Hydroxylated gedunin derivatives from Cedrela sinensis

Article abstract of DOI:10.1021/np068021f

Four new limonoids, 11α-hydroxygedunin (1), 11β-hydroxygedunin (2), 7-deacetoxy-7α,11α-dihydroxygedunin (3), and 7-deacetoxy-7α,11β-dihydroxygedunin (4), were isolated from the cortex of Cedrela sinensis, together with three known compounds, gedunin (5), 7-deacetoxy-7α-hydroxygedunin (6), and 11-oxogedunin (7). The structures of 1-4 were determined by a combination of 2D NMR experiments and chemical methods and by X-ray crystallography of 1 and 2.

Full text of DOI:10.1021/np068021f

1310
J. Nat. Prod. 2006, 69, 1310-1314
Hydroxylated Gedunin Derivatives from Cedrela sinensis
Kumiko Mitsui, Hiroaki Saito, Ryota Yamamura, Haruhiko Fukaya, Yukio Hitotsuyanagi, and Koichi Takeya*
School of Pharmacy, Tokyo UniVersity of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo, 192-0392, Japan
ReceiVed May 10, 2006
Four new limonoids, 11R-hydroxygedunin (1), 11â-hydroxygedunin (2), 7-deacetoxy-7R,11R-dihydroxygedunin (3),
and 7-deacetoxy-7R,11â-dihydroxygedunin (4), were isolated from the cortex of Cedrela sinensis, together with three
known compounds, gedunin (5), 7-deacetoxy-7R-hydroxygedunin (6), and 11-oxogedunin (7). The structures of 1-4
were determined by a combination of 2D NMR experiments and chemical methods and by X-ray crystallography of 1
and 2.
Cedrela sinensis Juss. (Meliaceae) has been used in mainland
China and Korea as a traditional medicine for the treatment of
enteritis, dysentery, and itching. Previously, we reported isolation
of five obacunone-type limonoids1 and 23 apotirucallane-type
triterpenoids2 from the seeds, leaves, and stems of this plant. In
the present study, we have isolated four new limonoids (1-4) along
with three known limonoids, from the cortex of C. sinensis and
have determined the structures of the new limonoids to be 11R-
hydroxygedunin (1), 11â-hydroxygedunin (2), 7-deacetoxy-7R,11R-
dihydroxygedunin (3), and 7-deacetoxy-7R,11â-dihydroxygedunin
(4). The three known limonoids were shown to be gedunin (5),
7-deacetoxy-7R-hydroxygedunin (6), and 11-oxogedunin (7).
at m/z 499.2293 (calcd for C28H35O8, 499.2332) in the HRESIMS.
Its NMR spectra generally resembled those of gedunin (5),
1
suggesting that 1 has a gedunin-type limonoid skeleton.3 The H
NMR spectrum of 1 showed the presence of five tertiary methyl
groups (δ 1.06, 1.08, 1.17, 1.35, and 1.38), one acetate methyl group
(δ 2.12, s), four oxymethine protons (δ 3.57, 4.45, 4.56, and 5.59),
and a â-substituted furan ring (δ 6.36, 7.42, and 7.42, 1H each)
(Table 1). The 13C NMR spectrum indicated the presence of six
methyls, two methylenes, 11 methines, and nine quaternary carbons,
of which two at δ 167.4 and 169.9 were assigned to ester carbonyl
carbons (Table 2). In the HMBC spectrum, the cross-peak between
δC 169.9 and δH 4.45 (H-7) demonstrated that the acetoxy group is
attached to C-7 (Figure 1). The 13C NMR signals of C-1 (δ 161.4),
C-2 (δ 123.9), and C-3 (δ 204.1) and the 1H NMR signal of a pair
of AB doublets at δ 5.79 and 8.18 (J ) 10.5 Hz) suggested that
the A-ring of 1 possesses a 1-en-3-one system. The cross-peaks
between H-15/C-14, H-15/C-16, H-17/C-13, and H-17/C-14 in the
HMBC spectrum suggested the presence of a δ-lactone group with
a 14,15-epoxide in the D-ring. The C-11 signal of 1 appeared at δ
65.7 (d), implying that it is an oxymethine carbon. TPAP oxidation4
of 1 afforded 11-oxogedunin (7), whereas acetylation of 1 afforded
diacetate 1a, which confirmed the presence of a hydroxyl group at
C-11 in 1. As regards the relative stereochemistry of 1, NOE
correlations detected between H-5/H-9, H-5/H3-28, H-7/H3-30, H-9/
H3-18, H-11/H3-19, H-11/H3-30, H-15/H3-18, H3-18/H-21, H3-18/
H-22, H3-19/H3-29, and H3-19/H3-30 showed that H-5, OAc-7, H-9,
OH-11, Me-18, and Me-28 are R-oriented, whereas the 14,15-
epoxide, H-17, Me-19, Me-29, and Me-30 groups are â-oriented
(Figure 2). From these observations, limonoid 1 was determined
to be 11R-hydroxygedunin. This structure was confirmed by X-ray
crystallographic analysis (Figure 3).
