Cytotoxic Chromenes from Myriactis Humilis

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Cytotoxic Chromenes from Myriactis humilis

Article in Planta Medica · May 2005 DOI: 10.1055/s-2005-864107 · Source: PubMed

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Jih-Jung Chen National Yang Ming University

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Jih-Jung Chen1, Chang-Yih Duh2, Ih-Sheng Chen3

Abstract

Two new chromenes, myriachromene and 2,2-dimethyl-6,7-methylenedioxy-2H-chromene, together with ten known compounds including a-tocopherylquinone, a-tocopherol acetate, diphenyl sulfone, b-carotene, chondrillasterol, b-sitosterol, b-si-

Letter tostenone, vanillin, and vanillic acid were isolated from the whole plant of Myriactis humilis. The structures of these new compounds were determined through spectral analyses. Among

the isolates, four compounds exhibited cytotoxicity (ED50 values <4mg/mL) against P-388 or HT-29 cell lines in vitro.

Myriactis humilis Merr. (Compositae) is a perennial herb found in the Philippines, Borneo, New Guinea, and throughout the medi- um-to-high altitude mountains of Taiwan [1]. The chemical constituents and biological activities of this plant have never been studied. In our continuing studies of the cytotoxic constituents of Formosan plants, over 1000 species have been screened for in vitro cytotoxicity to date, and Myriactis humilis has been found to be one of the active species. Investigation of chloroform-soluble fraction of the whole plant of this species has led to the isolation of two new (1 and 2) and ten known compounds. The structural elucidation of 1 and 2 and the cytotoxic properties of the isolates are described herein.

370 Myriachromene (1) was isolated as light yellow oil. The EI-MS af- ton [d = 6.36 (1H, s)]. Because the presence of NOESY correla- forded the positive ion [M]+ at m/z = 234, implying a molecular tions (Fig.1) could be observed between H-5 [d = 6.36 (1H, s)]

formula of C13H14O4, which was confirmed by the HR-EI-MS. The and H-4 [d = 6.22 (1H, d, J = 10.0 Hz)] and OMe-6 [d = 3.89 UV absorptions at 220, 279 and 314 nm were similar to those of (3H, s)], the methylenedioxy group [d = 5.94 (2H, s)] was reason- prunichromene A [2], and suggested the presence of a 6,7,8- ably assigned to C-7,8. According to the above data, the structure trioxygenated 2H-chromene nucleus. The presence of a methy- of 1 was elucidated as 6-methoxy-2,2-dimethyl-7,8-methylene- lenedioxy group in the molecule was revealed by the bands at dioxy-2H-chromene, which was further confirmed by 1H-1H 1055 and 940 cm-1 in the IR spectrum. An aromatic ring C = C COSY and NOESY experiments (Fig.1). The assignment of the stretch was revealed by IR absorptions at 1609 and 1503 cm±1. 13C-NMR resonances was confirmed by the HSQC and HMBC The 1H-NMR spectrum of 1 showed the presence of a dimethyl- techniques (Fig.1), which also supported the structure of 1. chromene ring [d = 1.44 (6H, s, Me2), 5.53, 6.22 (each 1H, each d, J = 10.0 Hz, H-3 and H-4)], a methoxy group [d = 3.89 (3H, s)], 2,2-Dimethyl-6,7-methylenedioxy-2H-chromene (2) was first a methylenedioxy group [d = 5.94 (2H, s)], and an aromatic pro- isolated from a natural source in this study, although it has been synthesized by Iyer et al. [3]. The structure of 2 was readily identified by comparison of its spectroscopic data (1H-NMR, IR, and Affiliation: 1 Department of Pharmacy, Tajen Institute of Technology, Pingtung, mass spectrometry data) with literature values [3]; and further Taiwan, Republic of China ´ 2 Institute of Marine Resources, National Sun Yat- 1 1 sen University, Kaohsiung, Taiwan, Republic of China ´ 3 School of Pharmacy, confirmed by H- H COSY, NOESY [Fig. 2 (a)], HSQC, and HMBC Kaohsiung Medical University, Kaohsiung, Taiwan, Republic of China [Fig. 2 (b)] experiments.

