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O-terpenoidal coumarins from Clausena anisum-olens
Biochemical Systematics and Ecology 36 (2008) 801–803
Contents lists available at ScienceDirect
Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco
O-terpenoidal coumarins from Clausena anisum-olens Yun-Song Wang, Rong Huang, Liang Li, Hong-Bin Zhang, Jing-Hua Yang* Key Laboratory of Medicinal Chemistry for Natural Resource, Ministry of Education, School of Chemical Science and Technology, Yunnan University, Kunming 650091, P.R. China
a r t i c l e i n f o Article history: Received 10 April 2008 Accepted 7 June 2008 Keywords: Rutaceae Clausena anisum-olens Hekumarin A and B Monoterpenoid coumarins
1. Subject and source The plants of the genus Clausena (Rutaceae) are shrubs widely distributed in south and southeast Asia and most of them are used as a Chinese folk medicine (Wu, 1995). Clausena anisum-olens is a shrub growing wild and cultivated from Philippines and South China through southeast Asia and the aerial parts of this plant have been used for the treatment of dysentery and arthritis (Wu, 1995). The leaves and twigs of C. anisum-olens were collected at Hekou County in Yunnan province, P.R. China, in May 2003 and identified by Professor De-Ding Tao of Kunming Institute of Botany. A voucher specimen (No. 02041705) is deposited in State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences. 2. Previous work Previous phytochemical studies indicate that carbazole alkaloids (Chakraborty et al., 1995; Wu et al., 1996, 1997) and coumarins (He et al., 2000; Ito et al., 2000; Nakamura et al., 1998) are common in the genus Clausena. To the best of our knowledge, no previous phytochemical studies on the species C. anisum-olens growing in Yunnan Province have been reported to date. 3. Present study The powdered plant material of C. anisum-olens (22.5 kg) was repeatedly extracted with EtOH at room temperature. The extract was then concentrated under reduced pressure to give a brown syrup, which was suspended in water and was partitioned with petroleum ether, ethyl acetate (EtOAc) and n-butanol (n-BuOH), successively. The EtOAc fraction (110.5 g) was subjected to silica gel column chromatography eluting with PE-AcOEt (4:1 / 2:3), EtOAc, EtOAc–MeOH (8:2 / 1:1), MeOH, * Corresponding author. Tel.: þ86 871 6598387; fax: þ86 871 5035538. E-mail address: [email protected] (J.-H. Yang). 0305-1978/$ – see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2008.06.005
802
Y.-S. Wang et al. / Biochemical Systematics and Ecology 36 (2008) 801–803
by which nine fractions (I–IX) were obtained. Fraction IV was resubmitted to silica gel column chromatography, Pharmadex LH-20 and RP C-18 to yield compounds 1/2 (6 mg) as a pair of inseparable epimers. Two new O-terpenoidal coumarins 1 and 2, named hekumarin A and B, were isolated from C. anisum-olens. We here report on their isolation and structural identification. Compounds 1/2 were isolated as a light yellow oil. Its molecular formula C20H20O7 was deduced from the HR-ESI-MS exhibiting the molecular ion at m/z 373.1298 [M]þ. Strong UV bands at lmax 256 and 318, an IR band at 1730 cm1, and two pairs of typical AB system doublets at dH 6.33 and 7.71 and dH 7.36, 6.97 in the 1H NMR spectrum (Table 1) indicated the presence of a 7,8-dioxygenated coumarin (Ito et al., 2000). The inspection of NMR data revealed the presence of a C-10 terpenoid side chain containing a g-lactone and a methoxyl group (dC 61.5q) besides the coumarin nucleus in 1/2 (Table 1). The 1H–13C long-range correlations between H-10 and C-20 , C-7 and C-30 proved that a C-10 terpenoid side chain was