A Novel C30 Sterol from Porana racemosa. LI Bo-Gang, CHEN Bin, WANG Ding-Yong, YE Qi, ZHANG Guo-Lin*. (Chengdu Institute of Biology, The Chinese ...
Acta Botanica Sinica 植 物 学 报
2004, 46 (3): 375－378
http://www.chineseplantscience.com
A Novel C30 Sterol from Porana racemosa LI Bo-Gang, CHEN Bin, WANG Ding-Yong, YE Qi, ZHANG Guo-Lin* (Chengdu Institute of Biology, The Chinese Academy of Sciences, Chengdu 610041, China)
Abstract: A novel C30 sterol, (22E, 24ξ)-24-n-propylcholest-7, 22-dien-3β-ol (racemosol, 1), along with scopoletin (2), scopolin (3), umbelliferone (4), methyl β-D-frucopyranoside (5), syringaresinol-4-O-β-Dglucopyranoside (6), quercetin-3-O-β-D-glucopyranoside (7), quercetin-3-O-α-L-rhamnopyranoside (8), eupatilin (9), kaempferol-3-O-β-D-glucopyranoside (10) and (E)-N-2-(2,3-dihydroxyphenyl) ethyl cinnamamide (11), was isolated from the whole plants of Porana racemosa Roxb. Their structures were elucidated predominantly by spectral evidence. Key words: Porana racemosa ; Convolvulaceae; C30 sterol; racemosol; flavonol glycoside Porana racemosa (Convolvulaceae) is widely distributed in South China (Delectis Florae Reipublicae Popularis Sinicae Agendae Academiae Sinicae Edita, 1979). Its whole plants are used in Chinese folk medicine for treatment of nameless swelling, overworked pain and severe fever (Jiangsu New Medical College, 2000). Previous study on P. discifera Schneid led to the isolation of ecdysteroids (Zhu et al., 2000), coumarins and flavanoids (Yu et al., 2003), whereas no phytochemical investigation on P. racemosa has been reported. In this study, eleven compounds were isolated from the whole plants of P. racemosa and determined to be (22E, 24ξ)-24-n-propylcholest-7, 22-dien-3β-ol (1), scopoletin (2), scopolin (3), umbelliferone (4) (Vilegas and Pozetti, 1993), methyl β-D-frucopyranoside (5), syringaresinol-4-O-β-D-glucopyranoside (6) (Zhu et al., 2001), quercetin-3-O-β-D-glucopyranoside (7) (Bergeron et al., 1998), quercetin-3-O-α-L-rhamnopyranoside (8) (Hemandez et al., 1996), eupatilin (9) (Kupchan et al., 1967), kaempferol-3-O-β-D-glucopyranoside (10) (Markham et al., 1978) and (E)-N-2-(2,3-dihydroxyphenyl) ethyl cinnamamide (11) (Tseng et al., 1992) on the basis of spectral data. Compound 1, named as racemosol, is a novel C30 sterol.