Correspondence: Dr. J. J. Chen ´ Department of Pharmacy ´ Tajen Institute of 25 Technology ´ Pingtung ´ Taiwan 907 ´ Republic of China ´ Fax: +886-8-762- The known isolates, including a-tocopherylquinone (3){[a]D : 5308 ´ E-mail: [email protected] 25 + 2.38 (c 0.16, CHCl3)} [4], a-tocopherol acetate (4){[a]D : + 3.18 Prof. Dr. I. S. Chen ´ School of Pharmacy ´ Kaohsiung Medical University ´ Kaoh- (c 0.23, CHCl )} [5], diphenyl sulfone (5) [6], a tetraterpenoid, b- siung ´ Taiwan 807 ´ Republic of China ´ Fax: +886-7-321-0683 ´ E-mail: 3 25 [email protected] carotene (6) [7], three steroids, chondrillasterol (7){a]D : ±3.88 (c 0.12, CHCl )} [8], b-sitosterol (8){[a]25: ±36.68 (c 0.15, CHCl )} [9], Received: July 12, 2004 ´ Accepted: December 12, 2004 3 D 3 26 b-sitostenone (9){[a]D :+81.28 (c 0.2, CHCl3)} [9], three benze- Bibliography: Planta Med 2005; 71: 370±372 ´ Georg Thieme Verlag KG noids, 4-prenyloxybenzaldehyde (10) [10], vanillin (11) [11], and Stuttgart ´ New York ´ DOI 10.1055/s-2005-864107 ´ ISSN 0032-0943 vanillic acid (12) [11], were easily identified by comparison of Table 1 Cytotoxic effects of compounds isolated from Myriactis humilis against P-388 and HT-29 cell linesa

±1 Compound ED50 [mgmL ] P-388 HT-29

Mithramycinb 0.06 0.08 Fig. 1 Significant NOESY (a) and HMBC (b) correlations of 1. Myriachromene (1) 3.85 2.43 2,2-Dimethyl-6,7-methylenedioxy- 11.2 3.10 2H-chromene (2) a-Tocopherylquinone (3) 2.65 7.15 a-Tocopherol acetate (4) > 50 > 50 Diphenyl sulfone (5) 3.79 10.4

b-Carotene (6) > 50 > 50 Letter Chondrillasterol (7) 16.4 25.9 b-Sitosterol (8) > 50 > 50 b-Sitostenone (9) 15.3 > 50 4-Prenyloxybenzaldehyde (10) 10.5 12.6 Fig. 2 Significant NOESY (a) and HMBC (b) correlations of 2. Vanillin (11) > 50 > 50 Vanillic acid (12) > 50 > 50

