Syntheses of two cytotoxic sinapyl alcohol derivatives and isolation of four new related compounds from Ligularia nelumbifolia

Article abstract of DOI:10.1021/np0200257

Phytochemical reinvestigation on Ligularia nelumbifolia afforded four novel sinapyl alcohol analogues named nelumols B-E (1-4) and three known sinapyl alcohol derivatives (5-7). Their structures were elucidated by NMR techniques. Total syntheses of cytotoxic geranyloxy sinapyl alcohol (6) and geranyloxy sinapyl aldehyde (7) were carried out via two different paths. The 4-O-benzyl-substituted analogues (20 and 27) as well as the 4-O-(2-methylbutenyl) derivatives (34 and 35) were also synthesized. The cytotoxicities of 6 and 7 were measured using A-549, HL-60, and KB cancer cell lines.

Full text of DOI:10.1021/np0200257

902
J . Nat. Prod. 2002, 65, 902-908
Syn th eses of Tw o Cytotoxic Sin a p yl Alcoh ol Der iva tives a n d Isola tion of F ou r
New Rela ted Com p ou n d s fr om Ligu la r ia n elu m bifolia
Yu Zhao,*,†,‡ Xiaojiang Hao,‡ Wei Lu,§ J unchao Cai,§ Hong Yu, Thierry Seve´net,| and Franc¸oise Gue´ritte|
Department of Traditional Chinese Medicine and Natural Drug Research, College of Pharmaceutical Sciences,
Zhejiang University, Hangzhou 310031, People’s Republic of China, Laboratory of Phytochemistry, Kunming Institute of
Botany, Chinese Academy of Sciences, Kunming 650204, People’s Republic of China, Shanghai Institute of Material Medica,
Chinese Academy of Sciences, Shanghai 200031, People’s Republic of China, Inmore Laboratory of Biotechnology,
Kunming 650011, People’s Republic of China, and Institut de Chimie des Substances Naturelles, CNRS,
Avenue de la Terrasse, 91198 Gif-sur-Yvette, France
Received J anuary 23, 2002
Phytochemical reinvestigation on Ligularia nelumbifolia afforded four novel sinapyl alcohol analogues
named nelumols B-E (1-4) and three known sinapyl alcohol derivatives (5-7). Their structures were
elucidated by NMR techniques. Total syntheses of cytotoxic geranyloxy sinapyl alcohol (6) and geranyloxy
sinapyl aldehyde (7) were carried out via two different paths. The 4-O-benzyl-substituted analogues (20
and 27) as well as the 4-O-(2-methylbutenyl) derivatives (34 and 35) were also synthesized. The
cytotoxicities of 6 and 7 were measured using A-549, HL-60, and KB cancer cell lines.
The genus Ligularia has been used medicinally for a long
time in China. Distributed in damp shadowy regions beside
brooks and sloping fields, the whole plant of Ligularia
nelumbifolia [(Bur. Et Franch) Hand.-Mazz] (family Com-
positae, Chinese folk name Lian Ye Tuo Wu) has been used
as folk medicine for pulmonary tuberculosis and apoplexy.1
Previous phytochemical examination of Ligularia species
revealed eremophilane derivatives.2-6 Interestingly, no
eremophilane derivatives were found in the species inves-
tigated by us; however, several sinapyl alcohol derivatives
and aromatic components were isolated.3 Thorough exami-
nation of this species has now afforded five further sinapyl
alcohol derivatives (1-5), four of which (1-4) are new
compunds. In the course of our continuing search for
pharmacologically active compounds, two major principles
of this species, geranyloxy sinapyl alcohol (6)3,7 and gera-
nyloxy sinapyl aldehyde (7), were found to be cytotoxic to
KB cell with an IC50 of 3.0 × 10-6 and 2.6 × 10-6 M,
suggesting a secondary OH group at the C-5′ position. This
was supported by an OH absorption band at 3399 cm-1 in
the IR spectrum of 1. The 13C NMR spectrum of 1 was in
complete accord with the proposed structure (Table 2).
Comparison of the 1H and 13C NMR spectra of 2 with
those of 1 indicated that 2 had an oxygenated C-6′, since
H-6′ was shifted downfield (from δ 2.06 to 4.55) when
compared to 1, thus disclosing that H-6′ was vicinal to the
7′(9′) double bond in the case of 2. Furthermore, the 1H
NMR spectrum of 2 revealed the presence of an ethoxy
group at C-6′. EIMS gave the molecular ion peak at m/z
390, which was consistent with the molecular formula
C23H34O5. Since ethanol was exclusive during the extraction
and isolation procedure, compound 2 might be derived
biosynthetically from precursor 6.
respectively. This prompted us to reinvestigate further
analogues in this plant and to synthesize compounds 6 and
7 as well as several analogues for further pharmacological
activity studies.
Resu lts a n d Discu ssion
Nelumol B (1) was obtained as colorless gum. EIMS and
elemental analysis indicated its molecular formula to be
C
21H30O5. Showing the molecular ion peak at m/z 362, the
EIMS of 1 also exhibited a base peak due to a sinapyl
1
alcohol fragment at m/z 210. The H and 13C NMR spectra
of 1 showed close similarities with those of the geranyloxy
1
sinapyl alcohol (6).3,7 In the H NMR spectrum (Table 1),
the only differences were the presence in 1 of an olefinic
methylene multiplet (H-9′) at δ 5.00 (1H, brs) and 4.98 (1H,
brs), as well as an olefinic methyl signal (H-8′) at δ 1.73
(brs, 3H) in place of the olefinic H-6′ and Me-9′ signals of
6. Furthermore, a signal was detected at δ 3.88 (m, 1H),
1
The H NMR spectrum of nelumol D (3) exhibited some
differences from that of geranyloxy sinapyl alcohol 6. The
1
methylene proton (H-5′) of 6 could not be found in the H
NMR spectrum of 3, while two olefinic hydrogens were
observable at δ 5.58 (m, 2H). Furthermore, the methyl
singlets appeared at δ 1.33 (s, 6H), somewhat higher field
* To whom correspondence should be addressed. Tel: (86)-571-87217313.
Fax: (86)-571-87217313. E-mail: dryuzhao@zjuem.zju.edu.cn or dryuzhao@
hotmail.com.
1
than those of 6 in the H NMR spectrum, suggesting that
†Zhejiang University.
an OH group was most likely connected to C-7′, in agree-
ment with the corresponding 13C resonance appearing at
δ 82.04 (s, C-7′). The olefinic carbons attributable to a
trisubstituted double bond at δ 140.16 (s) and 127.88 (d)
‡Kunming Institute of Botany, CAS.
§ Shanghai Institute of Material Medica, CAS.
Inmore Laboratory of Biotechnology.
| Institut de Chimie des Substances Naturelles, CNRS.
