Novel norsesquiterpenoids from the roots of Phyllanthus emblica

Article abstract of DOI:10.1021/np000135i

Three ester glycosides, named phyllaemblicins A (3), B (4), and C (5), and a methyl ester (2), of a highly oxygenated norbisabolane, phyllaemblic acid (1), were isolated from the roots of Phyllanthus emblica, along with 15 tannins and related compounds. The structures of 2-5 were established by spectral and chemical methods.

Full text of DOI:10.1021/np000135i

J . Nat. Prod. 2000, 63, 1507-1510
1507
Novel Nor sesqu iter p en oid s fr om th e Roots of P h ylla n th u s em blica
Ying-J un Zhang,† Takashi Tanaka,† Yoko Iwamoto,† Chong-Ren Yang,‡ and Isao Kouno*,†
Faculty of Pharmaceutical Sciences, Nagasaki University, Bunkyo Machi 1-14, Nagasaki 852-8521, J apan, and Laboratory of
Phytochemistry, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650204, People’s Republic of China
Received April 7, 2000
Three ester glycosides, named phyllaemblicins A (3), B (4), and C (5), and a methyl ester (2), of a highly
oxygenated norbisabolane, phyllaemblic acid (1), were isolated from the roots of Phyllanthus emblica,
along with 15 tannins and related compounds. The structures of 2-5 were established by spectral and
chemical methods.
Phyllanthus emblica L. (Euphorbiaceae), a shrub or tree
growing in subtropical and tropical areas of the People’s
Republic of China, India, Indonesia, and the Malay Pen-
insula, has been used widely for its antiinflammatory and
antipyretic effects in many local traditional medicinal
systems, such as Chinese herbal medicine, Tibetan medi-
cine, and Ayurvedic medicine. The minorities living in the
Southwest of China use the roots of P. emblica for the
treatment of eczema, and in Nepal it is used as an
astringent and hematostatic agent.1 In a previous com-
munication,2 we reported the structure elucidation includ-
ing the absolute stereochemistry of a novel highly oxygen-
ated norbisabolane, phyllaemblic acid (1), from the roots
of this plant. Further investigation has led to the isolation
of its methyl ester (2) and three novel ester glycosides,
phyllaemblicins A (3), B (4), and C (5), along with 15
tannins and related compounds. This paper deals with a
full account of the isolation and structural elucidation of
2-5.
Resu lts a n d Discu ssion
A 60% aqueous acetone extract of the air-dried roots of
P. emblica was partitioned between EtOAc and water, and
then the organic phase was subjected successively to
chromatography over Sephadex LH-20, MCI gel CH20P,
Chromatorex ODS, and silica gel to afford compounds 1-5
and 15 substances of known structure. By comparison of
the physical and spectral data with those of authentic
samples, the known compounds were identified as gallic
acid, pyrogallol, protocatechuic acid, corilagin,3 1,2,3,6-
tetra-O-galloyl-â-D-glucose,4 1,2,3,4,6-penta-O-galloyl-â-D-
glucose,5 (()-gallocatechin,6 (-)-epigallocatechin,7 (+)-
The structure of compound 1 was characterized previ-
catechin,7 (-)-epicatechin,7 (-)-epigallocatechin 3-O-gallate,7
ously as a novel norbisabolane-type terpenoid by spectro-
(-)-epicatechin 3-O-gallate,7 and epicatechin-(4âf8)-epi-
scopic and chemical means, and its absolute stereochem-
gallocatechin 3-O-gallate,8 respectively. Compounds 6 and
istry was determined by applying the modified Mosher’s
7 were characterized as epigallocatechin-(2âf7,4âf8)-
method to an opportune degradation product from our
gallocatechin (prodelphinidin A-1) and epicatechin-(4âf8)-
previous work.2
gallocatechin by negative-ion FABMS and 1H NMR com-
Compound 2 was obtained as a white amorphous powder.
parison, in turn, with proanthocyanidin A-19 and procyanidin
Comparison of the 1H and 13C NMR spectral data with
B-1.10 The structure of 7 was also confirmed by thiolysis
those of 1 enabled 2 to be assigned as a methyl ester of 1,
(mercaptoethanol-HCl), yielding gallocatechin and epicat-
and it was confirmed by the treatment of 1 with CH2N2,
echin-4-(2-hydroxy)ethylthio ether.11 The spectroscopic data
yielding 2. Hence, 2 was characterized as a methyl ester
have been assigned for the first time for compounds 6 and
of phyllaemblic acid.
7.
Compound 3 was obtained as a white amorphous powder.
Its molecular formula was assigned as C27H34O14 on the
* To whom correspondence should be addressed. Tel.: +81-95-847-1111.
basis of the NMR data, the positive-ion FABMS (m/z 583,
Fax: +81-95-848-4387. E-mail: ikouno@net.nagasaki-u.ac.jp.
