Insect growth inhibition, antifeedant and antifungal activity of compounds isolated/ derived from Zingiber officinale Roscoe (ginger) rhizomes

Article abstract of DOI:10.1002/ps.263

Fresh rhizomes of Zingiber officinale (ginger), when subjected to steam distillation, yielded ginger oil in which curcumene was found to be the major constituent. The thermally labile zingiberene-rich fraction was obtained from its diethyl ether extract. Column chromatography of ginger oleoresin furnished a fraction from which [6]-gingerol was obtained by preparative TLC. Naturally occurring [6]-dehydroshogaol was synthesised following condensation of dehydrozingerone with hexanal, whereas zingerone and 3-hydroxy-1-(4-hydroxy-3-methoxyphenyl)butane were obtained by hydrogenation of dehydrozingerone with 10% Pd/C. The structures of the compounds were established by ¹H NMR, ¹³C NMR and mass (EI-MS and ES-MS) spectral analysis. The test compounds exhibited moderate insect growth regulatory (IGR) and antifeedant activity against Spilosoma obliqua, and significant antifungal activity against Rhizoctonia solani. Among the various compounds, [6]-dehydroshogaol exhibited maximum IGR activity (EC₅₀ 3.55mg ml^-1) while dehydrozingerone imparted maximum antifungal activity (EC₅₀ 86.49mg litre^-1).

Full text of DOI:10.1002/ps.263

Pest Management Science
Pest Manag Sci 57:289±300 (2001)
Insect growth inhibition, antifeedant and
antifungal activity of compounds isolated/
derived from Zingiber of®cinale Roscoe (ginger)
rhizomes
Manjree Agarwal,1 Suresh Walia,1* Swaran Dhingra1 and Bhupinder PS Khambay2
1Division of Agricultural Chemicals, Indian Agricultural Research Institute, New Delhi - 110 012, India†
2Biological and Ecological Chemistry Department, IACR, Rothamsted, Harpenden, Hertfordshire AL5 2JQ, UK
Abstract: Fresh rhizomes of Zingiber of®cinale (ginger), when subjected to steam distillation, yielded
ginger oil in which curcumene was found to be the major constituent. The thermally labile zingiberene-
rich fraction was obtained from its diethyl ether extract. Column chromatography of ginger oleoresin
furnished a fraction from which [6]-gingerol was obtained by preparative TLC. Naturally occurring
[6]-dehydroshogaol was synthesised following condensation of dehydrozingerone with hexanal,
whereas zingerone and 3-hydroxy-1-(4-hydroxy-3-methoxyphenyl)butane were obtained by hydro-
genation of dehydrozingerone with 10% Pd/C. The structures of the compounds were established by 1H
NMR, 13C NMR and mass (EI-MS and ES-MS) spectral analysis. The test compounds exhibited
moderate insect growth regulatory (IGR) and antifeedant activity against Spilosoma obliqua, and
signi®cant antifungal activity against Rhizoctonia solani. Among the various compounds, [6]-
dehydroshogaol exhibited maximum IGR activity (EC50 3.55mg ml 1) while dehydrozingerone
imparted maximum antifungal activity (EC50 86.49mg litre 1).
# 2001 Society of Chemical Industry
Keywords: Zingiber of®cinale; gingerol; dehydrozingerone; dehydroshogaol; zingerone; insect growth inhibition;
antifeedant; antifungal
1
INTRODUCTION
¯avour and pleasant aroma.4 The characteristic
pleasant and aromatic odour of ginger is due to
constituents such as monoterpene and sesquiterpene
hydrocarbons and alcohols,5,6 which can be separated
from the essential oil isolated from unscraped rhi-
zomes by steam distillation. Ginger oleoresin, how-
ever, encompasses several closely related phenolic
alkanones such as gingerols, shogaols, zingerones,
paradols, gingerdiols and other pungent principles like
diarylheptanoids, gingerenones, dehydroshogaol and
cyclic diarylheptanoids.7±13 Many of these compounds
and crude extractives are known for their medic-
inal14,15 and anti-oxidative properties.16
The development of resistance to existing classes of
pesticide and the increasing public concern over
environmental pollution and toxicity generate a con-
tinuing need for new classes of pest control agents with
even higher activity against the target organisms, and
lower impact on the environment. Of particular
interest are insect growth regulators1 that target pro-
cesses unique to insects (eg moulting) and are there-
fore compatible with Integrated Pest Management
(IPM) and Integrated Crop Management (ICM)
programmes.2,3 During the last two decades, both
synthetic and botanical pesticides from a variety of
sources have been identi®ed and developed in this
regard. Neem extract (from Azadirachta indica A Juss)
is a particularly good example of the latter class that
also has prompted a renewed interest in botanical
pesticides. Other contributory factors include the
observation that botanical extracts often exhibit
activity against more than one class of pests.
Crude ginger extractives have earlier been reported
to exhibit antifeedant and insect growth disruption
activity against Spodoptera litura F larvae17,18 and to
control stored grain pests in cowpea, Callosobruchus
maculatus F,19 and the cowpea aphid Aphis craccivora
Koch.20 They are also active against several plant
pathogens, including those causing powdery mildew of
pea21 and cercospora leaf spot of mungbean,22 and
control seven seed-borne fungi of mustard.23 Crude
Zingiber of®cinale Roscoe (family Zingiberaceae),
commonly called ginger, is known for its pungency,
* Correspondence to: Suresh Walia, Division of Agricultural Chemicals, Indian Agricultural Research Institute, New Delhi - 110 012, India
E-mail: Suresh_walia@hotmail.com
† Contribution number 683
Contract/grant sponsor: University Grants Commission, New Delhi
Contract/grant sponsor: Biotechnology and Biological Sciences Research Council of the United Kingdom
(Received 19 January 2000; revised version received 21 June 2000; accepted 30 August 2000)
# 2001 Society of Chemical Industry. Pest Manag Sci 1526±498X/2001/$30.00
289