10.1021/np0200257 CCC: $22.00
© 2002 American Chemical Society and American Society of Pharmacognosy
Published on Web 06/07/2002

Two Cytotoxic Sinapyl Alcohol Derivatives
J ournal of Natural Products, 2002, Vol. 65, No. 6 903
Ta ble 1. 1H NMR Spectral Data [400 MHz, δH (J , Hz)] for Nelumols B-E (1-4) in CDCl3
position
1
2
3
4
2
6
6.59 s
6.59 s
6.60 s
6.60 s
6.61 s
6.61 s
6.61 s
6.61 s
7
8
9
1′
2′
4′
5′
6′
8′
9′
6.52 dt (15.8, 1.4)
6.28 dt (15.8, 5.8)
4.32 dd (5.8, 1.4)
4.54 br d (7.2)
5.66 tq (7.2, 1.0)
2.06 m
3.88 m
2.06 m
1.73 br s
5.00 br s
6.52 dt (15.9, 1.4)
6.28 dt (15.9, 5.8)
4.32 dd (5.8, 1.4)
4.54 br d (7.1)
5.58 tq (7.1, 1.0)
2.02 m
2.00 m
4.55 br dt (7.0, 1.5)
1.63 br s
4.92 br dd (1.5, 1.5)
4.83 br dd (1.5, 1.5)
1.65 d (1.0)
3.87 s
3.65 q (7.0)
6.55 dt (15.9, 1.4)
6.30 dt (15.9, 6.0)
4.32 dd (6.0, 1.4)
4.55 br d (7.0)
5.58 m
6.55 dt (16.0, 1.5)
6.30 dt (16.0, 6.0)
4.33 dd (6.0, 1.5)
4.54 br d (7.2)
5.61 tq (7.1, 1.0)
5.47 dt (2.0, 1.5)
2.74 m
5.58 m
1.31 s
1.31 s
2.74 dd (6.6, 2.0)
1.26 s
1.26 s
4.98 br s
1.65 d (1.0)
3.86 s
10′
OMe
OEt
1.63 d (0.9)
3.87 s
3.49 q (7.0)
1.22 t (7.0)
1.63 d (1.0)
3.86 s
3.32 q (7.0)
1.14 t (7.0)
1.24 t (7.0)
Ta ble 2. 13C NMR Spectral Data [100 MHz, δ (ppm)] for
Ta ble 3. IC50 of 6 and 7 on Some Selected Pharmacological
Models
a
Nelumols B-E (1-4) in CDCl3
C no.
1 (mult)
2 (mult)
3 (mult)
4 (mult)
A-549 cell
HL-60 cell
KB cell
1
2
3
4
5
6
7
8
136.5 s
103.5 d
153.7 s
139.8 s
153.7 s
103.5 d
131.2 d
129.0 d
63.6 t
136.3 s
103.3 d
153.6 s
141.0 s
153.6 s
103.3 d
131.1 d
127.8 d
63.5 t
136.6 s
103.3 d
153.7 s
139.8 s
153.7 s
103.3 d
131.2 d
127.9 d
63.7 t
138.0 s
103.4 d
153.7 s
140.0 s
153.7
103.4 s
131.2 d
127.9 d
63.7 t
6
7
3.4 × 10-5
M
M
6.7 × 10-6
M
M
3.0 × 10-6
M
M
2.2 × 10-5
1.2 × 10-5
2.6 × 10-6
reaction of the resulting methyl ester 9 with geranyl alcohol
led to the geranyl derivative 10.10 Reduction of 10 by
DIBAH afforded geranyloxy sinapyl alcohol 6 in an 86%
yield, while oxidation of 6 by magnesium dioxide gave
geranyloxy sinapyl aldehyde 7 in 92% yield (Scheme 1).
Another synthetic path started from methyl gallate (11)
(Scheme 2) Acetylation led to product 12, which was
subjected to a selective substitution reaction,11 during
which the 4-acetoxy group was replaced by a geranyl
moiety to yield compound 13b. The unexpected mono-
deacetylated product 13a was also formed in the reaction.
The reaction time and the temperature influenced the
yields of 13a and 13b. The mixture of 13a and 13b was
treated with aqueous K2CO3 to give 14, which was then
transformed to the methoxy derivative 15 (82% yield over
two steps). Reduction of 15 by LAH afforded primary
alcohol 16, which was oxidized to aldehyde 17 by pyri-
dinium chlorochromate in 86% yield. A Knoevenagel con-
densation of 17 with malonic acid in the presence of
piperidine afforded the E-form of acid 18. Reduction of 18
by LAH afforded, apart from the 80% yield of expected
target molecule 6, the 1,4-addition product 19 in 5% yield.
Finally, geranyloxy sinapyl aldehyde 7 was obtained by
manganese dioxide oxidation in 92% yield. The total yield
of 8 was 28%. Cytotoxic screening results of synthetic 6
and 7 against A-549, HL-60, and KB cell lines are shown
in Table 3.
To examine the importance of the C-4 side chain on
cytotoxicity, we designed another target molecule (20) with
a benzyl group attached to O-C(4). Furthermore, a five-
carbon side chain (compound 34) was also introduced to
extend the SAR concept. Two paths were examined to
synthesize these analogues, which are shown in Schemes
2 and 3. Cytotoxicity screening of 20, 27, 34, and 35 is
shown in Table 4. It was seen that compounds 20 and 27
were less cytotoxic to KB cells than 6 and 7, while the five-
carbon side chain derivatives 34 and 35 had cytotoxicities
to KB cells similar to those of 1 and 2.
9
1′
2′
3′
4′
5′
6′
7′
8′
9′
10′
OMe
OEt
69.2 t
69.2 t
69.3 t
69.4 t
121.4 d
132.4 s
39.5 t
88.9 d
28.7 t
143.8 t
17.1 q
114.1 t
16.1 q
56.1 q
120.6 d
132.3 s
35.4 t
121.2 d
132.3 s
42.2 t
140.2 s
127.9 d
70.8s
29.7 q
29.7 q
16.3 q
56.0 q
56.0 t
121.2 d
132.3 s
126.9 d
140.2 s
42.6 t
32.5 t
75.2 d
147.2 s
17.4 q
111.0 t
16.1 q
56.0 q
63.8 t
74.8 s
26.4 q
26.4 q
16.2 q
56.0 q
57.7 t
a
Assignment in the same column could be exchangeable.
were assigned to C-5′ and C-6′, respectively. This side chain
is similar to that of the sinapyl alcohol derivative 5,
previously isolated from Ligularia duciformis.8 However,
the molecular ion peak of 3 appearing at m/z 406, i.e., 44
mass units higher than that of 5, as well as the NMR data
all indicated that 3 was an C-5′-OEt derivative of 5 (Tables
1 and 2). Compound 3 might be another artifact or the
enzymatic derivative of 5, as mentioned above.
