Industrial Crops and Products 61 (2014) 331–337

Contents lists available at ScienceDirect

Industrial Crops and Products

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

Chemical constituents and biological activities of the Purple Perilla

essential oil against Lasioderma serricorne
Chun-xue You a , Ying Wang a , Wen-juan Zhang a , Kai Yang a , Yan Wu a,b , Zhu-feng Geng c ,
Hai-ping Chen d , Hai-yan Jiang d , Shu-shan Du a,∗ , Zhi-wei Deng c , Zhi-long Liu e
a
State Key Laboratory of Earth Surface Processes and Resource Ecology, Beijing Normal University, Haidian District, Beijing 100875, China
b
Technical Center of China Tobacco Guangxi Industrial Co. Ltd., Nanning 530001, Guangxi, China
c
Analytical and Testing Center, Beijing Normal University, Haidian District, Beijing 100875, China
d
College of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian 116600, Liaoning, China
e
Department of Entomology, China Agricultural University, Haidian District, Beijing 100193, China

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

Article history: Purple Perilla (Perilla frutescens (L.) Britt.) is an annual herb commonly used as a traditional Chinese
Received 26 February 2014 medicine for more than 1000 years. This work investigated the chemical composition, insecticidal and
Received in revised form 4 July 2014 repellent activities of the essential oil of Purple Perilla aerial parts against Lasioderma serricorne (Fabricius).
Accepted 9 July 2014
Moreover, we also isolated active constituents from the essential oil. The experiment was investigated in
2013 in Beijing. The essential oil of Purple Perilla was obtained by hydrodistillation and a total of 47 com-
Keywords:
ponents in the essential oil were identified with GC–MS. It was found that the main compounds included
Lasioderma serricorne
carvone (32.55%), perilla aldehyde (20.52%), caryophyllene (9.85%), 2-furyl methyl ketone (7.53%), 2,6-
Purple Perilla
Contact toxicity
dimethyl-6-(4-methyl-3-pentenyl)-2-norpinene (5.17%), and ␤-terpinyl acetate (3.41%). With a further
Fumigant toxicity isolation, four active constituents were obtained from the essential oil and identified as R-(+)-carvone,
Repellency perilla aldehyde, 2-furyl methyl ketone, and ␤-caryophyllene. In the progress of assay, it showed that the
Essential oil composition essential oil, R-(+)-carvone, perilla aldehyde, and 2-furyl methyl ketone exhibited stronger insecticidal
and repellent activities against L. serricorne than ␤-caryophyllene. The results indicate that the essential
oil of Purple Perilla aerial parts and isolated compounds have potential for development into natural
insecticides or repellents for control of insects in stored products.
© 2014 Elsevier B.V. All rights reserved.

1. Introduction hazards to warm-blooded animals, risk of environmental pollution,

The cigarette beetle, Lasioderma serricorne (Fabricius)
(Coleoptera: Anobiidae) is one of the most serious pest species
of stored botanicals worldwide. The cigarette beetle occurs
frequently in tropical and subtropical areas. They cause signif-
icant damage during storage of perishable food products such
as cereals, legumes, tobacco, and traditional Chinese medicinal
materials in warehouses (Abdelghany et al., 2010; Papadopoulou
and Athanassiou, 2004). Currently, recommended pest control
measures in durable stored products rely heavily on the use of
synthetic insecticides or fumigants which pose possible health
∗ Corresponding author. Tel.: +86 10 62208022; fax: +86 10 62208022.
E-mail addresses: youchunxue@mail.bnu.edu.cn (C.-x. You),
yingw150@163.com (Y. Wang), zwj0729@mail.bnu.edu.cn (W.-j. Zhang),
yangk 1988@mail.bnu.edu.cn (K. Yang), abelabel@126.com (Y. Wu),
gengzhufeng@bnu.edu.cn (Z.-f. Geng), apple90124@126.com (H.-p. Chen),
jhy880527@sina.cn (H.-y. Jiang), dushushan@bnu.edu.cn (S.-s. Du),
dengzw@bnu.edu.cn (Z.-w. Deng), zhilongliu@cau.edu.cn (Z.-l. Liu).
development of resistance by insects and pest resurgence (Zettler
and Arthur, 2000). The use of essential oils or their constituents
with low mammalian toxicity can effectively prevent insect pest
especially in storage (Isman, 2006). Investigations in several
countries confirm that some plant essential oils not only repel
insects, but also possess contact and fumigant toxicity against
stored product pests as well as exhibiting feeding inhibition or
harmful effects on the reproductive system of insects (Rajendran
and Srianjini, 2008). Essential oils and their constituents of many
plants such as medicinal herbs, spices, and fruits have been eval-
uated successfully for insecticidal or repellent activities against
stored product insects. Moreover, the essential oils and their
constituents have been proven more effective than traditionally
used pesticides (Ahmadi et al., 2013; Chu et al., 2013; Liang et al.,
2013; Liu et al., 2012a; Zhao et al., 2012).
Besides insecticidal and repellent activities, essential oils from
different plant sources have exhibited several biological activi-
ties, such as antibacterial and antifungal (Sakhanokho et al., 2013;
Wang et al., 2010), nematicidal (Bai et al., 2011; Li et al., 2011,

