Synthesis and biological evaluation of a natural ester sintenin and its synthetic analogues

Article abstract of DOI:10.1021/np0496441

Synthesis of 3-(3,4-dimethoxyphenyl)propyl-3-(3,4-dimethoxyphenyl) propanoate (18), a cytotoxic natural ester, was carried out by a convenient synthetic path with a total yield of 49%. Sixteen of its analogues (19-34) were also prepared. Seventeen unsaturated derivatives of 18, compounds 1-17, were also synthesized to examine the structure-activity relationship of this type of ester. All of the synthetic compounds were passed through the cytotoxicity screenings on human tumor cell lines, such as PC-3, Hela, A549, BEL7404, CNE, and KB. Some of the esters exhibited moderate inhibitory effects on these tumor cell lines. The phenolic derivatives exhibited the highest cytotoxicity among these derivatives, while the unsaturated esters were more cytotoxic than the saturated analogues. Some of the compounds also exhibited inhibition on α-glucosidase.

Full text of DOI:10.1021/np0496441

342
J. Nat. Prod. 2005, 68, 342-348
Synthesis and Biological Evaluation of a Natural Ester Sintenin and Its
Synthetic Analogues
Li Hong Hu,† Hong Bin Zou,† Jing Xu Gong,† Hai Bo Li,† Lei Xiang Yang,† Wei Cheng,† Chang Xin Zhou,†
Hua Bai,‡ Franc¸oise Gue´ritte,§ and Yu Zhao*,†
Department of Traditional Chinese Medicine and Natural Drug Research, College of Pharmaceutical Sciences,
Zhejiang University, Hangzhou 310031, People’s Republic of China, Zhejiang Hisun Naturelite Pharmaceutical R&D Co., Ltd.
19-G, Huazhe Plaza, Hangzhou 310006, People’s Republic of China, and Institut de Chimie des Substances Naturelles,
CNRS, 91198 Gif-sur-Yvette, France
Received November 9, 2004
Synthesis of 3-(3,4-dimethoxyphenyl)propyl-3-(3,4-dimethoxyphenyl) propanoate (18), a cytotoxic natural
ester, was carried out by a convenient synthetic path with a total yield of 49%. Sixteen of its analogues
(19-34) were also prepared. Seventeen unsaturated derivatives of 18, compounds 1-17, were also
synthesized to examine the structure-activity relationship of this type of ester. All of the synthetic
compounds were passed through the cytotoxicity screenings on human tumor cell lines, such as PC-3,
Hela, A549, BEL7404, CNE, and KB. Some of the esters exhibited moderate inhibitory effects on these
tumor cell lines. The phenolic derivatives exhibited the highest cytotoxicity among these derivatives,
while the unsaturated esters were more cytotoxic than the saturated analogues. Some of the compounds
also exhibited inhibition on R-glucosidase.
Table 1. Synthetic Saturated Esters 18-34
Cytotoxic compounds are crucial in the course of finding
new leads for antitumor drugs. Chen and co-workers
recently reported the isolation of a cytotoxic natural ester,
3-(3,4-dimethoxyphenyl)propyl-3-(3,4-dimethoxyphenyl) pro-
panoate (18), from the leaves and stems of Piper sintenense
scattered in Taiwan. This ester, named sintenin by Chen
et al., possessed selective cytotoxicity with an ED50 value
of 5.4 × 10-7 M against P-388 cells.1 However, the cyto-
toxicities on other common human tumor cell lines, e.g.,
KB, Hela, PC-3, CNE, A549, and BEL7404, were not
reported therein. To our knowledge, these kinds of esters
have wide biological activity. Lee et al. reported the
cytotoxicity of caffeic acid phenethyl ester analogues on oral
cancer cells, while Nagaoka et al. also indicated that the
caffeic acid phenethyl ester analogues have selective anti-
proliferative activity on the colon 26-L5 carcinoma cell
line.2,3 To further investigate the cytotoxic nature of
compound 18 and to preliminarily explore the structure-
activity relationship (SAR) of these types of esters, we have
designed a convenient synthetic route to prepare this
natural ester (18) and its derivatives (19-34) (Table 1)
along with their performance on KB, Hela, PC-3, CNE,
A549, and BEL7404 cell lines. Furthermore, the structure
of 18 is partially similar to nelumols B-D, the sinapyl
alcohol derivatives isolated from Ligularia nelumbifolia,
which were reported to be cytotoxic to A549, HL-60, and
KB cell lines.4,5 This further stimulated our interest to
synthesize the sinapyl sinapate analogues (1-17) (Table
2) and compare their influence to the cytotoxicity on the
six above-mentioned human tumor cell lines.
substituted groupa
compound
R1
R2
R3
R4
R5
R6
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
OCH3
H
OCH3
OCH3
H
OCH3
OCH3
OCH3
NH2
OMOM
OH
CH3
OMOM
H
H
H
OCH3
OCH3
H
OCH3
OCH3
H
OCH3
OCH3
OCH3
OCH3
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
OCH3
H
OCH3 OCH3
H
H
H
OCH3
H
OCH3
H
OCH3
H
H
H
H
H
H
H
H
H
H
CH3
H
H
H
OMOM
OMOM
H
OMOM OMOM
OMOM OMOM
H
H
H
H
OH
H
H
H
OH
H
H
H
OMOM
OH
H
H
OCH3
OCH3
H
H
OMOM
H
OMOM OMOM
a OMOM (OCH2OCH3).
OCH2OCH3 to prolong the substituents, and to remove the
OCH2OCH3 group to expose the phenolic OH group.
Further modifications were carried out by changing the
methoxy and hydrogen by an amine and nitro group to
make the aromatic ring more electron-rich or electron-poor.
All of the synthetic derivatives (1-34) were subjected to a
cytotoxic screening on PC-3, Hela, A549, BEL7404, CNE,
and KB cell lines, which correspond to the tumor diseases
found in China. Furthermore, the inhibitory effects of
selected compounds on R-glucosidase were also performed
to check the relationship between cytotoxicity and R-glu-
cosidase inhibition. The syntheses as well as the biological
evaluations of these compounds are reported herein.
Moreover, to detect the structure-activity relationship
of sintenin (18) derivatives, we selected different numbers
and patterns of substituents on the aromatic A and/or B
rings in the molecule, e.g., to replace the OCH3 group by
* To whom correspondence should be addressed. Tel: (86)-571-87217313.
Fax: (86)-571-85270026. E-mail: dryuzhao@zju.edu.cn or dryuzhao@
hotmail.com.
† Zhejiang University.
‡ Zhejiang Hisun Naturelite Pharmaceutical R&D Co., Ltd.
§ ICSN, CNRS.
10.1021/np0496441 CCC: $30.25
© 2005 American Chemical Society and American Society of Pharmacognosy
Published on Web 02/26/2005

Natural Ester Sintenin
Journal of Natural Products, 2005, Vol. 68, No. 3 343
Table 2. Synthetic Unsaturated Esters 1-17
Further hydrogenation of the unsaturated esters was
performed by palladium-charcoal catalysis and afforded a
series of saturated ester derivatives 19-24. The structures
of all the synthetic compounds were elucidated mainly on
1
the basis of their H and 13C NMR spectra as well as MS
spectral data.
