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Volume 16, No. 61, Winter 2017
Journal of Medicinal Plants
The Influence of Drying Treatments on the Essential Oil Content and
Composition of Melissa officinalis L. Compared with the Fresh
Sample
Mirahmadi SF (M.Sc.)1*, Norouzi R (M.Sc.)2, Ghorbani Nohooji M (Ph.D.)3
1- Department of Agriculture and Natural Resources, Horticulture science, Velayat
University, Iranshahr, Sistan & Balouchestan, Iran
2- Meshginshahr Faculty of Agriculture, University of Mohaghegh Ardabili,
Ardabil, Iran
3- Medicinal Plants Research Center, Institute of Medicinal Plants, ACECR, Karaj, Iran
* Corresponding author: Department of Agriculture Engineering, Horticulture
science, Velayat University, P.O.Box: 9911131311, Iranshahr, Sistan &
Balouchestan, Iran
Tel & Fax: +98 547-3312521
E-mail: Fazel.mirahmadi@gmail.com, f.mirahmadi@velayat.ac.ir
Received: 24 Oct. 2016 Accepted: 1 Feb. 2017
Abstract
Background: In the recent decade, artificial drying has been one of the most important needs of the
pharmaceutical industries. In addition, different drying methods have different effects on the quantity
and quality of the essential oils produced from medicinal plants.
Objective: The main objective of this study was to evaluate the effects of different drying
methods (shade and oven drying at 35 °C and 55 °C) in comparison with the fresh sample on the
essential oil yield and volatile composition of M. officinalis.
Methods: This experiment was conducted in completely randomized design with three
replicates. The essential oil samples were isolated by hydrodistillation in a Clevenger type
apparatus and analyzed using GC and GC–MS methods.
Results: Different drying treatments had a significant effect on the content of M. officinalis
essential oil (ranging from 0.08 to 0.3 % v/w; overall average of 0.22% v/w). The main
components of the essential oil of shade dried, oven-dried at 35 °C and oven-dried at 55 °C
samples were β-caryophyllene, geranial and γ-cadinene, respectively. Oven drying at 55°C
resulted in disappearance of neral, geranial and neryl acetate. Sesquiterpene hydrocarbons
constituted the principal fraction of all samples, followed by oxygenated monoterpenes, except the
oil of oven dried sample at 55°C in which oxygenated sesquiterpene represented as the second
main fraction.
Conclusion: The highest essential oil yield was obtained from oven drying at 35°C which
conserves the characteristic aroma of the spice, so this method seems to be more advisable for
drying of M. officinalis.
Keywords: Melissa officinalis, Drying method, Essential oil, GC and GC–MS
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The Influence of …
Introduction
Melissa officinalis L. (Family: Lamiaceae),
popularly known as lemon balm is one of the
oldest and most common aromatic and
medicinal herbs [1]. This Plant is native to
India, Africa and Southern Asia and nowadays
cultivated world-wide for its characteristic
lemon-scented leaves [2]. This perennial bushy
and upright herb, with local name of “varang-
bou” in Iran, puts out numerous herbaceous
stems reaching a height of about 1 m, and
grows widely in provinces of Alborz, Tehran,
Golestan, Azarbayjan, Lorestan and
Kermanshah [3, 4]. The leaves of
M. officinalis have been used traditionally to
treat in the folk medicine because of their
sedative, aromatic, digestive and
antispasmodic properties as well as additive in
food, herbal tea, and ingredient in cosmetics,
ornamental and medicinal usages [5]. It also
has antidepressant [6], neuro-protective [7],
anti-inflammatory [8], antioxidant, antifungal,
antibacterial [9], antiviral [10] and anti-
Alzheimer [11] effects. Nowadays, lemon
balm is gaining increasing interest in various
branches of industry (such as cosmetics,
pharmaceuticals, perfumes and food
industries) worldwide [12]. The essential oil of
lemon balm is considered the therapeutic
principle mainly responsible for most of the
biological activities mentioned, but plant
phenolic compounds (especially rosmarinic
acid and caffeic acid), are also considered to
contribute to the therapeutic potential of
M. officinalis [13]. To date, many
investigations have considered the volatile oil
composition of M. officinalis which contains
mainly monoterpenes, sesquiterpenes,
alcohols, aldehydes, ketones, esters and
miscellaneous compounds [13, 14].