linked with C-7 and the correlations between the protons of methoxyl group (dH 4.02) and C-8 (dC 136.3s) indicated that the methoxyl group was located at C-8. Two olefinic protons on terminal methylene at d 5.28, 5.43 (each 1H, s) were attributed to H-100 according to signal complexity and HMBC correlations between H-20 and C-30 and C-100 . The IR band at 3453 cm1 indicated the presence of hydroxyl groups. In the monoterpenoid side chain, a 13C NMR signals at dC 72.7d revealed the presence of hydroxyl-substituted carbon and the HMBC correlations between H-20 and C-30 , C-40 and C-100 indicated the hydroxyl group was attached to C-20 (Fig. 1). The difference between 1 and 2 was due to stereochemistry of hydroxyl group at C-20 . The NMR data of C-10, C-20 , C-30 and C-40 appeared in pairs (Table 1), indicating the presence of 1 and its C-20 stereoisomer 2. The NMR spectra established that 1 and 2 consisted of two epimers in a 3:1 ratio. In the NOESY spectrum, only the major epimer 1 showed a significant cross peak between H-2’ and H-4b0 and the same NOE correlations didn’t be observed in the minor epimer 2. This suggested cis arrangement of both functions whereas 2 exhibited the corresponding trans arrangement (Thuy et al., 1999). The evidences support the presence of a pair of epimers 1/2 instead of different conformations of one compound. On the basis of these results, the structures of epimers 1/2 were proposed as shown in Fig 1. The absolute configurations at C-20 and C-50 of these two coumarins remain to be determined. So far, the stereochemical problems of this type of O-terpenoidal coumarins reported previously remained open in the present study (Wu et al., 1997; Nakamura et al., 1998; Thuy et al., 1999; Ito et al., 2005). 3.1. Hekumarin A and B (1/2) 1 Light yellow oil. [a]21.8 3453, 2936, 2820, 1743, 1730, 1608; UV (MeOH) nm: D ¼ 19.08 (c 0.76, CH3OH). IRnmax (KBr) cm 387, 318, 256; 1H-NMR and 13C-NMR(d ppm, CDCl3, 500 MHz) see Table 1; EI-MS m/z 372 ([M]þ, 75), 342 (2), 275 (5), 206 (20), 192 (100), 164 (15); HR-ESI-MS m/z 373.1298 ([M þ 1]þ) (C20H20O7; calc. 373.1287).
4. Chemotaxonomic significance The plants of the Rutaceae family are one of the richest sources of coumarins (Phuwapraisirisan et al., 2006; Chlouchi et al., 2005; Na´jera and Yus, 2000). In this family, plants of Clausena genus are widely distributed in the south of China and the most Table 1 The 1H- and Position
2 3 4 5 6 7 8 9 10 10 a 10 b 20 30 40 a 40 b 50 60 70 80 90 100 a 100 b OMe
13
C-NMR data for Compounds 1–2 (d in ppm, J in Hz, in CD3Cl3) 1 (major isomer)
2 (minor isomer)
dH
dC
– 6.33 (d, J ¼ 9.8) 7.71 (d, J ¼ 9.8) 7.36 (d, J ¼ 8.6) 6.97 (d, J ¼ 8.6) – – – – 4.23 (m) 4.14 (m) 4.66 (m) – 2.63 (dd, J ¼ 14.6, 7.3) 2.54 (dd, J ¼ 14.6, 6.4) 5.25 (m) 7.22 (d, J ¼ 1.7) – – 1.98 (s) 5.43 (s) 5.28 (s) 4.02 (s)
160.4 113.6 143.6 123.0 110.4 154.3 136.3 114.1 147.9 72.8 72.8 72.7 142.0 36.4 36.4 79.8 148.7 130.0 174.0 10.5 116.2 116.2 61.5
(s) (d) (d) (d) (d) (s) (s) (s) (s) (t) (t) (d) (s) (t) (t) (d) (d) (s) (s) (q) (t) (t) (q)
dH
dC
– 6.33 (d, J ¼ 9.8) 7.71 (d, J ¼ 9.8) 7.36 (d, J ¼ 8.6) 6.97 (d, J ¼ 8.6) – – – – 4.24 (m) 4.17 (m) 4.68 (m) – 2.69 (dd, J ¼ 14.6, 5.1) 2.45 (dd, J ¼ 14.6, 8.1) 5.25 (m) 7.22 (d, J ¼ 1.7) – – 1.98 (s) 5.45 (s) 5.28 (s) 4.02 (s)
160.4 113.6 143.6 123.0 110.4 154.3 136.3 114.1 147.9 72.8 72.8 72.6 142.3 36.4 36.4 80.0 148.7 130.0 174.0 10.5 115.9 115.9 61.5
(s) (d) (d) (d) (d) (s) (s) (s) (s) (t) (t) (d) (s) (t) (t) (d) (d) (s) (s) (q) (t) (t) (q)
Y.-S. Wang et al. / Biochemical Systematics and Ecology 36 (2008) 801–803
803
Fig. 1. The structures, Key HMBC correlations and selected NOESY of 1 and 2.