Limonoid 2 was obtained as colorless prisms. Its molecular
formula, C28H34O8, as determined from the [M + H]+ peak at m/z
499.2312 (calcd for C28H35O8, 499.2332) in the HRESIMS, was
the same as that of 1, and the COSY and HMBC spectra were
quite similar to those of 1. The TPAP oxidation product of 2 was
11-oxogedunin (7), demonstrating that 1 and 2 have the same gross
structure. The NOE correlation detected between H-11 and H3-18
and the coupling constant of 4.6 Hz between H-9 and H-11 implied
that the hydroxyl group at C-11 in 2 is â-oriented (Figure 4).
Acetylation of 2 afforded a product, 2a, for which the spectroscopic
data were identical to those of the known compound 11â-
acetoxygedunin.5 Thus, 2 was determined to be 11â-hydroxyge-
dunin. This structure was also confirmed by X-ray crystallographic
analysis (Figure 5).
Limonoid 3 was obtained as an amorphous solid. From the [M
+ H]+ peak at m/z 457.2224 (calcd for C26H33O7, 457.2226) in the
HRESIMS, its molecular formula was determined to be C26H32O7.
The 1H and 13C NMR spectra of 3 showed a close resemblance to
those of 1, implying that both compounds are of the same basic
structure. The differences noted between the NMR spectra of these
Results and Discussion
By a series of separation steps including Diaion HP-20, activated
charcoal, and ODS column chromatography, and subsequent
purification by preparative HPLC, a MeOH extract of the cortex
of C. sinensis afforded four new limonoids, 1-4, along with three
known compounds, 5-7.
Limonoid 1 was isolated as colorless prisms. Its molecular
formula was determined to be C28H34O8 from the [M + H]+ peak
* Corresponding author. Tel: +81-42-676-3007. Fax: +81-42-677-1436.
E-mail: takeyak@ps.toyaku.ac.jp.
10.1021/np068021f CCC: $33.50
© 2006 American Chemical Society and American Society of Pharmacognosy
Published on Web 09/02/2006

Hydroxylated Gedunin DeriVatiVes from Cedrela sinensis
Journal of Natural Products, 2006, Vol. 69, No. 9 1311
Table 1. 1H NMR (500 MHz) Spectroscopic Data for Limonoids 1-4 in CDCl3
a
position
1
2
3
4
1
2
5
6R
6â
7
8.18 (d, 10.5)
5.79 (d, 10.5)
2.25 (m)
1.95 (dt, 15.0, 2.9)
1.76 (td, 15.0, 1.4)
4.45 (d, 1.4)
2.62 (d, 9.6)
4.56 (m)
1.51 (d, 14.6)
2.25 (m)
3.57 (s)
5.59 (s)
1.38 (s)
1.35 (s)
7.42 (s)
6.36 (s)
7.42 (s)
1.06 (s)
1.08 (s)
1.17 (s)
2.12 (s)
7.39 (d, 10.2)
5.91 (d, 10.2)
2.12 (m)
1.94 (m)
1.94 (m)
4.54 (t-like, 2.3)
2.35 (d, 4.6)
4.82 (m)
2.13 (m)
1.70 (m)
3.57 (s)
5.66 (s)
1.19 (s)
1.63 (s)
7.42 (s)
6.36 (s)
7.42 (s)
1.07 (s)
1.07 (s)
8.18 (d, 10.5)
5.78 (d, 10.5)
2.55 (dd, 13.4, 2.4)
1.67 (dt, 14.6, 3.0)
1.89 (td, 14.6, 1.5)
3.49 (br s)
2.63 (d, 9.7)
4.54 (m)
1.47 (d, 14.6)
2.23 (dd, 14.6, 6.9)
3.99 (s)
5.57 (s)
1.36 (s)
1.32 (s)
7.42 (s)
6.36 (s)
7.42 (s)
1.15 (s)
1.11 (s)
1.10 (s)
7.38 (d, 10.3)
5.90 (d, 10.3)
2.42 (dd, 13.2, 2.1)
1.67 (d-like, 13.7)
2.05 (td, 13.7, 1.7)
3.54 (d, 1.7)
2.39 (d, 4.7)
4.80 (m)
2.13 (dd, 13.4, 9.3)
1.68 (m)
3.84 (s)
5.65 (s)
1.19 (s)
1.60 (s)
7.41 (s)
6.36 (s)
7.42 (s)
1.15 (s)
1.10 (s)
1.31 (s)
9
11
12R
12â
15
17
18
19
21
22
23
28
29
30
OAc-7
1.38 (s)
2.07 (s)
a Chemical shifts are reported in ppm relative to residual CHCl3 resonance at 7.26 ppm. Multiplicity and J values in Hz are given in parentheses.