Based on the previous studies lemon balm
essential oils showed different chemotypes in
various parts of the world due to climatic,
geographical and plant genetic background
differences. The essential oil composition of
M. officinalis L. grown in Morocco show
citronellal and isogeraniol as the main
constituents [15]. In other reports, the major
component were cedrane and 2,2,8,8-
tetramethy-l-5-nonanone in Iranian lemon
balm [16] and β-pinene and sabinene in oil of
M. officinalis grown in Greece [17]. Citrals
(geranial + neral) and citronellal were reported
as the main constituents of essential oil in
Brazilian [18] and Serbian and Montenegrin
[9] lemon balm. Results by Mrlianova et al.
(2002) cited Citrals and β- Caryophyllene
oxide as the major compounds in this plant
[19].
In recent decades, the demand for high-
quality medicinal herbs is permanently
increasing all over the world. Herbs can be
marketed as fresh or dried products. Fresh
herbs cannot be supplied in a profitable way to
all world-wide locations. Drying is the most
common and oldest method for post-harvest
preservation and a fundamental requirement to
achieve a high quality product [20]. The main
aim of drying products is to allow longer
periods of storage, minimize packaging
requirements and reduce shipping weights and
improve shelf life in an uncomplicated manner
[21, 22]. Volatile constituents are the most
sensitive component in the process of food
drying. Previous studies showed that the
method of drying had a significant effect on
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Journal of Medicinal Plants, Volume 16,
No. 61, Winter 2017
Mirahmadi et al.
the quality and quantity of the essential oils
and the proportion of the various components
in medicinal plants [22-24]. The impact of
shade, sun and oven drying on the yield and
chemical composition of essential oil of
Satureja hortensis [23] and Chamaemelum
nobile [25] has been reported in the literatures.
Furthermore, it was also reported that essential
oil content and chemical profile of
M. officinalis L. were affected by different
drying methods [2, 26-27].
These results demonstrate that the influence
of drying methods on lemon balm essential oil
depends mainly on plants characteristics and
chemotypes as well as drying parameter.
However, there is no report on the effect of
different drying methods on lemon balm
volatile oil in Iran. Thus, the aim of this study
was to assess the influence of drying methods
(shade and oven drying at 35 °C and 55 °C) in
comparison with the fresh aerial part sample
on the essential oil content and composition of
M. officinalis cultivated Karaj, Alborz
Province, Iran.
Materials and Methods
Plant Material
The seeds of M. officinalis were provided
by Zardband Medicinal Plants Production Co.,
Tehran, Iran. Planting was done with 30 cm
row spacing in plant nursery in mid-February
2012. Transport of transplanting to the main
field (Garden of Medicinal Plants Research
Campus located in the Botanical Gardens of
College of Agriculture and Natural Resources
of University of Tehran, Karaj, Iran) was done
with 50 cm row spacing and 30 cm plant
distance in mid-May 2013. The aerial parts of
the plants were harvested at full flowering
stage by hand in mid-July 2013. A voucher
specimen was deposited at the Herbarium of
Horticultural Science Department, University
of Tehran, Karaj, Iran. To study the effect of
the drying method, different drying treatments
including shade-drying and two temperatures
of oven-drying at 35 °C and 55 °C in
compared with the fresh sample were
investigated.
