International Journal of Spectroscopy
Volume 2015 (2015), Article ID 901386, 6 pages
http://dx.doi.org/10.1155/2015/901386

Research Article

Secondary Metabolites of the Cuticular Abdominal Glands of Variegated Grasshopper (Zonocerus variegatus L.)

O. U. Igwe and D. E. Udofia

Department of Chemistry, Michael Okpara University of Agriculture, Umudike, P.M.B. 7267, Umuahia, Abia State, Nigeria

Received 2 June 2015; Accepted 29 July 2015

Academic Editor: Hakan Arslan

Copyright © 2015 O. U. Igwe and D. E. Udofia. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract.

Chemical compounds were extracted with petroleum ether from the cuticular abdominal glands of grasshopper (Zonocerus variegatus L.) and eleven compounds were characterised using Gas Chromatography/Mass Spectrometry (GC/MS) technique in combination with Fourier Transform-Infrared Spectroscopy (FT-IR). The compounds analysed were 2,7-dimethyloctane (3.21%), decane (5.33%), undecane (3.81%), tridecanoic acid methyl ester (4.76%), hexadecanoic acid (9.37%), 11-octadecenoic acid methyl ester (23.18%), pentadecanoic acid, 14-methyl-methyl ester (4.43%), (Z)-13-docosenoic acid (10.71%), dodecyl pentafluoropropionate (9.52%), 2-dodecyl-1,3-propanediol (6.38%), and 1,12-tridecadiene (19.30%). FT-IR analysis of the extract showed peaks at 1270.17 (C–O and C–F), 1641.48 (C=C), 2937.68 (C–H), and 3430.51 (O–H) cm⁻¹ indicating the presence of ether, alkene, alkane, alcohol, carboxylic acid, and fluoric compounds. These compounds consisted of 32.37% ester, 31.65% hydrocarbons, 20.08% fatty acid, 9.52% halogenated ester, and 6.38% alcohol. The highest component was 11-octadecenoic acid methyl ester followed by 1,12-tridecadiene. Since behavioural bioassays were not carried out, the consideration of these compounds to be pheromone semiochemicals remains a hypothesis.

1. Introduction

Chemicals play an important role in communication between insects. Chemicals that mediate interactions between organisms (inter- or intraspecific) are called semiochemicals [1]. The African grasshopper, Zonocerus variegatus (L.) is a tropical insect that belongs to the order Orthoptera and family Pyrgomorphidae. In Nigeria, it usually occurs on uncultivated land with the nymphs and adult stage sharing the same habitat which extends from rain forest zone to the Guinea Savannah in the north [2, 3]. Z. variegatus is a polyphagous insect that causes serious damage to both food and cash crops in West Africa [3–5]. In addition to wild plants it has been recorded as feeding on, and in most cases causing damage to, banana, cassava, Citrus, cocoa, coffee, cola, cotton, cowpea, maize, oil palm, okra, pawpaw, pepper, pineapple, plantain, soybean, teak, rice, various vegetables, and other cash crops such as melon. In southern Nigeria, the staple food, cassava (Manihot esculenta Crantz), is the major crop damaged [6]. Reports from West African countries invariably name this species as one of the major pests against which control measures (generally chemical insecticides) have been applied [7–9]. In 1970, Nigeria declared Z. variegatus a major pest, and subsequently it became a problem in Cote d’Ivoire, Ghana, Congo, Benin, Uganda, Senegal, and Burkina Faso [10].

Chemical insecticides cause or increase environmental pollution and constitute immediate or long-term health hazards. They are danger to human communities, domestic animals, wildlife and plants and they chronically disrupt food-chains. Insecticides eliminate natural enemies which might otherwise keep pest populations within manageable limits [10, 11]. Spraying chemicals on Z. variegatus is only effective in the short-term; hence alternative methods for use by farmers are necessary. It has been reported that both sexes and all instars of the variegated grasshopper, Z. variegatus, when molested, expel an odorous, milky secretions from a gland opening on the dorsal intersegment membrane between the first and second abdominal tergites [12, 13]. It has also been reported that, in studies involving the initiation of egg laying of Z. variegatus and the subsequent maintenance of the integrity of the egg laying sites, it was found that an odour given off by freshly laid pods plays a major role in attracting further mature males and females into the sites [12]. If this odour can be isolated and synthesised it could be used for attracting mature insects to simple traps baited with insecticide [14]. This idea forms the basis of this current research which attempts to extract and identify chemical constituents present in the abdominal glands of Z. variegatus in the hope that the synthetic forms of these compounds can be used to mass-trap the insects to locations baited with insecticides.

