Chemical Constituents and Toxicity of Essential Oils from
Three Asteraceae Plants against Tribolium confusum
Dalila Haouas, UR06AGR05, Ecole Supérieure d’Agriculture du Kef, Université de
Jendouba, 7119, Le Kef, Tunisia, Cioni Pier Luigi, Dipartimento di Farmacia,
Università di Pisa, Via Bonanno 33, 56126 Pisa, Italy, Monia Ben Halima-Kamel,
Institut Supérieur Agronomique de Chott Mariem, Université de Sousse, 4042
Sousse, Tunisia, Flamini Guido, Dipartimento di Farmacia, Università di Pisa, Via
Bonanno 33, 56126 Pisa, Italy, and Ben Hamouda Mohamed Habib, Institut
Supérieur Agronomique de Chott Mariem, Université de Sousse, 4042 Sousse,
Tunisia
__________________________________________________________________________
ABSTRACT
Haouas, D., Cioni, P.L., Ben Halima-Kamel, M., Flamini, G., and Ben Hamouda, M.H.
2014. Chemical constituents and toxicity of essential oils from three Asteraceae plants
against Tribolium confusum. Tunisian Journal of Plant Protection 9: 67-82.
Plants produce a high diversity of secondary metabolites with a prominent function in the protection
against pests and pathogens. In this work, we studied the chemical composition and the effect of six
essential oils extracted from three Asteraceae species (Heteromera fuscata, Coleostephus myconis and
Mauranthemum paludosum) on nutritional indexes, adult emergence and larva and adult toxicity of
Tribolium confusum. Flower and leaf essential oils were obtained by hydrodistillation. The study of
their chemical composition was carried out by GC-MS. The results showed that they are rich on monoand sesquiterpens. The consumption of essential oils mixed with artificial diet of T. confusum larvae
showed that H. fuscata leaf essential oil delayed the insect growth, reduced the food consumption and
exhibited significant food deterrent index (0.02 ± 0.01 mg/mg/j, 0.05 ± 0.02 mg/mg/j, and 71.20 ±
19.22%, respectively) on treated larvae. Topical application of Asteraceae essential oils on pupae less
than 24 h of age caused malformation on emerged adults. The highest level of malformation (18%) was
induced by C. myconis leaf essential. Topical application of those essential oils on T. confusum adults
(10 and 14 days-old) showed higher toxicity. The highest mortality of about 41% was recorded 7 days
post-treatment on adults treated with essential oils from M. paludosum leaves. These preliminary
results could represent the basis for further investigations on the susceptibility of the other
developmental stages and elucidation of the mode of action of mono- and sesquiterpenoids against
these insect pests.
Keywords: Asteraceae, essential oil, ingestion, topical application, Tribolium confusum
__________________________________________________________________________
Over 60 species insect feed on
stored grains. Care must be taken to
protect the stored products against
deterioration, especially loss of quality
and weight during storage. Control of
stored product insects relies heavily on
the use of synthetic insecticides, which
has led to problems such as
environmental disturbances, increasing
costs of application, pest resistance to
Corresponding author: Dalila Haouas
Email: [email protected]
Accepted for publication 21 June 2014
Tunisian Journal of Plant Protection
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Vol. 9, No. 1, 2014
research on other Tunisian species
highlights the insecticidal activity of
Plagius grandiflorus, G. segetum,
Heteromera fuscata, Chrysanthoglossum
trifurcatum, and Coleostephus macrotus
methanolic extracts against Spodoptera
littoralis caterpillars (12, 13). H. fuscata,
P. grandiflorus and G. coronaria
essentials oils composition was also
determined and studied for their
insecticidal activity against Tribolium
confusum (11). In this work, we
investigate the chemical composition and
the insecticidal activity of H. fuscata,
Coleostephus
myconis
and
Mauranthemum paludosum essential oils
against T. confusum, a serious pest in
stored grain and related products (3, 31)
and resistant to several traditional
insecticides (4).
pesticides and consequent resurgence and
lethal effects on non-target organisms, in
addition to direct toxicity to users (15,
40). In recent years, research has focused
on the use of plant oils as possible
alternatives to synthetic chemical
insecticides. Plant essential oils and their
components have been shown to possess
potential for development as new
insecticides and they may have
advantages in terms of low mammalian
toxicity, rapid degradation and local
availability (16). Botanical pesticides
have the advantage of providing novel
modes of action against insects that can
reduce the risk of cross-resistance as well
as offering new leads for design of targetspecific molecules (15, 16). In storedproduct insect pest control, essential oils
may have numerous types of effect: they
may have a fumigant activity, penetrate
inside the insect body as contact
insecticides, act as repellents or as
antifeedants or they may affect some
biological parameters such as growth rate
(5, 14, 18, 21, 24, 27, 36).
The Asteraceae family is very
common in Mediterranean basin countries
(19, 26, 29). Some species of these plants
are characterized by their insecticidal
propriety such as Glebionis coronaria,
Glebionis
segetum
and
C.
cinerareafolium (6, 10, 37). Recent new
MATERIALS AND METHODS
Plant material.
Chrysanthemum species were
collected from three different regions of
Tunisia characterized by different
climates (Table 1). Plant identity was
confirmed by experts of the Plant Biology
Department of the University of
Monastir. Voucher specimens were
deposited in the National Gene Bank of
Tunisia.
Table 1. Former and new classification, sampling dates and climate conditions of sites of Chrysanthemum
species used in this study
Sampling
Former classification (26)
New classification (20)
Site
Site climate
date
April
Gafsa
Chrysanthemum fuscatum
Heteromera fuscata
Arid
2007
(South)
Chrysanthemum myconis
Coleostephus myconis
Chrysanthemum paludosum
ssp. glabrum
Mauranthemum paludosum
Tunisian Journal of Plant Protection
68
March
2007
May
2007
Beja
(North)
Rades
(North)
Sub-humid
Semi-arid
Vol. 9, No. 1, 2014
library mass spectra built up from pure
substances and components of known oils
and MS literature data (2, 7, 17, 23, 33,
34). Moreover, all the molecular masses
of the identified substances were
confirmed by GC/CIMS (Chemical
Ionization Mass Spectrometry), using
MeOH as ionizing gas.
Essential oil extraction.
Essential oils were extracted from
fresh leaves and flowers by steam
distillation using a Clevenger apparatus
for 4 h. The essential oils were stored at
4°C until use.
Chemical analysis.