Isolation of the essential oil
The dried aerial parts of every treatment (50
g, three replicates) and the fresh aerial parts
(150 g, three replicates) were subjected to
hydrodistillation of 3 h using an all-glass
Clevenger-type apparatus. The oil was dried
over anhydrous sodium sulfate, and then, was
kept in a sealed vial at 4 ºC until analysis. The
percentage yields of the oils were calculated
based on the dried weight of plant material
according to volume/weight percent.
Oil analysis procedure
Gas chromatography analysis
GC analyses were performed using a
Perkin-Elmer gas chromatograph model 8700,
equipped with flame ionization detector (FID)
and HP-5MS capillary column (30 m x 0.25
mm, film thickness 0.25 μm). Oven
temperature was programmed from 80 °C to
220 °C at the rate of 4 °C/min; initial and final
temperatures were held for 3 and 10 minutes,
respectively. Detector (FID) temperature was
290 °C and injector temperature was 220°C.
Helium was used as carrier gas with a linear
velocity of 1.5 ml/ min. The percentages of
compounds were calculated by the area
normalization method, without considering
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The Influence of …
any response factors.
Gas chromatography/mass spectrometry
(GC-MS) analysis
GC–MS analyses were carried out in an
Agilent-Technologies (Little Falls, California,
USA) 6890N Network gas chromatographic
(GC) system equipped with a HP-5 MS fused
silica column (30 m × 0.25 mm i.d., film
thickness 0.25 μm); oven temperature was 80–
220 °C at a rate of 4 °C/min, transfer line
temperature 290 °C, carrier gas, helium, with a
linear velocity of 1.5 ml/ min, split ratio 1:100,
ionization energy 70 eV, scan time 1 s, and
mass range 50 –550 m/z.
Compounds identification
The components of the oils were identified
by comparison of their mass spectra with those
of a computer library or with authentic
compounds, and confirmed by comparison of
their retention indices, either with those of
authentic compounds or with data published in
the literature [28]. The mass spectra from the
literature were also compared [28]. The
retention indices were calculated for all
volatile constituents using a homologous series
of n-alkanes.
Statistical analysis
The experimental design was completely
randomized, with three replications. The
Analysis of variance (ANOVA) for essential
oil content affected by different drying
methods and fresh sample was conducted
using the SAS 9.1 software. Multiple
comparisons of means were carried out by the
Duncan’s Multiple Range Test. All statistical
significance was determined at the 1%
significance level.
Results
The analysis of variance for essential oil
content of M. officinalis is shown in Table 1.
Different drying treatments had a significant
effect (P ≤ 0.01) on the content of
M. officinalis essential oil. Mean comparisons
of different drying treatments on oil content
(Table 2) showed that dried plant materials in
oven-drying at 35 °C (0.3% v/w) and shade-
drying (0.3% v/w) yielded more essential oils
than the other materials. However, there was
no significant difference between these
treatments and oil content obtained from fresh
sample (0.23% v/w), so they were placed in
one group.
The eight important and common
constituents which are listed in Table 3 in
order of their elution on the HP-5MS column,
were the most abundant compounds in
different samples under study, representing
58.8-76.6% of the total volatiles concentration.
The percentages and relative concentrations of
these 8 compounds could be important in
determining the quality of the different dried
samples of lemon balm. The major
components obtained from fresh aerial part
were β-caryophyllene (21.8%), germacrene D
(15.5%), geranial (12.7%) and γ-cadinene
(12%).
The drying method caused some variation
on the relative proportions of the components.
The all major compounds showed no sharp
difference among the drying methods except
neral and geranial that did not exist in oil
obtained from oven dried at 55 °C.
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Journal of Medicinal Plants, Volume 16,
No. 61, Winter 2017
Mirahmadi et al.
