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Comparison of microwave-assisted
hydrodistillation and solvent-less microwave
extraction of essential oil from dry and fresh
Citruslimon (Eureka variety) peel

Mohammad-Taghi Golmakani & Mahsa Moayyedi

To cite this article: Mohammad-Taghi Golmakani & Mahsa Moayyedi (2016) Comparison of
microwave-assisted hydrodistillation and solvent-less microwave extraction of essential oil
from dry and fresh Citruslimon (Eureka variety) peel , Journal of Essential Oil Research, 28:4,
272-282, DOI: 10.1080/10412905.2016.1145606

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Journal of Essential Oil Research, 2016
VOL. 28, NO. 4, 272282
http://dx.doi.org/10.1080/10412905.2016.1145606

Comparison of microwave-assisted hydrodistillation and solvent-less microwave

extraction of essential oil from dry and fresh Citruslimon (Eureka variety) peel
Mohammad-Taghi Golmakania and Mahsa Moayyedib
a
Department of Food Science and Technology, School of Agriculture, Shiraz University, Shiraz, Iran; bDepartment of Food Science and Technology,
School of Agriculture, Varamin-Pishva Branch, Islamic Azad University, Varamin, Iran

ABSTRACT
This study investigated the effectiveness of microwave-assisted hydrodistillation (MAHD) and ARTICLE HISTORY
solvent-less microwave extraction (SLME) for extracting essential oil (EO) from lemon peels in Received 10 September 2014
comparison with conventional hydrodistillation (HD) in terms of extraction time, extraction yield, Accepted 19 January 2016
physical properties, chemical composition, operation cost, and antioxidant activity. The results
KEYWORDS
showed that MAHD and SLME methods required an extraction time of 15 minutes while the required Essential oil;
extraction time in HD was 120 minutes. There were no significant differences among physical hydrodistillation; lemon;
properties (refractive index, specific gravity, and color) of extracted EOs by the three methods. microwave-assisted
Gas chromatography analysis did not indicate any noticeable change in the composition of EOs hydrodistillation; solvent-less
extracted by microwave-assisted extraction methods (MAHD and SLME) and HD method. DPPH microwave extraction
analysis of the extracted EOs indicated that microwave irradiation did not adversely influence
antioxidant activity of the extracted EOs. The results of this study introduced MAHD and SLME as
rapid, economical, and environmentally friendly extraction methods.

1.Introduction contain volatile compounds with flavor and aroma pro-

Citrus (family Rutaceae), with annual production of
approximately 102 million metric tons, is the most impor-
tant fruit tree crop in the world (1, 2). The most impor-
tant citrus species are orange and mandarin, respectively.
The next important citrus species is lemon (Citruslimon),
with worldwide production of 4,200,000 metric tons per
year (3). Lemon is a member of the large family Rutaceae
which contains 130 genera in seven subfamilies, and has
many important varieties (4). Eureka is the most impor-
tant lemon variety of the world (2).
Lemon peel represents 3040% of the fruit weight, con-
stitutes its principal by-product and also a main source of
environmental pollution (1, 5). The growing use of this
by-product is mainly due to the great benefits that can be
obtained from lemon peel, as a source of health-promoting
compounds such as essential oil (EO), pectin, and flavo-
noids, processing (1).
EOs are complex mixtures of different volatile and
non-volatile compounds (6) derived from plants second-
ary metabolism (7). Citrus EOs are used in perfumes and
cosmetics and also in foods and beverages (8) to reduce
synthetic additives (9) and as flavoring agents, as they
files (10). For example, lemon EOs (annual consumption
of 1960.78 metric tons) are utilized as flavoring agents
in baked and confectionery products, desserts, and soft
drinks (10, 11).
Traditionally, recovery of Citrus EOs is performed by
cold pressing their peels. None of the mechanical or man-
ual processes used to extract EOs from Citrus peels are
able to recover the total quantity of EOs. For determining
the total EOs contents of Citrus, the Clevenger appara-
tus based on hydrodistillation (HD) is the best method
(10, 12, 13). EOs are thermally sensitive and vulnerable
to chemical changes (14), and thus conventional HD has
some disadvantages, for example, loss of some volatile
compounds, low extraction efficiency, degradation of
unsaturated and ester compounds through thermal and
hydrolytic effects (14). These shortcomings have led to
consideration of the use of methods for bioactive com-
pounds extraction, such as supercritical fluid extraction
(15), ultrasound-assisted extraction (16), microwave-as-
sisted hydrodistillation (MAHD) (17, 18), and solvent-free
microwave extraction (SFME) (14, 19). Generally, these
extraction methods, increase the yield of EOs, maintain
their qualities and use less energy.