http://dx.doi.org/10.1016/j.indcrop.2014.07.021
0926-6690/© 2014 Elsevier B.V. All rights reserved.
332 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337
2013), larvicidal against Aedes albopictus (Chen et al., 2013; Liu
et al., 2012b, 2013a), and acaricidal (Araujo et al., 2012). As a con-
sequence, this vast arsenal of bioactive compounds has attracted
significant and crescent attention of researchers in recent years
(Isman, 2000). During our screening program for new agrochemi-
cals from Chinese medicinal herbs and wild plants, the essential oil
of Purple Perilla (Perilla frutescens (L.) Britt.) aerial parts was found
to possess insecticidal and repellent activities against L. serricorne.
P. frutescens is an annual herb of Perilla genus in the Lamiaceae
family. It has been commonly used as a kind of traditional Chi-
nese medicine in China for more than 1000 years. It is an edible
plant, with its fresh leaves usually used as vegetables, and com-
monly used for seasoning pickles or as garnish for raw fish dishes
in Japan. It is a popular leafy vegetable in Korea, which is generally
consumed as a pickle or wrapping with roasted meats. The seeds
are ground and added to soup for seasoning in Korea (Huang et al.,
2011). There are both purple-leafed and green-leafed varieties of
P. frutescens, which are generally recognized as separate species by
botanists, and their Chinese names are Zisu and Baisu, respectively.
There are significant differences in the morphology and chemistry
between Zisu and Baisu. Both Zisu and Baisu were recorded in the
“Dictionary of Chinese Crude Drug” with different clinical usage.
However, in the Pharmacopoeia of People’s Republic of China, 2010,
Baisu was not recorded separately, just being used as Zisu. (JiangSu
New Medicinal College, 1977; Huang et al., 2011; Pharmacopoeia
Committee of Ministry of Health of People’s Republic of China,
2010). Zisu has significant anti-allergic, anti-inflammatory, and
strong antitumor-promoting activities (Banno et al., 2004). To date,
there are only two reports on the repellent activity of the essential
oil of Zisu (Purple Perilla) aerial parts against L. serricorne. However,
we use a different experimental method to investigate the repel-
lent activity of the essential oil against L. serricorne (Hori, 2003,
2004). And the literature survey has shown that there is no report
on insecticidal activity of Zisu (Purple Perilla) essential oil against L.
serricorne, thus we decided to investigate the insecticidal and repel-
lent activities of Zisu essential oil against L. serricorne and isolate
active compounds from the essential oil for the first time. Our Zisu
plant sample was identified by Dr. Liu Q.R. in 2013 (College of Life
Sciences, Beijing Normal University, Beijing, China) and a voucher
specimen (BNU-CMH-Dushushan-2013-07-09-008) was deposited
at the Herbarium (BNU) of College of Life Sciences, Beijing Normal
University.
2. Materials and methods
2.1. Plant material and essential oil extraction
Dried aerial parts (whole grass, 9.0 kg) of Purple Perilla were har-
vested in July 2013 from Beijing City (39.9◦ N latitude and 116.3◦ E
longitude), China. The aerial parts were air-dried for one week and
ground to a powder. The powder was subjected to hydrodistilla-
tion using a modified Clevenger-type apparatus for 6 h. Anhydrous
sodium sulphate was used to remove water after extraction. The
essential oil was stored in airtight containers in a refrigerator at
4 ◦ C for subsequent experiments.
2.2. Insects
The cigarette beetle, L. serricorne were obtained from laboratory
cultures maintained for the last 2 years in the dark in incubators
at 29 ± 1 ◦ C and 70–80% relative humidity. The insects were reared
in glass containers (0.5 L) containing wheat flour at 12–13% mois-
ture content mixed with yeast (10:1, w/w). Adults used in all the
experiments were about 7 ± 2 days old.
2.3. Gas chromatography and mass spectrometry
Components of the essential oil of Purple Perilla aerial parts
were identified by gas chromatography–flame ionization detection
(GC–FID) and gas chromatography–mass spectrometry (GC–MS)
using an Agilent 6890N gas chromatograph hooked to an Agilent
5973N mass selective detector. The same column and analysis con-
ditions were used for both GC–FID and GC–MS. They were equipped
with a HP-5MS (30 m × 0.25 mm × 0.25 ␮m) capillary column. The
column temperature was programmed at 50 ◦ C for 2 min, then
increased at 2 ◦ C/min to the temperature of 150 ◦ C and held for
2 min, and then increased at 10 ◦ C/min until the final temperature
of 250 ◦ C was reached, where it was held for 5 min. The injector
temperature was maintained at 250 ◦ C and the volume injected
was 0.1 mL of 1% solution (diluted in n-hexane). The carrier gas was
helium at flow rate of 1.0 mL/min. Spectra were scanned from 50 to
550 m/z. Most constituents were identified by comparison of their
retention indices with those reported in the literature. The reten-
tion indices were determined in relation to a homologous series of
n-alkanes (C5 –C36 ) under the same operating conditions. Further
identification was made by comparison of their mass spectra with
those stored in NIST 05 and Wiley 275 libraries or with mass spectra