Scheme 2 outlines the synthetic path of the compounds
9-17 as well as compounds 25-34. These compounds are
mainly substituted by OCH2OCH3 and free phenolic OH
groups, and therefore utilized a different synthetic sched-
ule. The unsaturated esters 9 and 15 as well as the
saturated esters 30 and 31 were selected to be examples
in the scheme. The protocol was initiated with 3,4-dihy-
droxybenzaldehyde, which was reacted with chloromethyl
methyl ether in the presence of potassium carbonate under
reflux to afford the disubstituted aldehyde 37 in 70%
yield.12 This aldehyde was subjected to a Wittig reaction
with ethyl(triphenylphosphoranylidene)acetate in benzene
to afford a substituted sinapic acid ethyl ester 38.13 The
ester was hydrolyzed by potassium hydroxide to afford the
substituted sinapic acid 39, which was then condensed with
cinnamic alcohol under the catalysis of dicyclohexylcarbo-
diimide (DCC) and 4-dimethylaminopyridine (DMAP) to
afford the unsaturated ester 9,14 which could be hydroge-
nated under palladium-charcoal (10% Pd-C) catalysis to
afford the saturated ester 30. Furthermore, treatment of
9 and 30 with 10% hydrochloric acid gave the correspond-
ing products 15 and 31, respectively, which contained free
phenolic hydroxyls in the aromatic ring.11,15
substituted groupa
compound
R1
R2
R3
R4
R5
R6
1
2
OCH3
H
OCH3
OCH3
H
H
H
H
OCH3
OCH3
H
OCH3
OCH3
H
OCH3
OCH3
OCH3
OCH3
OCH3
H
H
H
H
H
H
H
H
3
H
4
OCH3
H
OCH3
H
OCH3
OCH3
OCH3
OCH3
NO2
OCH3 OCH3
5
H
OCH3
H
H
H
OCH3
6
7
8
OCH3
OCH3 OCH3
OCH3
H
9
OMOM OMOM
H
H
H
H
H
H
H
H
H
H
H
H
H
H
10
11
12
13
14
15
16
17
H
OMOM
OMOM
OMOM
H
H
OMOM OMOM
H
H
OMOM
OH
OMOM
OH
OMOM
OH
H
H
H
H
H
OMOM OMOM
H
OH
H
H
H
H
H
H
OCH3
OCH3
H
H
H
a OMOM (OCH2OCH3).
Scheme 1: Synthetic Route of Compounds 1 and 18a
Cytotoxic Evaluation. All of the synthetic compounds
1-34 were subjected to in vitro cytotoxicity screenings on
six human tumor cell lines with the marketed agent
cisplatin (DDP) employed as a positive control. The tested
IC50 values for these active compounds are listed in Table
3. Meanwhile, those with IC50 values greater than 100 µg/
mL on all six tumor cell lines were considered inactive
compounds and thus are not included in Table 3. It could
be observed that compound 18 showed no significant
cytotoxicity to the six selected tumor cell lines. This result
is consistent with a former study, in which the natural
ester was found to be cytotoxic only to P-388 cells among
the three cell lines they tested.1 Furthermore, its analogues
possessing saturated carbon chains between two aromatic
rings, 19-24, also exhibited poor cytotoxicity except for
compound 21, which showed moderate cytotoxicity with
IC50 values of 1.0 × 10-4 and 1.3 × 10-4 M against PC-3
and KB cell lines, respectively. However, three analogues
with free phenolic hydroxyls in the aromatic ring, viz.,
compounds 26, 31, and 33, exhibited stronger and wider
cytotoxicity to the six tumor cell lines (Table 3).
Meanwhile, it could be found that the unsaturated esters
1-7 exhibited more visible cytotoxicity than those of their
saturated analogues 19-24. For example, unsaturated
ester 4 showed cytotoxicity to PC-3, Hela, A549, and
BEL7404 cell lines, while its corresponding saturated
precursor 21 exhibited weaker cytotoxicity limited to PC-3
and KB cells (Table 3). This suggested that the double
bonds conjugated to the two aromatic rings might be
desirable to promote the cytotoxicity of this type of esters.
a (i) 1: malonic acid, Py, piperidine, reflux, 1.5 h; 2: HCl, rt, 1 h; (ii)
LAH, THF, rt, 4 h; (iii) CDI, DBU, THF, 45 °C, 1 day; (iv) H2, Pd/C, EtOAc,
rt, 12 h.
Results and Discussion
Synthesis. The synthetic paths are outlined in Scheme
1 and Scheme 2. Scheme 1 describes the procedure of
preparing compounds 1 and 18. The synthesis started from
3,4-dimethoxybenzaldehyde, which was treated with ma-
lonic acid in pyridine catalyzed by piperidine (Knoevenagel
condensation) to furnish an allylic acid, 35.6,7 The allylic
alcohol 36 was achieved by reduction of 35 with LiAlH4 in
anhydrous THF at room temperature.8-10 The allylic acid
35 was treated with carbonyldiimidazole (CDI) to form a
CDI-adduct that contained an activated carbonyl group,
and thus facilitated the condensation of the adduct with
the allylic alcohol 36 in the presence of 1,8-diazabicyclo-
[5.4.0]undec-7-ene (DBU) to afford 1 with a yield of 76%.
This unsaturated ester 1 was further hydrogenated to give
compound 18 in an 84% yield.11
Moreover, almost all of the compounds containing OCH2-
OCH3 groups in the molecules did not exhibit cytotoxicity
except for compound 9, which exhibited an IC50 on KB of
2.0 × 10-4 M, and for compound 16, which showed
cytotoxicity to PC-3 and Hela cell lines with IC50 values of
2.3 × 10-4 and 4.6 × 10-5 M, respectively. However, after
the OCH2OCH3 groups were removed and the phenolic
hydroxyls were revealed, the cytotoxicity was elevated
Similar preparations were carried out as modeled in
Scheme 1, and different aromatic substituents were intro-
duced by changing the benzaldehyde starting materials.
A series of unsaturated esters 2-8 were thus obtained.

344 Journal of Natural Products, 2005, Vol. 68, No. 3
Scheme 2: Synthesis of Esters 9, 15, 30, and 31a
Hu et al.
a (i) MOMCl, K2CO3, acetone, reflux, 4 h, 70%; (ii) Ph3PdCHCO2Et, benzene, reflux, 6 h, 89%; (iii) KOH/H2O, EtOH, rt, 4 h, 80%; (iv) cinnamic alcohol,
DCC, DMAP, CH2Cl2, rt, 24 h, 63%; (v) H2, Pd/C, AcOEt, rt, 12 h, 82%; (vi) 10% HCl, MeOH, reflux, 0.5 h, 34%.
