Thirteen novel cycloartane-type triterpenes from Combretum quadrangulare

Article abstract of DOI:10.1021/np990336q

Thirteen novel cycloartane-type triterpenes were isolated from Combretum quadrangulare, a Vietnamese medicinal plant. The structures of the novel triterpenes were determined by spectroscopic methods as well as by chemical transformations. Among those compounds, quadrangularic acids F (1), G (2), and H (4) and 24-epiquadrangularic acid G (3) are the first examples of cycloartane-type triterpenes bearing carboxylic acid groups at both C-4 and C-20. Furthermore, norquadrangularic acid A (13) is the first example of a trinorcycloartane-type triterpene isolated from the genus Combretum.

Full text of DOI:10.1021/np990336q

J . Nat. Prod. 2000, 63, 57-64
57
Th ir teen Novel Cycloa r ta n e-Typ e Tr iter p en es fr om Com br etu m qu a d r a n gu la r e
Arjun H. Banskota,† Yasuhiro Tezuka,† Kim Qui Tran,‡ Ken Tanaka,§ Ikuo Saiki,† and Shigetoshi Kadota*,†
Institute of Natural Medicine, Toyama Medical and Pharmaceutical University, 2630-Sugitani, Toyama 930-0194, J apan,
National University Ho Chi Minh City, Ho Chi Minh City, Vietnam, and National Research Institute of Police Science,
6 Sanban-cho, Chiyoda-ku, Tokyo 102-0075, J apan
Received J uly 9, 1999
Thirteen novel cycloartane-type triterpenes were isolated from Combretum quadrangulare, a Vietnamese
medicinal plant. The structures of the novel triterpenes were determined by spectroscopic methods as
well as by chemical transformations. Among those compounds, quadrangularic acids F (1), G (2), and H
(4) and 24-epiquadrangularic acid G (3) are the first examples of cycloartane-type triterpenes bearing
carboxylic acid groups at both C-4 and C-20. Furthermore, norquadrangularic acid A (13) is the first
example of a trinorcycloartane-type triterpene isolated from the genus Combretum.
Combretum species (Combretaceae) are widely used as
folk medicine for the treatment of hepatitis, malaria,
respiratory infections, and even cancer in different parts
of Asia and Africa.1 Combretum quadrangulare Kurz is an
evergreen tree that grows widely in eastern Asia. Its seeds,
leaves, and the stem bark are used in Vietnamese tradi-
tional medicine as an antipyretic, antidysenteric, and
anthelmintic agent.2 In the course of the chemical inves-
tigation on Vietnamese medicinal plants,3,4 we have re-
cently reported the isolation of seven novel cytotoxic
cycloartane-type triterpenes from C. quadrangulare.3 In
this paper we describe the isolation and structure elucida-
tion of 13 additional novel cycloartane-type triterpenes
(1-13) from a MeOH extract of the leaves of C. quadran-
gulare.
Resu lts a n d Discu ssion
Air-dried leaves of C. quadrangulare were extracted with
MeOH at 80 °C. The dark-green MeOH extract showed a
potent hepatoprotective effect on lipopolysaccharide-in-
F igu r e 1. Significant correlations observed in the FG-pulsed HMBC
spectrum of compounds 1, 2, and 5. Compounds 2 and 5 also showed
the same significant correlations in rings A-D as compound 1.
duced liver injury in D-galactosamine-sensitized mice in
vivo.5 Interestingly, the same extract also had a cytotoxic
effect toward the liver-metastatic murine colon 26-L5
carcinoma in vitro, with an ED50 value of 75.9 µg/mL.
Hence, the MeOH extract was further fractionated into 11
fractions by Si gel column chromatography. Repeated
chromatography of fractions 8 and 9 on normal- and
reversed-phase Si gel columns, together with preparative
TLC, afforded 13 novel triterpenes, named quadrangularic
acid F (1), quadrangularic acid G (2), 24-epiquadrangularic
acid G (3), quadrangularic acid H (4), methyl quadrangu-
larate I (5), quadrangularic acid J (6), quadrangularic acid
K (7), quadrangularic acid L (8), 24-epiquadrangularic acid
L (9), quadrangularic acid M (10), 24-epiquadrangularic
acid M (11), 7â-hydroxy-23-deoxojessic acid (12), and
norquadrangularic acid A (13).
a set of cyclopropane methylene protons (δ 0.76 and 0.48,
both d, J ) 4.5 Hz), two oxymethine protons (δ 5.36, dd, J
) 12.0, 4.5 Hz; δ 3.77, br s), five tertiary methyls (δ 1.60,
1.56, 1.50, 1.34, 1.04), an ester methyl (δ 3.66), and two
trans-olefinic protons (δ 6.18, d, J ) 16.0 Hz; δ 6.07, dt, J
) 16.0, 7.5 Hz), suggesting that 1 is a cycloartane-type
triterpene bearing two hydroxyls and a trans-olefin. By
treatment with diazomethane, 1 gave a dimethyl ester 1a ,
indicating the presence of a free carboxylic acid group. The
1H and 13C NMR data of 1 were similar to those of methyl
quadrangularate B (14),3 except for the absence of an
aldehyde signal and the appearance of an additional
carbonyl signal (δ 177.8) in the 13C NMR spectrum. Thus,
1 was considered to be the C-20 oxidized derivative of 14.
The presence of a carboxylic acid function at C-20 was
further confirmed by the long-range correlations observed
in the FG-pulsed HMBC spectrum (Figure 1).
The stereochemistry of 1 was determined by NOE
experiments and the analysis of coupling constants. The
lack of any diaxial coupling of H-1 with H-2ax indicated that
the hydroxyl group at C-1 is located in the axial position.