The
GC
analyses
were
accomplished with a HP-5890 Series II
instrument equipped with HP-WAX and
HP-5 capillary columns (30 m × 0.25
mm, 0.25 mm film thickness), working
with the following temperature program:
60°C for 10 min, ramp of 5°C/min up to
220°C; injector and detector temperatures
250°C; carrier gas nitrogen (2 ml/min);
detector dual FID; split ratio 1:30;
injection of 0.5 µl). The identification of
the components was performed, for both
columns, by comparison of their retention
times with those of pure authentic
samples and by mean of their linear
retention indices (l.r.i.) relative to the
series of n-hydrocarbons.
GC/EIMS
(Electron
Impact
Ionization Mass Spectrometry) analyses
were performed with a Varian CP-3800
gas chromatograph equipped with a HP-5
capillary column (30 m × 0.25 mm, 0.25
mm film thickness) and a Varian Saturn
2000 ion-trap mass detector. Analytical
conditions: injector and transfer, line
temperatures
220
and
240°C,
respectively; oven temperature was
programmed from 60 to 240°C at
3°C/min; carrier gas was helium at 1
ml/min; injection volume was 0.2 ml
(10% hexane solution); split ratio was
1:30. Identification of the constituents
was based on comparison of the retention
times with those of authentic samples
comparing their linear retention indices
(l.r.i.) relative to the series of nhydrocarbons, and on computer matching
against commercial (2) and home-made
Tunisian Journal of Plant Protection
Insects.
Tribolium confusum larvae and
adults were obtained from laboratory
cultures maintained in the laboratory of
Entomology of the Higher Agronomic
Institute of Chott-Mariem in the dark in
incubators at 30 ± 1°C and 70-80% RH.
The two stages were reared on wheat
flour mixed with yeast (10/1, w/w).
Study of the effect of essential oils on
nutritional indexes and mortality.
Flour discs were prepared using a
previously described method (38). The
weights of the discs ranged from 35 to 39
mg. Each flour disc was treated with 5 µl
of 1% acetone solution of each essential
oil tested. Control discs were treated with
5 µl of acetone only. The discs were left
at room temperature for 15 min to allow
the solvent to evaporate. Pre-weighted
discs were placed in glass vials (5 cm
diameter). Each glass vial contained
either two untreated discs (control), or
one treated disc and one untreated disc
(choice test), or two treated discs (nochoice test). Ten group-weighed, 14 daysold T. confusum larvae were added
separately to each vial. The larvae were
starved for 24 h before starting the
experiment. Five replicates were used per
elementary treatment. The weights of the
flour discs and the number of live insects
were determined after 7 days of exposure.
Nutritional indices were calculated
according to the formulae of Manuwoto
and Scriber (22) and Farrar et al. (8) with
69
Vol. 9, No. 1, 2014
emerged adults from all live nymphs)
(25). The percentage of malformed new
emerged adults was calculated.
modifications as follows: relative growth
rate (RGR) = (A - B)/B × day-1, where A
= weight of live insects on the 7th day
(mg)/number of live insects on the 7th day
and B = initial weight of insects
(mg)/initial number of insects; relative
consumption rate (RCR) = D/B x day-1,
where
D
=
biomass
ingested
(mg)/number of live insects on the 7th
day; efficiency of conversion of ingested
food (ECI) (%) = (RGR)/(RCR) × 100.
Using the means of the amount of flour in
the control and treated discs consumed by
the insects, the percentage feeding
deterrence index (FDI) was calculated:
FDI (%) = (C - T)/C × 100, where C =
consumption of control discs and T =
consumption of treated discs. The insect
mortality (%) was recorded each seven
days during three weeks.
Statistical analyses.
In the whole experiment, the
essays were repeated 5 times to ensure the
reproducibility of the obtained results as
well as to make a correct statistical
analysis of each treatment in each
bioassay. Antifeedant and nutritional
indexes for the different treatments tested
were compared using analysis of variance
(ANOVA) followed by Duncan test for
multiple-comparison when significant
differences were observed at P < 0.05.
The recorded mortality data in
ingestion and topical toxicity tests were
adjusted for mortality in the control using
Abbott’s (1) formula then analysed by
one-way analysis of variance (ANOVA)
and means were compared using Duncan
multirange test at P < 0.05 using an SPSS
v.16.0 software package in Microsoft
Windows 7 operating system.
Insects were considered death
when tactile stimuli elicited no visible
normal reaction.
Mc = (Mo-Me)/(100-Me) × 100
with Mo: Mortality rate of treated adults,
Me: Mortality rate of control,
Mc: Adjusted mortality rate.
Contact toxicity by topical application
on adults.
A 1% acetone solution of essential
oils was prepared and 1 µl was topically
applied to the ventral surface of the
thoracic segments of the insects using a
Hamilton microsyringe. Controls were
treated with the solvent alone. After
treatment, insects were placed in an
incubator into plastic vials containing
food. Five replicates of 10 adults (10-14
days-old) were prepared. The mortality
(%) of insects was noted daily during
seven days (28).
RESULTS
Essential oil composition.
The chemical composition of all
used essential oils is listed in Table 2 and
a total of 159 compounds were counted.
The essential oil yield (v/w, volume/fresh
weight) was of 0.07% and 0.25% for C.
myconis leaves and flowers, respectively,
of 0.06% and 0.25% for M. paludosum,
respectively, and of 0.08% and 0.22% for
H. fuscata organs, respectively.
Contact toxicity by topical application
on pupae.
20 pupae of T. confusum, less than
24 h of age, were selected from breeding.
