Tirucallane triterpenoids from cornus walteri

Article abstract of DOI:10.1021/np100660v

Twelve new tirucallane triterpenoids, named cornusalterins A-L (1-12), and two known tirucallane triterpenoids, deoxyflindissone (13) and (-)-leucophyllone (14), were isolated from a MeOH extract of stems and stem bark of Cornus walteri. The structures of the new compounds were determined by spectroscopic methods, including 1D and 2D NMR analyses. Compounds 12 and 13, possessing a tetrahydrofuran ring in the side chain, exhibited significant cytotoxic activity against the A549, SK-OV-3, SK-MEL-2, and XF498 cell lines.

Full text of DOI:10.1021/np100660v

54
J. Nat. Prod. 2011, 74, 54–59
Tirucallane Triterpenoids from Cornus walteri
Ki Hyun Kim,† Sang Un Choi,‡ Young Choong Kim,§ and Kang Ro Lee*,†
Natural Products Laboratory, School of Pharmacy, Sungkyunkwan UniVersity, Suwon 440-746, Korea, Korea Research Institute of Chemical
Technology, Teajeon 305-600, Korea, and College of Pharmacy, Seoul National UniVersity, Seoul 151-742, Korea
ReceiVed September 16, 2010
Twelve new tirucallane triterpenoids, named cornusalterins A-L (1-12), and two known tirucallane triterpenoids,
deoxyflindissone (13) and (-)-leucophyllone (14), were isolated from a MeOH extract of stems and stem bark of Cornus
walteri. The structures of the new compounds were determined by spectroscopic methods, including 1D and 2D NMR
analyses. Compounds 12 and 13, possessing a tetrahydrofuran ring in the side chain, exhibited significant cytotoxic
activity against the A549, SK-OV-3, SK-MEL-2, and XF498 cell lines.
The genus Cornus comprises a group of 30-50 species of mostly
deciduous trees and shrubs that contain a variety of cytotoxic
constituents such as iridoid glycosides, polyphenols, triterpenoids,
and lignans from the fruits of Cornus kousa1-4 and the benzofura-
none derivative halleridone from Cornus controVersa.5 Cornus
walteri Wanger (Cornaceae) is a deciduous shrub that grows in
valley areas of Asia, particularly China and Korea.6 The fruits and
leaves are used to treat inflammation of the skin or boils caused by
lacquer poison in Chinese folk medicine.7 In Korean folk medicine,
the leaves have been used as an antidiarrheal.8 Previous phy-
tochemical investigations on C. walteri led to the isolation of gallic
acid and flavonoids.7,8 Extracts of C. walteri inhibited NO
production in lipopolysaccharide (LPS)-activated macrophages9 and
were reported to have elastase and tyrosinase inhibitory activity10
and antihyperglycemic and antiobesity effects.11
In our search for bioactive constituents from Korean medicinal
plants, we investigated a methanol extract of stems and stem bark
of C. walteri as the extract showed considerable cytotoxicity against
A549, SK-OV-3, and SK-MEL-2 cells. Bioassay-guided fraction-
ation of the MeOH extract resulted in the isolation of 12 new
tirucallane triterpenoids named cornusalterins A-L (1-12) and the
known tirucallane triterpenoids (13 and 14). Their structures were
determined using spectroscopic data, including 1D and 2D NMR.
Compounds 1-14 were evaluated for cytotoxic activity against four
human cancer cell lines. We report herein the isolation, structural
elucidation (1-12), and cytotoxicity of all of the isolated com-
pounds.
50.4, eight methylenes, five methines, including an oxygenated
methine at δC 79.4, and five quaternary carbons, including an
oxygenated quaternary carbon at δC 75.0. Comparison of its NMR
data with those of (-)-leucophyllone (14),12 and the fact that
tirucallane triterpenoids were previously isolated from this plant
genus,13 indicated that compound 1 was an analogue of 14. The
CD spectrum of 1 showed a negative (λmax 294 nm) Cotton effect
identical to that of 14, which was identical to the negative Cotton
effect of piscidinol A, a tirucallane triterpenoid.14 The NMR data
of 1 and 14 revealed that they possessed the same tetracyclic A-D
rings, and the only difference was the signal at C-3 in the NMR
data [δH 3.25 (1H, dd, J ) 11.5, 4.0 Hz); δC 79.4 in 1; δC 216.9 in
14]. This was confirmed by the HMBC experiment showing
correlations from the oxymethine proton (δH 3.25) to C-1, C-5,
C-28, and C-29. The HMBC spectrum indicated the presence of
an OCH3 group at C-25. The ꢀ-orientation of OH-3 was deduced
from correlations of H-3/H-28 and H-3/H-5 in the NOSEY
spectrum.
Cornusalterin B (2) had the molecular formula C31H52O2 by
HRFABMS. Inspection of the 1H and 13C NMR data of 2 revealed
that these data were again very similar to those of 14,12 except for
the chemical shifts of a methylene [δH 1.29 and 1.52 (each 1H,
m); δC 35.8] in 2 taking the place of C-7 in 14. This indicated that
the typical ∆7,8 double bond in 14 was saturated in 2. The structure
was supported by HMBC correlations from H-5 to C-7 and from
H-30 to C-8, and the configuration of 2 was determined by analysis
of the NOESY spectrum and CD data to be the same as 14.
1
Analysis of the H and 13C NMR data of 3 (C31H54O2, cornus-
alterin C) revealed that 3 possessed a tirucallane-type triterpenoid
skeleton similar to that of 2. The difference was that one oxygenated
methine [δH 3.22 (1H, dd, J ) 11.5, 4.0 Hz); δC 79.1] in 3 replaced
a ketone at C-3 in 2. The HMBC spectrum showed correlations
from H-3 to C-1, C-5, C-28, and C-29, supporting the presence of
an OH at C-3 in the structure.
Cornusalterin D (4) had the molecular formula C30H46O1.
Inspection of the 1H and 13C NMR data revealed that the tetracyclic
part of the molecule was nearly identical to that of 14,12 and the
side chain was similar to that of (23E)-cucurbita-5,23,25-triene-
3ꢀ,7ꢀ-diol.15 The (23E)-∆23,25-conjugated diene C-8 side chain of
4 was determined by the NMR data [δH 1.83 (3H, s, H-26), 4.84
(2H, br s, H-27), 5.61 (1H, ddd, J ) 15.5, 7.5, 7.5 Hz, H-23), and
6.09 (1H, d, J ) 15.5 Hz, H-24); δC 114.2 (C-27), 130.0 (C-23),
134.1 (C-24), and 142.4 (C-25)] and was confirmed by the HMBC
experiment showing correlations between H-23 and C-20, C-22,
and C-25 and between H-27 and C-24, C-25, and C-26.
Results and Discussion
Cornusalterin A (1), a white, amorphous powder, had the
molecular formula C31H52O2, as obtained from positive-ion HR-
FABMS. The IR spectrum of 1 indicated the presence of OH (3404
cm-1) and CdC double-bond groups (1658 cm-1). The H NMR
1
data (Table 1) showed the presence of seven tertiary methyl [δH
0.76, 0.85, 0.87, 0.98, 0.99, 1.26, and 1.26 (each 3H, s)], a secondary
methyl [δH 0.84 (3H, d, J ) 6.0 Hz)], an O-methyl [δH 3.16 (3H,
s)], an oxygenated methine [δH 3.25 (1H, dd, J ) 11.5, 4.0 Hz)],
and three olefinic [δH 5.27 (1H, d, J ) 2.0 Hz), 5.40 (1H, d, J )
16.0 Hz), and 5.53 (1H, ddd, J ) 16.0, 8.0, 5.5 Hz)] proton signals.
The 13C NMR spectrum indicated 31 carbon resonances, which were
classified by DEPT and HMQC experiments as one trisubstituted
double bond (δC 118.1 and 145.9), one disubstituted double bond
(δC 129.4 and 136.5), nine methyls, including an O-methyl at δC
* To whom correspondence should be addressed. Tel: +82-31-290-7710.
Fax: +82-31-290-7730. E-mail: krlee@skku.ac.kr.
† Sungkyunkwan University.