CONTACT Mohammad-Taghi Golmakani golmakani@shirazu.ac.ir

2016 Taylor & Francis

Microwave dielectric heating, as a substitute for the
conventional heating methods, has received much atten-
tion in many various processes in the food industry in
recent years. From among the developed microwave-as-
sisted extraction methods, MAHD has been investigated
with regard to its efficiency in extracting EOs from several
dried plant materials, including Buniumpersicum Boiss.
(20), Saturejahortensis, and Saturejamontana (21). SFME
is an original combination of microwave heating and dry
distillation of fresh plant materials (14). In addition, SFME
has been used to obtain EOs from Salviasomalensis (22).
The advantages of using microwave energy, as a non-con-
tact heat source, in extracting of EOs from aromatic plant
materials include more effective heating, faster energy
transfer, reduced thermal gradients, selective heating,
reduced equipment size, faster response to process heating
control, faster start-up, increased production, and elimi-
nation of process steps (13).
Despite the fact that many studies have investigated the
effect of different microwave extraction methods, no study
has yet compared MAHD and SLME (solvent-less micro-
wave extraction) with regard to their yield in extracting
EOs from fresh and dried lemon peels. Therefore, the
objective of this study was to investigate the potential of
MAHD and SLME for extraction of EOs from dried and
fresh Eureka lemon peels, respectively. An attempt was
also made to compare their extraction time, extraction
yield, scanning electron microscopy (SEM) images, physi-
cal properties, chemical composition, antioxidant activity,
and energy consumption with those of conventional HD
method.
2. Materials and methods
2.1.Chemicals
In order to conduct this study, 2, 2-diphenyl-1-picrylhy-
drazyl-free radical (DPPH), TBHQ (tert-Butylhydroqui-
none), methanol, and hexane supplied by Sigma-Aldrich
(St. Louis, MO).
2.2. Plants material
The Eureka lemons used in this study were Citruslimon
(L.) Burm. F. which were gathered from an indigenous
crop in Jahrom (Fars province, Iran) in March 2012. The
genus and variety of lemon were further approved by a
plant taxonomy expert of Department of Horticultural
Science, College of Agriculture, Shiraz University, Shiraz,
Iran. The lemons were peeled by hand, separating the
external part of the lemon (flavedo), giving a yield of 34.00
3.04% (w/w) of lemon peel with respect to the whole
fruit. The moisture contents of the fresh and dried lemon
Journal of Essential Oil Research 273
peels were measured in triplicate according to Golmakani
and Rezaei (17, 18) method. The initial moisture content
of the fresh lemon peel was 82.75 1.04% (w/w). The
lemon peel was then placed on a large screen tray for
3 days and dried under ambient conditions (35 5C).
Then, they were packed in polyethylene (PE) bags and
kept in a dark and cool place for further experiments. The
measured moisture content of dry peel was 5.78 0.06%
(w/w). All the obtained values have been reported on a
dry weight (moisture-free) basis.
2.3. Comparison of different extraction methods
2.3.1. Microwave-assisted hydrodistillation
In employing MAHD, a domestic microwave oven
(ME3410W, Samsung, Malaysia) was used. It operated at
2.45 GHz frequency with a maximum output power of
1000 W. Fifty gram of dried lemon peel and 450 of mL
distilled water (in a peel-to-water ratio of 1:9) were heated
using a fixed power of 1000 W (100%) for 15 minutes.
Every 5 minutes (i.e. 5, 10, and 15 minutes), the collected
EO was decanted from the condensate. Yields are reported
in grams of EOs per 100 g of dried lemon peel.
2.3.2. Solvent-less microwave extraction
SLME was employed through using a similar oven as the
one described above. The internal heating of the in situ
water within the lemon peel distends the EO glands and
leads to the rupture of the glands. This process thus frees
EO which is evaporated by the in situ water of the plant
materials. SLME is neither a modified HD, which uses an
Electromantle and a large quantity of water, nor a mod-
ified MAHD which uses a large quantity of water. In the
case of SLME, 250 g of fresh lemon peel and distilled water
(in a peel-to-water ratio of 1:1) were heated using a fixed
power of 1000 W. In SLME, we used an equal ratio of water
to fresh lemon peel in order to prevent peel from burning
and to ensure that sufficient amount of water would be
remained during refluxing. The extraction time was fixed
at 15 minutes until no more EO was obtained. During this
period, the collected EO was decanted from the conden-
sate in 5 minutes intervals.
2.3.3.Hydrodistillation
HD was used in a similar way to MAHD. However,
an Electromantle heater (EM2000/C, Electrothermal
Engineering Ltd., UK, 335 W) was used instead of the
microwave oven and the operation was performed for
120 minutes in 30 minutes intervals (i.e. 30, 60, 90, and
120 minutes). The EOs obtained from the three extraction
