HETEROCYCLES

Short Paper | Special issue | Vol. 84, No. 2, 2012, pp. 1265-1270
Received, 7th June, 2011, Accepted, 19th July, 2011, Published online, 1st August, 2011.
DOI: 10.3987/COM-11-S(P)31

■ Chisomicines D and E, Two New Limonoids from Chisocheton ceramicus

Ibrahim A. Najmuldeen, A. Hamid A. Hadi,* Khalit Mohamad, Khalijah Awang, Kamal Aziz Ketuly, Mat Ropi Mukhtar, Hairin Taha, Noraziah Nordin, Marc Litaudon, Françoise Guéritte, Alfarius Eko Nugroho, and Hiroshi Morita*

Department of Chemistry, Faculty of Science, University of Malaya, Pantai Valley, 50603 Kuala Lumpur, Malaysia

Abstract

Two new limonoids, chisomicines D and E (1 – 2) have been isolated from the bark of Malaysian Chisocheton ceramicus (Miq.) C. DC. Their structures were elucidated on the basis of 1D and 2D-NMR data analysis.

Meliaceae or the mahogany family comprises over 1400 species in 51 genera of woody plants distributed in subtropical and tropical regions worldwide. Within this family, two major subfamilies,1 Melioideae and Swietenioideae, have been recognized based on seed morphology,2 wood anatomy and molecular data.3 Melioideae forms a monophyletic group that includes 73% of the genera and 87% of the species of the family.3 Plants belonging to the Meliaceae family are known to produce a group of highly oxidized triterpenes named limonoids.
Information about a wide range of biological activities, such as antimurine P388 leukemia cell lines especially those with a 14,15-epoxy D ring,4 antifeedants,5 insecticides,6 antitumor,7 and antimalarial and growth regulating activities,8 as well as a variety of medicinal effects in animals and humans was shown for limonoids.9, 10 We have found the alcoholic extract of the bark of Chisocheton ceramicus (Miq.) C. DC. (Meliaceae) is a rich source of interesting limonoids, named as a series of ceramicines and chisomicines.11-14 In continuation of our research on the Meliaceae family,14-16 we have isolated two new phragmalin-type limonoids, chisomicines D (1) and E (2) from the bark of C. ceramicus. We now wish to report isolation and structure elucidation of 1 and 2.

Chisomicine D (1) was obtained as white amorphous solid and the HRESIMS displayed a pseudomolecular ion peak [M+H]+ at m/z 555.2551 corresponding to the molecular formula of C31H38O9 and IR absorptions (1735 and 1703 cm-1) implied the presence of carbonyl functionalities.
The 1H-NMR spectrum (CDCl3 700MHz) of 1 displayed signals for four tertiary methyls at δH 1.14 (H3-18), 1.02 (H3-19), 0.93 (H3-28), and 1.95 (H3-4’) as well as one methoxy at δH 3.71. The signals typical for a furan moiety at δH 6.45 (H-22), 7.42 (H-23), and 7.70 (H-21) were observed at downfield region.
The 13C NMR spectrum of 1 revealed the presence of thirty one carbon atoms in the molecule, and the observed signals were very similar to those of chisomicine B (3).14 Further analysis of the 13C NMR data indicated that the following signals at δC 135.3 (C-2’), δC 127.1 (C-3’), and δC 21.5 (C-4’) in place of δC 128.0 (C-2’), δC 139.3 (C-3’), δC 14.2 (C-4’), and δC 12.1 (C-5’) were observed, which implies the presence of a methacrylate group in 1 instead of a tiglate group in chisomicine B.

The previously mentioned assumption as well as the planar structure of 1 was confirmed by using the 2D-NMR correlations (HSQC, 1H–1H COSY, and HMBC) as shown in Figure 1. The HMBC correlations of H2-3’ to C-1’, C-2’, and C-4’ verified the presence of a methacrylate moiety, while the HMBC correlation of H-3 to C-1’ confirmed attachment of the methacrylate moiety to C-3.