The molecular formula of cornusalterin E (5) was C30H48O1. The
1H and 13C NMR data of 5 (Tables 1 and 2) were similar to those
of 4, except that the proton and carbon resonances of two double
‡ Korea Research Institute of Chemical Technology.
§ Seoul National University.
bonds in the side chain were absent, and resonances of the ∆1,2
-
10.1021/np100660v  2011 American Chemical Society and American Society of Pharmacognosy
Published on Web 12/23/2010

Tirucallane Triterpenoids from Cornus walteri
Journal of Natural Products, 2011, Vol. 74, No. 1 55
Chart 1
Table 1. 1H NMR Data of Compounds 1, 5-7, and 12 (CDCl3, 500 MHz, δ in ppm, J in Hz)a
H
1
5
6
7
12
1R
1.15, m
1.65, m
1.65, m
1.74, m
6.93, d (10.0)
1.43, m
1.97, m
2.24, ddd (14.5, 5.0, 5.0)
2.76, ddd (14.5, 14.5, 5.0)
1.44, m
1.97, m
2.23, ddd (14.5, 5.0, 5.0)
2.75, ddd (14.5, 14.5, 5.0)
1ꢀ
2R
2ꢀ
5.92, d (10.0)
5.80, d (10.0)
3R
5R
6
3.25, dd (11.5, 4.0)
1.32, dd (11.5, 6.0)
2.20, m, 2.14, m
5.27, d (2.0)
2.23, m
1.53, m
1.17, m
1.44, m
1.51, m
1.53, m
1.79, m
1.32, m
1.96, m
1.49, m
0.85, s
6.50, d (10.0)
2.12, m
2.22, m, 2.06, m
5.56, m
2.82, m
1.60, m
1.26, m
1.58, m
1.64, m
1.59, m
2.06, m
2.19, m, 2.08, m
5.58, m
2.55, m
1.57, m
1.09, m
1.53, m
1.62, m
1.54, m
1.70, m
1.56, m
1.81, m
1.32, m
0.98, s
1.74, m
2.11, m, 2.09, m
5.31, m
2.28, m
1.68, m
1.26, m
1.52, m
1.58, m
1.50., m
1.78, m
1.48, m
1.97, m
1.46, m
0.85, s
1.72, m
2.11, m, 2.09, m
5.31, m
2.31, m
1.61, m
1.25, m
1.52, m
1.58, m
1.50, m
1.79, m
1.46, m
1.87, m
1.46, m
7
9R
11R
11ꢀ
12R
12ꢀ
15R
15ꢀ
16R
16ꢀ
17
1.67, m
1.56, m
1.70, m
1.37, m
18
1.06, s
0.84, s
19
0.76, s
1.04, s
1.02, s
1.01, s
1.01, s
20
21
1.55, m
0.84, d (6.0)
1.53, m
1.06, d (6.5)
1.61, m
1.07, d (6.5)
1.70, m
0.90, d (6.5)
1.85, m
3.44, dd (9.0, 8.0)
3.91, dd (8.0, 7.5)
1.79, m, 1.65, m
4.56, m
5.19, br d (8.5)
1.68, s
22
23
24
26
27
28
2.39, m, 1.74, m
5.53, ddd (16.0, 8.0, 5.5)
5.40, d (16.0)
1.26, s
1.26, s
0.99, s
1.24, m, 1.09, m
1.22, m, 1.25, m
1.06, m, 1.10, m
0.87, d (6.5)
0.87, d (6.5)
1.14, s
1.25, m, 1.09, m
1.21, m, 1.24, m
1.08, m, 1.11, m
0.91, d (6.0)
0.91, d (6.0)
1.08, s
1.85, m, 0.89, m
3.94, ddd (9.0, 9.0, 2.0)
5.04, br d (9.0)
1.69, s
1.75, s
1.06, s
1.72, s
1.05, s
29
0.87, s
1.11, s
1.09, s
1.12, s
1.11, s
30
0.98, s
1.06, s
1.07, s
1.02, s
1.01, s
OMe
3.16, s
3.21, s
a The assignments were based on DEPT, HMQC, and HMBC experiments.
double bond [δH 6.93 (1H, d, J ) 10.0 Hz); δC 155.4 and δH 5.92
(1H, d, J ) 10.0 Hz); δC 126.0] were present in 5. In the HMBC
spectrum, correlations of H-1/C-3, C-5, C-19, H-2/C-4, C-10, H-28/
C-3, C-5, H-29/C-3, C-5, and H-19/C-1, C-5, C-9, C-10 revealed
the presence of a double bond at C-1/C-2 and a ketone at C-3.
HMBC correlations of H-21/C-17, C-22, H-26/C-24, C-25, H-27/
C-24, C-25, and H-30/C-8, C-13, C-15 were also observed. This
evidence confirmed that 5 and lanosta-1,7-dien-3-one shared the
same planar structure.16 However, the strongly negative optical
rotation of 5 suggested that it was a tirucallane-type triterpenoid,12,13
since lanosta-1,7-dien-3-one had a positive optical rotation value.16
This conclusion was supported by analysis of the NOESY spectrum
(correlations of H-19/H-30, H-30/H-17, H-5/H-9, H-9/H-18, and
H-17/H-21) and CD data.
Cornusalterin F (6) exhibited a molecular formula of C30H48O1.
Analysis of the 1H and 13C NMR data of 6 (Tables 1 and 2) revealed
that 6 possessed a tirucallane-type triterpenoid skeleton similar to
that of 5. The major differences were the chemical shifts of a double
bond [δH 5.80 and 6.50 (each 1H, d, J ) 10.0 Hz); δC 126.0 and
157.2] in 6 instead of those of the double bond [δH 5.92 and 6.93
(each 1H, d, J ) 10.0 Hz); δC 126.0 and 155.4] in 5. HMBC
correlations from H-19 (δH 1.02) to C-1 (δC 206.0), C-9 (δC 41.8),
and C-10 (δC 38.1), from H-28 (δH 1.08) to C-3 (δC 157.2), and

56 Journal of Natural Products, 2011, Vol. 74, No. 1
Kim et al.
Table 2. 13C NMR Data of Compounds 1, 5-7, and 12 (CDCl3, 125 MHz, δ in ppm)
carbon
1
5
6
7
12
1
2
3
4
5
6
7
8
37.4 t
27.9 t
79.4 d
39.2 s
50.8 d
24.1 t
118.1 d
145.9 s
49.1 d
35.1 s
18.3 t
34.0 t
43.8 s
51.4 s
34.1 t
28.5 t
53.2 d
22.4 q
13.3 q
36.3 d
19.1 q
38.5 t
129.4 d
136.5 d
75.0 s
26.0 q
26.3 q
27.8 q
14.9 q
27.5 q
50.4 q
155.4 d
126.0 d
205.1 s
45.2 s
48.4 d
23.9 t
117.7 d
145.3 s
44.4 d
38.3 s
17.4 t
32.6 t
42.0 s
55.2 s
32.3 t
29.4 t
55.2 d
23.1 q
13.6 q
35.6 d
21.5 q
37.9 t
24.4 t
38.2 t
29.4 d
22.7 q
22.7 q
24.5 q
21.5 q
25.9 q
206.0 s
126.0 d
157.2 d
35.2 s
47.1 d
24.5 t
116.7 d
146.6 s
41.8 d
38.1 s
19.9 t
33.1 t
42.3 s
55.3 s
32.3 t
29.5 t
55.3 d
23.5 q
23.8 q
35.6 d
22.9 q
37.1 t
24.9 t
38.3 t
29.2 d
22.7 q
22.7 q
25.2 q
24.9 q
25.9 q
38.7 t
35.2 t
217.0 s
48.0 s
52.5 d
24.6 t
117.9 d
146.2 s
48.6 d
35.1 s
18.5 t
38.4 t
34.8 t
216.7 s
47.8 s
52.4 d
24.3 t
118.1 d
145.5 s
48.4 d
35.1 s
17.8 t
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
OMe
34.2 t
32.2 t
43.7 s
51.5 s
34.1 t
43.9 s
50.5 s
34.3 t
28.6 t
27.3 t
54.1 d
22.3 q
13.0 q
32.6 d
18.9 q
41.7 t
75.3 d
127.4 d
134.7 s
18.3 q
26.0 q
24.8 q
21.8 q
27.6 q
55.9 q
51.0 d
22.5 q
12.7 q
43.1 d
72.9 t
38.6 t
75.7 d
126.1 d
135.7 s
18.0 q
25.8 q
24.5 q
21.5 q
27.2 q
from H-29 (δH 1.09) to C-3 (δC 157.2) revealed the presence of a
ketone at C-1 and a double bond at C-2/C-3.