2. Materials and Methods

2.1. Insect Collection

Colonies of Z. variegatus were collected from uncultivated farmland in Michael Okpara University of Agriculture, Umudike, Abia State, Nigeria, and housed in a wire cage (20 × 20 × 30 cm). The organism was identified and authenticated in the Zoology Department of the aforementioned university. About 85 adults of Z. variegatus were used for the investigation and they were maintained on fresh leaves of cassava (M. esculenta) until they were sacrificed for the analysis. Hereafter, the word “grasshopper” refers only to the adults of Z. variegatus unless otherwise stated.

2.2. Extraction of Chemical Constituents

Cuticular abdominal glands of Z. variegatus were excised with fine brand new razor blade after anaesthetising the organisms by cleaning with chloroform which also removed cuticular surface contaminants. Cleaning the insects with chloroform was done with the aid of a piece of foam (a polymeric material) sprinkled with a little amount of chloroform. The tissue was then extracted in petroleum ether for 20 min at room temperature. Extract was placed in screw cap vials and stored at −15°C until analysis.

2.3. Gas Chromatography/Mass Spectrometry (GC/MS) Analysis

GC analysis was carried out in SHIMADZU JAPAN Gas Chromatography 5890-11 with a fused GC column (OV-101) coated with polymethyl silicon (0.25 mm 50 m) and the conditions were as follows: temperature programming from 80–280°C held at 80°C for 1 min, at 200°C for 4 min (rate 10°C/min), and finally at 280°C for 5 min (rate 10°C/min). The injection temperature was 250°C. GC/MS analysis was conducted using GCMS-QP 2010 Plus (Shimadzu, Japan) with column oven temperature of 80°C. The carrier gas was helium with a pressure of 108.2 Kpa and linear velocity of 46.3 cm/s. Total flow was 6.2 mL/min, column flow was 1.58 mL/min, injection mode was split, flow control mode was linear velocity, purge flow was 3.0 mL/min, and split ratio was 1.0. Also, ion source temperature was 230°C, interface temperature was 250°C, solvent cut time was 2.5 min, detector gain was 0.00 KV, detector gain mode was relative, and the threshold was 1000. For the mass spectrometer, start time was 3.0 min, end time was 28.0 min, event time was 0.5 s, scan speed was 1250, and start was 40 while end was 600. The mass spectrometer was also equipped with a computer fed mass spectra data bank. Hermle Z 233 M-Z centrifuge, Germany, was used. All solvents used were of analytical grade and were procured from Merck, Germany.

2.4. Components Identification

The components of the extract were identified by matching the peaks with computer Wiley MS libraries and confirmed by comparing mass spectra of the peaks and those from literature as well as using the database of National Institute of Standards and Technology (NIST) [15].

2.5. FT-IR Analysis

FT-IR measurement of the extract was performed using FTIR-8400S Fourier Transform Infrared Spectrophotometer, Shimadzu, Japan, in a diffused reflectance mode at a resolution of 4 cm⁻¹ in sodium chloride (NaCl) pellets in the range 4500–400 cm⁻¹.

3. Results and Discussion

The chemical constituents of the abdominal glands of Z. variegatus were investigated using GC/MS technique and eleven compounds were characterised as shown by the chromatogram in Figure 1. The compositions of the compounds were 32.37% ester, 31.65% hydrocarbons, 20.08% fatty acid, 9.52% halogenated ester, and 6.38% alcohol. The highest component was 11-octadecenoic acid methyl ester followed by 1,12-tridecadiene. The FT-IR spectra of the extract from variegated grasshopper are shown in Figure 2. FT-IR analysis of the extract showed peaks at 1270.17 (C−O and C−F), 1641.48 (CC), 2937.68 (C−H), and 3430.51 (O−H) cm⁻¹ indicating the presence of ether, alkene, alkane, alcohol, carboxylic acid, and fluoric compounds. This is summarily shown in Table 1. Table 2 shows the nomenclatures, molecular formulae, molecular weights, retention times, peak areas, and the nature of the analysed compounds. The mass spectra of the eleven compounds are shown in Figures 3–13 while their structures are shown in Figure 14.

Table 1: FT-IR absorption of the extract from variegated grasshopper (Z. variegatus).

FT-IR absorption (cm⁻¹)
Functional group
Nature of compound

1
1270.17
C–O or C–F
Alcohol, ether, carboxylic acid, ester, fluorocompound

2
1641.48
CC
Alkene

3
2937.68
C–H
Alkane

4
3430.51
O–H
Alcohol, carboxylic acid

Table 2: Compounds identified from the GC-MS analysis of the abdominal extract of variegated grasshopper (Z. variegatus).