Table 2. 13C NMR (125 MHz) Spectroscopic Data for
a
Limonoids 1-4 in CDCl3
position
1
2
3
4
1
2
3
4
5
6
7
8
161.4
123.9
204.1
44.2
45.3
22.8
72.9
42.4
46.7
41.0
65.7
39.8
38.6
70.1
57.7
167.4
77.7
17.0
21.0
120.2
141.2
109.9
143.3
27.2
21.3
19.9
169.9
21.0
156.7
125.9
203.8
43.9
47.1
23.8
74.5
42.5
44.2
41.3
64.9
39.5
38.2
69.3
56.0
167.3
78.0
16.6
22.6
120.2
141.3
109.9
143.2
26.9
21.4
20.5
169.7
21.1
161.9
123.9
204.5
44.4
44.0
26.9
69.2
43.5
45.5
41.2
66.2
40.1
38.3
70.2
59.1
168.1
77.8
17.3
21.1
120.5
141.2
110.0
143.2
27.5
21.6
20.3
157.2
125.8
204.2
44.1
45.7
28.0
71.5
43.5
42.7
41.4
65.3
40.2
37.7
69.5
56.6
167.9
78.3
16.8
22.8
120.5
141.3
110.1
143.1
27.1
21.6
20.8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
28
29
30
Figure 1. Selected HMBC correlations for 1.
OAc-7 (CdO)
OAc-7 (Me)
a Chemical shifts are reported in ppm relative to the solvent resonance
at 77.03 ppm.
limonoids were that 3 had no signal due to an acetyl group and
that the C-7 signal of 3 (δ 69.2) was at a higher field than the
corresponding C-7 signal of 1 (δ 72.9), demonstrating that 3 has
no acetyl group at C-7. Deacetylation of 1 afforded a product that
was shown to be identical to natural 3 from its spectoscopic data.
From these facts, 3 was determined as 7-deacetoxy-7R,11R-
dihydroxygedunin.
Limonoid 4 was obtained as an amorphous solid. Its molecular
formula was determined to be C26H32O7 from the [M + H]+ peak
at m/z 457.2194 (calcd for C26H33O7, 457.2226) in the HRESIMS.
The NMR spectra of 4 were generally similar to those of 3,
suggesting that they have the same basic structure. The spectro-
scopic differences between 3 and 4 were analogous to those
observed between 1 and 2 (Tables 1 and 2). Deacetylation of 2
afforded a product whose spectroscopic data were shown to be
Figure 2. Selected NOE correlations for 1.
identical to those of natural 4. Thus, the structure of 4 was
determined to be 7-deacetoxy-7R,11â-dihydroxygedunin.
Compounds 5-7 were identified as gedunin,3,6 7-deacetoxy-7R-
hydroxygedunin,7 and 11-oxogedunin,8 respectively, by analysis of
their spectroscopic data. Full 1H and 13C NMR data for the known
compounds 6 and 7 are provided in the Experimental Section.
Compounds 2, 2a, and 4-7 all showed moderate cytotoxicity
against P-388 murine leukemia cells, with IC50 values of 5.4, 3.8,
7.8, 3.3, 4.5, and 3.0 µg/mL, respectively, whereas compounds 1,
1a, and 3 had very weak or no activity, with IC50 values of 71, 63,
and >100 µg/mL, respectively, suggesting that an oxygenated
functionality at the 11R-position has a detrimental effect on the
resultant cytotoxic activity.

