Isolation of ellagitannin monomer and macrocyclic dimer from castanopsis carlesii leaves

Article abstract of DOI:10.3987/COM-12-S(N)29

In a phytochemical and chemotaxonomical investigation of Castanopsis species (Fagaceae), new monomeric and dimeric ellagitannins, named carlesiins A (1) and B (2), were isolated from fresh leaves of Castanopsis carlesii along with 55 known compounds. Carl

Full text of DOI:10.3987/COM-12-S(N)29

HETEROCYCLES, Vol. 86, No. 1, 2012
381
HETEROCYCLES, Vol. 86, No. 1, 2012, pp. 381 - 389. © 2012 The Japan Institute of Heterocyclic Chemistry
Received, 9th June, 2012, Accepted, 20th July, 2012, Published online, 24th July, 2012
DOI: 10.3987/COM-12-S(N)29
ISOLATION OF ELLAGITANNIN MONOMER AND MACROCYCLIC
DIMER FROM CASTANOPSIS CARLESII LEAVES
a,b
*,a
a
a
Yong-Lin Huang,
Takashi Tanaka,
Yosuke Matsuo, Isao Kouno,
b
c
Dian-Peng Li, and Gen-ichiro Nonaka
a
Graduate School of Biomedical Sciences, Nagasaki University, 1-14
Bunkyo-Machi, Nagasaki 852-8521, Japan; t-tanaka@nagasaki-u.ac.jp
b
Guangxi Key Laboratory of Functional Phytochemicals Research and Utilization,
Guangxi Institute of Botany, Guilin 541006, China
c
Usaien Pharmaceutical Company, Ltd., 1-4-6 Zaimoku, Saga 840-0055, Japan
Abstract – In a phytochemical and chemotaxonomical investigation of
Castanopsis species (Fagaceae), new monomeric and dimeric ellagitannins,
named carlesiins A (1) and B (2), were isolated from fresh leaves of Castanopsis
carlesii along with 55 known compounds. Carlesiin A was identified as
1
-O-galloyl-4,6-(S)-tergalloyl-β-D-glucose. Carlesiin B is a macrocyclic
ellagitannin dimer with a symmetrical structure composed of two tergalloyl and
two glucopyranose moieties. Their structures were elucidated based on
spectroscopic and chemical evidence.
INTRODUCTION
The species in the Castanopsis (Fagaceae) genus are evergreen trees that are found in East Asia,
sometimes as the dominant species in a forest. These trees are often used as forestry or ornamental trees,
and the wood is an important construction material. There are about 120 species in the genus, but the
chemical compositions of only a few species have been studied. The leaves of Chinese and Japanese
Castanopsis sp. are rich sources of polyphenols, and each species has a unique phenolic composition. We
conducted a phytochemical and chemotaxonomical investigation of the Castanopsis sp. to determine how
1
,2
these plant resources could be effectively utilized. Our investigations of C. cuspidata var. sieboldii, C.
3
,4
5
6,7
hystrix, C. fissa, and C. sclerophylla, have revealed the presence of characteristic metabolites,
This paper is dedicated to Prof. Ei-ichi Negishi on the occasion of his 77th birthday.