Table 3. Cytotoxicities of Selected Compounds on Six Human
hydroxyls in the A ring, exhibited cytotoxicity to KB, BEL
Tumor Cell Lines (IC50 at 10-5 M)a
7404, and A549 cell lines with IC50 values of 5.4 × 10-5
,
9.1 × 10-5, and 7.1 × 10-5 M, respectively.
compound PC-3
Hela
A549 CNE BEL7404
KB
b
According to the cytotoxic evaluation results, it was
suggested that the analogues having one or more free
phenol groups are more toxic to human tumor cells than
the esters with alkyloxyl or alkyl substituents, and the
esters with nitro or amine groups in the aromatic rings. It
is clear that the number of methoxy substituents in the B
ring is related to the measure of cytotoxicity among the
batch of esters. However, the prolongation of the OCH3
group by OCH2OCH3 groups did not intensify the cytotox-
icity. In general, unsaturated esters showed more convinc-
ing cytotoxicity than their saturated analogues, which
might be due to the presence of a large π-π conjugative
system in the unsaturated molecules. These results could
serve as fundamental information for further SAR inves-
tigation on this type of natural-based esters.
1
9.2
18.1
8.0
8.67
16.7
6.4
18.1
/
/
/
/
13.6
13.3
b
b
b
b
b
b
2
/
/
/
b
4
17.2
21.2
/
b
b
b
7
21.2
18.9
/
/
/
16.5
20.6
20.2
b
8
4.3
18.0
5.0
/
b
b
b
b
b
b
9
/
/
/
/
/
/
b
13
14.3
12.3
22.6
8.8
16.0
17.1
4.6
12.0
7.1
16.2
21.4
/
15
9.1
5.4
b
b
b
b
16
/
/
/
/
17
9.5
7.2
4.0
19.3
14.3
12.4
12.9
12.5
82.0
b
b
b
b
21
10.4
32.2
18.9
28.3
0.3
/
/
/
/
26
31.7
17.1
31.7
13.8
18.6
0.3
26.8
23.5
28.7
0.2
21.7
b
31
/
b
33
/
10.8
0.7
cisplatin
0.1
0.04
a Key to cell lines: BEL7404, human hepatocellular carcinoma
line; CNE, nasopharyngeal carcinoma cell line; KB, human oral
epithelial cell line; A549, human lung adenocarcinoma cell line;
Hela, human cervical carcinoma cell line; PC-3, human prostate
cancer cell line. b IC50 values greater than 100 µg/mL were
considered inactive.
r-Glucosidase Inhibition. R-Glucosidase is a nutri-
tionally and biochemically important enzyme for dietary
carbohydrate digestion and post-translational processing
of glycoprotein.16 Inhibition of R-glucosidases causes ab-
normal functionality of glycoproteins because of incomplete
modification of glycans. Suppression of this processing is
to be expected for antiviral activity and decreasing of
growth rate of tumors.16 Some glucosidase inhibitors show
antitumor metastasis17 and anti-HIV activities18 and are
also clinically useful for treatment of diabetes.19
To the best of our knowledge, some of the cytotoxic
compounds also possess inhibition effects on R-glucosi-
dase.20 Therefore selected synthetic compounds were sub-
jected to an R-glucosidase inhibition assay. It was found
that compounds 8, 13, 15, and 31 exhibited IC50 values to
R-glucosidase at 1.1 × 10-4, 6.5 × 10-5, 9.7 × 10-5, and
7.3 × 10-5 M, respectively. Meanwhile the positive control,
acarbose, demonstrated a IC50 value to R-glucosidase of 6.8
× 10-4 M. This indicated some relationship between
cytotoxicity and R-glucosidase inhibition among this kind
of esters.
remarkably. This could be reflected from couples of com-
pounds, e.g., 13 vs 12, 15 vs 9, 17 vs 16, 31 vs 30, etc.
(Table 3).
By scrutiny of the initial results, it could also be observed
that the unsaturated esters with notable cytotoxicity are
those with two methoxy groups existing on the B ring (i.e.,
compounds 1, 2, 4, and 7). Nevertheless, compound 8,
which possessed a trimethoxy-substituted B ring and a
trimethoxy moiety in the A ring, exhibited a wide and
significant cytotoxicity to nearly all of the six tumor cell
lines.
Among these synthetic samples, ester 17, with free
phenols in the A ring and a methoxy group in the B ring,
showed the strongest cytotoxicity, especially on the cell
lines of CNE and PC-3 with IC50 values of 4.0 × 10-5 and
8.8 × 10-5 M, respectively. It should also be noticed that
the unsaturated ester 15, which contained two free phenolic

Natural Ester Sintenin
Journal of Natural Products, 2005, Vol. 68, No. 3 345
a
Table 4. 1H NMR Spectral Data [400 MHz, δH (J, Hz)] for Compounds 1-34 in CDCl3
compound
H-2
H-3
H-1′
H-2′
H-3′
Ar-H
1
2
6.36 d (16)
6.36 d (16)
6.50 d (16)
6.40 d (16)
6.35 d (16)
6.36 d (16)
6.58 d (16)
6.40 d (16)
6.37 d (16)
6.37 d (16)
6.37 d (16)
6.37 d (16)
6.36 d (16)
6.37 d (16)
6.42 d (16)
6.36 d (16)
6.37 d (16)
2.63 t (8.4)
2.62 t (8.4)
2.67 t (8.4)
2.66 t (8.0)
2.60 t (8.0)
2.63 t (8.0)
2.63 t (8.0)
2.65 t (8.0)
2.65 t (8.0)
2.70 t (8.0)
2.64 t (8.0)
2.63 t (8.0)
2.66 t (8.0)
2.64 t (8.0)
2.61 t (8.0)
2.64 t (8.0)
2.68 t (8.0)
7.68 d (16)
7.69 d (16)
7.75 d (16)
7.65 d (16)
7.69 d (16)
7.64 d (16)
7.75 d (16)
7.66 d (16)
7.66 d (16)
7.68 d (16)
7.65 d (16)
7.70 d (16)
7.69 d (16)
7.69 d (16)
7.52 d (16)
7.68 d (16)
7.67 d (16)
2.91 t (8.4)
2.90 t (8.4)
2.97 t (8.4)
2.90 t (8.0)
2.90 t (8.0)
2.90 t (8.0)
2.85 t (8.0)
2.91 t (8.0)
2.89 t (8.0)
2.93 t (8.0)
2.91 t (8.0)
2.89 t (8.0)
2.89 t (8.0)
2.83 t (8.0)
2.91 t (8.0)
2.88 t (8.0)