from the literature (Adams, 2001). Relative percentages of the indi-
vidual components of the essential oil were obtained by averaging
the GC–FID peak area% reports.
2.4. Purification and characterization of four compounds from
Purple Perilla essential oil
The crude essential oil (5 mL) was chromatographed on a silica
gel (Qingdao Marine Chemical Plant, Shandong Province, China)
column (35 mm i.d., 500 mm length) by gradient elution with n-
hexane first, then with n-hexane-ethyl acetate, and last with ethyl
acetate. Fractions (150 mL) were collected and concentrated at
35 ◦ C, and similar fractions according to thin layer chromatography
(TLC) profiles were combined to yield 25 fractions. Fractions (3–5,
9–11) that possessed contact toxicity, with similar TLC profiles,
were pooled and further purified by preparative silica gel column
chromatography (PTLC) until obtain the pure compounds for deter-
mining structure as R-(+)-carvone (1, 0.66 g), perilla aldehyde (2,
0.48 g), 2-furyl methyl ketone (3, 0.16 g), and ␤-caryophyllene (4,
0.22 g). The isolated compounds were elucidated based on nuclear
magnetic resonance. 1 H and 13 C NMR spectra were recorded on
Bruker Avance DRX 500 instruments using CDCl3 as solvent with
TMS as internal standard. The specific rotation ([˛]20 D ) was recorded
on Autopol III Rudolph Automatic polarimeter using CHCl3 as sol-
vent.
2.5. Isolated compounds from Purple Perilla essential oil
R-(+)-Carvone (1, Fig. 1). Slightly yellow oil, C10 H14 O. +50.62◦ . 1 H
NMR (CDCl3 , 500 MHz) ı (ppm): 6.78–6.77 (1H, m, H-6), 4.82 (1H,
s, H-9a), 4.77 (1H, s, H-9b), 2.73–2.27 (5H, m, H-3, 4, 5), 1.80 (3H,
s, CH3 ), 1.77 (3H, s, CH3 ); 13 C NMR (CDCl3 , 125 MHz) ı (ppm):
199.78 (C-2), 146.69 (C-8), 144.60 (C-6), 135.42 (C-1), 110.44 (C-9),
43.14 (C-3), 42.46 (C-4), 31.23 (C-5), 20.42 (C-10), 15.71 (C-7). The
1 H and 13 C NMR data were in agreement with the reported data
(Aggarwal et al., 2002; Lanfranchi et al., 2008).
Perilla aldehyde (2, Fig. 1). Slightly yellow oil, C10 H14 O. 1 H NMR
(CDCl3 , 500 MHz) ı (ppm): 9.46 (1H, s, H-7), 6.85 (1H, m, H-2), 4.80
(1H, s, H-9a), 4.76 (1H, s, H-9b), 2.52–2.45 (2H, m, H-6), 2.30–2.25
(2H, m, H-5), 2.17–2.11 (1H, m, H-4), 1.95–1.92 (1H, m, H-3a), 1.78
(3H, s, CH3 ), 1.51–1.43 (1H, m, H-3b); 13 C NMR (CDCl3 , 125 MHz)
ı (ppm): 193.93 (C-7), 150.65 (C-2), 148.36 (C-8), 141.27 (C-1),
109.53 (C-9), 40.70 (C-4), 31.74 (C-5), 26.35 (C-3), 21.58 (C-6), 20.71
 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337
Fig. 1. Compounds isolated from the essential oil of Purple Perilla aerial parts.
(C-10). The 1 H and 13 C NMR data were consistent with the literature
data (Zou et al., 2008).
2-Furyl methyl ketone (3, Fig. 1). Yellow oil, C6 H6 O2 . 1 H NMR
(CDCl3 , 500 MHz) ı (ppm): 7.59 (1H, d, J = 1.5 Hz, H-2), 7.19 (1H, d,
J = 3.5 Hz, H-4), 6.54 (dd, J = 3.5 Hz, 1.5 Hz, H-3), 2.46 (3H, s, CH3 );
13 C NMR (CDCl , 125 MHz) ı (ppm): 186.78 (C-6), 152.87(C-5),
3
146.41 (C-2), 117.21 (C-4), 112.24 (C-3), 26.01 ( CH3 ). The 1 H and
13 C NMR data were in accord with the reported data (Singh and
Singh, 2010).
ˇ-Caryophyllene (4, Fig. 1). Colorless oil, C15 H24 . 1 H NMR (CDCl3 ,
500 MHz) ı (ppm): 5.33 (1H, m, H-5), 4.97 (1H, s, H-12a), 4.85 (1H,
s, H-12b), 2.37 (1H, m, H-9), 2.33 (1H, m, H-7b), 2.23 (1H, m, H-7a),
2.11 (1H, m, H-1), 2.02 (1H, m, H-6b), 1.94 (1H, m, H-6a), 1.72 (1H,
m, H-2b), 1.69 (1H, m, H-3b), 1.65–1.67 (2H, m, H-10), 1.64 (3H, s,
15-CH3 ), 1.60 (1H, m, H-3a), 1.52 (1H, m, H-2a), 1.02 (3H, s, 12-CH3 ),
1.00 (3H, s, 13-CH3 ); 13 C-NMR (CDCl3 , 125 MHz) ı (ppm): 154.74
(C-8), 135.58 (C-4), 124.31 (C-5), 111.67 (C-12), 53.54 (C-1), 48.48
(C-9), 40.33 (C-10), 39.96 (C-3), 34.79 (C-7), 33.02 (C-11), 30.09 (C-
13), 29.36 (C-2), 28.38 (C-6), 22.66 (C-14), 16.32 (C-15). The 1 H and
13 C NMR data were in agreement with the reported data (Ragasa
et al., 2013).
2.6. Contact toxicity
The contact toxicity of the essential oil against L. serricorne
adults was measured as described by Liu and Ho (1999). Range-
finding studies were run to determine the appropriate testing
concentrations. A serial dilution of the essential oil/compounds
(five concentrations) was prepared in n-hexane. Aliquots of 0.5 ␮L
of the dilutions were applied topically to the dorsal thorax of
the insects. Controls were determined using n-hexane. Ten insects
were used for each concentration and control, and the experiment
was replicated five times. Both treated and control insects were
then transferred to glass vials with culture media and kept in incu-
bators. Mortality was recorded after 24 h. The observed mortality
data were corrected for control mortality using Abbott’s formula.
The LD50 values were calculated by using Probit analysis (IBM SPSS
V20.0) (Liu et al., 2012a; Sakuma, 1998; Yang et al., 2014). The posi-
tive control, pyrethrins (pyrethrin I, 24%; pyrethrin II, 13%; cinnerin
I, 2%; cinnerin II, 2%; jasmolin I, 1%; jasmolin II, 1%) was purchased
from Dr. Ehrenstorfer, Germany.
2.7. Fumigant toxicity
The fumigant activity of the essential oil against L. serricorne
adults was tested as described by Liu and Ho (1999). Range-finding
studies were run to determine the appropriate testing concen-