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HETEROCYCLES, Vol. 86, No. 1, 2012
including triterpene hexahydroxydiphenoyl (HHDP) esters, phenylpropanoid-substituted flavan-3-ols,
ellagitannins, and galloyl esters of quinic acid. In the present study, we examined the leaves of C. carlesii
collected in Southern China and isolated two new ellagitannins and 55 known compounds. The
differences in the phenolic composition compared to other species are also discussed.
RESULTS AND DISCUSSION
The fresh leaves were extracted with 80% aqueous acetone, and the extract was subjected to a
combination of column chromatography using Sephadex LH-20, Diaion HP20SS, MCI gel CHP 20P,
Toyopearl Butyl 650C, Chromatorex ODS, Toyopearl HW F40, and Avicel cellulose to afford 57
compounds, including the two new ellagitannins 1 and 2 (Figure 1). The known compounds were
identified by comparison of their spectroscopic data to data acquired from authentic samples and
literature data (see Experimental).
tergalloyl group
OH
OH
HO
HO
OH
OH
OH
O
C
HO
O
HO
HO
O
OH
OH
O
O
C
O
O
OH
OH
OH
HO
O
HO
OH
OH
C
O
O
C
O
OH
HOOC
O
COOH
O
O
HO
HO
HO
O
O
HO
O
OH
O
HO
HO
OH
C
O C
C
O
O
O
HO
O
C
OH
O
O
HO
OH
OH
O
O
HO
C
O
O
OH
OH
HO
OH
OH
HHDP unit
1
1a
2
Figure 1. Structures of 1, 2, and 1a
Carlesiin A (1) was obtained as a brown amorphous powder, which gave a dark blue color with ethanolic
FeCl
3
reagent. The molecular formula C34
H
26
O23 was determined by high-resolution fast atom
+
bombardment mass spectrometry (HR FAB-MS), which showed the [M+H] ion peak at m/z 803.0962
1
(
Calcd for C34
H
27
O
23, 803.0943). The proton nuclear magnetic resonance ( H-NMR) spectrum of 1 (Table
1
) showed an anomeric proton signal at  5.72 (d, J = 8.2 Hz, H-1). The large coupling constants of the
remaining sugar proton signals (J2,3, J3,4, and J4,5 = 9–10 Hz) indicated that this sugar was a
4

-glucopyranose in the C
1
conformation. Downfield shifts of the glucose H-1, H-4 and H-6 indicated
esterification of these positions. As for the acyl groups, the presence of a galloyl group was apparent from
1
13
the H [ 7.17 (2H, s)] and C NMR signals. Comparison among the chemical shifts of the remaining
three aromatic protons [ 6.55 (H-3'), 6.73 (H-3), 6.92 (H-6")] and 18 aromatic and three carboxyl carbons,

HETEROCYCLES, Vol. 86, No. 1, 2012
383
indicated the presence of a HHDP ester group
with an additional gallic acid moiety. Among the
aromatic carbon signals, the signal at  132.0 is
8
,9
characteristic of the C-2' of tergalloyl group.
This was confirmed by acid hydrolysis of 1,
which gave gallic acid and tergallic acid dilactone
(
1a). These compounds were also isolated from
the same extract in the present study. The
heteronuclear multiple bond connectivity
Figure 2. Selected HMBC correlations (H to C) of 1
(
HMBC) correlations between the glucose H-1
and galloyl carboxyl carbon indicated that the galloyl group was attached to an anomeric hydroxyl group
1
(
Figure 2). The large difference in the H NMR chemical shifts (Δδ 1.36) of the glucose C-6 methylene
protons suggests that the HHDP unit of the tergalloyl moiety forms macrocyclic esters with O-4 and O-6
of the glucose. This was confirmed by the HMBC correlations of H-4 ( 4.92) and H-6 ( 3.79 and 5.15)
with the tergalloyl C-7 ( 168.2) and C-7' ( 168.1) carboxyl carbons, respectively (Figure 2). The
orientation of the tergalloyl esters, that is, the location of the additional gallic acid moiety, was
determined by observation of the HMBC correlations of H-3' ( 6.55) with C-4' ( 149.4), C-5' ( 136.3)
and C-7'. Furthermore, atropisomerism of the tergalloyl biphenyl bond was revealed to be S, because the
5
circular dichroism (CD) spectrum exhibited a strong positive Cotton effect at 236 nm ([θ]236 +8.2×10 )
5
10
and a negative effect at 261 nm ([θ]261 -3.3×10 ). The glucose core was deduced to be in the D
configuration because coexisting known ellagitannins isolated from the same plant source have a
D-glucopyranose core. Based on these spectroscopic and chemical results, the structure of compound 1 was
confirmed to be 1-O-galloyl-4,6-O-(S)-tergalloyl--D-glucose and was named carlesiin A.
1
The H NMR spectrum of carlesiin B (2) was closely related to that of 1, showing three aromatic singlets
at  6.46, 6.70, and 7.00, and a set of signals attributable to 1,4,6-O-acylated β-glucopyranose (Table 1).
1
3
The presence of gallic acid trimer and glucopyranose residues was also apparent from the C NMR
signals. However, signals for the galloyl ester were not observed in the spectra of 2, and the matrix
+
assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) [M+Na] peak
was at m/z 1287. The results indicate that 2 is a dimeric ellagitannin with a symmetrical structure
composed of two glucopyranose and two triphenoyl groups. The C-2' of the triphenoyl group resonated at
8
,9

