®

Medicinal and Aromatic Plant Science and Biotechnology ©2009 Global Science Books

Chemical Composition and Insecticidal Activity

of Volatile Oil of Khaya grandifoliola
Abiodun Falodun1* • Rizwana Siraj2 • Muhammad Irfan Qadir2 •
Sheraz Ahmad Khan Tanoli2 • Muhammad Iqbal Choudhary2
1 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Benin, Benin City, Nigeria
2 H.E.J. Research Institute of Chemistry, International Center for Chemical Sciences, University of Karachi, Karachi 75270, Pakistan

Corresponding author: * [email protected], [email protected]

ABSTRACT
Khaya grandifoliola has been used in traditional medicine against infections, rheumatic and parasitic diseases. The essential oil (EO) from
the stem bark of K. grandifoliola was extracted with n-hexane, analysed by GC and GC-MS. The insecticidal activity of the EO was
evaluated against two insects, Rhyzopertha dominica and Tribolium castaneum using an established standard procedure. The chemical
composition of the EO obtained through GC-MS revealed 24 components. The major components were -pinene (10.56%), limonene
(1.25%), -carophyllene (3.87%), -pinene (7.80%), -phellandrene (7.45%), and citronellol (5.10%). The EO exhibited significant
insecticidal activity against Rhyzopertha dominica and Tribolium castaneum (85 and 80%, respectively).
_____________________________________________________________________________________________________________

Keywords: essential oil, plant extract, GC-MS, Rhyzopertha dominica, Tribolium castaneum

INTRODUCTION perties have been reported (Agbedahuns et al. 1998; Bumah

1998; Bickii et al. 2000; Makinde et al. 2006; Njifutie and
The use of plant products in the treatment of diseases and Njikam 2006). Some of the chemical constituents reported
infections is as old as man. About 80% of the world popu- include limonoids (Zhan et al. 2008). Recently, the effect of
lation relies on plants and plant extracts for their healthcare stem bark of the plant was evaluated against methicillin-
needs. A recent report indicates that of the top 150 drugs resistant Staphylococcus aereus and was observed to be
used in the USA 57% of them contain at least one major very effective against different strains of the organisms at
active compound derived from natural products (Rosenthal the tested doses (Uzoekwe et al. 2008).
1997; Joseph et al. 2008). Not only do higher plants con- Higher plants are a rich source of novel natural com-
tinue to serve as important sources of new drugs, but phyto- pounds that can be of great usefulness in developing envi-
chemicals derived from them are also extremely useful as ronmental safe methods for insect control (Amason et al.
lead structures for synthetic modification and optimization 1989). Insecticidal activity of many plants against several
of bioactivities. insect pests has been demonstrated (Isman 2000; Carlini
Tropical rainforests provide many medicinal plants with 2002).
potential medicinal properties but the ethno and phyto- Tribolium castaneum (Herbst) is considered as a major
pharmaceuticals of these medicinal plants have not been pest of stored grains (Howe 1965). Annual post harvest
fully exploited (Baladrin et al. 1993; Meena 2009). The pot- losses resulting from insect damages, microbial deteriora-
ential medicinal importance is due not only to the species tion are estimated to be 10-25% of worldwide production
richness of the tropical flora, but also to the diversity of (Adams 2001). The lesser grain borer is characterized as
pathogens, parasites and herbivores against which the plants both an internal and external feeder, considered to be a seri-
must provide a defensive mechanism. Many of the defense ous pest of both whole kernel stored grain and cereal pro-
chemicals secreted have been found to be useful in the ducts. The adults and larvae bore into undamaged kernels of
treatment and prevention of diseases affecting humans and grain, reducing them to hollow husks. They are also able to
other organisms as a result of their metabolic precision survive and develop in the accumulated “flour” produced as
(Farnsworth 1988). Essential oils (EOs) are made up of dif- the seeds are chewed up. The lesser grain borer is primarily
ferent volatile components and have been shown to possess a pest in stored wheat and corn, but it can infest tobacco,
alot of biological activities (Ahmet et al. 2005). nuts, beans, bird seed, biscuits, cassava, cocoa beans, dried
Khaya grandifoliola, family Meliaceae, commonly fruit, peanuts, spices, rodenticide baits, and dried meat and
called African mahogany (Hutchinson and Dalziel 1978), is fish (Koehler and Pereira 1994).
widely distributed across West Africa from the Guinea With a rising concern for environmental safety there has
coast to Cameroon and extending eastward through Congo been renewed interest in the use of naturally occurring in-
Basin to Uganda and some parts of Sudan. It grows up to 40 secticides including medicinal plant products (Isman 2006).
m high and 5 m in girth. The bark is grey in colour and Therefore the aim of this study was to determine the
exudes a bitter gum when wounded. The aqueous decoction chemical composition of the EO isolated from the stem of K.
of the plant is bitter and used to treat many diseases. grandifoliola by GC-MS analysis and to evaluate the insec-
Khaya spp. are a valuable indigenous traditional medi- ticidal activity of its EO against known insects causing
cine in West Africa. Its bitter bark is used for various ethno- severe damage to plants. To the best of our knowledge the
medicinal purposes such as fever, lumbago, cough, rheuma- separation and identification of the volatile components of
tism, stomach ache, gastric pains and diarrhoea in horses this plant using GC-MS and chemical methods have not
and camels (Dalziel 1994). Its antimalarial, antiulcer, anti- been previously examined.
anaemic and hypoglyceamic and hypocholesterolaemic pro-

