Two new steroidal glucosides from Ophiopogon japonicus (L.f.) Ker-Gawl

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Chinese Chemical Letters 19 (2008) 825–828 www.elsevier.com/locate/cclet

Two new steroidal glucosides from Ophiopogon japonicus (L.f.) Ker-Gawl Ya Jiuan Xu a, Tun Hai Xu b,c,*, Ling Zhu Hao a, Hong Feng Zhao a, Sheng Xu Xie a, Yun Shan Si a, Dong Han a, Dong Ming Xu a a

Academy of Traditional Chinese Medicine and Material Medica of Jilin Province, Changchun 130021,China b School of Traditional Chinese Medicine, Beijing University of Chinese Medicine, Beijing 100102,China c School of Traditional Chinese Medicine, Xinjiang University of Medicine, Urumchi 830011, China Received 29 December 2007

Abstract Two new steroidal glucosides, 26-O-b-D-glucopyranosyl (25S)-furost-5-ene-1b,3b,22a,26-tetraol 1-O-b-D-xylopyranosyl(1 ! 3)-[a-L-rhamnopyranosyl-(1 ! 2)]-b-D-fucopyranoside and (25R) spirost-5-ene-3b,14a-diol-3-b-O-b-L–rhamnopyranosyl(1 ! 2)-[b-D-xylopyranosyl(1 ! 4)]-b-D-glucopyranoside, were isolated from the Ophiopogon japonicus (L.f.) Ker-Gaw. Their structures were elucidated by spectroscopic methods. # 2008 Tun Hai Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. Keywords: Ophiopogon japonicus (L.f.) Ker-Gawl; Steroidal glucosides; Liliaceae

The tuber of Ophiopogon japonicus (L.f.) Ker-Gawl is a Chinese traditional medicine named ‘‘maidong’’. The tuber was recorded to have various medical functions for curing cardiovascular diseases and bacterial infections, especially heart diseases. Phytochemical studies on this plant were reported previously [1]. In the search for new and bioactive components from Chinese traditional medicine, we investigated the tubers of O. japonicus. In the present paper, we report the isolation and structure elucidation of two new steroidal glucosides by using 1D, 2D NMR techniques, ESI-MS analysis as well as chemical methods. Compound 1, amorphous powder, ½a20 D 29.5 (c 0.15, MeOH). Its IR spectrum (KBr, n) showed absorptions at 3612 (OH), 1625 (C C) cm1. An acidic hydrolysis of 1 with mineral acid afforded glucose, rhamnose, fucose and xylose as the sugar components. Its HRESI-MS showed [MH] at m/z 1033.52183 (calcd. 1033.52195), corresponding to the formula C50H82O22. Its ESI-MS showed significant ion peaks at m/z 887 (M-146-H), 901 (M132-H), 755 (M-132-146-H), and 609 (M-132-146-146-H). Its 1H NMR spectrum (Table 1) showed diagnostic signals of four methyl groups at d 0.90 (s, 3H, CH3-18), 1.39 (s, 3H, CH3-19), 1.22 (d, 3H, J = 7.0 Hz, CH3-21), 0.99 (d, 3H, J = 6.6 Hz, CH3-27), and three oxymethines at d 3.83 (m, 1H, H-1), 3.68 (m, 1H, H-3), 4.99 (m, 1H, H-16), one oxymethylene at d 3.47 (dd, 1H, J = 7.0, 9.5 Hz, H-26), 4.06 (m, 1H, H-26), and four anomeric protons at d 4.65 (d, 1H, J = 7.5Hz), 4.77 (d, 1H, J = 7.5Hz), 4.94 (d, 1H, J = 7.5 Hz), 6.34 (s, 1H). Its 13C NMR spectrum showed signals of

* Corresponding author. E-mail address: [email protected] (T.H. Xu). 1001-8417/$ – see front matter # 2008 Tun Hai Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. doi:10.1016/j.cclet.2008.04.033

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Table 1 13 C NMR and 1H NMR data of sugars 1 and 2 in C5D5N Compound 1

Compound 2

No.

dC

dH (J, Hz)

No.

dC

dH (J, Hz)

3-O-Fuc-1 2 3 4 5 6

100.4 74.7 85.5 72.7 71.6 17.1

4.65 (d, 7.5) 4.02 4.06 4.58 4.23 1.48 (d, 6.2)