Cornusalterin I (9) had the molecular formula C31H48O2. The 1H
and 13C NMR data of 9 were very similar to those of 8, except for
the presence of the additional signals assignable to a double bond
[δH 5.42 (1H, m, H-11); δC 114.0 (C-11) and 145.8 (C-9)] in 9.
This ∆9,11 double bond in 9 was confirmed by HMBC correlations
from H-7 to C-9 (δC 145.8), from H-11 to C-8 (δC 142.2), C-10
(δC 35.0), and C-13 (δC 43.5), and from H-19 to C-9 (δC 145.8).
Similarly, as described for 7, the absolute configuration of C-23 in
9 was determined using demethylation18 and modified Mosher’s
method,19 which proved the S-configuration for C-23 (see details
in the Supporting Information). In addition, the absolute configu-
ration of C-23 in 8 was also determined to be S because the chemical
shifts and coupling constants of H-23 and H-24 of 8 were very
similar to those of 9. This configuration of 8 was also supported
by NOESY correlations of O-methyl/H-17 and O-methyl/H-21.
The molecular formula of cornusalterin G (7) was established
to be C31H50O2. Its NMR data (Tables 1 and 2) resembled those of
14,12 except for some differences in the side chain. The chemical
shifts from C-20 to C-27 in the side chain were similar to those of
23-hydroxycycloart-24-en-3-one.17 The chemical shifts of C-23 in
7 [δH 3.94 (1H, ddd, J ) 9.0, 9.0, 2.0 Hz); δC 75.3] were shifted
as compared with those of C-23 in 23-hydroxycycloart-24-en-3-
one [δH 4.30-4.70 (1H, m); δC 65.8],17 indicating that an OCH3
group [δH 3.21 (3H, s); δC 55.9] instead of an OH group at C-23
in 7 was present. The side chain was confirmed by the HMBC
experiment (correlations of H-21/C-17, C-22, H-23/C-20, C-24,
C-25, and H-24/C-22, C-23, C-26, C-27). In addition, the HMBC
correlation between O-methyl (δH 3.21) and C-23 (δC 75.3)
indicated that the OCH3 group was located at C-23. Finally, the
absolute configuration of C-23 was determined using demethyla-
tion18 and modified Mosher’s method.19 Compound 7 was reacted
with trimethylsilyl iodide to give 3-oxotirucalla-7,24-dien-23-ol
(7a), and then treatment of 7a with (R)- and (S)-MTPA-Cl gave
the (S)- and (R)-MTPA esters 7s and 7r, respectively. The 1H NMR
signals of the two MTPA esters were assigned on the basis of their
1H-1H COSY spectra, and the ∆δH(S-R) values were then calculated
(see Supporting Information). The results indicated that the absolute
configuration of C-23 was R, which was also supported by NOSEY
correlations of H-23/H-17, H-21 and O-methyl/H-18.
Inspection of the 1H and 13C NMR data of 8 (C31H50O2,
cornusalterin H) revealed that these data were very similar to those
of 7, except for the chemical shifts from C-23 to C-25 [δH 3.93
(1H, ddd, J ) 9.5, 9.5, 4.0 Hz) and 4.93 (1H, br d, J ) 9.5 Hz);
δC 76.3 (C-23), 126.0 (C-24), and 136.9 (C-25) in 8; δH 3.94 (1H,
ddd, J ) 9.0, 9.0, 2.0 Hz) and 5.04 (1H, br d, J ) 9.0 Hz); δC 75.3
(C-23), 127.4 (C-24), and 134.7 (C-25) in 7]. Analysis of the 2D
NMR data of 8 (HMQC, HMBC, and NOESY) led to unambiguous
1H and 13C NMR assignments and confirmed 8 to be the 23-epimer
of 7.
The NMR data of 10 (C31H52O2, cornusalterin J) resembled those
of 7, except for signals of an oxygenated methine [δH 3.23 (1H,
dd, J ) 11.5, 4.0 Hz); δC 79.2] in 10. The HMBC spectrum showed
correlations from H-3 to C-1, C-5, C-28, and C-29, suggesting the
presence of an OH at C-3 in the structure. A ꢀ-orientation of OH-3
was determined by NOSEY correlations of H-3/H-28 and H-3/H-
5. The absolute configuration of C-23 was determined to be R on
the basis of the fact that the chemical shifts and splitting patterns
of H-23 and H-24 of 10 were very similar to those of 7.
Cornusalterin K (11) gave a molecular formula of C30H50O2. The
1H and 13C NMR data of 11 were compatible with 5, except that
the proton and carbon resonances of the ∆1,2 double bond were
absent, and resonances of one oxygenated methine [δH 3.52 (1H,
dd, J ) 12.0, 4.5 Hz); δC 78.9] and one methylene [δH 2.46 (1H,
dd, J ) 12.0, 4.5 Hz), 2.97 (1H, dd, J ) 12.0, 12.0 Hz); δC 45.6]
were present in 11. The structure of the tetracyclic A and B rings
was verified by HMBC correlations from H-1 to C-19, from H-2
to C-1, C-3, C-4, and C-10, from H-19 to C-1, C-5, and C-9, from
H-28 to C-3 and C-5, and from H-29 to C-3 and C-5, confirming
the presence of an OH at C-1, a ketone at C-3, and a ∆5,6 double
bond in the structure. The coupling constants of H-1 (dd, J1,2R
)

Tirucallane Triterpenoids from Cornus walteri
Journal of Natural Products, 2011, Vol. 74, No. 1 57
1
the A549, SK-OV-3, and SK-MEL-2 cell lines using a sulforhodamine
4.5 Hz and J1,2ꢀ ) 12.0 Hz) observed in the H NMR spectrum
indicated a ꢀ-orientation of OH-1, which was confirmed by NOESY
correlations of H-1/H-9 and H-1/H-28.
B
(SRB) assay. The n-hexane-soluble fraction was chromato-
graphed on a silica gel (230-400 mesh, 300 g) column eluted with
n-hexane-EtOAc (3:1 f 1:1, gradient system) to yield fractions
H1-H5. Fraction H1 (3.3 g) was chromatographed on an RP-C18 silica
gel (230-400 mesh, 100 g) column eluted with 100% MeOH to give
subfractions H11-H15. Fraction H13 (800 mg) was subjected to silica
gel CC (n-hexane-EtOAc, 7:1) and preparative RP-HPLC using 100%
MeOH to yield compounds 2 (10 mg) and 8 (5 mg). Fraction H14
(300 mg) was subjected to a LiChroprep Lobar-A Si gel 60 column
(n-hexane-EtOAc, 16:1) and then normal-phase HPLC using
n-hexane-EtOAc (12:1) to afford compounds 7 (12 mg) and 14 (30
mg). Compounds 5 (15 mg) and 6 (5 mg) were obtained from fraction
H15 (100 mg) by HPLC using n-hexane-EtOAc (20:1). Fraction H2
(2.5 g) was chromatographed on an RP-C18 silica gel (230-400 mesh,
100 g) column eluted with 100% MeOH to give subfractions H21-H28.
Fraction H24 (300 mg) was subjected to HPLC using n-hexane-EtOAc
(8:1) to yield compounds 12 (9 mg) and 13 (15 mg). Fraction H25 (80
mg) was separated by HPLC (n-hexane-EtOAc, 8:1) to give com-
pounds 9 (13 mg) and 10 (5 mg). Fraction H3 (1.7 g) was chromato-
graphed on an RP-C18 silica gel (230-400 mesh, 70 g) column eluted
with 100% MeOH to afford subfractions H31-H36. Fraction H33 (400
mg) was subjected to a LiChroprep Lobar-A Si gel 60 column
(n-hexane-EtOAc, 4:1) to give five subfractions, H331-H335. Fraction
H332 (100 mg) was purified by RP-HPLC using 100% MeOH to yield
4 (7 mg). Fraction H34 (120 mg) was fractionated by HPLC (n-
hexane-EtOAc, 3:1) to give 1 (30 mg), and fraction H35 (100 mg)
was separated by HPLC (n-hexane-EtOAc, 4:1) to obtain 11 (42 mg).
