Flavonol and lignan glycosides from datura metel l - Nguyen Thi Mai

Vietnam Journal of Science and Technology 55 (3) (2017) 263-270 DOI: 10.15625/2525-2518/55/3/8811 FLAVONOL AND LIGNAN GLYCOSIDES FROM Datura metel L. Nguyen Thi Mai1, Nguyen Thi Kim Cuc2, Tran Hong Quang2, Phan Van Kiem2, * 1University of Transport and Communications, 3 Cau Giay, Dong Da, Ha Noi, Viet Nam 2Institute of Marine Biochemistry, Vietnam Academy of Science and Technology (VAST), 18 Hoang Quoc Viet, Cau Giay, Ha Noi, Viet Nam *Email: phankiem@yahoo.com Received 26 October 2015; Accepted for publication: 21 February 2017 ABSTRACT Chemical investigation of an acidic methanol extract of the whole plants of D. metel resulted in the isolation of five compounds, including kaempferol 3-O-β-D-glucosyl(1→2)-β-D- galactoside 7-O-β-D-glucoside (1), kaempferol 3-O-β-glucopyranosyl(1→2)-β-glucopyranoside- 7-O-α-rhamnopyranoside (2), pinoresinol O-β-D-glucopyranoside (3), (7R,8S,7′S,8′R)-4,9,4′,7′- tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside (4), and (7S,8R,7′S,8′S)- 4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside (5). Their structures were elucidated by 1D and 2D NMR and MS spectroscopic analyses as well as comparing with the data reported in the literature. The absolute configurations of compounds 4 and 5 were determined by CD spectra. It is noted that (7R,8S,7′S,8′R)-4,9,4′,7′-tetrahydroxy- 3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside and (7S,8R,7′S,8′S)-4,9,4′,7′- tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside were isolated for the first time from the Datura genus. Keywords: Datura metel, Solanaceae, Flavonol glycoside, Lignan glycoside. 1. INTRODUCTION Datura metel L. is an annual herb of the Solanaceae family and is widely cultivated in many tropical and temperate regions. In the Vietnamese traditional medicine, D. metel has been used for the treatment of coughs, bronchial asthma, and rheumatism [1]. Its leaves have been used as anesthetics in surgery, a fumigant in bronchial asthma, and anti-contractive agents in the stomach ulcers [1]. The flowers of D. metel have been used widely in the Chinese traditional medicine for the treatment of asthma, convulsions, pain, and rheumatism for centuries [2]. Previous pharmacological studies have showed that the D. metel seeds have hypoglycemic activity in normal and alloxan-induced diabetic rats [3], the chloroform extract of D. metel exhibits an antifungal effect toward several pathogenic species of Aspergillus [4], and the seeds and fruit pulps of D. metel show high antioxidant activity [5]. Previous chemical studies have demonstrated that the major chemical components of D. metel are with anolide-type steroids [6 – 12], which have been shown to suppress NO production in lipopolysaccharide (LPS)-stimulated RAW264.7 cells [11, 12] and exhibit cytotoxicity against HCT-116, A549, DLD-1, BGC-823, Nguyen