of them are used as a Chinese folk medicine (Wu, 1995.). It is documented that coumarins are one of the major constituents in plants belonging to the genus Clausena (He et al., 2000; Ito et al., 2000; Nakamura et al., 1998). The result of the present study supports the conclusion that coumarins are characteristic constituents for the family Rutaceae, especially, the genus Clausena. Acknowledgements This work was supported by Science Foundation of Yunnan University (Grant No. 2004Q004A and 2005Z001A) and Science Foundation of Yunnan (Grant No. 2006B0003Q). References Chakraborty, A., Chowdhury, K., Bhattacharyya, P., 1995. Phytochemistry 40, 295. Chlouchi, A., Muyard, F., Girard, C., Waterman, P.G., Be´valot, F., 2005. Biochem. Syst. Ecol. 33, 967. He, H.P., Shen, Y.M., He, Y.N., Yang, X.S., Zhu, W.M., Hao, X.J., 2000. Heterocycles 53, 2067. Ito, C., Itoigawa, M., Katsuno, S., Omura, M., Tokuda, H., Nishino, H., Furukawa, H., 2000. J. Nat. Prod. 63, 1218. Ito, C., Itoigawa, M., Onoda, S., Hosokawa, A., Ruangrungsi, N., Okuda, T., Tokuda, H., Nishino, H., Furukawa, H., 2005. Phytochemistry 66, 567. Na´jera, C., Yus, M., 2000. Stud. Nat. Prod. Chem. 21, 373. Nakamura, K., Takemura, Y., Ju-ichi, M., Ito, C., Furukawa, H., 1998. Heterocycles 48, 549. Phuwapraisirisan, P., Surapinit, S., Sombund, S., Siripong, P., Tip-pyang, S., 2006. Tedrahedron Lett. 47, 3685. Thuy, T.T., Ripperger, H., Porzel, A., Sung, T.V., Adam, G., 1999. Phytochemistry 52, 511. Wu, T.S., Huang, S.C., Wu, P.L., 1996. Tedrahedron Lett. 37, 7819. Wu, T.S., Huang, S.C., Wu, P.L., 1997. Heterocycles 45, 969. Wu, Z.Y. (Ed.), 1995. Institutum Botanicum Kunmingense Academiae Sinicae Edita. Flora Yunnanica, vol. 6. Science Press, Beijing, p. 767.
Contents lists available at ScienceDirect
Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco
O-terpenoidal coumarins from Clausena anisum-olens Yun-Song Wang, Rong Huang, Liang Li, Hong-Bin Zhang, Jing-Hua Yang* Key Laboratory of Medicinal Chemistry for Natural Resource, Ministry of Education, School of Chemical Science and Technology, Yunnan University, Kunming 650091, P.R. China
a r t i c l e i n f o Article history: Received 10 April 2008 Accepted 7 June 2008 Keywords: Rutaceae Clausena anisum-olens Hekumarin A and B Monoterpenoid coumarins
1. Subject and source The plants of the genus Clausena (Rutaceae) are shrubs widely distributed in south and southeast Asia and most of them are used as a Chinese folk medicine (Wu, 1995). Clausena anisum-olens is a shrub growing wild and cultivated from Philippines and South China through southeast Asia and the aerial parts of this plant have been used for the treatment of dysentery and arthritis (Wu, 1995). The leaves and twigs of C. anisum-olens were collected at Hekou County in Yunnan province, P.R. China, in May 2003 and identified by Professor De-Ding Tao of Kunming Institute of Botany. A voucher specimen (No. 02041705) is deposited in State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences. 2. Previous work Previous phytochemical studies indicate that carbazole alkaloids (Chakraborty et al., 1995; Wu et al., 1996, 1997) and coumarins (He et al., 2000; Ito et al., 2000; Nakamura et al., 1998) are common in the genus Clausena. To the best of our knowledge, no previous phytochemical studies on the species C. anisum-olens growing in Yunnan Province have been reported to date. 3. Present study The powdered plant material of C. anisum-olens (22.5 kg) was repeatedly extracted with EtOH at room temperature. The extract was then concentrated under reduced pressure to give a brown syrup, which was suspended in water and was partitioned with petroleum ether, ethyl acetate (EtOAc) and n-butanol (n-BuOH), successively. The EtOAc fraction (110.5 g) was subjected to silica gel column chromatography eluting with PE-AcOEt (4:1 / 2:3), EtOAc, EtOAc–MeOH (8:2 / 1:1), MeOH, * Corresponding author. Tel.: þ86 871 6598387; fax: þ86 871 5035538. E-mail address: [email protected] (J.-H. Yang). 0305-1978/$ – see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2008.06.005