3-O-Glc-1 2 3 4 5 6

100.1 77.4 77.7 81.7 76.4 61.8

4.82 (d, 7.7) 4.11 4.20 4.08 3.95 4.50, 4.28

Rha-1 2 3 4 5 6

101.7 70.6 72.5 74.2 68.2 19.1

6.34 (s) 4.21 4.77 4.07 4.85 1.72 (d, 6.5)

Rha-1 2 3 4 5 6

102.1 72.6 73.0 74.3 69.8 18.8

6.23 (br, s) 4.79 4.65 4.42 4.48 1.46 (d, 5.6)

Xyl-1 2 3 4 5

106.6 74.9 78.3 70.7 67.03

4.94 (d, 7.5) 3.43 3.95 3.75 4.25, 3.68

Xyl-1 2 3 4 5

105.9 75.1 78.5 70.9 67.0

4.99 (d, 7.0) 3.92 4.06 4.15 4.12, 3.56

26-O-Glc-1 2 3 4 5 6

105.1 75.3 78.5 69.3 78.4 62.7

4.77 (d, 7.5) 4.06 4.21 4.77 3.99 4.52, 4.33

four angular methyl groups, four carbons bearing a hydroxyl group and four anomeric carbons. In a comparison of the 13 C NMR signals for aglycone of 1 (Table 2) with those of known saponin 26-O-b-D-glucopyranosyl-(25S)furost-5ene-1b,3b,22a,26-tetraol-1-O-b-D-fucopyranoside (compound 113) [2], all signals due to the aglycone of 1 were almost superimposable with those of compound 113, indicating the aglycone of 1 was same as that of compound 113 and its 1- and 26-hydroxy groups carried a sugar moiety, respectively. As described above, the sugar moiety of 1 consisted of glucose, rhamnose, fucose and xylose. The coupling constants of the anomeric protons revealed the b configurations for glucoses, fucose and xylose, and a configurations

Fig. 1. The structure and HMBC of compounds 1 and 2.

Y.J. Xu et al. / Chinese Chemical Letters 19 (2008) 825–828

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Table 2 13 C NMR and 1H NMR data of aglycone moieties 1 and 2 in C5D5N Compound 1

Compound 2

No.

dC

dH (J, Hz)

No.

dC

dH (J, Hz)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

84.4 37.1 67.0 43.8 139.5 124.8 32.0 32.1 50.5 42.8 24.0 40.5 40.8 57.1 32.7 81.1 63.9 17.0 15.0 40.8 16.3 110.6 38.0 28.2 34.4 75.2 17.4

3.83 2.68, 2.41 3.68 2.68, 2.55

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

37.9 32.1 78.5 40.7 140.4 122.6 26.9 35.8 43.8 37.7 20.2 30.8 45.2 86.6 39.1 82.0 60.1 20.5 19.5 42.2 15.5 109.7 30.3 29.4 30.8 66.9 17.5

1.80, 1.10 2.05, 1.76 3.95 2.25 (dd, 7.5, 7.5)

5.56 (br, d) 1.54 1.45 1.56 1.45, 1.32 2.18, 1.61 1.22 1.88, 1.14 4.99 1.94 (dd, 8.5, 7.5) 0.90 (s) 1.39 (s) 2.16 1.22 (d, 7.0) 2.41, 2.27 1.63, 1.65 1.92 3.43, 4.06 0.99 (d, 6.6)

5.16 (br, d, 4.9) 2.40, 2.33 1.93 1.71 1.47, 1.39 1.72, 1.12

2.68 4.06 2.64 0.95 1.00 1.99 1.06

(dd, 8.5, 7.5) (dd, 8.5, 7.7) (s) (s) (d, 6.9)