2.92 t (8.0)
4.86 d (6.4)
4.85 d (6.8)
4.89 d (6.4)
4.86 d (6.4)
4.84 d (6.4)
4.85 d (6.8)
4.88 d (6.4)
4.87 d (6.4)
4.87 d (6.4)
4.84 d (6.4)
4.84 d (6.4)
4.87 d (6.4)
4.88 d (6.4)
4.84 d (6.8)
4.80 d (6.4)
4.84 d (6.4)
4.87 d (6.4)
4.10 t (6.4)
4.09 t (6.4)
4.10 t (6.4)
4.10 t (6.4)
4.07 t (6.4)
4.09 t (6.4)
4.10 t (6.4)
4.11 t (6.4)
4.10 t (6.4)
4.09 t (6.4)
4.12 t (6.4)
4.08 t (6.4)
4.12 t (6.4)
4.09 t (6.4)
4.08 t (6.4)
4.07 t (6.4)
4.09 t (6.4)
6.23 dt (16, 6.4)
6.24 dt (16, 6.8)
6.38 dt (16, 6.4)
6.25 dt (16, 6.4)
6.26 dt (16, 6.4)
6.24 dt (16, 6.8)
6.24 dt (16, 6.4)
6.30 dt (16, 6.4)
6.36 dt (16, 6.4)
6.25 dt (16, 6.4)
6.25 dt (16, 6.4)
6.36 dt (16, 6.4)
6.35 dt (16, 6.4)
6.25 dt (16, 6.8)
6.30 dt (16, 6.4)
6.24 dt (16, 6.4)
6.36 dt (16, 6.4)
1.92 m
6.66 d (16)
6.65 d (16)
6.72 d (16)
6.66 d (16)
6.66 d (16)
6.66 d (16)
6.82 d (16)
6.64 d (16)
6.72 d (16)
6.66 d (16)
6.66 d (16)
6.72 d (16)
6.67 d (16)
6.63 d (16)
6.72 d (16)
6.66 d (16)
6.71 d (16)
2.61 t (8.4)
2.59 t (8.4)
2.65 t (8.4)
2.61 t (8.4)
2.58 t (8.0)
2.60 t (8.0)
2.58 t (8.0)
2.61 t (8.0)
2.61 t (8.0)
2.63 t (8.0)
2.53 t (8.0)
2.61 t (8.4)
2.61 t (8.4)
2.60 t (8.0)
2.59 t (8.0)
2.58 t (8.0)
2.57 t (8.4)
6.84-7.13 m
6.80-7.49 m
7.27-7.56 m
6.76-6.97 m
6.83-7.50 m
6.76-7.37 m
6.95-7.18 m
6.77 s, 6.64 s
6.85-7.62 m
7.00-7.48 m
6.99-7.38 m
7.04-7.51 m
6.83-7.45 m
7.00-7.50 m
6.77-7.48 m
6.83-7.49 m
6.94-7.43 m
6.69-6.80 m
6.68-7.14 m
7.16-7.32 m
6.43-6.80 m
6.82-7.14 m
6.42-7.73 m
6.45-7.02 m
6.96-7.31 m
6.75-7.31 m
7.05-7.12 m
6.95-7.15 m
6.79-7.27 m
6.80-7.31 m
6.61-7.31 m
6.83-7.15 m
6.74-7.26 m
6.73-7.14 m
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
1.91 m
1.95 m
1.91 m
1.91 m
1.91 m
1.91 m
1.94 m
1.92 m
1.92 m
1.90 m
1.91 m
1.91 m
1.92 m
1.90 m
1.92 m
1.91 m
a Only common characteristics of the synthetic compounds are listed. The individual 1H NMR spectral data of the different substituents
for compounds 1-34 were listed in the Experimental Section.
Experimental Section
DBU (0.16 g, 1.05 mmol) in dry THF (10 mL) was then added
quickly. The resulting mixture was stirred at 45 °C for 24 h.
The mixture was evaporated to afford a residue, which was
subjected to flash chromatography to provide 1 (0.28 g, 76%
yield) as a white crystalline solid: mp 78 °C (hexane); Rf
General Experimental Procedures. Melting points were
recorded on a Kofler hot-stage instrument and were uncor-
rected. ESIMS data were recorded on a Bruker Esquire 3000+
spectrometer. HRFABMS spectra were recorded on a VG ZAB-
HS spectrometer. The NMR spectra were obtained on a Bruker
AM-400 FT-NMR spectrometer with TMS as internal stan-
dard. Preparative TLC was performed using silica gel GF254
and RP-18 plates (Merck). Enzyme-linked immunosorbent
assays were recorded on a BIO-TEK ELX8000 ELISA reader.
R-Glucosidase was from Baker’s yeast (Sigma, 500 U/mL).
4-Nitrophenyl-R-D-glucopyranoside was from E. Merck. Acar-
bose was purchased from The First Hospital affiliated with
Zhejiang University.
3-(3,4-Dimethoxyphenyl)propenyl-3-(3,4-dimethoxyphen-
yl)allylate (1). To a stirred solution of malonic acid (3 g, 18
mmol) in Py (5 mL) at room temperature was added 3,4-
dimethoxybenzaldehyde (2.8 g, 27 mmol) in Py (7 mL).
Piperidine (2 mL) was added to the solution. The mixture was
heated at 118 °C for 4 h and was cooled to 0 °C. Then 3 N HCl
was added to adjust the pH value to 3, and the mixture was
stirred at 0 °C for another 1 h. After suction filtration, the
filtrate was crystallized in acetone to afford 35 as a white
crystalline solid. To a stirred suspension of LAH (56 mg, 1.5
mmol) in dry THF (8 mL) at room temperature was added a
solution of 35 (208 mg, 1 mmol) in THF (10 mL) under argon
atmosphere. The solution was stirred for 4 h and was quenched
by 8 mL of H2O. Then 15 mL of 1 N HCl was added, and the
mixture was extracted by ether. The ether layers were
combined and dried (MgSO4). Evaporation of the solvent
followed by CC (hexane/AcOEt, 6:1) afforded a yellow solid,
36 (109 mg, 56% yield). To a stirred solution of 35 (0.20 g,
0.96 mmol) in dry THF (10 mL) was added CDI (0.23 g, 1.44
mmol) in dry THF under argon. The mixture was stirred at
45 °C for 15 min. The solution of 36 (0.21 g, 1.05 mmol) and
1
(hexane/AcOEt, 3:1) 0.21; H NMR (400 MHz, CDCl3) δ 3.92
(6 H, s, OMe-6′ and OMe-6), 3.91 (6 H, s, OMe-7′ and OMe-7),
other data see Table 4; 13C NMR (100 MHz, CDCl3), see Table
5; ESIMS m/z 385 ([M + 1]+); HRFABMS m/z 384.1569 (calcd
for C22H24O6, 384.1573).
3-(3,4-Dimethoxyphenyl)propenyl-3-(3-methoxyphenyl)-
allylate (2). The method of preparation of 2 was similar to
that used for the preparation of 1: white crystalline solid; mp
66-67 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.22; 1H NMR (400
MHz, CDCl3) δ 3.91 (3 H, s, OMe-7), 3.89 (3 H, s, OMe-7′),
3.84 (3 H, s, OMe-6′), other data see Table 4; 13C NMR (100
MHz, CDCl3), see Table 5; ESIMS m/z 355 ([M + 1]+);
HRFABMS m/z 354.1468 (calcd for C21H22O5, 354.1467).
