Short communication
A New Neolignan Glycoside from the Roots of Adanitho^iani^x bi^iuek^j^^s
Hao-Bin Hu*, Xu-Dong Zheng, Huai-Sheng Hu and Yan Li
College of Chemistry and Chemical Engineering, Longdong Un^-versit-y, Qin^^ang 745000, Gansu Pro^vince, People's Republic o;f China
* Corresponding author: E-mail: hhb-88@126.com or huhaobin_88@^ahoo.com.cn; Tel.: +86-934-8631942, Fax: +86-934-8632822
Received: 19-09-2008
Abstract
A new neolignan glycoside, named as brachyposide A, was isolated from the EtOH extract of the roots of Acanthoma nax brachypus, together with nine known compounds. The structure of brachyposide A was characterized by spectros copic means as
noside. The known compounds were identified by comparing their spectral data with those of authentic samples or lite rature data.
Keywords: Araliaceae; Acanthopanax brachypus;; brachyposide A; (7S,8S)-A7 -2,9,9'-trihydroxy-7-O-3',8-O-4'-neolig nan-4-O-ß-D-apiofuranosyl-(1^6)-ß-D-glucopyranoside
1. Introduction
The Acanthopanax genus of the Araliaceae family includes 37 species around the world, and is widely distributed in Korea, Japan, China and the far-eastern region of Russia. Twenty-six species and 18 varieties grow in mainland China.1,2 The root and stem bark of these plants have been used clinically for a long time as a tonic and sedative, as well as for the treatment of rheumatism, diabetes, chronic bronchitis, hypertension, anti-stress and ischemic heart disease and gastric ulcers.3-6 As an endangered shrub in the wild due to overharvesting and loss of habitat through deforestation, Acanthopanax brachypus Harms is distributed in a narrow geographical area, mostly in the loess plateau of the Northwest of China.7,8 Research indi-
cates that the seeds of A. brachypus contain many kinds of micro-elements indispensable to the human body, can relax women's menopause syndrome and exhibit immuno-stimulatory and anticancer activities, and its rhizomatic extracts has also been successfully used in China and Korea for the inhibition of the various allergic responses.9-11 Nowadays, the other parts of this plant such as the roots, leaves and flowers are also employed for various therapeutic purposes.12-14 Although, the research has so far mainly concentrated on the reproductive biology and ecology, there have been a few studies on the chemical composition and biological activity.15,16 To further study its active constituents and provide the reference for effective utilization and quality control of the natural resources, our continuing phytochemical investigation on Acanthopanax
Figure 1: The structures of the isolated compound 1 from A. brachypus.
species17-21 has led to the identification of a new neolignan glycoside 1, named brachyposide A, along with nine known compounds, have been isolated for the first time from the roots of A. brachypus. In the present communication, we describe the isolation and structural elucidation of the new compound.
2. Results and Discussion