1
[M + H]+), and elemental analysis. The H and 13C NMR
† Nagasaki University.
‡ Kunming Institute of Botany.
spectral data of 3 were closely related to those of 1, except
10.1021/np000135i CCC: $19.00
© 2000 American Chemical Society and American Society of Pharmacognosy
Published on Web 09/30/2000

1508 J ournal of Natural Products, 2000, Vol. 63, No. 11
Zhang et al.
for an upfield shift of C-13 (δ 176.2) and the appearance of
signals due to a â-D-glucopyranose moiety. Chemical shifts
of the anomeric proton [δΗ 5.46 (d, J ) 7.5 Hz)] and carbon
(δ 95.64) signals indicated that the glucose moiety was
attached to the C-13 carboxyl group of 1 through an ester
linkage. This was supported by methanolysis with 2%
NaOMe yielding 2 and acidic hydrolysis affording glucose
and 1. On the basis of the above results, the structure of 3
was determined as an ester glucoside of 1 and named
phyllaemblicin A.
Several bisabolane glycosides, namely, phyllanthoside
and the phyllanthostatins,12,13 isolated from Phyllanthus
brasiliensis, have been found to inhibit strongly the growth
of the murine P-388 lymphocytic leukemia cell line. How-
ever, compounds 1 and 3, with a similar norbisabolane
skeleton, showed only weak activity for inhibition of protein
kinases18-20 and no activity on human cancer cell line panel
screening.21,22
Exp er im en ta l Section
Compound 4 showed the [M + H]+ ion peak at m/z 745
in the FABMS, which was 162 mass units larger than that
of 3. This difference corresponded to the mass of an
additional hexopyranose moiety. The 1H and 13C NMR
spectra of 4 resembled those of 3, which exhibited signals
due to two hexose moieties together with a set of signals
almost superimposable on those of 1. The sugar moieties
were both determined to be glucose by acidic hydrolysis
with 0.5 N H2SO4, and the configurations of the anomeric
positions were assigned as â on the basis of the J values of
their anomeric protons (δ 5.59, d, J ) 8.0 Hz and δ 4.18, d,
J ) 8.5 Hz). The HMBC spectrum of 4 showed long-range
correlations of the anomeric proton of the inner glucose (δ
5.59) with C-13 (δ 175.8) and the anomeric proton of the
terminal glucose (δ 4.18) with C-2 of the inner glucose (δ
83.1), which indicated that the additional glucose moiety
was attached to the C-2 position of the glucose moiety of 3
in structure 4. Accordingly, the structure of 4 was estab-
lished and named phyllaemblicin B.
Gen er a l Exp er im en ta l P r oced u r es. Optical rotations
were measured with a J ASCO DIP-370 digital polarimeter.
1H and 13C NMR spectra were recorded in CD3OD with Varian
Unity plus 500 and Varian Gemini 300 spectrometers operat-
ing at 500 and 300 MHz for 1H and 125 and 100 for 13C,
respectively. Coupling constants are expressed in hertz, and
chemical shifts are given on a δ (ppm) scale with tetrameth-
ylsilane as the internal standard. MS spectra were recorded
on a J EOL J MS DX-303 spectrometer, and glycerol was used
as a matrix for FABMS measurement. Column chromatogra-
phy was performed with Kieselgel 60 (70-230 mesh, Merck),
MCI-gel CHP 20P (75-150 µm, Mitsubishi Chemical Co.),
Sephadex LH-20 (25-100 µm, Pharmacia Fine Chemical Co.
Ltd.), and Chromatorex ODS (100-200 mesh, Fuji Silysia
Chemical Ltd.). TLC was performed on precoated Kieselgel 60
F254 plates (0.2 mm thick, Merck), and spots were detected by
ultraviolet (UV) illumination and by spraying with 10%
sulfuric acid reagent.
P la n t Ma ter ia l. The roots of Phyllanthus emblica L. were
collected at Wenshan, Yunnan, People’s Republic of China. A
voucher specimen is deposited in the Herbarium of Kunming
Institute of Botany, Chinese Academy of Sciences.
