Abstract
The metabolic alterations of Brassica rapa (L.) leaves attacked by larvae of the specialist Plutella xylostella L. (Lepidoptera: Yponomeutidae) and the generalist Spodoptera exigua Hubner (Lepidoptera: Noctuidae) were investigated with nuclear magnetic resonance (NMR) spectroscopy, followed by a multivariate data analysis. The principal component analysis (PCA) of 1H NMR spectra showed that metabolic changes in B. rapa leaves induced by the 2nd and the 4th instars were different from each other. However, the congestion of the one-dimensional 1H NMR spectrum made it difficult to identify discriminating metabolites. To overcome the spectral complexity, several two-dimensional NMR techniques were applied. Of those evaluated, J-resolved spectroscopy, which affords an additional coupling constant, provided a wide range of structure information on differentiating the metabolites. Based on the J-resolved spectra combined with PCA, the major signals contributing to the discrimination were alanine, threonine, glucose, sucrose, feruloyl malate, sinapoyl malate, and gluconapin.
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Baldwin, I. T. and Preston, C. A. 1999. The eco-physiological complexity of plant responses to insect herbivores. Planta 208:137–145.
Baldwin, I. T., Halitsche, R., Kessler, A., and Schittko, U. 2001. Merging molecular and ecological approaches in plant–insect interactions. Curr. Opin. Plant Biol. 4:351–358.
Choi, Y. H., Kim, H. K., Hazenkamp, A., Erkelens, C., Lefeber, A. W. M., and Verpoorte, R. 2004a. Metabolomic differentiation of Cannabis sativa cultivars using 1H NMR and principal component analysis. J. Nat. Prod. 67:953–957.
Choi, Y. H., Tapias, E. C., Kim, H. K., Lefeber, A. W., Erkelens, C., Verhoeven, J. T., Brzin, J., Zel, J., and Verpoorte, R. 2004b. Metabolic discrimination of Catharanthus roseus leaves infected by phytoplasma using 1H-NMR spectroscopy and multivariate data analysis. Plant Physiol. 135:2398–2410.
Doughty, K. J., Blight, M. M., Bock, C. H., Fieldsend, J. K., and Pickett, J. A. 1996. Release of alkenyl isothiocyanates and other volatiles from Brassica rapa seedlings during infection by Alternaria brassicae. Phytochemistry 43:371–374.
Daayf, F., Ongena, M., Boulanger, R., El Hadrami, I., and Belanger, R. R. 2000. Induction of phenolic compounds in two cultivars of cucumber by treatment of healthy and powdery mildew-infected plants with extracts of Reynoutria sachalinensis. J. Chem. Ecol. 26:1579–1593.
Foxall, P. J. D., Parkinson, J. A., Sadler, I. H., Lindon, J. C., and Nicholson, J. K. 1993. Analysis of biological fluids using 600 MHz proton NMR spectroscopy—application of homonuclear 2-dimensional J-resolved spectroscopy to urine and blood plasma for spectral simplification and assignment. J. Pharm. Biomed. 11:21–31.
Grant, G. G. and Langevin, D. 2002. Oviposition responses of four Choristoneura (Lepidoptera: Tortricidae) species to chemical an physical stimuli associated with host and nonhost foliage. Environ. Entomol. 23:224–456.
Geervliet, J. B. F., Posthumus, M. A., Vet, L. E. M., and Dicke, M. 1997. Comparative analysis of headspace volatiles from different caterpillar-infested or uninfested food plants of Pieris species. J. Chem. Ecol. 23:2935–2954.
Gouinguene, S., Degen, T., and Turlings, T. C. J. 2001. Variability in herbivore-induced odour emissions among maize cultivars and their wild ancestors (teosinte). Chemoecology 11:9–16.
Halkier, B. A. and Du, L. 1997. The biosynthesis of glucosinolates. Trends Plant Sci. 2:425–431.
Liang, Y.-S., Kim, H. K., Lefeber, A. W. M., Erkelens, C., Choi, Y. H., and Verpoorte, R. 2006. Identification of phenylpropanoids in Brassica rapa leaves treated with methyl jasmonate using two-dimensional nuclear magnetic resonance spectroscopy. J. Chromatogr. A 1112:148–155.
Maleck, K. and Dietrich, R. A. 1999. Defense on multiple fronts: How do plants cope with diverse enemies? Trends Plant Sci. 4:215–219.