Nelumol E (4) had a molecular ion peak and NMR data
similar to those of 3. Elemental analysis and a DEPT
spectrum revealed its molecular formula to be C23H34O6,
apparently isomeric with 3. Scrutiny of its H and 13C NMR
spectra with those of 3 led to the assignment of a 2′(3′),4′-
(5′)-diene system in compound 4 (Tables 1 and 2). A
COLOC experiment on 4 exhibited correlations of olefinic
H-4′ with C-2′ and C-10′, consistent with the presence of a
conjugated diene moiety in 4. This enol ether could be
either an artifact or a biosynthetic derivative, as discussed
above.
As 6 and 7 were cytotoxic to KB cells (Table 3) and
appeared as principle metabolites in L. nelumbifolia,
syntheses of further sinapyl alcohol derivatives become
interesting. Thus, 6 and 7 were selected to be totally
synthesized.
1
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. 1H NMR and 13C
NMR spectra were measured on Bruker AM-400 MHz and
Bruker AC-300 MHz NMR instruments, with TMS as internal
The first path used commercially available sinapinic acid
8 as starting material. After esterification,9 a Mitsunobu

904 J ournal of Natural Products, 2002, Vol. 65, No. 6
Zhao et al.
Sch em e 1a
a
(a) H2SO4, MeOH, reflux, 2 h, 98%; (b) geranyl alcohol, Ph3P, DEAD, 24 h, 50%; (c) DIBAH, THF, -78 °C, 2 h, 86%; (d) 1: PCC, CH2Cl2, rt, 6 h, 81%;
2: MnO2, EtOAc, rt, 92%.
Sch em e 2a
a
(a) H2SO4, MeOH, reflux, 2 h, 96%; (b) Ac2O, Py, rt, 12 h, 93%; (c) geranyl bromide, K2CO3, DMF, 0 °C, 24 h, 50% of 13b, 29% of 13a ; (d) K2CO3,
MeOH-H2O, rt, 0.5 h, 90%; (e) MeI, K2CO3, reflux, 3 h, 91%; (f) LAH, ether, 0 °C, 90%; (g) PCC, CH2Cl2, rt, 6 h, 86%; (h) malonic acid, piperidine, Py, reflux,
4 h, 86%; (i) LAH, ether, 0 °C, 80% of 6, 5% of 19; (j) MnO2, EtOAc, rt, 2 h, 92%.
Ta ble 4. IC50 of Compounds 20, 27, 34, and 35 on KB Cells
(mol/L)
and 35 mg of 7. F9 (3.2 g) was chromatographed (200 g of silica
gel gel, 200-300 mesh) using a CHCl3-Me2CO (20:1-1:1) step
gradient. Eluates 25-28 (150 mL each) were combined and
purified by PTLC (CHCl3-Me2CO, 3:1) to give 14 mg of 1 (Rf
) 0.46). Eluate 14 (120 mL) was evaporated and purified by
PTLC (C6H6-Me2CO, 4:1) to give 21 mg of 2. Eluate 17 (80
mL) contained 26 mg of 5, which was obtained by PTLC with
C6H6-Me2CO, 8:1 (Rf ) 0.65). F10 (6.6 g) was rechromato-
graphed over silica H (200 g) with a CHCl3-EtOAc (10:1-1:
2) solvent system. Eluates 16-17 (125 mL each) were com-
bined and evaporated, and the residue (86 mg) was purified
through PTLC (CHCl3-MeOH, 8:1) to afford 17 mg of 3 (Rf )
0.57) and 15 mg of 4 (Rf ) 0.49).
4-O-[(2E)-3,7-Dim eth yl-2,7-octa d ien -5-ol]sin a p yl a lco-
h ol (1): gum; IR (KBr) νmax 3399 (OH), 3349 (OH), 2977, 1659,
1583, 1504, 1459, 1420, 1332, 1241, 1127, 963 cm-1; EIMS m/z
(rel int) 362 [M]+ (16), 347 (3), 344 (5), 329 (6), 306 (14), 277
(10), 252 (18), 238 (50), 210 (100), 182 (36), 167 (42), 154 (18);
1H NMR (CDCl3) data, see Table 1; 13C NMR (CDCl3) data,
see Table 2; anal. C 69.56%, H 8.27%, calcd for C21H30O, C
69.61%, H 8.29%.
20
27
34
35
8.6 × 10-4
6.4 × 10-4
7.8 × 10-6
5.3 × 10-6
standard. HREIMS and EIMS were performed on a VG Auto
Spec-3000 MS instrument. EIMS: direct inlet, 70 eV. Solvents
and reagents were purified according to standard laboratory
techniques. IR spectra were recorded on a Perkin-Elmer 577
spectrometer.
P la n t Ma ter ia l. The material plant was collected in August
2000, Zhaotong County, Yunnan Province, China, and identi-
fied by Prof. Hua Peng. A voucher specimen (no. 20000806) is
deposited in the Laboratory of Phytochemistry, Kunming
Institute of Botany, Chinese Academy of Sciences, Kunming,
Yunan Province, China.
Extr a ction a n d Isola tion . Air-dried roots of Ligularia
nelumbifolia [(Bur. Et Franch) Hand.-Mazz] (2.0 kg) were
powdered and extracted with petroleum ether (60-90 °C)-
Et2O-MeOH (1:1:1) at room temperature (3 days × 3) to give
85 g of crude extract, which was subjected to column chroma-
tography on 1 kg of silica gel with petroleum ether containing
gradually increasing amounts of EtOAc (1:0-1:1). Ten crude
fractions (F1-F10) were obtained. F1-F7 contained, by TLC,
mainly the same products reported previously.3 F8 (2.1 g)
afforded, after repeated column chromatography, 86 mg of 6
4-O -[(2E )-3,7-D i m e t h y l-6-e t h o x y -2,7-o c t a d i e n e ]-
sin a p yl a lcoh ol (2): gum; IR (KBr) νmax 3398 br (OH), 3072,
2939, 1653, 1583, 1504, 1456, 1418, 1333, 1241, 1128, 992, 904,
629 cm-1; EIMS m/z (rel int) 390 [M]+, (15), 375 (22), 349 (18),
344 (16), 277 (10), 210 (100), 182 (55), 167 (43), 137 (16), 121
(14), 113 (20), 69 (72), 46 (23); 1H NMR (CDCl3) data, see Table

Two Cytotoxic Sinapyl Alcohol Derivatives
J ournal of Natural Products, 2002, Vol. 65, No. 6 905
Sch em e 3a
a
(a1) Benzyl bromide, DMF, 0 °C, 24 h; (a2) 2-methylbutenyl bromide, K2CO3, DMF, 0 °C, 10 h; (b) K2CO3, MeOH-H2O, rt, 0.5 h; (c) MeI, K2CO3, reflux,
3 h; (d) LAH, ether, 0 °C; (e) PCC, CH2Cl2, rt, 6 h, (f) malonic acid, piperidine, Py, reflux, 4 h; (g) LAH, ether, 0 °C; (h) MnO2, EtOAc, rt, 2 h (MB )
2-methylbutenyl).