This was also supported by the NOE enhancement from
H-1 to H-19. The H-3 signal was observed as a double
doublet due to diaxial (J ) 12.0 Hz) and axial-equatorial
Quadrangularic acid F (1) was obtained as a colorless
amorphous solid, and its molecular formula was deter-
mined as C31H48O8 by HRFABMS. Absorption bands at
3400 and 1700 cm-1 in the IR spectrum of 1 indicated the
presence of hydroxyl and carbonyl groups, respectively. The
1H NMR spectrum of 1 displayed characteristic signals of
* To whom correspondence should be addressed. Tel.: 81-76-434-7625.
Fax: 81-76-434-5059. E-mail: kadota@ms.toyama-mpu.ac.jp.
† Institute of Natural Medicine, Toyama Medical and Pharmaceutical
University.
‡ National University, Ho Chi Minh City.
§ National Research Institute of Police Science.
10.1021/np990336q CCC: $19.00
© 2000 American Chemical Society and American Society of Pharmacognosy
Published on Web 11/24/1999

58 J ournal of Natural Products, 2000, Vol. 63, No. 1
Banskota et al.
Sch em e 1. ∆δRS () δR - δS) Values Obtained from the 1H NMR Spectra of the MTPA Esters 2c and 2d
1
coupling (J ) 4.5 Hz), suggesting the axial nature of H-3.
Irradiation of the methyl protons at δ 1.60 caused an NOE
increase of H-19 and vice versa, indicating that the methyl
group at C-4 should be in the â position, that is at C-29.
Additionally, irradiation of H-3 gave enhancement of H-5,
placing them in a 1,3-diaxial arrangement in a chair
conformation. Finally, the structure of quadrangularic acid
F, including the configuration at C-20, was confirmed to
be 1 by sodium chlorite oxidation6 of 14 to 1.
gested the molecular formula to be C30H44O7. The H and
13C NMR spectra of 4 were similar to those of 2 and 3, but
they showed a new signal for a ketone-carbonyl at δ 201.2
and the disappearance of the signal assignable to the
hydroxymethylene (C-24) found in 2 and 3. In addition,
correlations between the ketone carbon and the protons H2-
26 and H3-27 were observed in the long-range 1H-13C
COSY spectrum, suggesting that 4 should have a ketone
group at C-24 instead of a hydroxyl group as in 2 and 3.
This was further confirmed by esterification of 4 with
diazomethane to the R,â-unsaturated ketone 4a .
Quadrangularic acid G (2), a colorless amorphous solid,
was obtained as a monomethyl ester and was easily
converted into dimethyl ester 2a with diazomethane. The
molecular formula of 2 (C31H48O7) was calculated from the
quasimolecular ion peak [M + Na]+ at m/z 555.3272 in the
Quadrangularic acid I (5), a colorless amorphous solid,
showed a quasimolecular ion at m/z 541.3506 in the
HRFABMS, being consistent with the molecular formula
1
1
HRFABMS. In the H NMR spectrum of 2, the signals of
C31H50O6. The H NMR spectrum of 5 was similar to those
four tertiary methyls, an ester methyl, and two exo-olefinic
protons were observed, in addition to the signals of two
characteristic cyclopropane methylene protons. Further-
more, three signals of oxymethine protons at δ 5.33, 4.46,
of 2 and 3. Differences between 5 and 2 and 3 were
apparent only in the signals due to the ring D side chain.
In the 1H and 13C NMR spectra of 5, the signals of the
oxymethylene group appeared at δH 4.09 (dd, J ) 11.0, 3.0
Hz), δH 3.85 (dd, J ) 11.0, 5.0 Hz), and δC 62.0, instead of
a signal for a carboxylic acid group at C-20 as in 2 and 3.
These data and the long-range correlations between
H2-21 and C-20 in the FG-pulsed HMBC spectrum (Fig-
ure 1) indicated that there should be a hydroxymeth-
ylene group at C-20 in 5. The stereochemistry in rings A-D
of 5 was determined by NOE difference experiments and
found to be the same as 1, while that of C-24 was con-
1
and 3.75 in the H NMR spectrum suggested the presence
of three hydroxyl groups, which was confirmed by acety-
lation of the dimethyl ester 2a into a triacetate 2b. Among
the three hydroxyl groups, two were considered to be
located at C-1 (δ 3.75) and C-3 (δ 5.33) by comparing the
1H NMR spectrum with that of 1. Detailed analysis of the
1H-1H, 1H-13C, and long-range 1H-13C COSY spectra
indicated that the third hydroxyl group should be located
at C-24 and that the ester (δ 178.1) and carboxylic acid (δ
178.6) groups were at C-4 and C-20, respectively (Figure
1). These and other long-range correlations established the
planar structure of quadrangularic acid G (2).
1
cluded as S by comparing the H and 13C NMR data with
those for 2 (24S) and 3 (24R) (Table 1 and Experimental
Section).
Quadrangularic acid J (6), a colorless amorphous solid,
was isolated as an acid and gave a monomethyl ester 6a
with diazomethane. The molecular formula of 6 was
determined as C31H50O5 by HRFABMS, and its IR spec-
trum showed the presence of hydroxyl (3400 cm-1) and
The stereochemistry of rings A-D in 2 was determined
to be the same as 1, based on the coupling constants and
the result of NOE experiments. The configuration at the
chiral center C-24, on the other hand, was determined
through the NMR study of MTPA esters of the dimethyl
1
carbonyl (1700 cm-1) groups. The H and 13C NMR spectra
1
ester 2a . In the H NMR spectrum of the (R)-MTPA ester
of 6 showed almost identical signals to those of 1, but the
signals of a secondary methyl appeared at δH 0.96 (d, J )
6.5 Hz) and δC 18.6 instead of a signal due to a carboxylic
acid group in 1, suggesting that C-21 should be a methyl
group. The quaternary carbon signal assignable to C-25
was shifted to δ 74.8, indicating the presence of a methoxyl
group (δH 3.20, δC 50.1). These were further confirmed by
the FG-pulsed HMBC spectrum (Figure 2). From these
data and NOE difference experiments, the structure of
quadrangularic acid J was determined as 6.