Received: 16 June, 2009. Accepted: 1 December, 2009.

Short Communication
 Medicinal and Aromatic Plant Science and Biotechnology 3 (Special Issue 1), 61-63 ©2009 Global Science Books

MATERIALS AND METHODS test was carried out in which several doses were chosen among
those having no killing effect on the experiment population to a
Collection and identification of plant material minimal of 100% death of this population. Using a micropipette
(Rainin magnetic assist), a precise volume of EO was added to
Fresh stem bark of K. grandifoliola was collected from a local ethanol (negative control) and diluted to 5 ml. From this 0.5 ml of
government area in Benue State, Nigeria around April, 2008. The solution was uniformly applied to a 9 cm disk of filter paper
plant was identified by Mr. Sunny A of the Department of Pharma- (Whatman No.1) and placed in a Petri dish. Twenty adult insects,
cognosy, Faculty of Pharmacy, University of Benin, Benin City, less than 1 month old, were introduced into the dish 5 min later
Nigeria. and the dish was covered. A control with ethanol alone was used.
For each preparation five replications were made. The number of
Extraction and isolation of EO dead insects was determined 24 h after application.
The percentage mortality was calculated by:
The plant was cut into smaller pieces, dried and reduced to a fine
powder with the aid of a mechanical grinder. The powdered sam- § No. of insects alive in test ·
Mortality (%) ¨ ¸ u 100
ple (200 g) was extracted with n-hexane by maceration for 48 h at © No. of insects alive in control ¹
room temperature. The extract was evaporated to dryness using a
rotary evaporator at reduced pressure. The EO was passed over Data analysis
anhydrous Na2SO4 to remove moisture. The fraction obtained was
stored in a refrigerator at -4°C in the dark. Statistical analyses were performed using SAS statistical package
1994. Users guide, Version 8.2. SAS Institute Inc., Cary, NC, USA.
Identification of chemical components Data are expressed as means ± standard error (SEM). Comparison
was done using one-way Analysis of Variance. The value of 0.05
Identification of the chemical constituents was done by a computer level of probability was chosen as a criterion for significance.
library MS searches using retention indices as a pre-selection rou-
tine and visual inspection of mass spectra from the literature for RESULTS AND DISCUSSION
confirmation (British Pharmacopoeia 1980; Julian and Konig
1988; Adams 1989, 2001). The EO obtained from the stem bark of K. grandifoliola has
a light yellow colour and fragrant aroma. The EO yield is
Gas chromatography 1.56% of dry plant material. The chemical components pre-
sented in Table 1 were identified unambiguously by direct
Quantitative and qualitative data were determined by GC and GC- comparison (mass fragmentation and retention index) with
MS respectively. The EO was injected onto a Shimadzu GC-17A reported data as well as a computer library search. The com-
system, equipped with an AOC-20i autosampler and a split/split- ponents identified were mainly oxygenated monoterpenes
less injector. The column used was a DB-5 (Optima-5), 30 m, 0.25 and sesquiterpenes hydrocarbons. The sequiterpene hydro-
mm i.d., 0.25 m df, coated with 5% diphenyl - 95% polydi- carbon accounted for 51.35% of the total while the oxy-
methylsiloxane, operated with the following oven temperature genated components were 19.47%. The remaining com-
programme: 50°C, held for 1 min, rising at 3°C/min to 250°C, pounds were fatty acids. As shown in Table 1, 24 com-
held for 5 min, rising at 2°C/min to 280°C, held for 3 min; injec- pounds were identified in the EO.
tion temperature and volume, 250°C and 1.0 l, respectively; The EO also showed significant insecticidal activity
injection mode, split; split ratio, 30: 1; carrier gas, nitrogen at 30 (Table 2) against Rhyzopertha dominica and Tribolium cas-
cm/s linear velocity and inlet pressure 99.8 KPa; detector tempera- taneum with 80 and 85% activity, respectively.
ture, 280°C; hydrogen, flow rate, 50 ml/min; air flow rate, 400 The insecticidal activity observed is likely due to the
ml/min; make-up (H2/air), flow rate, 50 ml/min; sampling rate, 40 relatively high concentrations of limonene in the EO and
ms. Data were acquired by means of GC solution software (Shi- also partly due to the presence of -pinene, -pinene and -
madzu). caryophyllene in the EO. The synergistic activity of the div-