trations. A serial dilution of the essential oil/compounds (five
concentrations) was prepared in n-hexane. A Whatman filter paper
333
(diameter 2.0 cm) was impregnated with 10 ␮L dilution and then
placed on the underside of the screw cap of a glass vial (diameter
2.5 cm, height 5.5 cm, volume 25 mL). The solvent was allowed to
evaporate for 20 s before the cap was placed tightly on the glass vial,
each of which contained 10 insects inside to form a sealed cham-
ber. Fluon (ICI America Inc.) was used inside the glass vial to prevent
insects from contacting the treated filter paper. Preliminary exper-
iments demonstrated that 20 s was sufficient for the evaporation of
solvents. n-Hexane was used as a control. Five replicates were car-
ried out for all treatments and controls, and they were incubated
under the same conditions as rearing. Mortality was determined
after 24 h of treatment. The observed mortality data were corrected
for control mortality using Abbott’s formula. The LC50 values were
calculated using Probit analysis (IBM SPSS V20.0) (Liu et al., 2012a;
Sakuma, 1998; Yang et al., 2014).
2.8. Repellency tests
The repellent activity to L. serricorne adults was tested using the
area preference method (Zhang et al., 2011). Petri dishes (9 cm in
diameter) were used to confine cigarette beetles during the exper-
iment. The crude essential oil and the isolated compounds were
diluted in n-hexane to five concentrations (39.32, 7.86, 1.57, 0.31,
and 0.06 nL/cm2 ), and n-hexane was used as the control. Filter
paper (9 cm in diameter) was cut in half and 500 ␮L of each concen-
tration was applied separately to half of the filter paper as uniformly
as possible with a micropipette. The other half (control) was treated
with 500 ␮L of n-hexane. Both the treated half and the control half
were then air-dried to evaporate the solvent completely (30 s). A
full disk was carefully remade by attaching the tested half to the
negative control half with tape. Care was taken so that the attach-
ment did not prevent free movement of insects from the one half to
the other, but the distance between the filter paper halves remained
sufficient to prevent seepage of test samples from one half to the
other. Each remade filter paper after treatment with solid glue was
placed in a Petri dish with the seam oriented in one of four ran-
domly selected different directions to avoid any insecticidal stimuli
affecting the distribution of insects. Twenty insects were released
in the center of each filter paper disk, and a cover was placed over
the Petri dish. Five replicates were used and the experiment was
repeated three times. Counts of the insects present on each strip
were made after 2 and 4 h. The percent repellency (PR) of each
volatile oil/compound was then calculated using the formula:
N − N 
c t
PR (%) = × 100
Nc + Nt
where Nc is the number of insects present in the negative con-
trol half while Nt is the number of insects present in the treated
half. Analysis of variance (One-Way ANOVA and GLM Univariate)
334 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337
Table 1
The scale to be assign repellency of the essential oil of Purple Perilla aerial parts and its constituents.
Class Percent repellency Class
0 >0.01 to <0.1 II
I 0.1–20 III
and Tukey’s test were conducted by using SPSS 20.0 for Windows
2007. Percentage was subjected to an arcsine square-root transfor-
mation before variance and Tukey’s tests. The averages were then
assigned to different classes (0 to V) in Table 1 (Liu and Ho, 1999).
A commercial repellent, DEET (N,N-diethyl-3-methylbenzamide),
was purchased from Dr. Ehrenstorfer, Germany and used as a pos-
itive control.
3. Results and discussion
3.1. Chemical composition of the Purple Perilla essential oil
The Purple Perilla essential oil was yellow with a yield of 0.079%
(v/w) and density of 0.89 g/mL. A total of 47 components of the
essential oil of Purple Perilla were identified, accounting for 93.62%
of the total oil (Table 2). The main compounds in the essential
oil were carvone (32.55%), perilla aldehyde (20.52%), caryophyl-
lene (9.85%), and 2-furyl methyl ketone (7.53%), followed by
2,6-dimethyl-6-(4-methyl-3-pentenyl)-2-norpinene (5.17%) and
␤-terpinyl acetate (3.41%).
The chemical composition of the essential oil of Purple Perilla
aerial parts in the present study was not same as that reported
in previous studies. For example, perilla aldehyde, caryophyllene,
limonene, and (Z, E)-a-farnesene were the main volatile compo-
nents of Zisu harvested in June from Yangpu District, Shanghai
Province. Moreover, the content and composition of the volatile
oil of Zisu were various with different extraction methods (Huang
et al., 2011). However, ␤-caryophyllene, 2-hexanoylfuran, ␤-
farnesene, 1,4,7-cycloundecatriene-1,5,9,9-tetramethyl-zzz, and
1-cyclohexane-1-carboxaldehyde were common constituents in
Zisu collected from herb markets in mainland China and Hong Kong,
and content and composition of the volatile oil of Zisu were vari-
ous in different three medicinal parts of P. frutescens (Liu et al.,
2013b). These differences of chemical content and composition of
the essential oils might have been due to harvest time and local,