A volume of 1 µl of the same dilution of
the essential oils was applied directly on
each pupa. Pupae were placed in Petri
dishes and glass uncovered. The mortality
of pupae and the emergence of new adults
were recorded daily for 7 days (total
Tunisian Journal of Plant Protection
70
Vol. 9, No. 1, 2014
Table 2. Chemical composition of the essential oils from leaves and flowers of Heteromera fuscata,
Coleostephus myconis and Mauranthemum paludosum and percentage content of components
H. fuscata
C. myconis
M. paludosum
No
Compounda
RIb
Flower
Leaves
Flowers
Leaves
Flowers
Leaves
s
n-hexanol
871
tr
0.2
1
santolina triene
910
0.2
2
α-thujene
933
0.1
0.3
3
α-pinene
941
2.4
2.8
tr
tr
0.5
0.1
4
camphene
955
0.5
0.1
tr
0.8
5
benzaldehyde
963
tr
1.0
tr
tr
6
sabinene
978
2.2
0.8
7
β-pinene
981
4.4
tr
0.3
8
6-methyl-5986
0.9
tr
1.4
0.7
9
hepten-2-one
myrcene
992
7.1
2.2
0.2
10
yomogi alcohol
999
9.6
tr
11
limonene
1032
20.6
12.9
0.3
0.6
12
β-phellandrene
1033
6.5
13
lavender lactone
1040
3.6
tr
14
phenyl
1049
tr
tr
0.4
tr15
acetaldehydede
(E)-β-ocimene
1051
1.5
1.2
16
γ-terpinene
1064
tr
0.1
17
cis -linalool oxyde
1073
0.2
0.7
18
trans -linalool
1087
tr
0.5
19
oxyde
terpinolene
1089
0.2
0.4
20
linalool
1101
tr
0.8
tr
tr
0.5
21
nonanal
1104
tr
0.4
0.1
22
1-octen-3-yl1113
tr
1.2
1.0
0.8
1.1
23
acetate
cis-p-menth-2-en1123
0.3
0.3
tr
24
1-ol
(E)-myroxide
1145
tr
0.4
25
camphor
1145
0.2
26
isopulegol
1150
0.3
27
borneol
1167
0.3
tr
28
pinocarvone
1168
0.2
tr
29
3-thujanol
1169
0.9
30
p-mentha-1,51170
0.2
31
dien-8-ol
4-terpineol
1179
0.3
32
isopinocamphone
1181
0.5
33
cryptone
1186
1.1
0.6
34
α-terpineol
1189
tr
0.2
tr
35
n-decanal
1206
tr
tr
36
trans-piperitol
1212
0.3
0.3
0.2
37
cumin aldehyde
1241
0.1
tr
38
neral
1243
tr
tr
39
benzene acetic
1247
0.2
40
acid, ethyle ester
geraniol
1256
0.7
0.4
41
cis-chrysanthenyl
1265
0.4
0.4
0.3
42
acetate
Tunisian Journal of Plant Protection
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Vol. 9, No. 1, 2014
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
geranial
p-menth-1-en-7-al
isobornyl acetate
γ-terpinen-7-al
cumin alcohol
silphiperfol-5-ene
7-epi-silphiperfol5-ene
α-terpinyl acetate
citronellyl acetate
neryl acetate
α-copaene
β-maaliene
β-cubebene
β-caryophyllene
β-cedrene
β-gurjunene
β-copaene
(2Z,6E)-dodeca2,6-dien-1-al
α-humulene
(E)-β-farnesene
alloaromadendrene
dehydroaromadendrane
dehydrosesquicineole
geranyl npropanoate
γ-muurolene
germacrene D
a-himachalene
β-selinene
trans-muurola4(14),5-diene
valencene
bicyclogermacrene
α-muurolene
(E,E)-α-farnesene
trans-γ-cadinene
geranyl
isobutyrate
cubenol
δ-cadinene
(Z)-nerolidol (=cis
nerolidol)
Cis-calamenene
elemol
(E)-nerolidol
geranyl n-butyrate
ledol
trans-longipinanol
dendrolasin
(Z)-3-hexenyl
1272
1276
1286
1291
1291
1329
0.9
1.3
0.2
0.3
0.3
0.9
tr
tr
tr
tr
tr
tr
-
0.3
0.2
-
0.4
-
1348
-
0.2
0.2
-
-
-
1349
1353
1362
1377
1382
1388
1420
1421
1432
1432
0.5
1.6
1.9
1.6
1.7
4.2
0.5
tr
-
0.1
0.8
0.4
0.7
3.5
0.1
0.1
-
tr
tr
0.3
tr
1.2
0.2
tr
0.3
tr
tr
-
0.4
-
0.5
-
1447
-
-
-
-
0.3
-
1455
1460
2.7
-
1.7
-
5.6
1.0
-
-
1461
-
-
tr
0.3
-
-
1463
-
0.5
-
-
-
-
1472
-
-
-
0.2
-
-
1478
-
0.5
-
-
-
-
1480
1481
1483
1487
tr
10.4
-
tr
2.9
tr
0.2
0.3
1.4
tr
0.3
0.3
tr
-
0.3
-
1493
0.1
tr
-
-
-
-
1493
1495
1499
1507
1513
1.5
0.5
0.3
0.3
0.3
tr
0.3
5.5
tr
0.2
tr
0.6
0.2
-
-
1515
0.2
2.2
-
-
-
-
1515
1524
0.4
1.4
1.0
0.5
-
-
0.3
1533
-
-
0.6
-
1.3
-
1540
1550
1563
1564
1567
1569
1570
1572
0.2
-
0.2
0.4
0.5
-
0.2
1.8
0.7
0.7
-
5.1
0.3
-
tr
-
0.4
1.2
0.6
Tunisian Journal of Plant Protection
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Vol. 9, No. 1, 2014
benzoate
α-cedrene epoxide
1575
spathulenol
1577
caryophyllene
1582
91
oxide
globulol
1585
92
viridiflorol
1593
93
geranyl-2-methyl
1601
94
butanoate
geranyl-isovalerate 1607
95
(Z)1607
96
sesquilavandulol
5-epi-7-epi-α1608
97
eudesmol
humulune epoxide
1608
98
II
trans-arteannuic
1613
99
alcohol
1,10-di-epi1615
100
cubenol
humulane-1,61620
101
dien-3-ol
10-epi- γ1624
102
eudesmol
1-epi-cubenol
1629
103
10-epi-g-eudesmol
1631
104
γ-eudesmol
1632
105
α-acorenol
1633
106
β-acorenol
1637
107
6-methyl-6-(3methylphenyl)1641
108
heptan-2-one
T-cadinol
1642
109
T-muurolol
1643
110
cubenol
1647
111
α-muurolol
1647
112
α- eudesmol
1647
113
himachalol
1654
114
α-cadinol
1655
115
3-thujopsanone
1655
116
valerianol
1657
117
α-bisabolol oxide
1658
118
B
valeranone
1675
119
Heilifolenol B
1679
120
epi-α-bisabolol
1685
121
α-bisabolone oxide
1686
122
A
Z-trans- α 1691
123
bergamotol
2-pentadecanone
1695
124
(Z,E)-farnesyl
1701
125
acetate
14-hydroxy- α1714