3-Phenylpropenyl-3-phenylallylate (3). The method of
preparation of 3 was similar to that used for the preparation
of 1: white crystalline solid; mp 60-61 °C (hexane); Rf (hexane/
AcOEt, 3:1) 0.63; 1H NMR (400 MHz, CDCl3), see Table 4; 13
C
NMR (100 MHz, CDCl3), see Table 5; ESIMS m/z 265 ([M +
1]+); HRFABMS m/z 264.1153 (calcd for C18H16O2, 264.1150).
3-(3,4-Dimethoxyphenyl)propenyl-3-(2,3,4-trimethoxy-
phenyl)allylate (4). The method of preparation of 4 was
similar to that used for the preparation of 1: white crystalline
solid; mp 77-78 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.27; 1H
NMR (400 MHz, CDCl3) δ 3.88 (6 H, s, OMe-6′ and OMe-6),
3.86 (6 H, s, OMe-7′ and OMe-7), 3.85 (3 H, s, OMe-8), other
data see Table 4; 13C NMR (100 MHz, CDCl3), see Table 5;
ESIMS m/z 437 ([M + Na]+); HRFABMS m/z 414.1674 (calcd
for C23H26O7, 414.1679).
3-(3-Methoxyphenyl)propenyl-3-(3-methoxyphenyl)al-
lylate (5). The method of preparation of 5 was similar to that
used for the preparation of 1: white crystalline solid; mp 63-

346 Journal of Natural Products, 2005, Vol. 68, No. 3
Hu et al.
Table 5. 13C NMR Spectral Data [100 MHz, δ (ppm)] for Selected Compounds in CDCl3
C no.
1
2
3
4
10
11
12
18
19
25
27a
28
29
1
167.0
120.1
144.9
134.2
109.2
151.0
149.0
115.4
122.7
65.2
167.0
115.5
133.8
127.0
127.0
114.2
161.3
114.2
127.0
64.9
121.3
129.2
129.7
108.7
148.9
144.6
111.1
120.1
166.7
117.8
145.0
134.3
126.6
130.3
128.8
130.3
126.6
65.1
128.0
128.5
134.3
126.6
130.3
128.8
130.3
126.6
166.9
110.8
134.9
121.0
127.0
147.1
148.7
153.3
117.1
63.9
167.0
121.4
144.2
133.7
129.6
116.2
158.9
116.2
129.6
65.2
127.8
132.5
133.7
129.6
116.2
157.1
116.2
129.6
166.8
121.5
144.5
130.1
115.6
149.1
157.1
116.1
128.8
65.2
167.0
115.7
144.6
123.3
126.6
116.4
158.9
116.4
126.6
65.0
126.6
128.0
134.1
123.3
129.7
128.0
139.7
123.3
172.9
36.1
173.0
36.1
173.0
36.1
173.0
36.0
173.0
36.1
172.9
36.0
2
3
30.5
30.3
30.1
30.3
30.3
30.4
4
133.0
111.5
148.7
147.4
111.1
120.0
63.7
132.5
113.8
148.7
147.1
111.4
120.1
63.7
133.9
129.3
116.2
155.6
116.2
129.3
63.8
135.5
129.0
128.2
155.3
128.2
129.0
63.8
133.9
129.2
116.2
155.4
116.2
129.2
63.7
134.5
116.6
147.1
145.5
116.6
129.3
63.7
5
6
7
8
9
1′
2′
3′
4′
5′
6′
7′
8′
9′
127.2
129.1
134.2
108.5
151.0
149.0
110.8
121.1
55.7
111.4
129.8
129.7
104.9
139.8
144.7
107.5
111.0
55.7
123.4
123.4
133.8
127.8
116.3
147.3
116.3
127.8
56.2
30.3
31.4
30.3
31.6
30.1
32.1
30.2
31.4
30.3
31.2
30.2
31.2
133.6
111.5
148.7
147.4
111.1
120.1
55.7
133.7
113.8
157.9
147.1
111.0
129.2
141.1
128.4
128.4
126.0
128.4
128.4
137.9
129.0
128.2
155.3
128.2
129.0
134.5
129.2
116.2
155.6
116.2
129.2
134.9
129.3
116.2
155.4
116.2
129.3
56.1
OMe-6
OMe-7
55.8
55.3
55.8
56.0
56.0
56.1
55.8
55.2
55.9
55.9
55.9
OMe-8
56.0
OMe-6′
55.7
55.8
55.7
55.8
55.7
55.7
55.8
55.7
55.8
OMe-7′
55.8
56.0
94.3
94.3
56.3
95.1
94.3
55.9
94.9
94.9
55.9
95.3
94.4
OCH2O-6
OCH2O-7
OCH2O-6′
OCH2O-7′
94.0
94.4
95.5
94.4
a Methyl signals: 21.0.
64 °C (hexane); Rf (hexane/AcOEt, 6:1) 0.70; 1H NMR (400
MHz, CDCl3) δ 3.84 (6 H, s, OMe-7′ and OMe-7), other data
see Table 4; ESIMS m/z 324 ([M]+); HRFABMS m/z 324.1358
(calcd for C20H20O4, 324.1362).
3-(3-Methoxyphenyl)propenyl-3-(2,3,4-trimethoxyphen-
yl)allylate (6). The method of preparation of 6 was similar
to that used for the preparation of 1: white crystalline solid;
mp 62-63 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.21; 1H NMR
(400 MHz, CDCl3) δ 3.90 (3 H, s, OMe-6), 3.88 (6 H, s, OMe-7′
and OMe-7), 3.78 (3 H, s, OMe-8), other data see Table 4;
ESIMS m/z 385 ([M + 1]+); HRFABMS m/z 384.1579 (calcd
for C22H24O6, 384.1573).
evaporated. The residue was purified by column chromatog-
raphy eluted with 10% AcOEt/hexane to afford white solid 9
(120 mg, 63% yield): mp 32-33 °C (hexane); Rf (hexane/AcOEt,
1
3:1) 0.53; H NMR (400 MHz, CDCl3) δ 5.27 (2 H, s, OCH2O-
7), 5.26 (2 H, s, OCH2O-6), 3.53 (3 H, s, OMe-7), 3.52 (3 H, s,
OMe-6), other data see Table 4; ESIMS m/z [M + Na]+ 407;
HRFABMS m/z 384.1566 (calcd for C22H24O6, 384.1573).
3-(3-Methoxymethoxyphenyl)propenyl-3-(3-meth-
oxymethoxyphenyl)allylate (10). The method of prepara-
tion of 10 was similar to that used for the preparation of 9:
white crystalline solid; mp 30 °C (hexane); Rf (hexane/AcOEt,
3:1) 0.60; 1H NMR (400 MHz, CDCl3) δ 5.21 (4 H, s, OCH2O-7′
and OCH2O-7), 3.48 (6 H, s, OMe-7′ and OMe-7), other data
see Table 4; 13C NMR (100 MHz, CDCl3), see Table 5; ESIMS
m/z 384 ([M]+); HRFABMS m/z 384.1563 (calcd for C22H24O6,
384.1573).
