Tumor promoting diterpenes from Euphorbia leuconeura L. Gerd Vogg a , Elke Mattes a , Jan Rothenburger a , Norbert Hertkorn b , Stefan Achatz b , Heinrich Sandermann Jr. a, * a Institut fu Èr Biochemische P¯anzenpathologie, GSF Ð Forschungszentrum fu Èr Umwelt und Gesundheit GmbH, Ingolsta Èdter Landstraûe 1, D-85764 Oberschleiûheim, Germany b Institut fu Èr O È kologische Chemie, GSF Ð Forschungszentrum fu Èr Umwelt und Gesundheit GmbH, Ingolsta Èdter Landstraûe 1, D-85764 Oberschleiûheim, Germany Received 26 October 1998; received in revised form 17 December 1998; accepted 17 December 1998 Abstract Diterpene esters of the phorbol and ingenol types are known to be highly active tumor promoting agents that typically occur in members of the Euphorbiaceae. In the present work, Euphorbia leuconeura, a rare indoor plant, is analyzed for its tumor promoting potential. Latex as well as total leaf extracts exhibited Epstein±Barr-virus (EBV) inducing activity comparable to 12- O-tetradecanoyl-phorbol-13-O-acetate, a well known tumor promoter. The activity of individual fractions correlated with their ingenol ester content. Three ingenol esters with EBV inducing activity could be isolated and identi®ed. They belong to the milliamine type of diterpene esters that contain aromatic peptidyl groups. Two of them (milliamines L and M) are already known from E. milii. The third compound is identi®ed as an isomer of milliamine F with a novel 3,20-diester arrangement. The data show a close relationship between E. leuconeura and the more popular indoor plant E. milii whose latex is also used as a powerful molluscicide. # 1999 Elsevier Science Ltd. All rights reserved. Keywords: Euphorbia leuconeura; Indoor plant; Euphorbiaceae; Ingenol esters; Tumor promotion; Epstein±Barr-virus induction 1. Introduction In two-stage chemical carcinogenesis, tumor promotors such as phorbol and ingenol diterpenes strongly enhance tumor formation by carcinogenic chemicals (Evans & Soper, 1978; Hecker, 1968). A pre- liminary screening of several available Euphorbiaceae (spurge) indoor plant species had shown that Euphorbia leuconeura contained high EBV-inducing ac- tivity for human lymphocytes. This method is generally accepted to test unknown substances and extracts for their tumor promoting potential (Kloz, Hergenhahn, Fellhauer, & Hecker, 1989; Polack et al., 1992). The assay is based on the combination of the luciferase reporter gene with an EBV early promoter (Polack et al., 1992). E. leuconeura is a succulent plant with attractive leaves. It is easy to cultivate and is occasionally used as an indoor plant. Its origin is in Madagascar and it belongs to the Euphorbia lophogona group (section Goniostema Baill). The tumor promoting compounds are now chemically characterized. E. leuconeura is re- lated to Euphorbia milii which is also known among gardeners as E. splendens or `crown of thorns'. The latex of E. milii contains a unique group of ingenol esters that were ®rst described by Uemura and Hirata (1971) and are termed milliamines. The latter are pep- tidyl esters containing substituted di- or tri-anthrani- loyl groups. At least 14 dierent milliamines have been identi®ed (Marston & Hecker, 1984a, 1984b; Zani, Marston, Hamburger, & Hostettmann, 1993). Some of the milliamines possess high molluscicidal activity, and the latex of E. milii ®nds practical application Phytochemistry 51 (1999) 289±295 0031-9422/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved. PII: S0031-9422(99)00016-3 * Corresponding author. Tel.: +49-89-3187-2285; fax +49-89- 3187-3383. E-mail address: sandermann@gsf.de (H. Sandermann Jr.) Abbreviations: EBV, Epstein±Barr-virus; LC±MS, liquid chroma- tography/mass spectroscopy; NMR, nuclear magnetic resonance; TPA, 12-O-tetradecanoyl-phorbol-13-O-acetate.