The 1H and 13C NMR spectra of compound 5 were closely
related to those of 4 except for the appearance of a set of
additional sugar signals, indicating that 5 is also a glyco-
side of 1. The FABMS (m/z 877, [M + H]+) and elemental
analysis enabled the molecular formula C33H44O19 to be
assigned to 5. The presence of three sugar residues in 5
was indicated by the appearance of three anomeric proton
signals at δ 5.59 (d, J ) 8.5 Hz), 4.14 (d, J ) 8.0 Hz), and
Extr a ction a n d Isola tion . The air-dried roots (8.0 kg)
were extracted with 60% aqueous acetone four times at room
temperature to give an extract (450 g), which was partitioned
between EtOAc (1 L) and H2O (1 L) four times. After
concentration in vacuo, the EtOAc layer (46.7 g) was chro-
matographed on Sephadex LH-20 (60%-100% MeOH then 60%
acetone) to afford five fractions (fractions 1-5). Fraction 1 (7.76
g) was separated successively by passage over MCI gel CHP
20P (H2O-MeOH, 1:0-0:1), Chromatorex ODS (10-70%
MeOH), and Si gel (CHCl3-MeOH-H2O, 9:1:0.1-6:4:1) to
afford compounds 2 (518 mg), 3 (190 mg), 4 (430 mg), and 5
(133 mg). Fraction 2 (8.66 g) was chromatographed over
Chromatorex ODS (40-100% MeOH), MCI gel CHP 20P (0-
30% MeOH), and silica gel (CHCl3-MeOH-H2O, 9:1:0.1-8:
2:0.2) to give 1 (3.4 g), gallic acid (226 mg), and pyrogallol (472
mg). Fractions 3 (6.0 g) and 4 (8.31 g) were chromatographed
repeatedly on MCI gel CHP 20P (10-80% MeOH), Sephadex
LH-20 (EtOH and 60% MeOH), and Chromatorex ODS (5-
50% MeOH) to afford protocatechuic acid (396 mg), corilagin
(35 mg), 1,2,3,6-tetra-O-galloyl-â-D-glucose (68 mg), 1,2,3,4,6-
penta-O-galloyl-â-D-glucose (102 mg), (()-gallocatechin (122
mg), and (-)-epigallocatechin (89 mg) from fraction 3, and (+)-
catechin (19 mg), (-)-epicatechin (116 mg), (-)-epigallocatechin
3-O-gallate (35 mg), (-)-epicatechin 3-O-gallate (105 mg), and
6 (64 mg) from fraction 4, respectively. Fraction 5 (14.59 g)
was chromatographed over MCI gel CHP 20P (10-90% MeOH)
and Chromatorex ODS (10-65% MeOH) to give epicatechin-
(4âf8)-epigallocatechin 3-O-gallate (456 mg) and 7 (58 mg).
1
4.47 (d, J ) 7.0 Hz) in the H NMR spectrum. Taking the
molecular weight into account, the sugar moieties were
concluded to be two hexoses and a pentose unit. Acidic
hydrolysis of 5 with 0.5 N H2SO4 afforded D-glucose ([R]D
+16.3°) and L-arabinose ([R]D +22.3°). The coupling con-
stants of the anomeric protons of the two glucose moieties
indicated their anomeric positions to be in the â configu-
ration, and coupling constants (J 1,2 ) 7.0 Hz, J 2,3 ) 9.0 Hz,
J 3,4 < 1 Hz, J 4,5 ) 3.0 and <1 Hz) of the L-arabinose moiety
suggested it to be in the R-pyranose form. Connectivities
of the sugar moieties were revealed by the HMBC correla-
tions of C-13 with H-1 of the inner glucose, C-2 of the inner
glucose with H-1 of the middle glucose, and C-2 of the
middle glucose with H-1 of the arabinose, which suggested
strongly that the additional arabinose moiety was attached
to the C-2 position of the terminal glucose of 4 in structure
5. This was further confirmed by complete assignments of
the sugar proton signals observed in the 1H NMR spectrum
of the peracetate 5a , in which neither of the C-2 methine
protons of the glucose units (δ 3.85 and 3.56) showed any
acylation shift. On the basis of the above results, the
structure of 5 was determined and named phyllaemblicin
C.