1312 Journal of Natural Products, 2006, Vol. 69, No. 9
Mitsui et al.
Figure 3. Crystal structure of 1.
80% MeOH fraction (198 g) was subjected to activated charcoal (450
g) column chromatography, eluting with MeOH, CHCl3-MeOH (1:
9), and CHCl3-MeOH (1:1), with 30 L of each solvent used. The
CHCl3-MeOH (1:9) fraction (10 g) was further subjected to RP-18
(100 g) column chromatography, eluting with MeOH-H2O (60:40, 80:
20, 100:0, each 1 L), to afford three fractions (1-3). After removal of
the solvent to dryness, fraction 1 (3.0 g) was subjected to HPLC using
MeOH-H2O (55:45) to afford two limonoid-containing fractions, each
of which was subsequently purified by HPLC using MeCN-H2O (35:
65) to give 3 (1.5 mg) and 4 (1.2 mg), respectively. Fraction 2 (4.0 g)
was subjected to HPLC using MeOH-H2O (60:40) to give four
limonoid-containing fractions. When subsequently purified by HPLC
using MeCN-H2O (40:60), the first limonoid-containing fraction gave
1 (191.8 mg) and 2 (42.7 mg), the second limonoid-containing fraction
afforded 11-oxogedunin (7) (28.2 mg), and the third fraction yielded
7-deacetoxy-7R-hydroxygedunin (6) (66.0 mg). Fraction 3 (3.0 g) was
subjected to HPLC using MeOH-H2O (75:35) to give a limonoid-
containing fraction, which was then purified by HPLC using MeCN-
H2O (43:57) to give gedunin (5) (286.7 mg).
Figure 4. Selected NOE correlations for 2.
Gedunin (5) is reported to have in vitro antimalarial activity
against chloroquine-resistant K1 strain of Plasmodium falciparum9
and thus has been regarded as a possible lead for new antimalarial
drugs. Compounds 1-4, having the same basic structure with a
hydroxyl group at C-11, may be useful as a starting material for
the preparation of new analogues of gedunin-related antimalarial
drugs.
11R-Hydroxygedunin (1): colorless prisms (CHCl3-MeOH); mp
240-243 °C; [R]25 +18.5 (c 0.3, CHCl3); UV (MeOH) λmax (log ꢀ)
D
215 (4.20) nm; IR (film) νmax 3650, 2965, 1795, 1701, 1679, 1651,
1539 cm-1 1H and 13C NMR, see Tables 1 and 2; HRESIMS m/z
;
499.2293 ([M + H]+, calcd for C28H35O8, 499.2332).
11â-Hydroxygedunin (2): colorless prisms (CHCl3-MeOH); mp
294-297 °C; [R]25 +7.8 (c 0.1, CHCl3); UV (MeOH) λmax nm (log
D
Experimental Section
ꢀ) 216 (4.20); IR (film) νmax 3650, 2950, 1795, 1700, 1650, 1539 cm-1
;
General Experimental Procedures. Optical rotations were mea-
sured on a JASCO P-1030 digital polarimeter. UV spectra were taken
on a JASCO V-530 spectrophotometer, IR spectra on a JASCO FT/IR
620 spectrophotometer, NMR spectra on a Bruker DRX-500 spectrom-
eter at 300 K, and mass spectra on a Micromass LCT spectrometer.
Preparative HPLC was performed on a Shimadzu LC-6AD system
equipped with a SPD-10A UV detector (at 205 nm) and a reversed-
phase column, Wakosil-II 5C18HG prep (5 µm, 20 × 250 mm), using
mixed solvent systems of MeOH-H2O or MeCN-H2O, at a flow late
of 10 mL/min. Single-crystal X-ray analysis was carried out on a Mac
Science DIP diffractometer with Mo KR radiation (λ ) 0.71073 Å).
Plant Material. The cortex of C. sinensis was collected in Jilin
Province, People’s Republic of China, in September 2000, and the
botanical origin was identified by Professor Soo-Cheol Kim of the
Agricultural College of Yanbian University. A voucher specimen
(00CHI005) has been deposited in the herbarium of Tokyo University
of Pharmacy and Life Science.
1H and 13C NMR, see Tables 1 and 2; HRESIMS m/z 499.2312 ([M +
H]+, calcd for C28H35O8, 499.2332).