Gas chromatography-mass spectrometry

Table 1 Chemical composition (%) of EO of K. grandifoliola.
Agilent 6890N GC was interfaced with a VG Analytical 70-250s No RI* Compounds Relative %
double - focusing Mass spectrometer. Helium was used as the 1 923 Thujene 2.56
carrier gas. The MS operating conditions were: Ionization voltage 2 945 Camphene 2.67
70eV, ion source 250°C. The GC was fitted with a 30 m × 0.32 3 970 Į- Pinene 10.56
mm fused capillary silica column coated with DB-5. The GC ope- 4 977 ȕ- Pinene 7.80
rating parameters were identical with those of the GC analysis. 5 984 ȕ-Myrecene 3.26
6 995 -Phellandrene 7.45
Insects 7 1005 3-Carene 3.45
8 1012 p-cymene 2.42
Insects (Tribolium castaneum, Rhyzopertha dominica) used for the 9 1019 Limonene 1.25
test were reared in glass jars at 27 ± 1°C and 60 ± 5% relative 10 1049 Terpene 3.57
humidity (RH) in the laboratory of the H.E.J Research Institute of 11 1464 ȕ- Carophyllene 3.87
Chemistry in Karachi, Pakistan. 12 1467 Carophyllene oxide 1.34
13 1130 Camphor trace
Insecticidal activity 14 1161 ȕ-Terpinol-4-ol 0.80
15 1172 - Terpineol 4.56
To test insectividal activit, test insects’ volatile organic solvent 16 1212 Citronellol 5.10
(ethanol), standard insecticide (Permethrin), Petri dishes (9 cm 17 1260 - Terpenylacetate 1.35
diameter), micropipette (1000 μl), growth chamber, test sample, 18 1377 Isoledene 0.84
filter paper, glass vials and a brush are required. 19 1389 ȕ –cubebene trace
20 1477 Naphthalene 4.52
Test: The EO of K. grandifoliola was screened for insecticidal pro- 21 1565 ȕ-Carophyllene oxide 5.54
perty against R. dominica and T. castaneum in comparison with 22 1787 Tetradecanoic acid 2.39
commercial insecticide Permethrin and ethanol (negative control). 23 1821 1,8-cineole 0.78
Insecticidal activity was determined using established standard 24 1963 Hexadecanoic acid 12.32
experimental procedures (Atta-ur-Rahman et al. 1997). Total 88.40
In evaluating the insecticidal activity of the EO, a preliminary *Retention indices of DB-5 fused column