Meth yl ester of p h ylla em blic a cid (2): white amorphous
powder; [R]25 +19.3° (c 0.85, MeOH); UV (MeOH) λmax (log ꢀ)
D
272 (3.00), 279 (2.93) nm; IR (KBr) νmax 3406, 2954, 1777, 1718,
1603, 1450, 1277, 1114, 1006 cm-1 1H NMR (CD3OD, 300
;
MHz) δ 8.14 (2H, dd, J ) 8.0, 1.5 Hz, H-3′, 7′), 7.61 (1H, tt,
J ) 1.5, 8.0 Hz, H-5′), 7.48 (2H, br t, J ) 8.0 Hz, H-4′, 6′), 5.30
(1H, q, J ) 3.0 Hz, H-10), 4.25 (1H, br d, J ) 1.8 Hz, H-5),
4.03 (1H, t, J ) 11.4 Hz, H-12a), 3.89 (1H, br s, H-1), 3.61
(3H, s, OMe), 3.58 (1H, dd, J ) 11.4, 4.2 Hz, H-12b), 2.79 (1H,
tt, J ) 12.9, 3.0 Hz, H-3) 2.34 (1H, dd, J ) 14.7, 3.3 Hz, H-9a),
2.23 (1H, br d, J ) 14.4 Hz, H-4a), 2.18 (1H, m, H-11), 1.95
(1H, dd, J ) 14.7, 3.3 Hz, H-9b), 1.86 (1H, m, H-2a), 1.82 (1H,
Although several norbisabolenes with a skeleton repre-
sentative of the loss of one of the terminal dimethyl carbons
are so far known,16,17 compounds 1-5 with a highly
oxygenated aglycon are the first norbisabolanes biosyn-
thesized by oxidative removal of the central methyl car-
bon.16

Norsesquiterpenoids from Phyllanthus
J ournal of Natural Products, 2000, Vol. 63, No. 11 1509
m, H-4b), 1.69 (1H, dt, J ) 12.9, 2.4 Hz, H-2b), 0.91 (3H, d,
J ) 6.9 Hz, H-14); 13C NMR (CD3OD, 75 MHz) δ 213.8 (C-7),
177.7 (C-13), 167.9 (C-1′), 134.0 (C-5′), 132.1 (C-2′), 130.8 (C-
3′, 7′), 129.4 (C-4′, 6′), 100.5 (C-8), 76.3 (C-5), 75.6 (C-6), 71.4
(C-1), 71.2 (C-10), 63.5 (C-12), 52.1 (OMe), 34.2 (C-11), 33.0
(C-2), 32.5 (C-9), 31.8 (C-3), 29.1 (C-4), 13.1 (C-14); FABMS
m/z 435 [M + H]+.
(glc C-3), 70.8 (glc C-4), 62.3 (glc C-6); terminal glucose, 105.9
(glc C-1), 77.7 (glc C-3), 77.6 (glc C-5), 75.9 (glc C-2), 70.7 (glc
C-4), 62.0 (glc C-6); FABMS m/z 767 [M + Na]+ (18), 745
[M + H]+ (31), 583 (29), 421 (28), 299 (49), 105 (31); anal. C
50.19%, H 6.25%, calcd for C33H44O19‚5/2 H2O, C 50.13%, H
5.91%. Acidic hydrolysis of 4 and TLC analysis in a manner
similar to that described for
3 showed the presence of
Meth yla tion of 1. A solution of 1 (15 mg) in MeOH (2 mL)
was treated with CH2N2/Et2O at room temperature. After
concentration, the mixture was purified by Si gel column
chromatography (CHCl3-MeOH-H2O, 80:20:2) to afford 2 (8
mg).
compound 1 and glucose in the products.
P h ylla em blicin C (5): yellowish amorphous powder; [R]25
D
+14.5° (c 0.52, MeOH); UV (MeOH) λmax (log ꢀ) 272 (3.20), 279
(3.12) nm; IR (KBr) νmax 3349, 2931, 1779, 1711, 1603, 1452,
1
1279, 1171, 1073, 1027 cm-1; H NMR (CD3OD, 500 MHz) δ
P h ylla em blicin A (3): white amorphous powder; [R]25
aglycon: 8.18 (2H, dd, J ) 7.5, 1.0 Hz, H-3′, 7′), 7.69 (1H, t,
J ) 7.5 Hz, H-5′), 7.57 (2H, t, J ) 7.5 Hz, H-4′, 6′), 5.39 (1H,
d, J ) 3.0 Hz, H-10), 4.31 (1H, br s, H-5), 4.03 (1H, t, J ) 11.0
Hz, H-12a), 3.94 (1H, br s, H-1), 3.57 (1H, br d, J ) 11.0 Hz,
H-12b), 2.93 (1H, tt, J ) 13.5, 2.5 Hz, H-3), 2.45 (1H, dd, J )
13.5, 2.5 Hz, H-4a), 2.24 (1H, dd, J ) 15.0, 3.0 Hz, H-9a), 2.18
(1H, m, H-11), 2.07 (1H, dd, J ) 15.0, 3.0 Hz, H-9b), 1.99 (1H,
dd, J ) 13.5, 2.0 Hz, H-2a), 1.88 (1H, dt, J ) 13.5, 3.5 Hz,
H-4b), 1.79 (1H, dt, J ) 13.5, 2.5 Hz, H-2b), 0.88 (3H, d, J )