3-(3,4-Dimethoxyphenyl)propenyl-3-(3-nitrophenyl)al-
lylate (7). The method of preparation of 7 was similar to that
used for the preparation of 1: yellow crystalline solid; mp 89-
90 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.26; 1H NMR (400
MHz, CDCl3) δ 3.91 (3 H, s, OMe-6′), 3.89 (3 H, s, OMe-7′),
other data see Table 4; ESIMS m/z 392 ([M + Na]+); HR-
FABMS m/z 369.1211 (calcd for C20H19NO6, 369.1212).
3-(2,3,4-Trimethoxyphenyl)propenyl-3-(2,3,4-trimethox-
yphenyl)allylate (8). The method of preparation of 8 was
similar to that used for the preparation of 1: white crystalline
solid; mp 60-62 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.27; 1H
NMR (400 MHz, CDCl3) δ 3.89 (3 H, s, OMe-6′), 3.88 (3 H, s,
OMe-6), 3.86 (6 H, s, OMe-7′ and OMe-7), 3.83 (6 H, s, OMe-8
and OMe-8′), other data see Table 4; ESIMS m/z 467 ([M +
Na]+); HRFABMS m/z 444.1769 (calcd for C24H28O8, 444.1784).
3-Phenylpropenyl-3-(3,4-dimethoxymethoxyphenyl)al-
lylate (9). To a stirred solution of 3812,13 (98 mg, 0.33 mmol)
in 5 mL of ethanol was added a solution of potassium
hydroxide (65 mg) in 4 mL of H2O. The mixture was heated to
reflux for 4 h. Evaporation of ethanol was followed by adding
2 mL of water. Then 5 mL of 1 N HCl was added to get a white
suspension. The mixture was then extracted by ether, and the
ether layers were washed with brine and dried (MgSO4). The
solution was evaporated to afford 39 as a white solid (70 mg,
80% yield). To a stirred solution of 39 (0.13 g, 0.50 mmol) in
dry CH2Cl2 (8 mL) was added DCC (0.11 g, 0.55 mmol) in dry
CH2Cl2 (5 mL) under argon. The mixture was stirred at room
temperature for 5 min until a white suspension occurred. A
solution of cinnamic alcohol (0.07 g, 0.55 mmol) and DMAP
(0.01 g, 0.10 mmol) in dry CH2Cl2 was then added to the
mixture. The suspension was stirred at room temperature for
24 h. The mixture was filtered through a Celite layer and
3-(3-Methoxymethoxyphenyl)propenyl-3-(3,4-dimeth-
oxymethoxyphenyl)allylate (11). The method of prepara-
tion of 11 was similar to that used for the preparation of 9:
white crystalline solid; mp 32-33 °C (hexane); Rf (hexane/
1
AcOEt, 3:1) 0.32; H NMR (400 MHz, CDCl3) δ 5.27 (2 H, s,
OCH2O-7), 5.26 (2 H, s, OCH2O-6), 5.18 (2 H, s, OCH2O-7′),
3.53 (3 H, s, OMe-7), 3.52 (3 H, s, OMe-6), 3.48 (3 H, s, OMe-
7′), other data see Table 4; 13C NMR (100 MHz, CDCl3), see
Table 5; ESIMS m/z 467 ([M + Na]+); HRFABMS m/z 444.1781
(calcd for C24H28O8, 444.1784).
3-Phenylpropenyl-3-(3-methoxymethoxyphenyl)allyl-
ate (12). The method of preparation of 12 was similar to that
used for the preparation of 9: white crystalline solid; mp 52-
53 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.54; 1H NMR (400
MHz, CDCl3) δ 5.22 (2 H, s, OCH2O-7), 3.49 (3 H, s, OMe-7),
other data see Table 4; 13C NMR (100 MHz, CDCl3), see Table
5; ESIMS m/z 347([M + Na]+); HRFABMS m/z 324.1369 (calcd
for C20H20O4, 324.1362).
3-Phenylpropenyl-3-(3-hydroxyphenyl)allylate (13). The
method of preparation of 13 was similar to that used for the
preparation of 9: colorless oil; Rf (hexane/AcOEt, 3:1) 0.19;
1H NMR (400 MHz, CDCl3) δ 5.87 (1 H, s, OH-7), other data
see Table 4; ESIMS m/z 303 ([M + Na]+); HRFABMS m/z
280.1087 (calcd for C18H16 O3, 280.1099).
3-(3,4-Dimethoxymethoxyphenyl)propenyl-3-(3-meth-
oxymethoxyphenyl) allylate (14). The method of prepara-
tion of 14 was similar to that used for the preparation of 9:
white crystalline solid; mp 65-66 °C (hexane); Rf (hexane/

Natural Ester Sintenin
Journal of Natural Products, 2005, Vol. 68, No. 3 347
1
1
AcOEt, 3:1) 0.38; H NMR (400 MHz, CDCl3) δ 5.26 (2 H, s,
AcOEt, 3:1) 0.23; H NMR (400 MHz, CDCl3) δ 3.84 (6 H, s,
OCH2O-7), 5.24(2 H, s, OCH2O-7′), 5.21 (2 H, s, OCH2O-6′),
3.53 (3 H, s, OMe-7), 3.52 (3 H, s, OMe-7′), 3.48 (3 H, s, OMe-
6′), other data see Table 4; ESIMS m/z 445 ([M + 1]+);
HRFABMS m/z 444.1776 (calcd for C24H28O8, 444.1784).
3-Phenylpropenyl-3-(3,4-dihydroxyphenyl)allylate (15).
To a solution of 14 (0.04 g, 0.10 mmol) in CH3OH (5.0 mL)
was added 10% HCI (1.0 mL). The resulting mixture was
refluxed for 30 min and then poured into cold water and
extracted with AcOEt. The organic phase was washed with a
saturated solution of NaHCO3 and brine and then dried (Na2-
SO4). After removal of the solvent, the residue was chroma-
tographyed over silica gel. Elution with petroleum ether/AcOEt
(8:1) yielded 15 (10 mg, 34% yield) as a colorless oil: Rf
OMe-7 and OMe-7′), 3.81 (3 H, s, OMe-6), 3.78 (3 H, s, OMe-
8), other data see Table 4; ESIMS m/z 406 ([M + H2O]+);
HRFABMS m/z 388.1882 (calcd for C22H28O6, 388.1886).
3-(3,4-Dimethoxyphenyl)propyl-3-(3-aminophenyl)pro-
panoate (24). The method of preparation of 24 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
1
AcOEt, 3:1) 0.13; H NMR (400 MHz, CDCl3) δ 5.30 (2 H, s,
NH2-7), 3.87 (3 H, s, OMe-6′), 3.86 (3 H, s, OMe-7′), other data
see Table 4; ESIMS m/z 344 ([M + 1]+); HRFABMS m/z
343.1782 (calcd for C20H25NO4, 343.1784).
