ORIGINAL ARTICLE Metabolomics demonstrates divergent responses of two Eucalyptus species to water stress Charles R. Warren • Ismael Aranda • F. Javier Cano Received: 12 September 2010 / Accepted: 11 March 2011 / Published online: 19 March 2011 Ó Springer Science+Business Media, LLC 2011 Abstract Past studies of water stress in Eucalyptus spp. generally highlighted the role of fewer than five ‘‘impor- tant’’ metabolites, whereas recent metabolomic studies on other genera have shown tens of compounds are affected. There are currently no metabolite profiling data for responses of stress-tolerant species to water stress. We used GC–MS metabolite profiling to examine the response of leaf metabolites to a long (2 month) and severe (W predawn \ -2 MPa) water stress in two species of the perennial tree genus Eucalyptus (the mesic Eucalyptus pauciflora and the semi-arid Eucalyptus dumosa). Polar metabolites in leaves were analysed by GC–MS and inor- ganic ions by capillary electrophoresis. Pressure–volume curves and metabolite measurements showed that water stress led to more negative osmotic potential and increased total osmotically active solutes in leaves of both species. Water stress affected around 30–40% of measured metab- olites in E. dumosa and 10–15% in E. pauciflora. There were many metabolites that were affected in E. dumosa but not E. pauciflora, and some that had opposite responses in the two species. For example, in E. dumosa there were increases in five acyclic sugar alcohols and four low- abundance carbohydrates that were unaffected by water stress in E. pauciflora. Re-watering increased osmotic potential and decreased total osmotically active solutes in E. pauciflora, whereas in E. dumosa re-watering led to further decreases in osmotic potential and increases in total osmotically active solutes. This experiment has added several extra dimensions to previous targeted analyses of water stress responses in Eucalyptus, and highlights that even species that are closely related (e.g. congeners) may respond differently to water stress and re-watering. Keywords Drought Á Metabolome Á Osmotic adjustment Á Water relations Á GC–MS Á Plant Á Water stress 1 Introduction Maintenance of turgor is a necessary prerequisite for cell enlargement and growth, and thus limited water supply can cause large reductions in plant growth (Hsiao 1973; Kramer and Boyer 1995). Nearly all plants can acclimate to water deficits, at least to some extent, by employing a range of responses to maintain cell turgor and growth under more negative soil water potentials than would otherwise be possible. These responses affect various aspects of physi- ology, biochemistry and molecular biology (Shao et al. 2009). In the case of the large Eucalyptus genus, for example, responses to water deficits include changes in biomass allocation (Rawat and Banerjee 1998), stomatal control (Macfarlane et al. 2004), cell wall reinforcement (Ladiges 1975), cell wall water storage (Tuomela 1997) and cellular osmolarity (Myers and Neales 1986; Lemcoff Electronic supplementary material The online version of this article (doi:10.1007/s11306-011-0299-y) contains supplementary material, which is available to authorized users. C. R. Warren (&) School of Biological Sciences, Heydon-Laurence Building A08, University of Sydney, Sydney, NSW 2006, Australia e-mail: charles.warren@sydney.edu.au I. Aranda Á F. J. Cano Centro de Investigacio ´n Forestal, Instituto Nacional de Investigacio ´n y Tecnologı ´a Agraria y Alimentaria (INIA), Carretera de la Corun ˜a km 7.5, 28040 Madrid, Spain F. J. Cano Unidad Docente de Anatomı ´a, Fisiologı ´a y Gene ´tica Forestal, E.T.S.I. Montes, Universidad Polite ´cnica de Madrid (UPM), Ciudad Universitaria s/n, 28040 Madrid, Spain 123 Metabolomics (2012) 8:186–200 DOI 10.1007/s11306-011-0299-y