2c, H2-26 and H3-27 appeared shielded, whereas H2-23 and
H2-22 were deshielded, in comparison to analogous data
for (S)-MTPA ester 2d (Scheme 1). Thus, H2-26 and H3-27
in the (R)-MTPA ester 2c were more affected by the phenyl
ring of the MTPA part; that is, the configuration at C-24
should be S.7,8 The configuration at C-20 of 2 was assumed
to be the same as that of 1 because both compounds were
isolated from the same plant part.
24-Epiquadrangularic acid G (3), a colorless amorphous
solid, showed the same molecular formula as 2 (C31H48O7)
Quadrangularic acid K (7) was isolated as a colorless
amorphous solid, and its molecular formula was deter-
mined as C30H48O5 by HRFABMS. The 1H and 13C NMR
data (Table 1 and Experimental Section) of 7 were similar
to those of 6, and correlations observed in the long-range
1H-13C COSY spectrum (Figure 2) indicated that 7 has a
hydroxyl group instead of a methoxyl group at C-25 as in
1
in the HRFABMS. The H and 13C NMR spectra of 3 were
similar to those of 2, except for a slight downfield shift of
one of the olefinic protons (3, δ 5.31; 2, δ 5.20) and small
differences in the carbon chemical shifts from C-24 to C-27
(Table 1). Thus, 2 and 3 were considered to be epimers at
C-24; that is, 3 has a 24R configuration. This was confirmed
by the fact that their methyl esters 2a and 3a gave the
same R,â-unsaturated ketone 4a by MnO2 oxidation.9
Quadrangularic acid H (4) was also obtained as a
colorless amorphous solid, and the HRFABMS data sug-
1
6. In the H NMR spectrum of 7, the signals of two olefinic
protons appeared as a broad singlet at δ 5.94, suggesting
a cis configuration of the double bond. The cis nature of
the double bond was further supported by comparison of

Triterpenes from Combretum
J ournal of Natural Products, 2000, Vol. 63, No. 1 59
Ta ble 1. 13C NMR Data (100 MHz) of Compounds 1-13 in Pyridine-d5
position
1
2
3
4
5
6
7
8
9
10
11
12
13
1
2
3
4
5
6
7
8
72.1
38.6
70.4
56.0
37.9
23.2
26.0
47.8
20.8
30.3
25.7
30.6
45.7
48.9
35.4
27.3
49.2
18.1
29.6
49.5
177.8
37.8
127.5
137.5
81.1
25.3
24.9
178.1
9.4
72.1
38.5
70.4
56.0
37.8
23.1
27.5
47.8
20.8
30.2
26.0
30.6
45.6
48.9
34.0
25.7
49.2
18.1
29.6
49.6
178.6
29.5
35.4
75.6
149.4
110.6
17.5
178.1
9.4
72.1
38.5
70.4
56.0
37.8
23.2
27.5
47.9
20.8
30.3
26.0
30.6
45.7
48.9
33.9
25.7
49.0
18.3
29.8
49.6
178.5
29.2
35.4
74.8
149.6
110.0
18.1
178.1
9.5
72.3
38.5
70.6
55.5
37.5
23.1
27.2
47.8
20.7
30.3
25.7
30.5
45.5
48.9
35.2
25.6
49.4
18.0
29.6
48.5
178.5
27.6
35.6
201.2
144.3
124.6
17.5
180.1
9.6
72.2
38.6
70.4
56.0
37.9
23.4
27.7
48.3
20.9
30.1
26.2
32.5
45.5
49.1
35.9
25.8
47.0
18.7
29.7
43.6
62.0
26.4
32.4
76.2
149.5
110.2
17.9
178.1
9.5
72.5
38.8
70.7
55.7
37.7
23.4
28.3
48.1
20.8
30.3
26.2
33.2
45.5
49.2
35.9
25.7
52.2
18.4
29.7
36.6
18.6
39.6
128.3
137.5
74.8
26.5
26.0
180.0
9.7
72.4
38.6
70.5
55.5
37.6
23.3
28.1
48.0
20.7
29.8
26.0
33.0
45.4
48.9
35.7
25.7
52.1
18.3
29.6
36.7
18.4
39.3
124.4
141.3
69.6
30.6
30.0
180.0
9.6
72.6
38.8
70.7
55.7
37.7
23.4
28.4
48.2
20.8
30.3
26.2
33.3
45.5
49.1
35.9
25.8
52.9
18.4
29.8
36.3
18.6
34.1
28.9
79.0
72.7
26.1
25.9
180.0
9.7
75.5
38.8
70.7
55.7
37.7
23.4
28.3
48.2
20.8
30.3
26.2
33.3
45.5
49.1
35.9
25.9
52.8
18.4
29.8
36.3
18.8
34.5
29.4
79.8
72.7
26.1
26.0
180.0
9.7
72.5
38.8
70.7
55.7
37.7
23.4
28.4
48.2
20.8
30.3
26.2
33.3
45.5
49.1
36.3
25.9
52.6
18.4
29.8
36.4
18.7
32.7
35.9
75.6
149.6
110.0
18.2
180.1
9.8
72.5
38.8
70.7
55.7
37.7
23.4
28.4
48.2
20.8
30.3
26.2
33.3
45.5
49.1
36.3
25.9
52.6
18.4
29.8
36.4
18.7
32.7
35.9
76.1
149.6
110.4
17.7
180.1
9.8
72.5
38.8
70.5
55.4
36.7
34.0
69.5
54.9
20.9
30.7
26.7
33.3
46.0
49.2
37.5
28.8
52.0
17.7
27.8
36.4
18.7
35.4
31.6
156.7
34.1
22.0
21.9
179.9
9.7
72.5
37.7
70.7
55.7
38.8
23.4
28.2
48.1
20.8
30.3
26.2
33.2
45.5
49.1
35.8
25.9
52.5
18.3
29.7
36.0
18.1
32.1
32.0
176.5
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
25
27
28
29
30
31
MeO-25
MeO-28
180.0
9.7
19.4
19.4
19.4
19.4
19.3
19.7
19.4
19.3
19.5
19.5
19.5
19.5
19.0
106.6
50.1
51.4
51.4
51.4
51.5
Quadrangularic acid L (8), a colorless amorphous solid