7-Deacetoxy-7R,11R-dihydroxygedunin (3): amorphous solid; [R]25
D
+54.5 (c 0.06, CHCl3); UV (MeOH) λmax (log ꢀ) 215 (4.14) nm; IR
(film) νmax 3650, 2946, 1795, 1737, 1650, 1539 cm-1; 1H and 13C NMR,
see Tables 1 and 2; HRESIMS m/z 457.2224 ([M + H]+, calcd for
C26H33O7, 457.2226).
7-Deacetoxy-7R,11â-dihydroxygedunin (4): amorphous solid; [R]25
D
+47.0 (c 0.04, CHCl3); UV (MeOH) λmax (log ꢀ) 215 (4.12) nm; IR
(film) νmax 3650, 2931, 1795, 1737, 1650, 1539 cm-1; 1H and 13C NMR,
see Tables 1 and 2; HRESIMS m/z 457.2194 ([M + H]+, calcd for
C26H33O7, 457.2226).
Gedunin (5): colorless crystalline powder (CHCl3-MeOH); mp
194-198 °C; [R]27 +42.8 (c 0.1, CHCl3); HRESIMS m/z 483.2386
D
([M + H]+, calcd for C28H35O7, 483.2383).
7-Deacetoxy-7R-hydroxygedunin (6): colorless crystalline powder
1
(CHCl3-MeOH); mp 270-275 °C; [R]28 +60.7 (c 0.1, CHCl3); H
D
Extraction and Isolation. The cut and air-dried cortex of C. sinensis
(24 kg) was extracted with hot MeOH (3 × 90 L). After the removal
of the solvent, the MeOH extract (3.2 kg) was placed on a column of
HP-20 (8.0 kg) and fractionated into five fractions, by eluting with
H2O, 50% MeOH, 80% MeOH, MeOH, and acetone (each 90 L). The
NMR (CDCl3, 500 MHz) δ 7.41 (1H, s, H-21), 7.40 (1H, d, J ) 1.3
Hz, H-23), 7.10 (1H, d, J ) 10.2 Hz, H-1), 6.35 (1H, s, H-22), 5.84
(1H, d, J ) 10.2 Hz, H-2), 5.60 (1H, s, H-17), 3.91 (1H, s, H-15),
3.57 (1H, br s, H-7), 2.53 (1H, m, H-9), 2.48 (1H, m, H-5), 1.97 (1H,
m, H-11R), 1.96 (1H, m, H-6â), 1.80 (1H, m, H-11â), 1.72 (1H, m,

Hydroxylated Gedunin DeriVatiVes from Cedrela sinensis
Journal of Natural Products, 2006, Vol. 69, No. 9 1313
Figure 5. Crystal structure of 2.
H-12â), 1.63 (1H, m, H-6R), 1.55 (1H, m, H-12R), 1.23 (3H, s, H3-
18), 1.20 (3H, s, H3-19), 1.14 (3H, s, H3-28), 1.09 (6H, s, H3-29 and
H3-30); 13C NMR (CDCl3, 125 MHz) δ 204.6 (C, C-3), 168.2 (C, C-16),
157.7 (CH, C-1), 143.0 (CH, C-23), 141.2 (CH, C-21), 125.8 (CH,
C-2), 120.7 (C, C-20), 110.0 (CH, C-22), 78.5 (CH, C-17), 70.0 (C,
C-14), 69.8 (CH, C-7), 57.9 (CH, C-15), 44.6 (CH, C-5), 44.2 (C, C-4),
43.7 (C, C-8), 40.2 (C, C-10), 38.4 (C, C-13), 38.0 (CH, C-9), 27.3
(CH3, C-28), 27.3 (CH2, C-6), 26.4 (CH2, C-12), 21.5 (CH3, C-29),
19.9 (CH3, C-19), 18.7 (CH3, C-30), 17.8 (CH3, C-18), 15.1 (CH2,
C-11); HRESIMS m/z 441.2316 ([M + H]+, calcd for C26H33O6,
441.2277).