802
Y.-S. Wang et al. / Biochemical Systematics and Ecology 36 (2008) 801–803
by which nine fractions (I–IX) were obtained. Fraction IV was resubmitted to silica gel column chromatography, Pharmadex LH-20 and RP C-18 to yield compounds 1/2 (6 mg) as a pair of inseparable epimers. Two new O-terpenoidal coumarins 1 and 2, named hekumarin A and B, were isolated from C. anisum-olens. We here report on their isolation and structural identification. Compounds 1/2 were isolated as a light yellow oil. Its molecular formula C20H20O7 was deduced from the HR-ESI-MS exhibiting the molecular ion at m/z 373.1298 [M]þ. Strong UV bands at lmax 256 and 318, an IR band at 1730 cm1, and two pairs of typical AB system doublets at dH 6.33 and 7.71 and dH 7.36, 6.97 in the 1H NMR spectrum (Table 1) indicated the presence of a 7,8-dioxygenated coumarin (Ito et al., 2000). The inspection of NMR data revealed the presence of a C-10 terpenoid side chain containing a g-lactone and a methoxyl group (dC 61.5q) besides the coumarin nucleus in 1/2 (Table 1). The 1H–13C long-range correlations between H-10 and C-20 , C-7 and C-30 proved that a C-10 terpenoid side chain was linked with C-7 and the correlations between the protons of methoxyl group (dH 4.02) and C-8 (dC 136.3s) indicated that the methoxyl group was located at C-8. Two olefinic protons on terminal methylene at d 5.28, 5.43 (each 1H, s) were attributed to H-100 according to signal complexity and HMBC correlations between H-20 and C-30 and C-100 . The IR band at 3453 cm1 indicated the presence of hydroxyl groups. In the monoterpenoid side chain, a 13C NMR signals at dC 72.7d revealed the presence of hydroxyl-substituted carbon and the HMBC correlations between H-20 and C-30 , C-40 and C-100 indicated the hydroxyl group was attached to C-20 (Fig. 1). The difference between 1 and 2 was due to stereochemistry of hydroxyl group at C-20 . The NMR data of C-10, C-20 , C-30 and C-40 appeared in pairs (Table 1), indicating the presence of 1 and its C-20 stereoisomer 2. The NMR spectra established that 1 and 2 consisted of two epimers in a 3:1 ratio. In the NOESY spectrum, only the major epimer 1 showed a significant cross peak between H-2’ and H-4b0 and the same NOE correlations didn’t be observed in the minor epimer 2. This suggested cis arrangement of both functions whereas 2 exhibited the corresponding trans arrangement (Thuy et al., 1999). The evidences support the presence of a pair of epimers 1/2 instead of different conformations of one compound. On the basis of these results, the structures of epimers 1/2 were proposed as shown in Fig 1. The absolute configurations at C-20 and C-50 of these two coumarins remain to be determined. So far, the stereochemical problems of this type of O-terpenoidal coumarins reported previously remained open in the present study (Wu et al., 1997; Nakamura et al., 1998; Thuy et al., 1999; Ito et al., 2005). 3.1. Hekumarin A and B (1/2) 1 Light yellow oil. [a]21.8 3453, 2936, 2820, 1743, 1730, 1608; UV (MeOH) nm: D ¼ 19.08 (c 0.76, CH3OH). IRnmax (KBr) cm 387, 318, 256; 1H-NMR and 13C-NMR(d ppm, CDCl3, 500 MHz) see Table 1; EI-MS m/z 372 ([M]þ, 75), 342 (2), 275 (5), 206 (20), 192 (100), 164 (15); HR-ESI-MS m/z 373.1298 ([M þ 1]þ) (C20H20O7; calc. 373.1287).