methods were collected, dried with anhydrous sodium
sulfate and stored in amber vials at 4C until they were
used for further analysis.
274 M.-T. Golmakani and M. Moayyedi
2.4. Physical properties
Specific gravity and the refractive index of the EOs
extracted by the three different extraction methods from
the lemon peels were measured at 25 and 20C, respec-
tively, according to the methods suggested by Food
Chemical Codex (FCC) [10]. Color of the EOs (L*: bright-
ness, a*: redness-greenness, and b*: blueness-yellowness)
was determined according to the method described by
Afshari-Jouybari and Farahnaky (23). In addition, color of
the EOs was determined visually as directed in FCC [11].
In the L*a*b* color space, color difference can be expressed
as a single numerical value, Eb, which indicates the size
of the color difference but not in what way the colors are
different Eab of EOs was calculated according to the fol-
lowing Equation (24):
Eb = (L)2 + (a)2 + (b)2 (1)
2.5. Scanning electron microscopy
SEM images of lemon peels were obtained for the untreated
sample as well as the treated samples after HD (for 120
minutes), MAHD (for 15 minutes), and SLME (for 15
minutes). The lemon peels were freeze-dried using a freeze
drier (Armfield, UK) and fixed on the specimen holder
with aluminum tape and then sputtered with gold in a
sputter coater (Polaron SC7640, UK). All the specimens
were examined with an SEM (Cambridge, UK) under high
vacuum conditions at an accelerating voltage of 20.0 kV
and a working distance of 1316 mm (i.e. the distance
between the surface of the sample and the microscope
lens).
2.6. Gas chromatography (GC)
Identification of EO constituents (qualitative analysis)
was made using a GC (Agilent Technologies 7890A, USA)
coupled with a mass spectrometer (Agilent Technologies
5975C, USA), operating at 70 eV ionization energy, 0.5
s/scan, and the mass range of 35400 atomic mass unit
(amu), equipped with a HP-5MS capillary column (5%
Phenyl Polysilphenylene-siloxane; 30 m 0.25 mm ID;
0.25 m film thickness). The injector and detector tem-
peratures were both 280C. The oven temperature was
programmed to start at 60C. The oven was heated to a
temperature of 210C (rate: 3C/minute). Then the tem-
perature increased to 240C (rate: 20C/minute) and,
here, the temperature was held constant for 8.5 minutes.
One L of sample was injected into the GC in the split
mode (split ratio: 1/100). Helium was used as the carrier
gas with a flow rate of 0.9 mL/minute. MSD ChemStation
(G1701EA, E.02.01.1177 USA) software was used to han-
dle mass spectra and chromatograms. Retention indices
(RIs) were calculated through retention times (RTs) of
C5C28 n-alkanes that were injected after the EOs under
the same chromatographic conditions. The compounds
were identified by comparing their mass spectral frag-
mentation patterns with those stored in the data bank
(Wiley/NBS) and with mass spectra literature data (25).
Also, quantitative analysis of EO constituents was made
under the same chromatographic conditions using a GC
coupled with a flame ionization detector (FID). Figure 1
shows the GC/FID chromatogram of C. limon peel EO
extracted by hydrodistillation method. As it was done
in study by Gavahian et al. (26) relative percentage data
were obtained from electronic integration of chromato-
gram peak areas.
2.7. Antioxidant activity: DPPH scavenging
activity
Therefore, antioxidant activity of EOs was evaluated
using DPPH free radical (DPPH) scavenging method
as described by Brand-Williams et al. (27). The effect of
antioxidant on DPPH is a result of antioxidant hydrogen
donating ability or radical-scavenging activity. Mixing the
DPPH solution with a substrate used for donating hydro-
gen atom leads to the reduced form (non-radical DPPH)
of this reagent and simultaneous change of its color from
violet to pale yellow. Four mL of different concentrations
of the EOs (2, 4, 6, and 10 mg/mL) in methanol was added
to 2 mL of 0.2 mM methanol solution of DPPH. The mix-
tures were shaken vigorously and then they were placed at
room temperature for 60 minutes in the dark. Then, the
absorbance values were recorded at 517 nm against the
blank (methanol). The percentage inhibition of DPPH by
the EO sample was calculated according to the following
Equation (28):
Inhibition% = ((AC AS )AC ) 100 (2)
where AC is the absorbance of the control reaction (with-
out any EO) and AS is the absorbance of the EO sample
after 60 minutes. Also, TBHQ was used as a positive con-
trol on antioxidant activity. DPPH scavenging activity
is shown by IC50 value, defined as the concentration of
the antioxidant required for the 50% inhibition of the
DPPH activity (20). IC50 value was determined through
the graph plotting percentage of remaining DPPH against
EO concentrations.
2.8. Statistical analysis
All the extractions by HD, MAHD, and SLME and also
evaluation of chemical and physical properties of extracted
EOs were done in triplicate and a general linear model
(GLM) procedure in SAS (Statistical Analysis Software,