M Agarwal et al
ginger aqueous extract has also been reported to
inhibit growth of Aspergillus ¯avus Link, A niger Van
Tiegh and Rhizopus stolonifer (Ehrenberg) Linder in
orange and pineapple juices,24 while its combination
with the synthetic fungicide thiophanate-methyl
(TPM) has been found to be active against TPM-
resistant Penicillium digitatum Sacc causing green
mould of mosambi.25 Ginger oil has somewhat
different properties and been found to control growth
of Fusarium moniliforme Sheldon on mung seed26 and
repel the cockroach Periplaneta americana (L).27
Furthermore, the non-volatile diarylheptenone con-
stituents of ginger oleoresin have been found to be
active against the rice fungus Pyricularia oryzae Cav.11
Since most of the studies on the evaluation of ginger
products for pest control potential have been centred
around crude extracts/mixtures, the present study was
aimed at evaluating some of the major constituents
and their synthetic derivatives for insect growth
inhibitory, antifeedant and antifungal activity.
2.2.3 Gas Chromatography-mass spectroscopy (GC-
MS)
Gas chromatography-mass spectroscopy (electron
impact mass spectra) [GC-MS (EI-MS)] was carried
out on a HRGC-MEGA 2 series GC coupled with a
Fisons TRIO-1000 ion trap mass spectrometer at an
ionization potential of 70eV. The GC was ®tted with a
SE-54 capillary column (15mÂ0.25mm ID). The
initial temperature was kept at 60°C for 1min and
1
then heated at 15°C min up to 250°C (10min).
Helium was used as the carrier gas.
2.2.4 Electrospray mass spectroscopy (ES-MS)
Electrospray mass spectroscopy was carried out on a
VG platform quadruple mass spectrometer (Fison
Instruments) by ¯ow injection analysis via a Rheodyne
injection loop (10ml) into a stream of acetonitrile‡
water (1‡1 by volume) ¯owing at 5ml min 1 into the
electrospray probe. The spectrometer was scanned in
positive ion mode from 100 to 800amu.
2.3 Extraction of essential oil and isolation of
curcumene and zingiberene
2
MATERIALS AND METHODS
2.1 Chemicals and other plant materials
Fresh ginger rhizomes (1kg) were ground, loaded into
a 3-litre ¯ask containing water (1.5 litre) and subjected
to steam distillation to furnish a golden coloured oil
(1.8g). This was preserved in the refrigerator for
further use.
Laboratory grade reagents and solvents were locally
procured. The solvents were distilled and, wherever
required, dried before use. Hexanal and sodium
bis(trimethylsilyl)amide were purchased from Aldrich
Chemicals. The dried and fresh rhizomes of Z of®cinale
procured from the local market were identi®ed by the
Department of Botany, University of Delhi, Delhi.
Ready-made potato-dextrose-agar (PDA) medium
was obtained from Hi-Media Lab, Mumbai. The
culture of Rhizoctonia solani Kuhn was obtained from
Division of Mycology and Plant Pathology, IARI, New
Delhi.
2.3.1 Curcumene
Steam-distilled ginger oil (1g) was adsorbed on silica
gel (3g) and the material loaded onto a column
(45cmÂ1cm ID) packed with activated silica gel (60±
140 mesh). The column was eluted with hexane and
50 fractions (10ml each) were collected. The ®rst ®ve
fractions of the same Rf value were combined and the
solvent removed under vacuum to yield curcumene
(200mg, 20% yield, 91% purity). GC-MS (EI-MS)
2.2 Chromatography and spectroscopy
2.2.1 Thin layer chromatography (TLC)
‡
m/z (relative intensity): 202 (45%, M ), 159 (10%),
145 (55%), 133 (29%), 132 (99%), 119 (100%), 105
(93%), 91 (92.7%), 77 (52%) and 55 (79%).
Activated silica gel coated plates (6cmÂ20cm) of
0.25mm thickness were used. Plates were developed
in suitable solvents, air dried and visualised with iodine
vapour or by spraying with 10% sulfuric acid solution.
Preparative TLC was performed on silica gel coated
glass plates (20cmÂ20cm) of 0.25mm thickness. The
spots on preparative TLC plates were marked by
comparing each one with the reference plate, and then
scraped and extracted with a suitable solvent. The
solvent was evaporated under vacuum to obtain the
compounds.
2.3.2 Zingiberene
Fresh ginger rhizomes (500g) were chopped into small
pieces and stirred with diethyl ether (2Â1 litre) for 6h.
The combined ether extract was ®ltered and solvent
evaporated at room temperature. The oil (500mg) so
obtained was subjected to preparative TLC (hexane‡
diethyl ether, 1‡1 by volume). The uppermost layer
was scraped and extracted with diethyl ether to yield a
compound (75mg, 15% yield) which was identi®ed as
zingiberene (75% purity) by its mass spectral data.
GC-MS (EI-MS) m/z (relative intensity): 204 (16%,
2.2.2 Nuclear magnetic resonance spectroscopy (NMR)
1H NMR spectra of dehydrozingerone, dehydro-
shogaol, zingerone and 3-hydroxy-1-(4-hydroxy-3-
methoxyphenyl)butane were recorded on a Jeol
400MHz instrument using deuterated chloroform as
solvent and tetramethylsilane as internal standard.
The spectrum of [6]-gingerol was recorded on a
Bruker 300 AC (300MHz) instrument. 13C NMR
was performed on a Jeol (100MHz) instrument.
‡
M ), 189 (31%), 161 (12%), 162 (14%), 147 (18%),
133 (20%), 119 (79%), 69 (100%).
2.4 Extraction of oleoresin from fresh/dry ginger
Fresh/dry ginger rhizomes (1kg each) were sliced/
ground, and stirred in hexane (3Â2 litre) for 24h. The
combined hexane extract was ®ltered and concen-
290
Pest Manag Sci 57:289±300 (2001)

