Two new ent-kaurane diterpenoids from Albizia mollis (Wall.) Boiv

SHORT REPORT

Two new ent-kaurane diterpenoids from Albizia mollis (Wall.) Boiv

Zhong-Quan Cheng^I,II; Dan Yang^I; Yu-Qing Liu^I; Jiang-Miao Hu^I; He-Zhong Jiang^I; Peng-Cheng Wang^I; Ning Li^III; Jun Zhou^I; You-Xing Zhao^I,*

^IState Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, the Chinese Academy of Sciences, Kunming 650204, P. R. China

^IIGraduate School of the Chinese Academy of Sciences, Beijing 100049, P. R. China

^IIISchool of Life Sciences, Anhui University, Hefei 230039, P. R. China

ABSTRACT

Two new kaurane diterpenoids, 3α, 16β, 17-trihydroxy-ent-kaurane 3-O-β-D-glucopyranoside and 2β,3α-dihydroxy-ent-kaur-15-en-17-oic acid 3-O-β-D-glucopyranoside, were isolated from the bark of Albizia mollis (Wall.) Boiv. The structures of two new compounds were elucidated by extensive 1D- and 2D-NMR spectroscopic methods in combination with MS experiments.

Keywords:Albizia mollis, kaurane diterpenoids, mollisside A, mollisside B

RESUMO

Dois novos diterpenos com esqueleto caurano, 3a,16b-17-tri-hidróxi-ent-caurano 3-O-β-D-glucopiranosídeo e ácido 2β,3α-di-hidróxi-ent-caur-15-eno-17-óico 3-O-β-D-glucopiranosídeo, foram isolados das cascas de Albizia mollis (Wall.) Boiv. As estruturas destes dois cauranos foram elucidadas por métodos espectroscópicos uni e bidimensionais e por espectrometria de massas.

Indroduction

Plant species of Albizia are known for their traditional medicinal uses as medicaments of mind unrest, insomnia, physique damage, carbuncle gall in many parts of tropic zone. In China, Albizia mollis, popularly named "maoyehehuan" is one of representative plant species of this family. This plant is well known for its sedative and sleeping pill properties.¹ The previous chemical investigations on Albizia species have led to the isolation of lignans,^2-5 flavonoids,^6-9 saponins,¹⁰ alkaloids,¹¹ together with other secondary metabolites.¹² In search for new and bioactive compounds from A. mollis, the chemical investigation of this plant, collected from Kunming Botanical Garden, Yunnan Province, People's Republic of China, led to the isolation of two new compounds. This report describes the isolation and structural elucidation of two new ent-kaurane diterpenoids named mollissides A (1) and B (2).

Results and Discussion

Mollisside A (1) was isolated as white powder. Its molecular formula was determined to be C₂₆H₄₄O₈ with five unsaturation degrees by HRFABMS (m/z 483.2959 [M-1]^-) and ¹³C NMR spectra. The ¹H NMR spectrum of compound 1 clearly showed signals to three methyls at δ 0.83 (3H, s), 1.03 (3H, s), 1.07 (3H, s), and one anomeric hydrogen of β-type sugar at δ 4.30 (1H, d, J 7.8 Hz). The ¹³C NMR ({¹H} and DEPT) spectroscopic data (Table 1) revealed the presence of 3 methyls, 10 methylenes, 9 methines and 4 quaternary carbons. Carbon signals at δ 106.8 (d), 75.6 (d), 77.7 (d), 71.6 (d), 78.2 (d), 62.8 (t) suggested the presence of one glucose, which were further confirmed by its MS fragmentation peaks at m/z 323 [M-1-160 = C₆H₈O₅ formed by fragmentation involving glc moiety C₆H₁₀O₅]^- and the acid hydrolysis of 1. Comparison of these NMR data with those similar data reported in the literature¹³ showed that compound 1 had the same ent-kaurane skeleton. The linkage of glucose moiety to C-3 was determined by the HMBC correlations from H-1' (δ 4.30), H-18 (δ 1.03) and H-19 (δ 0.83) to C-3 (δ 90.9) (Figure 2). The attachment of one hydroxyl at C-17 in 1 was established by HMQC and HMBC spectra revealing correlations of H-17 (δ 3.27) with C-16 (δ 80.7), C-13 (δ 42.3) and C-15 (δ 53.1). Besides, the relative configuration of 1 was elucidated by a ROESY experiment and by comparison of the NMR data with those reported in the literature.¹⁴ The key ROESY correlations of H-3/H-18, H-3/H-5 and H-9/H-5 (Figure 3), indicated α-orientation of C-3 glycoside and β-orientations of H-5 and H-9. The α-orientation of CH₂OH-17 (δc 70.6) in 1 was elucidated by comparison of the ¹³C NMR data with those reported δc 69.8 for α-orientation and δc 66.2 for β-orientation at C-17.¹⁴ Based on the above evidences, compound 1 was identified as 3α, 16β, 17-trihydroxy-ent-kaurane 3-O-β-D- glucopyranoside, named Mollisside A.