Table 1- Analysis of variance for essential oil content of Melissa officinalis
Source of variation DF MS
Treatment 3 0.034**
Error 8 0.002
Total 11 -
CV=22; **significant at P ≤ 0.01
Table 2- Mean comparisons of different drying treatments and fresh sample on essential oil content of Melissa
officinalis
Treatment Means of oil content (% v/w)
shade-drying 0.3
a
oven-drying at 35 °C 0.3 a
oven-drying at 55 °C 0.08
b
Fresh sample 0.23 a
Similar letters in means column are not statistically different at 1%
level of probability using Duncan's multiple range tests.
Table 3- Chemical composition of essential oils (%) in Melissa officinalis using different drying treatments
No. Compound RI
Shade-drying
(%)
Oven-drying at 35
°C (%)
Oven-drying at 55
°C (%)
Fresh sample
(%)
1 Neral 1245 8.1 5.3 tr 5.4
2 Geranial 1271 13.0 18.9 tr 12.7
3 Neryl acetate 1365 1.6 1.9 tr 1.6
4 β-Caryophyllene 1421 14.9 17.9 12.6 21.8
5 α-Caryophyllene 1455 1.5 1.8 2.2 2.5
6 Germacrene D 1489 11.0 10.5 11.6 15.5
7 γ -Cadinene 1514 8.0 11.3 20.6 12.0
8 Caryophyllene oxide 1583 2.5 6.5 11.5 5.2
Oxygenated monoterpenes 21.1 24.2 0.0 18.1
Sesquiterpen hydrocarbon 37.0 43.4 47 53.4
Oxygenated Sesquiterpenes 2.5 6.5 11.5 5.2
RI, retention indices in elution order from HP-5MS column. tr, less than 0.05%
Discussion
In almost all plant species of the
Lamiaceae, the essential oil is synthesized in
specialized secretory organs such as glandular
trichomes [29]. Considerable levels of
essential oil yield (>2%) were recorded for
genera Mentha, Lavandula, Origanum, Salvia
and Satureja as essential to the oil-rich plant
[30, 31]. The total oil content in
M. officinalis is relatively low [27]. The
essential oil content of M. officinalis in our
study (range, 0.08 –0.3 % v/w; overall average
of 0.22% v/w) was relatively higher than when
compared with literature reports in India [32]
Turkey [33] and Germany [34]. Also, in
another study that was conducted among the
three methods of drying (shadow drying, sun
drying and fresh sample) the highest oil
content (0.084% and 0.122% w/w, from two
locations) was obtained from shadow drying,
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The Influence of …
and the lowest (0.099% and 0.06% w/w, from
two locations) was observed from fresh plant
[35]. The Essential oil yield obtained from
fresh material for this species in present study
is comparable to results reported by Abdellatif
et al (2015) [14] and Adinee et al. (2009) [36]
for the oil yield extracted from the fresh lemon
balm, but still lower than those recorded by
Blank et al. (2005) [37] and Pino et al. (1999)
[38] for fresh and air-dried sample,
respectively. Although, the samples dried in
oven at 35 °C and shade had considerable oil
yield, there were no significant differences
between essential oil content of these
treatments with oil content of fresh sample.
Sefidkon et al. (2006) revealed that the oven
drying at 45 °C and shade drying had no
significant effect on oil yield of Satureja
hortensis [23]. These findings contradict those
obtained by Sellami et al. (2011) in Laurus
nobilis [39] Ghasemi Pirbalouti et al. (2013) in
Satureja bachtiarica [20], and Rahimmalek
and Goli (2013) in Thymys daenensis [22]
which reported the essential oil yields of fresh
sample and those dried by oven temperature
(at 35- 45 °C) and shade were not similar and
they had significant differences.
Increase in the essential oil content of dried
plant materials compared with the oil yielded
from the fresh sample can be attributed to the
structural changes of specific cells containing
the essential oil with special structures and
lignified cell walls. Such cells are located in
the leaves of parenchymatous tissue [40].