Biological properties of Zingiber of®cinale Roscoe isolates
trated under vacuum to yield essential oil. Fresh ginger
rhizomes afforded a golden yellow oil (3.2%), whereas
the dry ginger yielded a light brown oil (2.5%). The
de-oiled material was further extracted with dichloro-
methane (3Â2 litre) and the combined dichloro-
methane extract was concentrated under vacuum to
yield oleoresin (2.2% from fresh ginger and 2.7% from
dry ginger).
the reaction. After neutralisation with dilute hydro-
chloric acid, the mixture was extracted with ethyl
acetate. The organic phase was washed with water,
and after evaporating the solvent, the resultant
product was dissolved in ethanol and kept in a
refrigerator for crystallisation to produce shiny light
yellow crystals of dehydrozingerone (4-hydroxy-3-
methoxybenzylideneacetone) (2.0g, 83% yield), mp
128±129°C, Rf 0.46 in chloroform and 0.23 in diethyl
ether ‡hexane, 4‡1 by volume. 1H NMR (400MHz)
d: 2.37 (3H, s, ÐCOCH3), 3.91 (3H, s, ÐOCH3),
6.286 (1H, s,ÐOH Ar), 6.567 (1H, d, J =16Hz, H-2,
[ÐC(O)-CH=CH-Ar], 6.94±6.97 (1H, d, J =13Hz,
H-5', ortho-coupled), 7.055 (1H, s, H-2'), 7.07±7.09
(1H, m, H-6', ortho and meta coupled), 7.43±7.47
(1H, d, J =16Hz, H-1 [ÐC(O)-CH=CH-Ar]. 13C
NMR d: 27.267 (C-4), 55.94 (OCH3), 109.426
(C-2'), 114.92 (C-5'), 123.54 (C-2), 124.88 (C-6'),
126.836 (C-1'), 143.96 (C-1), 146.999 (C-4'),
148.407 (C-3'), 198.67 (C-3). ES-MS m/z (relative
intensity): 615 (2%), 599 (5%), 423 (4%), 408 (5%),
2.4.1 [6]-Gingerol
Ginger oleoresin (1g) from fresh ginger rhizome was
adsorbed on silica gel (3g), and the resultant material
loaded onto a column (75cmÂ1.5cm ID) packed
with silica gel. It was eluted with hexane, hexa-
ne‡ethyl acetate (as gradient) and ethyl acetate. A
total of 80 fractions of 20ml each was collected. After
removal of the solvent under vacuum, each fraction
was spot-tested by TLC and the fractions possessing
the same Rf values were combined. Fractions 20±25
eluted with hexane‡ethyl acetate (8‡2 by volume)
were comparatively pure and were combined to yield
gingerol mixture (240mg). The ES-MS of this fraction
showed the presence of peaks typical of [6]-, [8]-, [10]-
and [12]-gingerols (Table 1). [6]-Gingerol (47mg,
39.2% yield with 85% purity) was isolated by
preparative TLC (hexane‡ethyl acetate, 7‡3 by
volume) of the gingerol mixture (120mg). Rf 0.47 in
benzene‡ethyl acetate, 7‡3 by volume, and 0.25 in
diethyl ether ‡hexane, 4‡1 by volume. 1H NMR
(300MHz) d: 0.88 (3H, t, J =6.64Hz, H-10), 1.27
(6H, m, H-7, H-8, H-9), 1.38 (2H, m, H-6), 2.50±
2.53 (1H, d, J =9.93Hz, H-4b), 2.54±2.56 (1H, d
J =6.39Hz, H-4a), 2.72±2.75 (2H, dd, J =6.8 and
6.96Hz, H-2), 2.82±2.85 (2H, dd, J =6.8 and 6.7Hz,
H-1), 3.85 (3H, s, ÐOCH3), 5.69 (1H, brs,ÐOH),
4.02 (1H, m, H-5), 6.67 (1H, dd, J =3 and 8Hz,
H-6', ortho and meta coupled), 6.70 (1H, s, H-2'), 6.81
(1H, d, J =8Hz, H-5', ortho coupled). ES-MS m/z
(relative intensity): 611 (2%), 588 (2.5%), 340 (3%),
‡
407 (11%), 215 (7%), 193 (100%, M‡H ), 175
(13%).
2.5.2 [6]-Dehydroshogaol
Dehydrozingerone (10.4mmol; 2.0g) solution in dry
tetrahydrofuran (THF; 15ml) was slowly added under
an atmosphere of nitrogen to a stirred solution of
sodium bis(trimethylsilyl)amide (20.8mmol; 7.0ml)
in THF. After 1h, hexanal (10.4mmol) was added and
stirring continued for another 3h at 0°C. After
completion of the reaction (TLC), ethyl acetate was
added and the resulting mixture was washed with
dilute hydrochloric acid (5%), and then with saturated
aqueous sodium chloride solution. The organic layer
was dried over anhydrous sodium sulfate and con-
centrated under vacuum. The residue was puri®ed by
¯ash column chromatography to afford [6]-dehy-
droshogaol as a yellow viscous product (1.25gm; yield
45%). Rf 0.65 (hexane‡ethyl acetate, 1‡1 by
‡
333 (2%), 317 (8%), 295 (26%, M‡H ), 277 (61%),
1
178 (25%), 177 (25%) and 137 (100%).
volume). H NMR (400MHz) d: 0.89±0.92 (3H, t,
J =6.86Hz, H-10), 1.317±1.35 (4H, m, H-8 and
H-9), 1.50±1.53 (2H, m, H-7), 2.26±2.37 (2H, q,
J =6.86Hz, H-6), 3.93 (3H, s, ÐOCH3), 5.31 (1H, s,
4'-OH Ar), 6.43±6.47 (1H, d, J =16Hz, H-4), 6.80±
6.84 (1H, d, J =16Hz, H-2), 6.89±6.91 (1H, d,
J =8Hz, H-5', ortho coupled), 6.98±7.028 (1H, dt,
J =6.86 and 16Hz, H-5), 7.09 (1H, d, J =1.92Hz,
H-2', meta coupled), 7.14 (1H, dd, J =1.9 and 8Hz,
H-6', ortho-meta coupled), 7.57 (1H, d, J =16Hz,
2.5 Synthesis of ginger constituents and related
compounds
2.5.1 Dehydrozingerone
The solution of vanillin (16mmol; 2.4g) in distilled
acetone (100ml) was stirred with a magnetic stirrer,
and aqueous sodium hydroxide (100g litre 1; 70ml)
added dropwise during 30min. The stirring was
continued for another 3h to ensure completion of
Mass (relative intensity, %)
‡
‡
‡
Compound
M‡H
M‡H
H2O
M‡Na
Tropylium ion
[6]-Gingerol
[8]-Gingerol
[10]-Gingerol
295 (8)
323 (3)
277 (27)
305 (8)
317 (100)
345 (16)
373 (8)
137 (100)
137 (100)
137 (100)
137 (100)
351 (2.5)
369 (2.5)
333 (9)
[12]-Gingerol
351 (2.5)
391 (2)
Table 1. Detection of various gingerols by ES-MS
Pest Manag Sci 57:289±300 (2001)
291