Fraction H4 (1.3 g) was chromatographed on an RP-C18 silica gel
(230-400 mesh, 50 g) column eluted with 100% MeOH to afford
subfractions H41-H45. Fraction H42 (200 mg) was subjected to a silica
gel CC (n-hexane-EtOAc, 3:1) and purified by preparative HPLC (n-
hexane-EtOAc, 2:1) to yield 3 (10 mg).
The molecular formula of cornusalterin L (12) was determined
to be C30H46O2. The 1H and 13C NMR data of 12 (Tables 1 and 2)
suggested that 12 had the same triterpene skeleton as 13.13,20 The
chemical shifts of H-21 [δH 3.44 (1H, dd, J ) 9.0, 8.0 Hz) and
3.91 (1H, dd, J ) 8.0, 7.5 Hz)] and H-23 [δH 4.56 (1H, m)]
suggested the presence of an epoxy function at these positions,
forming a tetrahydrofuran ring in the side chain.20 The structure
was confirmed by HMBC correlations from H-21 to C-17, C-22,
and C-23, from H-23 to C-20, C-21, and C-25, and from H-24 to
C-22, C-26, and C-27. The chemical shifts of H-21 and H-23 in
the side chain of 12 differed from those of 13, indicating that 12 is
a stereoisomer of 13. The NOESY spectrum of 13 showed a
correlation from H-20 to H-23, but that of 12 did not. Moreover,
the CD spectrum of 12 showed a positive (λmax 258 nm) Cotton
effect and that of 13 showed a negative (λmax 259 nm) Cotton effect.
The remaining configuration of 12 was determined to be the same
as 13 by analysis of the NOESY and CD data. In addition, full
NMR data of 13 were assigned using 2D NMR data (including
HMQC, HMBC, and NOESY) because the published 13C NMR
data of 21,23-epoxytirucalla-7,24-diene-3-one were not in good
agreement with those of 13.13
The known compound 14 was identified by comparison of
spectroscopic data with reported data.12 Compound 14 showed the
strongly negative specific optical rotation value [R]2D5 -102.3 (c
0.93, CHCl3) and a negative (λmax 295 nm) Cotton effect.
Cytotoxic activities of compounds 1-14 against the A549, SK-
OV-3, SK-MEL-2, and XF498 human cancer cell lines were
evaluated using the SRB assay.21 Compounds 12 and 13, possessing
a tetrahydrofuran ring in the side chain, exhibited significant
cytotoxic activity against the A549, SK-OV-3, SK-MEL-2, and
XF498 cell lines (IC50(12): 4.29, 3.82, 4.73, and 5.81 µM, and
IC50(13): 4.02, 3.64, 6.07, and 5.10 µM, respectively). The other
compounds were essentially noncytotoxic (see Supporting Informa-
tion).
Cornusalterin A (1): white, amorphous powder; mp 115.0-116.0
°C; [R]2D5 -98.6 (c 0.85, CHCl3); CD (MeOH) λmax (∆ε) 294 (-18.0),
279 (+5.3) nm; IR (KBr) νmax 3404, 2949, 1738, 1658, 1461, 1374,
1216, 1032, 696 cm-1; 1H NMR data, see Table 1; 13C NMR data, see
Table 2; FABMS m/z 456 [M]+; HRFABMS m/z 456.3966 [M]+ (calcd
for C31H52O2, 456.3967).
Cornusalterin B (2): white, amorphous powder; mp 131.5-132.5
°C; [R]2D5 -31.8 (c 0.40, CHCl3); CD (MeOH) λmax (∆ε) 292 (-16.4),
272 (+2.5) nm; IR (KBr) νmax 2948, 1705, 1657, 1374, 1031, 731 cm-1
;
1H NMR (CDCl3, 500 MHz) δ 5.51 (1H, ddd, J ) 16.0, 8.0, 5.5 Hz,
H-23), 5.41 (1H, d, J ) 16.0 Hz, H-24), 3.16 (3H, s, OCH3-25), 2.65
(1H, ddd, J ) 14.5, 14.5, 5.0 Hz, H-2ꢀ), 2.23 (1H, m, H-22a), 2.06
(1H, ddd, J ) 14.5, 5.0, 5.0 Hz, H-2R), 1.84 (1H, m, H-1ꢀ), 1.75 (1H,
m, H-6a), 1.72 (1H, m, H-22b), 1.71 (2H, m, H-12ꢀ, H-16ꢀ), 1.59
(1H, m, H-20), 1.57 (1H, m, H-15ꢀ), 1.55 (1H, m, H-16R), 1.53 (1H,
m, H-6b), 1.52 (2H, m, H-7a, H-11R), 1.50 (1H, m, H-17), 1.42 (1H,
m, H-8), 1.35 (1H, m, H-9R), 1.34 (1H, m, H-1R), 1.29 (2H, m, H-7b,
H-15R), 1.27 (6H, s, H-26, H-27), 1.22 (1H, m, H-12R), 1.12 (3H, s,
H-28), 1.08 (2H, m, H-5R, H-11ꢀ), 1.08 (3H, s, H-29), 0.97 (3H, s,
H-19), 0.94 (3H, s, H-30), 0.87 (3H, d, J ) 6.0 Hz, H-21), 0.85 (3H,
s, H-18); 13C NMR (CDCl3, 125 MHz) δ 216.2 (C-3), 136.7 (C-24),
129.2 (C-23), 75.0 (C-25), 55.2 (C-17), 53.2 (C-5), 50.4 (OCH3-25),
49.5 (C-9), 48.7 (C-14), 47.2 (C-4), 46.7 (C-13), 42.6 (C-8), 38.4 (C-
22), 38.3 (C-1), 37.7 (C-20), 35.8 (C-7), 34.8 (C-2), 33.1 (C-15), 32.3
(C-10), 29.6 (C-16), 26.3 (C-27), 26.0 (C-26), 25.1 (C-12), 23.8 (C-
28), 21.5 (C-11), 21.0 (C-29), 18.5 (C-6), 18.3 (C-21), 16.1 (C-30),
14.8 (C-18), 14.7 (C-19); FABMS m/z 457 [M + H]+; HRFABMS
m/z 457.4048 [M + H]+ (calcd for C31H53O2, 457.4046).
Cornusalterin C (3): white, amorphous powder; mp 151.0-152.0
°C; [R]2D5 -20.5 (c 0.45, CHCl3); CD (MeOH) λmax (∆ε) 295 (-18.2)
nm; IR (KBr) νmax 3390, 2948, 2838, 1657, 1373, 1031, 722 cm-1; 1H
NMR (CDCl3, 500 MHz) δ 5.65 (1H, ddd, J ) 16.0, 8.0, 5.5 Hz, H-23),
5.51 (1H, d, J ) 16.0 Hz, H-24), 3.22 (1H, dd, J ) 11.5, 4.0 Hz,
H-3R), 3.17 (3H, s, OCH3-25), 2.23 (1H, m, H-22a), 1.72 (3H, m, H-6a,
H-12ꢀ, H-22b), 1.69 (1H, m, H-2ꢀ), 1.67 (1H, m, H-16ꢀ), 1.60 (1H,
m, H-2R), 1.57 (2H, m, H-16R, H-20), 1.56 (1H, m, H-1ꢀ), 1.53 (2H,
m, H-7a, H-15ꢀ), 1.51 (2H, m, H-6b, H-11R), 1.48 (1H, m, H-17),
1.44 (1H, m, H-8), 1.34 (1H, m, H-9R), 1.28 (2H, m, H-7b, H-15R),
1.28 (6H, s, H-26, H-27), 1.27 (1H, m, H-1R), 1.22 (1H, m, H-12R),
1.15 (3H, s, H-28), 1.06 (1H, m, H-11ꢀ), 0.99 (6H, s, H-18, H-30),
0.87 (3H, s, H-29), 0.86 (3H, d, J ) 6.0 Hz, H-21), 0.79 (3H, s, H-19),
0.77 (1H, m, H-5R); 13C NMR (CDCl3, 125 MHz) δ 136.4 (C-24),
Experimental Section
General Experimental Procedures. Melting points were determined
on a Gallenkamp melting point apparatus and are uncorrected. Optical
rotations were measured on a JASCO P-1020 polarimeter. CD spectra
were measured on a JASCO J-715 spectropolarimeter. IR spectra were
recorded on a Bruker IFS-66/S FT-IR spectrometer. UV spectra were
obtained on a Varian Cary 5000 UV-vis-NIR spectrophotometer.