Sch em e 4a
a
(a1) Benzol, Ph3P, DEAD, 24 h, 65%; (a2) 2-methylbutenol, Ph3P, DEAD, 24 h, 60%; (b) DIBAH, THF, -78 °C, 2 h; 88% of 20, 80% of 34; (c) 1: PCC,
CH2Cl2, rt, 6 h; 83% of 27, 81% of 35; 2: MnO2, EtOAc, rt, 92% of 27, 94% of 35 (MB ) 2-methylbutenyl).
1; 13C NMR (CDCl3) data, see Table 2; anal. C 70.73%, H
8.72%, calcd for C23H34O5, C 70.77%, H 8.72%.
solution was stirred overnight and then refluxed for 0.5 h. The
cooled solution was partitioned between H2O (30 mL) and
EtOAc (30 mL × 3) and dried (MgSO4). After filtration, the
solvent was evaporated and the residue was subjected to CC
(petroleum ether-Et2O, 5:1-2:1); 186 mg of 10 was isolated
(50%).
4-O-[(2E,5E)-3,7-Dim eth yl-5-eth oxy-2,5-octa d ien e-7-ol]-
sin a p yl a lcoh ol (3): gum; IR (KBr) νmax 3408 br (OH), 2967,
2926, 1665, 1582, 1504, 1459, 1417, 1332, 1240, 1127, 969, 914,
744 cm-1; EIMS m/z (rel int) 406 [M]+, (8), 391 (2), 389 (5),
360 (6), 314 (15), 264 (3), 210 (100), 197 (3), 182 (25), 167 (42),
4-Ger a n yl sin a p ic a cid m eth yl ester (10): gum; 1H NMR
(CDCl3, 400 MHz) 7.57 (1H, d, J ) 16.0 Hz, H-7), 6.72 (2H, s,
H-2, H-6), 6.32 (1H, d, J ) 15.8 Hz, H-8), 5.53 (1H, brt, J )
7.0 Hz, H-2′), 5.05 (1H, m, H-6′), 4.55 (2H, d, J ) 7.1 Hz, H-1′),
3.86 (6H, s, OMe-3, OMe-5), 3.79 (3H, s, CO2Me), 2.03 (4H, m,
H-4′, H-5′), 1.65 (3H, s, H-8′), 1.63 (3H, s, H-9′), 1.57 (3H, s,
H-10′); 13C NMR (CDCl3, 100 MHz) δ 167.3 (s, C-9), 153.82 (s,
C-3, C-5), 144.90 (d, C-7), 141.6 (s, C-4), 139.0 (s, C-2), 131.5
(s, C-3′), 129.7 (s, C-7), 123.92 (d, C-6′), 119.97 (d, C-2′), 116.77
(d, C-8), 105.17 (d, C-2, C-6), 69.50 (t, C-1′), 56.09 (q, OMe-3,
OMe-5), 51.62 (q, CO2Me), 39.57 (t, C-4′), 26.39 (t, C-5′), 25.62
(t, C-8′), 17.61 (q, C-9′), 16.31 (q, C-10′); EIMS m/z 374 [M]+
(1), 343 (1), 305 (2), 266 (1), 248 (1), 238 (100), 223 (3), 207
(8), 175 (3), 163 (2), 135 (2), 69 (13); HREIMS m/z 374.2082
(calcd for C22H30O5, 374.2093).
1
154 (16), 69 (18), 46 (48); H NMR (CDCl3) data, see Table 1;
13C NMR (CDCl3) data, see Table 2; anal. C 67.90%, H 8.31%,
calcd for C23H34O6, C 67.98%, H 8.37%.
4-O-[(2E,4E)-3,7-Dim eth yl-5-eth oxy-2,4-octa d ien -7-ol]-
sin a p yl a lcoh ol (4): gum, IR (KBr) νmax 3402 br (OH), 3349,
2973, 2933, 1673, 1582, 1503, 1457, 1418, 1333, 1240, 1128,
969, 844 cm-1; EIMS m/z (rel int) 406 [M]+, (12), 391 (4), 389
(8), 374 (4), 360 (2), 343 (5), 210 (100), 197 (6), 182 (38), 167
(44), 154 (23), 128 (6), 69 (18), 46 (36); 1H NMR (CDCl3) data,
see Table 1; 13C NMR (CDCl3) data, see Table 2; anal. C,
67.90%, H, 8.31%, calcd for C23H34O6, C, 67.98%, H, 8.37%.
Sin a p ic Acid Meth yl Ester (9). NMR and physical data
were identical with a previous publication.9 EIMS: m/z 238
[M]+ (100), 223 (9), 207 (95), 175 (33), 163 (11), 119 (10), 91
(6). HREIMS: 238.0856 (calcd for C12H14O5, 238.0841).
Eth er ifica tion of 9. To a stirred solution of 313 mg (1.2
mmol) of Ph3P and 240 mg of 9 (1.0 mmol) in dry THF (10
mL) was added 150 mg of geraniol (1.0 mmol) and DEAD (262
µL, 1.2 mmol) at room temperature under nitrogen. The
Red u ction of 10. To a stirred solution of 374 mg (1.0 mmol)
of 10 in dry Et2O (10 mL) was added DIBAH (1.0 mL, 1.0 M
in hexane) at -78 °C under nitrogen. The solution was stirred
for 0.5 h, 3 mL of H2O was added at -78 °C to quench the
reaction, and the solution was allowed to warm to room

906 J ournal of Natural Products, 2002, Vol. 65, No. 6
Zhao et al.
temperature. Ten milliliters of 1 M HCl was added, and the
solution was extracted with EtOAc (15 mL × 3). The organic
layers were combined and dried (MgSO4). Purification by flash
column afforded 299 mg of 6 (86%). Physical and NMR data
for compound 6 have been reported in an earlier publication.3
Allylic Oxid a tion of 6 by Mn O2. To a stirred suspension
of 105 mg (1.2 mmol) of MnO2 in EtOAc (15 mL) was added
345 mg (1.0 mmol) of 6 in EtOAc (5 mL) at room temperature,
and the solution was stirred for 4 h. After filtration, the eluate
was evaporated to dryness and was partitioned between H2O
(20 mL) and Et2O (60 mL). The organic layer was combined
and dried (MgSO4), and the solvent was evaporated to afford
7 (317 mg, 92%). Physical and NMR data for compound 7 have
been reported in an earlier publication.3
Dea cetyla tion of 12. To a stirred solution of 15.5 g of 12
(50 mmol) in dry DMF (150 mL) was added 20.7 g of K2CO3
(150 mmol) at 0 °C. The solution was stirred for 20 min and
10.85 g (9.9 mL) of geranyl bromide (60 mmol) in dry DMF
(60 mL) was added in 10 min. The solution was stirred for 10
h. After suction filtration, 300 mL of H2O was added. The
mixture was extracted with EtOAc (600 mL), followed by Et2O
(600 mL). The organic layers were combined, washed with
brine (100 mL), and dried (MgSO4). The solution was evapo-
rated, and the residue was subjected to CC (hexane-Et2O, 5:1)
to afford 10.11 g (25 mmol) of 13b (50%) and 5.25 g (14.5 mmol)
of 13a (29%). Also, 925 mg (3.0 mmol) of 12 (6%) was recovered.