climatic and seasonal factors, storage duration of medicinal herbs
as well as extraction method, and these differences may result in
different biological activities (Huang et al., 2011; Liu et al., 2013b).
However, it is only our conjectures that need a further study.
3.2. Insecticidal activities
The essential oil of Purple Perilla aerial parts exhibited
contact toxicity against L. serricorne adults with LD50 value
of 3.13 ␮g/adult. When compared with the positive control,
pyrethrins (LD50 = 0.24 ␮g/adult), the essential oil demonstrated
13 times less toxic against L. serricorne adults (Table 3). However,
compared with the other essential oils in the literature, the essen-
tial of Purple Perilla aerial parts possessed stronger contact toxicity
against L. serricorne adults, e.g., the essential oil of Litsea cubeba
(LD50 = 27.33 ␮g/adult) (Yang et al., 2014). Moreover, perilla alde-
hyde possessed almost 2, 3, and 9 times more toxicity than 2-furyl
methyl Ketone, R-(+)-carvone, and ␤-caryophyllene, respectively.
It is suggested that perilla aldehyde is a major contributor to the
contact activity of the essential oil against L. serricorne adults. How-
ever, compared with pyrethrins (positive control), perilla aldehyde
showed 16 times less toxicity.
2-Furyl methyl ketone (LC50 = 0.86 mg/L air) exhibited stronger
fumigant toxicity against L. serricorne adults than R-(+)-carvone
Percent repellency Class Percent repellency
20.1–40 IV 60.1–80
40.1–60 V 80.1–100
(LC50 = 1.83 mg/L air), perilla aldehyde (LC50 = 3.03 mg/L air), and
the crude essential oil (LC50 = 4.16 mg/L air). The results are
presented in Table 3. However, ␤-caryophyllene did not show
fumigant toxicity at the tested concentrations. 2-Furyl methyl
ketone showed almost 2, 4, and 5 times stronger fumigant toxic-
ity than R-(+)-carvone, perilla aldehyde, and the crude essential
oil against L. serricorne adults. However, compared with the
other essential oils in the literature, the essential of Purple Per-
illa aerial parts possessed stronger fumigant toxicity against L.
serricorne adults, e.g., the essential oil of Agastache foeniculum
(LC50 = 21.57 ␮L/L air) (Ebadollahi et al., 2010), the essential oil
of L. cubeba (LC50 = 22.97 mg/L air) (Yang et al., 2014). Moreover,
compared with other previous studies on essential oils, we iso-
lated the compounds and investigated their insecticidal activities.
It is beneficial to develop active compounds into natural insecti-
cides (Chaubey, 2013; Ebadollahi et al., 2010). The currently used
fumigants are synthetic insecticides and the most effective fumi-
gants (e.g., phosphine and methyl bromide) are also highly toxic to
humans and other non-target organisms. The essential oil of Pur-
ple Perilla aerial parts and its isolated compounds show potential
to be developed as possible natural fumigants or insecticides for
the control of L. serricorne. However, for the practical application
of the essential oil and the isolated constituents as novel insecti-
cides or fumigants, further studies on the safety of the essential oil
to humans and on development of formulations are necessary to
improve the efficacy and stability and to reduce costs.
3.3. Repellent activity
Purple Perilla essential oil and isolated constituents exhibited
comparable repellent activity against L. serricorne. The results are
presented in Fig. 2. Data showed that at tested concentration of
39.32 nL/cm2 , the crude essential oil, R-(+)-carvone, and perilla
aldehyde showed strong (class V) repellency (80%, 82%, and 82%,
respectively) and moderate (Class IV) repellency (72%, 78%, and
74%, respectively) against L. serricorne at 2 h and 4 h after expo-
sure, whereas 2-furyl methyl ketone exhibited moderate (Class IV)
repellency (70% and 76%, respectively) against L. serricorne at 2 h
and 4 h after exposure.
Compared with the positive control, DEET, perilla aldehyde
exhibited stronger repellency against L. serricorne at 2 h after expo-
sure. It is due to the reason that at the concentrations of 39.32,
7.86, 1.57, and 0.31 nL/cm2 , perilla aldehyde and DEET (P = 0.345,
0.664, 0.386, and 0.091) exhibited the same level of repellency
against L. serricorne at 2 h after exposure. Whereas at the concentra-
tion of 0.06 nL/cm2 , perilla aldehyde exhibited stronger repellency
than DEET (P = 0.031) against L. serricorne at 2 h after exposure.
In addition, at the concentrations of 39.32, 7.86, 1.57, 0.31, and
0.06 nL/cm2 , the crude essential oil (P = 0.434, 0.932, 0.066, 0.211,
and 0.236), R-(+)-carvone (P = 0.345, 0.408, 0.573, 0.091, and 0.236),
and DEET exhibited the same level of repellency against L. serricorne
at 2 h after exposure. Moreover, at the concentrations of 1.57, 0.31,
and 0.06 nL/cm2 , 2-furyl methyl ketone exhibited stronger repel-
lency than DEET (P = 0.016, 0.003, and 0.003) against L. serricorne at
2 h after exposure. However, at the concentrations of 39.32 and
7.86 nL/cm2 , ␤-caryophyllene exhibited weaker repellency than
DEET (P = 0.000 and 0.000) against L. serricorne at 2 h after exposure.
Many essential oils and their constituents have been evaluated for
 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337 335