1.96, 1.77 1.77, 1.78 1.58 3.40, 3.55 0.56 (d, 3.7)

for rhamnose [3,4]. The positions of the sugar residues in 1 were defined unambiguously by the HMBC experiment (Fig. 1). HMBC correlations between H-1 (d 4.65) of the fucose and C-1 (d 84.4) of the aglycone, H-1 (d 6.34) of rhamnose and C-2 (d 74.7) of the fucose, H-1 (d 4.94) of the xylose and C-3 (d 85.5) of fucose indicated that a trisaccharide moiety 1-O-b-D-xylopyranosyl-(1 ! 3)-[a-L-rhamnopyranosyl-(1 ! 2)]-b-D-fucopyranoside was linked to C-1 of the aglycone. Additionally, a HMBC correlation between H-1 (d 4.77) of glucose and C-26 (d 75.2) of the aglycone indicated that the glucose was linked to C-26 of the aglycone. On the basis of the above evidence, the structure of 1 was elucitated as 26-O-b-D-glucopyranosyl (25S)-furost-5-ene-1b,3b, 22a,26-tetraol 1-O-b-D-xylopyranosyl-(1 ! 3)[a-L-rhamnopyranosyl-(1 ! 2)]-b-D-fucopyranoside.  Compound 2 amorphous powder, ½a20 D 38.9 (c 0.15, MeOH). Its HRESI-MS showed [MH] at m/z 869.4547 (calcd. 869.4534), corresponding to the formula C44H70O17. Its ESI-MS showed significant ion peaks at m/z 737 (M132-H), 591 (M-132-146-H). The 1H NMR spectra of two showed signals ascribable three tertiary methyl groups (dH 0.56, 1.06, 0.95, 1.00), a olefinic proton (dH 5.16), three anomeric protons (dH 4.82, 6.23, 4.99), and a characteristic doublet of the 6-deoxyhexopyranosyl group (dH 1.46). The 13C NMR spectrum data displayed 44 resonance lines, 17 of which could be due to the monosaccharide components and 27 due to the aglycone part, including two olefinic carbons, three oxygenated methines, and three anomeric carbons. These observations suggested that 2 was steroidal saponin. Additionally, the 13C NMR spectral data of the aglycone of 2 were almost consistent with those of known sapogenin (25R) spirost-5-ene-3b,14a-diol (compound 59) [2], except that the signal of C-3 (d 78.3) was deshielded by Dd 7.0, and signals of C-2 (d 32.1) and C-4 (d 42.2) were shielded by Dd 0.6 and 1.3, respectively. Thus, the structure of the aglycon as (25R) spirost-5-ene-b,14a-diol, and the 3-b-hydroxy group of the aglycone was glycosidated.

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An acidic hydrolysis of 2 with mineral acid afforded glucose, rhamnose and xylose as the sugar components. The signals of three anomeric protons [d 4.82 (1H, d, J = 7.7 Hz, glc H1), 4.99 (1H, d, J = 7.6 Hz, xyl H1), 6.23 (1H, br s, rha H1)], and a secondary methyl signals ascribable to 6-deoxyhexose [d 1.46 (3H, d, J = 5.6 Hz)], indicated that 2 had three sugars including a 6-deoxyhexose. These coupling constants indicated that the glycosidic linkage of glucose and xylose were b-configuration [3,4], and rhamnose was a-configuration [3,4]. The positions of the sugar residues in 2 were defined unambiguously by HMBC experiment (Fig. 1). HMBC correlation between H-1 (d 4.82) of glucose and C-3 (d 78.3) of the aglycon, H-1 (d 6.23) of rhamnose and C-2 (d 77.4) of glucose, H-I (d 4.99) of xylose and C-4 (d 81.7) of glucose. These assignments showed that a trisaccharide moiety, 3-O-b-L-rhamnopyranosyl(1 ! 2)-[b-Dxylopyranosyl(1 ! 4)]-b-D-glucopyranoside, was inked to the at C-3. On the basis of all of these evidences, 2 was identified as (25R) spirost-5-ene-3b,14a-diol-3-b-O-b-L-rhamnopyranosyl(1 ! 2)-[b-D-xylopyranosyl(1 ! 4)]-bD-glucopyranoside. Acknowledgments The authors are grateful to the National Nature Science Foundation of China (No. 30772890) and the Program of Progressing Beijing New Medicine Subject Group (XK100270569) and the Key Project of Chinese Ministry of Education (No. 108132). References [1] [2] [3] [4]

G. Xiao, Modern Chinese Materia Medica, vol. I, Chemical Industry Press, Beijing, 2001, p. 481. P.K. Agrawal, D.C. Jain, A.K. Pathak Pathak, et al. Magentic Resonance in Chemistry 33 (1995) 923. P.K. Angarawa, Phytochemistry 31 (10) (1992) 3307. P.K. Agrawal, D.C. Jain, R.K. Gupta, et al. Phytochemistry 24 (11) (1985) 2479.