Eucosterol Oligoglycosides Isolated from Scilla scilloides and Their Anti-tumor Activity
Chemical and pharmaceutical bulletin
Scilla scilloides (LIND.) DRUCE (Liliaceae) is a traditional Chinese medicine used for the treatment of antidote, blood circulatory activator, and abscess reducer. 1) Eucosterol oligoglycosides 2) isolated from S. scilloides have been proved to have inhibitory effect on 12-O-tetradecanoylphobol-13-acetate (TPA) stimulated 32 P incorporation into phospholipids of HeLa cells. 3,4) On the course of our research for anti-tumor activity from natural sources, we studied the constituents of S.
... uents of S. scilloides, and isolated two new eucosterol oligoglycosides (1-2) and four known compounds (3-6). Herein we present the structural elucidation of new compounds, as well as anti-tumor activities of six eucosterol oligoglycosides. Results and Discussion Repeat column chromatography of the butanol-soluble fraction of the MeOH extract of S. scilloides on Sephadex LH 20 and reversed-phase silica gel columns let to the isolation of six compounds (1-6). The known compounds were identified as scillascillosides E-1 (3), E-2 (4), E-3 (5) and G-1 (6), which had been isolated from this plant, on the basis of their spectral and physical data in comparison with those reported in literature (Chart 1). 2) Compound 1 (scillanoside L-1) was isolated as a white amorphous powder, [a] D Ϫ57.1°. A molecular formula of C 58 H 94 O 28 was established by high-resolution fast atom bombardment mass spectrometry (HR-FAB-MS), which gave a mass of m/z 1239.6012 (Calcd for C 58 H 95 O 28 , 1239.6010). The IR spectrum was showed with the presence of hydroxyl group (3350 cm Ϫ1 ) and a carbonyl group (1725 cm Ϫ1 ). The aglycon of 1 was shown to be a eucosterol derivative from 1 H-NMR signals for a methylene at d H 2.51 (q, Jϭ7.3 Hz), three tertiary methyls (d H 0.90, 1.04, and 1.46), a primary methyl at d H 1.00 (t, Jϭ7.3 Hz), a secondary methyl at d H 1.01 (d, Jϭ6.6 Hz), and two hydroxymethyls at d H 4.34 and 4.61 (each 2H, s). The 13 C-NMR spectrum, in combination with distortion less enhancement by polarization transfer (DEPT) and 1 H-detected multiple quantum coherence (HMQC) experiments, showed signals for a carbonyl carbon at d C 212.6, two sp 2 carbons at d C 135.4 and 136.0, a methylene at d C 32.4, a primary methyl carbon at d C 7.7, and three oxygen-bearing carbons at d C 81.6, 82.1 and 97.1, including two hydroxymethyl carbons at d C 61.1 and 62.8. These data indicated spirocyclic nortriterpene close to the structure of scillascilloside E-1 (3) except for the presence a hydroxy methyl at C-30 (d H 4.34, d C 61.1) in 1. The connectivity of the hydroxy methyl group was confirmed by heteronuclear multiple-bond correlation (HMBC) observed between d H 4.34 (H-30) and d C 48.3 (C-4). On acid hydrolysis, 5) 1 afforded the monosaccharide units which were identified by co-TLC with authentic samples such as glucose, arabinose, and rhamnose; their absolute configurations were determined as D-glucose , L-rhamnose, and L-arabinose by liquid chromatography method developed by Oshima et al. 6) In the acid hydrolysate of 1, D-glucose, Lrhamnose, and L-arabinose were confirmed by comparison of the retention times of their 1-[(S)-N-acetyl-a-methylbenzylamino]-1-deoxyalditol acetate (AMBA) derivatives with those of authentic monosaccharides prepared in similar way, which showed retention times of 33.4, 20.0, and 26.3 min in the HPLC, respectively. In the correlation spectroscopy (COSY), HMQC, and total correlation spectroscopy (TOCSY) spectra of 1, correlations were observed between the anomeric signals at d H 5.84 and signals d H 4. 61, 4.54, 4.25, 4.92, and 1.78. The series of signals were considered to be due to the protons of a rhamnose, because correlations were found between the methyl signal at d 18.8 and two of the proton signals at d 4.25 (rhaЉЉ-H-4) and 4.92 (rhaЉЉ-H-5) in the HMBC spectrum. Similarly, the spin system for the glucoses and arabinose were assigned, since the anomeric proton signals at d 5.02, 5.11, 5.14, and 5.28 were correlated with the signals as Table 2 . Furthermore, the connectivity of the sugars were determined on the presence of correlations between a proton signal at d H 5.02 (glc-H-1Ј) and a carbon signal at d C 82.