127.9, which is different from that of tergalloyl group ( 132.0). The chemical shift was related to
7
9
those of valoneoyl (C-2': about  126) and macaranoyl groups (C-2': about  124), which are positional
isomers of the tergalloyl group (Figure 4).

384
HETEROCYCLES, Vol. 86, No. 1, 2012
1
13
a
Table 1. H-(500 MHz) and C-(125 MHz) NMR data for 1 and 2 (in acetone-d +D O)
6
2
1
2
1
13
1
13
Position
H
C
Position
H
C
Anomer
Anomer
1
2
3
4
5
6
5.72 (d, 8.2)
3.71 (dd, 8.2, 9.8)
3.84 (t, 9.8)
4.92 (t, 9.8)
4.11 (dd, 5.5, 9.8)
95.8
74.4
75.2
72.6
72.9
64.0
1
2
3
4
5
6
5.54 (d, 8.0)
3.49 (dd, 8.0, 9.8)
3.52 (t, 9.8)
4.84 (t, 9.8)
4.07 (dd, 6.3, 9.8)
96.5
74.1
75.2
73.1
72.6
63.5
5.15 (dd, 6.2, 13.3)
.79 (d, 13.3)
5.21 (dd, 6.3, 13.4)
3.76 (d, 13.4)
3
Tergalloyl
Tergalloyl
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
3
4
5
6
7
116.3
125.5
108.1
145.1
136.7
144.2
168.2
115.6
132.0
108.0
149.4
136.3
149.4
168.1
114.7
139.7
139.2
140.2
142.2
108.3
170.9
1
2
3
4
5
6
7
1'
2'
3'
4'
5'
6'
7'
1"
2"
3"
4"
5"
6"
7"
115.4
125.1
107.6
145.3
136.7
145.4
171.1
115.4
127.9
109.2
145.3
136.2
144.6
168.0
113.8
142.7
139.6
139.6
140.2
108.6
166.1
6.73 (s)
6.70 (s)
'
'
'
'
'
'
'
"
"
"
"
"
"
"
6.55 (s)
6.46 (s)
6.92 (s)
7.00 (s)
Galloyl
1
120.4
110.1
146.0
139.4
165.5
2
3
4
,6
,5
7.17 (2H, s)
7
a
Chemical shifts are given in δ values; multiplicities and coupling constants (J in Hz) in parentheses.
The acyl group was chemically confirmed by hydrolysis of 2, which gave tergalloyl acid dilactone (1a).
Furthermore, methylation and subsequent methanolysis of 2 afforded a trimethyl ester 2a, which was
2
D
3
identified as (S)-trimethyl octamethyltergallate ([α]
-20.5) (Figure 4). The S-biphenyl bond was also