7.0 Hz, H-14); inner glucose, 5.59 (1H, d, J ) 8.5 Hz, glc H-1),
3.90 (1H, dd, J ) 12.0, 2.5, glc H-6a), 3.76 (1H, dd, J ) 12.0,
5.0 Hz, glc H-6b), 3.57 (1H, dd, J ) 9.5, 9.0 Hz, glc H-3), 3.51
(1H, t, J ) 9.0 Hz, glc H-4), 3.41 (1H, m, glc H-4), 3.31 (1H,
dd, J ) 8.5, 9.0 Hz, glc H-2), middle glucose: 4.14 (1H, d, J )
8.0 Hz, glc H-1), 3.46 (1H, dd, J ) 12.0, 2.0 Hz, glc H-6a), 3.52
(1H, dd, J ) 12.0, 4.0 Hz, glc H-6b), 3.37 (1H, m, glc H-5),
3.29 (1H, dd, J ) 8.5, 9.0 Hz, glc H-4), 3.24 (1H, dd, J ) 8.0,
9.5, glc H-2), 2.48 (1H, br t, J ) 9.5 Hz, glc H-3); arabinose,
4.47 (1H, d, J ) 7.0 Hz, ara H-1), 3.99 (1H, dd, J ) 12.0, 3.0
Hz, ara H-5a), 3.64 (1H, ss, J ) 9.0, 7.0 Hz, ara H-2), 3.63
(1H, br d, J ) 12.0 Hz, ara H-5b), 3.57 (1H, br d, J ) 9.0 Hz,
ara H-3), 3.82 (1H, br s, ara H-4); 13C NMR (CD3OD, 125 MHz)
δ 213.5 (C-7), 175.7 (C-13), 167.7 (C-1′), 134.5 (C-5′), 132.1 (C-
2′), 130.9 (C-3′, 7′), 130.0 (C-4′, 6′), 100.5 (C-8), 76.3 (C-5), 75.4
(C-6), 71.5 (C-1), 70.8 (C-10), 63.3 (C-12), 34.3 (C-11), 32.8 (C-
9), 32.1 (C-2, 3), 29.6 (C-4), 13.1 (C-14); inner glucose, 93.5
(glc C-1) 84.5 (glc C-2), 78.9 (glc C-5), 77.5 (glc C-3), 70.0 (glc
C-4), 62.3 (glc C-6); middle glucose, 104.8 (glc C-1), 85.2 (glc
C-2), 77.7 (glc C-5), 76.9 (glc C-5), 70.1 (glc C-4), 61.4 (glc C-6);
arabinose, 107.2 (ara C-1), 74.0 (ara C-3), 73.7 (ara C-2), 69.5
(ara C-4), 67.7 (ara C-5); FABMS m/z 899 [M + Na]+ (13), 877
[M + H]+ (20), 583 (5), 421 (32); anal. C 48.56%, H 6.33%,
calcd for C38H52O23‚7/2H2O, C 48.41%, H 5.89%.
D
+27.9° (c 0.27, MeOH); UV (MeOH) λmax (log ꢀ) 273 (2.99), 279
(2.91) nm; IR (KBr) νmax 3292, 2930, 1778, 1720, 1603, 1452,
1286, 1124, 1012 cm-1; 1H NMR (CD3OD, 500 MHz) δ aglycon
8.13 (2H, d, J ) 7.5 Hz, H-3′, 7′), 7.63 (1H, br t, J ) 7.5 Hz,
H-5′), 7.54 (2H, br t, J ) 7.5 Hz, H-4′, 6′), 5.32 (1H, br d, J )
3.5 Hz, H-10), 4.28 (1H, br s, H-5), 4.01 (1H, t, J ) 11.5 Hz,
H-12a), 3.91 (1H, br s, H-1), 3.57 (1H, dd, J ) 11.5, 4.5 Hz,
H-12b), 2.95 (1H, t, J ) 13.0 Hz, H-3), 2.31 (1H, br d, J ) 15.0
Hz, H-4a), 2.27 (1H, dd, J ) 15.0, 3.5 Hz, H-9a), 2.18 (1H, m,
H-11), 1.99 (1H, dd, J ) 15.0, 3.5 Hz, H-9b), 1.93 (1H, br d,
J ) 13.0 Hz, H-2a), 1.90 (1H, dt, J ) 15.0, 4.0 Hz, H-4b), 1.78
(1H, dt, J ) 13.0, 2.0 Hz, H-2b), 0.90 (3H, d, J ) 7.0 Hz, H-14),
glucose: 5.46 (1H, d, J ) 8.0 Hz, glc H-1), 3.89 (1H, br d, J )
12.0 Hz, glc H-6a), 3.73 (1H, dd, J ) 12.0, 3.5 Hz, glc H-6b),
3.39 (1H, m, glc H-5), 3.31 (2H, m, glc H-3, 4), 3.25 (1H, t,
J ) 8.0 Hz, glc H-2); 13C NMR (CD3OD, 125 MHz) δ 213.8
(C-7), 176.2 (C-13), 168.0 (C-1′), 134.4 (C-5′), 132.0 (C-2′), 130.7
(C-3′, 7′), 129.7 (C-4′, 6′), 100.5 (C-8), 76.2 (C-5), 75.5 (C-6),
71.2 (C-1), 71.1 (C-10), 63.4 (C-12), 34.3 (C-11), 32.8 (C-2), 32.6
(C-9), 32.3 (C-3), 29.0 (C-4), 13.1 (C-14); glucose, 95.6 (glc C-1)
78.8 (glc C-5), 78.1 (glc C-3), 74.0 (glc C-2), 71.0 (glc C-4), 62.4
(glc C-6); FABMS m/z 605 [M + Na]+ (22), 583 [M + H]+ (8),
421 [M - glc]+ (63), 299 (52), 105 (100); anal. C 53.60%, H
6.08%, calcd for C27H34O14‚5/4 H2O, C 53.62%, H 5.93%.