4-Ger a n oyl-3,5-d ia cetoxyben zoic a cid m eth yl ester
(13b): gum; 1H NMR (CDCl3, 300 MHz) δ 7.64 (2H, s, H-2,
H-6), 5.42 (1H, brt, J ) 7.0 Hz, H-2′), 5.09 (1H, m, H-6′), 4.59
(2H, d, J ) 7.2 Hz, H-1′), 3.89 (3H, s, CO2Me), 2.36 (3H, s,
OCOCH3), 2.09 (4H, m, H-4′, H-5′), 1.70 (3H, s, H-8′), 1.68 (3H,
s, H-9′), 1.62 (3H, s, H-10′); HREIMS m/z 404.1818 (calcd for
1.64 (3H, s, C-9′), 1.60 (3H, s, C-10′); 13C NMR (CDCl3, 75 MHz)
δ 166.8 (s, C-7), 153.4 (s, C-3, C-5), 141.9 (s, C-1), 141.2 (s,
C-4), 131.6 (s, C-3′), 125.1 (s, C-7′), 123.9 (d, C-6′), 119.9 (s,
C-2′), 109.6 (d, C-2, C-6), 69.4 (t, C-1′), 56.2 (q, OMe-3, OMe-
5), 52.2 (q, CO2Me), 39.6 (t, C-4′), 26.4 (t, C-5′), 25.6 (q, C-8′),
17.6 (q, C-9′), 16.3 (q, C-10′); HREIMS δ 348.1925 (calcd for
C
20H28O5, 348.1937).
4-Ger a n oyl-3,5-d im eth oxyben zyl Alcoh ol (16). To a
stirred suspension of LAH (49 mg, 1.28 mmol) in Et2O (50 mL)
at 0 °C was added a solution of 15 (280 mg, 0.8 mmol) in dry
Et2O (20 mL) under argon atmosphere. The solution was
stirred for 10 min and was quenched by H2O (8 mL). Then 50
mL of 1 N HCl was added, and the mixture was extracted by
Et2O (150 mL). The ether layers were combined and dried
(MgSO4). Evaporation of the solvent followed by PTLC afforded
1
230 mg of 16 (0.72 mmol, 90%): gum; H NMR (CDCl3, 300
MHz) δ 6.62 (2H, brs, H-2, H-6), 5.60 (1H, brt, J ) 7.0 Hz,
H-2′), 5.05 (1H, m, H-6′), 4.69 (2H, brs, H-7), 4.52 (2H, d, J )
7.1 Hz, H-1′), 3.89 (6H, s, OMe-3, OMe-5), 2.10 (4H, m, H-4′,
H-5′), 1.70 (3H, s, H-8′), 1.68 (3H, s, H-9′), 1.63 (3H, s, H-10′);
HREIMS m/z 320.1966 (calcd for C19H28O4, 320.1988).
4-Ger an oyl-3,5-dim eth oxyben zaldeh yde (17). To a stirred
suspension of PCC (225 mg, 1.04 mmol) in CH2Cl2 (30 mL) at
0 °C was added 208 mg of 16 (0.65 mmol) in CH2Cl2 (10 mL).
The solution was stirred at 0 °C for 5 h. The suspension was
filtered and washed by Et2O (60 mL) and partitioned between
Et2O (90 mL) and H2O (30 mL). The ether layers were
combined, dried (MgSO4), and evaporated to afford a residue.
PTLC of the residue afforded finally 178 mg of 17 (0.56 mmol,
1
86%): gum; H NMR (CDCl3, 300 MHz) δ 9.86 (1H, s, H-7),
7.13 (2H, s, H-2, H-6), 5.60 (1H, brt, J ) 6.9 Hz, H-2′), 5.05
(1H, m, H-6), 4.77 (2H, brd, J ) 7.0 Hz, H-1′), 3.94 (6H, s,
OMe-3, OMe-5), 2.04 (4H, m, H-4′, H-5′), 1.64 (3H, s, H-8′),
1.63 (3H, s, H-9′), 1.57 (3H, s, H-10′); 13C NMR (CDCl3, 75
MHz) δ 191.2 (s, C-7), 154.2 (s, C-3, C-5), 142.5 (s, C-1), 142.3
(s, C-4), 131.7 (s, C-3′), 131.8 (s, C-7′), 123.9 (d, C-6′), 119.78
(d, C-2′), 106.6 (d, C-2, C-6), 69.6 (t, C-1′), 56.3 (q, OMe-3, OMe-
5), 39.6 (t, C-4′), 26.4 (t, C-5′), 25.7 (q, C-8′), 17.7 (q, C-9′), 16.4
(q, C-10′); HREIMS m/z 318.1822 (calcd for C19H26O4, 318.1831).
4-Ger a n oylsin a p ic Acid (18). To a stirred solution of
malonic acid (156 mg, 1.5 mmol) in Py (15 mL) at room
temperature was added 475 mg (1.5 mmol) of 17 in Py (10
mL). Piperidine (20 mg) was added to the solution. The mixture
was heated at 120 °C for 4 h. The solvent was evaporated and
dried (MgSO4), evaporated, and followed by CC (CHCl3-
MeOH, 8:1) to afford 463 mg of 18 (1.3 mmol, 86%): gum; 1H
NMR (CDCl3, 400 MHz) δ 7.69 (1H, d, J ) 15.9 Hz, H-7), 6.75
(2H, s, H-2, H-6), 6.34 (d, J ) 15.8 Hz, H-8), 5.53 (1H, brt, J
) 7.2 Hz, H-2′), 5.05 (1H, m, H-6′), 4.57 (2H, brd, J ) 7.1 Hz,
H-1′), 3.87 (6H, s, OMe-3, OMe-5), 2.03 (4H, m, H-4′, H-5′),
1.65 (3H, s, H-8′), 1.64 (3H, s, H-9′), 1.57 (3H, s, H-10′); 13C
NMR (CDCl3, 100 MHz) δ 172.1 (s, C-9), 154.0 (s, C-3, C-5),
147.1 (d, C-7), 141.8 (s, C-4), 139.4 (s, C-1), 131.6 (s, C-3′), 129.4
(s, C-7′), 134.0 (d, C-6′), 120.0 (d, C-2′), 116.2 (d, C-8), 105.5
(d, C-2, C-6), 69.6 (t, C-1′), 56.2 (q, OMe-3, OMe-5), 39.6 (t,
C-4′), 26.4 (t, C-5′), 25.7 (q, C-8′), 17.7 (q, C-9′), 16.4 (q, C-10′);
EIMS m/z 360 [M]+, (3), 345 (1), 331 (11), 316 (3), 224 (100),
209 (4), 198 (26), 181 (4), 69 (23); HREIMS m/z 360.1927 (calcd
for C21H28O5, 360.1937).