HETEROCYCLES, Vol. 86, No. 1, 2012
385
confirmed by observation of a positive Cotton
6
effect at 235 nm ([θ]235 +1.5×10 ) and
negative Cotton effect at 261 nm ([θ]261
5
10
-
3.0×10 ) in the CD spectrum of 2. Similar
glucose proton signals for 2 and 1 suggested
that the HHDP unit of the triphenoyl group
was located at the O-4 and O-6 positions. This
was confirmed by the HMBC correlations of
ester carboxyl carbons C-7 and C-7' with
glucose H-4 and H-6, respectively (Figure 3).
The HMBC spectrum also revealed the
Figure 3. Selected HMBC correlations (H to C) of 2.
remaining gallic acid unit of the triphenoyl group was attached to the glucose anomeric position. Thus,
the structure of carlesiin B was determined (2 in Figure 1). To the best of our knowledge, this compound is
the first macrocyclic ellagitannin dimer with tergalloyl esters. The upfield shift of the tergalloyl C-2' was
probably caused by steric strain, and the upfield sifts of the glucose protons compared to those of 1 could
be explained by anisotropic effect of the aromatic rings.
OH
OH
HO
OH
O
OH
OH
OH
HOOC
HO
MeO
MeO
MeO MeO OMe
MeO
OMe
OH
OH
O
C
O
COOH
HO
HO
OH HO
O
C
O
O
O
O
C
O
HO
O
HO
O
COOMe
OMe
C
O
HO
HO
OH
O
HOOC
C
O
HO
O
C
C O
C
O
OH
HO
HO
HO
MeOOC COOMe
OH
OH
OH
HO HO OH OH
2
a
3
valoneoyl group
macaranoyl group
Figure 4. Structures of 2a, platycaryanin B (3), and valoneoyl and macaranoyl groups.
CONCLUSION
1
1
Although dimeric and oligomeric ellagitannins have been isolated from many plant families, only two
Castanopsis plants have been shown to contain the ellagitannin dimers. One is C. sclerophylla, which
contains rugosin E with a valoneoyl group (Figure 4) as one of the major polyphenolic constituents
7
(
0.50% from fresh leaves). The other is C. carlesii, which was examined in this study, although the yield
of the dimeric ellagitannin 2 is very low (isolation yield 0.0007% from fresh leaves). C. carlesii contains
monomeric ellagitannins as the major constituents, pedunculagin [2,3;4,6-bis-(S)-HHDP-D-glucose;

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HETEROCYCLES, Vol. 86, No. 1, 2012
0.14%], 1(β)-O-galloyl pedunculagin (0.42% from fresh leaves), and platycaryanin
B
[2,3-(S)-HHDP-4,6-(S)-tergalloyl-D-glucose (3); 0.18%]. These were accompanied by their degradation
products, ellagic acid (0.49%) and tergallic acid dilactone (1a) (0.08%). Unlike C. sclerophylla, C.
carlesii contains ellagitannins with a tergalloyl group, such as platycaryanin B (3). This is an important
characteristic feature of this plant from the viewpoint of chemotaxonomy of the genus Castanopsis.
Carlesiins A (1) and B (2) are biogenetically related to platycaryanin B, and it should be noted that the
symmetrical macrocyclic structure with tergalloyl ester groups of 2 is the first example in the gallic acid
metabolites. Triterpene HHDP esters isolated from C. cuspidata var. sieboldii, C. hystrix, and C. fissa,
and phenylpropanoid-substituted flavan-3-ols isolated from C. hystrix, and C. sclerophylla were not
detected in the leaves of C. carlesii. Further phytochemical and chemotaxonomical studies on
Castanopsis species are now underway.
EXPERIMENTAL
IR spectra were obtained with a JASCO FT/IR-410 spectrophotometer, UV spectra were obtained using a
JASCO V-560 UV/Vis spectrophotometer, and optical rotations were measured with a JASCO DIP-370
1
13
6
digital polarimeter (Jasco Co., Tokyo, Japan). H- and C-NMR spectra were measured in acteone-d or
1
13
3
CD OD at 27 °C using a Varian Unity plus 500 spectrometer (500 MHz for H and 125 MHz for C)
1
13
(
(
(
Varian, Palo Alto, CA) or a JEOL JNM-AL 400 spectrometer (400 MHz for H and 100 MHz for C)
JEOL Ltd., Tokyo, Japan). Coupling constants are expressed in Hz and chemical shifts are given in δ
ppm). MS were recorded on a Voyager DE-PRO (Applied Biosystems, Foster City, CA) and a JEOL
JMS-700N spectrometer (JEOL Ltd.). 2,5-Dihydroxybenzoic acid and glycerol were used as the matrices
for MALDI-TOF-MS and FAB-MS measurements, respectively. Column chromatography was performed
using a Diaion HP20SS (Mitsubishi Chemical, Tokyo, Japan), MCI gel CHP 20P (75–150 μm; Mitsubishi
Chemical), Sephadex LH-20 (25–100 μm; GE Healthcare Bio-Science AB, Uppsala, Sweden), Toyopearl
Butyl 650C (Tosoh Corp., Tokyo, Japan), Toyopearl HW F40 (Toshoh Corp.), Avicel Cellulose
(
Funakoshi Co., Tokyo, Japan), and Chromatorex ODS (100–200 mesh; Fuji Silysia Chemical, Tokyo,
Japan) columns. Thin-layer chromatography was performed on precoated Kieselgel 60 F254 plates (0.2
mm thick; Merck, Darmstadt, Germany) and Cellulose plates (0.1 mm thickness; Merck), using solvent
systems of toluene-ethyl formate-formic acid (1:7:1, v/v/v) and 2% acetic acid, respectively. Spots were
3
detected by UV illumination (254 nm) and by spraying with a 2% ethanolic FeCl and 10% sulfuric acid
reagent, followed by heating.
Plant material
The leaves of C. carlesii were collected at Guangxi Institute of Botany, Guangxi, China, in August 2009,