126
humulène
pentadecanal
1716
127
Tunisian Journal of Plant Protection
89
90
2.5
0.3
3.2
1.4
0.9
1.1
1.8
0.9
3.6
1.6
4.3
1.6
7.6
0.6
-
0.4
-
0.8
0.3
0.4
0.4
-
0.4
-
2.2
7.0
-
-
-
-
-
9.3
-
-
-
-
-
-
0.7
-
0.2
-
-
-
-
0.1
-
-
-
2.5
0.5
0.4
-
0.7
-
3.8
-
-
1.3
0.4
-
-
1.3
-
-
-
-
-
-
-
0.4
-
-
0.3
-
-
-
0.3
-
0.3
0.3
0.9
0.2
1.3
-
2.1
3.1
-
-
-
1.6
-
-
-
-
-
0.7
0.7
0.9
1.4
0.3
5.5
0.2
1.6
-
0.5
0.6
0.5
1.6
-
18.4
1.7
9.4
3.8
0.2
0.3
1.0
1.0
-
0.8
0.6
1.0
2.6
-
-
-
3.1
6.7
-
-
-
0.7
-
4.8
1.1
1.2
18.0
-
-
-
-
-
0.2
-
-
-
-
-
0.5
-
-
-
-
-
0.2
-
-
-
-
3.5
-
-
-
-
-
0.5
-
-
-
-
73
-
-
0.4
Vol. 9, No. 1, 2014
128
129
130
131
132
133
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
-
pentadecanal
methyl
tetradecanoate
(E,E)-farnésol
oplopanone
(E,Z)-farnesol
α-bisabolol oxide
A
benzyl benzoate
tetradecanoic acid
14-oxy-αmuurolene
β-bisabolenal
α-bisabolyle
acetate
14-hydroxy-δcadinene
2-ethylhexylsalicylate
hexahydrofarnesyl
acetone
6,10,14-triméthyl
pentadécanone
Z-lanceol
α-chenopodiol
benzyl salicylate
oplopanonyl
acetate
11-hydroxyeudesm-4-en-3one
n-hexadecanoic
acid
11acétoxyeudésman4-α-ol
Phytol
hexadecanoate
ethyle
methyl linoleate
n-heineicosane
linoleic acid
1-docosene
n-docosane
n-tricosane
n-tetracosane
n-pentacosane
1717
-
0.3
-
-
-
0.2
1724
0.2
-
0.5
-
0.2
0.2
1725
1740
1746
tr
-
0.5
0.5
0.9
0.4
-
-
-
-
1749
-
-
-
0.6
-
-
1760
1764
-
0.1
-
-
0.8
-
1.1
1769
-
-
1.6
-
1.5
1.5
1770
-
-
-
0.4
-
-
1798
-
-
1.1
-
-
1804
0.2
-
-
-
-
-
1807
-
-
-
-
0.5
-
1843
0.2
-
-
0.7
-
1.7
1856
1857
1866
-
-
0.4
0.2
1.4
-
-
-
1888
-
-
-
0.5
-
-
1928
-
-
-
0.2
-
-
1940
-
-
0.7
1.0
-
19.1
1940
-
-
-
-
1.4
-
1943
-
-
0.3
-
-
-
1993
-
-
0.2
-
0.3
-
2096
2100
2141
2190
2200
2300
2400
2500
tr
0.6
tr
0.8
tr
tr
0.7
0.3
0.7
tr
tr
-
0.2
0.1
0.2
3.4
0.3
1.4
0.6
0.8
0.3
0.8
2.1
2.5
1.7
2.0
5.2
3.7
26.9
45.5
20.8
0.5
0.8
1.6
-
8.6
37.8
0.6
-
3.3
0.5
27.4
12.2
15.9
1.9
1.9
0.9
9.7
21.8
32.3
68.4
7.4
16.5
1843
Mononterpene
hydrocarbons
Oxygenated
mononterpenes
Sesquiterpene
hydrocarbons
Oxygenated
sesquiterpenes
Tunisian Journal of Plant Protection
0.5
74
0.7
Vol. 9, No. 1, 2014
Non-terpene
1.4
1.7
3.0
3.1
9.2
hydrocarbons
Oxygenated non2.9
2.4
0.9
0.3
3.2
terpenes
a
Compounds listed in order of elution.
b
RI: retention index relative to n-alkane under conditions listed in the experimental section.
tr = traces < 0.1.
-
0.6
the used organs. The leaves of H. fuscata
gave an essential oil especially rich in
limonene (20.6%), germacrene D (10.4%)
and myrcene (7.1%). In the case of C.
myconis, T-muurolol (18.4%), epi-αbisabolol (18.0%) and α-cadinol (9.4%)
were the main compounds detected in
flowers and (E)-β-farnesene (5.6%) and
epi-α-bisabolol (4.8%) were the most
detected in leaves. The essential oil of the
flowers of M. paludosum was particularly
rich in n-pentacosane (26.9%) and nhexadecanoic acid (19.1%) while the oil
from the leaves mainly contained npentacosane (2.5%) and n-tetracosane
(2.1%) (Table 2).
Chemical analyses of the six
samples shared a similar qualitative
composition but with some quantitative
differences. Substantially, all the essential
oils were mainly constituted by monoand
sesquiterpenoids.
Non-terpene
derivatives varied from 0.3 to 52.0%.
These compounds were almost equally
distributed between flowers and leaves in
H. fuscata (4.1% vs. 4.3%, respectively)
and C. myconis (3.3% vs 3.9%,
respectively),
while
they
were
consistently higher in flowers than leaves
in M. paludosum samples (52.0% vs.
9.2%,
respectively).
Monoterpene
hydrocarbons were not detected in M.
paludosum flowers (0.0% vs. 1.6%)
whereas they were detected in higher
amounts in H. fuscata leaves (45.5% vs.
20.8%, respectively), and were almost
equal in C. myconis ones (0.5% vs. 0.8%,
respectively). Oxygenated monoterpenes
were minor compounds in C. myconis
(0.0% and 0.6% in flowers and leaves,
respectively) and M. paludosum (0.5%
and 3.3% in flowers and leaves,
respectively). In contrast, they reached
important percentages in H. fuscata,
particularly in flowers (8.6% vs. 37.8%,
respectively).
Sesquiterpene
hydrocarbons were minor in all essential oil
where the highest amount was detected in
H. fuscata leaves (27.4%). Oxygenated
sesquiterpenes followed an opposite trend
in C. myconis (68.4% and 32.3% in
flowers and leaves, respectively), H.
fuscata (9.7% and 21.8%, respectively)
and M. paludosum (16.5% and 7.4%,
respectively). The main volatiles were
different, both for the three species and
Tunisian Journal of Plant Protection
52.0
Effect of essential oils on nutritional
indexes and insect mortality.