1 Results and Discussion Compound 1, obtained as colorless needles, gave a quasi-molecular ion peak at m/z 425 [M－H]－ in the negative ESI-MS spectrum. The molecular ion peak at m/z 426.385 2 in the HREI-MS spectrum of compound 1 indicated a molecular formula C30H50O. Thirty signals were observed in the 13C-NMR spectrum (Table 1). In the 1HNMR spectrum, six methyls resonate at δ 0.58 (s, 3H), 0.82 (s, 3H), 1.05 (d, 3H, J = 6.4 Hz), 0.84 and 0.88 (each d, 3H, J
Received 23 Jul. 2003
= 6.6 Hz), and 0.83 (t, 3H, J = 7.3 Hz). An E-disubstituted double bond was supported by 1H-NMR signals at δ 5.17 (dd, 1H, J = 15.1, 8.8 Hz) and 5.05 (dd, 1H, J = 15.1, 8.7 Hz) and their corresponding 13C-NMR signals at δ 138.4 (d) and 129.6 (d) in HMQC experiment. The HMQC correlation between δ H 5.18 (t, 1H, J = 6.3 Hz) and δC 117.9 (d), combined with the 13C-NMR signal at δ 139.8 (s), indicated the presence of a trisubstituted double bond. By comparing the 13C-NMR data of compound 1 with those of spinasterol (12, Table 1) (Raymond and Jose, 1974; Toshihiro et al., 1981), it could be concluded that compound 1 is a sterol, possessing one more CH 2 than spinasterol. The HMBC correlations of H-19 (δ 0.82, s, 3H), H-3 (δ 3.63, m, 1H) and H-7 (δ 5.18, t, 1H) with C-5 (δ 41.1, d) confirmed the presence of 7, 8-double bond. The E-disubstituted double bond could be assigned to C-22 and C-23 in view of the HMBC correlations (Fig.1) derived from H-22 (δ 5.17, dd, 1H, J = 15.1, 8.7 Hz) and H-21 (δ 1.05, d, 3H, J = 6.4 Hz) with C-17 (δ 56.1, d). An n-propyl group could be located at C-24, based on the HMBC correlation H-23 (δ 5.05, dd, 1H, J = 15.1, 8.7 Hz) with C-28 (δ 25.6, t), and H24 (δ 2.04, m, 1H) with C-29 (δ 21.8, t). The IR absorption at νmax 3 419 and 1 045 cm－1 displayed the presence of hydroxyl group. The ion peak at m/z 255 [M+−H2O−C11H21] in its EIMS spectrum showed not only the presence of a C11H21 side chain with a double bond, but also a hydroxyl group at sterol nucleus in compound 1. The 13C-NMR signal at δ 71.3 (d) for C-3 suggested that 3OH should be β-configurated (Hanne et al., 1976). Thus, the structure of compound 1 was determined to be (22E, 24ξ)-24-n-propylcholest-7, 22-dien-3β-ol (racemosol) as shown in Fig.1.
Accepted 1 Dec. 2003
* Author for correspondence. Tel: +86 (0)28 85229742; Fax: +86 (0)28 85225401; E-mail: .
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Ａｃｔａ Ｂｏｔａｎｉｃａ Ｓｉｎｉｃａ 植物学报 Ｖｏｌ．４６ Ｎｏ．３ ２００４
Fig.1.
The structures of compounds 1 and 12 (→, HMBC correlation).
2 Experimental 2.1 General experimental procedures Melting points (uncorrected) were determined on XRC1 micro-melting point apparatus. Optical rotation was determined on Perkin-Elmer 341 polarimeter. IR and UV spectra were recorded on PROTEGE 460 spectrometer and ANTHELIE advanced spectrometer, respectively. NMR spectra were recorded on a Bruker Avance 600 or Bruker Avance 500 spectrometer, TMS as internal standard. EIMS and HREI-MS spectra were obtained on VG AutoSpec3000 mass spectrometer and ESI-MS spectra on LC-QDeca mass spectrometer. Silica gel (160−200 or 200−300 mesh) for column chromatography (CC) and silica gel G for TLC were purchased from Qingdao Ocean Chemical Factory, Shandong, China. 