Table 2
Chemical composition of the essential oil of Purple Perilla aerial parts.

Compound RIa Relative content (%)

3,6,6-Trimethyl-bicyclo[3.1.1]hept-2-ene 1087 0.34

␣-Pinene 1190 0.12
6-Ethyl-2-methyl-octane 1205 0.11
2,2-Dimethyl-heptane 1208 0.12
2,2,4,6,6-Pentamethyl-heptane 1240 0.26
o-Cymene 1251 0.67
␤-Terpinyl acetate 1278 3.41
2,3,6,7-Tetramethyl-octane 1301 0.11
4,6-Dimethyl-undecane 1309 0.76
2,4,6-Trimethyl-decane 1315 0.38
2,3,6,7-Tetramethyl-4-octene 1333 0.06
5-Methyl-dodecane 1349 0.06
5,7-Dimethyl-undecane 1358 0.27
2-Furyl methyl ketone 1373 7.53
␣-Linalool 1425 0.50
6-Ethyl-2-methyl-decane 1470 0.09
p-Menth-1-en-8-ol 1631 0.15
Decahydro-naphthalen-1-ol 1721 0.05
Carvone 1757 32.55
Germacrene D 1816 1.33
Perilla aldehyde 1821 20.52
8-Isopropenyl-1,5-dimethyl-1,5-cyclodecadiene 1841 0.20
trans-Shisool 1855 0.10
Caryophyllene 1872 9.85
p-Mentha-1,8-dien-7-ol 1883 1.64
trans-3,6-Diethyl-3,6-dimethyl-tricyclo[3.1.0.0(2,4)]hexane 1911 2.15
␣-Caryophyllene 1916 1.17
1-Methylene-2b-hydroxymethyl-3,3-dimethyl-4b-(3-methylbut-2-enyl)-cyclohexane 1931 0.06
3,5-Dimethyl-octane 1938 0.08
p-Mentha-2,4(8)-diene 1956 0.29
4-Isopropylidene-1-vinyl-o-menth-8-ene 1973 0.44
2,6-Dimethyl-6-(4-methyl-3-pentenyl)-2-norpinene 1981 5.17
Cadina-3,9-diene 2010 0.08
Eugenol 2016 0.14
6-(3-Methyl-3-cyclohexenyl)-2-methyl-2,6-heptadienol 2029 0.06
2-[(E)-(3 -Methylbutadien-2 -yl) methylidene]-1,3,3-trimethylcyclohexanol 2076 0.06
1-Methylene-2b-hydroxymethyl-3,3-dimethyl-4b-(3-methylbut-2-enyl)-cyclohexane 2082 0.07
3-Ethyl-3-methyl-decane 2101 0.22
4,7-Dimethyl-4,4a,5,6-tetrahydrocyclopenta[c]pyran-1,3-dione 2143 0.07
3,7,11-Trimethyl-3-hydroxy-6,10-dodecadien-1-yl acetate 2236 0.28
Pentadecane 2299 0.07
(1-Butylheptyl)-benzene 2329 0.13
(1-Ethylnonyl)-benzene 2368 0.06
Hexadecane 2373 0.09
(1-Propylnonyl)-benzene 2567 0.07
3,7,11,15-Tetramethyl-2-hexadecen-1-ol 2824 0.72
Phytol 2921 0.96
Total 93.62
a
RI, retention index as determined on a HP-5MS column using the homologous series of n-hydrocarbons as reference.

Table 3
Contact toxicity and fumigant toxicity of the essential oil of Purple Perilla aerial parts and its constituents against Lasioderma serricorne adults.

Activity Treatment LD50 95% FLa Slope ± SE Chi square (2 )

LC50

Contact toxicity (␮g/adult) Purple Perilla 3.13 2.65–3.62 1.99 ± 0.30 7.93
R-(+)-Carvone 11.41 10.05–12.82 2.69 ± 0.34 14.40
Perilla aldehyde 3.82 3.36–4.31 2.52 ± 0.32 12.28
2-Furyl methyl Ketone 6.67 6.27–7.10 3.33 ± 0.38 19.77
␤-Caryophyllene 35.52 31.89–39.54 3.07 ± 0.37 15.41
Pyrethrins 0.24 0.16–0.35 1.31 ± 0.20 17.36