All the studied essential oils had
negative effects on nutritional indexes of
T. confusum larvae. Statistical analysis
showed that the oils obtained from the
leaves affected significantly the relative
growth rate (RGR), more than those
obtained from the flowers, except for
those from C. myconis that exhibited the
same effect. Concerning their activity on
relative consumption rate (RCR), the H.
fuscata
leave
essential
oil
has
significantly (P < 0.05) the highest
inhibitory activity (0.05 mg/mg/d) and the
highest inhibition level on efficiency of
conversion of ingested food (ECI).
Nevertheless the most significant
antifeedant effect on T. confusum larvae
was recorded on individuals treated with
H. fuscata leaf essential oils (71.20%)
while the flower essential oils exhibited
the lowest feeding deterrence index
(Table 3).
75
Vol. 9, No. 1, 2014
Table 3. Effect of six Asteraceae essential oils on nutritional indexes
es of Tribolium confusum larvae treated
during 7 days at concentration of 1% †
RGR
RCR
ECI (%)
FDI (%)
Species
Organ
(mg/mg/d)
(mg/mg/d)
0.12 ± 0.02 a
0.21 ± 0.00 a
73.09 ± 1.33 a
Control
Flowers
0.09 ± 0.03 b
0.15 ± 0.03 b
58.15 ± 11.62 c
15.91 ± 8.87 g
Heteromera
fuscata
Leaves
0.02 ± 0.01 f
0.05 ± 0.02 g
39.99 ± 9.77 f
71.20 ± 19.22 a
Flowers
0.06 ± 0.01 d
0.14 ± 0.05 c
40.91 ± 10.96 e
41.63 ± 23.93 e
Coleostephus
myconis
Leaves
0.06 ± 0.03 d
0.13 ± 0.04 d
41.53 ± 17.71 d
55.33 ± 22.53 b
Flowers
0.07 ± 0.02 c
0.10 ± 0.05 e
67.77 ± 16.73 b
52.69 ± 19.50 c
Mauranthemum
paludosum
Leaves
0.05 ± 0.01 e
0.09 ± 0.01 f
58.12 ± 13.83 c
40.33 ± 17.65 f
RGR: relative growth rate, RCR: relative consumption rate, ECI: efficiency of conversion of ingested food,
FDI: feeding deterrence index,
†
Means in the same column followed by the same letters are not significantly (P
( < 0.05) different as
determined by Duncan’s multirange test.
Mortality of T. confusum larvae
was affected by exposure duration and the
origin of the essential oil. Seven days
after consumption of flour discs treated
with essential oil from C. myconis leaves,
they showed statistically significant (P <
0.05) mortality (49.6%) (Fig. 1). At the
same time, essential oil extracted from H.
fuscata flowers caused the lowest
mortality and reached -6.0% after
mortality correction using Abbott’s
formula (Fig. 1). In the second week, the
percentage of mortality caused by H.
fuscata leaf essential oil increased
significantly from 8.7 to 65.0% in
comparison with the other essential oils
tested. After three weeks of treatment,
larvae fed on flour discs treated with
essential oils from C. myconis leaves
showed the highest mortality records
(85.3%) (Fig. 1).
Fig. 1. Corrected mortality (%) of treated Tribolium confusum larvae after consumption of treated
discs with six Asteraceae essential oils used at a concentration of 1%.
Tunisian Journal of Plant Protection
76
Vol. 9, No. 1, 2014
paludosum flowers induced the highest
mortality in treated pupae (23.26%) noted
seven days post-treatment
post
(Fig. 2j). No
significant differences
d
were observed
between C. myconis and H. fuscata leaf
and C. myconis flower essential oils.
Furthermore, M. paludosum leaf essential
oil induced the lowest toxicity (6.34%) on
T. confusum pupae (Fig. 2j).
Contact toxicity by topical application
on pupae.
T. confusum pupae (less than 24 hold) were treated with topical applications
of essential oils at 1% and then daily
followed-up for toxicity for seven days.
The results showed that all essential oils
exhibited variable insecticidal effets (Fig.
2j). In fact, the essential oil from M.
j
k
a
b
c
c
c
d
e
Fig. 2. Corrected mortality (j) and malformation (k) level (%) of treated Tribolium confusum pupae with six
Asteraceae essential oils used at a concentration of 1%.
HFF: Heteromera fuscata Flowers; MPL: Mauranthemum paludosum Leaves; HFL: Heteromera fuscata
Leaves; CMF: Coleostephus myconis Flowers; MPF: Mauranthemum paludosum Flowers; CML:
Coleostephus myconis Leaves
After emergence of the new adults,
an occurrence of fatal aberation was
recorded where the highest level was
registred on T. confusum pupae treated
with C. myconis leaf essential oils (18%).
These results increased the effect of this
essential oil and the new mortality
calculated was of about 35%. The
essential oil extracted from M. paludosum
flowers exibited similar toxicity against
Tunisian Journal of Plant Protection
T. confusum once the the pupae mortality
(23%) added to the deadly aberation
(12%) (Fig. 2k). Among these anomalies,
it is necessary to note the persistence of
nymphal characters with apparition of
legs (Fig. 3b) and the presence of new
adults with elytra malformation (Fig. 3c).
These anomalies
an
were also fatal and
enhanced the inhibitory potential of the
essential oils by topical application.
77
Vol. 9, No. 1, 2014
a
b
c
Fig. 3. Morphological effect of Asteraceae essential oils topically applied against Tribolium
confusum pupae at a concentration of 1% as compared to the untreated control.
(a) Control, (b) Persistence of nymphal characters, (c) Elytra
lytra malformation (arrows).
T. confusum adults (10-14 daysold) were treated with topical applications
of essential oils at 1% and then daily
surveyed for toxicity for seven days. The
results showed that all essential oils have
a variable insecticidal activity (Fig. 4). In
fact, essential oil from M. paludosum
leaves induced the highest mortality in
treated adults (41%) followed by the
those from C. myconis flowers and leaves
(23 and 20%, respectively). The lowest
toxicity (10%) was recored on individuals
treated with M. paludosum flower
essential oil (Fig. 4).
Fig. 4. Corrected mortality level (%) of treated Tribolium confusum adults with studied essential
oils applied at a concentration of 1%.
