Classification of Aristolochia species based on GC–MS and chemometric analyses of essential oils Carla S. Francisco a , Gisele B. Messiano a , Lucia M.X. Lopes a, * , Aristeu G. Tininis a , Jose ´ E. de Oliveira a , Lindolpho Capellari Jr. b a Instituto de Quı ´mica, Universidade Estadual Paulista, UNESP, C.P. 355, 14801-970 Araraquara, SP, Brazil b Escola Superior de Agricultura ‘‘Luiz de Queiroz’’ (ESALQ), USP, C.P. 09, 13418-900 Piracicaba, SP, Brazil Received 18 January 2007; received in revised form 28 May 2007 Available online 20 August 2007 Abstract Essential oils were obtained from roots of 10 Aristolochia species by hydrodistillation and analysed by GC–MS. A total of 75 com- pounds were identified in the analysed oils. Multivariate analyses of the chemical constituents of the roots enabled classification of the species into four morphological groups. These forms of analysis represent an aid in identification of further specimens belonging to these species. Ó 2007 Elsevier Ltd. All rights reserved. Keywords: Aristolochia; Aristolochiaceae; Essential oils; Multivariate analysis; Principal component analysis; Morphology; Monoterpene; Sesquiterpene 1. Introduction Brazilian Aristolochiaceae is represented by three genera: Aristolochia, which consists of approximately 100 species, and Holostylis and Euglypha, each of which consists of only one species Holostylis reniformis and Euglypha rojasiana (Hoehne, 1942). The latter have been suggested to be included in the Aristolochia genus (Gonza ´lez, 1997). With regard to synonyms, more than 300 names have been pro- posed for these species. Aristolochia and Euglypha are also represented by both small and large vines, whereas Holosty- lis reniformis is a shrub. In Brazilian traditional medicine, these plants are known as ‘‘one thousand men’’, and have been mainly used as abortifacients, stomachics, antiophidi- ans, antiasthmatics, expectorants, and, recently, in slim- ming therapies (Lopes et al., 2001). They are generally sold at popular Brazilian markets, under their common names. Thus, there has been little distinction made among the species. In addition to the Aristolochia species and Lepi- dopteras belonging to the specialist Troidini tribe (Papilion- idae and Papilioninae) which feed on such plants, aristolochic acids have been detected in Chinese phytomed- icines and dietary supplements used in slimming therapy (Ioset et al., 2003). These acids have been associated with Chinese herb nephropathy, which is a kind of severe kidney disease caused by intake of excessive aristolochic acids (Bal- achandran et al., 2005; Cui et al., 2005; Ishibe et al., 2005; Zang et al., 2005). Therefore, it is essential, for health safety and quality control of related Brazilian herbal medicines, to develop efficient methods for species identification. In our continuous studies on the Aristolochiaceae fam- ily, we have shown that the chemical constituents of Brazil- ian species differ significantly. Although allantoin, sugars, aristolochic acids, and furofuran and butyrolactone lignans occur in most of these species (Lopes et al., 2001), they also contain specific types of compounds, such as bicubebins in Aristolochia lagesiana and Aristolochia pubescens (Pascoli et al., 2006), aryltetralone lignans in H. reniformis (da Silva and Lopes, 2004, 2006), benzofuran neolignans, and diter- pene ester of aristolochic acids in A. pubescens (Nascimento et al., 2000; Nascimento and Lopes, 1999, 2000, 2003). 0031-9422/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.phytochem.2007.07.007 * Corresponding author. Tel.: +55 16 3301 6663; fax: +55 16 3301 6692. E-mail address: lopesxl@iq.unesp.br (L.M.X. Lopes). www.elsevier.com/locate/phytochem Available online at www.sciencedirect.com Phytochemistry 69 (2008) 168–175 PHYTOCHEMISTRY