M Agarwal et al
H-1). 13C NMR d: 13.99 (C-10), 22.51 (C-9), 27.93
(C-8), 31.47 (C-7), 32.79 (C-6), 56.00 (ÐOCH3),
110.22 (C-2'), 115.205 (C-5'), 122.465 (C-6'),
123.48 (C-2, C-4), 127.5 (C-5), 129.05 (C-1'),
144.009 (C-1), 147.0 (C-4'), 148.535 (C-3') and
189.7 (C-3). ES-MS m/z (relative intensity): 571
(4%), 313.2 (7%), 297 (5%), 276 (30%), 275.3
relative humidity. Every day, fresh and clean leaves
were provided in each jar after removing the excreta
and remaining leaves. The fully grown, about to
pupate, larvae were transferred to clean jars having
thick layer of sterilised soil for pupation. One week
later, emergent moths were transferred to clean jars
containing a honey solution food source and a paper
oviposition substrate. The freshly hatched larvae of the
same batch were removed and maintained separately
on fresh tender castor leaves to produce third-instar
larvae. These were then exposed separately to the test
compounds for studying their insect growth inhibition
and antifeedant activity.
‡
(100%, M‡H ), 177 (3%), 137 (2%).
2.5.3 Hydrogenation of dehydrozingerone
To the solution of dehydrozingerone (1g) in ethyl
acetate (25ml), palladised charcoal (10% Pd/C,
100mg) was added. The mixture was ®rst degassed
and then stirred at room temperature under an
atmosphere of hydrogen for 3h. The solution was
then ®ltered and concentrated under vacuum. TLC
examination of the product showed it to be a mixture
of two components which were separated by prepara-
tive TLC (methanol‡dichloromethane, 1‡99 by
volume). The upper and lower layers on the TLC
plate corresponded to zingerone and 3-hydroxy-1-
(4-hydroxy-3-methoxyphenyl)butane. Zingerone: 1H
NMR (400MHz) d: 2.12 (3H, s, ÐCOCH3), 2.70±
2.74 (2H, t, J =6.86Hz, H-2), 2.80±2.83 (2H, t,
J =6.6Hz, H-1), 3.84 (3H, s, ÐOCH3), 5.76 (1H, s,
Ar-OH, D2O exchangeable), 6.64±6.66 (1H, dd, J =2
and 8Hz, H-6'); 6.69 (1H, d, J =2Hz, H-2', meta
coupled), 6.80 (1H, d, J =8Hz, H-5', ortho coupled).
13C NMR d: 29.45 (C-4), 30.11 (C-2), 45.52 (C-1),
55.86 (ÐOCH3), 111.117 (C-2'), 114.409 (C-5'),
120.709 (C-6'), 132.88 (C-1'), 143.935 (C-4'),
146.496 (C-3'), 208.437 (C-3). ES-MS m/z (relative
intensity): 410 (2%), 409.1 (6%), 233 (3%), 217
(3%), 137 (100%).
2.6.2 Stock and test solutions
The test materials (200mg) were dissolved in a small
amount of distilled acetone, and further distilled
acetone added to make the volume up to 10ml, giving
1
a 20g litre stock solution, from which various con-
centrations (10, 7, 5, 3 and 1g litre 1) were prepared
by serial dilution with aqueous emulsi®er (5ml of
Triton-X in 1 litre of distilled water).
2.6.3 Insect growth inhibition activity
Discs (6cm diameter) were cut from the centre of
similar-sized washed and dried castor leaves which had
been collected from the same plot. These were treated
with individual concentrations of a compound and one
leaf disc each was transferred to each rearing bottle
containing a moist ®lter paper. Twenty third-instar
larvae (7 (Æ1) days old, Average weight. 30±40mg per
larva) were released separately in each jar and each
treatment was replicated three times. Insects were
exposed to test chemicals for 24h, after which normal
leaves were fed. Every day, the left-over leaf discs and
excreta of the insects were removed and fresh and
clean untreated leaves were provided in each jar. Leaf
discs treated with acetone-emulsi®ed water served as a
control. Regular counting of larval mortality, larval
abnormality, larval-pupal/pupal-adult intermediates,
pupal mortality and abnormal adults were taken until
adult emergence. Corrected growth inhibition (%) was
calculated by Abbott's formula:28
1
3-hydroxy-1-(4-hydroxy-3-methoxyphenyl)butane: H
NMR (400MHz) d: 1.21 (3H, d, J =6.3Hz, H-4),
1.70±1.75 (2H, m, H-2), 2.61±2.68 (2H, m, H-1),
3.81 (1H, m, H-3), 3.85 (3H, s, ÐOCH3), 5.66 (1H,
brs,ÐOH), 6.675±6.694 (1H, dd, J =8 and 1.93Hz
H-6'), 6.7 (1H, s, H-2'), 6.814 and 6.834 (1H, d,
J =8Hz, H-5'). 13C NMR d: 23.619 (C-4), 31.82
(C-2), 41.11 (C-1), 55.85 (ÐOCH3), 67.555 (C-3),
111.017 (C-2'), 114.309 (C-5'), 120.856 (C-6'),
133.969 (C-1'), 143.67 (C-4'), 146.45 (C-3'). ES-MS
m/z (relative intensity): 607.3 (13%), 413.2 (100%),
235 (5%), 219 (10%), 193 (12%), 179 (8%), 149.5
(15%), 137 (82%).
Corrected growth inhibition ꢀ%†
ˆ ‰ꢀT C†=ꢀ100 C†Š  100
where T =total growth inhibition in treatment and
C =total growth inhibition in control.
2.6 Insect growth regulatory and antifeedant
activity against Spilosoma obliqua Walker
2.6.1 Rearing of Spilosoma obliqua
S obliqua is a signi®cant insect pest of vegetables,
oilseeds and other crops. Its third-instar larval stage,
being more voracious, was used for bioassay work.
This stage is optimum for bioassay work, as the
subsequent fourth- and ®fth-instar stages become
more resistant to various control measures. The larvae
were collected from the ®eld and reared in the
laboratory on castor leaves (Ricinus communis L) in
glass jars (15cmÂ20cm) at 27(Æ1)°C and 60%
2.6.4 Antifeedant activity
Twenty third-instar larvae were exposed to treated leaf
discs as in Section 2.6.3. After 24h, the amount of leaf
consumed in each case was estimated using squared
graph paper. Leaf discs treated with acetone-
emulsi®ed water were used as control. The corrected
Antifeedant Index was calculated as follows:29
Antifeedant index ꢀ%† ˆ ‰ꢀC T†=ꢀC ‡ T†Š  100
Pest Manag Sci 57:289±300 (2001)
292