Chromatogram peak
Compound name
Molecular formula
Molecular weight
Retention time (min)
Peak area (%)
Nature of compound

1
2,7-Dimethyloctane
C₁₀H₂₂
142
5.703
3.21
Hydrocarbon

2
Decane
C₁₀H₂₂
142
7.078
5.33
Hydrocarbon

3
Undecane
C₁₁H₂₄
156
8.455
3.81
Hydrocarbon

4
Tridecanoic acid methyl ester
C₁₄H₂₈O₂
228
17.151
4.76
Ester

5
Hexadecanoic acid (palmitic acid)
C₁₆H₃₂O₂
256
18.526
9.37
Fatty acid

6
11-Octadecenoic acid methyl ester
C₁₉H₃₆O₂
296
20.196
23.18
Ester

7
Pentadecanoic acid, 14-methyl-methyl ester
C₁₇H₃₄O₂
270
20.535
4.43
Ester

8
(Z)-13-Docosenoic acid (erucic acid)
C₂₂H₄₂O₂
338
21.288
10.71
Fatty acid

9
Dodecyl pentafluoropropionate
C₁₅H₂₅F₅O₂
332
22.753
9.52
Halogenated ester

10
2-Dodecyl-1,3-propanediol
C₁₅H₃₂O₂
244
24.145
6.38
Alcohol

11
1,12-Tridecadiene
C₁₃H₂₄
180
24.572
19.30
Hydrocarbon

Figure 1: GC-MS chromatogram of extract from abdominal glands of variegated grasshopper (Z. variegatus).

Figure 2: FT-IR spectrum of extract from the abdominal glands of variegated grasshopper (Z. variegatus).

Figure 3: 2,7-Dimethyloctane.

Figure 4: Decane.

Figure 5: Undecane.

Figure 6: Tridecanoic acid methyl ester.

Figure 7: Hexadecanoic acid.

Figure 8: Mass spectra of 11-octadecenoic acid methyl ester.

Figure 9: Pentadecanoic acid, 14-methyl-methyl ester.

Figure 10: (z)-13-Docosenoic acid.

Figure 11: Dodecyl pentafluoropropionate.

Figure 12: 2-Dodecyl-1,3-propanediol.

Figure 13: Mass spectra of 1,12-tridecadiene.

Figure 14: Structures of compounds identified from the GC/MS result of variegated grasshopper (Z. variegatus) extract.

This current investigation explicitly shows that variegated grasshopper contained many fractions in the cuticular abdominal regions which constituted esters, alkenes, alkanes, alcohol, carboxylic acids, and fluoric compounds. However, esters and hydrocarbons constituted the bulk of the extract from variegated grasshopper. Some of the identified constituents were fatty acids and alcohols. The fatty acids include palmitic and erucic acids. The hydrocarbon constituents of many insects are synthesized from fatty acids in a series of steps that involve chain shortening or elongation and desaturation and also through modification of the functional group by reduction, acetylation, or, sometimes, oxidation [16]. Hydrocarbons have been documented as sex pheromones of some insects; for instance, Peschke [17] reported that cuticular hydrocarbons served as cues for sexual recognition in a rove beetle (Staphylinidae). Also, other hydrocarbons, methylalkanes, and (Z)-9-tricosene have been reported as the sex pheromones in housefly [18, 19]. This investigation provides evidence suggesting that the constituents in variegated grasshopper are multicomponents and the constituents may be synergists to one another. In some other insects like termites, it has been reported that some sex pheromones produced by the reproductive sterna gland are identical to the worker trail pheromones but are secreted in much higher quantities, especially in females whose sternal gland is larger [20]. It is therefore worthy to state that the use of some of these compounds extracted from variegated grasshopper as sex, trail, or aggregating pheromones is a hypothesis. A cooperative and mutual stimulation of these compounds may enable effective communication and behavioural pattern of the insect.

4. Conclusion

The chemical compound extracted from the abdominal glands of variegated grasshopper was analysed with GC/MS and FT-IR techniques which revealed the presence of esters, hydrocarbons, fatty acids, and alcohols. Although behavioural bioassays were not carried out to determine whether these compounds are pheromones, development of the synthetic analogue of the compounds and further investigating the insect response to them might provide a green alternative method of variegated grasshopper control, leading to increased agricultural produce. Further research is therefore required.

Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.

Acknowledgment

The authors are grateful to Dr. O. O. Okore of Zoology Department, Michael Okpara University of Agriculture, Umudike, Abia State, Nigeria, for her kindness in identifying and authenticating their insect sample.