Opposite results may be due to the differences
of plant species, the secretory structures and
their position in plant body, and the chemical
composition of essential oil [41]. In this study,
the lowest amount of essential oil yield was
resulted through oven drying at 55 °C which
indicates that increasing the drying
temperature would significantly decrease the
essential oil content. At high temperatures, the
biological structure of the oil glands of
aromatic plants can be affected significantly,
and the epithelial cells in the dried samples of
some sensible plants can collapse resulting in
more volatile oil diffusion during the drying,
which could explain the loss of essential oil in
high temperatures [42, 43]. In the same way,
Khangholil and Rezaeinodehi (2008) [41],
Braga et al. (2005) [44], and Argyropoulos
and Müllera (2011) [2] held that increasing the
drying temperature would result in a
significant decrease in the essential oil content.
Concerning the essential oil extracted from
the fresh sample, β-Caryophyllene and
Germacrene D were the two major
components. According to previous reports
regarding this species, this combination of
major compounds was rare for lemon balm in
other regions. To the best of our knowledge,
there is only one published case report [35]
which cited β-Caryophyllene and Germacrene
D as the main constituents of essential oil of
fresh lemon balm. Our results confirm
previous reports which cite different
chemotypes for the species M. officinalis. The
chemical composition of medicinal and
aromatic plants can be altered in response to
different environmental conditions and
developmental stages or in different plant parts
[45]. Many researchers have reported that the
main components of fresh lemon balm are
neral and geranial [13, 14, 33, 37, 46, 47]. In
other reports, the major compounds of fresh
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Journal of Medicinal Plants, Volume 16,
No. 61, Winter 2017
Mirahmadi et al.
sample of M.officinalis essential oils were
geraniol and citronellol in Iran [36], Nerol and
Citral in Morocco [8], and Citronellal and
Citronellol in China [26].
Drying of lemon balm caused not only the
quality deterioration of the essential oil, but
also resulted in some quantitative variations of
relative proportion of the components. The
chemical compounds fluctuation in different
drying treatments (such as shade or oven
drying) was reported previously in lemon balm
[26, 27], Satureja bachtiarica [20], Laurus
nobilis [39] and sage [42].
In the present study, some of the
components were missing in the essential oil
of oven dried sample at 55°C. The components
of the essential oils that are lost in the dried
samples are those stored on or near the leaf
and stem surfaces [48]. Also Sellami et al.
(2011) reported that relatively long period of
drying may cause oxidation process and
chemical rearrangements which lead to the
disappearance of some oil constituents [39].
Besides, in accordance with our results,
Rahimmalek and Goli (2013) reported that the
oven drying resulted in the loss of some
components in thyme compared with the shade
dried and the fresh sample. On the other hand,
some compounds seem to have more affinity
to the water fraction contained in thyme leaves
and thereby, they were lost with water during
the drying process.
In general, although the behavior of the
various compounds during different drying
methods is not fully understood, it can be
hypothesized that compounds with lower
boiling points (retention time) will show a
lower share in the total oil at higher drying
temperatures [27].
Sesquiterpene hydrocarbons constituted the
principal fraction of all samples, followed by
oxygenated monoterpenes, except oil of oven
dried sample at 55°C in which the oxygenated
sesquiterpene represented the second main
fraction. Lemon balm belongs to the
Lamiaceae family of plants, which are known
to store their essential oils on or near the leaf
surfaces [48]. Besides, increasing the
temperature to 55◦C, would increase the
chance to collapse the cuticle layer and
damage the extensive cell of the epidermis
which Causes monoterpenes (the low
molecular components) to leave the plant
organ more rapidly compared to
sesquiterpenes [27].
Overall, the development of the essential oil
sector has a direct relation to the improvement
of postharvest process especially the drying
technology in industrial plants. Results
obtained from experimental data could be
recommended to their uses in functional food
and pharmaceutical applications. In this study,
oven drying at 35°C was faster than other
treatments and resulted in appreciable essential
oil yield, and helped to conserve the
characteristic aroma of the spice, so would
seem to be more advisable for drying the
lemon balm.
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