HETEROCYCLES, Vol. 86, No. 1, 2012
387
and identified by Prof. Wei Fa-nan. The voucher specimen (CC20090826) was deposited in the Herbarium
of Guangxi Institute of Botany, China.
Extraction and separation
2
The fresh leaves of C. carlesii (7.60 kg) were cut into small pieces and extracted with acetone-H O (8:2,
v/v) at room temperature. After filtration, the plant debris remaining on the filter paper was extracted with
the same solvent twice. The filtrate was concentrated under reduced pressure to give a dark green
precipitate consisting mainly of chlorophylls and waxes, which were removed by filtration. The filtrate
2
was then concentrated to give an extract (610 g). The extract was partitioned between Et O and water. The
aqueous layer was subjected to Sephadex LH-20 column chromatography (10 cm i. d. × 40 cm) with water
containing increasing proportions of methanol (MeOH) (0–100%, 10% stepwise elution) and finally 60%
acetone to give the following eight fractions: 1 (153.3 g), 2 (35.5 g), 3 (33.9 g), 4 (82.0 g), 5 (122.5 g), 6
(17.9 g), 7 (18.9 g), and 8 (42.8 g). Fraction 5 gave a precipitate in aqueous MeOH, which was collected
by filtration and identified as ellagic acid (37.2 g). The soluble part of the fraction was separated by a
Diaion HP20SS column (6 cm i. d. × 40 cm) with 0–100% MeOH (10% stepwise elution) to give 9
subfractions. Subfraction 5-4 (15.7 g) was further separated by column chromatography using Sephadex
LH-20 (0–100% MeOH, 10% stepwise elution), Toyopearl Butyl 650C (0–100% MeOH, 10% stepwise
elution), Avicel Cellulose (2% AcOH), and MCI-gel CHP-20P (10–100% MeOH, 10% stepwise elution)
to yield 1 (159 mg), 2 (56 mg), (–)-epicatechin (122 mg), pedunculagin (10.5 g), platycaryanin B (13.5 g),
1
-O-galloyl-4,6-O-(S)-valoneoyl-β-D-glucose (38 mg), flavogallonic acid (24 mg), valoneic acid dilactone
(
3.31 g), tergallic acid dilactone (6.13 g), and isoquercitrin (34 mg).
The known compounds isolated from other fractions by similar chromatographic separation were as
follows: 5-O-(β-apiosyl-(1→2)-O-β-xylopyranosyl)gentisic acid (192 mg), gentisic acid
5
-O-β-D-glucoside (46 mg), and benzyl-α-L-rhamnopyranosyl(1→6)-β-D-glucoside (172 mg) from
fraction 1; 1-O-galloyl-β-D-glucose (77 mg), 4,6-(S)-HHDP-D-glucose (494 mg), gentisic acid
-O-β-D-xylopyranoside (160 mg), 1,6-di-O-galloyl-β-D-glucose (484 mg), brevifolin carboxylic acid
131 mg), and 6,7-dihydroxycoumarin (10 mg) from fraction 2; 4,6-O-(S)-valoneoyl-D-glucose (600 mg),
gallic acid (725 mg), 6-O-galloyl-glucose (26 mg), (2R,3R)-taxifolin (191 mg), gentisic acid
-O-β-D-(6'-O-galloyl)glucopyranoside (9 mg), 4-hydroxy-3-methoxyphenol l-O-β-D-(6'-O-galloyl)-
glucoside (161 mg), 4-hydroxy-2-methoxyphenol 1-O-β-D-(6'-O-galloyl)glucoside (133 mg),
,4-dihydroxyphenethyl alcohol 4-O-β-D-(6"-O-galloyl)glucopyranoside (13 mg), 4-methoxycatechol (2
mg), 4-O-(6'-O-galloyl-β-glucopyranosyl)-cis-p-coumaric acid (9 mg), and 2R,3R-dihydromyricetin
-O-β-L-arabinopyranoside (25 mg) from fraction 3; sanguiin H-5 (29 mg), casuariin (1.78 g), platycariin
5
(
5
3
3