with the molecular formula C30H50O6, showed hydroxyl and
carbonyl group absorption in the IR spectrum. The 1H NMR
1
spectrum of 8, analyzed with the aid of the H-1H COSY
spectrum, showed signals due to a cyclopropane methylene,
three oxymethines, five tertiary methyls, and a secondary
methyl. Based on a comparison of the 1H and 13C NMR
data with those of 7, 8 was considered to be a C-24 hydroxyl
derivative, which was confirmed by the long-range 1H-
13C COSY spectrum (Figure 2). The orientation of the
hydroxyl groups at C-1 and C-3 in 8 were concluded to be
1
R and â, respectively, by comparison of the H NMR data
with those of 1-7. The stereochemistry of the rest of the
molecule was determined by NOE experiments. For the
determination of the absolute configuration at C-24, the
(R)-MTPA ester 8b and the (S)-MTPA ester 8c were
prepared from the methyl ester 8a . The ∆δRS () δR - δS)
values of H2-23, H3-26, and H3-27, however, were all
negative, and thus the advanced Mosher’s method7 could
not be applied. This could result from their unusual
conformation due to the presence of the C-25 hydroxyl
group. Thus, the MTPA esters 8b and 8c were dehydrated
with POCl3-pyridine11 into the respective olefins, 10b and
10c (Scheme 2). In the case of 10b and 10c, H2-26 and H3-
F igu r e 2. Significant correlations observed in the long-range 1H-
13C COSY spectrum of compounds 6, 7, and 8. Compounds 7 and 8
also showed the same significant correlations in rings A-D as com-
pound 6.
the chemical shifts of methyl ester 7a in CDCl3 (C-23, δ
125.5; C-24, δ 139.4) with those of (23Z)-3â-acetoxycycloart-
23-en-25-ol, a compound having the same side chain (C-
23, δ 125.6; C-24, δ 139.3) as 7.10
Sch em e 2. ∆δRS () δR-δS) Values Obtained from the MTPA Esters of Methyl Quadrangularate L (8A) and Methyl Quadrangularate
M (10A)

60 J ournal of Natural Products, 2000, Vol. 63, No. 1
Banskota et al.
1H-1H COSY analysis and was further confirmed by the
long-range correlations observed in the FG-pulsed HMBC
spectrum (Figure 3a).
F igu r e 3. (a) Significant correlations observed in the FG-pulsed
HMBC spectrum of compound 12, and (b) NOEs observed in the NOE
difference experiments of compound 12.
27 of (R)-MTPA ester 10b resonated downfield as compared
to (S)-MTPA ester 10c, while H2-23 of 10b resonated
upfield when compared to 10c. This indicated that H2-23
in the (R)-MTPA ester 10b was more affected by the phenyl
ring of the MTPA part; that is, C-24 should have an R
configuration.7,8 The structure of quadrangularic acid L was
thus determined as 8.
24-Epiquadrangularic acid L (9), a colorless amorphous
solid, had the same molecular formula C30H50O6 as 8. The
1
IR, H NMR, and 13C NMR spectra of 9 were almost the
same as those of 8, but a slight difference was found in
the chemical shifts of H-24 (9, δ 3.71; 8, δ 3.76) and C-24
1
(9, δ 79.8; 8, δ 79.0). Analysis of the H-1H COSY, HMQC,
and HMBC spectra indicated that 9 had the same planar
structure as 8, while the NOE spectra revealed the pres-
ence of the same stereochemistry on rings A-D as 8. Thus,
9 was concluded to be the 24S epimer of 8.
Quadrangularic acid M (10) and 24-epiquadrangularic
acid M (11) were obtained as an epimeric mixture (10:11
) 4:3 from the 1H NMR spectrum), and their molecular
formulas were determined as C30H48O5 based on FABMS.
Analysis of the 1H and 13C NMR spectra of the mixture
through the 1H-1H, 1H-13C, and long-range 1H-13C COSY
spectra enabled the assignment of the signals to each
epimer. The assigned data of 10 and 11 differed only in
the chemical shift of one of the olefinic protons (10, δ 5.27;
1
11, δ 5.22) in the H NMR spectrum, while in the 13C NMR
spectrum, they clearly differed at three signals, C-24, C-26,
and C-27 (Table 1). Thus, they were considered to be
1
epimers at C-24, and, by comparing their H and 13C NMR
data with those of 2 and 3, 10 and 11 were assigned with
the 24R and 24S configuration, respectively. This was
confirmed by the fact that methylation with diazomethane,
followed by esterification with (R)-MTPA chloride, gave a
mixture of 10b and its 24S epimer (10c) (4:3).