Figure 1. Chromatogram of lemon peel essential oil obtained by hydrodistillation.
Version 9.1, SAS Institute Inc., Cary, NC) software was
used to compare the means.
3. Results and discussion
3.1. Comparison of temperature profiles of the
extraction methods
In order to determine the effect of HD, MAHD, and
SLME, temperature profile during EO extractions from
lemon peels was examined (Figure 2). The extraction
temperature in HD, MAHD, and SLME methods is equal
to the boiling point of water (100C) at the atmospheric
pressure. With regard to all the extractions, the initial tem-
perature of samples was 25C. The extraction temperature
marks the first droplets of extractions (23.0 minutes in
HD, 3.5 minutes in MAHD, and 3.0 minutes in SLME).
Yet, two phases were observed in the process of increas-
ing extraction temperature in HD, MAHD, and SLME
methods. The first part was represented by an increasing
line until the temperature reached 100C and thus distil-
lation of the first EO droplets was observed. In the second
part, the extraction temperature was equal to the boiling
temperature (100C) of water at the atmospheric pressure
until the end of extraction. The rate of temperature rise
was obtained through determining the slope of the linear
part of the temperature profile (Figure 2a).
Table I1 shows that the rates of temperature rise in
MAHD and SLME methods were nearly seven-fold greater
than that in HD method. The data also indicate that there
was a significant difference in the rate of temperature rise
between microwave oven and Electromantle. These results
Journal of Essential Oil Research 275
can be due to a slighter energy loss in microwave oven
than in the Electromantle. Still the most important reason
is that MAHD and SLME utilize three ways of heat trans-
fer within the samples, namely irradiation, conduction,
and convection while in HD, heat transfer only takes place
through conduction and convection (17, 18). The cells
were thermally stressed during irradiation and there was
a steady rise in the temperature of the cells. Consequently,
cell walls and EO glands ruptured faster and the induc-
tion stage was almost eliminated. The results can also be
attributed to the more powerful effect of microwaves on
water, as a solvent with a high dielectric constant (21).
Lucchesi et al. (19) showed that the required heating time
to reach the extraction temperature (100C) and to obtain
the first EO droplets of ajowan, cumin, and star anise was
only 5 minutes in SFME method while it was 90 minutes
in HD method. Golmakani and Rezaei (17, 18) observed
that the boiling point (100C) was reached in 7 minutes
in MAHD. However, the starting time of EO extraction
from Thymusvulgaris L. and Zatariamultiflora Boiss. took
place after 30 minutes in HD.
3.2. Comparison of extraction kinetics and
extraction yield
Table 1 shows the effect of different extraction methods
on starting time of EO accumulation, total extraction
time, extraction duration (i.e. the difference between the
total extraction time and the extraction starting time),
yield, and rate of EO accumulation. Extraction in MAHD
and SLME started at much earlier time (after 3.5 and 3
276 M.-T. Golmakani and M. Moayyedi
Figure 2. Extraction (a) temperature and (b) yield as a
function of time in hydrodistillation (HD), microwave-assisted
hydrodistillation (MAHD), and solvent-less microwave extraction
(SLME) of essential oil from lemon peels. Different letters are
significantly different (p < 0.05).
minutes, respectively) than extraction in HD (after 23
minutes). This might be due to the fact that a more effi-
cient heat flow was provided by the microwave oven.
The extraction time in MAHD and SLME was faster
than that in the conventional HD. The extraction pro-
cess took 15 minutes in MAHD and SLME while it took
120 minutes in HD. Lucchesi et al. (14) found that less
time was required for EO extraction from basil, garden
mint, and thyme in SFME (30 minutes) than in HD (270
minutes). Also, Golmakani and Rezaei (17, 18) presented
similar findings in their study on EO extraction from
Thymusvulgaris L. and Zatariamultiflora Boiss. through
HD (4 hours) and MAHD (2 hours).
Table 1. The effects of different essential oil (EO) extraction methods (hydrodistillation (HD), microwave-assisted hydrodistillation
(MAHD), and solvent-less microwave extraction (SLME)) from lemon peels on extraction kinetics and energy consumption.
Extraction parameters
Rate of temperature increase
Starting time of EO accumulation (min)
Total extraction time (min)
Extraction duration (min)
Yield (%, w/w)
Rate of EOs accumulation (g/min)
Electric consumed (kWh)
Relative electric consumption (kWh/g EOs)
CO2 rejected (g)
Relative CO2 rejection (g/g EOs)
Note: *In each row means with different letters are significantly different (p < 0.05).
The changes in the extraction yield of EOs in HD,
MAHD, and SLME, are shown in Figure 2b. In the three
cases, the extraction yield increased by as increase in the
extraction time. Extraction of EOs started after 23 minutes
in HD when extraction in MAHD or SLME had been
completed. After 15 minutes, the use of MAHD and SLME
methods resulted in significantly equivalent yields (1.12
0.06 and 1.26 0.08% w/w, respectively) in comparison to
the yield obtained in HD after 120 minutes (1.18 0.14%
w/w). Since the EO quantity of samples was constant and
since we had no EO loss caused by evaporation, the final
yields in the three methods (HD, MAHD, and SLME)
were equal when the extraction processes were com-
pleted. Ferhat et al. (13) extracted EO from orange peel
and reported that the yields by HD and SFME were 0.39
0.02 and 0.42 0.02% w/w, respectively. They observed
that there were no significant differences between the
yields by HD and SLME. Furthermore, Bousbia et al. (12)
investigated the effect of microwave-hydrodiffusion and
gravity on total extraction time and yield of EOs extracted
from Citruslimon. They found that the yields in HD and
microwave-hydrodiffusion and gravity were 0.8 and 0.7%
w/w, respectively and there were no significant differences
in the yields obtained through HD and microwave-hy-
drodiffusion and gravity.
The extraction yield in SLME after 10 minutes (1.24
0.08% w/w) was similar to that in the conventional HD
after 120 minutes (1.18 0.14% w/w) and in MAHD after
15 minutes (1.12 0.06% w/w). In SLME, large amounts
of interior heating resulted in increased moisture vapor
generation inside the sample which created high interior
pressure and concentration gradient (29). In addition, the
extraction yields by MAHD and SLME (1.08 0.06 and
1.21 0.07% w/w, respectively) after 5 minutes were sig-
nificantly equal to the yield obtained by the conventional
HD after 120 minutes.
The rates of EO accumulation in HD, MAHD, and
SLME are shown in Table 1. This index was obtained by
dividing the amount of extracted EOs (g) to the corre-
sponding total extraction time (minute), which was equal
HD MAHD SLME
3.26 0.23c* 21.43 1.51b 25.00 1.76a
23.00a 3.50b 3.00b
120.00a 15.00b 15.00b
97.00a 11.50b 12.00b
1.18 0.14a 1.12 0.06a 1.26 0.08a
0.010 0.00c 0.070 0.00b 0.080 0.00a
0.67a 0.25b 0.25b
0.56a 0.22b 0.20b
532.80a 200.00b 200.00b
451.53a 178.57b 158.73b

