the presence of the extraction, usually a toxic organic solvent. In the last 
decade there has been an increasing demand for new extraction techniques, amenable to 
automation, with shortened extraction times and reduced organic solvent consumption, preventing 
pollution and reducing sample preparation costs [4].  
Solid-phase microextraction (SPME) is a solvent-free sampling and effective technique 
that introduced in the early 1990s as a new sampling and sample-preparation method. In this 
method require only small amounts of sample and the aromatic oils from the sample are adsorbed 
directly onto an adsorbent coated fused-silica fiber and then either thermally desorbed directly into 
a gas chromatograph (GC) injection port or high performance liquid chromatography (HPLC) 
injection valve. In addition, SPME has been widely applied to the sampling and analysis of 
environmental, food, aroma, forensic, and pharmaceutical samples [5]. Headspace SPME (HS-
SPME) and ultrasonic-assisted (UA) extraction has been successfully applied to determine volatile 
compounds from plant materials [6-10].  
The aim of this research work was to extract essential oil from C. carvi as a famous 
Iranian essential oil-bearing material by UA-HS-SPME method. 
 
2. Experimental 
 
The Carum carvi L. fruits at ripening stage were harvested from plants growing in the 
gardens of the Agricultural College of Razi University, during May 2009. About 100 g of fruits 
dried in room temperature by spreading them on a clean aluminum foil in laboratory. 10 g portions 
of air-dried sample were subjected to a household coffee grinder in order to make them to a coarse 
powder. The ground samples were stored in nylon bags and placed in refrigerator until use and 
further analysis. 
The SPME fiber, 100 µm polydimethylsiloxane (PDMS) was purchased from Supelco 
(Bellefonte, PA, USA) and conditioned prior to use according to suppliers prescriptions. Helium, 
99.999%, used as carrier gas, was purchased from Roham Gas Company (Tehran, Iran). The 
alkane mixture consisting of the C8-C26 alkanes (concentration of 40 mg/mL in hexane) was 
purchased from Fluka. All other chemicals were of the highest purity available from Merck or 
Fluka. Doubly distilled deionized water was used. 
The aerial parts of the plant were dried at room temperature by spreading them on clean 
aluminum foil in the laboratory. 10-g portions of air-dried sample were ground to a coarse powder 
using a household coffee grinder. The ground samples were stored in nylon bags and placed in a 
refrigerator prior to analysis. Extraction of the volatiles from the plant sample using SPME fibers 
was achieved by placing 0.5 g of ground sample into a 40-ml vial to which 500 μl double-distilled 
water was added as a matrix modifier, the vial was then vigorously shaken by hand to ensure 
homogeneous dispersal of the spiked water. The sample vial was then placed into an ultra-
sonicator and incubated for 15 min to allow the volatiles to equilibrate between the headspace and 
sample matrix, during which time the sample was heated to 70 ºC. The actual SPME extraction of 
volatile compounds was accomplished by incubation with a polydimethylsiloxane (PDMS) fiber at 
70 ºC for 40 min. Ultrasonic irradiation (18 kHz, 450 W) was applied by means of a PFO100 5RS 
Series ultrasonicator (Italy) equipped with a water bath in which the extraction vials were placed. 
The samples were sonicated to create stress in the sample matrix to facilitate the release of the 
analytes, and control the temperature during the extraction process. The alkane mixture (C8-C26), 
40 mg/mL in hexane) was used for the calculation of retention indices (RIs). Loading the alkane 
mixture onto the fiber was carried out using a 5-min head space extraction from the 10-ml SPME 
vial including 1 ml double-distilled water spiked with 10 μl of the above-mentioned mixture. 
Gas chromatography was performed with a Shimadzu model GC-17A (Kyoto, Japan) 
instrument equipped with a Shimadzu Quadropole-MS (QMS) model QP5050 detector. Separation 
was achieved using a 30 m × 0.25 mm I.D capillary fused silica column 6 coated with a 0.25 μm 
film of DB5-MS (5% Phenyl-95% Polydimethyl Siloxane), and a split/splitless injector with a 1 
mm internal diameter glass liner. Ten-, 20- and 40-milliliter sample vials sealed with PTFE coated 
silicone septa (Supelco) were used for extraction. SPME fibers with PDMS (100 μm, non-bonded) 
coating, were used as commercial fibers for the extraction of analyses. The fibers were handled 
using a manual SPME fiber holder provided by Supelco (Bellefonte, PA, USA). Analyses 
639 
 