Mauricio, R. and Rausher, M. D. 1997. Experimental manipulation of putative selection agents provides evidence for the role of natural enemies in the evolution of plant defense. Evolution 51:1435–1444.
Nicholson, J. K., Foxall, P. J. D., Spraul, M., Farrant, R. D., and Lindon, J. C. 1995. 750 MHz 1H and 1H–13C NMR spectroscopy of human blood plasma. Anal. Chem. 67:793–811.
Nilsson, M., Duarte, I. F., Almeida, C., Delgadillo, I., Goodfellow, B. J., Gil, A. M., and Morris, G. A. 2004. High-resolution NMR and diffusion-ordered spectroscopy of port wine. J. Agric. Food Chem. 52:3736–3743.
Peiwu, L., Guangming, L., Wen, Z., Mei, Y., and Jianqiang, Z. 1999. Investigations on dominant glucosinolates in rapeseed germplasm collected in China. Proceedings of the 10th International Rapeseed Congress, Canberra, Australia. The Regional Institute, Ltd., Australia.
Ratzka, A., Vogel, H., Kliebenstein, D. J., Mitchell-Olds, T., and Kroymann, J. 2002. Disarming the mustard oil bomb. Proc. Natl. Acad. Sci. U.S.A. 99:11223–11228.
Rostas, M., Bennett, R., and Hilker, M. 2002. Comparative physiological responses in Chinese cabbage induced by herbivory and fungal infection. J. Chem. Ecol. 28:2449–2462.
Sharma, B. K. and Singh, U. P. 2003. Ferulic acid may prevent infection of Cicer arietinum by Sclerotium rolfsii. World J. Microb. Biotechnol. 19:123–127.
Shonle, I. and Bergelson, J. 2000. Evolutionary ecology of the tropane alkaloids of Datura stramonium L. (Solanaceae). Evolution 54:778–788.
Stotz, H. U., Kroymann, J., and Mitchell-Olds, T. 1999. Plant–insect interactions. Curr. Opin. Plant Biol. 2:268–272.
Stout, J. and Chapple, C. 2004. The phenylpropanoid pathway in Arabidopsis: Lessons learned from mutant in sinapate ester biosynthesis, Ch. 3, pp. 39–67, in J. T. Romeo (ed.). Secondary Metabolism in Model Systems. Elsevier, Amsterdam, The Netherlands.
Stratmann, J. W and Ryan, C. A. 1997. Myelin basic protein kinase activity in tomato leaves is induced systemically by wounding and increases in response to systemin and oligosaccharide elicitors. Proc. Natl. Acad. Sci. U.S.A. 93:11085–11089.
Sumner, L. W., Mendes, P.,and Dixon, R. A. 2003. Plant metabolomics: Large-scale phytochemistry in the functional genomics era. Phtyochemistry 62:817–836.
Viant, M. R. 2003. Improved methods for the acquisition and interpretation of NMR metabolomic data. Biochem. Biophys. Res. Commun. 310:943–948.
Vuorinen, T., Reddy, G. V. P., Nerg, A. M., and Holopainen, J. K. 2004. Monoterpene and herbivore-induced emissions from cabbage plants grown at elevated atmospheric CO2 concentration. Atmos. Environ. 38:675–682.
Ward, J. L., Harris, C., Lewis, J., and Beale, M. H. 2003. Assessment of 1H NMR spectroscopy and multivariate analysis as a technique for metabolite fingerprinting of Arabidopsis thaliana. Phytochemistry 62:949–957.
Zasada, I. A. 2003. Sensitivity of Meloidogyne javanica and Tylenchulus semipenetrans to isothiocyanates in laboratory assays. Phytopathology 93:747–750.
Acknowledgment
Financial support from the Netherlands Education Centre (STUNED Scholarship 2003) for Heru Tri Widarto is highly acknowledged. We also acknowledge Henk Nell for rearing S. exigua larvae.
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Widarto, H.T., Van Der Meijden, E., Lefeber, A.W.M. et al. Metabolomic Differentiation of Brassica rapa Following Herbivory by Different Insect Instars using Two-Dimensional Nuclear Magnetic Resonance Spectroscopy. J Chem Ecol 32, 2417–2428 (2006). https://doi.org/10.1007/s10886-006-9152-6
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DOI: https://doi.org/10.1007/s10886-006-9152-6