C
22H28O7, 404.1835).
4-Ger a n oyl-3-a cetoxy-5-h yd r oxysin a p ic a cid m eth yl
ester (13a ): gum; 1H NMR (CDCl3, 300 MHz) δ 7.52 (1H, brs,
H- 2), 7.36 (1H, brs, H-6), 5.90 (1H, brs, OH-5), exchanged in
D2O), 5.48 (1H, t, J ) 7.0 Hz, H-2′), 5.08 (1H, m, H-6′), 4.63
(1H, d, J ) 7.1 Hz, H-1′), 3.90 (3H, s, CO2Me), 2.36 (3H, s,
OCOCH3), 2.10 (4H, m, H-4′, H-5′), 1.70 (3H, s, H-8′), 1.66 (3H,
s, H-9′), 1.61 (3H, s, H-10′); HREIMS m/z 362.1709 (calcd for
C
20H26O6, 362.1729).
Dea cetyla tion of 13 (13a a n d 13b) (e.g., 13a ). To a stirred
solution of 13a (2.91 g, 8.0 mmol) in MeOH (200 mL) at 0 °C
was added 5.66 g (43.2 mmol) of K2CO3 in H2O (60 mL) in 10
min. The solution was stirred for 20 min, and the solvent was
evaporated. Then 1 M HCl was added to adjust the pH value
to 2, and the aqueous solution was extracted by EtOAc (300
mL). The organic layers were combined and dried (MgSO4),
and the solvent was evaporated to afford 2.32 g (7.2 mmol) of
14 (90%).
4-Ger a n oyl-3,5-h yd r oxyben zoic a cid m eth yl ester (14):
gum; 1H NMR (300 MHz, CDCl3) δ 7.25 (2H, s, H-2, H-6), 5.91
(1H, s, exchanged in D2O, ArOH), 5.60 (1H, brt, J ) 7.0 Hz,
H-2′), 5.09 (1H, m, H-6′), 4.66 (2H, d, J ) 7.1 Hz, H-1′), 3.90
(3H, s, CO2Me), 2.08 (4H, m, H-4′, H-5′), 1.70 (3H, s, H-8′),
1.66 (3H, s, H-9′), 1.61 (3H, s, H-10′); 13C NMR (75 MHz,
CDCl3) δ 166.9 (s, C-7), 149.2 (s, C-3, C-5), 145.2 (s, C-1), 137.4
(s, C-4), 132.1 (s, C-3′), 126.1 (s, C-7′), 123.5 (s, C-6′), 118.7 (s,
C-2′), 109.5 (d, C-2, C-6), 69.9 (t, C-1′), 52.2 (q, CO2Me), 39.6
(t, C-4′), 26.2 (t, C-5′), 25.6 (q, C-8′), 17.7 (q, C-9′), 16.4 (q,
C-10′); HREIMS δ 320.1622 (calcd for C18H24O5, 320.1624).
4-Ger a n oyl-3,5-m eth oxyben zoic a cid m eth yl ester (15).
To a stirred solution of 14 (320 mg, 1.0 mmol) in dry DMF (30
mL) was added 8 mg of K2CO3 (6.0 mmol) at room temperature
under argon, then 0.312 mL (5.0 mmol) of MeI in DMF (5 mL)
was added. The solution was heated at 100 °C for 3 h and was
cooled to 25 °C. After suction filtration, the filtrate was
partitioned between H2O (120 mL) and EtOAc-ether (100 mL/
100 mL). The organic layers were combined and dried (MgSO4).
The solution was evaporated under reduced pressure, and the
residue was subjected to PTLC; 315 mg (0.91 mmol) of 15 was
obtained (91%): gum; 1H NMR (CDCl3, 300 MHz) δ 7.30 (2H,
s, H-2, H-6), 5.55 (1H, brt, J ) 7.0 Hz, H-2′), 5.05 (1H, m, H-6′),
4.59 (1H, d, J ) 7.0 Hz, H-1′), 3.93 (3H, s, H-9′), 3.89 (6H, s,
OMe-3, OMe-5), 2.04 (4H, m, H-4′, H-5′), 1.66 (3H, s, C-8′),
4-Ger a n oyl-7,8-d ih yd r osin a p ic Acid (19). To a stirred
solution of LAH (41 mg, 1.09 mmol) in dry Et2O (15 mL) at 0
°C was added 195 mg (0.54 mmol) of 18 in dry Et2O (10 mL)
under argon. The mixture was stirred for 1 h at 0 °C and was
quenched by H2O (6 mL). Then 1 N HCl (10 mL) was added
and extracted by Et2O (30 mL). The ether layer was dried
(MgSO4), evaporated, and subjected to CC (petroleum ether-
Et2O, 1:2) to afford 150 mg of 7 (0.43 mmol, 80%) and 10 mg
of 19 (0.03 mmol, 5%): gum; 1H NMR (CDCl3, 300 MHz) δ
6.46 (2H, brs, H-2, H-6), 5.58 (1H, brt, J ) 7.0 Hz, H-2′), 5.07
(1H, m, H-6′), 4.55 (2H, brd, J ) 7.0 Hz, H-1′), 3.86 (2H, brt,
J ) 7.6 Hz, H-9), 3.90 (6H, s, OMe-3, OMe-5), 2.79 (2H, brt, J
) 7.6 Hz, H-7), 2.01-1.94 (2H, m, H-8); HREIMS m/z 348.2298
(calcd for C21H32O4, 348.2300).
4-O-Ben zyl-3,5-d ia cetoxyben zoic Acid Meth yl Ester
(21). The method of preparation of 21 was similar to that used

Two Cytotoxic Sinapyl Alcohol Derivatives
J ournal of Natural Products, 2002, Vol. 65, No. 6 907
for the preparation of 13. The yield 21 from 12 was 67%. This
compound was identical to that reported by Pearson et al.11 It
was noticeable that no mono-deacetylated compound was
obtained in this reaction.
OMe-5), 1.72 (3H, s, H-4′), 1.64 (3H, s, H-5′); HREIMS m/z
280.1300 (calcd for C15H20O5, 280.1311).