7â-Hydroxy-23-deoxojessic acid (12) was isolated as
colorless crystals having a melting point of 219 °C, and
its molecular formula was determined as C31H50O5 by
HRFABMS. The IR spectrum of 12 showed a broad absorp-
tion band at 3400 cm-1 and a sharp absorption band at
1700 cm-1, suggesting the presence of hydroxyl and car-
bonyl groups, respectively. The 1H and 13C NMR data of
12 were similar to those of 23-deoxojessic acid3 (15) except
that 12 had one more hydroxyl group. The presence of three
free hydroxyl groups was confirmed by acetylation of the
methyl ester 12a to a triacetate 12b. The position of the
additional hydroxyl group was determined to be at C-7 by
The configurations of the hydroxyl groups at C-1 and C-3
were concluded to be R and â, respectively, on the basis of
coupling constants of H-1 (br s) and H-3 (dd, J ) 12.0, 4.5
Hz). The diaxial coupling of H-7 with H-6ax (11.8 Hz) and
H-8ax (8.5 Hz) suggested the â configuration of OH-7. These
were further confirmed by NOEs observed in NOE differ-

Triterpenes from Combretum
J ournal of Natural Products, 2000, Vol. 63, No. 1 61
Sch em e 3. Possible Biogenetic Pathway of 1-13 from the Hypothetical Precursor 16 or Mollic Acid (17)
ence experiments (Figure 3b). The cyclopropane methylene
protons of 7â-hydroxycycloartane-type triterpenes have
been reported to appear in the usual range; that is, δ 0.35-
0.39 and δ 0.66-0.70.12,13 It should be noted here that,
contrary to previous reports, these protons in 12 were
deshielded to resonate at δ 0.55 and 1.11.
Norquadrangularic acid A (13) was obtained as a color-
less amorphous solid and gave a dimethyl ester 13a by
methylation with diazomethane. It showed a quasimolecu-
lar ion peak at m/z 485.2883 in HRFABMS corresponding
to the molecular formula C27H42O6. The IR spectrum of 13
suggested the presence of hydroxyl (3400 cm-1) and car-
bonyl group (1700 cm-1) absorptions. The 1H NMR spec-
trum of 13 displayed the signals of two cyclopropane
methylene protons at δ 0.82 and 0.54 (both d, J ) 4.5 Hz)
along with three tertiary methyls, one secondary methyl,
and two oxymethine protons. These were identical to those
of the other cycloartane-type triterpenes isolated from C.
quadrangulare. The 13C NMR spectrum of 13, however,
displayed only 27 carbon signals, and thus 13 was assigned
as a trinorcycloartane-type triterpene. The 13C NMR
spectrum of 13 showed signals of two carbonyl carbons (δ
176.5 and 180.0) due to two free carboxylic acid groups.
F igu r e 4. Significant correlations observed in the FG-pulsed HMBC
spectrum of compound 13.
range correlations observed in the FG-pulsed HMBC
spectrum (Figure 4), the positions of the carboxylic acid
groups were determined as C-23 and C-4. The positions of
the two hydroxyl groups were determined as C-1R and C-3â
by comparing the chemical shifts and the coupling con-
stants of H-1 (δ 3.91, br s) and H-3 (δ 5.57, dd, J ) 12.0,
4.5 Hz) with those of 1-12 and were consistent with the
results of the NOE difference experiments. Accordingly, the
1
On the basis of the H-13C COSY spectrum and the long-

62 J ournal of Natural Products, 2000, Vol. 63, No. 1
Banskota et al.
MeOH-CHCl3 eluate, 15.0 g; fraction 5, 5% MeOH-CHCl3
eluate, 39.4 g; fraction 6, 5% MeOH-CHCl3 eluate, 23.2 g;
fraction 7, 10% MeOH-CHCl3 eluate, 10.2 g; fraction 8, 10%
MeOH-CHCl3 eluate, 12.9 g; fraction 9, 20% MeOH-CHCl3
eluate, 23.2 g; fraction 10, 30% MeOH-CHCl3 eluate, 45.7 g,
and fraction 11, 50% MeOH-CHCl3 eluate, 84.5 g).
structure of norquadrangularic acid A was established as
13.
The cycloartane-type triterpenes (1-13) are all new and
are characterized by the presence of 1R,3â-dihydroxy and
28-carboxyl groups, which seems to be typical for Combre-
tum species among this class of compounds.14-18 Previous
literature reports reveal the presence of a carboxylic acid
group either at C-414-18 or at C-20 in cycloartane-type
triterpenes.19-21 Compounds 1-4, however, have two car-
boxylic acid groups at both C-4 and C-20. Furthermore,
norquadrangularic acid A (13) is a trinorcycloartane-type
triterpene. Previously, this type of triterpene was reported
from Wrightia tinctoria22 and Euphorbia broteri,23 and the
commercial drug “Cimicifuga Rhizoma”,24 but there is no
previous report of their presence in Combretum species.
In our previous work, we observed that few of the cycloar-
tane-type triterpenes isolated from C. quadrangulare pos-
sessed potent cytotoxicity toward the liver-metastatic
murine colon 26-L5 carcinoma cells.3,28 The triterpenes
Fractions 7 and 8 were combined (23.0 g) and chromato-
graphed on a Cosmosil 75C18-OPN with H2O-MeOH-CH3CN
(1:1:1) to give 12 subfractions. Further Si gel column chroma-
tography and preparative TLC of subfraction 3 yielded qua-
drangularic acid F (1, 9.1 mg). Fraction 9 (20.0 g) was also
applied on a Cosmosil 75C18-OPN column with H2O-MeOH-
CH3CN (1:1:1) and eight subfractions were collected. Further
Si gel column chromatography and preparative TLC of the
subfractions 2-7 yielded the following compounds: fraction
2, norquadrangularic acid A (13, 20.6 mg), quadrangularic acid
G (2, 120.2 mg), quadrangularic acid H (4, 15.5 mg), 24-
epiquadrangularic acid L (9, 32.0 mg); fraction 3, 24-epiqua-
drangularic acid G (3, 74.3 mg), methyl quadrangularate I (5,
24.0 mg), quadrangularic acid L (8, 32.0 mg); fraction 5, a
mixture of quadrangularic acid M (10) and 24-epiquadrangu-
laric acid M (11) (63.3 mg); fraction 7, quadrangularic acid J
(6, 73.0 mg), quadrangularic acid K (7, 15.5 mg), and 7â-
hydroxy-23-deoxojessic acid (12, 62.1 mg).