Biological properties of Zingiber of®cinale Roscoe isolates
where C =consumption of leaf in control and T =con-
sumption of leaf in treatment.
the fungus in the control. EC50 values were calculated
as in Section 2.6.
EC50 values (effective concentration for 50% inhi-
bition; mg ml 1) were calculated for inhibition of
growth and of feeding using a Basic LD50 programme,
version 1.1.30
3
RESULTS AND DISCUSSION
The GC-MS analysis of the steam-distilled volatile oil
of ginger rhizome showed the presence of curcumene
(Fig 1; 1) as the major constituent, which was
separated by column chromatography. Its structure
was assigned on the basis of its mass fragmentation
2.7 Antifungal activity against Rhizoctonia solani
Kuhn
‡
R solani is a soil fungus causing damping off of
seedlings, root/stem rot and stem canker in many
vegetables, ¯owers and other ®eld crops. Screening for
antifungal activity was carried out by the poisoned
food technique using potato-dextrose-agar (PDA)
medium.31 Cultures of the test fungus were main-
tained on PDA slants at 25°C, and were subcultured
in Petri dishes prior to testing. The ready-made
medium (39g) was suspended in distilled water
(1000ml) and heated to boiling until it had dissolved
completely. The medium and the Petri dishes were
autoclaved at 775.5mmHg for 30min. Stock solutions
were prepared by dissolving the test materials (130mg)
in acetone (2ml), and dilution of 1, 0.5, 0.25 and
0.125ml of this solution in 65ml of PDA medium gave
test solutions of approximately 1000, 500, 250 and
125mg litre 1, respectively. Acetone (1ml) served as
control. The medium was poured into a set of two
Petri dishes (two replications) under aseptic condi-
tions in a laminar ¯ow chamber. When the medium in
the plates was partially solidi®ed, a 5-mm thick disc of
fungus (spores and mycelium) cut from earlier sub-
cultured Petri dishes was placed at the centre of the
semi-solid medium and the lids of the dishes were
replaced. The treated and control dishes were kept in a
BOD incubator at 26 (Æ2)°C till the fungal growth in
the control dishes was almost complete (2±3 days).
The mycelial growth of fungus (mm) in both treated
(T) and control (C) Petri dishes was measured
diametrically in three different directions and the
growth inhibition (I) was calculated using the for-
mula:32
pattern, which gave a molecular ion peak at M 202,
along with other characteristic fragment ion peaks.
The bisabolene type sesquiterpene, zingiberene, which
has been reported earlier as a major constituent
obtained by a supercritical ¯uid extraction technique
using liquid carbon dioxide,5,33 could not be obtained
in suf®cient yield, possibily due to its transformation to
curcumene during steam distillation.34 Zingiberene
(Fig 1; 2) was, however, obtained from the diethyl
ether extract of fresh ginger rhizome and identi®ed by
its mass fragmentation pattern, which showed the
‡
molecular ion peak (M ) at m/z 204.
3.1 Isolation and characterization of gingerols
Ginger oleoresin, obtained from the dichloromethane
extract of fresh ginger rhizome, gave a mixture of
closely-related gingerols when subjected to column
chromatography. Preparative TLC of this mixture
yielded [6]-gingerol of satisfactory purity (85%). Its
spectral data (1H NMR, 13C NMR and ES-MS) were
in agreement with those previously reported for this
molecule.35 The compound was thus identi®ed as
5-hydroxy-1-(4-hydroxy-3-methoyxphenyl)decan-3-
one (Fig 1; 3).
3.2 Synthesis of dehydrozingerone and
[6]-dehydroshogaol
Dehydrozingerone (4) was prepared by reacting
vanillin with acetone in the presence of dilute alkali
(Fig 2). The earlier reported method36 which required
four days to complete the reaction was modi®ed to
obtain the product within 3h. The 1H NMR spectrum
of dehydrozingerone showed the presence of peaks
characteristic of COCH3 (d3.91), D2O-exchangeable
Ar-OH (d6.286), 1,3,4-trisubstituted aromatic ring
(d6.94, 7.06 and 7.07), and a,b-unsaturated methine
protons (d6.57 and 7.43). The 13C NMR spectrum
showed the presence of eleven peaks, indicating an
eleven carbon skeleton of the molecule. The structure
was further con®rmed by ES-MS, which showed a
Iꢀ%† ˆ ‰ꢀC T†=CŠ  100
The corrected inhibition (IC) was then calculated as
follows:
IC ˆ ‰ꢀI CF†=ꢀ100 CF†Š  100
where CF =[(90 Co)/Co]Â100; 90 is the diameter
(mm) of the Petri dish, and Co is the growth (mm) of
Figure 1. Structures of 1 curcumene, 2
zingiberene and 3 5-hydroxy-1-(4-
hydroxy-3-methoxyphenyl)decan-3
one.
Pest Manag Sci 57:289±300 (2001)
293

M Agarwal et al
Figure 2. Synthesis of dehydroshogaol,
zingerone and related products.
NaN(TMS)2: sodium
bis(trimethylsilyl)amide.
base peak at m/z 193 corresponding to the protonated
‡
spectrum at d 3.84 and 5.76, corresponding to
methoxy and aromatic hydroxyl (D2O exchangeable)
groups. As with dehydrozingerone, the presence of a
1,3,4-tri-substituted benzene nucleus was evident
from the presence of signals located at d 6.64, 6.69
and 6.80. The two down®eld methine protons, as
observed in dehydrozingerone, appeared up®eld in
zingerone at d 2.70 and 2.80, suggesting its hydro-
genation. In the 13C NMR spectrum, sp2 hybridized
carbons, which appeared down®eld between d 110 and
140 in dehydrozingerone, were now located up®eld
between d 30.00 and 45.00 due to hydrogenation of
the double bond. The presence of eleven peaks in the
13C NMR spectrum were in agreement with an eleven
carbon skeleton of the molecule. Its structure was
further con®rmed by ES-MS, wherein a peak appear-
ing at m/z 137 (base peak) was attributed to a tro-
molecular ion (M‡H ), and other characteristic
‡
peaks at m/z 215 and 175 corresponding to M‡Na
‡
and M‡H -H2O ion respectively.
[6]-Dehydroshogaol (5) was prepared by conden-
sing dehydrozingerone with hexanal in the presence of
sodium bis(trimethylsilyl)amide.12 The physical and
spectral data, from H NMR and 13C NMR, were in
1
accordance with the proposed structure. Its structure
was further con®rmed by ES-MS, which showed a
base peak at m/z 275.3 due to the protonated
molecular ion, along with a signature peak at m/z
137 due to a substituted tropylium ion. Other peaks at
‡
‡
m/z 571, 313, 297 corresponded to MÂ2 ‡Na ,
‡
‡
M‡K and M‡Na ions respectively. The com-
pound was thus assigned the structure 1-(4-hydroxy-
3-methoxyphenyl)dec-1,4-diene-3-one.
‡
pylium ion. Other peaks located at m/z 233 (M‡K ),
‡
3.3 Synthesis and characterization of zingerone
and 3-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-
butane (dihydrozingerone)
Hydrogenation of dehydrozingerone with 10% palla-
dised charcoal yielded a mixture of two compounds,
which were separated and puri®ed by preparative TLC
(Fig 2). The ®rst compound, with the higher Rf value,
showed two characteristic peaks in its 1H NMR
217 (M‡Na ) were also characteristic of this com-
pound. The compound was thus identi®ed as zinger-
one and was assigned the structure 1-(4-hydroxy-3-
methoxyphenyl)-3-butanone (6).
1
The H NMR spectrum of the comparatively polar
second compound was very similar to that of
zingerone, except that the singlet peak at d 2.12,
corresponding to the ÐCOCH3 proton in zingerone,
294
Pest Manag Sci 57:289±300 (2001)