4-O-(2-Meth yl-2-bu ten yl)-3,5-d im eth oxyben zyl Alcoh ol
(31). The method of preparation of 31 was similar to that used
for the preparation of 16. The yield of 31 from 30 was 89%:
gum; 1H NMR (CDCl3, 400 MHz) δ 6.53 (2H, s, H-2, H-6), 5.51
(1H, brt, J ) 7.1 Hz, H-2′), 4.56 (2H, s, H-7), 4.42 (1H, d, J )
7.2 Hz, H-1′), 3.79 (6H, s, OMe-3), OMe-5), 1.69 (3H, s, H-5′),
1.62 (3H, s, H-4′); EIMS m/z 252 [M]+ (6), 239 (6), 235 (5), 226
(28), 211 (10), 205 (33), 184 (100), 182 (2), 167 (14), 155 (12),
153 (8), 127 (8), 123 (12), 109 (9), 69 (18); HREIMS δ 252.1374
(calcd for C14H20O4, 252.1362).
4-O-Ben zyl-3,5-d ih yd r oxyben zoic Acid Meth yl Ester
(22). The method of preparation of 22 was similar to that used
for the preparation of 14. The yield of 22 from 21 was 92%:
1
gum; H NMR (CDCl3, 300 MHz) δ 7.43-7.36 (5H, m, H-3′-
H-7′), 7.25 (2H, s, H-2, H-6), 5.80 (brs, exchanged in D2O,
ArOH), 5.16 (2H, s, H-1′), 3.90 (3H, s, CO2Me); HREIMS m/z
274.0870 (calcd for C15H14O4, 274.0841). This compound was
first reported by Pearson et al.16
4-O-Ben zyl-3,5-d im eth oxyben zoic Acid Meth yl Ester
(23). The method of preparation of 23 was similar to that used
for the preparation of 15. The yield of 23 from 22 was 90%:
gum; 1H NMR (CDCl3, 300 MHz) δ 7.25-7.5 (5H, m, H-2′-
H-7′), 5.10 (2H, s, H-1′), 3.93 (3H, s, Me-8), 3.90 (3H, s, OMe-
3, OMe-5); 13C NMR (CDCl3, 75 MHz) δ 166.8 (s, C-7), 153.3
(s, C-3, C-5), 141.0 (s, C-1), 137.5 (s, C-4), 128.5 (d, C-3′, C-7′),
128.27 (d, C-4′, C-6′), 128.1 (d, C-5′), 125.4 (s, C-2′), 106.9 (d,
C-2, C-6), 75.0 (t, C-1′), 56.3 (q, OMe-3, OMe-5), 52.3 (q,
CO2Me); HREIMS m/z 302.1133 (calcd for C17H18O5, 302.1154).
This compound was first reported by J urd et al.14
4-O-Ben zyl-3,5-d im et h oxyb en zyl Alcoh ol (24). The
method of preparation of 24 was similar to that used for the
preparation of 16. The yield of 24 from 23 is 94%. This
compound was identical to that reported by Battersby et al.15
4-O -Ben zyl-3,5-dim eth oxyben zaldeh yde (25).Themethod
of preparation of 25 was similar to that used for the prepara-
tion of 17. The yield of 25 from 24 was 88%. This compound
was identical to that reported by Battersby et al.16
4-O-Ben zylsin a p ic Acid (26). The method of preparation
of 26 was similar to that used for the preparation of 18. The
yield of 26 from 25 was 90%. This compound was identical to
that reported by Kametani et al.17
4-O-Ben zylsin a p yl Alcoh ol (20). The method of prepara-
tion of 20 was similar to that used for the preparation of 7.
The yield of 20 from 26 was 87%: gum; 1H NMR (CDCl3, 300
MHz) δ 7.51-7.20 (5H, m, H-3′-H-7′), 6.56 (2H, brs, H-2, H-6),
6.50 (1H, d, J ) 15.8 Hz, H-7), 6.28 (1H, dt, J ) 15.8, 5.7 Hz,
H-8), 5.06 (2H, brs, H-1′), 3.88 (6H, s, OMe-3, OMe-5); HREIMS
m/z 300.1341 (calcd for C18H20O4, 300.1362).
4-O-Ben zylsin a p a ld eh yd e (27). The method of prepara-
tion of 27 was similar to that used for the preparation of 7.
The yield of 27 from 20 was 94%: gum; 1H NMR (CDCl3, 300
MHz) δ 9.68 (1H, d, J ) 7.5 Hz, H-9), 7.52-7.22 (6H, m, H-3′-
H-7′, H-7), 6.74 (2H, brs, H-2, H-6), 6.61 (1H, dd, J ) 15.8, 7.5
Hz, H-8), 5.09 (2H, brs, H-1′), 3.90 (6H, s, OMe-3, OMe-5);
HREIMS m/z 298.1229 (calcd for C18H18O4, 298.1205).
4-O-(2-Met h yl-2-b u t en yl)-3,5-d ia cet oxyb en zoic Acid
Meth yl Ester (28). The method of preparation of 28 was
similar to that used for the preparation of 13. The yield of 28
from 12 was 60%: gum; 1H NMR (CDCl3, 400 MHz) δ 7.65
(2H, s, H-2, H-6), 5.37 (1H, t, J ) 7.0 Hz, H-2′), 4.48 (2H, d, J
) 7.25 Hz, H-1′), 3.86 (3H, s, CO2Me), 2.31 (6H, s, OCOCH3-3,
OCOCH3-5), 1.74 (3H, s, H-4′), 1.64 (3H, s, H-5′); EIMS m/z
336 [M]+ (1), 321 (1), 295 (1), 281 (2), 286 (6), 253 (2), 237 (4),
226 (41), 195 (3), 184 (60), 153 (5), 121 (4), 85 (14), 69 (100);
HREIMS m/z 336.1208 (calcd for C17H20O7, 336.1209).
4-O-(2-Meth yl-2-bu ten yl)-3,5-d ih yd r oxyben zoic Acid
Meth yl Ester (29). The method of preparation of 29 was
similar to that used for the preparation of 14. The yield of 29
from 28 was 91%: gum; 1H NMR (CDCl3, 400 MHz) δ 7.20
(2H, s, H-2, H-6), 5.96 (1H, brs, exchanged in D2O, ArOH),
5.50 (1H, t, J ) 7.0 Hz, H-2′), 4.60 (2H, d, J ) 7.0 Hz, H-1′),
3.86 (3H, s, CO2Me), 1.75 (3H, s, H-4′), 1.63 (3H, s, H-5′); EIMS
m/z 252 [M]+ (21), 235 (8), 226 (75), 211 (33), 205 (18), 184
(44), 167 (5), 153 (46), 149 (8), 69 (100); HREIMS m/z 252.0978
(calcd for C13H16O5, 252.0998).