Tunisian Journal of Plant Protection
78
Vol. 9, No. 1, 2014
Yang et al. (2005) also highlighted that
essential oil composition can vary with
geographical distribution, harvesting
time, growing conditions and method of
extraction (39).
Concerning
the
insecticidal
activity of the essential oils used against
T. confusum in this study, it should be
noted that these oils exhibited a negative
effect on nutritional indices which was
more accentuated when diet was treated
with essential oil extracted from H.
fuscata leaves. Based on chemical
analysis, this last essential oil was found
to be rich in monoterpens especially in
limonene (20.6%). According to Raina et
al. (30), this compound can significantly
reduce the insect feeding (30). Moreover,
this compound is the most toxic one for
all T. confusum stages and particularly for
the larval stage (32).
In contact method (topical
application), this study also demonstrated
that M. paludosum leaf essential oils had
a lethal effect on T. confusum. Its
chemical analyses showed that there are
no major compounds in this essential oil,
so this effect can be attributed to their
high lipophilicity that permits them to
rapidly penetrate into insect bodies and
interfere
with
their
physiological
functions.
In conclusion, this research
revealed that each essential oil of the
three Asteraceae species has its specific
mode of action on the same beetle, T.
confusum. In fact, the essential oils
extracted from H. fuscata can affect the
nutritional behavior and led to the death
of insect by starvation. Those from M.
paludosum induced an insecticidal
activity by contact whereas those of C.
myconis can contribute to the death of the
confused beetle by hormone perturbation
(lethal malformation). Thus, these
essential oils may be used as sources of
DISCUSSION
Plant essential oils are considered
to be an alternative means of controlling
many insect pests (35). In integrated
stored product protection, phytochemicals
may be used for pest prevention, early
pest detection or pest control (21).
This study reported the chemical
composition of six essential oils extracted
from three species belonging to the
Asteraceae family (H. fuscata, C. myconis
and M. paludosum) collected from
different region in Tunisia. All these
essential oils were screened for their
potential insecticidal activity against T.
confusum larvae and pupae. Based on
these findings, these three species
presented
a
similar
qualitative
composition but a very different
distribution of the main volatiles in their
essential oils depending on used organs.
Chemical analysis revealed that these
essential oils were rich on mono- and
sesquiterpenes at different levels. The
distribution of the main volatiles
compounds depended on species tested
and organs used. Few studies are reported
concerning the essential oil composition
of H. fuscata, C. myconis and M.
paludosum, however, analyzes reported in
a previous study by Haouas et al. (11)
showed a similar qualitative composition
between G. coronaria, H. fuscata, and P.
grandiflorus samples and a variation in
the percentage of each volatile
compound. Nevertheless, the percentage
of volatile compounds can vary for the
same species depending on sampling
sites. This hypothesis was confirmed by
Flamini et al. (9) who demonstrated that
essential oil extracted from G. coronaria
flowers growing in Italy contained more
camphor (22.1%) and cis-chrysanthenyl
acetate (19.9%) than that extracted from
the same species and collected from
Tunisia (camphor 12.6%, 13.4%) (11).
Tunisian Journal of Plant Protection
79
Vol. 9, No. 1, 2014
actives biomolecules for
environmentally friendly
biopesticides.
ACKNOWLEDGMENTS
The authors are grateful to the International
Foundation for Science (IFS grant N°. F/3968-1)
and to the Organization for the Prohibition of
Chemical Weapons (OPCW) for financial support.
They also thank Prof. Fethia Skhiri-Harzallah for
her help in the identification of Chrysanthemum
species.
developing
and safe
__________________________________________________________________________
RESUME
Haouas D., Cioni P.L., Ben Halima-Kamel M., Flamini G. et Ben Hamouda M.H. 2014.
Constituants chimiques et toxicité des huiles essentielles de trois plantes Asteraceae
contre Tribolium confusum. Tunisian Journal of Plant Protection 9: 67-82.
Les plantes produisent une grande diversité de métabolites secondaires qui les protègent contre les
ravageurs et les agents pathogènes. Dans ce travail, nous avons étudié la composition chimique et
l’effet de six huiles essentielles extraites de trois espèces d’Asteraceae (Heteromera fuscata,
Coleostephus myconis et Mauranthemum paludosum) sur l'indice nutritionnel, l'émergence des adultes
et la toxicité contre les larves et des adultes de Tribolium confusum. Les huiles essentielles des fleurs et
des feuilles ont été obtenues par hydrodistillation. L'étude de leur composition chimique a été réalisée
par CG-SM. Les résultats ont montré qu'elles sont riches en mono-et sesquiterpènes. La consommation
des huiles essentielles mélangées avec le milieu artificiel des larves de T. confusum a montré que
l’huile essentielle de H. fuscata a retardé la croissance des insectes, réduit la prise de la nourriture et a
présenté l'indice d’anti-appétence le plus important contre les larves traitées (respectivement de 0,02 ±
0,01 mg/mg/j, 0,05 ± 0,01 mg/mg/j et 71,20 ± 19,22). L'application topique des huiles essentielles de
ces trois Asteraceae contre les nymphes âgées de moins de 24 heures a provoqué des malformations
chez les nouveaux adultes émergés. Le plus haut taux de malformation (18%) a été induit par l’huile
essentielle des feuilles de C. myconis. L'application topique de ces huiles essentielles sur les adultes de
T. confusum (âgés de 10 et 14 jours) a montré une toxicité importante. La plus forte mortalité d’environ
41% a été enregistrée, 7 jours après le traitement, chez les insectes traités par l’huile essentielle des
feuilles de M. paludosum. Ces résultats préliminaires pourraient constituer la base pour d'autres
investigations sur la sensibilité des autres stades de développement et pour l’élucidation du mode
d'action des mono- et sesquiterpènes contre ces insectes.
Mots clés: Application topique, Asteraceae, huile essentielle, ingestion, Tribolium confusum
___________________________________________________________________________
‫ ا ( ' ت‬.2014 .‫َ دة‬
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$% ‫و‬# ‫و‬
‫ س‬1 ‫ة ا‬+2 #3 (Asteraceae) ‫ا ّ ت‬
0%
Tunisian Journal of Plant Protection 9: 67-82.
‫ ت‬- !'