388
HETEROCYCLES, Vol. 86, No. 1, 2012
(
210 mg), tamarixetin 3-O-α-rhamnopyranoside (525 mg), myricitrin (568 mg), mearnsitrin (3.43 g),
gemin D (2.57 g), 1-O-galloyl-4,6-(S)-HHDP-β-D-glucose (880 mg), (+)-catechin (608 mg), and
(
(
(
+)-gallocatechin (8 mg) from fraction 4; flosin A (1.11 g), 1,2-di-galloyl-4,6-O-(S)-HHDP-β-D-glucose
20 mg), alnusnin B (156 mg), (–)-epigallocatechin 3-O-gallate (597 mg), (–)-epicatechin 3-O-gallate
314 mg), eschweilenol A (12 mg), and myricetin 3-O-(3''-O-galloyl)-α-rhamnopyranoside (69 mg) from
fraction 6; 1,2,3,6-tetra-O-galloyl-β-D-glucose (48 mg), and nobotanin D (53 mg) from fraction 7; and
(β)-O-galloyl-pedunculagin (32.1 g), platycaryanin A (311 mg), 1,2,3,4,6-penta-O-galloyl-β-D-glucose
12 mg), 1-desgalloyl eugeniin (17 mg), eugeniin (466 mg), cuspinin (305 mg), and rugosin C (16 mg)
1
(
from fraction 8.
2
D
4
Carlesiin A (1). Brown amorphous powder, [α]
–123.3 (c 0.14, MeOH); HR FAB-MS m/z: 803.0962
+
-1
[
M+H] (Calcd for C34
H
27
O
23: 803.0943); IR νmax cm : 3348, 1711, 1563, 1228; UV λmax (MeOH) nm (log
5
5

): 261 (4.53), 212 (5.07); CD (MeOH) [θ] (nm): +8.2×10 (236), 0 (251), –3.3×10 (261), 0 (275),
4
+ 1
+
6 2
8.6×10 (283), 0 (292); MALDI-TOF-MS m/z: 825 [M+Na] ; H-NMR (500 MHz, acetone-d +D O) and
1
3
6 2
C-NMR (125 MHz, acetone-d +D O). See Table 2.
2
D
4
Carlesiin B (2). Brown amorphous powder, [α]
+12.0 (c 0.13, MeOH); MALDI-TOF-MS m/z: 1287
+
+
-1
[
M+Na] ; HR FAB-MS m/z: 1265.1334 [M+H] (Calcd for C54
H
41
O
36: 1265.1378); IR νmax cm : 3350,
6
1702, 1560, 1214; UV λmax (MeOH) nm (log ): 263 (4.58), 212 (5.10); CD (MeOH) [θ] (nm): +1.5×10
5
5
1
(
6 2
235), 0 (253), –3.0×10 (261), 0 (271), +2.2×10 (279), 0 (292); H-NMR (500 MHz, acetone-d +D O)
1
3
6 2
and C-NMR (125 MHz, acetone-d +D O). See Table 2.
Acid hydrolysis of 1 and 2
2 4
A solution of 1 (1.6 mg) in 0.5 mol/L H SO (0.2 mL) was kept at 90–100 °C for 2 h. An equal volume of
MeOH was added to dissolve the resulting precipitates, and the products were analyzed by HPLC under
the following conditions: column, Cosmosil 5C AR II (250×4.6 mm i.d., Nacalai Tesque Inc., Kyoto,
1
8
3 4
Japan); solvent, MeCN in 50 mmol/L H PO ; solvent gradient, 4–30% MeCN (39 min) and 30–75% MeCN
(
15 min); flow rate, 0.8 mL/min; and detector, Jasco photodiode array detector MD-910. The peaks at 8.69
min and 24.57 min corresponded to the derivatives prepared from gallic acid and tergallic acid dilactone,
respectively. Hydrolysis of 2 (1.0 mg) was also achieved in a similar manner, and the HPLC showed a
peak for the tergallic acid dilactone.
Production of 2a from 2
2 2
A solution of 2 (5.2 mg) in MeOH (0.5 mL) was treated with an ether solution of CH N at 0 °C for 2 h.