to the average rates of EO accumulation (g/min). The
results show that the average rates of EO accumulation in
MAHD and SLME were at least seven times greater than
that in HD. This theme (the presence of any significant
difference between the microwave extraction and the con-
ventional HD method) was further examined with regard
to total extraction time. Additionally, the findings revealed
that the less total extraction time in MAHD and SLME
was not only caused by the earlier extraction starting times
in MAHD and SLME, but also by the higher extraction
rates in MAHD and SLME. Unlike the classical conduc-
tive heating methods, microwave extraction methods
can heat the entire sample almost simultaneously and at
a higher rate (30). Moreover, the higher pressure gradient
was formed inside the plant materials during microwave
heating in MAHD and especially in SLME.
3.3. Structural changes during the extraction of
essential oil
In lemon, the development of the fruit consists in the for-
mation of three strata: the exocarp (flavedo), the meso-
carp (albedo), and the endocarp. In the outer layers of
the exocarp, groups of cells become secretory-forming
cavities (glands) filled of EOs rich in terpenes (31). The
SEM images of the untreated and treated lemon peels after
EO extraction by HD, MAHD, and SLME are shown in
Figure 3. The untreated lemon peel (Figure3a) displayed
a regular and compact surface structure and the cell
walls were intact. The three extraction methods resulted
in apparent physical changes in the lemon peel glands.
Images of the lemon peels which were used in HD (with
a total extraction time of 120 minutes), in MAHD (with
a total extraction time of 15 minutes) and in SLME (with
a total extraction time of 15 minutes) are presented, for
the sake of comparison, through Figures 3b, 3c, and 3d,
respectively. While the glands in MAHD and SLME were
completely destroyed after 15 minutes, the extraction in
HD had not been started yet. This indicates that micro-
wave heating causes the faster rupture of the glandular
walls. Wang et al. (32) in their study on EO extraction
from Mango flower through MAHD and HD, presented
similar findings. They reported that microwaves caused
the glandular walls to rupture faster, resulting in high
extraction efficiency in less time.
According to Figure 3, glands in the HD method were
wrinkled while the glands in the microwave-assisted
methods (especially those in SLME) were smoother.
Such a difference can be attributed to different rate of
heat transfer among the three different extraction meth-
ods. Also, Golmakani and Rezaei (17, 18) in their study
on the extraction of EOs from Thymusvulgaris L. and
Zatariamultiflora Boiss. reported similar results.
Journal of Essential Oil Research 277
When the glands were under a great thermal stress, as
it was the case with SLME (Figure 3d) and MAHD (Figure
3c), the localized high pressure build-up within the glands
exceeded the capacity of the glands for expansion and, as
a result, a faster rupture of the cell walls, in comparison
to the conventional HD method, was observed (13). By
Pare and Belanger (33) stated that explosion occurred at
the cell level as a consequence of a sudden temperature
rise, generated in that case by located hot spots caused by
microwave heating.
3.4. Evaluation of physical properties
The mean values for the physical properties (refractive
index, specific gravity, and color) of EOs extracted from
lemon peels by HD, MAHD, and SLME methods are
shown in Table 2. There were no significant differences
between the extracted EOs through HD, MAHD, and
SLME methods in specific gravities, refractive indices, and
color. Specific gravity, refractive index, and color percep-
tions of all the samples were within the range indicated by
FCC standards (11). The results of the physical examina-
tion of the extracted EOs from lemon peels revealed that
MAHD and SLME, as novel and fast extraction methods,
did not introduce any noticeable changes to the measured
physical properties of EOs extracted from lemon peels.
Eab of EOs extracted by MAHD and SLME (in compar-
ison to HD as a reference method) were 8.35 and 5.61,
respectively. There was a clear similarity in the color of
EOs extracted by HD or MAHD or SLME. Ferhat et al.
(13) observed no significant difference between HD and
SFME in terms of the physical properties of EOs extracted
from orange peels. Also, Wang et al. (32) found that the
specific gravities and refractive indices of Mango flower
EOs extracted by HD and MAHD were similar, with the
only difference being that the color of EOs extracted by
MAHD was lighter than that of the HD EOs.
3.5.GC
Table 3 presents qualitative and quantitative description
of different EO extraction methods. This table includes
the retention indices and percentages of the areas under
curves of 38 identified constituents. The constituents are
grouped into four classes: oxygenated terpenes, mono-
terpene hydrocarbons, sesquiterpene hydrocarbons, and
aliphatic aldehydes.
Results show that there are many similarities in the
quality of EOs extracted by HD, MAHD, and SLME.
Based on the obtained values by GC, it was indicated
that there were no significant differences in the quan-
tity of the constituents with relative peak areas more
than 2% (the total percentage of these constituents was
278 M.-T. Golmakani and M. Moayyedi