NMR spectra were recorded on a Varian UNITY INOVA 500 NMR
spectrometer operating at 500 MHz (1H) or 125 MHz (13C), with
chemical shifts given in ppm (δ). FAB and HR-FAB mass spectra were
obtained on a JEOL JMS700 mass spectrometer. Preparative HPLC
was performed using a Gilson 306 pump with a Shodex refractive index
detector. Chromatographic separation was performed on an Apollo
Silica 5 µm column (250 × 10 mm i.d.) or Econosil RP-18 10 µm
column (250 × 10 mm i.d.). Silica gel 60 and RP-C18 silica gel (Merck
Co., Germany, 70-230 and 230-400 mesh) were used for column
chromatography. TLC was performed using Merck precoated silica gel
F254 plates and RP-18 F254s plates. Low-pressure liquid chromatography
was performed over a LiChroprep Lobar-A Si gel 60 (240 × 10 mm
i.d.) column with a FMI QSY-0 pump (ISCO).
Plant Material. Stems and stem bark of C. walteri were collected
on Jeju Island, Korea, in October 2005, and the plant was identified
by one of the authors (K.R.L.). A voucher specimen (SKKU 2005-
10a) was deposited in the herbarium of the School of Pharmacy,
Sungkyunkwan University, Suwon, Korea.
Extraction and Isolation. Stems and stem bark of C. walteri (2.5
kg) were dried, chopped, and extracted with 80% MeOH (2 × 6 h)
under reflux and filtered. The filtrate was evaporated under vacuum to
obtain a MeOH extract (220 g), which was suspended in distilled H2O
(7.2 L) and then successively partitioned with n-hexane, CHCl3, and
n-BuOH, yielding 9.5, 25.0, and 43.0 g of residue, respectively. The
n-hexane-soluble fraction showed significant cytotoxic activity against

58 Journal of Natural Products, 2011, Vol. 74, No. 1
Kim et al.
128.1 (C-23), 79.1 (C-3), 75.6 (C-25), 56.1 (C-5), 54.5 (C-17), 50.9
(C-9), 50.7 (C-14), 50.2 (OCH3-25), 45.3 (C-13), 40.6 (C-8), 39.2 (C-
22), 39.2 (C-4), 37.1 (C-1), 35.7 (C-20), 35.4 (C-7), 33.0 (C-15), 31.7
(C-10), 28.2 (C-16), 27.6 (C-2), 27.6 (C-28), 26.4 (C-29), 26.1 (C-
27), 25.8 (C-26), 25.1 (C-12), 21.4 (C-11), 18.4 (C-6), 18.4 (C-21),
15.7 (C-30), 15.7 (C-19), 15.5 (C-18); FABMS m/z 459 [M + H]+;
HRFABMS m/z 459.4209 [M + H]+ (calcd for C31H55O2, 459.4202).
Cornusalterin D (4): white, amorphous powder; mp 105.0-106.0
°C; [R]2D5 -136.0 (c 0.32, CHCl3); CD (MeOH) λmax (∆ε) 293 (-21.5),
269 (+11.8) nm; IR (KBr) νmax 2949, 2836, 1706, 1657, 1453, 1216,
Hz, H-23), 3.23 (3H, s, OCH3-23), 2.74 (1H, ddd, J ) 14.5, 14.5, 5.5
Hz, H-2ꢀ), 2.27 (1H, ddd, J ) 14.5, 5.5, 5.5 Hz, H-2R), 2.24 (1H, m,
H-12ꢀ), 2.11 (1H, m, H-6a), 2.10 (1H, m, H-6b), 2.02 (1H, m, H-12R),
2.00 (1H, m, H-5R), 1.95 (1H, m, H-1ꢀ), 1.81 (1H, m, H-22a), 1.79
(1H, m, H-16ꢀ), 1.78 (3H, s, H-27), 1.73 (3H, s, H-26), 1.69 (1H, m,
H-20), 1.66 (1H, m, H-15ꢀ), 1.50 (1H, m, H-15R), 1.47 (1H, m, H-16R),
1.46 (2H, m, H-1R, H-17), 1.11 (3H, s, H-29), 1.06 (3H, s, H-28),
1.02 (3H, s, H-30), 1.01 (3H, s, H-19), 0.91 (3H, d, J ) 6.5 Hz, H-21),
0.89 (1H, m, H-22b), 0.79 (3H, s, H-18); 13C NMR (CDCl3, 125 MHz)
δ 216.9 (C-3), 145.8 (C-9), 142.2 (C-8), 133.8 (C-25), 129.8 (C-24),
117.8 (C-7), 114.0 (C-11), 73.5 (C-23), 54.2 (OCH3-23), 52.3 (C-17),
51.3 (C-5), 48.4 (C-14), 47.8 (C-4), 43.5 (C-13), 42.2 (C-22), 36.4
(C-1), 35.1 (C-12), 35.0 (C-10), 34.9 (C-2), 33.9 (C-20), 33.6 (C-15),
27.4 (C-16), 27.2 (C-30), 25.7 (C-27), 24.5 (C-6), 24.3 (C-28), 22.2
(C-18), 21.6 (C-29), 18.7 (C-21), 18.3 (C-26), 12.7 (C-19); FABMS
m/z 453 [M + H]+; HRFABMS m/z 453.3734 [M + H]+ (calcd for
C31H49O2, 453.3733).
1
1030, 726 cm-1; UV (MeOH) λmax (log ε) 227 (3.2) nm; H NMR
(CDCl3, 500 MHz) δ 6.09 (1H, d, J ) 15.5 Hz, H-24), 5.61 (1H, ddd,
J ) 15.5, 7.5, 7.5 Hz, H-23), 5.31 (1H, d, J ) 2.5 Hz, H-7), 4.84 (2H,
br s, H-27), 2.74 (1H, ddd, J ) 15.0, 15.0, 5.5 Hz, H-2ꢀ), 2.31 (1H,
m, H-9R), 2.23 (1H, ddd, J ) 15.0, 5.5, 5.5 Hz, H-2R), 2.10 (1H, m,
H-6a), 2.08 (1H, m, H-6b), 2.01 (1H, m, H-22a), 1.95 (1H, m, H-16ꢀ),
1.92 (1H, m, H-1ꢀ), 1.83 (3H, s, H-26), 1.76 (1H, m, H-15ꢀ), 1.75
(1H, m, H-22b), 1.74 (1H, m, H-5R), 1.63 (1H, m, H-11R), 1.56 (1H,
m, H-12ꢀ), 1.54 (1H, m, H-15R), 1.53 (1H, m, H-20), 1.52 (1H, m,
H-12R), 1.48 (1H, m, H-17), 1.43 (2H, m, H-1R, H-16R), 1.23 (1H,
m, H-11ꢀ), 1.11 (3H, s, H-29), 1.04 (3H, s, H-28), 1.01 (3H, s, H-30),
1.00 (3H, s, H-19), 0.84 (3H, s, H-18), 0.83 (3H, d, J ) 6.5 Hz, H-21);
13C NMR (CDCl3, 125 MHz) δ 217.0 (C-3), 146.1 (C-8), 142.4 (C-
25), 134.1 (C-24), 130.0 (C-23), 118.1 (C-7), 114.2 (C-27), 53.4 (C-
17), 52.5 (C-5), 51.5 (C-14), 48.6 (C-9), 48.1 (C-4), 43.8 (C-13), 38.8
(C-22), 38.7 (C-1), 36.6 (C-20), 35.2 (C-2), 35.1 (C-10), 34.2 (C-12),
33.9 (C-15), 28.5 (C-16), 27.7 (C-30), 24.8 (C-28), 24.6 (C-6), 22.5
(C-18), 21.8 (C-29), 19.1 (C-26), 18.9 (C-21), 18.5 (C-11), 13.0 (C-
19); FABMS m/z 423 [M + H]+; HRFABMS m/z 423.3628 [M + H]+
(calcd for C30H47O1, 423.3627).