.$/
‫نو‬
‫ د و ر‬،‫اس‬
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‫رزة‬
‫ذات وظ‬
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‫ ّ& ت‬% ‫ ا‬0 1
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‫ ز ت رو‬6 7 8 ‫" ! ا‬# ‫ ا‬9‫را‬: % ; ،= > ‫ا‬
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‫ ت‬A ! ‫; ت وا‬# ‫ ا‬D0$
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‫و‬# ‫ ت ا‬K ‫ ا‬D0$ ‫ ل‬1G ‫ ا‬M' .Tribolium confusum
9‫ و إط ر درا‬. % #' ‫اد‬
%W ‫ ت‬K ‫ه ا‬,‫( أن ھ‬7 % ‫ت ا‬# ‫ أظ‬.GC-MS %O' ‫ > ل‬9 7 8 ‫" ! ا‬#' 9‫درا‬
‫ ء‬6 %5 ‫ ا‬G ‫وج‬K H. fuscata !%
‫و‬# ‫ ا‬R K ‫ك ا‬3 9‫زت ا & رب أن ا‬# ‫ أ‬،‫ات‬#FG0 ‫ة‬: ! ‫ ا‬0$
0,01 ± ) ‫ة‬#FG ‫ ا‬B :G ‫ ا‬#BC D0$‫& = أ‬6'‫اء و‬,A0 "3 9‫ ا‬:G ‫ة وا‬#FG ‫ا‬
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Tunisian Journal of Plant Protection
80
Vol. 9, No. 1, 2014
‫و‬# ‫ ت ا‬K ‫ ا‬R>.‫ و‬:%$ .( ‫ ا ا‬D0$ ،%19,22 ± 71,20 ‫ م و‬/]0 /]0 0,02±0,05 ،‫ م‬/]0 /]0 0,02
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‫ ا‬H ‫ا‬#F ‫ ا‬D0$ ‫ة‬#B ! ‫! ' ت‬% ‫ه ا‬,
>. ‫ ا‬H !E ‫( أن ا‬7 % ‫زت ا‬# ‫ أ‬.(%18) C. myconis !%
‫و‬# ‫ ا‬R K ‫ ا‬$ 4 ‫ ّ ه‬F' !6 D0$‫ أ‬R "‫و‬
!6% R " ` ، ّ 9 !6 D0$‫& = أ‬6' D ‫( أدى إ‬
14 ‫ و‬10) ‫ة‬#FG ‫ا‬
‫ ت‬A ! ‫ ا‬D0$ ‫و‬# ‫ ت ا‬K ‫? ا‬0
‫و‬+‫( ا‬7 % ‫ه ا‬,‫ أن ' = ھ‬8 . 0 > ‫ا‬
‫ أ م‬7 :> ،%41 M. paludosum ! ‫ھ ر‬K
‫و‬# ‫ ا‬R K ‫ ا‬D ‫إ‬
D0$ % # ‫ = ا اد ا‬$ ‫ق‬#‫ط‬
H > ‫ ? ا‬,"‫ات و‬#FG ‫ة ا‬
‫ى‬#J+‫ا = ا‬# ‫ ا‬0 ; ‫ ث‬G! ‫ ا‬: K 9 9‫أ‬
.‫ات‬#FG ‫ه ا‬,‫ھ‬
Tribolium confusum ،Asteraceae ‫ّ& ت‬% ‫ ا‬، ‫ ز ت رو‬، >. H !E' ،‫ع‬3 ‫ ا‬:
‫ ت‬0"
___________________________________________________________________________
LITERATURE CITED
1. Abbott, W.S. 1925. A method of computing the
effectiveness of an insecticide. J. Econom.
Entomol. 18: 265-267.
2. Adams, R.P. 1995. Identification of essential oil
components by gas chromatography mass
spectroscopy. Carol Stream, Illinois: Allured
Publ. Corp.
3. Aitken, A.D. 1975. Insect travelers, I: Coleoptera.
Technical bulletin 31, H.M.S.O, London
4. Arthur, F.H. 1996. Grain protectants: current
status and prospects for the future. J. Stored
Prod. Res. 32: 293-302.
5. Benzi, V., Stefanazzi, N., and Ferrero, A. 2009.
Biological activity of essential oils from leaves
and fruits of pepper tree (Schinusmolle L.) to
control rice weevil (Sitophilus oryzae L.).
Chilean J. Agric. Res. 69: 154-159.
6. Cũnat, P., Primo, E., Sanz, I., March, M.C.,
Bowers, W.S., and Martinez-Pardo, R., 1990.
Biocidal
activity
of
some
Spanish
Mediterranean plants. J. Agric. Food Chem. 38:
497-500.
7. Davies, N.W. 1990. Gas chromatographic
retention indices of monoterpenes and
sesquiterpenes on methyl silicone and carbowax
20 M phases. J. Chromatogr. A 503: 1-24.
8. Farrar, R.R., Barbour, J.D., and Kenedy, G.G.
1989. Quantifying food consumption and
growth in insects. Ann. Entomol. Soc. Am. 82:
593-598.
9. Flamini, G., Cioni, P.L., and Morelli, I. 2003.
Differences in the fragrances of pollen, leaves,
and floral parts of garland (Chrysanthemum
coronarium) and composition of the essential
oils from flowerheads and leaves. J. Agric.
Food Chem.51: 2267-2271.
10. Gao, Y., Wu, W.L., Wu, Y.L., Ye, B., and Zhou,
R. 1998. A straight forward synthetic approach
to the spiroketal-eno ethers synthesis of natural
antifeeding compound Tonghaosu and ist
analogs. Tetrahedron 54: 12523-12538.
11. Haouas, D., Cioni, P.L., Ben Halima-Kamel, M.,
Flamini, G., and Ben Hamouda M.H. 2012.
Tunisian Journal of Plant Protection
Chemical composition and bioactivity of three
Chrysanthemum essential oils against Tribolium
confusum (du Val) (Coleoptera: Tenebrionidae).
J. Pest Sci. 85: 367-379.
12. Haouas D., Flamini G., Ben Halima-Kamel M.,
and Ben Hamouda M.H. 2011. Identification of
an insecticidal polyacetylene derivative from
Chrysanthemum macrotum leaves. Indust. Crop.
Prod. 34: 1128-1134.
13. Haouas, D., Flamini G., Ben Halima-Kame, M.,
and Ben Hamouda, M.H. 2010. Feeding
perturbation
and
toxic
activity
of
Chrysanthemum
crude
extract
against
Spodoptera littoralis (Boisduval) (Lepidoptera;
Noctuidae). Crop Prot. 29: 992-997.
14. Huang, Y., Tan, J., Kini, R., and Ho, S. 1997.
Toxic and antifeedant action of nutmeg oil
against Tribolium castaneum (Herbst) and
Sitophilus zeamais Motsch. J. Stored Prod. Res.