1-13, however, showed only very weak cytotoxicity (ED50
:
5, 29.4; 6, 86.1; 7, 82.6; 12, 37.3; mixture of 10 and 11,
88.4 µg/mL; others >100 µg/mL).
Qu a d r a n gu la r ic a cid F (1): colorless amorphous solid;
Most of the cycloartane-type triterpenes isolated from
C. quadrangulare differ in the side chain attached to ring
D. These compounds might be biosynthesized via photo-
oxygenation of olefinic precursors such as 16 or mollic acid14
(17) (Scheme 3). It is well-known that molecular oxygen
reacts with olefins to form allylic hydroperoxides.25-27
When the methyl proton of olefin 16 or 17 takes part in
the reaction, a hydroperoxy group will be generated at
C-24, which, on reduction, gives 2 and 3 (by 16) or 10 and
11 (by 17). If a methylene proton is involved, a hydroper-
oxide is generated at C-25. Then, 14 will be formed from
16, which on oxidation gives 1 or on reduction gives methyl
quadrangularate B (18).2 By photooxygenation dioxetane
may also be formed25 and would lead to diols 8 and 9
through a reduction of the O-O bond or to trinorcycloar-
tane 13 through a cleavage of O-O and C-C bonds.
[R]25 +15.7° (c 0.03, MeOH); IR νmax (KBr) 3400, 1720, 1440,
D
1250 cm-1; H NMR (pyridine-d5) δ 6.18 (1H, d, J ) 16.0 Hz,
1
H-24), 6.07 (1H, dt, J ) 16.0, 6.0 Hz, H-23), 5.36 (1H, dd, J )
12.0, 4.5 Hz, H-3), 3.77 (1H, br s, H-1), 3.66 (3H, s, MeO-28),
3.23 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.40 (1H, ddd, J ) 13.0,
4.5, 4.0 Hz, H-2), 2.19 (1H, ddd, J ) 13.0, 12.0, 3.5 Hz, H-2),
1.60 (3H, s, H3-29), 1.56 (3H, s, H3-26), 1.50 (3H, s, H3-27),
1.34 (3H, s, H3-18), 1.04 (3H, s, H3-30), 0.76 (1H, d, J ) 4.5
Hz, H-19), 0.48 (1H, d, J ) 4.5 Hz, H-19); HRFABMS m/z
571.3239 (calcd for C31H48O8Na [M + Na]+, 571.3247).
Qu a d r a n gu la r ic a cid G (2): colorless amorphous solid;
[R]25 +73.4° (c 0.09, MeOH); IR νmax (KBr) 3450, 1700, 1260,
D
1040 cm-1; 1H NMR (pyridine-d5) δ 5.33 (1H, dd, J ) 12.0, 4.0
Hz, H-3), 5.20 (1H, br s, H-26), 4.93 (1H, br s, H-26), 4.46 (1H,
t, J ) 5.0 Hz, H-24), 3.75 (1H, br s, H-1), 3.64 (3H, s, MeO-
28), 3.21 (1H, dd, J ) 12.5, 4.5 Hz, H-5), 2.66 (2H, m, H-17,
H-11), 2.52 (1H, m, H-20), 2.37 (1H, dt, J ) 13.0, 4.0 Hz, H-2),
2.17 (1H, ddd, J ) 13.0, 12.0, 3.5 Hz, H-2), 1.89 (3H, s, H3-
27), 1.63 (1H, br t, J ) 8.0 Hz, H-8), 1.58 (3H, s, H3-29), 1.35
(3H, s, H3-18), 1.03 (3H, s, H3-30), 0.75 (1H, d, J ) 4.5 Hz,
H-19), 0.41 (1H, d, J ) 4.5 Hz, H-19); HRFABMS m/z 555.3272
(calcd for C31H48O7Na [M + Na]+, 555.3298).
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. Melting points were
determined on a Yanaco micromelting point apparatus and
are uncorrected. Optical rotations were recorded on a J ASCO
DIP-140 digital polarometer. IR spectra were measured with
a Shimadzu IR-408 spectrophotometer in KBr disks. NMR
spectra were taken on a J EOL GX-400 spectrometer or a J EOL
J NM-LA400WB spectrometer with tetramethylsilane (TMS)
as the internal standard, and chemical shifts are expressed
in δ values. HRFABMS measurements were carried out on a
J EOL J MS-700T spectrometer, and glycerol was used as a
matrix. Column chromatography was performed with normal-
phase (Fuji Silysia, BW-820 MH) or reversed-phase Si gel
(Cosmosil 75C18-OPN, Nacalai Tesque Inc., Kyoto, J apan).
Analytical and preparative TLC were carried out on precoated
Merck Kieselgel 60F254 plates (0.25 or 0.50 mm thickness).