Cornusalterin J (10): white, amorphous powder; mp 125.5-126.5
°C; [R]2D5 -16.4 (c 0.25, CHCl3); CD (MeOH) λmax (∆ε) 294 (-14.3)
nm; IR (KBr) νmax 3406, 2970, 1739, 1657, 1368, 1216, 1056, 1033
1
cm-1; H NMR (CDCl3, 500 MHz) δ 5.25 (1H, d, J ) 2.0 Hz, H-7),
5.03 (1H, br d, J ) 8.5 Hz, H-24), 3.94 (1H, ddd, J ) 8.5, 8.5, 2.0 Hz,
H-23), 3.23 (1H, dd, J ) 11.5, 4.0 Hz, H-3R), 3.20 (3H, s, OCH3-23),
2.21 (1H, m, H-9R), 2.19 (1H, m, H-6a), 2.15 (1H, m, H-6b), 1.90
(1H, m, H-16ꢀ), 1.84 (1H, m, H-22a), 1.76 (2H, m, H-2ꢀ, H-15ꢀ),
1.74 (3H, s, H-27), 1.71 (1H, m, H-20), 1.69 (1H, m, H-2R), 1.67 (3H,
s, H-26), 1.66 (1H, m, H-1ꢀ), 1.55 (1H, m, H-15R), 1.53 (1H, m,
H-12ꢀ), 1.51 (1H, m, H-11R), 1.49 (1H, m, H-17), 1.44 (1H, m, H-12R),
1.34 (1H, m, H-5R), 1.30 (1H, m, H-16R), 1.17 (2H, m, H-1R, H-11ꢀ),
0.97 (6H, s, H-28, H-30), 0.90 (3H, d, J ) 6.5 Hz, H-21), 0.90 (1H,
m, H-22b), 0.86 (3H, s, H-29), 0.84 (3H, s, H-18), 0.74 (3H, s, H-19);
13C NMR (CDCl3, 125 MHz) δ 145.6 (C-8), 133.7 (C-25), 129.9 (C-
24), 117.9 (C-7), 79.2 (C-3), 75.6 (C-23), 55.6 (OCH3-23), 53.1 (C-
17), 51.2 (C-14), 50.6 (C-5), 48.8 (C-9), 43.5 (C-13), 42.0 (C-22), 38.9
(C-4), 37.1 (C-1), 34.9 (C-10), 33.9 (C-15), 33.7 (C-12), 32.7 (C-20),
29.1 (C-16), 27.6 (C-28), 27.5 (C-2), 27.4 (C-30), 26.0 (C-27), 23.9
(C-6), 22.2 (C-18), 18.9 (C-21), 18.7 (C-11), 18.6 (C-26), 14.7 (C-
29), 13.1 (C-19); FABMS m/z 456 [M]+; HRFABMS m/z 456.3967
[M]+ (calcd for C31H52O2, 456.3967).
Cornusalterin E (5): white, amorphous powder; mp 102.5-103.5
°C; [R]2D5 -84.1 (c 0.63, CHCl3); CD (MeOH) λmax (∆ε) 296 (-19.0),
256 (+4.1) nm; IR (KBr) νmax 2949, 1669, 1658, 1454, 1029, 724 cm-1
;
1
UV (MeOH) λmax (log ε) 223 (3.9) nm; H NMR data, see Table 1;
13C NMR data, see Table 2; FABMS m/z 425 [M + H]+; HRFABMS
m/z 425.3789 [M + H]+ (calcd for C30H49O1, 425.3783).
Cornusalterin F (6): white, amorphous powder; mp 100.0-101.0
°C; [R]2D5 -72.1 (c 0.41, CHCl3); CD (MeOH) λmax (∆ε) 292 (-12.5),
260 (+8.7) nm; IR (KBr) νmax 2950, 1670, 1656, 1454, 1029, 732 cm-1
;
1
UV (MeOH) λmax (log ε) 224 (3.9) nm; H NMR data, see Table 1;
13C NMR data, see Table 2; FABMS m/z 425 [M + H]+; HRFABMS
m/z 425.3780 [M + H]+ (calcd for C30H49O1, 425.3783).
Cornusalterin K (11): white, amorphous powder; mp 132.5-133.5
°C; [R]2D5 -71.7 (c 0.98, CHCl3); CD (MeOH) λmax (∆ε) 294 (-15.4),
250 (+3.4) nm; IR (KBr) νmax 3398, 2928, 2867, 1703, 1462, 1375,
1
Cornusalterin G (7): white, amorphous powder; mp 127.0-128.0
1251, 1041, 822 cm-1; H NMR (CDCl3, 500 MHz) δ 5.51 (1H, m,
°C; [R]2D5 -14.3 (c 0.60, CHCl3); CD (MeOH) λmax (∆ε) 294 (-22.3)
H-6), 3.52 (1H, dd, J ) 12.0, 4.5 Hz, H-1R), 2.97 (1H, dd, J ) 12.0,
12.0 Hz, H-2ꢀ), 2.46 (1H, dd, J ) 12.0, 4.5 Hz, H-2R), 2.07 (1H, m,
H-7a), 1.80 (1H, m, H-16ꢀ), 1.67 (1H, m, H-12ꢀ), 1.61 (1H, m, H-15ꢀ),
1.58 (2H, m, H-7b, H-16R), 1.56 (1H, m, H-15R), 1.55 (1H, m, H-11R),
1.53 (1H, m, H-20), 1.51 (1H, m, H-12R), 1.45 (1H, m, H-8), 1.41
(1H, m, H-9R), 1.30 (1H, m, H-17), 1.28 (1H, m, H-22a), 1.26 (1H,
m, H-23a), 1.23 (1H, m, H-23b), 1.22 (1H, m, H-11ꢀ), 1.12 (1H, m,
H-24a), 1.12 (3H, s, H-30), 1.09 (1H, m, H-22b), 1.09 (3H, s, H-19),
1.06 (1H, m, H-24b), 1.05 (3H, s, H-29), 1.04 (3H, s, H-28), 1.03 (3H,
d, J ) 6.5 Hz, H-21), 1.02 (3H, s, H-18), 0.91 (6H, d, J ) 6.5 Hz,
H-26, H-27); 13C NMR (CDCl3, 125 MHz) δ 213.3 (C-3), 146.1 (C-
5), 116.5 (C-6), 78.9 (C-1), 55.2 (C-17), 55.2 (C-14), 50.8 (C-4), 49.0
(C-8), 45.6 (C-2), 42.0 (C-13), 41.3 (C-9), 39.6 (C-10), 38.3 (C-24),
37.7 (C-22), 35.5 (C-20), 33.1 (C-12), 32.3 (C-15), 29.7 (C-16), 29.5
(C-7), 28.1 (C-25), 25.9 (C-28), 25.0 (C-29), 23.8 (C-23), 23.7 (C-
18), 23.5 (C-21), 22.8 (C-26), 22.8 (C-27), 18.8 (C-11), 15.6 (C-19),
12.2 (C-30); FABMS m/z 443 [M + H]+; HRFABMS m/z 443.3895
[M + H]+ (calcd for C30H51O2, 443.3889).
nm; IR (KBr) νmax 2969, 1708, 1657, 1370, 1216, 1056, 1033 cm-1
;
1H NMR data, see Table 1; 13C NMR data, see Table 2; FABMS m/z
455 [M + H]+; HRFABMS m/z 455.3884 [M + H]+ (calcd for
C31H51O2, 455.3889).