Compound 1 was obtained as a white amorphous powder from MeOH. Its molecular formula C29H36O15 was determined by positive ion HRFAB-MS (m/z: 625.2138 [M+H]+, calcd. 625.2132, A 0.6 nnu), corresponding to 12 degrees of unsaturation. The UV spectrum showed absorption bands at 209 and 266 nm, and its IR spectrum revealed the presence of hydroxy (3327 cm-1), olefinic carbons (1628 cm-1) and phenyl (1602, 1515 cm-1) moieties. The 1H and 13C NMR spectra showed the presence of a 1,3,4-trisubstiuted benzene ring [5H 6.93 (1H, d, J = 1.7 Hz), 6.82 (1H, d, J = 8.2 Hz) and 6.88 (1H, dd, J = 1.7, 8.2 Hz), 5C 110.9, 117.3 and 118.2], an asym-
trisubstituted benzene ring [5H 6.43 (1H, d, J = 2.4 Hz), 6.45 (1H, dd, J = 7.9, 2.4 Hz) and 6.96 (1H, d, J = 7.9 Hz), 5C 104.0, 108.5 and 116.7], an (£)-coniferyl alcohol [5H 4.02 (2H, br d, J = 5.7 Hz), 6.38 (1H, d, J = 15.3 Hz) and 6.19 (1H, dd, J = 15.3, 5.7 Hz), 5C 61.6, 128.6 and 126.5],22 two methines [5d 4.76 (1H, d, J = 8.0 Hz) and 4.30 (1H, dq, J = 8.0, 6.4 Hz), 5C 80.2 and73.8], one phenolic hydroxy [5d 9.70 (1H, s), 5C 155.0] and a hydroxy-methyl [5d 5.18 (1H, s) and 3.76 (2H, br d, J = 11.2 Hz), 5C 60.8], and two sugar anomeric protons [^g 4.82 (1H, d, J = 7.5 Hz, H-1") and 5.27 (1H, d, J = 2.2 Hz, H-1'''), the corresponding anomeric carbon signals at 5C 104.6 (C-1") and 111.0 (C-1''')]. The 13C NMR and DEPT spectra of 1 clearly displayed 29 carbon signals (5 x CH2, 17 x CH, 7 X C), of which 11 could be assigned to a glucose unit (5C 104.6, 74.7, 77.5, 71.0, 77.1, 68.0) and an apiose unit (5C 111.0, 77.9, 80.4, 75.1, 65.7), and the remaining 18 carbon signals were assigned to the aglycone. Comparison of the 1H and 13C NMR data of 1 with those of eusiderin E,23 indicated that 1 is a 7-0-3',8-0-4'-neolignan glycoside. In the HMBC and NOESY spectra, the correlations between 5C 145.9 (C-4) and 5d 4.82 (H-1")/6.43 (H-3)/6.45 (H-
Table 1: 'H (400 MHz) and 13C NMR (100 MHz) spectral data of compounds 1 (DMSO-rf6 TMS)*.
No.	^H (■/)(Hz)	Sc	DEPT	HMBC(H^C)	ROESY(H^H)
1	-	131.1	C	H-3, H-5, H-6, H-7, H-8, HO-2	
2	-	155.0	C	H-3, H-6, H-7, HO-2	
3	6.43 (d, 2.4)	104.0	CH	HO-2, H-5	
4	-	145.9	C	H-1'', H-3, H-5, H-6	
5	6.45 (dd, 7.9, 2.4)	108.5	CH	H-3, H-6	H-1"
6	6.96 (d, 7.9)	116.7	CH	H-5, H-7	
7	4.76 (d, 8.0)	80.2	CH	H-6, H-8	H-9
8	4.30 (dq, 8.0, 6.4)	73.8	CH	H-7, H-9	
9	3.76 (br d, 11.2)	60.8	CH2	H-8, HO-9	H-7
1'	-	131.3	C	H-2', H-5', H-6', H-7', H-8'	
2'	6.93 (d, 1.7)	110.9	CH	H-6', H-7'	H-7'
3'	-	143.4	C	H-7, H-2', H-5'	
4'	-	136.7	C	H-8, H-6', H-5'	
5'	6.82 (d, 8.2)	117.3	CH	H-6'	
6'	6.88 (dd, 1.7, 8.2)	118.2	CH	H-2', H-5', H-7'	H-8'
7'	6.38 (d, 15.3)	128.6	CH	H-2', H-6', H-8'	
8'	6.19 (dd, 15.3, 5.7)	126.5	CH	H-7', H-9'	H-6'
9'	4.02 (br d, 5.7)	61.6	CH2	H-8', HO-9'	
HO-2	9.70 (s)	-	-		
HO-9	5.18 (s)	-	-		
Glc-1''	4.82 (d, 7.5)	104.6	CH	H-2''	H-6, H-3'', H-5''
2''	3.82 (dd, 9.1, 7.4)	74.7	CH	H-1''	H-4''
3''	3.78 (dd, 9.1, 8.5)	77.5	CH		H-1'', H-5''
4''	3.94 (dd, 9.9, 8.5)	71.0	CH		H-2''
5''	3.81 (ddd, 9.9, 6.0, 1.6)	77.1	CH		H-1'', H-3''
6''	4.06 (dd, 11.3,1.6)	68.0	CHj	H-1'''	
	3.94 (dd, 11.3, 6.0)				
Api-1'''	5.27 (d, 2.2)	111.0	CH	H-6'', H-2'''	
2'''	3.98 (d, 2.2)	77.9	CH	H-4''', H-5'''	
3'''	-	80.4	C	H-2'''	
4'''	3.77 (d, 9.4)/3.95 (d, 9.4)	75.1	CH2		
5'''	3.68 (s)	65.7	CH2		
Figure 2: The key HMBC (H ^ C) correlations of compound 1.