Thi Mai, Nguyen Thi Kim Cuc, Tran Hong Quang, Phan Van Kiem 264 and K562 cancer cell lines [6, 7, 10]. In addition, some megastigmane sesquiterpenes and amide alkaloids from D. metel were also reported [13, 14]. In the present study, we report the isolation and structural elucidation of two flavonol glycosides (1 and 2) and three lignan glycosides (3-5) from the acidic methanol extract of the whole plants of D. metel. 2. MATERIAL AND METHODS 2.1. General experimental procedures All NMR spectra were recorded on a Bruker AM500 FT-NMR spectrometer (500 MHz for 1H and 125 MHz for 13C-NMR), chemical shifts are reported in ppm using TMS as an internal standard. ESIMS spectra were recorded on Agilent 1100 spectrometer. Circular dichroism (CD) spectra were measured on a Chirascan CD spectrometer (Applied Photophysics Ltd., Surrey, UK). Column chromatography (CC) was performed on silica gel 230 - 400 mesh or reversed phase (RP) C18 resins (150 µm, Fuji Silysia Chemical Ltd.). Compounds were visualized by spraying with aqueous 10 % H2SO4 and heating for 5 minutes. 2.2. Plant material The whole plants of D. metel were collected in Thai Binh province, Vietnam during May 2015, and identified by Dr. Bui Van Thanh, Institute of Ecology and Biological Resources. A voucher specimen (NCCT-CDM-5.2015) was deposited at the Herbarium of the Institute of Marine Biochemistry, VAST. 2.3. Extraction and isolation The dried whole plants of D. metel (5 kg) were ground and extracted with MeOH/acetic acid (pH = 5.0) under sonication at room temperature. After concentration under reduced pressure, the MeOH extract (300 g) was suspended in water and partitioned with CHCl3 to give CHCl3- and water-soluble fractions. The water-soluble fraction was alkalinized by adding NH4OH until pH = 9.0, and then partitioned successively with CH2Cl2 and EtOAc to provide CH2Cl2-, EtOAc-, and water-soluble fractions, respectively. The water-soluble fraction was neutralized and subjected to fractionation through a Diaion HP-20 column, eluted with a gradient of MeOH in water (25–100 %) to give four fractions (DMW1–DMW4). Fraction DMW2 was separated using RP C18 column chromatography (CC), eluting with MeOH-H2O (1:3, v/v) to yield subfractions DMW21–DMW25. Subfraction DMW22 was then separated by silica gel CC, eluting with CH2Cl2-MeOH-H2O (5:1:0.05, v/v/v) to give 1 (25 mg) and 4 (6 mg). Subfraction DMW23 was separated by silica gel CC, eluting with CH2Cl2-MeOH-H2O (6:1:0.05, v/v/v) and further purified by a RP C18 CC, eluting with MeOH-H2O (1:3, v/v) to yield 5 (6 mg). Subfraction DMW25 was separated using silica gel CC, eluting with CH2Cl2-MeOH-H2O (6:1:0.05, v/v/v) and subsequently purified by RP C18 CC, eluting with MeOH-H2O (1:2, v/v) to release 2 (20 mg). Fraction DMW42 was separated by silica gel CC, eluting with CH2Cl2- MeOH-H2O (6:1:0.05, v/v/v) to provide four subfractions (DMW421–DMW424). Subfraction DMW424 was separated by RP C18 CC, eluting with acetone-H2O (1:3, v/v) to obtain 3 (5 mg). Kaempferol 3-O-β-D-glucosyl(1→2)-β-D-galactoside 7-O-β-D-glucoside (1): yellow, amorphous powder; C33H40O21, M = 772; ESI-MS m/z: 795 [M+Na]+; 1H NMR (DMSO-d6, 500 MHz) δH: 6.43 (d, J = 2.0 Hz, H-6), 6.80 (d, J = 2.0 Hz, H-8), 8.11 (d, J = 8.5 Hz, H-2′ and H-6′), 6.90 (d, J = 8.5 Hz, H-3′ and H-5′), 5.68 (d, J = 7.5 Hz, H-1′′), 4.58 (d, J = 8.0 Hz, H-1′′′), 5.07 (d, J = 7.0 Hz, H-1′′′′); 13C NMR (DMSO-d6, 125 MHz): see Table 1. Flavonol and lignan glycosides from Datura metel L. 265 Table 1. 13C NMR data (500 MHz) for compounds 1−5. C 1 2 3 4 5 δC#1 δCa δC#2 δCb δC#3 δCb δC#4 δCa,b δC#5 δCa,b 1 53.7 55.5 133.5 138.5 133.8 137.8 2 156.2 156.2 159.5 159.4 85.1 87.5 110.6 111.7 110.9 111.8 3 133.2 133.2 135.2 135.1 147.4 150.8 147.4 150.9 4 177.7 177.7 180.0 179.7 71.0 72.6 145.8 147.1 145.6 147.6 5 160.9 160.9 163.0 163.0 53.5 55.3 115.1 117.8 115.1 117.8 6 99.4 99.4 100.7 100.5 84.9 87.0 118.9 120.0 118.9 120.4 7 162.8 162.8 163.6 163.4 83.4 84.6 82.8 85.4 8 94.5 94.5 95.6 95.5 71.0 72.6 53.9 53.7 51.9 55.9 9 156.0 156.0 158.2 158.0 61.6 62.4 60.4 63.3 10 105.6 105.6 107.6 107.5 1′ 120.8 120.8 122.7 122.5 132.3 133.7 138.4 136.0 138.3 136.2 2′ 131.2 131.2 132.5 132.4 110.6 110.9 111.1 111.5 110.4 111.4 3′ 115.4 115.4 116.4 116.3 146.0 147.0 148.7 148.9 148.7 149.0 4′ 160.2 160.2 161.7 161.8 147.5 149.1 145.8 147.3 145.8 148.5 5′ 115.4 115.4 116.4 116.3 115.2 116.0 114.9 115.9 114.7 115.9 6′ 131.2 131.2 132.5 132.4 118.6 119.7 119.4 120.7 118.7 120.8 7′ 74.8 76.6 73.7 77.4 8′ 50.6 50.8 49.3 52.7 9′ 69.5 71.6 69.5 71.3 1′′ 98.3 98.4 100.9 100.8 135.4 136.0 100.7 102.9 100.2 102.8 2′′ 80.5 80.5 82.8 82.6 110.8 111.5 73.3 74.9 73.3 74.9 3′′ 73.4 73.4 78.0 77.8 146.0 147.4 77.1 77.8 77.0 77.8 4′′ 67.7 67.7 71.3 71.2 149.1 150.9 69.7 71.3 69.7 71.3 5′′ 75.9 75.9 78.4 78.1 115.6 117.9 76.9 78.2 76.9 78.2 6′′ 60.0 60.0 62.6 62.4 118.2 120.0 60.7 62.5 60.7 62.5 1′′′ 104.3 104.3 104.8 104.7 100.4 102.8 2′′′ 74.5 74.4 75.7 75.5 73.3 74.8 3′′′ 76.5 76.5 78.0 77.8 76.8 77.8 4′′′ 69.7 69.7 71.4 71.2 70.0 71.3 5′′′ 77.0 77.0 78.3 78.2 77.0 78.2 6′′′ 60.7 60.7 62.7 62.5 60.7 62.4 1′′′′ 99.8 99.8 99.9 99.7 2′′′′ 73.1 73.1 71.8 71.6 3′′′′ 76.6 76.6 72.2 72.0 4′′′′ 69.7 69.7 73.7 73.5 5′′′′ 77.2 77.2 71.3 71.1 6′′′′ 60.8 60.9 18.1 18.0 OCH3 55.7 55.9 56.3 56.7 55.7 55.8 56.7 56.4 55.6 55.7 56.7 56.4 a Recorded in DMSO-d6, b in CD3OD; #1 δC of 3-O-β-D-glucosyl(1→2)-β-D-galactoside 7-O-β-D-glucoside in DMSO-d6 [16]; #2 δC of kaempferol 3-O-β-D-glucopyranosyl(1→2)-β-D-glucopyranoside-7-O-α-L- rhamnopyranoside in CD3OD [17]; #3 δC of pinoresinol O-β-D-glucopyranoside in DMSO-d6 [1]; #4 δC of (7R,8S,7′S,8′R)-4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside in DMSO-d6 [15]; #5 δC of (7S,8R,7′S,8′S)-4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside in DMSO-d6 [15]. Nguyen Thi Mai, Nguyen Thi Kim Cuc, Tran Hong Quang, Phan Van Kiem 266 Kaempferol 3-O-β-D-glucopyranosyl(1→2)-β-D-glucopyranoside-7-O-α-L-rhamnopyranoside (2): yellow, amorphous powder; C33H40O20, M = 756; ESI-MS m/z: 779 [M+Na]+; 1H NMR (CD3OD, 500 MHz) δH: 6.47 (s, H-6), 6.76 (s, H-8), 8.08 (d, J = 8.5 Hz, H-2′ and H-6′), 6.93 (d, J = 8.5 Hz, H-3′ and H-5′), 5.50 (d, J = 7.5 Hz, H-1′′), 4.79 (d, J = 7.5 Hz, H-1′′′), 5.59 (br s, H- 1′′′′), 1.27 (d, J = 6.5 Hz, H3-6′′′′); 13C NMR (CD3OD, 125 MHz): see Table 1. Pinoresinol O-β-D-glucopyranoside (3): white, amorphous powder; C26H32O11, M = 520; ESI- MS m/z: 543 [M+Na]+; 1H NMR (CD3OD, 500 MHz) δH: 4.57 (H-2), 4.73 (H-6), 6.97 (s, H-2′), 6.95 (d, J = 8.5 Hz, H-5′), 6.79 (d, J = 8.5 Hz, H-6′), 7.05 (s, H-2′′), 7.17 (d, J = 8.5 Hz, H-5′′), 6.84 (d, J = 8.5 Hz, H-6′′), 4.85 (d, J = 7.5 Hz, H-1′′′), 3.87 and 3.89 (each s, OCH3); 13C NMR (CD3OD, 125 MHz): see Table 1. (7R,8S,7'S,8'R)-4,9,4',7'-Tetrahydroxy-3,3'-dimethoxy-7,9'-epoxylignan-4-O-β-D- glucopyranoside (4): white, amorphous powder; C26H34O12, M = 538; ESI-MS m/z: 561 [M+Na]+; CD (MeOH) λmax (∆ε) 279 (−7.23) and 231 (−16.11) nm [15]: λmax (∆ε) 277 (−1.84) and 231 (−5.34) nm]; 1H NMR (CD3OD, 500 MHz) δH: 7.03 (d, J = 2.0 Hz, H-2), 7.16 (d, J = 8.0 Hz, H-5), 6.94 (dd, J = 2.0, 8.0 Hz, H-6), 4.70 (d, J = 7.0 Hz, H-7), 1.91 (m, H-8), 3.33 (m, H-9a), 3.88 (m, H-9b), 6.88 (d, J = 2.0 Hz, H-2′), 6.76 (d, J = 8.0 Hz, H-5′), 6.74 (dd, J = 2.0, 8.0 Hz, H-6′), 4.49 (d, J = 8.5 Hz, H-7′), 2.55 (m, H-8′), 3.98 (m, H-9′a), 4.28 (dd, J = 4.5, 8.0 Hz, H-9′b), 4.91 (d, J = 7.5 Hz, H-1′′), 3.88 (s, 3-OCH3), 3.83 (s, 3′-OCH3); 13C NMR (CD3OD, 125 MHz): see Table 1. (7S,8R,7'S,8'S)-4,9,4',7'-Tetrahydroxy-3,3'-dimethoxy-7,9'-epoxylignan-4-O-β-D- glucopyranoside (5): white, amorphous powder; C26H34O12, M = 538; ESI-MS m/z: 561 [M+Na]+; CD (MeOH) λmax (∆ε) 284 (+0.52), 233 (+17.45) nm [15]: λmax (∆ε) 281 (+0.88) and 234 (+1.19) nm]; 1H NMR (CD3OD, 500 MHz) δH: 7.03 (br s, H-2), 7.16 (d, J = 8.5 Hz, H-5), 6.93 (br d, J = 8.5 Hz, H-6), 4.60 (d, J = 8.5 Hz, H-7), 2.29 (m, H-8), 3.67 (m, H2-9), 7.01 (br s, H-2′), 6.80 (d, J = 8.0 Hz, H-5′), 6.85 (dd, J = 1.5, 8.0 Hz, H-6′), 4.50 (d, J = 9.0 Hz, H-7′), 2.64 (m, H-8′), 3.67 (m, H-9′a), 3.77 (dd, J = 6.5, 9.0 Hz, H-9′b), 4.86 (d, J = 7.5 Hz, H-1′′), 3.89 (s, 3-OCH3 and 3′-OCH3); 13C NMR (CD3OD, 125 MHz): see Table 1. 3. RESULTS AND DISCUSSION Compound 1 was obtained as a yellow, amorphous powder. Its molecular formula was established as C33H40O21 by an ion peak [M+Na]+ at m/z 795 in the ESIMS and the 13C NMR spectroscopic analysis. The 1H NMR of 1 showed signals for two meta coupled aromatic protons at δH 6.43 (d, J = 2.0 Hz, H-6) and 6.80 (d, J = 2.0 Hz, H-8) and a para-substituted aromatic ring at δH 8.11 (d, J = 8.5 Hz, H-2′ and H-6′) and 6.90 (d, J = 8.5 Hz, H-3′ and H-5′). The 1H NMR spectrum