3-Phenylpropyl-3-(3-methoxymethoxyphenyl)pro-
panoate (25). The method of preparation of 25 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
1
1
(hexane/AcOEt, 3:1) 0.11; H NMR (400 MHz, CDCl3) δ 9.43
AcOEt, 3:1) 0.55; H NMR (400 MHz, CDCl3) δ 5.15 (2 H, s,
(1 H, s, OH-7), 8.99 (1 H, s, OH-6), other data see Table 4;
ESIMS m/z 319 ([M + Na]+); HRFABMS m/z 296.1058 (calcd
for C18H16 O4, 296.1049).
OCH2O-7), 3.47 (3 H, s, OMe-7), other data see Table 4; 13C
NMR (100 MHz, CDCl3), see Table 5; ESIMS m/z 329 ([M +
1]+); HRFABMS m/z 328.1671 (calcd for C20H24O4, 328.1675).
3-Phenylpropyl-3-(3-hydroxyphenyl)propanoate (26).
The method of preparation of 26 was similar to that used for
the preparation of 18: colorless oil; Rf (hexane/AcOEt, 3:1)
3-(3-Methoxyphenyl)propenyl-3-(3-methoxymethoxy-
phenyl)allylate (16). The method of preparation of 16 was
similar to that used for the preparation of 9: white crystalline
solid; mp 49-50 °C (hexane); Rf (hexane/AcOEt, 3:1) 0.20; 1H
NMR (400 MHz, CDCl3) δ 5.21 (2 H, s, OCH2O-7), 3.82 (3 H,
s, OMe-7′), 3.48 (3 H, s, OMe-7), other data see Table 4; ESIMS
m/z 354 ([M]+); HRFABMS m/z 354.1474 (calcd for C21H22O5,
354.1467).
1
0.21; H NMR (400 MHz, CDCl3) δ 5.71 (1 H, s, OH-7), other
data see Table 4; ESIMS m/z 285 ([M + 1]+); HRFABMS m/z
284.1420 (calcd for C18H20O3, 284.1412).
3-(3-Methylphenyl)propyl-3-(3-methylphenyl)pro-
panoate (27). The method of preparation of 27 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
3-(3-Methoxyphenyl)propenyl-3-(3-hydroxyphenyl)al-
lylate (17). The method of preparation of 17 was similar to
that used for the preparation of 9: colorless oil; Rf (hexane/
1
AcOEt, 3:1) 0.26; H NMR (400 MHz, CDCl3) δ 2.34 (3 H, s,
Me-7′), 2.33 (3 H, s, Me-7), other data see Table 4; 13C NMR
(100 MHz, CDCl3), see Table 5; ESIMS m/z 314 ([M + H2O]+);
HRFABMS m/z 296.1768 (calcd for C20H24O2, 296.1776).
3-(3-Methoxymethoxyphenyl)propyl-3-(3-methoxy-
methoxyphenyl)propanoate (28). The method of prepara-
tion of 28 was similar to that used for the preparation of 18:
colorless oil; Rf (hexane/AcOEt, 3:1) 0.62; 1H NMR (400 MHz,
CDCl3) δ 5.16 (2 H, s, OCH2O-7), 5.15 (2 H, s, OCH2O-7′), 3.48
(3 H, s, OMe-7), 3.47 (3 H, s, OMe-7′), 2.91 (2 H, t, J ) 8.0 Hz,
H-3), other data see Table 4; 13C NMR (100 MHz, CDCl3), see
Table 5; ESIMS m/z 406 ([M + H2O]+); HRFABMS m/z
388.1876 (calcd for C22H28O6, 388.1886).
1
AcOEt, 3:1) 0.18; H NMR (400 MHz, CDCl3) δ 5.90 (1 H, s,
OH-7), 3.93 (3 H, s, OMe-7′), other data see Table 4; ESIMS
m/z 310 ([M]+); HRFABMS m/z 310.1212 (calcd for C19H18O4,
310.1205).
3-(3,4-Dimethoxyphenyl)propyl-3-(3,4-dimethoxyphen-
yl)propanoate (18). The mixture of 1 (0.10 g, 0.26 mmol) and
10% Pd/C (0.01 g) was suspended in AcOEt (15 mL) and was
pressurized by H2 gas overnight. The resulted mixture was
filtered through kieselguhr and evaporated. The residue was
purified by column chromatography eluted with 25% AcOEt/
hexane to afford 18 (83 mg, 84%) as a colorless oil: Rf (hexane/
1
AcOEt, 3:1) 0.23; H NMR (400 MHz, CDCl3) δ 3.87 (6 H, s,
3-(3-Methoxymethoxyphenyl)propyl-3-(3,4-dimethoxy-
methoxyphenyl)propanoate (29). The method of prepara-
tion of 29 was similar to that used for the preparation of 18:
colorless oil; Rf (hexane/AcOEt, 3:1) 0.35; 1H NMR (400 MHz,
CDCl3) δ 5.23 (2 H, s, OCH2O-6), 5.20 (2 H, s, OCH2O-7), 5.16
(2 H, s, OCH2O-7′), 3.52 (3 H, s, OMe-6), 3.51 (3 H, s, OMe-7),
3.48 (3 H, s, OMe-7′), other data see Table 4; 13C NMR (100
MHz, CDCl3), see Table 5; ESIMS m/z 466 ([M + H2O]+);
HRFABMS m/z 448.2090 (calcd for C24H32O8, 448.2097).
3-Phenylpropyl-3-(3,4-dimethoxymethoxyphenyl)pro-
panoate (30). The mixture of 14 (0.05 g, 0.13 mmol) and 10%
Pd/C (0.01 g) was suspended in AcOEt (15 mL) and was
pressurized by H2 gas overnight. The resulted mixture was
filtered through kieselguhr and evaporated. The residue was
purified by column chromatography eluted with 25% AcOEt/
hexane to afford 30 (47 mg, 93%) as a colorless oil; Rf (hexane/
OMe-6′ and OMe-6), 3.86 (3 H, s, OMe-7), 3.85 (3 H, s, OMe-
7′), other data see Table 4; 13C NMR (100 MHz, CDCl3), see
Table 5; ESIMS m/z 406 ([M + H2O]+); HRFABMS m/z
388.1880 (calcd for C22H28O6, 388.1886).
3-(3,4-Dimethoxyphenyl)propyl-3-(3-methoxyphenyl)-
propanoate (19). The method of preparation of 19 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
1
AcOEt, 3:1) 0.25; H NMR (400 MHz, CDCl3) δ 3.88 (3 H, s,
OMe-6′), 3.86 (3 H, s, OMe-7), 3.78 (3 H, s, OMe-7′); 13C NMR
(100 MHz, CDCl3), see Table 5; ESIMS m/z 376 ([M + H2O]+);
HRFABMS m/z 358.1783 (calcd for C21H26O5, 358.1780).
3-Phenylpropyl-3-phenylpropanoate (20). The method
of preparation of 20 was similar to that used for the prepara-
tion of 18: colorless oil; Rf (hexane/AcOEt, 3:1) 0.66; 1H NMR
(400 MHz, CDCl3), see Table 4; ESIMS m/z 286 ([M + H2O]+);
HRFABMS m/z 268.1460 (calcd for C18H20O2, 268.1463).