Biological properties of Zingiber of®cinale Roscoe isolates
was no longer present and was replaced by a doublet at
d 1.21, suggesting the presence of a hydrogen at the
adjacent carbon. As expected, a new multiplet at d
3.81 appeared for the H-3 proton which was absent in
zingerone. The comparison of the 13C NMR spectra of
zingerone and its further hydrogenated product
revealed some minor variations. The peak located at
d 208.4 corresponding to C-3 carbonyl in zingerone
now appeared up®eld at d 67.55 as a result of its
conversion to alcohol. Since keto groups cannot be
hydrogenated with Pd/C, zingerone is ®rst converted
to its enol form, and the double bond so created is
hydrogenated to produce 3-hydroxy-1-(4-hydroxy-3-
methoxyphenyl)butane (7). Its structure was further
con®rmed by ES-MS, which exhibited a base peak at
413.2, corresponding to the possible formation of an
adduct of dihydrozingerone, zingerone (as a minor
inseparable impurity) and one sodium ion
Table 2. Effect of feeding of ginger constituents and their related products on insect growth regulatory activity against Spilosoma obliqua
Larval
pupal
Larval
pupal
Corrected
Abnormal
Total growth total growth
Concentration mortality, intermediate, pupa/adults, Normal
inhibition,
inhibition
(%)
EC50
(mg ml
1
Treatment
(%)
a (%)
b (%)
c (%)
adults (%) a‡b‡c (%)
)
Ginger oil*
1.0
0.7
0.5
0.3
0.1
25
20
20
15
10
15
15
10
20
15
20
20
15
5
40
45
55
60
70
60
55
45
40
30
52.9
9.620
9.550
Ð
47.06
35.29
29.41
17.65
5
Curcumene (1)*
1.0
0.7
0.5
0.3
0.1
20
20
15
10
5
20
10
15
20
15
20
20
15
5
40
50
55
65
70
60
50
45
35
30
52.9
41.2
35.29
23.53
17.65
10
Ginger oleoresin*
1.0
0.7
0.5
0.3
0.1
20
15
15
10
5
15
10
15
10
15
20
20
10
15
10
45
55
60
65
70
55
45
40
35
30
47.06
35.29
29.41
23.53
17.65
Gingerol*
1.0
0.7
0.5
0.3
0.1
30
25
25
15
10
15
20
15
10
15
15
10
5
40
45
55
60
70
60
55
45
40
30
52.9
9.772
5.528
3.548
5.012
2.128
47.06
35.29
29.41
17.65
15
5
Dehydrozingerone (4)**
Dehydroshogaol (5)**
1.0
0.7
0.5
0.3
0.1
20
20
15
10
5
35
30
25
20
15
10
10
10
15
20
35
40
50
55
60
65
60
50
45
40
61.11
55.56
44.44
38.89
33.33
1.0
0.7
0.5
0.3
0.1
20
15
15
10
10
40
35
40
35
25
20
25
10
5
20
25
35
50
60
80
75
65
50
40
77.78
72.22
61.11
44.44
33.33
5
Zingerone (6)
‡ dihydrozingerone
(7)**
1.0
0.7
0.5
0.3
0.1
30
25
25
15
10
35
30
25
20
15
5
10
5
30
35
45
55
60
70
65
55
45
40
66.67
61.11
50.00
38.89
33.33
10
15
Azadirachtin*
1.0
0.7
0.5
0.3
0.1
50
50
45
45
20
5
10
10
5
40
20
20
15
20
5
20
25
25
50
95
80
75
65
50
94.12
76.47
70.58
58.82
41.17
10
*Control
10
10
5
0
0
0
85
90
15
10
**Control
Pest Manag Sci 57:289±300 (2001)
295

M Agarwal et al
‡
(196‡194‡Na ), and another characteristic signa-
ture peak at m/z 137 (tropylium ion) along with other
peaks created as a result of proton and sodium
adducts/dimers etc.
derivatives have therefore been evaluated for their
insect growth regulatory (IGR) and antifeedant
activity against S obliqua and antifungal activity against
R solani. The results are presented in Tables 2 to 4.
When leaves coated with known concentrations of test
materials (10 to 1g litre 1) were fed to third-instar
larvae of S obliqua, some of the test compounds caused
signi®cant shrinkage of larvae within two days, and
these ultimately succumbed to death, while the
remainder of the test compounds caused moderate
inhibition of insect growth, the deformities mostly
being in the form of larval-pupal mortality, larval-
3.4 Insect growth regulatory, antifeedant and
antifungal activity
Except for a report of antifungal gingerenones A, B
and C and isogingerenone B,11 most of the earlier
work on the pest-control properties of ginger has been
primarily centred around crude solvent extrac-
tives.17±27 Some of the ginger constituents and their
Corrected
Concentration
1
Feeding
feeding
EC50
(mg ml
1
Treatment
(g litre
)
inhibition (%)
inhibition (%)
)
Ginger oil (steam distilled)
10
7
65
60
50
45
35
48
43
33
29
21
Ð
9.6
3.1
Ð
5
3
1
Curcumene (1)
Ginger oleoresin
Gingerol
10
7
70
60
45
40
30
54
43
29
25
18
5
3
1
10
7
90
85
70
60
50
82
74
54
43
33
5
3
1
10
7
40
35
20
10
0.0
25
21
11
5.3
0.0
5
3
1
Dehydrozingerone (4)
Dehydroshogaol (5)
10
7
30.0
20.0
10.0
0.0
17.6
11.1
5.3
Ð
5
3
0.0
1
0.0
0.0
10
7
20.0
10.0
5.0
11.1
5.3
2.6
0.0
0.0
Ð
5
3
0.0
1
0.0
Zingerone ‡
dihydrozingerone
(6‡7)
10
7
45.0
40.0
30.0
20.0
20.0
29.0
25.0
17.6
11.1
11.1
Ð
5
3
1
Azadirachtin
10
7
100.0
90.0
85.0
70.0
65.0
100.0
81.8
73.9
53.8
48.1
1.1
5
3
1
Table 3. Antifeedant activity of ginger
constituents and their related products
Control
0.0
0.0
against Spilosoma obliqua
296
Pest Manag Sci 57:289±300 (2001)

