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column chromatography (4.5 cm i.d.ꢅ26 cm) using H2O containing an in-
creasing proportion of MeOH (10% stepwise elution from 0 to 100%, each
300 ml) to yield six fractions. The first fraction mainly contained sugars and
was not examined further. Fraction 2 was applied to a MCI-gel CHP 20P
column (3.0 cm i.d.ꢅ20 cm) and eluted with H2O–MeOH (10% stepwise
elution from 0 to 60%, each 100 ml) to give 1 (2.1 g). Fraction 3 (571 mg)
was successively subjected to chromatography over Chromatorex ODS
(1.5 cm i.d.ꢅ18 cm, 5% stepwise elution from 0 to 60% MeOH, each
100 ml), and Sephadex LH-20 (2.0 cm i.d.ꢅ32 cm, 10% stepwise elution
from 30 to 100% MeOH, each 100 ml) to yield a crude sample of 4. Purifi-
cation of 4 was achieved by gel filtration chromatography over Sephadex
LH-20 (1.0 cm i.d.ꢅ13 cm) using 8 M urea in 60% acetone30) followed by re-
moval of the urea using a MCI-gel CHP-20P column (1.0 cm i.d.ꢅ13 cm, 0
to 100% MeOH) to give 4 (22.9 mg). Fraction 4 was separated by Chroma-
torex ODS column chromatography (1.5 cm i.d.ꢅ18 cm, 5% stepwise elu-
tion from 0 to 60% MeOH, each 100 ml) to afford 2 (712 mg). Fraction 6
(310 mg) was subjected to Sephadex LH-20 column chromatography (2.0 cm
i.d.ꢅ32 cm, 10% stepwise elution from 50 to 100% MeOH, each 100 ml) to
yield 3 (99.3 mg). Fraction 5 (218.9 mg) contained 2 and 3, and was not sep-
arated further.
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Chem., 49, 5785—5789 (2001).
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Chem., 50, 2142—2148 (2002).
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Kouno I., J. Nat. Prod., 65, 1582—1587 (2002).
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59, 7939—7947 (2003).
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(2005).
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138—147 (1999).
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P., J. Sci. Food Agric., 74, 401—408 (1997).
Theaflavate
C Red amorphous powder. [a]D ꢀ358.8° (cꢄ0.05,
MeOH). IR nmax cmꢀ1: 3381, 1704, 1607, 1520, 1471. UV lmax (MeOH) nm
(log e): 279 (4.7) and 405 (4.2). HR-FAB-MS m/z: 1263.2267 [MꢁH]ꢁ
(Calcd for C64H47O28: 1263.2252). H- and 13C-NMR data: see Table 1. As-
1
signments of the signals were achieved with the aid of 1H–1H COSY, HSQC,
and HMBC spectra.
Bistheaflavate A Brown amorphous powder. [a]D ꢀ419.2° (cꢄ0.12,
MeOH). IR nmax cmꢀ1: 3406, 1699, 1631, 1610, 1519, 1470. UV lmax
(MeOH) nm (log e): 276 (4.6) and 354 (4.1). HR-FAB-MS m/z: 1721.3114
[MꢁH]ꢁ (Calcd for C86H65O39: 1721.3101). 1H- and 13C-NMR data: see
Table 2. Assignments of the signals were achieved with the aid of 1H–1H
COSY, HSQC, and HMBC spectra.
Acknowledgments The authors are grateful to Mr. K. Inada and Mr. N.
Yamaguchi for NMR and MS measurements. This work was supported by a
Grant-in-aid for Scientific Research No. 18510189 from the Japan Society
for the Promotion of Science.
25) Sang S., Tian S., Meng X., Stark R. E., Rosen R. T., Yang C. S., Ho
C.-T., Tetrahedron Lett., 43, 7129—7133 (2002).
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Chem., 12 3009—3017 (2004).
27) Roberts E. A. H., Chem. Ind., 1957, 1354—1355 (1957).
28) Guyot S., Vercauteren J., Cheynier V., Phytochemistry, 42, 1279—
1288 (1996).
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