Extraction and Isolation. The seeds of Ziziphus jujuba var. spinosa were extracted with 95% aqueous EtOH (3 ꢃ 20 L).
After removal of the solvent under reduced pressure, the extract was dissolved in water and partitioned with petroleum, ethyl
acetate, and n-BuOH, successively. The n-BuOH extract (252 g) was fractioned by HP-20 (2.5 kg) eluted with 20%, 50%, and
80% aqueous EtOH to give fractions A (20%), B (50%), and C (80%), respectively. Fractions B (30 g) was further fractionated
by a silica gel column (250 g) eluted with gradient CHCl –MeOH (10:1ꢄ8:2ꢄ6:4ꢄ2:8) to give four subfractions (B 1-4).
3
Subfraction B2 was subjected to column chromatography on polyamide eluted with gradient CHCl –MeOH to give fractions
3
B 2-1 to B 2-3. Fraction B 2-2 was further separated on a RP-18 column using gradient aqueous MeOH, followed by a
Sephadex LH-20 column eluted with aqueous MeOH to afford 1 (4 mg), 2 (85 mg), 3 (230 mg), and 4 (18 mg), respectively.
Hydrolysis and GC Chromatography. Two mg of compound 1 was refluxed with 5 mL 2M HCl for 2 h at 100ꢅC.
The reaction mixture was extracted with ethyl acetate. The residue was evaporated to dryness. The gas-liquid chromatography
experiment was finished using the reported method [6].
Compound 1 (6ꢀꢀꢀ-vanilloylspinosin), yellow powder, mp 225–228ꢅC. UV spectrum (MeOH,
, nm): 276, 335;
max
(NaOMe,
, nm): 276, 392; (NaOAc,
, nm): 276, 393; (AlCl ,
, nm): 284, 302, 351, 385; (AlCl -HCl,
, nm):
max
max
3
max
3
max
–1
284, 302, 351, 385. IR spectrum (KBr, ꢆ , cm ): 3340, 2928, 1725, 1608, 1585, 1502, 1483, 1380, 1345, 1278, 1200, 1175,
max
–
+
1093, 1078, 1021. ESI-MS m/z 757 [M – H] , HR-ESI-MS m/z 781.1953, calcd for C H O Na [M + Na] 781.1956. For
36 38 18
1
13
H NMR (400 MHz, DMSO-d ) (300 K) spectral data, see Table 2, and for C NMR (100 MHz, DMSO-d ) (300 K) spectral
6
6
data, see Table 1.
–
Compound 2 (spinosin), yellow powder, mp 237–240ꢅC. UV (MeOH,
, nm): 270, 334; ESI-MS m/z 607 [M – H] .
max
1
13
For H NMR (400 MHz, DMSO-d ) (300 K) spectral data, see Table 2, and for C NMR (100 MHz, DMSO-d ) (300 K)
6
6
spectral data, see Table 1.
Compound 3 (6ꢀꢀꢀ-feruloylspinosin), yellow powder, mp 224ꢅC (dec.). UV (MeOH,
, nm): 273, 328; ESI-MS m/z
max
13
–
1
783 [M – H] . For H NMR (400 MHz, DMSO-d ) (300 K) spectral data, see Table 2, and for C NMR (100 MHz, DMSO-d )
6
6
(300 K) spectral data, see Table 1.
Compound 4 (isospinosin), yellow powder, mp 215–216ꢅC. UV (MeOH,
, nm): 268, 330. ESI-MS m/z 607
max
–
1
13
[M – H] . For H NMR (400 MHz, DMSO-d ) (300 K) spectral data, see Table 2, and for C NMR (100 MHz, DMSO-d )
6
6
1
(300 K) spectral data, see Table 1. For comparison with the literature [3], we modified its H NMR spectral data.
ACKNOWLEDGMENT
The authors are grateful for the support of Tianjin Zhongxin Pharmaceuticals (China).
REFERENCES
1.
M. Yoshikawa, T. Murakami, A. Ikebata, S. Wakao, N. Murakami, H. Matsuda, and J. Yamahara, Chem. Pharm. Bull.,
45, 1186 (1997).
2.
3.
4.
5.
6.
H. Matsuda, T. Murakami, A. Ikebata, J. Yamahara, and M. Yoshikawa, Chem. Pharm. Bull., 47, 1744 (1997).
G. Cheng, Y. J. Bai, Y. Y. Zao, J. Tao, Y. Liu, G. Z. Tu, L. B. Ma, N. Liao, and X. J. Xu, Tetrahedron, 56, 8915 (2000).
B. H. Han, M. H. Park, and Y. N. Han, Phytochemistry, 29, 3315 (1997).
M. H. Park, D. Y. Suh, and B. H. Han, Phytochemistry, 43, 701 (1996).
S. Hara, H. Okabe, and K. Mihashi, Chem. Pharm. Bull., 35, 501 (1987).
372