33: 289-298.
15. Isman, M.B. 2006. Botanical insecticides,
deterrents, and repellents in modern agriculture
and an increasingly regulated world. Ann. Rev.
Entomol. 51: 45-66.
16. Isman, M.B. 2000. Plant essential oils for pest
and disease management. Crop Prot. 19: 603608.
17. Jennings, W. and Shibamoto, T. 1980.
Qualitative analysis of flavor and fragrance
volatiles by glass capillary chromatography,
Academic Press Edition, New York, 467 pp.
18. Kordali, S., Aslan, I., Almasur, O., and Cakir, A.
2006. Toxicity of essential oils isolated from
three Artemisia species and some of their major
components to granaryweevil, Sitophilus
granarius (L.) (Coleoptera: Curculionidae).
Indust. Crop. Prod. 23: 162-170.
19. Lee, K.D., Yang, M.S., Ha, T.J., Park, K.M., and
Park, K.H. 2002. Isolation and identification of
dihydrochrysanolide and its 1-epimer from
Chrysanthemum coronarium L. Biosci.
Biotechnol. Biochem. 66: 862-865.
81
Vol. 9, No. 1, 2014
20. Le Floc'h, E., Boulos, L., Véla, E., GhrabiGammar, Z., Daoud-Bouattour, A., Ben SaadLimam, S., Martin, R., Muller, S.D., Reduron,
J.P., and Tison, J.M. 2010. Flore de Tunisie.
Catalogue synonymique commenté. Ministère
de l’environnement et du développement
Durable et Banque Nationale de Gènes
Editions, Tunisie, 500 pp.
21. Lόpez, M., Jordán, M., and Pascual-Villalobos,
M. 2008. Toxic compounds in essential oils of
coriander, caraway and basil active against
stored rice pest. J. Stored Prod. Res. 44: 273278.
22. Manuwoto, S. and Scriber, J.M. 1982.
Consumption and utilization of three maize
genotypes by the southern armyworm. J.
Econom. Entomol. 75: 163-167.
23. Massada, Y. 1976. Analysis of essential oils by
gas chromatography and mass spectrometry.
John Wiley & Sons Editions, New York, 334
pp.
24. Nerio, L., Olivero-Verbel, J., and Stashenko, E.
2009. Repellent activity of essential oils from
seven aromatic plants grown in Colombia
against Sitophilus zeamais Mostschulsky
(Coleoptera). J. Stored Prod. Res. 45: 212-214.
25. Pascual-Villalobos, M.J. and Robledo A. 1999.
Anti-insect activity of plant extracts from the
wild flora in southeastern Spain. Biochem. Syst.
Ecol. 27: 1-10.
26. Pottier-Alapetite, G. 1981. La flore de la
Tunisie :
Angiosperme-Dicotylédones
Gamopétales. Ministère de l’Enseignement
Supérieur et de la Recherche Scientifique et le
Ministère de l’Agriculture Editions, Tunisie,
1189 pp.
27. Pungitore, C., Garcia, M., Gianello, J., Tonn, C.,
and Sosa, M. 2005. Lethal and sublethal effects
of
triterpenes
from
Junellia
aspera
(Verbenaceae) on the grain storage insect
Tribolium
castaneum
(Coleoptera:
Tenebrionidae). Revista de la Sociedad.
Entomologica Argentina 64: 45-51.
28. Pungitore, C.R., Garcıa, M., Gianello, J.C., Sosa,
M.E., and Tonn, C.E. 2005. Insecticidal and
antifeedant effects of Junellia aspera
(Verbenaceae) triterpenes and derivatives on
Sitophilus oryzae (Coleoptera: Curculionidae).
J. Stored Prod. Res. 41: 433-443.
29. Quezel, P. and Santa, S. 1981. Nouvelle flore de
l’Algérie
et
des
régions
désertiques
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
méridionales, Tome II, Centre national de la
recherche scientifique Editions, Algérie, 1170
pp.
Raina, A.K., Bland, J., Dollittle, M., Lax, A.,
Boopathy, R., and Lolkins, M. 2007. Effect of
orange oil extract on the formosan subterranean
termite (Isoptera: Rhinotermitidae). J. Econom.
Entomol. 100: 880-885.
Rees, D.P. 1995. Coleoptera. Pages 1-39. In:
Integrated management of insects in stored
products. Subramanyam, B.H., Hagstrum,
D.W., Ed. Marcel Dekker, New York.
Stamopoulos, D.C., Damos, P., and
Karagianidou, G. 2007. Bioactivity of five
monoterpenoid vapours to Tribolium confusum
(du Val) (Coleoptera: Tenebrionidae). J. Stored
Prod. Res. 43: 571-577.
Stenhagen, E., Abrahamsson, S., and
McLafferty, F.W. 1974. Registry of mass
spectral data. John Wiley & Sons Editions, New
York, 3136 pp.
Swigar, A.A. and Silverstein, R.M. 1981.
Monoterpenes. Milwaukee: Aldrich Chemical
Company, USA, 130 pp.
Tripathi, A., Upadhyay, S., Bhuiyan, M., and
Bhattacharya, P. 2009. A review on prospects of
essential oils as biopesticide in insect-pest
management. J. Pharmacognosy Phytother. 1:
52-63.
Ukeh, D., Birkett, M., Pickett, J, Bowman, A.,
and Mordue Luntz, A. 2009. Repellent activity
of alligator pepper, Aframomum melegueta, and
ginger, Zingiber offcinale, against the maize
weevil, Sitophilus zeamais. Phytochemistry 70:
751-758.
Valente, M.E., Borreani, G., Caredda, S.,
Cavallarin, L., and Sulas, L. 2003. Ensiling
forage garland (Chrysanthemum coronarium L.)
at two stages of maturity and at different wilting
levels. Anim. Feed Sci.Technol. 108: 181-190.
Xie, Y.S., Bodnaryk, R.P., and Fields, P.G.
1996. A rapid and simple flour disk bioassay for
testing natural substances active against storedproduct insects. Can. Entomol. 128: 865-875.
Yang, P., Ma, Y., and Zheng, S. 2005.
Adulticidal activity of five essential oils against
Culex pipiens Quinque faciatus. J. Pest Sci. 30:
84-89.
Zettler, J.L. and Arthur, F.H. 2000. Chemical
control of stored product insects with fumigants
and residual treatments. Crop Prot. 19: 577-582.
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