Figure 1

Mollisside B (2) was determined to have a molecular formula C₂₆H₄₀O₉ on the basis of the positive HRFABMS (m/z 519.2565 [M+Na]⁺) and ¹³C NMR (DEPT) spectra, which possessed seven unsaturation degrees. Its IR spectrum showed a broad band (3518-2930 cm^-1) and absorption 1691 cm^-1 for a carboxyl and 1607 cm^-1 for a double bond. The ¹H NMR spectrum of compound 2 exhibited three methyls at δ 0.88 (3H, s), 1.12 (3H, s), 1.16 (3H, s), one anomeric proton of β-type sugar at δ 4.32 (1H, d, J 7.8 Hz) and one olefinic proton at δ 6.48 (1H, s). The ¹³C NMR spectroscopic data (Table 1) revealed 26 carbon atoms, including one carboxyl (C-17 at δ 164.1) and two olefinic carbons (C-16 at δ 139.9, C-15 at δ 154.7). Comparison of these NMR data with mollisside A showed that compound 2 had the similar skeleton of compound 1 except for the distinct differences of chemical shift of C-2 (δ 68.1) adjacent one oxygenated group and C-17, C-15 and C-16 conforming the α,β-unsaturation ketone system. The linkage of the glucose moietiy to C-3 (δ 96.1) was determined by the HMBC correlations from H-1' (δ 4.32), H-18 (δ 1.12) and H-19 (δ 0.88) to C-3 (Figure 2). Besides, the relative configuration of 2 was elucidated by a ROESY experiment and by comparison of the NMR data with 1. The α-orientation of H-2 was established by key ROESY correlations of H-2/H-1α, H-2/H-19 and H-2/H-20 (Figure 3) and the coupling constant J 9.3 Hz observed in the signal corresponding to H-3 (interaction axial- axial with H-2). Moreover, the β-orientations of H-3, H-5 and H-9 were confirmed by ROESY between H-3/H-5, H-3/H-18 and H-5/H-9. Based on the above evidences, the structure of 2 was elucidated as 2β,3α-dihydroxy-ent-kaur-15-en-17-oic acid 3-O-β-D-glucopyranoside, named Mollisside B.

Experimental

General experimental procedures

Melting points were measured on a XRC-1 micro-melting point apparatus and were uncorrected. MS spectra were obtained on a VG Auto Spec-3000 mass spectrometer. 1D and 2D NMR spectra were recorded on Bruker AM-400 MHz and DRX-500 MHz spectrometers, with chemical shifts (d) in ppm relative to TMS as internal standard and coupling constants in hertz (Hz). IR spectra were measured with a Bio-Rad FTS-135 spectrometer with KBr pellets. UV spectra were measured on a Hitachi UV-3210 spectrophotometer. Silica gel (200-300 mesh) for column chromatography was product of the Qingdao Marine Chemical Ltd., Qingdao, P. R. China. Sephadex LH-20 for chromatography was purchased from Amersham Biosciences. Reversed-phase chromatography was with RP-18 (LiChroprep, 40-63 µm, Merck, Darmstadt, Germany).

Plant material

The bark of A. mollis was collected in Kunming Botanical Garden, Yunnan Province, People's Republic of China, in September 2007, and authenticated by Professor Xun Gong. A voucher specimen (ZM080920) has been deposited in the Herbarium of Kunming Institute of Botany, Chinese Academy of Sciences.

