Natural Product Isolation DOI: 10.1002/anie.201310923 EBC-219: A New Diterpene Skeleton, Crotinsulidane, from the Australian Rainforest Containing a Bridgehead Double Bond** Lidiya A. Maslovskaya, Andrei I. Savchenko, Elizabeth H. Krenske, Carly J. Pierce, Victoria A. Gordon, Paul W. Reddell, Peter G. Parsons, and Craig M. Williams* Dedicated to Professor Armin de Meijere on the occasion of his 75th birthday Abstract: EBC-219 (4), isolated from Croton insularis (Baill), was established by spectroscopic and DFT methods as the first member of a new diterpene skeletal class, uniquely defined by the presence of a bicyclo[10.2.1] bridgehead olefin. The proposed biogenetic pathway to 4 from the co-isolated natural products EBC-131 (1), EBC-180 (2) and EBC-181 (3) is highly likely. EBC-180 (2) and EBC-181 (3) showed moderate to strong cytotoxic activity against various cancer cell lines. Croton is a well-known plant genus, which has delivered many classes of bioactive compounds, for example phorbols, [1] ent-kauranes, [2] clerodanes, [3] and halimanes [4] to name a few. This rich history prompted our interest in pursuing the minor constituents of croton, particularly Croton insularis (Baill), [5, 6] from Australias rain forest. [7] We herein report the isolation of a novel diterpene, EBC-219 (4) (Figure 1), which defines a new diterpene skeletal class, in addition to possessing a rare naturally occurring bridgehead olefin (anti-Bredt system). The term anti-Bredt, arising from the work of Julius Bredt in the early 1900s, [8] is bestowed upon unsaturated bridgehead bicyclic caged systems. [8] This rule, although initially derived by Bredt utilizing monoterpenes, has been mostly contested by synthetic and physical organic chemists. [9] Nevertheless, there are some rare, but famous, natural product examples, such as taxol [10] and the phomoidrides, [11] along with other uniquely sized ring systems [12, 13] that have appeared in the literature. Lastly, reassignments of natural product structures based on Bredts rule have been made by our group [14] and by Fraga et al. [15] EBC-131 (1), EBC-180 (2), EBC-181 (3), and EBC-219 (4) (Figure 1) were extracted from the stems of Croton insularis (Baill) and separated by silica gel chromatography followed by HPLC, as guided by a combination of bioassays and 1 H NMR spectroscopy. The high-resolution mass spectrum of EBC-131 (1) gave the molecular formula C 20 H 30 O 3 suggesting six ring and/or double-bond equivalents (RDBE). The 1 H NMR spectrum revealed two methyl groups situated on double bonds (d H = 1.58, 1.69 ppm), and a third which was adjacent to a carbonyl group (d H = 1.93 ppm). This, together with 13 C NMR shifts, corresponded to the data of a casbane reported by Filho et al. [16] EBC-180 (2), EBC-181 (3) and EBC-219 (4) all possess the same molecular formula (C 20 H 28 O 3 ), which constitutes seven RDBE. For EBC-180 (2) the 1 H NMR and 13 C NMR spectra revealed three methyl groups (d H = 1.57, 1.85, and 1.97 ppm) each placed on a double bond, one of which is isolated while the other two are adjacent to two carbonyls, suggesting the remainder of the skeleton to be bicyclic. Considering the similarity to EBC-131 (1), compar- ison to literature data confirmed EBC-180 (2) as another known casbane. [17] However, a slight reassignment of the reported structure containing a 6-Z double bond was required Figure 1. EBC-131 (1), EBC-180 (2), EBC-181 (3), and EBC-219 (4). Numbering of the casbane system used for 4. [*] Dr. A. I. Savchenko, Dr. E. H. Krenske, Assoc.Prof. C. M. Williams School of Chemistry and Molecular Biosciences University of Queensland Brisbane, 4072, Queensland (Australia) E-mail: c.williams3@uq.edu.au Homepage: http://www.scmb.uq.edu.au/homepages/williams/ index.html Dr. L. A. Maslovskaya, C. J. Pierce, Prof. P. G. Parsons Queensland Institute of Medical Research, Brisbane (Australia) Dr. V. A. Gordon, Dr. P. W. Reddell EcoBiotics Limited, Yungaburra (Australia) [**] We thank EcoBiotics Ltd. and the University of Queensland (UQ) for financial support as well as Dr. L. Lambert and Dr. G. Pierens from CAI UQ for assistance with NMR measurements. Computational resources at NCI NF and UQ RCC are gratefully acknowledged along with the Australian Research Council for Future Fellowship awards to E.H.K. and C.M.W. Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/anie.201310923. A ngewandte Chemi e 1 Angew. Chem. Int. Ed. 2014, 53,1–5  2014 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim These are not the final page numbers! Ü Ü