2.2 Plant material The whole plants of Porana racemosa Roxb. were collected in December, 1999, in Xishuangbanna, Yunnan Province, China and identified by Prof. CUI Jing-Yun in Xishuangbanna Tropical Botanic Garden, The Chinese Academy of Sciences, where a voucher specimen (No. W991021) is deposited. 2.3 Extraction and isolation The whole plants of P. racemosa (7.5 kg) were soaked three times with 95% EtOH at room temperature. After evaporation, 550 g residue was obtained, which was suspended in water (5 L), and then partitionated successively with petroleum ether, CHCl3, EtOAc and n-BuOH to give corresponding extracts 75 g, 60 g, 100 g and 110 g. The petroleum ether extract (75 g) was chromatographied with petroleum ether (60－90 ℃):acetone (100:1→5:1) to give fractions P1－6. Fraction P5 (10 g) was separated by CC eluted with petroleum ether:EtOAc (30:1), then recrystallized in
CHCl3 to give compound 1 (755 mg). The CHCl3 extract (60 g) was separated by CC eluted with petroleum ether (60－90 ℃):acetone (15:1; 10:1; 5:1, each 2 L) to give fractions C1－3. Fraction C1 (10 g) was subjected to CC eluted with petroleum ether (60－90 ℃): CHCl3:EtOAc (3:2:1) to give compounds 2 (420 mg) and 4 (35 mg). Compound 11 (153 mg) was obtained from fraction C3 (15 g) by CC eluted with CHCl3: MeOH (40:1). The EtOAc extract (100 g) was separated by CC eluted with CHCl3:MeOH (30:1; 20:1; 10:1, each 3 L) to give fractions E1－3. Fraction E1 (15 g) was subjected to CC eluted with CHCl3:MeOH (30:1) to give compound 9 (380 mg). Compound 3 (152 mg) was isolated from fraction E2 (35 g) by CC eluted with CHCl3:MeOH (20:1). The separation of fraction E3 (12 g) was carried out on CC eluted with CHCl3: MeOH (15:1) leading to compounds 10 (128 mg) and 8 (370 mg). The n-BuOH extract (110 g) was absorbed on macroporous resin D101 in CC and the resin was eluted with ethanol after being washed with H2O till no sugar eluted out. The ethanol eluate was evaporated to dryness under reduced pressure to give a residue (21 g), which was subjected to CC eluted with CHCl3:MeOH (10:1; 5:1) to give fractions B1-2. Fraction B1 (8 g) was further separated by CC eluted with CHCl3:MeOH (10:1) to give compounds 5 (280 mg) and 6 (100 mg). Compound 7 (260 mg) was obtained from fraction B2 (12 g) by CC (polyamide, 80－100 mesh) eluted with MeOH:H2O (1:2). 2.4 Identification Racemosol (1) Colorless needles (CHCl3), mp 120.0− －1 121.5 ℃. [α]25 D −5.0 ° (c 0.06, CHCl 3 ). IR νmax (KBr) cm : 3 419, 2 926, 2 858, 1 657, 1 463, 1 376, 1 159, 1 045, 971. Negative ESI-MS m/z: 425 [M−H]−. EI-MS m/z (%): 426 (4),
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LI Bo-Gang et al.: A Novel C30 Sterol from Porana racemosa Table 1 The 13C-NMR data of compound 1 and spinasterol (12) in CDCl3 1a 12 b C-atom δ 1 37.3 (t) 36.8 2 31.7 (t) 31.4 3 71.3 (d) 71.0 4 38.2 (t) 38.0 5 41.1 (d) 40.1 6 29.9 (t) 29.5 7 117.7 (d) 117.3 8 139.8 (s) 139.5 9 49.6 (d) 49.3 10 34.4 (s) 34.2 11 23.2 (t) 21.5 12 39.7 (t) 39.4 13 43.5 (s) 43.3 14 55.3 (d) 55.1 15 23.8 (t) 23.0 a, at 150 MHz, multiplicity determined by DEPT; b, at 25.2 MHz.