24-Ep iqu a d r a n gu la r ic a cid G (3): colorless amorphous
solid; [R]25D +103.5° (c 0.05, MeOH); IR νmax (KBr) 3450, 1700,
1440, 1250 cm-1 1H NMR (pyridine-d5) δ 5.33 (1H, dd, J )
;
12.0, 4.0 Hz, H-3), 5.31 (1H, br s, H-26), 4.95 (1H, br s, H-26),
4.50 (1H, t, J ) 5.0 Hz, H-24), 3.75 (1H, br s, H-1), 3.64 (3H,
s, MeO-28), 3.21 (1H, dd, J ) 12.5, 4.5 Hz, H-5), 2.72 (1H, m,
H-17), 2.65 (1H, m, H-11), 2.52 (1H, m, H-20), 2.38 (1H, dt, J
) 13.0, 4.0 Hz, H-2), 2.16 (1H, ddd, J ) 13.0, 12.0, 3.5 Hz,
H-2), 1.89 (3H, s, H3-27), 1.58 (3H, s, H3-29), 1.35 (3H, s, H3-
18), 1.03 (3H, s, H3-30), 0.75 (1H, d, J ) 4.5 Hz, H-19), 0.41
(1H, d, J ) 4.5 Hz, H-19); HRFABMS m/z 555.3272 (calcd for
C
31H48O7Na [M + Na]+, 555.3298).
Qu a d r a n gu la r ic a cid H (4): colorless amorphous solid;
[R]25 +14.3° (c 0.03, MeOH); IR νmax (KBr) 3450, 1710, 1450,
D
1040 cm-1; 1H NMR (pyridine-d5) δ 5.97 (1H, br s, H-26), 5.64
(1H, br s, H-26), 5.52 (1H, dd, J ) 12.5, 4.5 Hz, H-3), 3.79
(1H, br s, H-1), 3.37 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.98 (2H,
m, H2-23), 2.65 (2H, m, H-11, H-20), 2.47 (2H, m, H-2, H-17),
2.24 (1H, ddd, J ) 12.5, 12.0, 3.0 Hz, H-2), 1.86 (3H, s, H3-
27), 1.69 (3H, s, H3-29), 1.35 (3H, s, H3-18), 1.04 (3H, s, H3-
30), 0.80 (1H, d, J ) 4.5 Hz, H-19), 0.42 (1H, d, J ) 4.5 Hz,
H-19); HRFABMS m/z 539.2980 (calcd for C30H44O7Na [M +
Na]+, 539.2984).
P la n t Ma ter ia l. Leaves of Combretum quadrangulare Kurz
were purchased at a local market at Ho Chi Minh City,
Vietnam, in 1995. A voucher sample (TMPW 18999) is
preserved in the Museum for Materia Medica, Toyama Medical
and Pharmaceutical University, Toyama, J apan, as a refer-
ence.
Extr a ction a n d Isola tion . Air-dried leaves (2.65 kg) were
extracted with MeOH (16L, 3 h × 3) at 80 °C. The filtrate was
evaporated under reduced pressure to yield a dark green
MeOH extract (610 g). A part of the MeOH extract (400 g)
Meth yl qu a d r a n gu la r a te I (5): colorless amorphous solid;
was chromatographed over Si gel with
a CHCl3-MeOH
[R]25D +137.0° (c 0.02, MeOH); IR νmax (KBr) 3400, 1710, 1450
1
gradient system to give 11 fractions (fraction 1, 2% MeOH-
CHCl3 eluate, 6.1 g; fraction 2, 2% MeOH-CHCl3 eluate, 11.3
g; fraction 3, 5% MeOH-CHCl3 eluate, 20.4 g; fraction 4, 5%
cm-1; H NMR (pyridine-d5) δ 5.37 (1H, dd, J ) 12.5, 4.5 Hz,
H-3), 5.23 (1H, br s, H-26), 4.95 (1H, br s, H-26), 4.42 (1H, t,
J ) 5.0 Hz, H-24), 4.09 (1H, dd, J ) 11.0, 3.0 Hz, H-21), 3.85

Triterpenes from Combretum
J ournal of Natural Products, 2000, Vol. 63, No. 1 63
(1H, dd, J ) 11.0, 5.0 Hz, H-21), 3.84 (1H, br s, H-1), 3.65
(3H, s, MeO-28), 3.24 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.75
(1H, ddd, J ) 13.0, 8.0, 4.0, H-11), 2.42 (1H, ddd, J ) 13.0,
4.5, 4.0 Hz, H-2), 2.21 (2H, m, H-2, H-17), 1.94 (3H, s, H3-27),
1.61 (3H, s, H3-29), 1.12 (3H, s, H3-18), 1.01 (3H, s, H3-30),
0.76 (1H, d, J ) 4.5 Hz, H-19), 0.50 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 541.3502 (calcd for C31H50O6Na [M + Na]+,
541.3506).
J ) 12.0, 4.5 Hz, H-3), 4.86 (1H, br s, H-31), 4.85 (1H, br s,
H-31), 4.12 (1H, ddd, J ) 11.0, 8.5, 4.0 Hz, H-7), 4.00 (1H, br
s, H-1), 3.71 (1H, dd, J ) 12.5, 4.5 Hz, H-5), 2.63 (1H, ddd, J
) 13.0, 8.0, 4.0 Hz, H-11), 2.54 (1H, ddd, J ) 13.0, 4.5, 4.0
Hz, H-2), 2.34 (1H, ddd, J ) 13.0, 12.0, 3.5 Hz, H-2), 2.10 (1H,
d, J ) 8.5 Hz, H-8), 1.77 (3H, s, H3-29), 1.31 (3H, s, H3-30),
1.15 (3H, s, H3-18), 1.11 (1H, d, J ) 4.5 Hz, H-19), 1.06 (3H,
d, J ) 7.0 Hz, H3-26), 1.05 (3H, d, J ) 7.0 Hz, H3-27), 0.98
(3H, d, J ) 5.0 Hz, H3-21), 0.55 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 525.3593 (calcd for C31H50O5Na [M + Na]+,
525.3556).
