Tigliane Diterpenoids from the Stem Bark of Neoboutonia macrocalyx Peter G. Kirira, ²,‡ Geoffrey M. Rukunga, ‡ Alphonse W. Wanyonyi, ² Charles N. Muthaura, ‡ Geoffrey M. Mungai, § Alex K. Machocho, ² and Isaiah O. Ndiege* ,² Department of Chemistry, School of Pure and Applied Sciences, Kenyatta UniVersity, P.O. Box 43844, Nairobi 00100, Kenya, Center for Traditional Medicine and Drug Research, Kenya Medical Research Institute, P.O. Box 54840, Nairobi 00200, Kenya, and East Africa Herbarium, National Museums of Kenya, P.O. Box 40658, Nairobi 00100, Kenya ReceiVed August 23, 2006 Neoboutonia macrocalyx is a plant used by traditional healers among the Meru community in Kenya. Chromatographic fractionation of the petroleum ether and dichloromethane extracts of this plant yielded one known (1) and three new tigliane-type diterpenoids (2-4). The chemical structures of the isolated compounds were established through spectroscopic data interpretation. Neoboutonia macrocalyx Pax. (Euphorbiaceae), commonly known as ‘‘Mutuntuki” (Kimeru), is found in central and eastern Kenya. The stem bark is used to treat headache and fever in traditional medicine. Extracts from the stem bark of this plant have shown antiplasmodial activity. 1 Several sterols and daphnane and tigliane diterpenoids have been isolated from other Neoboutonia species. 2,3 As part of a program on phytochemical investigations on Kenyan medicinal plants, we describe herein the isolation of a known compound, 6R,7R-epoxy-5-hydroxy-12-deoxyphorbol-13-tetrade- canoate (6R,7R-epoxy-4,5,9R,20-tetrahydroxy-13R-tetradecanoate- 1-tiglien-3-one) (1), 4,5 and the structure elucidation of three new tigliane diterpenoid compounds (2-4) from the dichloromethane and petroleum ether extracts of the stem bark of N. macrocalyx. The structure of the known compound, 6R,7R-epoxy-5-hydroxy- 12-deoxyphorbol-13-tetradecanoate (1), was determined by com- parison of generated spectroscopic data with values reported in the literature. 4,5 The spectroscopic data for this compound is included in Table 1 for comparison with other derivatives that were isolated together with 1. Comparison of the IR data for 1 and 2 revealed minor differences. Unlike 1, which had one ester group (ν max 1713 cm -1 ), the IR spectrum of 2 revealed the presence of two ester groups (ν max 1735, 1717 cm -1 ). The 1 H NMR spectrum of 2 (Table 1) showed minor differences including the presence of six methyl groups at δ 0.89 (t, J ) 6.3 Hz, Me-12′′), 0.85 (t, J ) 7.0 Hz, Me-16′), 0.93 (d, J ) 6.7 Hz, H-18), 1.78 (s, H-19), 1.19 (s, H-16), and 1.09 (s, H-17) instead of five in 1. Unlike in 1, the two diastereotopic methylene protons attached to the oxygenated carbon (C-20) in 2 were observed distinctly at δ 4.80 (1H, d, J ) 12.0 Hz, H-20a) and 3.82 (1H, d, J ) 12.0 Hz, H-20b), suggesting esterification at this carbon. The region between δ 1.26 (br, m) and 1.60 (m) was more complicated in 2 due to an increase in the number of CH 2 groups in the ester groups (C-4′-C-15′, C-4′′-C-12′′). The presence of two signals at δ 2.30 (2H, t, J ) 7.6 Hz, H-2′) and 2.30 (2H, t, J ) 7.5 Hz, H-2′′), superimposed on each other, confirmed the presence of two ester groups. The 1 H NMR data for the cyclopentyl and cyclohexyl rings of 2 were similar to those of 1, 4,5 mellerin A, 2 and 12-deoxyphorbol-13-hexadecanoate, 6 while the data for the cycloheptyl ring were similar to those of montanin. 3 The rest of the 1 H NMR signals for 2 were similar to those of 1. The 13 C NMR spectrum of 2 (Table 1) revealed minor differences from that of 1 and confirmed the presence of two ester groups (δ 175.3, C-1′; 173.4, C-1′′). The presence of an ester group at C-20 was further confirmed by the shift of the signal to δ 65.7 in 2 from δ 64.8 in 1. The signals of the gem-dimethylcyclopropane skeleton were as reported for 1 7,8 and other tigliane diterpenoids. The 17 carbon signals at δ 29.1-29.7 (C-4′-C-13′ and C-4′′-C-10′′) were assigned to the CH 2 groups in the two pendent ester chains. In addition to the two ester groups, the other part of the molecule contained 20 carbons and was proposed to be a tigliane-type diterpenoid skeleton on the basis of literature spectroscopic data. 2,8 COSY, HMBC, and HMQC spectra confirmed the existence of two side chain ester moieties in 2 (hexadecanoyl at C-13 and dodecanoyl at C-20). The connectivity of the methyl, methylene, methine, and quaternary carbons was determined on the basis of 1 H- 13 C NMR long-range correlation signals in the HMBC spectrum. The EIMS fragments observed at m/z 239 and 185 were assigned to loss of a hexadecanoyl and a lauroyl ion fragment, respectively. The mo- lecular formula, C 48 H 80 O 9 , for 2 was deduced from the quasi- molecular ion adducts observed at m/z 829 [M + C 2 H 5 ] + and 801 [M + H] + in the CIMS and confirmed by HREIMS (m/z 661.4830 for C 43 H 65 O 5 ). Consequently, the structure of 2 was confirmed as * Corresponding author. Tel: +254-20-810901, +254-20-811622. Fax: +254-20-811575. E-mail: indiege@icipe.org. ² Kenyatta University. ‡ Kenya Medical Research Institute. § East Africa Herbarium. 842 J. Nat. Prod. 2007, 70, 842-845 10.1021/np0604174 CCC: $37.00 © 2007 American Chemical Society and American Society of Pharmacognosy Published on Web 03/24/2007