The relative structure of 1 was presumed to be the same as chisomicine B because of the high similarity of the NMR chemical shifts of the backbone skeleton, and the NOESY correlations further supports the presumption. NOESY correlations (Figure 2) of H-2/H-29b and H-3/H3-28 and H-29b suggested β orientation of H3-28 and α orientation of H-2 and H-3 while those of H-5/H-14 and H-3’b and H-14/H-30 indicated β orientation of H-5, H-14, and H-30. And finally the relative configurations of C-10, C-13, and C-14 were deduced from the NOESY correlations of H-14/H-10 and H3-18, implying α orientation of H-10, H-14, and H3-18.

Chisomicine E (2) was obtained as white amorphous solid and the HRESIMS displayed a pseudomolecular ion peak [M+H]+ at m/z 557.2693 corresponding to the molecular formula of C31H40O9, and IR absorptions (1735 cm-1) implied the presence of carbonyl functionalities. The 1H and 13C NMR spectra of 2 are similar to those of 1 except for the chemical shifts of a side chain at position 3 (Table 1). Based on the NMR data and the HRESIMS data, the structure of 2 was presumed to be chisomicine E with an isobutyrate group at C-3 instead of a methacrylate group. The COSY correlations of H-2’/H3-3’ and H3-4’ and the HMBC correlations of H3-3’ and H-3 to C-1’ confirmed the existence of the isobutyrate unit at C-3 (Figure 1).
The relative structure of 2 was assumed to be the same as chisomicine B based on the chemical shifts similarity, and was also confirmed by the observed NOESY correlations which are similar to those of 1.
The isolated compounds were tested for cytotoxicity against HL-60 cell line. Both compounds were found to be inactive (IC50 > 50 µM).

EXPERIMENTAL
General Experimental Procedures. Optical rotations were measured on a JASCO DIP-1000 automatic digital polarimeter. UV spectra were obtained on an Ultrospec 2100 pro spectrophotometer and IR spectra were recorded on a JASCO FT/IR-4100 spectrophotometer. High-resolution ESI MS were obtained on a LTQ Orbitrap XL (Thermo Scientific). 1H and 2D NMR spectra were recorded on a Bruker AV700 spectrometer and chemical shifts were referenced to the residual solvent peaks (δH 7.26 and δC 77.0 for chloroform-d). Standard pulse sequences were employed for the 2D NMR experiments. 1H-1H COSY, and NOESY spectra were measured with spectral widths of both dimensions of 4800 Hz, and 32 scans with two dummy scans were accumulated into 1K data points for each of 256 t1 increments. NOESY spectra in the phase sensitive mode were measured with a mixing time of 800 ms. For HSQC spectra in the phase sensitive mode and HMBC spectra, a total of 256 increments of 1K data points were collected. For HMBC spectra with Z-axis PFG, a 50 ms delay time was used for long-range C-H coupling. Zero-filling to 1K for F1 and multiplication with squared cosine-bell windows shifted in both dimensions were performed prior to 2D Fourier transformation.
Plant Material. The barks of C. ceramicus were collected in 2000 from Hutan Simpan Bukit Enggang, Malaysia. The plant species was identified by Mr. Teo Leong Eng, with a voucher specimen (No. KL 4973) and the herbarium specimen was deposited in the herbarium of the Chemistry Department, University of Malaya.
Extraction and Isolation. The dried and powdered bark of C. ceramicus (900 g) was extracted successively with methanol and the methanol extract (200 g) was partitioned with 10% MeOH aq. and EtOAc. EtOAc soluble materials (10 g) were subjected to a silica gel column (hexane/ EtOAc, 1:0→0:1), in which a fraction eluted with hexane/EtOAc (1:4) was further purified on a silica gel column with the mixture of EtOAc/acetone/hexane (65:10:25). The first sub-fraction has been subjected to an π-NAP HPLC (H2O/MeCN 77:23, 2.4 mL/min, 210 nm) to give chisomicine D (1, 0.5 mg 0.005%) and E (2, 0.5 mg 0.005%).
Chisomicine D (1): white amorphous solid; [α]D20 -15o (c 0.3, MeOH); IR (KBr) νmax 3391, 2972, 1735, 1703 and 1268 cm–1; UV (MeOH) λmax (log ε) 203 (3.82) and 274 (2.69); CD (MeOH) λmax (Δε) 226 (0.03), 230 (0) and 244 (0.04); 1H and 13C NMR data (Table 1); ESIMS m/z 577 (M+Na)+; HRESIMS m/z 555.2551 (M+H; calcd for C31H39O9, 555.2589).
Chisomicine E (2): white amorphous powder; [α]D20 +18o (c 0.2, MeOH); IR (KBr) νmax 3391, 2972, 1735 and 1268 cm–1; UV (MeOH) λmax (log ε) 203 (3.77) and 272 (2.92); CD (MeOH) λmax (Δε) 222 (0.27), 257 (0.04), 275 (0) and 299 (0.07); 1H and 13C NMR data (Table 1); ESIMS m/z 579 (M+Na)+; HRESIMS m/z 557.2693 (M+H; calcd for C31H41O9, 557.2745).