Fumigant toxicity (mg/L air) Purple Perilla 4.16 3.61–4.91 2.05 ± 0.29 10.90
R-(+)-Carvone 1.83 1.64–2.03 3.21 ± 0.39 18.00
Perilla aldehyde 3.03 2.63–3.46 2.21 ± 0.30 16.65
2-Furyl methyl Ketone 0.86 0.80–0.93 2.72 ± 0.31 17.79
␤-Caryophyllene – – – –
Phosphine 9.23 × 10−3 7.13 × 10−3 –11.37 × 10−3 2.12 ± 0.27 11.96
a
Fiducial limits.
336 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337
Fig. 2. Percentage repellency (PR) of the essential oil from Purple Perilla aerial parts and its constituents against L. serricorne at 2 h (A) and 4 h (B) after exposure. a means in
the same column followed by the different letters differ significantly (P < 0.05) in ANOVA and Tukey’s tests. PR was subjected to an arcsine square-root transformation before
ANOVA and Tukey’s tests.
repellency against insects (Nerio et al., 2010; Caballero-Gallardo
et al., 2011).
Comparing the above isolated compounds, their insecticidal and
repellent activities varied greatly. This might be attributed to the
number and position of the double bond in chemical structures. The
more double bonds the compounds had, the better insecticidal and
repellent activities they usually possessed. However, it is only our
conjectures that need a further study.
The above results indicated that the bioactivity properties of the
essential oil are related to the synergistic effects of its diverse major
and minor components. Hence, it is significant to isolate chemical
constituents in both high and low percentages for their appreciable
bioactivity. In addition, further investigations that focus on effi-
ciency and safety of the pure compounds should be conducted,
while structure modification is a considerable method.
4. Conclusions
The work indicates that the essential oil of Purple Perilla aerial
parts and the isolated compounds possess significant insecticidal
and repellent activities against L. serricorne and show potential for
development as natural insecticides or repellents for stored prod-
uct insects. The development of natural pesticides would help to
decrease the negative effect of synthetic insecticides and repellents,
such as pesticide residues, resistance, and environmental pollution.
Moreover, the natural resources of Purple Perilla are abundant, so
they have a very good perspective of application. However, further
studies are required to investigate the insecticidal and repellent
mechanism of Purple Perilla essential oil.
Acknowledgments
This project was supported by State Key Laboratory of Earth Sur-
face Processes and Resource Ecology and National Natural Science
Foundation of China (No. 81374069).
References
Abdelghany, A.Y., Awadalla, S.S., Abdel-Baky, N.F., El-Syrafi, H.A., Fields, P.G., 2010.
Stored-product insects in botanical warehouses. J. Stored Prod. Res. 46, 93–97.
Adams, R.P., 2001. Identification of Essential Oil Components by Gas Chromatogra-
phy/Quadrupole Mass Spectroscopy. Allured, Carol Stream, IL, USA.
Aggarwal, K.K., Khanuja, S.P.S., Ahmad, A., Santha Kumar, T.R., Gupta, V.K., Kumar,
S., 2002. Antimicrobial activity profiles of the two enantiomers of limonene and
carvone isolated from the oils of Mentha spicata and Anethum sowa. Flavour Frag.
J. 17, 59–63.
Ahmadi, M., Abd-alla, A.M., Moharramipour, S., 2013. Combination of gamma radi-
ation and essential oils from medicinal plants in managing Tribolium castaneum
contamination of stored products. Appl. Radiat. Isot. 78, 16–20.
 C.-x. You et al. / Industrial Crops and Products 61 (2014) 331–337
Araujo, M.J., Camara, C.A., Born, F.S., Moraes, M.M., Badji, C.A., 2012. Acaricidal activ-
ity and repellency of essential oil from Piper aduncum and its components against
Tetranychus urticae. Exp. Appl. Acarol. 57, 139–155.
Bai, C.Q., Liu, Z.L., Liu, Q.Z., 2011. Nematicidal constituents from the essential oil of
Chenopodium ambrosioides aerial parts. E-J. Chem. 8, 143–148.
Banno, N., Akihisa, T., Tokuda, H., Yasukawa, K., Higashihara, H., Ukiya, M., Watanabe,
K., Kimura, Y., Hasegawa, J., Nishino, H., 2004. Triterpene acids from the leaves of
Perilla frutescens and their anti-inflammatory and antitumor-promoting effects.
Biosci. Biotechnol. Biochem. 68, 85–90.
Caballero-Gallardo, K., Olivero-Verbel, J., Stashenko, E.E., 2011. Repellent activity
of essential oils and some of their individual constituents against Tribolium
castaneum Herbst. J. Agric. Food Chem. 59, 1690–1696.
Chaubey, M.K., 2013. Biological activities of Zingiber officinale (Zingiberaceae) and
Piper cubeba (Piperaceae) essential oils against Pulse Beetle, Callosobruchus chi-
nensis (Coleoptera: Bruchidae). Pak. J. Biol. Sci. 16, 517–523.
Chen, X.B., Liu, X.C., Zhou, L., Liu, Z.L., 2013. Essential oil composition and larvicidal
activity of Clinopodium gracile (Benth) Matsum (Labiatae) aerial parts against
the Aedes albopictus mosquito. Trop. J. Pharm. Res. 12, 799–804.
Chu, S.S., Liu, S.L., Liu, Q.Z., Jiang, G.H., Liu, Z.L., 2013. Chemical composition and
insecticidal activities of the essential oil of the flowering aerial parts of Aster
ageratoides. J. Serb. Chem. Soc. 78, 209–216.
Ebadollahi, A., Safaralizadeh, M.H., Pourmirza, A.A., Gheibi, S.A., 2010. Toxicity of
essential oil of Agastache foeniculum (Pursh) Kuntze to Oryzaephilus surinamensis
L. and Lasioderma serricorne F. J. Plant Prot. Res. 50, 215–219.
Hori, M., 2003. Repellency of essential oils against the cigarette beetle, Lasioderma
serricorne (Fabricius) (Coleoptera: Anobiidae). Appl. Entomol. Zool. 38, 467–473.
Hori, M., 2004. Repellency of shiso oil components against the cigarette beetle, Lasio-
derma serricorne (Fabricius) (Coleoptera: Anobiidae). Appl. Entomol. Zool. 39,