411 (21), 396 (5), 367 (7), 300 (6), 271 (38), 255 (20), 213 (9), 187 (5), 159 (15), 147 (21), 133 (19), 119 (22), 107 (41), 95 (39), 81 (79), 69 (60), 55 (100). HREI-MS m/z: 426.385 2 ([M]+, calcd. for C30H50O, 426.386 2). 1H-NMR (600 MHz, CDCl3） δ: 3.63 (1H, m, H-3), 5.18 (1H, t, J = 6.3 Hz, H-7), 5.17 (1H, dd, J = 15.1, 8.8 Hz, H-22), 5.05 (1H, dd, J = 15.1, 8.7 Hz, H-23), 2.04 (1H, m, H-24), 0.58 (3H, s, H-18), 0.82 (3H, s, H-19), 1.05 (3H, d, J = 6.4 Hz, H-21), 0.84 (3H, d, J = 6.6 Hz, H-26), 0.88 (3H, d, J = 6.6 Hz, H-27), 0.83 (3H, t, J = 7.3 Hz, H-30). 13CNMR (150 MHz, CDCl3) data are listed in Table 1. Scopoletin (2) Light green needles (MeOH), mp 230.5− 232.0 ℃, it was identified by comparison of the mixed melting point and Rf value (on TLC) with an authentic sample. Scopolin (3) Light green powder, mp 202.3−204.5 ℃， it was identified by comparison of the mixed melting point and Rf value (on TLC) with an authentic sample. Umbelliferone (4) Yellow powder, mp 225.0−227.5 ℃, its spectral data are consistent with those reported. Methyl β-D-frucopyranoside (5) White powder, mp 193.0−195.3 ℃, it was identified by comparing its spectral data with those reported. Syringaresinol-4-O-β-D-glucopyranoside (6) White powder, mp 167.0−168.5 ℃，its spectral data are in agreement with those reported. Quercetin-3-O-β-D-glucopyranoside (7) Yellow powder, mp 230−231.5 ℃， it was identified by comparing its spectral data with those reported. Quercetin-3-O-α-L-rhamnopyranoside (8) Yellow needles (MeOH), mp 265.0−267.5 ℃, it was identified by comparing its spectral data with those reported. Eupatilin (9) Yellow prisms (MeOH), mp 235.0−237.5
C-atom 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30
1a δ 29.8 (t) 56.1 (d) 12.3 (q) 13.3 (q) 40.5 (d) 21.3 (q) 138.4 (d) 129.6 (d) 51.5 (d) 32.1 (d) 21.6 (q) 19.2 (q) 25.6 (t) 21.8 (t) 12.5 (q)
12 b 28.5 55.9 12.1 13.0 40.8 21.1 138.1 129.4 51.2 31.9 21.5 19.0 25.4 12.3
℃，it was identified by comparing its spectral data with those reported. Kaempferol-3-O-β-D-glucopyranoside (10) Yellow powder, mp 192.0−194.5 ℃, it was identified by comparing its spectral data with those reported. (E)-N-2-(2, 3-Dihydroxyphenyl)ethyl cinnamic amide (11) White powder, mp 189.0−191.5 ℃， it was identified by comparing its spectral data with those reported. References: Bergeron C, Marston A, Antus S, Robert G, Kurt H. 1998. Flavonoids from Pyrola elliptica. Phytochemistry, 49:233−236. Delectis Florae Reipublicae Popularis Sinicae Agendae Academiae Sinicae Edita . 1979. Flora Reipublicae Popularis Sinicae. Tomus 64. No. 1. Beijing: Science Press. 34−35. (in Chinese) Hanne E, Craig L V, Norman S B, Carl D. 1976. Carbon-13 nuclear magnetic resonance spectra of hydroxy steroids. J Org Chem, 41:71−78. Hemandez P M, Hernandez T, Gomez G, Isabel E, Rosa M R. 1998. Phenolic composition of the “Mocan”(Visuea mocanera L. f.). J Agric Food Chem, 44:3512−3515. Jiangsu New Medical College . 2000. Dictionary of Chinese Traditional Medicine. Vol. 2. Shanghai: Shanghai Publishing House of Science and Technology. 640. (in Chinese) Jung J H, Kim C O, Kang S S. 1996. New bioactive cerebrosides from Arisaema amurense. J Nat Prod, 59:319−322. Kupchan S M, Sigel C W, Hemingway R T. 1969. Tumor inhibitors- Ⅻ cytotoxic flavones from Eupatorium species. Tetrahedron, 25:1603−1605. Markham K R, Ternal B, Stanley R. 1978. 13C-NMR studies of flavonoids-Ⅲ. Tetrahedron, 34:1389−1398.
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