3-(3,4-Dimethoxyphenyl)propyl-3-(2,3,4-trimethoxyphen-
yl)propanoate (21). The method of preparation of 21 was
similar to that used for the preparation of 18: colorless oil; Rf
1
AcOEt, 3:1) 0.54; H NMR (400 MHz, CDCl3) δ 5.23 (2 H, s,
OCH2O-6), 5.20 (2 H, s, OCH2O-7), 3.52 (3 H, s, OMe-6), 3.51
(3 H, s, OMe-7), other data see Table 4; ESIMS m/z 406 ([M +
H2O]+); HRFABMS m/z 388.1869 (calcd for C22H28O6, 388.1886).
3-(3-Hydroxyphenyl)propyl-3-(3-hydroxyphenyl)pro-
panoate (31). The mixture of 15 (0.06 g, 0.22 mmol) and 10%
Pd/C (0.01 g) was suspended in AcOEt (15 mL) and was
pressurized by H2 gas overnight. The resulting mixture was
filtered through kieselguhr and evaporated. The residue was
purified by column chromatography eluted with 25% AcOEt/
hexane to afford 31 (54 mg, 82%) as a colorless oil; Rf (hexane/
1
(hexane/AcOEt, 3:1) 0.28; H NMR (400 MHz, CDCl3) δ 3.87
(3 H, s, OMe-6), 3.86 (3 H, s, OMe-6′), 3.84 (6 H, s, OMe-7′ and
OMe-7), 3.81 (3 H, s, OMe-8), other data see Table 4; ESIMS
m/z 402 ([M + H2O]+); HRFABMS m/z 384.1569 (calcd for
C23H30O7, 384.1573).
3-(3-Methoxyphenyl)propyl-3-(3-methoxyphenyl)pro-
panoate (22). The method of preparation of 22 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
1
AcOEt, 3:1) 0.13; H NMR (400 MHz, CDCl3) δ 5.95 (1 H, s,
1
AcOEt, 6:1) 0.71; H NMR (400 MHz, CDCl3) δ 3.78 (6 H, s,
OH-6), 5.77 (1 H, s, OH-7), other data see Table 4; ESIMS m/z
318 ([M + H2O]+); HRFABMS m/z 300.1353 (calcd for C18H20O4,
300.1362).
OMe-7′ and OMe-7), other data see Table 4; ESIMS m/z 346
([M + H2O]+); HRFABMS m/z 328.1676 (calcd for C20H24O4,
328.1675).
3-(3-Methoxyphenyl)propyl-3-(3-methoxymethoxyphen-
yl)propanoate (32). The method of preparation of 32 was
similar to that used for the preparation of 18: colorless oil; Rf
3-(3-Methoxyphenyl)propyl-3-(2,3,4-trimethoxyphenyl)-
propanoate (23). The method of preparation of 23 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
1
(hexane/AcOEt, 3:1) 0.22; H NMR (400 MHz, CDCl3) δ 5.15

348 Journal of Natural Products, 2005, Vol. 68, No. 3
Hu et al.
(2 H, s, OCH2O-7), 3.79 (3 H, s, OMe-7′), 3.29 (3 H, s, OMe-7),
other data see Table 4; ESIMS m/z 376 ([M + H2O]+);
HRFABMS m/z 358.1786 (calcd for C21H26O5, 358.1780).
3-(3-Methoxyphenyl)propyl-3-(3-hydroxyphenyl)pro-
panoate (33). The method of preparation of 33 was similar
to that used for the preparation of 18: colorless oil; Rf (hexane/
AcOEt, 3:1) 0.20; 1H NMR (400 MHz, CDCl3) δ 5.38 (1 H, brs,
OH-7), 3.79 (3 H, s, OMe-7′), other data see Table 4; ESIMS
m/z 332 ([M + H2O]+); HRFABMS m/z 314.1523 (calcd for
C19H22O4, 314.1518).
50 µL of 1 M Na2CO3 was added to the incubation solution to
stop the reaction. The negative control was prepared by adding
phosphate buffer instead of the sample in the same way as
test. Acarbose was utilized as positive control. The blank was
prepared by adding phosphate buffer instead of the R-glucosi-
dase in the same way as test. The inhibition rates (%) )
[(ODnegative control
-
ODblank
)
-
(ODtest
-
ODtest blank)]/
(ODnegative blank - ODblank) × 100%. IC50 value of sample was
calculated by the IC50 calculative software.
3-(3,4-Dimethoxymethoxyphenyl)propyl-3-(3-meth-
oxymethoxyphenyl)propanoate (34). The method of prepa-
ration of 34 was similar to that used for the preparation of
Acknowledgment. The authors would like to thank Dr.
X. Hao (KIB, CAS) and Prof. Dr. J. Sto¨ckigt (an der Uni Mainz)
for helpful suggestions. One of the authors (Y.Z.) expresses
his thanks to the Chinese Ministry of Education as well as to
Mr. K. Li for the “Cheung Kong Scholar Chair Professorship”
at Zhejiang University.
1
18: colorless oil; Rf (hexane/AcOEt, 3:1) 0.39; H NMR (400
MHz, CDCl3) δ 5.20 (2 H, s, OCH2O-6′), 5.23 (2 H, s, OCH2O-
7′), 5.14 (2 H, s, OCH2O-7), 3.46 (3 H, s, OMe-6′), 3.63 (3 H, s,
OMe-7′), 3.52 (3 H, s, OMe-7), other data see Table 4; ESIMS
m/z 466 ([M + H2O]+); HRFABMS m/z 448.2067 (calcd for
C24H32O8, 448.2097).
References and Notes
Cytotoxicity Assay. The cytotoxicity activity of test com-
pounds was performed in 96-well microtiter plates by the
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
(MTT) assay. After the cells attached to the substratum,
monolayers of cells were incubated with culture medium
containing the compounds at different concentrations for 72
h. Control cells were treated with the same volume of deionized
water (DI) as the stock of the test agent for examination of
the solvent effect. Then 10 µL of MTT (final concentration 5
mg/mL) was added to each well. After 3 h incubation, the
supernatant was then removed and the cells were dissolved
in DMSO (200 µL/well). The optical density was measured
spectrophotometrically at 570 nm on an enzyme-linked im-
munosorbent assay reader. The 50% cytotoxic concentration
(CC50) was calculated as the compound concentration required
to reduce the MTT signal by 50% compared with untreated
control cultures.
r-Glucosidase Inhibition Assay. The activities of glu-
cosidase were determined in a 96-well plate, and the assay
was based on the cleavage of an R-1,6 glucosidic bond in the
substrate to release p-nitrophenol, which is detected by its
absorbance at 405 nm. A reaction mixture consisted of 25 µL
of R-glucosidase (0.2 U/mL), 25 µL of 23.2 mM substrate (p-
nitrophenyl-R-D-glucopyranoside) solution, 25 µL of various
concentration of samples, and 175 µL of 67 mM phosphate
buffer (pH 6.8). The final volume of the reaction was 250 µL.
The reaction mixture was incubated for 15 min at 37 °C, then
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