Biosynthetic origins of the isoprene units of gaudichaudianic acid in Piper gaudichaudianum (Piperaceae) Adriana A. Lopes a , Debora C. Baldoqui a , Silvia N. Lo ´pez a , Massuo J. Kato b , Vanderlan da S. Bolzani a , Maysa Furlan a, * a Instituto de Quı ´mica, Universidade Estadual Paulista, C.P. 355, CEP 14801-970 Araraquara, SP, Brazil b Instituto de Quı ´mica, Universidade de Sa ˜o Paulo, C.P. 26077, CEP 05599-970 Sa ˜o Paulo, SP, Brazil Received 19 October 2006; received in revised form 13 April 2007 Available online 14 June 2007 Abstract The biosynthesis of (2S)-2-methyl-2-(4 0 -methyl-3 0 -pentenyl)-8-(3 00 -methyl-2-butenyl)-2H-1-benzopyran-6-carboxylic acid (gaudichau- dianic acid), the major metabolite in leaves and roots of Piper gaudichaudianum Kunth (Piperaceae), has been investigated employing [1- 13 C]-D-glucose as precursor. The labelling pattern in the isolated gaudichaudianic acid was determined by quantitative 13 C NMR spec- troscopy analysis and was consistent with involvement of both mevalonic acid and 2-C-methyl-D-erythritol-4-phosphate pathways in the formation of the dimethylallyl- and geranyl-derived moieties. The results confirmed that both plastidic and cytoplasmic pathways are able to provide isopentenyl diphosphate units for prenylation of p-hydroxybenzoic acid. Ó 2007 Elsevier Ltd. All rights reserved. Keywords: Piper gaudichaudianum; Piperaceae; Biosynthesis; Chromene; Gaudichaudianic acid; Mevalonate pathway; 2-C-Methyl-D-erythritol-4- phosphate pathway; Isopentenyl diphosphate units 1. Introduction The isoprenoids, when considered together with the ste- roids, constitute the largest class of secondary metabolites comprising more than 30,000 known compounds (Dictio- nary of Natural Products, 2000). The common biosynthetic building block of this group, the so-called isoprene unit, is derived from isopentenyl diphosphate (IPP) and dimethyl- allyl diphosphate (DMAPP). The biosynthesis of IPP can proceed via two different pathways, namely the long- established mevalonic acid (MVA) and the 2-C-methyl-D- erythritol-4-phosphate (MEP) pathways, the reaction sequence of which has been characterized (Adam et al., 1999). In plants, the MEP pathway appears generally oper- ative for formation of monoterpenoids, diterpenoids, phy- tols and carotenoids (Hirai et al., 2000; Umlauf et al., 2004; Bouvier et al., 2005). The key regulatory step of the MVA pathway involves reduction of 3-hydroxy-3-methylglutaryl CoA (HMG- CoA) to mevalonate, with this being catalysed by the cyto- solic enzyme HMG-CoA reductase (HMGR; Fig. 1). Plastidic IPP is derived, however, from MEP formed via the condensation of pyruvate and glyceraldehyde-3-phos- phate (G3P) catalysed by 1-deoxy-D-xylulose-5-phosphate synthase (DXS; Fig. 1) (Bouvier et al., 2005; Enfissi et al., 2005). The two pathways to IPP are thus compart- mentalized in the cytosol and the plastid, respectively; how- ever, they may function in the formation of certain isoprenoids, in a cooperative process that results from met- abolic cross-talk within the plastidic envelope membrane (Enfissi et al., 2005). A quantitative assessment of the differential contribu- tion of the two IPP pathways in the biosynthesis of an individual isoprenoid may be deduced from the 13 C-label- ling pattern, as determined by 13 C NMR spectroscopy, fol- lowing incorporation of [1- 13 C]-D-glucose into product (Rohmer et al., 1993; Umlauf et al., 2004). As shown in 0031-9422/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.phytochem.2007.04.025 * Corresponding author. Tel.: +55 16 3301 6679; fax: +55 16 3322 2308. E-mail address: maysaf@iq.unesp.br (M. Furlan). www.elsevier.com/locate/phytochem Phytochemistry 68 (2007) 2053–2058 PHYTOCHEMISTRY