further showed signals for three anomeric protons at δH 5.68 (d, J = 7.5 Hz, H-1′′), 4.58 (d, J = 8.0 Hz, H-1′′′), and 5.07 (d, J = 7.0 Hz, H-1′′′′), revealing that 1 has three sugar units. Analysis of 13C NMR and HSQC spectra indicated the presence of 33 carbons, including one carbonyl carbon at δC 177.7 (C-4), eight non-protonated aromatic carbons (of which five were oxygenated), and six aromatic methine carbons, suggesting that 1 possesses the flavonol skeleton (Table 1). The 18 remaining carbons were assigned to three sugar units, which were identified as two glucopyranoses and one galactopyranose by comparison with those reported in the literature [16]. The relatively large spin couplings of the three anomeric protons (J ≥ 7.0 Hz) are characteristic features of the β-configurations for the glucose and galactose units. In the HMBC spectrum, the HMBC correlations from δH 4.58 (H-1′′′) to δC 80.5 (C-2′′) and from δH Flavonol and lignan glycosides from Datura metel L. 267 5.68 (H-1′′) to δC 98.4 (C-3) suggested that the β-glucopyranosyl-(1→2)]-β-D-galactopyranoside sugar chain was located at C-3 position (Figure 2). The remaining β-glucopyranose was attached to C-7 by the HMBC correlation observed from δH 5.07 (H-1′′′′) to δC 162.8 (C-7). On the basis of the above analysis, along with comparison with those of the reported flavonol glycoside [16], the structure of 1 was established as kaempferol 3-O-β-D-glucosyl(1→2)-β-D-galactoside 7-O-β- D-glucoside. Figure 1. Chemical structures of compounds 1−5 from D. metel. The molecular formula of compound 2 was determined to be C33H40O20 by the presence of an ion [M+Na]+ at m/z 779 in the ESIMS. The 1H NMR spectrum contained signals for an AX spin system [δH 6.47 (s, H-6) and 6.76 (s, H-8)] and an AA′BB′ pattern at δH 8.08 (d, J = 8.5 Hz, H-2′ and H-6′) and 6.93 (d, J = 8.5 Hz, H-3′ and H-5′). The signals for three anomeric protons at δH 5.50 (d, J = 7.5 Hz, H-1′′), 4.79 (d, J = 7.5 Hz, H-1′′′), and 5.59 (br s, H-1′′′′) observed in the 1H NMR spectrum indicated the presence of three sugars in the structure. The 13C NMR spectrum comprised 33 carbon signals, including 15 carbons of the aglycone and 18 carbons belonging to the sugar moiety (Table 1). Comparison of the 1H and 13C NMR data of 2 with those of 1 revealed that these compounds have the same aglycone but different sugar moieties. The sugar moiety of 2 was found to consist of two glucose units and one rhamnose by detailed analysis of 13C NMR and HSQC spectra in comparison with the previously reported values [17]. The β-configurations for the anomeric protons of the glucopyranoses were deduced based on the relatively large coupling constants (J = 7.5 Hz), while the α-oriented anomeric proton of the rhamose was determined by its carbon chemical shift values of C-3 and C-5 positions [19]. The sugar chain at C-3 of the aglycone was identified as