ACKNOWLEDGEMENTS
This work was supported by a Grant from a collaborative research between ICSN, CNRS, France and University of Malaya and a Grant from CENAR. This work was also partly supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan, and a grant from the Open Research Center Project.

References

1. B. Oliver-Bever, Medicinal Plants in Tropical West Africa, Cambridge University Press: 1986; p. 375. CrossRef
2. H. Harms, Die natuerlichen Pflanzenfamilien. Leipzig: Engelmann, 1940.
3. A. N. Muellner, R. Samuel, M. W. Chase, and A. Coleman, Taxon, 2008, 57, 1.
4. G. R. Pettit, D. H. R. Barton, C. L. Herald, J. Polonsky, J. M. Schmidt, and J. D. Connolly, J. Nat. Prod., 1983, 46, 379. CrossRef
5. M. Nakatani In Limonoids from Meliaceae and their biological activities, 2001; Taylor & Francis Ltd.: 2001; p. 527.
6. C. L. Cespedes, J. S. Calderon, and E. Aranda In Insecticidal effects of limonoids from Mexican Cedrela spp (meliaceae), 2000; American Chemical Society: 2000; AGRO-078.
7. J. Wang, M. Dong, and Y. Guo, Guoji Yaoxue Yanjiu Zazhi, 2009, 36, 321.
8. A. Roy and S. Saraf, Biol. Pharm. Bull., 2006, 29, 191. CrossRef
9. P. S. Kumar, D. Mishra, G. Ghosh, and C. S. Panda, Ann. Biol. Res., 2010, 1, 24.
10. M. Miyake and Y. Ifuku, Foods Food Ingredients J. Jpn., 1996, 168, 83.
11. K. Mohamad, Y. Hirasawa, C. S. Lim, K. Awang, A. H. A. Hadi, K. Takeya, and H. Morita, Tetrahedron Lett., 2008, 49, 4276. CrossRef
12. K. Mohamad, Y. Hirasawa, M. Litaudon, K. Awang, A. H. A. Hadi, K. Takeya, W. Ekasari, A. Widyawaruyanti, N. C. Zaini, and H. Morita, Bioorg. Med. Chem., 2009, 17, 727. CrossRef
13. C. P. Wong, M. Shimada, Y. Nagakura, A. E. Nugroho, Y. Hirasawa, T. Kaneda, K. Awang, A. H. A. Hadi, K. Mohamad, M. Shiro, and H. Morita, Chem. Pharm. Bull., 2011, 59, 407. CrossRef
14. I. A. Najmuldeen, A. H. A. Hadi, K. Awang, K. Mohamad, K. A. Ketuly, M. R. Mukhtar, S.-L. Chong, G. Chan, M. A. Nafiah, N. S. Weng, O. Shirota, T. Hosoya, A. E. Nugroho, and H. Morita, J. Nat. Prod., 2011, 74, 1313. CrossRef
15. I. A. Najmuldeen, H. A. H. Abdul, K. Awang, K. Mohamad, and S. W. Ng, Acta Crystallogr., Sect. E: Struct. Rep. Online, 2010, E66, o1927.
16. K. Awang, C. S. Lim, K. Mohamad, H. Morita, Y. Hirasawa, K. Takeya, O. Thoison, and A. H. A. Hadi, Bioorg. Med. Chem., 2007, 15, 5997. CrossRef