11-Oxogedunin (7): amorphous solid; [R]28D -38.1 (c 0.1, CHCl3);
1H NMR (CDCl3, 500 MHz) δ 7.45 (1H, s, H-21), 7.44 (1H, s, H-23),
7.31 (1H, d, J ) 10.2 Hz, H-1), 6.35 (1H, s, H-22), 5.85 (1H, d, J )
10.2 Hz, H-2), 5.68 (1H, s, H-17), 4.68 (1H, t-like, J ) 2.5 Hz, H-7),
3.66 (1H, s, H-15), 3.35 (1H, s, H-9), 2.47 (1H, d, J ) 19.0 Hz, H-12R),
2.30 (1H, d, J ) 19.0 Hz, H-12â), 2.14 (3H, s, OAc-7), 2.07 (1H, dd,
J ) 13.3, 2.0 Hz, H-5), 1.97 (1H, m, H-6R), 1.84 (1H, ddd, J ) 14.9,
13.3, 1.9 Hz, H-6â), 1.47 (3H, s, H3-19), 1.45 (3H, s, H3-18), 1.17
(3H, s, H3-30), 1.08 (3H, s, H3-28), 1.07 (3H, s, H3-29); 13C NMR
(CDCl3, 125 MHz) δ 205.5 (C, C-11), 203.2 (C, C-3), 169.4 (C,
OCOCH3-7), 166.3 (C, C-16), 158.0 (CH, C-1), 143.7 (CH, C-23), 141.3
(CH, C-21), 125.9 (CH, C-2), 119.4 (C, C-20), 109.4 (CH, C-22), 77.1
(CH, C-17), 72.8 (CH, C-7), 68.5 (C, C-14), 56.0 (CH, C-9), 55.6 (CH,
C-15), 45.8 (CH2, C-12), 45.5 (CH, C-5), 44.1 (C, C-4), 43.6 (C, C-8),
38.9 (C, 2C, C-10 and C-13), 27.1 (CH3, C-28), 23.4 (CH2, C-6), 21.2
(CH3, C-29), 21.1 (CH3, C-19), 21.0 (CH3, OCOCH3-7), 20.2 (CH3,
C-30), 18.5 (CH3, C-18); HRESIMS m/z 497.2187 ([M + H]+, calcd
for C28H33O8, 497.2175).
Oxidation of 1. Solid TPAP (tetrapropylammonium per-ruthenate,
0.6 mg, 0.0017 mmol) was added in one portion to a stirred mixture of
1 (9.0 mg, 0.018 mmol), 4-methylmorpholine N-oxide (3.2 mg, 0.027
mmol), and powdered 4 Å molecular sieves (9.0 mg) in CH2Cl2 (2
mL), and the whole was stirred at room temperature for 7 h. The mixture
was diluted with CHCl3 and washed sequentially with aqueous Na2-
SO3 and brine. The organic layer was dried (MgSO4), filtered, and
concentrated. The residue was subjected to ODS-HPLC with MeCN-
H2O (43:57) to afford an oxidation product (8.5 mg, 95%), which was
identified as 11-oxogedunin (7) by comparison of their NMR and mass
spectra.
Oxidation of 2. TPAP oxidation of 2 (2.2 mg) by the procedure
described above gave an oxidation product (2.0 mg, 91%), which was
shown to be identical to 11-oxogedunin (7) by comparison of their
NMR and mass spectra.
Acetylation of 1. Acetic anhydride (1 mL) was added to a solution
of 1 (7.0 mg) in pyridine (1 mL), and the mixture was left at room
temperature for 24 h. After removal of the volatiles in vacuo, the residue
was subjected to ODS-HPLC with MeOH-H2O (60:40) to afford 1a
(6.5 mg, 86%) as an amorphous solid: [R]25D +6.1 (c 0.3, CHCl3); 1H
NMR (CDCl3, 500 MHz) δ 7.41 (2H, s, H-21 and H-23), 7.36 (1H, d,
J ) 10.5 Hz, H-1), 6.32 (1H, s, H-22), 5.81 (1H, d, J ) 10.5 Hz, H-2),
5.59 (1H, s, H-17), 5.55 (1H, t-like, J ) 9.1 Hz, H-11), 4.49 (1H, br
s, H-7), 3.59 (1H, s, H-15), 2.87 (1H, d, J ) 10.6 Hz, H-9), 2.30 (1H,
dd, J ) 15.3, 7.5 Hz, H-12â), 2.24 (1H, m, H-5), 2.13 (3H, s, OAc-7),
2.06 (3H, s, OAc-11), 1.98 (1H, d-like, J ) 14.6 Hz, H-6R), 1.78 (1H,
t-like, J ) 14.6 Hz, H-6â), 1.46 (1H, d, J ) 15.3 Hz, H-12R), 1.33
(3H, s, H3-18), 1.29 (3H, s, H3-19), 1.22 (3H, s, H3-30), 1.09 (3H, s,
H3-29), 1.06 (3H, s, H3-28); 13C NMR (CDCl3, 125 MHz) δ 203.4 (C,
C-3), 169.8 (C, 2C, OCOCH3-7 and -11), 167.0 (C, C-16), 158.3 (CH,
C-1), 143.4 (CH, C-23), 141.3 (CH, C-21), 124.9 (CH, C-2), 120.0
(C, C-20), 109.6 (CH, C-22), 77.5 (CH, C-17), 72.7 (CH, C-7), 69.7
(C, C-14), 68.5 (CH, C-11), 57.6 (CH, C-15), 45.1 (CH, C-5), 44.4
(C, C-4), 43.9 (CH, C-9), 42.4 (C, C-8), 40.8 (C, C-10), 38.3 (C, C-13),
36.6 (CH2, C-12), 27.3 (CH3, C-28), 22.7 (CH2, C-6), 21.7 (CH3,
OCOCH3-11), 21.2 (CH3, C-29), 21.0 (CH3, OCOCH3-7), 20.9 (CH3,
C-19), 19.9 (CH3, C-30), 17.4 (CH3, C-18); HRESIMS m/z 541.2410
([M + H]+, calcd for C30H37O9, 541.2438).