Extraction and isolation

The air-dried and powdered bark of A. mollis (18.0 kg) was extracted three times each with 20 L of 95% EtOH under reflux for 3 h. The extract was evaporated and the residue (1050 g) was resuspended in 15 L of H₂O and partitioned successively with EtOAc (3×5 L) and n-BuOH (3×5 L) to yield EtOAc extract (300 g), n-BuOH extract (640 g), respectively. Part of n-BuOH extract (500 g) was applied to a silica gel column chromatography (200-300 mesh) eluted with CHCl₃/CH₃OH/H₂O (10:3:0.5, v/v) to give seven fractions. Fraction 3 (45 g) was purified by column chromatography silica gel with CHCl₃/CH₃OH/H₂O (5:3:1, v/v) to give four fractions, fraction 2 were purified on RP-18 with 40% →100% aqueous CH₃OH, and on Sephadex LH-20 with CH₃OH/CH₃Cl 1:1 to afford compound 1 (17 mg), fraction 3 were purified on RP-18 with 30% →70% aqueous CH₃OH, and on Sephadex LH-20 with CH₃OH to afford compound 2 (15 mg).

Mollisside A

White powder, mp 292-294 ºC; [α]_D²¹ +6.3116 (c 0.101, CH₃OH); UV (CH₃OH) λ_max/nm (log ε): 193(3.587), 205 (3.829), 277 (3.250); IR (KBr) ν_max/cm^-1: 3420, 2937, 1166, 1074, 1020; ¹H NMR (400 MHz, CD₃OD) and ¹³C NMR (100 MHz, CD₃OD) see Table 1; HRESIMS (negative ion) m/z 483.2959 [M-1]^- (Calc. for C₂₆H₄₄O₈, 483.2957); FABMS (negative ion) m/z(%) 483[M-1]^- (100), 391 (5), 323 (15), 282 (5), 159 (6), 113 (5).

Mollisside B

White powder, mp 270-271 ºC; [α]_D²¹ -1.5989 (c 0.246, CH₃OH); UV (CH₃OH) λ_max/nm (log ε): 196 (3.393), 203 (4.108), 276 (2.890); IR (KBr) ν_max/cm^-1: 3518, 3380, 3190, 2980, 2930, 1691, 1607, 1073, 1037; ¹H NMR (500 MHz, CD₃OD) and ¹³C NMR (125 MHz, CD₃OD) see Table 1; HRESIMS (positive ion) m/z 519.2565 [M+Na]⁺ (Calc. for C₂₆H₄₀O₉Na, 519.2570); FABMS (negative ion) m/z(%): 495[M-1]^- (100), 400 (5), 335 (5), 281(6), 123 (4).

Acid hydrolysis of compounds 1 and 2

Compounds 1 (3 mg) and 2 (3mg) in 50% MeOH (2 mL) containing 5% HCl were each heated in a boiling water-bath for 5 h, then cooled for few minutes. The mixture was washed with EtOAc (2 × 2 mL). The H₂O layer was concentrated in vacuo to give a residue. The residue and authentic sugar were dotted to the plate developed with CHCl₃/CH₃OH/H₂O (5/3/0.5, v/v), and phenylenediamine-aniline-phosphoric acid used as spray reagent, followed by heating at 80 ºC. From compounds 1 and 2 glucose was detected, R_f: glucose 0.45.

Supplementary Information

Supplementary data including ¹H, ¹³C NMR (DEPT), HMQC, HMBC, COSY, and ROSEY MS for compounds 1 and 2 are available free of charge at http://jbcs.sbq.org.br as PDF file.

Acknowledgments

This research was supported by the fund (P2008-ZZ22) of State Key Laboratory of Phytochemistry and Plant Resources in West China. We thank the analytical group of the State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences, for measurements of all spectra.

Submitted: January 11, 2010

Published online: May 14, 2010

Supplementary Information

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Publication Dates

• Publication in this collection
  10 Sept 2010
• Date of issue
  2010

History

• Received
  11 Jan 2010
• Accepted
  14 May 2010