HETEROCYCLES, Vol. 86, No. 1, 2012
389
After concentration under reduced pressure, the residue was dissolved in 1% NaOH in 50% MeOH (1.0
mL) and heated at 80 °C for 1 h. The reaction mixture was acidified with 0.5 mol/L HCl (1.5 mL) and
partitioned between ethanol and water. The organic layer was dried with anhydrous Na
treated with an ether solution of CH at 0 °C for 2 h. The reaction mixture was concentrated, and the
product was purified by silica gel chromatography with toluene-acetone (100:1–92:8, 1% stepwise
2 4
SO , and then
2
N
2
2
D
3
elution) to yield (S)-trimethyl octa-O-methyl tergallate (2a) (2.5 mg) as a colorless syrup, []
–20.5 (c
) : 3.35, 3.59, 3.60, 3.61
×2), 3.74, 3.79, 3.86 (×2), 3.89, 3.92 (each 3H, s, OMe), 7.16, 7.32, 7.37 (each 1H, s).
+
1
0
3 3
.13, CHCl ); MALDI-TOF-MS m/z: 683 [M+Na] ; H-NMR (400 MHz, CDCl
(
ACKNOWLEDGEMENTS
The authors are grateful to Mr. K. Inada, Mr. N. Yamaguchi and Mr. N. Tsuda (Nagasaki University Joint
Research Center) for NMR and MS measurements.
REFERENCES AND NOTES
1
2
3
4
5
. M. Ageta, K. Ishimaru, G. Nonaka, and I. Nishioka, Chem. Pharm. Bull., 1988, 36, 870.
. M. Ageta, G. Nonaka, and I. Nishioka, Chem. Pharm. Bull., 1988, 36, 1646.
. H. F. Chen, T. Tanaka, G. Nonaka, T. Fujioka, and K. Mihashi, Phytochemistry, 1993, 32, 1457.
. H. F. Chen, T. Tanaka, G. Nonaka, T. Fujioka, and K. Mihashi, Phytochemistry, 1993, 33, 183.
. Y. L. Huang, T. Tsujita, T. Tanaka, Y. Matsuo, I. Kouno, D. P. Li, and G. Nonaka, Phytochemistry,
2
011, 72, 2006.
. Y. L. Huang, Y. Matsuo, T. Tanaka, I. Kouno, D. P. Li, and G. Nonaka, Heterocycles, 2011, 83,
321.
. Y. L. Huang, T. Tanaka, Y. Matsuo, I. Kouno, D. P. Li, and G. Nonaka, Phytochemistry Letters,
012, 5, 158.
6
7
2
2
8
9
1
. T. Tanaka, S. Kirihara, G. Nonaka, and I. Nishioka, Chem. Pharm. Bull., 1993, 41, 1708.
. J. H. Lin, M. Ishimatsu, T. Tanaka, G. Nonaka, and I. Nishioka, Chem. Pharm. Bull., 1990, 38, 1844.
0. T. Okuda, T. Yoshida, T. Hatano, T. Koga, N. Toh, and K. Kuriyama, Tetrahedron Lett., 1982, 23,
3
937.
1
1. T. Okuda, T. Yoshida, and T. Hatano, Prog. Chem. Org. Nat. Prod., 1995, 66, 1.