Acetylation of 2. Acetylation of 2 (3.0 mg) by the procedure
described above gave 2a (2.8 mg, 86%) as an amorphous solid, which
1
was identified as 11â-acetoxygedunin by comparing its H NMR and
mass spectra and optical rotations with those in the literature:5 [R]25
D
1
+32.9 (c 0.2, CHCl3); H NMR (CDCl3, 500 MHz) δ 7.41 (1H, s,
H-23), 7.40 (1H, s, H-21), 7.12 (1H, d, J ) 10.3 Hz, H-1), 6.33 (1H,
s, H-22), 5.91 (1H, d, J ) 10.3 Hz, H-2), 5.80 (1H, m, H-11), 5.61
(1H, s, H-17), 4.59 (1H, br s, H-7), 3.59 (1H, s, H-15), 2.54 (1H, d, J
) 4.8 Hz, H-9), 2.32 (1H, dd, J ) 14.1, 9.8 Hz, H-12R), 2.14 (1H, m,
H-5), 2.14 (3H, s, OAc-11), 2.08 (3H, s, OAc-7), 1.94 (2H, m, H-6R
and H-6â), 1.54 (1H, m, H-12â), 1.45 (3H, s, H3-19), 1.37 (3H, s,
H3-30), 1.24 (3H, s, H3-18), 1.07 (3H, s, H3-28), 1.06 (3H, s, H3-29);
13C NMR (CDCl3, 125 MHz) δ 203.2 (C, C-3), 169.9 (C, OCOCH3-
11), 169.6 (C, OCOCH3-7), 166.9 (C, C-16), 155.0 (CH, C-1), 143.4
(CH, C-23), 141.3 (CH, C-21), 126.5 (CH, C-2), 119.9 (C, C-20), 109.8
(CH, C-22), 78.0 (CH, C-17), 74.3 (CH, C-7), 68.9 (C, C-14), 66.4
(CH, C-11), 55.7 (CH, C-15), 47.2 (CH, C-5), 43.9 (C, C-4), 43.4 (CH,
C-9), 42.4 (C, C-8), 40.7 (C, C-10), 37.8 (C, C-13), 36.1 (CH2, C-12),
27.0 (CH3, C-28), 23.8 (CH2, C-6), 21.8 (CH3, C-19), 21.6 (CH3,
OCOCH3-11), 21.4 (CH3, C-29), 21.1 (CH3, OCOCH3-7), 20.2 (CH3,
C-30), 16.9 (CH3, C-18); HRESIMS m/z 541.2421 ([M + H]+, calcd
for C30H37O9, 541.2438).
Deacetylation of 1. Potassium carbonate (1.0 mg) was added to a
solution of 1 (2.0 mg) in MeOH (1 mL), and the mixture was stirred
at room temperature for 24 h. The mixture was diluted with CHCl3,
washed sequentially with H2O and brine, dried (Na2SO4), filtered, and
concentrated under reduced pressure. The residue was purified by ODS-
HPLC using MeCN-H2O (40:60) to give a product (1.6 mg, 87%)
that was shown to be identical to natural 3 by comparison of their NMR
and mass spectra.
Deacetylation of 2. Deacetylation of 2 (1.1 mg) by the procedure
described above gave a product (0.9 mg, 89%) whose NMR and mass
spectra showed that the product was identical to natural 4.