Figure 3. Scanning electron micrographs of lemon peels: (a) untreated, (b) after hydrodistillation (HD) for 120 minutes, (c) after
microwave-assisted hydrodistillation (MAHD) for 15 minutes, and (d) after solvent-less microwave extraction (SLME) for 15 minutes.

Table 2.Physical properties of essential oils extracted from lemon peels by hydrodistillation (HD), microwave-assisted hydrodistillation
(MAHD), and solvent-less microwave extraction (SLME).
Physicalproperties FCC1 HD MAHD SLME
Specific gravity (25C) 0.8420.856 0.853 0.021 0.853 0.006 0.850 0.026
Refractive index (20C) 1.4701.475 1.474 0.001 1.473 0.000 1.473 0.000
Appearance Pale yellow Pale yellow Pale yellow Pale yellow
L*2 65.50 1.00 64.75 0.50 65.00 0.00
a* -12.25 0.50 -8.25 0.50 -9.75 0.50
b* 10.75 0.50 3.25 0.50 5.75 0.50
Eab3 8.53 5.61

Notes: 1Standard physical property of lemon essential oil according to FCC (11); 2L*: brightness; a*: redness-greenness; b*: blueness-yellowness;
3
Relative to hydrodistillation method.

75.1182.69%) in the different extraction methods.
Limonene, a monoterpene hydrocarbon, appeared to
comprise the highest percentage among the constit-
uents in the EO when extraction was done by HD
(65.05%), MAHD (72.26%), and SLME (68.15%). From
the EOs of lemon peel, two other constituents were also
extracted by HD, MAHD, and SLME. These two were
notably -terpinene (6.536.77%) and -pinene (3.42
3.69%). There were no significant differences among
the amounts of limonene, -terpinene, and -pinene
extracted by the three methods. Bousbia et al. (12)
reported similar findings with regard to the constit-
uents of C. limon (Eureka variety). They showed that
limonene was the main constituent of EOs extracted
from lemon, with almost equal amounts when extracted
by HD (65.25%), cold pressing (68.81%), and micro-
wave-hydrodiffusion and gravity (69.65%).
Monoterpene hydrocarbons accounted for 80.19
87.97% of the total identified constituents in the ana-
lyzed EOs. Moreover, monoterpene hydrocarbons in EOs
extracted by MAHD and SLME were more in amount than
those extracted by the HD. However, this appears to con-
trast with the findings by Ferhat et al. (13). They showed
that monoterpene hydrocarbons in EOs extracted by the
SFME (85.7%) can have lower concentrations compared
to the condition when the EO was extracted by the HD
(89.8%).
The concentrations of oxygenated constituents such
as oxygenated terpenes and aliphatic aldehydes reached
3.839.49% and 0.020.84%, respectively. The mentioned
 Journal of Essential Oil Research 279