β-D-glucopyranosyl(1→2)-β-D- glucopyranoside by the HMBC correlations from δH 4.79 (H-1′′′) to δC 82.6 (C-1′′) and from δH 5.50 (H-1′′) to δC 135.1 (C-3) (Figure 2). The location of the rhamnose at C-7 position was deduced by the HMBC cross-peak between δH 5.59 (H-1′′′′) to δC 163.4 (C-7). Thus, the Nguyen Thi Mai, Nguyen Thi Kim Cuc, Tran Hong Quang, Phan Van Kiem 268 structure of 2 was identified as kaempferol 3-O-β-D-glucopyranosyl(1→2)-β- D- glucopyranoside-7-O-α-L-rhamnopyranoside. Compound 3 was isolated as a white, amorphous powder. Its molecular formula, C26H32O11 was deduced by the observation of an ion peak [M+Na]+ at m/z 543 in the ESIMS and 13C NMR spectroscopic analysis. The 1H NMR spectrum exhibited signals for two ABX spin systems at δH 6.97 (s, H-2′), 6.95 (d, J = 8.5 Hz, H-5′), 6.95 (d, J = 8.5 Hz, H-6′), 7.05 (s, H-2′′), 7.17 (d, J = 8.5 Hz, H-5′′), and 6.84 (d, J = 8.5 Hz, H-6′′) and two methoxy groups at δH 3.87 and 3.89 (each s, 4′-OCH3 and 4′′-OCH3). The signal of an anomeric proton at δH 4.85 (d, J = 7.5 Hz, H-1′′′) observed in the 1H NMR of 3 implied the presence of a sugar unit. The 13C NMR and DEPT spectra displayed 26 carbon signals, of which six nonprotonated aromatic carbons (including four were oxygenated), six aromatic methines, two oxymethines at δC 87.5 (C-2) and 87.0 (C-6), two oxymethylenes at δC 72.6 (C-4 and C-8), two methines at δC 55.5 (C-1) and 55.3 (C-5), suggesting that 3 is a lignan derivative (Table 1). The six remaining carbon signals at δC 102.8, 74.8, 77.8, 71.3, 78.2, and 62.4 could be assigned to a glucopyranose. Comparison of the 1H and 13C NMR data of 3 with those of the reported lignan, pinoresinol O-β-D-glucopyranoside, revealed that the structures of these compounds are identical [18]. Therefore, the structure of compound 3 was identified as shown in Figure 1. Figure 2. Selected HMBC correlations of compounds 1, 2, 4, and 5. The ESIMS of compound 4 exhibited an ion [M+Na]+ at m/z 561, corresponding with the molecular formula C26H34O12. The 1H NMR spectrum showed signals for two ABX spin systems at δH 7.03 (d, J = 2.0 Hz, H-2), 7.16 (d, J = 8.0 Hz, H-5), 6.94 (dd, J = 1.5, 8.5 Hz, H-6), 6.88 (d, J = 2.0 Hz, H-2′), 6.76 (d, J = 8.0 Hz, H-5′), and 6.74 (dd, J = 2.0, 8.0 Hz, H-6′) and two methoxy groups at δH 3.88 (s, 3-OCH3) and 3.83 (3′-OCH3). Compound 4 was found to have one sugar unit by the observation of an anomeric proton at δH 4.91 (d, J = 7.5 Hz, H-1′′) in the 1H NMR spectrum. Analysis of 13C NMR and HSQC spectra indicated the presence of 26 carbons, including six non-protonated aromatic carbons (of which four were oxygenated), six aromatic methines, two oxymethines at δH 4.70/δC 84.6 (C-7) and δH 4.49/δC 76.6 (C-7′), two oxymethylenes at δH 3.33 and 3.88/δC 62.4 (C-9) and δH 3.98 and 4.28/δC 71.6 (C-9′), suggesting that 4 belongs to the lignan skeleton (Table 1). The sugar was suggested to be