X-ray Crystallographic Study of 1. C28H34O8, M ) 498.55, 0.50
× 0.30 × 0.30 mm, orthorhombic, P212121, a ) 11.7970(4) Å, b )

1314 Journal of Natural Products, 2006, Vol. 69, No. 9
Mitsui et al.
12.0160(4) Å, c ) 34.2630(5) Å, V ) 4856.9(2) Å,3 Z ) 8, Dx )
1.364 Mg m-3, µ(Mo KR) ) 0.099 mm-1, 5911 measured reflections,
5911 unique reflections, 4698 observed reflections [I > 2σ(I)], R1 )
0.0434, wR2 ) 0.1026 (observed data), GOF ) 0.913; R1 ) 0.0522,
wR2 ) 0.1053 (all data). The absolute structure could not be determined
crystallographically.
X-ray Crystallographic Study of 2. C28H34O8, M ) 498.55, 0.50
× 0.44 × 0.20 mm, monoclinic, P21, a ) 9.6670(11) Å, b ) 12.593-
(2) Å, c ) 10.2860(16) Å, â ) 103.447(9)°, V ) 1217.9(3) Å,3 Z )
2, Dx ) 1.360 Mg m-3, µ(Mo KR) ) 0.099 mm-1, 2791 measured
reflections, 2791 unique reflections, 2019 observed reflections [I >
2σ(I)], R1 ) 0.0411, wR2 ) 0.0954 (observed data), GOF ) 0.912;
R1 ) 0.0561, wR2 ) 0.0995 (all data). The absolute structure could
not be determined crystallographically.
References and Notes
(1) Mitsui, K.; Maejima, M.; Fukaya, H.; Hitotsuyanagi, Y.; Takeya, K.
Phytochemistry 2004, 65, 3075-3081.
(2) Mitsui, K.; Maejima, M.; Saito, H.; Fukaya, H.; Hitotsuyanagi, Y.;
Takeya, K. Tetrahedron 2005, 61, 10569-10582.
(3) Khalid, S. A.; Duddeck, H.; Gonzalez-Sierra, M. J. Nat. Prod. 1989,
52, 922-926.
(4) Ley, S. V.; Norman, J.; Griffith, W. P.; Marsden, S. P. Synthesis
1994, 639-666.
(5) Connolly, J. D.; McCrindle, R.; Overton, K. H.; Feeney, J. Tetra-
hedron 1966, 22, 891-896.
(6) Akisanya, A.; Bevan, C. W. L.; Hirst, J.; Halsall, T. G.; Taylar, D.
A. H. J. Chem. Soc. 1960, 3827-3829.
(7) Lavie, D.; Levy, E. C.; Jain, M. K. Tetrahedron 1971, 27, 3927-
3939.
(8) Taylor, D. A. H. J. Chem. Soc. C 1970, 336-340.
(9) Bray, D. H.; Warhurst, D. C.; Connolly, J. D.; O’Neill, M. J.;
Phillipson, J. D. Phytother. Res. 1990, 4, 29-35.
The structures were solved by direct methods using the maXus
crystallographic software package10 and refined by full-matrix least-
squares on F2 using the program SHELXL-97.11
CCDC 604386 and 604387 contain the supplementary crystal-
lographic data for compounds 1 and 2, respectively, from this paper.
These data can be obtained free of charge via http://www.ccdc.ca-
m.ac.uk/data_request/cif, by e-mailing data_request@ccdc.cam.ac.uk,
or by contacting The Cambridge Crystallographic Data Centre, 12 Union
Road, Cambridge CB2 1EZ, UK; fax: +44 1223 336033.
(10) Mackay, S.; Gilmore, C. J.; Edwards, C.; Stewart, N.; Shankland,
K. MaXus: Computer Program for the Solution and Refinement of
Crystal Structures; Bruker Nonius/MacScience/The University of
Glasgow: The Netherlands/Japan/The University of Glasgow, 1999.
(11) Sheldrick, G. M. SHELXL97: Program for the Refinement of Crystal
Structures; University of Go¨ttingen: Go¨ttingen, Germany, 1997.
(12) Kim, I. H.; Takashima, S.; Hitotsuyanagi, Y.; Hasuda, T.; Takeya,
K. J. Nat. Prod. 2004, 67, 863-868.
Cytotoxicity Assays. Evaluation of cytotoxicity against P-388 murine
leukemia cells was performed as described previously.12
NP068021F