Biological properties of Zingiber of®cinale Roscoe isolates
Concentration
1
Growth in
Inhibition
(%)
Corrected
EC50
1
Treatment
(mg litre
)
treatment (cm)
inhibition * (%) (mg litre )
Ginger oil
2000
1000
500
1.3
1.5
2.1
3.0
4.0
5.2
86
84
77
67
55
42
86
84
77
67
55
42
90
97
250
125
62.5
Curcumene (1)
2000
1000
500
1.2
2.2
2.9
3.8
4.1
4.6
87
75
68
57
54
48
87
75
68
57
54
48
250
125
62.5
Ginger oleoresin
(fresh ginger)
2000
1000
500
2.6
3.5
4.3
5.4
71
61
52
40
71
61
52
40
475
210
250
Ginger oleoresin
(dry ginger)
2000
1000
500
2.1
2.6
3.1
4.4
5.2
77
71
65
51
42
77
71
65
51
42
250
125
Gingerol
2000
1000
500
2.0
2.4
3.5
4.7
5.5
78
73
61
48
39
78
73
61
48
39
261
86
250
125
Dehydrozingerone (4)
Dehydroshogaol (5)
1000
500
250
125
62.5
1.5
2.3
3.4
4.1
4.6
83
74
62
54
49
83
74
62
54
49
1000
500
250
125
2.4
3.6
5.7
6.5
73
60
36
27
73
60
36
27
383
980
Zingerone ‡
2000
1000
500
3.7
4.0
6.2
9.0
59
55
38
0.0
59
55
38
dihydrozingerone
Table 4. Antifungal activity of ginger
constituents and their related products
against Rhizoctonia solani
(6‡7)
250
0.0
* CF =0, as growth in control, was 90mm.
pupal intermediates and abnormal adults. Perusal of
the data (Table 2, Fig 3) revealed that, among the
synthetic compounds, dehydroshogaol was the most
active exhibiting 61 and 78% growth inhibition at 5
dehydroshogaol and the mixture of zingerone and
dihydrozingerone exhibited more larval-pupal inter-
mediates, while ginger oil, ginger oleoresin and
gingerol imparted more larval-pupal mortality.
1
and 10g litre respectively, (EC50, 3.5mg ml 1). It
Ginger oleoresin exhibited pronounced feeding
inhibition, causing 82 and 74% inhibition at 10 and
7g litre 1, respectively (EC50, 3.1mg ml 1). Other
test materials were however, less active (Table 3). The
activity of ginger oleoresin has been attributed to the
possible synergistic action of its various constituents.
The next in order of antifeedant action was steam-
distilled ginger oil and its major constituent curcu-
was followed in activity by the hydrogenated mixtures
of zingerone and dihydrozingerone (EC50, 5.0mg
ml 1) and dehydrozingerone (EC50, 5.5mg ml 1).
The IGR activity of these materials was, however, less
than that of azadirachtin (EC50, 2.1mg ml 1). The
activities of natural constituents like gingerol, curcu-
mene, ginger oil and ginger oleoresin were signi®cantly
lower, with EC50 values ranging from 9.6 to 9.8mg
ml 1. The results revealed that dehydrozingerone,
1
mene, which at 10g litre level caused 48 and 54%
feeding inhibition, respectively.
Pest Manag Sci 57:289±300 (2001)
297

M Agarwal et al
Figure 3. Insect growth regulatory activity of ginger constituents and their related products.
The IGR and antifeedant activity of ginger oleoresin
and its constituents such as gingerols, dehydroshogaol
and related phenolic alkenones can be accounted for
by the presence of a phenolic moiety in these com-
pounds which reportedly generates an oxygen radical
during oxidation, disruption of membrane metabolism
in the insect gut epithelium.37 Such compounds may
also affect P450-dependent ecdysone 20-monoxygen-
ase activity, causing disruption in ecdysteroid and
juvenile hormone levels and inhibition of chitin
synthesis and antioxidant enzyme system, and hence
dysfunction of the insect gut and endocrine sys-
tem.17,37 Although a large number of plant secondary
metabolites are known to affect insect growth devel-
opment and reproduction, their exact mechanisms of
action remain to be established.
inhibited in a dose-dependent manner (Table 4, Fig
4). Among the most active compounds, curcumene
1
and ginger oil at 2000mg litre showed 87 and 86%
inhibition of the test fungus (EC50 97.1 and 90.1mg
1
litre respectively). The pronounced antifungal ac-
tivity of steam-distilled ginger oil was attributed to its
major constituent, curcumene, which originated as a
result of the thermal conversion or oxidative decom-
position of zingiberene during steam distillation.5,34
Earlier reports have indicated complete control of F
moniliforme on mung seed by ginger oil at 2100mg
1
litre without affecting seed quality26 and also the
control of A ¯avus, A niger and R stolonifer in orange
and pineapple juices by aqueous extract of ginger at
1 24
.
30ml litre
The comparatively more polar ginger
oleoresin was, however, less active than ginger oil.
Furthermore, ginger oleoresin from dry ginger was
The mycelial growth of the test fungus R solani was
Figure 4. Antifungal activity of ginger constituents and their related products.
298
Pest Manag Sci 57:289±300 (2001)