357–362.
Huang, B.K., Lei, Y.L., Tang, Y.H., Zhang, J.C., Qin, L.P., Liu, J., 2011. Comparison of HS-
SPME with hydrodistillation and SFE for the analysis of the volatile compounds
of Zisu and Baisu, two varietal species of Perilla frutescens of Chinese origin. Food
Chem. 125, 268–275.
Isman, M.B., 2000. Plant essential oils for pest and disease management. Crop Prot.
19, 603–608.
Isman, M.B., 2006. Botanical insecticides, deterrents, and repellents in modern agri-
culture and an increasingly regulated world. Annu. Rev. Entomol. 51, 45–66.
JiangSu New Medicinal College, 1977. Dictionary of Chinese Herbal Medicine.
Shanghai Science & Technology Press, Shanghai.
Lanfranchi, D.A., Blanc, M.C., Vellutini, M., Bradesi, P., Casanova, J., Tomi, F., 2008.
Enantiomeric differentiation of oxygenated p-menthane derivatives by 13 C NMR
using Yb(hfc)3 . Magn. Reson. Chem. 46, 1188–1194.
Li, H.Q., Bai, C.Q., Chu, S.S., Zhou, L., Du, S.S., Liu, Z.L., Liu, Q.Z., 2011. Chemical
composition and toxicities of the essential oil derived from Kadsura heteroclita
stems against Sitophilus zeamais and Meloidogyne incognita. J. Med. Plants Res.
5, 4943–4948.
Li, H.Q., Liu, Q.Z., Liu, Z.L., Du, S.S., Deng, Z.W., 2013. Chemical composition and
nematicidal activity of essential oil of Agastache rugosa against Meloidogyne
incognita. Molecules 18, 4170–4180.
Liang, Y., Li, J.L., Xu, S., Zhao, N.N., Zhou, L., Cheng, J., Liu, Z.L., 2013. Evaluation
of repellency of some Chinese medicinal herbs essential oils against Liposcelis
bostrychophila (Psocoptera: Liposcelidae) and Tribolium castaneum (Coleoptera:
Tenebrionidae). J. Econ. Entomol. 106, 513–519.
Liu, Z.L., Ho, S.H., 1999. Bioactivity of the essential oil extracted from Evodia
rutaecarpa Hook f. et Thomas against the grain storage insects, Sitophilus
337
zeamais Motsch, and Tribolium castaneum (Herbst). J. Stored Prod. Res. 35,
317–328.
Liu, Z.L., Zhao, N.N., Liu, C.M., Zhou, L., Du, S.S., 2012a. Identification of insecticidal
constituents of the essential oil of Curcuma wenyujin rhizomes against Liposcelis
bostrychophila Badonnel. Molecules 17, 12049–12060.
Liu, Z.L., Liu, Q.Z., Du, S.S., Deng, Z.W., 2012b. Mosquito larvicidal activity of alka-
loids and limonoids derived from Evodia rutaecarpa unripe fruits against Aedes
albopictus (Diptera: Culicidae). Parasitol. Res. 111, 991–996.
Liu, X.C., Dong, H.W., Zhou, L., Du, S.S., Liu, Z.L., 2013a. Essential oil composition and
larvicidal activity of Toddalia asiatica roots against the mosquito Aedes albopictus
(Diptera: Culicidae). Parasitol. Res. 112, 1197–1203.
Liu, J., Wan, Y.K., Zhao, Z.Z., Chen, H.B., 2013b. Determination of the content of
rosmarinic acid by HPLC and analytical comparison of volatile constituents by
GC–MS in different parts of Perilla frutescens (L.) Britt. Chem. Cent. J. 7, 1–11.
Nerio, L.S., Olivero-Verbel, J., Stashenko, E., 2010. Repellent activity of the essential
oils: a review. Bioresour. Technol. 101, 372–378.
Papadopoulou, S.C., Athanassiou, C.G., 2004. Lariophagus distinguendus (F.) (Hyme,
Chalcidoidea, Pteromalidae), an ectoparasitoid of Lasioderma serricorne (F.) (Col.,
Anobiidae), found for the first time in tobacco stores in Greece. J. Pest Sci. 77,
183–184.
Pharmacopoeia Committee of Ministry of Health of People’s Republic of China, 2010.
Pharmacopoeia of People’s Republic of China, vol. 1. China Medical Science and
Technology Press, Beijing, China.
Ragasa, C.Y., Espineli, D.L., Agoo, E.M.G., Fierro, R.S., 2013. Chemical constituents of
Cinnamomum cebuense. Chin. J. Nat. Med. 11, 264–268.
Rajendran, S., Srianjini, V., 2008. Plant products as fumigants for stored-product
insects control. J. Stored Prod. Res. 44, 126–135.
Sakhanokho, H.F., Sampson, B.J., Tabanca, N., Wedge, D.E., Demirci, B., Baser, K.H.,
Bernier, U.R., Tsikolia, M., Agramonte, N.M., Becnel, J.J., Chen, J., Rajasekaran, K.,
Spiers, J.M., 2013. Chemical composition, antifungal and insecticidal activities
of Hedychium essential oils. Molecules 18, 4308–4327.
Sakuma, M., 1998. Probit analysis of preference data. Appl. Entomol. Zool. 33,
339–347.
Singh, V., Singh, V., 2010. IBX, an excellent reagent for oxidation of 2-furyl carbinols:
a new and general method for preparation of furyl ketones. Synth. Commun. 40,
1280–1291.
Wang, J.H., Zhao, J.L., Liu, H., Zhou, L., Liu, Z.L., Han, J.G., Zhu, Y., Yang, F.Y., 2010. Chem-
ical analysis and biological activity of the essential oils of two Valerianaceous
species from China: Nardostachys chinensis and Valeriana officinalis. Molecules
15, 6411–6422.
Yang, K., Wang, C.F., You, C.X., Geng, Z.F., Sun, R.Q., Guo, S.S., Du, S.S., Liu, Z.L.,
Deng, Z.W., 2014. Bioactivity of essential oil of Litsea cubeba from China and
its main compounds against two stored product insects. J. Asia-Pac. Entomol.
17, 459–466.
Zettler, J.L., Arthur, F.H., 2000. Chemical control of stored product insects with fumi-
gants and residual treatments. Crop Prot. 19, 577–582.
Zhang, J.S., Zhao, N.N., Liu, Q.Z., Liu, Z.L., Du, S.S., Zhou, L., Deng, Z.W., 2011. Repel-
lent constituents of essential oil of Cymbopogon distans aerial parts against two
stored-product insects. J. Agric. Food Chem. 59, 9910–9915.
Zhao, N.N., Zhou, L., Liu, Z.L., Du, S.S., Deng, Z.W., 2012. Evaluation of the toxicity of the
essential oils of some common Chinese spices against Liposcelis bostrychophila.
Food Control 26, 486–490.
Zou, Y., Wang, Q., Goeke, A., 2008. Organocatalytic multicomponent ␣-
methylenation/Diels–Alder reactions: a versatile route to substituted cyclohex-
enecarbaldehyde derivatives. Chem. Eur. J. 14, 5335–5345.