Table 3.Chemical compositions of essential oils extracted by different extraction methods from lemon peels using gas chromatography.
No. Compounds RI1 Relative peak area[%]
HD2 MAHD3 SLME4
Monoterpene hydrocarbons
1 -Thujene 924 0.33 0.36 0.34
2 -Pinene 932 0.60 0.64 0.60
3 Camphene 946 0.10 0.10 0.08
4 Sabinene 971 1.19 1.28 1.14
5 -Pinene 977 3.53 3.69 3.42
6 -Myrcene 989 1.62 1.73 1.66
7 -Phellandrene 1004 0.14 0.15 0.15
8 -Terpinene 1015 0.12 0.10 0.11
9 p-Cymene 1024 0.31 0.28 0.22
10 Limonene 1036 65.05 72.26 68.15
11 Cis--Ocimene 1038 0.03 0.03 0.04
12 Trans--Ocimene 1046 0.12 0.12 0.17
13 -Terpinene 1059 6.53 6.74 6.77
14 Terpinolene 1087 0.52 0.49 0.50
Aliphatic aldehydes
15 n-Octanal 1001 0.79 ND ND
16 n-Nonanal 1102 0.05 0.02 0.02
Oxygenated terpenes
17 Linalool 1098 0.39 0.12 0.14
18 Cis-Limonene oxide 1131 0.15 0.10 0.05
19 Trans-Limonene oxide 1135 0.08 0.03 0.04
20 Camphore 1142 0.06 ND5 ND
21 Citronellal 1150 0.58 0.29 0.42
22 Terpinen-4-ol 1174 0.55 0.04 0.05
23 -Terpineol 1187 0.96 0.17 0.28
24 Nerol 1225 0.23 0.05 0.09
25 Neral 1238 1.38 0.69 0.32
26 Carvone 1240 0.24 0.03 0.06
27 Geraniol 1251 0.69 0.12 0.38
28 Geranial 1268 1.30 0.38 0.93
29 Citronellyl acetate 1350 0.52 0.34 0.29
30 Neryl acetate 1362 1.00 0.62 0.70
31 Geranyl acetate 1381 1.36 0.85 0.84
Sesquiterpene hydrocarbons
32 Cis--Bergamotene 1411 1.41 1.18 1.62
33 (E)-Caryophyllene 1415 0.92 0.78 1.12
34 Trans--Bergamotene 1432 0.56 0.47 0.69
35 Valencene 1489 0.61 0.21 0.59
36 Bicyclogermacrene 1492 0.60 0.61 0.90
37 Cis--Bisabolene 1499 0.58 0.50 0.62
38 -Bisabolene 1505 1.13 0.90 1.38
Monoterpene hydrocarbons 80.19 87.97 83.35
Aliphatic aldehydes 0.84 0.02 0.02
Oxygenated terpenes 9.49 3.83 4.59
Sesquiterpene hydrocarbons 5.81 4.65 6.92

Notes: 1Retention index; 2Hydrodistillation; 3Microwave-assisted hydrodistillation; 4Solvent-less microwave extraction; 5Not detected.