References

A. Hendrikse, Sex-pheromone communication and reproductive isolation in small ermine moths [Ph.D. thesis], University of Leiden, Leiden, The Netherlands, 1990.

K. O. Ademolu, B. A. Idowu, and O. A. Oke, “Impact of reproductive activities on the tissues of Zonocerus variegatus grasshopper adults (Orthoptera: Pygomorphidae),” Florida Entomologist, vol. 94, no. 4, pp. 993–997, 2011.

A. Youdeowei, The Dissection of the Variegated Grasshopper: Zonocerus variegatus (L.), Oxford University Press, Ibadan, Nigeria, 1974.

S. A. Toye, “Studies on the biology of the grasshopper pest Zonocerus variegatus (L.) (Orthoptera:Pyrgomorphidae) in Nigeria: 1911–1981,” International Journal of Tropical Insect Science, vol. 3, no. 1, pp. 1–7, 1982.

R. F. Chapman, W. W. Page, and A. R. McCaffery, “Bionomics of the variegated grasshopper (Zonocerus variegatus) in West and Central Africa,” Annual Review of Entomology, vol. 31, pp. 479–505, 1986.

W. W. Page, “The biology and control of the grasshopper Zonocerus variegatus,” PANS, vol. 24, no. 3, pp. 270–277, 1978.

S. Blanford, M. B. Thomas, and J. Langewald, “Thermal ecology of Zonocerus variegatus and its effects on biocontrol using pathogens,” Agricultural and Forest Entomology, vol. 2, no. 1, pp. 3–10, 2000.

G. Bani, “The acridid situation in the Congo,” in Biological Control of Locusts and Grasshoppers, C. J. Lomer and C. Prior, Eds., pp. 79–81, CAB International, Willingford, UK, 1992.

A. Niassy, “Senegal grasshopper and locust control in Senegal,” in Biological control of Locusts and Grasshoppers, C. J. Lomer and C. Prior, Eds., pp. 90–96, CAB International, Willingford, UK, 1992.

W. W. D. Modder, “Control of the variegated grasshopper Zonocerus variegatus (L.) on cassava,” African Crop Science Journal, vol. 2, no. 4, pp. 391–406, 1994.

R. Van den Bosch, The Pesticide Conspiracy, Doubleday and Company, Garden City, NY, USA, 1st edition, 1978.

A. B. Idowu, “The structure of the repellent gland is Zonocerus variegatus (Orthoptera: Pyrgomorphidae) 1,” African Zoology, vol. 109, pp. 247–252, 1995.

A. B. Idowu and O. A. Idowu, “Pharmacological properties of the repellent secretion of Zonocerus variegatus (Orthoptera: Prygomorphidae),” Revista de Biologia Tropical, vol. 47, no. 4, pp. 1015–1020, 1999.

A. O. Bamidele and W. A. Muse, “Geographical variation of the pyrgomorphid grasshopper, Zonocerus variegatus L. (Orthoptera: Pyrgomorphidae) in southern Nigeria,” Journal of Entomology and Zoology Studies, vol. 2, no. 2, pp. 72–75, 2014.

O. U. Igwe and D. E. Okwu, “GC-MS evaluation of bioactive compounds and antibacterial activity of the oil fraction from the stem bark of Brachystegia eurycoma harms,” International Journal of Chemical Sciences, vol. 11, no. 1, pp. 357–371, 2013.

R. F. Chapman, The Insects: Structure and Function, Cambridge University Press, 1998.

K. Peschke, “Cuticular hydrocarbons regulate mate recognition, male aggression, and female choice of the rove beetle, Aleochara curtula,” Journal of Chemical Ecology, vol. 13, no. 10, pp. 1993–2008, 1987.

E. C. Uebel, P. E. Sonnet, and R. W. Miller, “House fly sex pheromone: enhancement of mating strike activity by combination of (Z)-9-tricosene with branched saturated hydrocarbons,” Environmental Entomology, vol. 5, no. 5, pp. 905–908, 1976.

W. M. Rogoff, G. H. Gretz, P. F. Sonnet, and M. Schwarz, “Responses of male house flies to muscalure and to combinations of hydrocarbons with and without muscalure,” Environmental Entomology, vol. 9, no. 5, pp. 605–606, 1980.

J. M. Pasteels and C. Bordereau, “Releaser pheromones in termites,” in Pheromone Communication in Social Insects: Ants, Wasps, Bees, and Termites, R. K. VanderMeer, M. D. Breed, M. L. Winston, and K. E. Espalie, Eds., pp. 193–215, Westview Press, Oxford, UK, 1998.