62
 Analysis of essential oil of K. grandifoliola. Falodun et al.

Table 2 Insecticidal activity of volatile oil of K grandifoliola. Bumah VV, Essien EU, Agbedahunsi JM, Ekah OU (1988) Effects of Khaya
Treatments Conc. (µg/cm3 ) Effect of sample (%) grandifoliola (Meliaceae) on some biochemical parameters in rats. Journal of
EO of kg a 1019.1 85 ± 1.34 (T.C b) Ethnopharmacology 102, 446 -449
a Carlini CR, Grossi-de Sáa MF (2002) Plant toxic proteins: A review on their
EO of kg 1019.1 80 ± 1.00 (R D c)
potentialities as bioinsecticides. Toxiconomy 40, 1515-1539
Permethrin (positive control) 237.5 100 ± 0.00 Dalziel JM (1994) The useful Plants of West Tropical Africa, Crown Agents for
Ethanol (negative control) - the Colonies, London, 612 pp
a
Khaya grandifoliola Deba F, Xuan TD, Yasuda M, Tawata S (2008) Chemical composition and
b
Tribolium castaneum
c Antioxidant, antibacterial and antifungal activities of essential oils from
Rhyzopertha dominica
Bidens pilosa Linn, var. Radiata. Food Control 19, 346-352
Farnsworth NR (1988) Screening plants for new medicines. In: Wilson EO,
Peter FM (Eds) Biodiversity, National Academy Press, Washington DC, pp
ersity of major and minor chemical constituents present in 83-96
Filopiwicz N, Karminski M, Kurlenda J, Asztemborska M (2003) Antibacte-
the EO is important in considering the desired biological rial and antifungal activity of Juniper berry oil and its selected components.
activity. Phytotherapy Research 17, 227-231
The insecticidal activity of the EO is dependent on the Griffo F, Rosenthal J (1997) Biodiversity and Human Health, Island Press,
active chemical constituents and the gross sensitivity of the Washington DC, pp 7-38
target insects to the active chemical principle (Obeng et al. Howe RW (1965) Losses caused by insects and mites in stored foods and food-
1997; Rachid et al. 2006). The EO of K. grandifoliola con- stuffs. Nutrition Abstracts and Review 35, 285-302
tains limonene. Limonene is an active insecticidal chemical Hutchinson J, Dalziel JM (1978) Flora of West Tropical Africa (Part 1, 2nd
constituent, which has been reported to have toxic effect on Edn), Crown Agents for Overseas Government and Administration, London,
Coleopterans (Tanpojou et al. 2002). Enatiomers of -pinene pp 345-520
Isman MB (2000) Plant essential oils for pest and disease management. Crop
and -pinene and limonene have been reported to have Protection 19, 603-608
potent antibacterial and insecticidal activity (Megiatis 1999; Isman MB (2006) Botanical Insecticides, deterents and repellents in modern
Tzakou et al. 2001; Filopiwicz 2003). - and -Pinene are agriculture and an increasing regulated world wide. Annual Review of Ento-
able to destroy cellular integrity, and thereby inhibit respira- mology 51, 45-56
tion and ion transport processes. The antimicrobial activity Joseph NO, Kennedy MMH, Herbert VL, Rogasian LAM (2008) Multiplant
of caryophyllene and caryophyllene oxide has been de- or single plant extracts, which is the most effective for local healing in Tanza-
monstrated (Ulubelen 1994; Azaz 2002; Deba et al. 2008) nia? African Journal of Traditional, Complimentary and Alternative Medi-
although insecticidal action is yet to be determined. cines 5, 165-172
Julain D, Konig WA (1988) The Atlas of Spectral Data of Sesquiterpene
ACKNOWLEDGEMENTS Hydrocarbons, Verlag, Hamburg, pp 8-20
Koehler PG, Pereir RM (1994) Lesser Grain Borer, Rhyzopertha dominica
The corresponding author is grateful to Third World Academy of (Coleoptera, Bostrichidae) Entomology and Nematology Department, Coope-
rative Extension Service, Institute of Food and Agricultural Sciences, Uni-