Acid ic Hyd r olysis of 3. A solution of 3 (1 mg) in 0.5 N
H2SO4 (0.5 mL) was heated at 80 °C for 1 h. The mixture was
neutralized with Amberlite IRA-400 (OH- form) resin and
concentrated to dryness. TLC analysis indicated the presence
of compound 1 and glucose [CHCl3-MeOH-H2O, 7:3:0.5, for
glucose (Rf 0.1), and 9:1:0.1 for 1 (Rf 0.5)].
Meth a n olysis of 3. A solution of 3 (6 mg) in 2% NaOMe in
MeOH (1 mL) was left standing at room temperature for 30
min. After neutralization with Amberlite IR-120B (H+ form)
resin, the solution was concentrated to dryness and applied
to a Si gel column to afford 2 (2 mg).
Acid ic Hyd r olysis of 5. A solution of 5 (10 mg) in 0.5 N
H2SO4 (1 mL) was heated at 80 °C for 21 h. The mixture was
neutralized with Amberlite IRA-400 (OH- form) resin, con-
centrated in vacuo, and then chromatographed over Si gel with
P h ylla em blicin B (4): yellowish amorphous powder; [R]25
D
+10.4° (c 0.58, MeOH); UV (MeOH) λmax (log ꢀ) 272 (2.99), 279
(2.91) nm; IR (KBr) νmax 3205, 2933, 1778, 1718, 1604, 1452,
CHCl3-MeOH-H2O (7:3:0.5) to yield D-glucose [(3.9 mg) [R]25
D
+16.3° (c 0.3, H2O), Rf 0.1 (CHCl3-MeOH-H2O, 7:3:0.5)] and
1294, 1124, 1012 cm-1; H NMR (CD3OD, 500 MHz) δ agly-
L-arabinose [(2.1 mg), [R]25 +22.3° (c 0.16, H2O), Rf 0.2
1
D
con: 8.16 (2H, dd, J ) 7.5, 1.5 Hz, H-3′, 7′), 7.66 (1H, br t,
J ) 7.5 Hz, H-5′), 7.57 (2H, br t, J ) 7.5 Hz, H-4′, 6′), 5.36
(1H, q, J ) 3.0 Hz, H-10), 4.29 (1H, br s, H-5), 4.03 (1H, t,
J ) 11.0 Hz, H-12a), 3.93 (1H, br s, H-1), 3.58 (1H, br d, J )
11.0 Hz, H-12b), 2.94 (1H, tt, J ) 13.5, 2.5 Hz, H-3), 2.36 (1H,
br d, J ) 13.5 Hz, H-4a), 2.28 (1H, dd, J ) 15.0, 3.0 Hz, H-9a),
2.18 (1H, m, H-11), 2.01 (1H, dd, J ) 15.0, 3.0 Hz, H-9b), 2.04
(1H, br d, J ) 13.5 Hz, H-2a), 1.90 (1H, dt, J ) 13.5, 4.0 Hz,
H-4b), 1.77 (1H, dt, J ) 13.5, 2.5 Hz, H-2b), 0.89 (3H, d, J )
7.0 Hz, H-14); inner glucose, 5.59 (1H, d, J ) 8.0 Hz, glc H-1),
4.08 (1H, dd, J ) 9.0, 8.0, glc H-2), 3.89 (1H, dd, J ) 12.0, 2.5
Hz, glc H-6a), 3.75 (1H, dd, J ) 12.0, 5.0 Hz, glc H-6b), 3.62
(1H, t, J ) 9.0 Hz, glc H-3), 3.46 (1H, dd, J ) 9.5, 9.0 Hz, glc
H-4), 3.39 (1H, m, glc H-5), terminal glucose: 4.18 (1H, d, J )
8.0 Hz, glc H-1), 3.62 (1H, dd, J ) 12.0, 2.5 Hz, glc H-6a), 3.56
(1H, dd, J ) 12.0, 4.5 Hz, glc H-6b), 3.25 (1H, dd, J ) 9.5, 9.0
Hz, glc H-4), 3.23 (1H, dd, J ) 9.5, 9.0 Hz, glc H-4), 3.11 (1H,
dd, J ) 9.0, 8.0, glc H-2), 2.76 (1H, m, glc H-5); 13C NMR (CD3-
OD, 125 MHz) δ 213.7 (C-7), 175.8 (C-13), 167.8 (C-1′), 134.4
(C-5′), 132.1 (C-2′), 130.8 (C-3′, 7′), 129.9 (C-4′, 6′), 100.5 (C-
8), 76.3 (C-5), 75.4 (C-6), 71.5 (C-1), 70.9 (C-10), 63.4 (C-12),
34.3 (C-11), 32.7 (C-9), 32.2 (C-2, 3), 29.3 (C-4), 13.1 (C-14);
inner glucose, 93.7 (glc C-1) 83.1 (glc C-2), 78.9 (glc C-5), 77.8
(CHCl3-MeOH-H2O, 7:3:0.5)].