Cornusalterin H (8): white, amorphous powder; mp 127.5-128.5
°C; [R]2D5 -5.1 (c 0.23, CHCl3); CD (MeOH) λmax (∆ε) 296 (-20.1)
nm; IR (KBr) νmax 2949, 2836, 1707, 1657, 1452, 1030, 713 cm-1; 1H
NMR (CDCl3, 500 MHz) δ 5.31 (1H, m, H-7), 4.93 (1H, br d, J ) 9.5
Hz, H-24), 3.93 (1H, ddd, J ) 9.5, 9.5, 4.0 Hz, H-23), 3.23 (3H, s,
OCH3-23), 2.76 (1H, ddd, J ) 14.5, 14.5, 5.0 Hz, H-2ꢀ), 2.29 (1H, m,
H-9R), 2.25 (1H, ddd, J ) 14.5, 5.0, 5.0 Hz, H-2R), 2.11 (1H, m, H-6a),
2.09 (1H, m, H-6b), 1.97 (1H, m, H-1ꢀ), 1.95 (1H, m, H-16ꢀ), 1.84
(1H, m, H-22a), 1.79 (1H, m, H-15ꢀ), 1.79 (3H, s, H-27), 1.73 (3H, s,
H-26), 1.73 (1H, m, H-5R), 1.70 (1H, m, H-20), 1.68 (1H, m, H-11R),
1.58 (1H, m, H-12ꢀ), 1.53 (1H, m, H-12R), 1.50 (1H, m, H-15R), 1.47
(2H, m, H-16R, H-17), 1.44 (1H, m, H-1R), 1.26 (1H, m, H-11ꢀ), 1.13
(3H, s, H-29), 1.02 (3H, s, H-28), 1.01 (6H, s, H-19, H-30), 0.91 (1H,
m, H-22b), 0.86 (3H, d, J ) 6.0 Hz, H-21), 0.79 (3H, s, H-18); 13C
NMR (CDCl3, 125 MHz) δ 217.0 (C-3), 146.2 (C-8), 136.9 (C-25),
126.0 (C-24), 118.0 (C-7), 76.3 (C-23), 55.4 (OCH3-23), 53.9 (C-17),
52.5 (C-5), 51.5 (C-14), 48.7 (C-9), 48.1 (C-4), 43.6 (C-13), 41.2 (C-
22), 38.7 (C-1), 35.2 (C-2), 35.1 (C-10), 34.3 (C-12), 34.1 (C-15), 33.0
(C-20), 28.5 (C-16), 27.7 (C-30), 26.1 (C-27), 24.8 (C-28), 24.6 (C-
6), 22.2 (C-18), 21.8 (C-29), 19.4 (C-21), 18.7 (C-26), 18.5 (C-11),
13.0 (C-19); FABMS m/z 455 [M + H]+; HRFABMS m/z 455.3890
[M + H]+ (calcd for C31H51O2, 455.3889).
Cornusalterin L (12): white, amorphous powder; mp 140.5-141.5
°C; [R]2D5 -89.6 (c 0.45, CHCl3); CD (MeOH) λmax (∆ε) 294 (-24.7),
265 (+4.2), 258 (+3.7) nm; IR (KBr) νmax 2969, 1739, 1706, 1657,
1
1370, 1216, 1031, 695 cm-1; H NMR data, see Table 1; 13C NMR
data, see Table 2; FABMS m/z 439 [M + H]+; HRFABMS m/z
439.3576 [M + H]+ (calcd for C30H47O2, 439.3576).
Deoxyflindissone (13): white, amorphous powder; mp 143.0-144.0
°C; [R]2D5 -41.0 (c 0.60, CHCl3); CD (MeOH) λmax (∆ε) 294 (-27.2),
266 (+6.8), 259 (-4.5) nm; IR (KBr) νmax 2967, 1706, 1657, 1462,
1
Cornusalterin I (9): white, amorphous powder; mp 120.0-121.0
°C; [R]2D5 -83.5 (c 0.65, CHCl3); CD (MeOH) λmax (∆ε) 294 (-22.5)
nm; IR (KBr) νmax 2948, 2835, 1707, 1658, 1452, 1029, 732 cm-1; 1H
NMR (CDCl3, 500 MHz) δ 5.42 (1H, m, H-11), 5.31 (1H, m, H-7),
4.94 (1H, d, J ) 10.0 Hz, H-24), 3.94 (1H, ddd, J ) 10.0, 10.0, 4.0
1380, 1057, 725 cm-1; H NMR (CDCl3, 500 MHz) δ 5.32 (1H, m,
H-7), 5.21 (1H, br d, J ) 9.0 Hz, H-24), 4.66 (1H, m, H-23), 4.01
(1H, dd, J ) 7.5, 7.5 Hz, H-21a), 3.27 (1H, dd, J ) 9.5, 7.5 Hz, H-21b),
2.76 (1H, ddd, J ) 14.5, 14.5, 5.0 Hz, H-2ꢀ), 2.33 (1H, m, H-9R),
2.25 (1H, ddd, J ) 14.5, 5.0, 5.0 Hz, H-2R), 2.11 (1H, m, H-6a), 2.09

Tirucallane Triterpenoids from Cornus walteri
Journal of Natural Products, 2011, Vol. 74, No. 1 59
(1H, m, H-6b), 2.01 (1H, m, H-1ꢀ), 1.87 (1H, m, H-16ꢀ), 1.83 (1H,
m, H-20), 1.78 (1H, m, H-15ꢀ), 1.77 (1H, m, H-22a), 1.75 (1H, m,
H-5R), 1.73 (3H, s, H-27), 1.70 (3H, s, H-26), 1.66 (1H, m, H-22b),
1.60 (1H, m, H-11R), 1.58 (1H, m, H-12ꢀ), 1.51 (1H, m, H-15R), 1.50
(1H, m, H-12R), 1.48 (1H, m, H-16R), 1.45 (1H, m, H-1R), 1.44 (1H,
m, H-17), 1.27 (1H, m, H-11ꢀ), 1.13 (3H, s, H-29), 1.06 (3H, s, H-28),
1.03 (6H, s, H-19, H-30), 0.85 (3H, s, H-18); 13C NMR (CDCl3, 125
MHz) δ 216.7 (C-3), 145.5 (C-8), 134.9 (C-25), 126.8 (C-24), 118.1
(C-7), 75.7 (C-23), 73.5 (C-21), 52.4 (C-5), 50.5 (C-14), 50.4 (C-17),
48.4 (C-9), 47.8 (C-4), 43.9 (C-13), 42.1 (C-20), 38.4 (C-1), 37.3 (C-
22), 35.1 (C-10), 34.9 (C-2), 34.3 (C-15), 32.3 (C-12), 27.4 (C-16),
27.2 (C-30), 25.7 (C-27), 24.5 (C-28), 24.3 (C-6), 22.6 (C-18), 21.5
(C-29), 18.0 (C-26), 17.8 (C-11), 12.7 (C-19); FABMS m/z 439 [M +
H]+.
cultured human cancer cell lines.21 The assays were performed at the
Korea Research Institute of Chemical Technology. The cell lines used
were A549 (non small cell lung adenocarcinoma), SK-OV-3 (ovarian
cancer cells), SK-MEL-2 (skin melanoma), and XF498 (human CNS
cancer). Etoposide was used as a positive control. The cytotoxicities
of etoposide against the A549, SK-OV-3, SK-MEL-2, and XF498 cell
lines were IC50 1.85, 1.81, 1.17, and 1.72 µM, respectively.
Acknowledgment. This research was supported by the Basic Science
Research Program through the National Research Foundation of Korea
(NRF) funded by the Ministry of Education, Science and Technology
(20100029358). We thank Drs. E. J. Bang, S. G. Kim, and J. J. Seo at
the Korea Basic Science Institute for their aid in obtaining the NMR
and mass spectra.
(-)-Leucophyllone (14): white, amorphous powder; mp 123.0-124.0
1
Supporting Information Available: H and 13C NMR spectra for
°C; [R]2D5 -102.3 (c 0.93, CHCl3); CD (MeOH) λmax (∆ε) 295 (-19.5),
255 (+6.6) nm; IR (KBr) νmax 2948, 1708, 1657, 1373, 1031, 727 cm-1
;
1-13, 2D NMR (HMQC, HMBC, NOESY) data for 4-7 and 11-13,
1
FABMS m/z 455 [M + H]+; H and 13C NMR data were identical to
CD spectrum for 1, 7, and 12-14, HMBC correlations for 1-2, 4-7,
reported data.12
1
and 9-12, NOESY correlations for 1, 5, 7, 8, and 11-13, partial H
Demethylation of Compounds 7 and 9 (ref 18). To 4.5 mg of
compound 7 in an NMR tube was added 0.5 mL of CDCl3 to give a
clear solution. Then 0.1 mL of trimethylsilyl iodide was added. The
solution was maintained at room temperature, and the reaction was
monitored by NMR periodically. The reaction was apparently complete
after 24 h, as indicated by the absence of a signal for the OCH3 protons
in the NMR spectrum and the presence of a signal at δ 2.09 for protons
of CH3I. The brown-purple solution was filtered, and 0.5 mL of MeOH
was added to the filtrate. After 2 h, the solvent was evaporated under
vacuum to give a brown gum (6.0 mg), which was suspended in H2O
(3 mL) and partitioned with CHCl3 to yield a purple extract (5.5 mg).