5)/6.96 (H-6), 5C 131.3 (C-1') and 5H 6.38 (H-7')/6.88 (H-6')/6.93 (H-2'), AC 155.0 (C-2) and 5H 6.43 (H-3)/6.96 (H-6), 5C 104.0 (C-3)/131.1 (C-1) and 5H 9.70 (HO-2), 5H 4.82 (H-1") and 5H 6.45 (H-5), 6.88 (H-6') and 5H 6.1H (H-8') as well as 5H 6.93 (H-2') and 5H 6.38 (H-7'), suggested that the disaccharide chain, (£)-coniferyl alcohol side-chain and hydroxyl groups were connected to C-4, C-1' and C-2 of the aglycone, respectively.
Upon acid hydrolysis, compound 1 gave ö-glucose and ö-apiose, according to co-TLC with authentic samples and rotational analysis.24 In addition, this was also confirmed by the FAB-MS spectral observation of fragment ions at m/z 493 [M+H-132]+ and m/z 331 [M+H-132-162]+, arising from the elimination of an apiose and a glucose unit, indicating the apiose was terminal sugar and the glucose was attached to the aglycone. Comparison of 13C NMR data of the sugar moieties with literature values,25 revealed that the glucose was present in pyranoside form and the apiose was in furanoside form. The HMBC correlations (Figure 2) of H-1'" (5^ 5.27) with C- 6" (5C 68.0) and H-6" (5h 4.06/3.94) witli C-1''' (5C 111.0), suggested an apiose-(1^6)-glucose linkage. The ^-configuration of apiose was confirmed by comparing the 13C-NMR spectra of 1 with those of a-D-(5C 104.5) and ^-D-apiofuranosides (5C 111.5), respectively,26 and the glucose had the ^-configuration according to the coupling constant (J = 7.5 Hz) of H-1" of glucose. The coupling constants (J = 15.3 Hz) between H-7' and H-8' suggested that the (£)-coniferyl alcohol side-chain had a tran^-configuration. The signals of H-7 (5H 4.76) and H-8 (5h 4.30) at slightly lower fields, with a larger coupling
constant (J = 8.0 Hz), along with the NOESY correlations between 5H 4.76 (H-7) and 5H 3.76 (H-9), indicated a trans-orientation of H-7 and H-8 pair in 1.27 By comparison of CD value of 1 with that of the known (7S,8S)-A7'-2,4-dihydroxy-7-O-3',8-O-4'-neolignan,28 suggested the plausible configurations of C-7 and C-8 as S and S, respectively. On these grounds, the structure of 1 was determined as (7S,8S)-A7'-2,9,9'-trihydroxy-7-0-3',8-0-4'-neolignan-4- O-ß-D-apiofuranosyl-(1 ^6)-ß-D-glucop-yranoside, and named brachyposide A.
The known compounds 2-10 were identified as quercetin-3-O-neohesperidoside (2),29 echioidin (3),30 maltol ß-D-glucopyranoside (4),31 isoandrographolide (5),32 2-methoxyphenyl ß-D-glucopyranoside (6),33 (-)-syringaresinol-4,4'-bis-O-ß-D-glucopyranoside (7),34,35 acantrifoside A (8),6 ß-sitosterol (9)36 and daucosterol (10)37 by comparing their spectroscopic data with values reported in the literature.
3. Experimental
3. 1. General
Melting points (uncorrected) were observed with a Chinese X-4 melting point apparatus. Optical rotations were measured with Perkin-Elmer 241 digital polarimeter. UV and IR (KBr disks) spectra were obtained on Shimad-zu UV-300 (double beam) and Alpha-Centari FT-IR spectrophotometers. CD spectra were recorded on a Jasco J-715 spectropolarimeter. 1H and 13C NMR (DEPT) spectra were recorded on a Bruker AM-400 NMR spectrometer. Mass spectra were carried out on ZAB-HS and MAT-112 mass spectrometers, respectively. Separation and purification were performed by column chromatography on silica
Figure 3: The key NOESY (H^H) correlations of compound 1.
gel (100-200, 200-300 mesh). TLC was performed on silica gel GF254 plates. The spots were visualized by UV (254 nm) and EtOH-H2SO4.