Acetyla tion of 5. Compound 5 (32 mg) was treated with
pyridine (1 mL) and Ac2O (1 mL) at room temperature
overnight. After concentration in vacuo, the residue was
subjected to passage over a Si gel column to give 5a (19.3
mg): white powder, [R]25 +20.8° (c 0.27, CHCl3); 1H NMR
D
(CDCl3, 500 MHz) δ 8.13 (2H, dd, J ) 8.0,1.0 Hz, H-3′, 7′),
7.58 (1H, tt, J ) 1.0, 8.0 Hz, H-5′), 7.49 (2H, br t, J ) 8.0 Hz,
H-4′, 6′), 5.33 (1H, q, J ) 3.0 Hz, H-10), 4.94 (1H, br s, H-1),
4.38 (1H, br s, H-5), 3.64 (1H, dd, J ) 4.5,11.0 Hz, H-12b),
2.59 (1H, tt, J ) 3.0, 12.5 Hz, H-3), 2.34 (1H, br d, J ) 13.0
Hz, H-4a), 2.27 (1H, dd, J ) 3.0, 15.0 Hz, H-9a), 2.24 (3H, s,
CH3CO-1), 2.23 (1H, m, H-11), 2.10 (1H, dd, J ) 3.0, 15.0 Hz,
H-9b), 2.07, 2.05, 2.01, 1.99 (each 6H, s, CH3CO), 1.97 (3H, s,
CH3CO), 1.94-2.02 (2H, m, H-2a, 4b), 1.83 (1H, ddd, J ) 2.5,
3.0, 15.0 Hz, H-2b), 0.97 (3H, d, J ) 7.0 Hz, H-14); inner
glucose, 5.69 (1H, d, J ) 7.5 Hz, glc H-1), 5.20 (1H, dd, J )
9.5, 8.5 Hz, glc H-3), 5.00 (1H, dd, J ) 9.5, 10.0 Hz, glc H-4),
4.31 (1H, dd, J ) 4.5, 13.0 Hz, glc H-6a), 4.07 (1H, dd, J )
2.0, 13.0 Hz, glc H-6b), 3.99 (1H, ddd, J ) 2.0, 4.5, 10.0 Hz,
glc H-5), 3.85 (1H, dd, J ) 7.5, 8.5 Hz, glc H-2); middle glucose,
5.06 (1H, dd, J ) 9.0, 9.5 Hz, glc H-3), 4.84 (1H, dd, J ) 9.0,
10.0 Hz, glc H-4), 4.60 (1H, d, J ) 7.5 Hz, glc H-1), 4.26 (1H,

1510 J ournal of Natural Products, 2000, Vol. 63, No. 11
Zhang et al.
dd, J ) 5.5, 12.5 Hz, glc H-6a), 4.03 (2H, m, H-12a, glc H-6b),
3.62 (1H, ddd, J ) 3.0, 5.5, 10.0 Hz, glc H-5), 3.56 (1H, dd,
J ) 7.5, 9.5 Hz, glc H-2); arabinose, 5.17 (1H, dd, J ) 3.5, 4.0
Hz, ara H-4), 5.15 (1H, dd, J ) 7.5, 10.0 Hz, ara H-2), 4.91
(1H, dd, J ) 3.5, 10.0 Hz, ara H-3), 4.46 (1H, d, J ) 7.5 Hz,
ara H-1), 4.04 (1H, dd, J ) 4.0, 12.5 Hz, ara H-5a), 3.53 (1H,
dd, J ) 1.5, 12.5 Hz, ara H-5b).
Refer en ces a n d Notes
(1) Xia, Q.; Xiao, P.-G.; Wang, L.-W.; Kong, J . Zhongguo Zhongyao Zazhi
1997, 22, 515-518.