extracted onto the fiber were injected into the injection port of the GC system. The fiber was kept 
in the injection port for additional 2 min after injection to ensure the complete adsorption of the 
compounds from the fiber. Every 10 analyses on a GC run were carried out in the presence of the 
fiber but without sampling to assure complete adsorption. The injector was set at 220 ºC. The 
carrier gas was helium and flowed at a rate of 1 ml/min. The GC was operated in a splitless mode. 
The column temperature was initially set at 40 ºC and increased to 200 ºC at a rate of 4 ºC/min, 
and remain at 200 ºC for 1 min, resulting in a total GC run time of 45 min. The temperature of the 
ion source was kept at 220 ºC, and the transfer line temperature at 250 ºC. The mass fragments 
were collected in the range from m/z 40 to 450 with an acquisition rate of 1000 to provide a 
satisfactory number of points per peak for effective spectral resolution. The ionization energy of 
70 eV and the detector voltage of 1700 V were applied to the QMS detector.  
Most constituents were identified by gas chromatography through comparison of their 
retention indices (RIs) with those of the literature [11] or with those of authentic compounds 
available in our laboratories. The retention indices (RIs) were determined in relation to a 
homologous series of n-alkanes (C8–C26) under the same operating conditions. Further 
identification was made by comparison of their mass spectra on both columns with those stored in 
NIST 98 and Wiley 5 Libraries or with mass spectra from literature [11-13]. Component relative 
concentrations were calculated based on GC peak areas without using correction factors. 
 
3. Results and discussion 
 
The chemical composition of the oil of the C. carvi is listed in Table 1, in which the 
percentage and retention indices of components are given. The components were listed in order of 
their elution on the DB-5 column.  
 
Table 1. Chemical composition of C. carvi oil extracted by UA-HS-SPME. 
 
No. RT (min) a RI cal 
b Compounds Relative content (%) 
1 14.05 1000 Myrcene 0.48 
2 16.29 1055 Limonene 35.5 
3 18.68 1112 α-Terpinolene 0.1 
4 20.40 1157 Trans Limonene Oxide 2.01 
5 23.26 1231 Trans Dihydro Carvone 0.33 
6 24.59 1266 Trans Carveol 0.31 
7 25.68 1295 Carvone 57.7 
8 26.31 1312 Perilla Alcohol 1.48 
9 26.74 1324 Carvacrol 0.34 
10 31.04 1447 β-Caryophyllene 0.41 
Total content fraction of determinates compounds 98.66 
a RT: retention time  b RI cal: calculated retention indices using kovats formula 
 
Ten components were identified in the essential oil of C. carvi extracted using UA-HS-
SPME method, represented 98.66% f the total components, the major compounds were carvone 
(57.7%) and limonene (35.5%). These results were accordance to previous studies that reported 
carvone and limonene were the main components of C. carvi oil [3, 4, 15]. Also, the essential oil of C. 
carvi was characterized by high contents of oxygenated monoterpenes (62.17%), monoterpenes 
(36.08%) and sesquiterpenes (0.41%), respectively (Figure 1). The fraction of oxygenated 
monoterpenes and monoterpene were enriched, mainly due to an increase in carvone and limonene 
percentage. 
In order to provide a complete peak separation of extracted compounds, some preliminary 
SPME-GC/MS experiments were performed using ground C. carvi sample utilizing a PDMS fiber. 
From different recorded chromatograms, it becomes clear that the best GC program was as which 
mentioned previously (the column initial temperature: 40 ºC, with a rate of 4 ºC/min increased to 
200 ºC, and remain at 200 ºC for 1 min). The affecting experimental parameters such as fiber's 
coating type, sonication time, extraction time and temperature, desorption time, and water content 
640 
 
of sample were optimized. The optimization of different affecting parameters was accomplished 
by using simplex method. The optimal conditions were as fiber's coating type: PDMS, sonication 
time: 15 min, extraction time: 40 min, extraction temperature: 70 ºC, adsorption time: 2 min, and 
water content: 500 μl per 0.5 g of ground sample. The use of a simplex optimization method was 
of paramount importance in order to select the best working conditions of interrelated variables. 
A number of SPME fibers of different polarity and coating thickness are commercially 
available and have been used for extraction of the volatile compounds in medicinal plants [13]. 
Among the fibers, PDMS or PDMS-based mixed fibers are most commonly used. Using ultrasonic 
assist with headspace solid phase microextraction, the highest extraction efficiency was achieved 
with a 100 μm polydimethylsiloxane (PDMS) fiber and it was found that the oxygenated 
monoterpene fractions increased but the amount of sesquiterpenes and monoterpenes decreased.  
In order to get access to the absolute mass percentage of the identified compounds, the 
essential oil of C. carvi was analyzed after extraction by SPME. Our experience showed that 
SPME could give the exact mass percentage of the constituents of volatile compounds in 
comparison with reported hydrodistillation method [3, 4, 15].  
In conclusion, the study of the essential oil from C. carvi seeds showed the presence of 
carvone and limonene after extraction by SPME. Thus, the experimental parameters such as 
extraction time, irradiation power and ultra sound effects can be optimized for the particular aim of 
the SPME, either to obtain a high yield of essential oil, or to obtain essential oils of differing 
composition [4]. However, SPME is capable to analyze the volatiles with the least extraction time, 
sample amount, and sample preparation steps. In addition, significant ability of trapping and 
extracting of compounds which are more volatile.  
 
  
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Identification of essential oil constituents of caraway (Carum carvi) using ultrasonic assist with headspace solid phase microextraction (UA-HS-SPME),” Digest

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Identification of essential oil constituents of caraway (Carum carvi) using ultrasonic assist with headspace solid phase microextraction (UA-HS-SPME),” Digest

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