4. Chemotaxonomic significance The plants of the Rutaceae family are one of the richest sources of coumarins (Phuwapraisirisan et al., 2006; Chlouchi et al., 2005; Na´jera and Yus, 2000). In this family, plants of Clausena genus are widely distributed in the south of China and the most Table 1 The 1H- and Position
2 3 4 5 6 7 8 9 10 10 a 10 b 20 30 40 a 40 b 50 60 70 80 90 100 a 100 b OMe
13
C-NMR data for Compounds 1–2 (d in ppm, J in Hz, in CD3Cl3) 1 (major isomer)
2 (minor isomer)
dH
dC
– 6.33 (d, J ¼ 9.8) 7.71 (d, J ¼ 9.8) 7.36 (d, J ¼ 8.6) 6.97 (d, J ¼ 8.6) – – – – 4.23 (m) 4.14 (m) 4.66 (m) – 2.63 (dd, J ¼ 14.6, 7.3) 2.54 (dd, J ¼ 14.6, 6.4) 5.25 (m) 7.22 (d, J ¼ 1.7) – – 1.98 (s) 5.43 (s) 5.28 (s) 4.02 (s)
160.4 113.6 143.6 123.0 110.4 154.3 136.3 114.1 147.9 72.8 72.8 72.7 142.0 36.4 36.4 79.8 148.7 130.0 174.0 10.5 116.2 116.2 61.5
(s) (d) (d) (d) (d) (s) (s) (s) (s) (t) (t) (d) (s) (t) (t) (d) (d) (s) (s) (q) (t) (t) (q)
dH
dC
– 6.33 (d, J ¼ 9.8) 7.71 (d, J ¼ 9.8) 7.36 (d, J ¼ 8.6) 6.97 (d, J ¼ 8.6) – – – – 4.24 (m) 4.17 (m) 4.68 (m) – 2.69 (dd, J ¼ 14.6, 5.1) 2.45 (dd, J ¼ 14.6, 8.1) 5.25 (m) 7.22 (d, J ¼ 1.7) – – 1.98 (s) 5.45 (s) 5.28 (s) 4.02 (s)
160.4 113.6 143.6 123.0 110.4 154.3 136.3 114.1 147.9 72.8 72.8 72.6 142.3 36.4 36.4 80.0 148.7 130.0 174.0 10.5 115.9 115.9 61.5
(s) (d) (d) (d) (d) (s) (s) (s) (s) (t) (t) (d) (s) (t) (t) (d) (d) (s) (s) (q) (t) (t) (q)
Y.-S. Wang et al. / Biochemical Systematics and Ecology 36 (2008) 801–803
803
Fig. 1. The structures, Key HMBC correlations and selected NOESY of 1 and 2.
of them are used as a Chinese folk medicine (Wu, 1995.). It is documented that coumarins are one of the major constituents in plants belonging to the genus Clausena (He et al., 2000; Ito et al., 2000; Nakamura et al., 1998). The result of the present study supports the conclusion that coumarins are characteristic constituents for the family Rutaceae, especially, the genus Clausena. Acknowledgements This work was supported by Science Foundation of Yunnan University (Grant No. 2004Q004A and 2005Z001A) and Science Foundation of Yunnan (Grant No. 2006B0003Q). References Chakraborty, A., Chowdhury, K., Bhattacharyya, P., 1995. Phytochemistry 40, 295. Chlouchi, A., Muyard, F., Girard, C., Waterman, P.G., Be´valot, F., 2005. Biochem. Syst. Ecol. 33, 967. He, H.P., Shen, Y.M., He, Y.N., Yang, X.S., Zhu, W.M., Hao, X.J., 2000. Heterocycles 53, 2067. Ito, C., Itoigawa, M., Katsuno, S., Omura, M., Tokuda, H., Nishino, H., Furukawa, H., 2000. J. Nat. Prod. 63, 1218. Ito, C., Itoigawa, M., Onoda, S., Hosokawa, A., Ruangrungsi, N., Okuda, T., Tokuda, H., Nishino, H., Furukawa, H., 2005. Phytochemistry 66, 567. Na´jera, C., Yus, M., 2000. Stud. Nat. Prod. Chem. 21, 373. Nakamura, K., Takemura, Y., Ju-ichi, M., Ito, C., Furukawa, H., 1998. Heterocycles 48, 549. Phuwapraisirisan, P., Surapinit, S., Sombund, S., Siripong, P., Tip-pyang, S., 2006. Tedrahedron Lett. 47, 3685. Thuy, T.T., Ripperger, H., Porzel, A., Sung, T.V., Adam, G., 1999. Phytochemistry 52, 511. Wu, T.S., Huang, S.C., Wu, P.L., 1996. Tedrahedron Lett. 37, 7819. Wu, T.S., Huang, S.C., Wu, P.L., 1997. Heterocycles 45, 969. Wu, Z.Y. (Ed.), 1995. Institutum Botanicum Kunmingense Academiae Sinicae Edita. Flora Yunnanica, vol. 6. Science Press, Beijing, p. 767.