(2) Zhang, Y.-J .; Tanaka, T.; Iwamoto, Y.; Yang, C. R.; Kouno, I.
Tetrahedron Lett. 2000, 41, 1781-1784.
(3) Seikel, M.; Hillis, W. E. Phytochemistry 1970, 9, 1115-1128.
(4) Nishizawa, M.; Yamagishi, Y.; Nonaka, G.; Nishioka, I. J . Chem. Soc.,
Perkin Trans. 1 1983, 961-965.
Ep iga lloca tech in -(2âf7,4âf8)-ga lloca tech in (p r od el-
(5) Nishizawa, M.; Yamagishi, Y.; Nonaka, G.; Nishioka, I. J . Chem. Soc.,
Perkin Trans. 1 1982, 2963-2968.
p h in id in A-1) (6): tan amorphous powder; [R]16 +25.3°
D
(c 0.4, MeOH); 1H NMR (CD3COCD3, 300 MHz) δ 6.76 (2H, s,
B ring H-2, 6), 6.61 (2H, s, B′ ring H-2, 6), 6.15 (1H, s, H-6′),
5.98, 6.07 (each 1H, d, J ) 2.4 Hz, H-6, 8), 4.57 (1H, d, J ) 9
Hz, H-2′), 4.20, 4.13 (each 1H, d, J ) 3.3 Hz, H-3, 4), 4.13 (1H,
m, H-3′), 3.05 (1H, dd, J ) 16.5, 6 Hz, H-4′), 2.57 (1H, dd, J )
16.5, 9.3 Hz, H-4′); negative FABMS m/z 607 [M-H]-; anal.
C 54.17%, H 4.74%, calcd for C30H24O14‚3H2O, C 54.38%, H,
4.56%.
(6) Nonaka, G.; Muta, M.; Nishioka, I. Phytochemistry 1983, 22, 237-
241.
(7) Nonaka, G.; Kawahara, O.; Nishioka, I. Chem. Pharm. Bull. 1983,
31, 3906-3914.
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1981, 781-783.
Ep ica tech in -(4âf8)-ga lloca tech in (7): tan amorphous
powder; [R]16D +17.0° (c 0.4, MeOH); 1H NMR (CD3COCD3, 300
MHz) δ 6.99 (1H, br s, B ring H-2), 6.76 (1H, d, J ) 8.1 Hz, B
ring H-5), 6.71 (1H, dd, J ) 8.1, 2.1 Hz, B ring H-6), 6.52 (2H,
s, B′ ring H-2, 6), 6.03, 5.96 (each 1H, d, J ) 2.4 Hz, H-6, 8),
5.97 (1H, s, H-6′), 5.06 (1H, br s, H-2), 4.64 (2H, br s, H-2′, 3′),
4.02 (1H, m, H-3′), 3.94 (1H, br s, H-4), 2.84 (1H, dd, J ) 16.2,
5.4 Hz, H-4′), 2.58 (1H, dd, J ) 16.5, 7.8 Hz, H-4′); negative
FABMS m/z 593 [M-H]-; anal. C 54.69%, H 5.04%, calcd for
(11) Tanaka, T.; Takahashi, R.; Kouno, I.; Nonaka, G. J . Chem. Soc.,
Perkin Trans. 1 1994, 3013-3022.
(12) Kupchan, S. M.; LaVoie, E. J .; Branfman, A. R.; Fei, B. Y.; Bright,
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C
30H26O13‚3.5H2O, C 54.80%, H, 5.06%.
Th iolysis of 7. A mixture of 7 (1 mg), mercaptoethanol (0.2
(16) Bohlmann, F.; Zdero, C. Phytochemistry 1978, 17, 759-761.
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mL), concentrated HCl (0.01 mL), and EtOH (0.5 mL) was
heated at 50 °C for 1 h and analyzed by reversed-phase HPLC
[Cosmosil 5C18-AR (2.5 mm i.d. × 250 mm); gradient elution
of 10%f20% (30 min) CH3CN in 50 mM H3PO4]. The peaks
corresponding to gallocatechin (7.84 min) and epicatechin 4-(2-
hydroxyethyl)thio ether (28.4 min) were detected and con-
firmed by co-HPLC with authentic samples.
(18) Fukazawa, H.; Li, P.-M.; Mizuno, S.; Uehara, Y. Anal. Biochem. 1993,
212, 106-110.
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Ack n ow led gm en t. The authors are grateful to Mr. K.
Inada and Mr. N. Yamaguchi (Nagasaki University) for NMR
and MS measurements, and Prof. T. Yamori (J apanese Foun-
dation for Cancer Research) for carrying out the bioassays.
One of the authors (Y.J .Z.) is grateful to the J apanese
Government for the award of a scholarship.
NP000135I