Two New ent-Kaurane Diterpenoids from Albizia mollis ( Wall . ) Boiv .

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. The previous chemical investigations on Albizia species have led to the isolation of lignans, flavonoids, saponins, alkaloids, together with other secondary metabolites. 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 C 26 H 44 O 8 with five unsaturation degrees by HRFABMS (m/z 483.2959 [M-1] -) and 13 C NMR spectra.The 1 H NMR spectrum of compound 1 clearly showed signals to three methyls at d 0.83 (3H, s), 1.03 (3H, s), 1.07 (3H, s), and one anomeric hydrogen of b-type sugar at d 4.30 (1H, d, J 7.8 Hz).The 13 C NMR ({ 1 H} and DEPT) spectroscopic data (Table 1) revealed the presence of 3 methyls, 10 methylenes, 9 methines and 4 quaternary carbons.Carbon signals at d 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 = C 6 H 8 O 5 formed by fragmentation involving glc moiety C 6 H 10 O 5 ] -and the acid hydrolysis of 1.Comparison of these NMR data with those similar data reported in the literature 13 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′ (d 4.30), H-18 (d 1.03) and H-19 (d 0.83) to C-3 (d 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 (d 3.27) with C-16 (d 80.7), C-13 (d 42.3) and C-15 (d 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. 14The key ROESY correlations of H-3/H-18, H-3/H-5 and H-9/H-5 (Figure 3), indicated a-orientation of C-3 glycoside and b-orientations of H-5 and H-9.The a-orientation of CH 2 OH-17 (dc 70.6) in 1 was elucidated by comparison of the 13 C NMR data with those reported dc 69.8 for a-orientation and dc 66.2 for b-orientation at C-17. 14 Based on the above evidences, compound 1 was identified as 3a, 16b, 17-trihydroxy-ent-kaurane 3-O-b-Dglucopyranoside, named Mollisside A.
Mollisside B (2) was determined to have a molecular formula C 26 H 40 O 9 on the basis of the positive HRFABMS (m/z 519.2565 [M+Na] + ) and 13 C 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 1 H NMR spectrum of compound 2 exhibited three methyls at d 0.88 (3H, s), 1.12 (3H, s), 1.16 (3H, s), one anomeric proton of b-type sugar at d 4.32 (1H, d, J 7.8 Hz) and one olefinic proton at d 6.48 (1H, s).The 13 C NMR spectroscopic data (Table 1) revealed 26 carbon atoms, including one carboxyl (C-17 at d 164.1) and two olefinic carbons (C-16 at d 139.9, C-15 at d 154.7).Comparison of these NMR data with mollisside A showed that compound 2 had the similar skeleton of compound

General experimental procedures
Melting points were measured on a XRC-1 micromelting 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 H 2 O and partitioned successively with EtOAc (3×5 L) and n-BuOH