ranges of percentages result from the three different
extraction methods that make the EOs of lemon peel
appear with various concentrations of the constituents.
Oxygenated terpenes and aliphatic aldehydes extracted
by HD were present in higher amounts in comparison
with those of microwave-assisted extraction methods.
When extracted by the HD method, some constituents
were observed to occur in higher amounts than when
extracted by MAHD and SLME. These constituents are
namely alcohols (such as linalool, terpinen-4-ol, -terpi-
neol, nerol, and geraniol), aldehydes (such as citronellal,
neral, and geranial), and esters (such as citronellyl ace-
tate, neryl acetate, and geranyl acetate). These differences
result from the longer total extraction duration (being
8 times more) in the HD method than the MAHD and
SLME methods. The longer total extraction duration can
help with the extraction of heavier EO fractions such as
oxygenated terpenes and aliphatic aldehydes, as observed
toward the ending of each extraction. Contrary to our
results, Ferhat et al. (13) showed that the oxygenated con-
stituents in the EO extracted by HD (7.9%) occurred in
lower amounts compared to when the EO was extracted
by the SFME (11.7%).
Based on the obtained values by GC, monoter-
pene hydrocarbons were found in more considerable
amounts than oxygenated terpenes in HD, MAHD, and
SLME. Monoterpene hydrocarbons are less valuable
than oxygenated terpenes as they make a small contri-
bution to the fragrance of the EO. Conversely, oxygen-
ated terpenes are highly odoriferous and hence they
280 M.-T. Golmakani and M. Moayyedi
are the most valuable constituents of EOs (19). Ferhat
et al. (13) showed that substantially larger amounts of
monoterpene hydrocarbons (89.8% in HD and 85.7%
in SFME) and smaller amounts of oxygenated fraction
(7.9% in HD and 11.7% in SFME) were present in the
EOs extracted from orange peels extracted by HD and
SFME. Lucchesi et al. (19) indicated that the constitu-
ents of basil, garden mint, and thyme EOs extracted by
HD and SFME were similar. Furthermore, Golmakani
and Rezaei (17, 18) reported that chemical composi-
tions of the EOs of two genotypes of thyme extracted
by MAHD were similar to chemical compositions
obtained by the conventional HD method. In all of the
above mentioned applications, microwave irradiation
greatly accelerated the extraction process without caus-
ing any considerable change to the EO profile.
3.6. Cost, cleanliness and up-scaling
The reduced time extraction is clearly advantageous for
the proposed microwave extraction in terms of cost and
energy. For starting extraction, HD required 23 minutes
(at 335 W). Furthermore, evaporation of water and EO
took 97 minutes in HD (at 335 W). MAHD required only
3.5 minutes in the initial stage and an additional 11.5 min-
utes to pass the final stage (at 1000 W). The use of this
method resulted in a significant saving of time. Besides,
for the extraction to start in SLME, no longer than 3 min-
utes was required (at 1000 W) and evaporation of the in
situ water and EO of the lemon peels took 12 minutes (at
1000 W). The required energy for the complete extraction
of EOs, based on the total period of a full recovery and the
maximum power consumption of the Electromantle (in
HD) and microwave oven (in both MAHD and SLME),
was 0.67 kWh for HD and 0.25 kWh for MAHD and SLME
(Table 1). The energy consumption for the production of
1 g of EO was estimated as well. This theme (the energy
consumption for the production of 1 g of EO) in HD,
MAHD, and SLME was 0.56, 0.22, and 0.20 kWh/g EO,
respectively (Table 1). Therefore, there was a significant
decrease in the extraction cost when MAHD and SLME
were used rather than HD. Lucchesi et al. (14) reported
that the energy required for extraction of EOs from basil,
garden mint, and thyme was 4.5 kWh in HD and it was
0.25 kWh in SFME. Also, Golmakani and Rezaei (17, 18)
showed that the energy which was required for the extrac-
tion of thyme EOs to take place was 2.00 kWh in HD and
1.24 kWh in MAHD.
Regarding environmental impact of pollution, the find-
ings indicated that the amount of carbon dioxide emit-
ted to the atmosphere via HD (532.80 g CO2) was larger
than that in MAHD and SLME (200.00 g CO2).These
measurements were carried out based on the procedures
mentioned in the previous studies: to obtain 1 kWh of
energy from coal or fuel, 800 g of CO2 will be released
into the atmosphere during combustion of fossil fuels (13).
The amount of carbon dioxide released into the atmos-
phere for the production of 1 g of EO was larger in HD
(451.53 g/g of EO) than that in MAHD (178.57 g/g of
EO) and SLME (158.73 g/g of EO). Lucchesi et al. (14)
showed that the amount of carbon dioxide released into
the atmosphere was dramatically larger in HD (3600 g
CO2/g of EO) than that in SFME (200 g CO2/g of EO).
Golmakani and Rezaei (17, 18) found that the amount of
carbon dioxide which was released into the atmosphere
was great in HD (1600 g CO2/g of EO) than that in MAHD
(990 g CO2/g of EO).
3.7. Antioxidant activity: DPPH scavenging
activity
The method of Brand-Williams et al. (27) was used to
compare the antioxidant activities of the lemon peel EOs
extracted by HD, MAHD, and SLME with that of known
synthetic antioxidant, TBHQ. A lower IC50 value reflects
a better protective action. The IC50 values of the EOs
extracted by HD, MAHD, and SLME were 60.46, 61.40,
and 76.45 mg/mL, respectively and there was no signifi-
cant difference among different extraction methods in this
regard. As Di Vaio et al. (34) found, antioxidant activity
might be mainly related to terpenes constituents; however,
a small contribution was also made by other substances
with radical scavenging activities which exist in the EOs.
Since the qualitative and quantitative results revealed no
significant differences among EOs constituents, it was pre-
dicted that there would not be any significant difference in
the IC50 values of the extracted EOs by the three different
methods. These results are in agreement with the findings
of Mazidi et al. (20). They showed that there were no sig-
nificant differences between the IC50 values of Black Zira
EOs extracted by HD and MAHD.
4.Conclusions
Two green methods for the extraction of EO from lemon
peels were used and compared in this study: MAHD and
SLME. These alternative extraction methods were examined
using dried and fresh lemon peels. Based on the findings,
MAHD and SLME methods have some important advan-
tages over the conventional HD method, advantages such
as less extraction time, and thus, less energy consumption
and lower extraction cost. SEM images of treated lemon
peels emphasized the differences among different extrac-
tion methods. Glands ruptured more rapidly in MAHD and
SLME methods than in the conventional HD method. No
significant differences were found in the physical properties

(refractive index, specific gravity, and color) of EOs extracted
by HD, MAHD, and SLME. The EOs which were extracted
by the three extraction methods had the same compositions.
At the same time, the amount of carbon dioxide which was
released into the atmosphere, as a result of the EO extraction
process, was dramatically higher in HD than that in MAHD
and SLME. DPPH analysis of the extracted EOs indicated
that microwave irradiation did not adversely influence anti-
oxidant activity of the extracted EOs. Such advantages can
make MAHD and especially SLME as excellent and environ-
mentally friendly alternative EO extraction methods from
dry and fresh samples, respectively.
Acknowledgments
This research project was financially supported by Shiraz Uni-
versity. We also like to thank Mohsen Hamedpour-Darabi for
his native English editing service.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding
This work was supported by Shiraz University.
ORCID
Mohammad-Taghi Golmakani http://orcid.org/0000-0001-
5173-1178
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