Sciences (TWAS) and H.E.J Research Institute of Chemistry, versity of Florida, Gainesville, pp 1-2
University of Karachi, Pakistan, for the Postdoctoral Fellowship Makinde JM, Awe SO, Agbedahunsi JM (2006) Effect of Khaya grandifoliola
Award. Mr. Simon Peters is also acknowledged for the collection extract on Plasmodium berghei in mice. Phytotherapy Research 2, 30-32
and authentication of plant materials. Meena M (2009) Non-conventional sources, antioxidative properties, and novel
medicinal use of plant derivatives. Journal of Medicinal Plants Research 3, 8
REFERENCES Megiatis P, Melliou E, Skaltsounis AL, Chinou IB, Mitaku S (1999) Che-
mical composition and antimicrobial activity of the essential oils of Pistacia
Adams RP (1989) Identification of Essential oils by Ion Trap Mass Spectros- lentiscus var. Chia. Planta Medica 65, 749-752
copy, Academic Press, New York, 302 pp Njifutie N, Njikam N (2006) Curative dose of K. grandifoliola stem bark for
Adams RP (2001) Identification of Essential Oils by Gas Chromatography the treatment of gastric ulcers using Wistar rats. Pharmaceutical Biology 44,
Quadrupole Mass Spectrometry, Allured Publishing Corp., Carol Stream, 152-155
USA, 1126 pp Obeng OD, Reichmuth C, Bekele J, Hassanali A (1997) Biological activity of
Agbedahunsi JM, Elujoba AA, Makinde JM, Oduda AJM (1998) Antimala- 1,8 cineole, a major component of essential oil of Ocimim kenyense (Ayoban-
rial Activity of Khaya grandifoliola stem bark. Pharmaceutical Biology 36, gira) against stored product beetles. Journal of Applied Entomology 121,
8-12 237-243
Ahmet C, Saban K, Hamdullah K, Ercan K (2005) Antifungal properties of Rachid J, Abdeslam E, Fouad S (2006) Insecticidal activity of four medicinal
essential oil and crude extracts of Hypericum linariodes Boss. Biochemical plant extracts against Tribolium castaneum (Herbst) (Coleoptera: Tenebrioni-
Systematic Ecology 33, 245-256 dae). African Journal of Biotechnology 5, 936-940
Amason JT, Philogne BJR, Morand P (1989) Insecticides of Plant Origin, Stephen UA, Abiodun F, Osahon O, Ewaen E (2009) Phytochemical analysis
American Chemical Society Symposium Series (Vol 387), Washington D.C., and antibacterial activity of Khaya grandifoliola stem bark. Journal of Biolo-
pp 136-149 gical Science 9, 63-67
Atta-ur-Rahman, Choudhary MI, William JT (1997) Bioassay Techniques Tanpojou LA, Adler C, Bouda H, Fountern DA (2002) Efficacy of powder
for Drug Development, Harvard Academic Publisher, pp 67-68 and essential oil from Chenopodium ambrosioides leaves as post-harvest
Azaz D, Dermeci F, Satil F, Kurkcouglu M, Baser KH (2002) Antimicrobial grain protectants against six-stored product beetles. Journal of Stored Pro-
activity of Satureja essential oils. Zeitschrift für Naturforschung 57c, 817- ducts Research 38, 395-402
821 Tzakou O, Pitarokili D, Chinou IB, Harvala C (2001) Composition and anti-
Baladrin MF, Kinghorn AD, Farnsworth NR (1993) Plants derived natural microbial activity of the essential oil of Salvia ringens. Planta Medica 67,
products in drug discovery and development. In: Baladrin MF (Ed) Human 81-83
Medicinal Agents from Plants, Symposium Series 534, American Chemical Ulubelen A, Topcu G, Eris C, Sonmez U, Kartal M, Kurucu S, Bozok
Society, Washington DC, pp 2-12 Johasson C (1994) Diterpenoids from the roots of Salvia hypargeia.
Bickii JJA, Njifutie LK, Basco P, Ringwald J (2000) In-vitro antimalarial Phytochemistry 36, 971-974
activity of limonoids from Khaya grandifoliola C D.C (Meliaceae). Journal Zhang H, Odeku OA, Wang XN, Yue JM (2008) Limonoids from the stem
of Ethnopharmacology 69, 27 -33 bark of Khaya grandifoliola. Phytohemistry 69, 271-275

63