The extract was further purified by a silica gel Waters Sep-Pak Vac 6
cc using a solvent system of n-hexane-EtOAc (10:1) to give 7a (4.0
mg), 3-oxotirucalla-7,24-dien-23-ol. Compound 9 (4.0 mg) was also
reacted with trimethylsilyl iodide as described above to afford 9a (3.5
mg), 3-oxotirucalla-7,9,24-trien-23-ol.
NMR data of the (S)- and (R)-MTPA esters of 7a and 9a, and cytotoxic
activities of compounds 1-14. This material is available free of charge
via the Internet at http://pubs.acs.org.
References and Notes
(1) Lee, D. Y.; Yoo, K. H.; Chung, I. S.; Kim, J. Y.; Chung, D. K.; Kim,
D. K.; Kim, S. H.; Baek, N. I. Arch. Pharm. Res. 2008, 31, 830–833.
(2) Vareed, S. K.; Schutzki, R. E.; Nair, M. G. Phytomedicine 2007, 14,
706–709.
(3) Lee, D. Y.; Song, M. C.; Yoo, K. H.; Bang, M. H.; Chung, I. S.;
Kim, S. H.; Kim, D. K.; Kwon, B. M.; Jeong, T. S.; Park, M. H.;
Baek, N. I. Arch. Pharm. Res. 2007, 30, 402–407.
(4) Lee, D. Y.; Jung, L.; Park, J. H.; Yoo, K. H.; Chung, I. S.; Baek,
N. I. Chem. Nat. Compd. 2010, 46, 142–145.
(5) Nishino, C.; Kobayashi, K.; Fukushima, M. J. Nat. Prod. 1988, 51,
1281–1282.
(6) Lee, T. B. Coloured Flora of Korea; Hyangmoonsa: Seoul, 2006; p
851.
(7) Choi, W. H.; Park, W. Y.; Hwang, B. Y.; Oh, G. J.; Kang, S. J.; Lee,
K. S.; Ro, J. S. Kor. J. Pharmacogn. 1998, 29, 217–224.
(8) Yook, C. S. Coloured Medicinal Plants of Korea; Academybook:
Seoul, 1993; p 368.
(9) Yang, E. J.; Yim, E. Y.; Song, G.; Kim, G. O.; Hyun, C. G. Interdisc.
Toxicol. 2009, 2, 245–249.
(10) Moon, J. Y.; Yim, E. Y.; Song, G.; Lee, N. H.; Hyun, C. G. EurAsia.
J. BioSci. 2010, 4, 41–53.
(11) Lim, C. S.; Li, C. Y.; Kim, Y. M.; Lee, W. Y.; Rhee, H. I. J. Korean
Soc. Appl. Biol. Chem. 2005, 48, 103–108.
(12) Benosman, A.; Richomme, P.; Sevenet, T.; Perromat, G.; Hadi, A. H.;
Bruneton, J. Phytochemistry 1995, 40, 1485–1487.
(13) Bhakuni, R. S.; Shukla, Y. N.; Thakur, R. S. Phytochemistry 1987,
26, 2607–2610.
(14) Mcchesney, J. D.; Dou, J.; Sindelar, R. D.; Goins, D. K.; Walker,
L. A.; Rogers, R. D. J. Chem. Crystallogr. 1997, 27, 283–290.
(15) Chang, C. I.; Chen, C. R.; Liao, Y. W.; Cheng, H. L.; Chen, Y. C.;
Chou, C. H. J. Nat. Prod. 2006, 69, 1168–1171.
(16) Van Tamelen, E. E.; Murphy, J. W. J. Am. Chem. Soc. 1970, 92, 7204–
7206.
(17) Furlan, M.; Roque, N. F.; Filho, W. W. Phytochemistry 1993, 32,
1519–1522.
(18) Jung, M. E.; Lyster, M. A. J. Org. Chem. 1977, 42, 3761–3764.
(19) Wang, B. G.; Ebel, R.; Wang, C. Y.; Wray, V.; Proksch, P.
Tetrahedron Lett. 2002, 43, 5783–5787.
(20) Kumar, V.; Niyaz, N. M. M.; Wickramaratne, D. B. M.; Balasubra-
maniam, S. Phytochemistry 1991, 30, 1231–1233.
(21) Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; MaMahon, J.;
Vistica, D.; Warren, J. T.; Bokesch, H.; Kenney, S.; Boyd, M. R.
J. Natl. Cancer Inst. 1990, 82, 1107–1112.
1
7a: colorless gum; [R]D25 -45.6 (c 0.20, CHCl3); H NMR (CDCl3,
500 MHz) δ 5.34 (1H, m, H-7), 5.17 (1H, br d, J ) 9.0 Hz, H-24),
4.34 (1H, ddd, J ) 9.0, 9.0, 2.0 Hz, H-23), 2.75 (1H, ddd, J ) 14.5,
14.5, 5.0 Hz, H-2ꢀ), 2.24 (1H, ddd, J ) 14.5, 5.0, 5.0 Hz, H-2R), 1.76
(3H, s, H-27), 1.71 (3H, s, H-26), 1.70 (1H, m, H-20), 1.12 (3H, s,
H-29), 1.06 (3H, s, H-28), 1.02 (3H, s, H-30), 1.01 (3H, s, H-19), 0.97
(3H, d, J ) 6.0 Hz, H-21), 0.83 (3H, s, H-18); FABMS m/z 441 [M +
H]+.
1
9a: colorless gum; [R]D25 -21.8 (c 0.17, CHCl3); H NMR (CDCl3,
500 MHz) δ 5.41 (1H, m, H-11), 5.29 (1H, m, H-7), 5.10 (1H, br d, J
) 10.0 Hz, H-24), 4.33 (1H, ddd, J ) 10.0, 10.0, 4.0 Hz, H-23), 2.74
(1H, ddd, J ) 14.5, 14.5, 5.0 Hz, H-2ꢀ), 2.25 (1H, ddd, J ) 14.5, 5.0,
5.0 Hz, H-2R), 1.77 (3H, s, H-27), 1.72 (3H, s, H-26), 1.70 (1H, m,
H-20), 1.11 (3H, s, H-29), 1.06 (3H, s, H-28), 1.02 (3H, s, H-30), 1.01
(3H, s, H-19), 0.95 (3H, d, J ) 6.0 Hz, H-21), 0.81 (3H, s, H-18);
FABMS m/z 439 [M + H]+.
Preparation of the (R)- and (S)-MTPA Ester Derivatives of
Compounds 7a and 9a (ref 19). To a stirred solution of 7a (2.0 mg)
in pyridine (400 µL) were added 4-(dimethylamino)pyridine (2 mg)
and (S)-(+)-R-methoxy-R-(trifluoromethyl)phenylacetyl chloride (MTPA-
Cl, 10 µL). The mixture was stirred at room temperature for 16 h. The
reaction mixture was then passed through a silica gel Waters Sep-Pak
Vac 6 cc and eluted with n-hexane-EtOAc (15:1) to give the respec-
tive (R)-Mosher ester 7r. Treatment of 7a (2.0 mg) with (R)-MTPA-
Cl (10 µL) as described above yielded the corresponding (S)-MTPA
ester 7s. Similarly, treatment of 9a with (S)- and (R)-MTPA-Cl afforded
the respective Mosher esters 9r and 9s.
In Vitro Cytotoxicity Test. A sulforhodamine B (SRB) bioassay
was used to determine the cytotoxicity of each compound against four
NP100660V