4-O -(2-Me t h y l-2-b u t e n y l)-3,5-d im e t h o x y b e n za ld e -
h yd e (32). The method of preparation of 32 was similar to
that used for the preparation of 17. The yield of 32 from 31
was 85%: gum; 1H NMR (CDCl3, 400 MHz) δ 9.81 (1H, s, H-7),
7.07 (2H, s, H-2, H-6), 5.49 (1H, t, J ) 7.1 Hz, H-2′), 4.56 (2H,
d, J ) 7.3 Hz, H-1′), 3.87 (6H, s, OMe-3, OMe-5), 1.69 (3H, s,
H-4′), 1.62 (3H, s, H-5′); EIMS m/z 250 [M]+ (1), 235 (1), 226
(16), 196 (2), 182 (100), 167 (8), 153 (2), 139 (4), 125 (5), 110
(6), 95 (7); HREIMS m/z 250.1199 (calcd for C14H18O4, 250.1205).
4-O-(2-Meth yl-2-bu ten yl)sin a p ic Acid (33). The method
of preparation of 33 was similar to that used for the prepara-
1
tion of 18. The yield of 33 from 32 was 88%: gum; H NMR
(CDCl3, 400 MHz) δ 7.68 (1H, d, J ) 15.7 Hz, H-7), 6.75 (1H,
s, H-2, H-6), 6.34 (1H, d, J ) 15.8 Hz, H-8), 5.53 (1H, dt, J )
7.2, 1.3 Hz, H-2′), 4.57 (2H, d, J ) 7.2 Hz, H-1′), 3.86 (6H, s,
OMe-3, OMe-5), 1.72 (3H, s, H-4′), 1.65 (3H, s, H-5′); EIMS
m/z 292 [M]+ (6) 277 (2), 265 (5), 250 (3), 224 (100), 209 (40),
197 (3), 195 (3), 181 (10), 163 (12), 149 (8), 135 (9), 121 (15),
69 (69); HREIMS m/z 292.1303 (calcd for C16H20O5, 292.1311).
4-O-(2-Meth yl-2-bu ten yl)sin apyl Alcoh ol (34).The method
of preparation of 34 was similar to that used for the prepara-
tion of 6 from 18. The yield of 34 from 33 was 81%, while the
yield of byproduct 36 was 8%: gum; 1H NMR (CDCl3, 400
MHz) δ 6.59 (2H, s, H-2, H-C6), 6.52 (1H, d, J ) 16.0, H-7),
6.27 (1H, dt, J ) 15.6, 5.7 Hz, H-8), 5.54 (1H, m, H-2′), 4.47
(2H, brd, J ) 7.1 Hz, H-1′), 4.30 (2H, brd, J ) 5.7 Hz, H-9),
3.84 (6H, s, OMe-3, OMe-5), 1.72 (3H, H-4′), 1.65 (3H, s, H-5′);
HREIMS m/z 278.1532 (calcd for C16H22O4, 278.1518).
4-O-(2-Meth yl-2-bu ten yl)sin apaldeh yde (35). The method
of preparation of 35 was similar to that used for the prepara-
1
tion of 7 from 6. The yield of 35 from 34 was 91%: gum; H
NMR (CDCl3, 400 MHz) δ 9.66 (1H, d, J ) 7.6 Hz, H-9), 7.40
(1H, d, J ) 15.9 Hz, H-7), 6.77 (2H, br. s, H-2, H-6), 6.60 (1H,
dd, J ) 15.9, 7.6 Hz, H-8), 5.54 (1H, brt, J ) 7.2 Hz, H-2′),
4.56 (2H, brd, J ) 7.2 Hz, H-1′), 3.89 (6H, s, OMe-3, OMe-5),
1.72 (3H, s, H-4′), 1.65 (3H, s, Me-5′); HREIMS m/z 276.1351
(calcd for C16H20O4, 276.1362).
4-O-(2-Meth yl-2-bu ten yl)-7,8-d ih yd r osin a p yl a lcoh ol
1
(36): gum; H NMR (CDCl3, 400 MHz) δ 6.48 (2H, brs, H-2,
H-6), 5.53 (1H, brt, J ) 7.2 Hz, H-2′), 4.51 (2H, brd, J ) 7.1
Hz, H-1′), 3.90 (2H, brt, J ) 7.5 Hz, H-9), 3.88 (6H, s, OMe-3,
OMe-5), 2.81 (2H, brt, J ) 7.5 Hz, H-7), 2.03-1.96 (2H, m,
H-8), 1.71 (3H, s, H-4′), 1.65 (3H, s, H-5′); HREIMS m/z
278.1509 (calcd for C16H22O4, 278.1518).
Cytotoxicity Assa y. KB cells were obtained from the
American type culture collection.12 Effects of compounds on
the growth of the cells were monitored at the Laboratoire de
Cultures Cellulaires, ICSN, Gif-sur-Yvette, France. The IC50
values refer to the concentration of drug corresponding to 50%
growth inhibition after 72 h incubation.13 The assays of A-549
and HL-60 were carried out at the Institute of Shanghai
Material Medica and were performed according to published
techniques.18-20
Ack n ow led gm en t. This work was financially supported
by the Life Science Special Fund of the Chinese Academy of
Sciences supported by the Ministry of Finance (STZ-00-24);
the Yunnan Province Foundation of Applied and Basic Re-
search (2000C0072M); the Foundation for Visiting Professor
from the State Key Laboratory of New Drug Research at
SIMM; a grant from the Dean of KIB, CAS; and the Chine-
France PRA (PRA BT01-02). One of the authors (Y.Z.) ex-
4-O-(2-Meth yl-2-bu ten yl)-3,5-d im eth oxyben zoic Acid
Meth yl Ester (30). The method of preparation of 30 was
similar to that used for the preparation of 15. The yield of 30
from 29 was 92%: gum; 1H NMR (CDCl3, 400 MHz) δ 7.26
(2H, s, H-2, H-6), 5.52 (1H, brt, J ) 7.2 Hz, H-2′), 4.55 (2H, d,
J ) 7.3 Hz, H-1′), 3.89 (3H, s, CO2Me), 3.88 (3H, s, OMe-3,

908 J ournal of Natural Products, 2002, Vol. 65, No. 6
Zhao et al.
(9) Fujita, M.; Yamada, M.; Nakajima, S.; Kawai, K.-I.; Nagai, M. Chem.
Pharm. Bull. 1984, 32, 2622-2627.
presses his thanks to the Chinese Ministry of Education as
well as to Mr. Ka-Shing Lee for a “Cheung Kong Scholar Chief
Professorship” at Zhejiang University, and the EGIDE founda-
tion (France) for a fellowship at ISCN, CNRS at Gif-sur-Yvette,
where this work was finally accomplished. We also thank
Christiane Gaspard for conducting the cytotoxicity assays.
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