a An ED value £ 4.0 mgmL±1 is considered to indicate significant cytotoxicity [12]. 1 50 physical and spectroscopic data (UV, IR, H-NMR, [a]D, and mass b Positive control. spectrometry data) with those of corresponding authentic sam- ples or literature values. mg) was chromatographed on silica gel (17.5 g) and eluted with The cytotoxic effects of the isolates from the whole plant of For- n-hexane-EtOAc (3:1) to give 11 (3.5 mg) and 12 (4.3 mg). mosan M. humilis were tested in vitro against P-388 and HT-29 cell lines. The cytotoxicity data are shown in Table 1. The clinical- Cytotoxic assays were carried out in accordance with methods ly applied anticancer agent, mitramycin, was used as the refer- discussed in the literature [13], [14]. ence compound. An ED50 value £ 4.0 mg/mL is considered to indicate significant cytotoxicity [12]. Compounds 1, 2, 3, and 5 exhib- Myriachromene (1): Colorless oil; UV (MeOH): lmax (log e) = 220 ited cytotoxicity with ED50 values < 4 mg/mL against P-388 and/or (4.60), 279 (4.15), 314 (4.00) nm; IR:umax = 1609, 1503 (aromatic ±1 HT-29 cell lines. The chromene compounds 1 and 2 were more ring C = C stretch), 1055, 940 cm (OCH2O); EI-MS: m/z (rel. 371 effective against the P-388 cell line than the HT-29 cell line. int.) = 234 (M+, 22), 219 (100), 204 (7), 189 (5), 159 (6), 131 (11), + 77 (10); HR-EI-MS: m/z = 234.0892 [M] , calcd. for C13H14O4 : 1 234.0892; H-NMR (CDCl3, 500 MHz): d = 1.44 (6H, s, Me-22), Materials and Methods 3.89 (3H, s, OMe-6), 5.53 (1H, d, J = 10.0 Hz, H-3), 5.94 (2H, s, 13 OCH2O), 6.22 (1H, d, J = 10.0 Hz, H-4), 6.36 (1H, s, H-5); C-NMR

The whole plant of M. humilis was collected from Southern-Tawu (CDCl3,125MHz):d = 27.8 (Me-22), 56.0 (OMe), 77.9 (C-2), 97.1 mountain, Pingtung County, Taiwan, in August, 2001, and identi- (C-5), 99.9 (OCH2O), 113.1 (C-4a), 117.2 (C-4), 127.8 (C-3), 136.2 (C- fied by Dr. I. S. Chen. A voucher specimen (Chen 6019) was depos- 7), 149.3 (C-8), 149.5 (C-6), 153.9 (C-8a). ited in the herbarium of the School of Pharmacy, Kaohsiung Med- ical University, Kaohsiung, Taiwan, Republic of China. 2,2-Dimethyl-6,7-methylenedioxy-2H-chromene (2): Yellow nee-

dles from CH2Cl2-MeOH, m.p. 82±848C; HR-EI-MS: m/z = + The dried whole plant (2.0 kg) was extracted with cold MeOH, 204.0786 [M] , calcd. for C12H12O3 :204.0787. and the extract concentrated under reduced pressure. The

MeOH extract (180 g), when partitioned between H2O-CHCl3

(1:1), afforded a CHCl 3-soluble fraction (fr. A, 26.4 g). Fr. A (26.4 Acknowledgements g) was chromatographed on silica gel (980 g) and eluted with n- hexane. Polarity was gradually increased with EtOAc to give 14 This study was supported by a grant (NSC 90-2320-B-127-001) frs. Fr. A2 (512 mg) was chromatographed on silica gel (18 g) from the National Science Council of the Republic of China and eluted with n-hexane-EtOAc (20 :1) to give 5 (3.4 mg). Fr. A3 (415 mg) was chromatographed on silica gel (15.5 g) and eluted with n-hexane-EtOAc (9:1) to obtain 4 (7.8 mg) and 10 References (4.8 mg). Fr. A4 (388 mg) was chromatographed on silica gel (11 1 Peng CI, Chung KF, Li HL. Compositae in Flora of Taiwan. 2nd edn. Edi- g) and eluted with n-hexane-CH2Cl2 (2:1) to give 1 (2.5 mg), 2 (3.3 mg), and 9 (5.3 mg). Fr. A5 (674 mg) was chromatographed torial Committee of the Flora of Taiwan, Taipei, Taiwan: 1993; Vol. 4: pp 1017±9 on silica gel (24 g) and eluted with n-hexane-EtOAc (3:1) to ob- 2 Ober AG, Urbatsch LE, Fischer NH. Guaianolides and chromenes from tain 3 (4.5 mg), 6 (3.7 mg), 7 (5.2 mg), and 8 (6.8 mg). Fr. A6 (529 Calea species. Phytochemistry 1985; 24: 795 ±9

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Letter ¼ Planta Med2005; 71: 370±372

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