Biological properties of Zingiber of®cinale Roscoe isolates
from rhizomes of Zingiber of®cinale. Phytochemistry 30:3647±
3651 (1991).
more active than ginger oleoresin from fresh ginger.
Like feeding inhibition activity, the marked antifungal
activity of ginger oleoresin can be attributed to the
synergistic action of the various constituents in the
oleoresin. Two of its constituents, gingerol (EC50
261mg litre 1) and dehydroshogaol (EC50 383mg
litre 1), showed considerable antifungal activity.
11 Endo K, Kanno E and Oshima Y, Structures of antifungal
diarylheptenones, gingerenones A, B, C and isogingerenone B,
isolated from the rhizomes of Zingiber of®cinale. Phytochemistry
29:797±799 (1990).
12 Wu TS, Wu YC, Wu PL, Chern CY, Leu YL and Chan YY,
Structure and synthesis of (n)-dehydroshogaols from Zingiber
of®cinale. Phytochemistry 48:889±891 (1998).
Among synthetic constituents, dehydrozingerone
1
13 Kikuzaki H and Nakatani N, Cyclic diarylheptanoids frrom
rhizomes of Zingiber of®cinale. Phytochemistry 43:273±277
(1996).
showed 83% inhibition at 1000mg litre
(EC50
86mg litre 1). A mixture of zingerone and dihydro-
zingerone was however, less active (EC50 980mg
litre 1). The diminished activity of hydrogenated
products may be suggestive of the signi®cance of
unsaturation in retaining antifungal activity.
14 Denyer CV, Jackson P and Loakes DM, Isolation of antirhino-
viral sesquiterpenes from ginger (Zingiber of®cinale). J Nat Prod
57:658±662 (1994).
15 Katiyar SK, Agarwal R and Mukhtar H, Inhibition of tumor
promotion in sencar mouse skin by ethanol extract of Zingiber
of®cinale rhizome. Cancer Res Baltimore 56:1023±1030 (1996).
16 Kikuzaki H and Nakatani N, Antioxidant effect of some ginger
constituents. J Food Sci 58:1407±1410 (1993).
Though ginger constituents and their derivatives
themselves possess only modest biological activity,
they may be useful as lead products for quantitative
structure-activity relationships studies. Derivatization
of the hydroxyl group, substitution in the aromatic
nucleus and modi®cations in the alkyl/alkenyl side-
chain may result in compounds with profound pest-
control properties.
17 Sahayaraj K, Antifeedant Effect of some plant extracts on the
Asian armyworm, Spodoptera litura (Fabricius). Current Sci
74:523±525 (1998).
18 Sahayaraj K and Sekar R, Ef®cacy of plant extracts against
tobacco caterpillar larvae in groundnut. Int Arachis Newslett
16:38 (1996).
19 Echendu TNC, Ginger, cashew and neem as surface protectants
of cowpeas against infestations and damage by Callosobruchus
maculatus (Fab). Trop Sci 31:209±211 (1991).
ACKNOWLEDGEMENTS
20 Ofuya TI and Okuku IR, Insecticidal effect of some plant extract
on the cowpea aphid Aphis craccivora Koch (Homoptera:
Aphididae). Anzeiger-fuer-schaedlingakunde-p¯anzenschutz-um-
weltschutz (Germany) 67:127±129 (1994).
The authors thank the University Grants Commission
(UGC), New Delhi for the award of a scholarship to M
Agarwal, and the Head of the Division of Agricultural
Chemicals, IARI, New Delhi for providing necessary
facilities. IACR-Rothamsted receives grant-aided sup-
port from the Biotechnology and Biological Sciences
Research Council of the United Kingdom.
21 Singh UP, Srivastava BP, Singh KP and Mishra GD, Control of
powdery mildew of pea by ginger extract. Indian Phytopath
44:55±59 (1991).
22 Maroon CJM, Opina OS and Molina AB, Evaluation of crude
plant extracts as fungicides for the control of Cercospora
leafspot of mungbean (Vigna radiata (L) Wojizeckj). Philippine
Phytopath 20:27±38 (1995).
23 Zaman MA, Saleh AKM, Rahman GMM and Islam MT, Seed-
borne fungi of mustard and their control with indigenous plant
extracts. Bangladesh J Plant Pathol 13:28±29 (1997).
24 Akpomedaye DE and Ejechi BO, The hurdle effect of mild heat
and two tropical spice extracts on the growth of three fungi in
fruit juices. Food Res Internat 31:339±341 (1998).
25 Khilare VC and Gangawane LV, Application of medicinal plant
extracts in the management of thiophanate methyl resistant
Penicillium digitatum causing green mold of Mosambi. J Mycol
Pl Path 27:134±137 (1997).
REFERENCES
1 Wing KD and Aller HE, Ecdysteroid agonists as novel insect
growth regulators, in Pesticides and Alternatives: Innovative
Chemical and Biological Approaches to Pest Control, ed by Casida
JE, Elsevier, Amsterdam, pp 251±257 (1990).
2 Sukh Dev and Koul O, Insecticides of Natural Origin, Harwood
Academic Publishers, Amsterdam, The Netherlands, 365 pp
(1997).
3 Ishaaya I and Horowitz AR, Insecticides with novel modes of
action: an overview, in Insecticides with Novel Modes of Action,
ed by Ishaaya I and Degheele D, Narosa Publishing House,
New Delhi, pp 1±24 (1998).
26 Mishra D, Seed protectant property of essential oil of Zingiber
of®cinale Roscoe. Indian Perfumer 34:266±268 (1990).
27 Ahmad FBH, Mackeen MM, Ali AM, Mashirun SR and Yaacob
MM, Repellency of essential oils against the domiciliary
cockroach, Periplaneta americana. Insect Sci Applic 16:391±
393 (1995).
4 AnonymousWealth of India. A dictionary of Indian raw materials
and industrial products, Vol XI: X-Z and cumulative indexes,
PID, CSIR, New Delhi, pp 89±105 (1969).
5 Bartley JP and Foley P, Supercritical ¯uid extraction of
Australian-grown Ginger (Zingiber of®cinale). J Sci Food Agric
66:365±371 (1994).
28 Abbott WS, A method of computing the effectiveness of an
insecticide. J Econ Entomol 18:265±267 (1925).
29 Blaney WM, Simmonds MSJ, Ley SV, Anderson JC, Smith SC
and Wood A, Effect of azadirachtin-derived decalin (per-
hydronaphthalene) and dihydrofuranacetal (furo[2,3-b]pyran)
fragments on the feeding behaviour of Spodoptera littoralis.
Pestic Sci 40:169±173 (1994).
6 Connel DW, The chemistry of the essential oil and oleoresin of
ginger (Zingiber of®cinale Roscoe). The Flavour Industry 1:677±
693 (1970).
7 Connell DW and Sutherland MD, A re-examination of gingerol,
shogaol, and zingerone, the pungent principles of ginger
(Zingiber of®cinale Roscoe). Aust J Chem 22:1033±1043 (1969).
8 Connell DW, Natural pungent compounds III. The paradols and
associated compounds. Aust J Chem 23:369±376 (1970).
9 Kikuzaki H, Tsai SM and Nakatani N, Gingerdiol related
compounds from the rhizomes of Zingiber of®cinale. Phyto-
chemistry 31:1783±1786 (1992).
30 Trevors JT, A basic programme for estimating LD
50 values using
the IBM-PC. Bull Environ Contam Toxicol 37:18±26 (1986).
31 Nene YL and Thapliyal PN, Evaluation of fungicides, in Fungi-
cides in Plant Disease Control, ed by Nene YL and Thapliyal PN,
Oxford and IBH Publishing Co, New Delhi, pp 406±428
(1979).
10 Kikuzaki H, Kobayashi M and Nakatani N, Diarylheptanoids
32 Sztejnberg A, Azaizia H and Chet I, The possible role of phenolic
Pest Manag Sci 57:289±300 (2001)
299

M Agarwal et al
compounds in resistance of horticulture crops to Dematophora
36 Denniff P, Mcleod I and Whiting DP, Synthesis of the (I)
-[n]-gingerols (Pungent principles of ginger) and related com-
pounds through regioselective aldol condensation. I. Relative
pungency assays. J Chem Soc, Perkin Trans 1, 82±87 (1981).
37 Koul O, Insect growth inhibition and regulation by allelochemi-
cals: biochemical basis and role in integrated pest manage-
ment, in Ecological Agriculture and Sustainable Development, vol
2, ed by Dhalwal GS, Randhawa NS, Arora R and Dhawan
AK, Indian Ecological Society and Centre for Research in
Rural and Industrial Development, India, pp 93±110 (1998).
necatrix Hartig. Phytopath Z 107:318±326 (1983).
33 Chen CC and Ho CT, Gas chromatographic analysis of volatile
components of ginger oil (Z of®cinale Roscoe) extracted with
liquid carbon dioxide. J Ag Food Chem 36:322±328 (1988).
34 Connell DW and Jordan RA, Composition and distinctive
volatile ¯avour characteristics of the essential oil of Australian
grown ginger (Zingiber of®cinale). J Sci Food Agric 22:93±95
(1971).
35 Bartley JP, A new method for the determination of pungent
compounds in ginger (Zingiber of®cinale). J Sci Food Agric
68:215±222 (1995).
300
Pest Manag Sci 57:289±300 (2001)