β-glucopyranose by the observation of six carbon signals at δC 102.9, 74.9, 77.8, 71.3, 78.2, and 62.5 and the large coupling constant of the anomeric proton (J = 7.5 Hz). Comparison of the 1H and 13C NMR data of 4 with those of the reported lignan glycoside, tetrahydroxy-3,3′-dimethoxy-7,9′- epoxylignan-4-O-β-D-glucopyranoside resulted in the close similarity [15]. In the HMBC Flavonol and lignan glycosides from Datura metel L. 269 spectrum, the HMBC correlations from δH 4.70 (H-7) to δC 138.5 (C-1), 111.7 (C-2), and 120.0 (C-6) and from δH 4.49 (H-7′) to δC 136.0 (C-1′), 111.5 (C-2′), and 120.7 (C-6′) allowed to fix the location of the two 1,3,4-trisubstituted aromatic rings at C-7 and C-7′, respectively (Figure 2). The HMBC correlations from δH 3.88 to δC 150.8 (C-3) and from δH 3.83 to δC 148.9 (C-3′) indicated that the two methoxyl groups are located at C-3 and C-3′ positions. The position of the β-glucopyranose was determined to be at C-4 based on the HMBC correlations observed from δH 6.94 (H-2) to δC 147.1 (C-4) and from δH 4.91 (H-1′′) to δC 147.1 (C-4). Based on the above analysis, the planar structure of 4 was established. The CD spectrum of 4 showed the negative Cotton effects at 279 nm (∆ε -7.23) and 231 nm (∆ε -16.11), which were in good agreement with those of the related compound, indicating the absolute configuration of 4 to be 7R,8S,7′S,8′R [15]. Thus, the structure of 4 was established as (7R,8S,7′S,8′R)-4,9,4′,7′-tetrahydroxy-3,3′- dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside. The molecular formula of compound 5, C26H34O12 was deduced by its ESIMS ion at m/z 561 [M+Na]+ and 1H and 13C NMR spectra. The 1H and 13C NMR data of 5 were found to be very similar with those of 4, except for the carbon chemical shift values of C-8 (5: δC 55.9 vs 4: δC 53.7) and C-8′ (5: δC 52.7 vs 4: δC 50.8), suggesting that these compounds are stereoisomers at C-8 and C-8′ (Table 1). This was supported by comparing the CD spectrum of 5 with that of the reported lignan glycoside, (7S,8R,7′S,8′S)-4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′- epoxylignan-4-O-β-D-glucopyranoside [5: λmax (∆ε) 284 (+0.52) and 233 (+17.45) nm vs. λmax (∆ε) 281 (+0.88) and 234 (+1.19) nm [15]. Hence, the structure of 5 was identified as shown in Figure 1. 4. CONCLUSION Our chemical study of the acidic methanol extract of the D. metel whole plants led to the isolation and identification of five compounds, namely: kaempferol 3-O-β-D-glucosyl(1→2)-β- D-galactoside 7-O-β-D-glucoside (1), kaempferol 3-O-β-D-glucopyranosyl(1→2)-β-D- glucopyranoside-7-O-α-L-rhamnopyranoside (2), pinoresinol O-β-D-glucopyranoside (3), (7R,8S,7′S,8′R)-4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D-glucopyranoside (4), and (7S,8R,7′S,8′S)-4,9,4′,7′-tetrahydroxy-3,3′-dimethoxy-7,9′-epoxylignan-4-O-β-D- glucopyranoside (5). 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