3. 2. Plant Material
The roots of A. brachypus were collected in August 2007 from Qingyang of Gansu Province, China, and were identified by Prof. Xiao-Qiang Guo, Department of Life Sciences, Longdong University. A voucher specimen (No. 12107) was deposited in the Herbarium of the Department of Life Sciences, Longdong University, People's Republic of China.
3. 3. Extraction and Isolation
The air-dried and powered roots of A. brachypus (5.5 kg) were soaked in 95% EtOH (15 L, 7 dx3) at room temperature. After removing the solvent, the extract (298 g) was suspended in warm water and partitioned with petroleum ether (60-90 °C), CHCl3, EtOAc and n-BuOH, successively. The «-BuOH-soluble fraction was evaporated to give 86.7 g of residues, which was isolated on a silica gel column eluting with CHCl3-MeOH (6:0^1:6) in increasing polarity and combined by monitoring with TLC to give four fractions (A, B, C and D). Fraction A (4.6 g) was further fractionated over silica gel column and eluted with CHCl3-MeOH (4:1) to obtain 3 (17 mg). Fraction B (3.1 g) was purified on a silica gel column using CHCl3-MeOH gradient (3:0^0:3) as eluent to afford 2 (11 mg). Fraction C (5.4 g) was rechromatographed over a silica gel column eluting with acetone-MeOH (3:0^1:3) to yield 1 (11 mg) and two subfractions (C1-C2). Subfrac-tion C1 was further purified by preparative TLC (silica gel) and developed with CHCl3-MeOH (1:1) to provide compounds 4 (8 mg) and 5 (10 mg). Subfraction C2 was further purified on silica gel column eluting stepwise with CHCl3-MeOH (from 5:1 to 1:8, V/V), and then on Sepha-dex LH-20 eluting with MeOH to obtain compounds 6 (17 mg) and 7 (9 mg), respectively. Fraction D (5.4 g) was purified on a silica gel column using CHCl3-MeOH gradient (2:1^1:8) as eluent to afford three subfractions (D1-D3). Subfraction D1 was rechromatographed over silica gel and further purified by preparative TLC (MeOH/CHCl3/n-he-xane, 1:5:2, V/V) to afford compounds 8 (24 mg) and 9 (11 mg). Subfraction D3 was chromatographed on Sepha-dex LH-20 with MeOH-H2O (1:1, V/V) to afford compound 10 (19 mg).
3. 4. Characterization Data
Compound 1: White amorphous powder (MeOH), mp. 212-215°C; [a]20 -10.8° (c 0.45, MeOH); HRFAB-MS: m/z 625.2138 [M+H]+ (C29H36O15 calcd. 625.2132, A 0.6 nnu); UV XM^OH (nm): 209, 26(5; (^D (MeOH, c 2.45 x 10-5 g/mL), Ae18 (nm): +10.58 (223.5), 0 (237.5), -2.43
(258.5); IR v^Bx (cm-1): 3327 (OH), 1628 (olefinic C=C), 1602, 1515 (i)henyl); FAB-MS: m/z 625 [M+H]+, 493 [M+H-132]+ and 331 [M+H-132-162]+; the 1H and 13C NMR data see Table 1.
4. Acknowledgements
This work was supported by the Support Program for Longyuan Scientific and Technical Innovation Talented Person and the Scientific Research Project of Longdong University (Grant No. XYKZ0802).
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Povzetek
Novi, neolignanski glikozid imenovan brahipozid A, je bil skupaj s še devetimi, že znanimi spojinami, izoliran iz EtOH ekstrakta korenin Acanthopanax brachypus. Struktura brahipozida A je bila s spektroskopskimi metodami določena kot (7S,8S)-A7'-2,9,9'-trihidroksi-7-O-3',8-O-4'-neolignan-4-O-ß-D-apiofuranozil-(1^6)-ß-D-glukopiranozid. Ostale znane spojine so bile identificirane s pomočjo primerjave njihovih spektroskopskih podatkov s podatki za avtentične vzorce ali literaturnimi vrednostmi.