Qu a d r a n gu la r ic a cid J (6): colorless amorphous solid;
[R]25 +8.4° (c 0.02, MeOH); IR νmax (KBr) 3400, 1710, 1450
D
cm-1; 1H NMR (pyridine-d5) δ 5.65 (1H, ddd, J ) 15.5, 8.5, 6.0
Hz, H-23), 5.54 (2H, m, H-3, H-24), 3.90 (1H, br s, H-1), 3.40
(1H, dd, J ) 12.0, 4.5 Hz, H-5), 3.20 (3H, s, MeO-25), 2.74
(1H, ddd, J ) 13.0, 9.0, 8.0 Hz, H-11), 2.48 (1H, dt, J ) 13.0,
4.0 Hz, H-2), 2.28 (2H, m, H-2, H-22), 1.72 (3H, s, H3-29), 1.32
(6H, s, H3-26, H3-27), 1.05 (3H, s, H3-18), 0.99 (3H, s, H3-30),
0.96 (3H, d, J ) 6.5 Hz, H3-21), 0.84 (1H, d, J ) 4.5 Hz, H-19),
0.55 (1H, d, J ) 4.5 Hz, H-19); HRFABMS m/z 525.3524 (calcd
for C31H50O5Na [M + Na]+, 525.3556).
Nor qu a d r a n gu la r ic a cid A (13): colorless amorphous
solid; [R]25D +200.6° (c 0.01, MeOH); IR νmax (KBr) 3400, 1710,
1550, 1470 cm-1 1H NMR (pyridine-d5) δ 5.57 (1H, dd, J )
;
12.0, 4.5 Hz, H-3), 3.91 (1H, br s, H-1), 3.43 (1H, dd, J ) 11.5,
4.5 Hz, H-5), 2.74 (1H, m, H-11), 2.62 (1H, m, H-23), 2.52 (1H,
m, H-23), 2.50 (1H, m, H-2), 2.30 (1H, ddd, J ) 12.5, 11.0, 2.0
Hz, H-2), 2.10 (1H, m, H-22), 1.92 (1H, m, H-7), 1.74 (3H, s,
H3-29), 1.03 (3H, s, H3-18), 0.98 (3H, s, H3-30), 0.95 (3H, d, J
) 5.0 Hz, H3-21), 0.82 (1H, d, J ) 4.5 Hz, H-19), 0.54 (1H, d,
Qu a d r a n gu la r ic a cid K (7): colorless amorphous solid;
[R]25D +133.7° (c 0.03, MeOH); IR νmax (KBr) 3450, 1700, 1460,
1380 cm-1; 1H NMR (pyridine-d5) δ 5.94 (2H, m, H-23, H-24),
5.57 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 3.92 (1H, br s, H-1), 3.41
(1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.74 (1H, ddd, J ) 13.0, 9.0,
8.0 Hz, H-11), 2.52 (1H, ddd, J ) 13.0, 4.5, 4.0 Hz, H-2), 2.29
(2H, m, H-2, H-23), 1.73 (3H, s, H3-29), 1.55 (6H, s, H3-26, H3-
27), 1.04 (3H, s, H3-18), 0.98 (3H, s, H3-30), 0.95 (3H, d, J )
6.5 Hz, H3-21), 0.83 (1H, d, J ) 4.5 Hz, H-19), 0.55 (1H, d, J
) 4.5 Hz, H-19); HRFABMS m/z 511.3392 (calcd for C30H48O5-
Na [M + Na]+, 511.3400).
Qu a d r a n gu la r ic a cid L (8): colorless amorphous solid;
[R]25D +100.4° (c 0.03, MeOH); IR νmax (KBr) 3450, 1700, 1370,
1040 cm-1; 1H NMR (pyridine-d5) δ 5.56 (1H, dd, J ) 12.0, 4.5
Hz, H-3), 3.91 (1H, br s, H-1), 3.76 (1H, dd, J ) 8.0, 2.5 Hz,
H-24), 3.42 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.75 (1H, ddd, J )
13.0, 9.0, 8.0 Hz, H-11), 2.50 (1H, ddd, J ) 12.5, 4.5, 4.0 Hz,
H-2), 2.29 (1H, ddd, J ) 12.5, 12.0, 2.0 Hz, H-2), 1.73 (3H, s,
H3-29), 1.54 (3H, s, H3-26), 1.52 (3H, s, H3-27), 1.06 (3H, s,
H3-18), 1.00 (3H, d, J ) 6.5 Hz, H3-21), 0.97 (3H, s, H3-30),
0.83 (1H, d, J ) 4.5 Hz, H-19), 0.55 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 529.3476 (calcd for C30H50O6Na [M + Na]+,
529.3495).
J
C
) 4.5 Hz, H-19); HRFABMS m/z 485.2883 (calcd for
27H42O6Na [M + Na]+, 485.2879).
Oxid a tion of Meth yl Qu a d r a n gu la r a te B (14) to Qu a -
d r a n gu la r ic a cid F (1). To a stirred solution of 14 (2 mg,
3.75 µmol) in a mixture of CH3CN (0.5 mL), aqueous NaH2-
PO4 (0.1 mg/mL, 0.5 mL), 30% H2O2 (40 µL), and an aqueous
solution of NaClO2 (0.4 mg/mL, 125 µL) were added dropwise
at 10 °C, and the mixture was stirred for 2 h at 10 °C. After
Na2SO3 (1 mg) was added, the mixture was subjected to
preparative TLC with 20% MeOH-CHCl3 to yield 1 (1.2 mg,
58.4%).
Oxid a tion of Dim eth yl Qu a d r a n gu la r a te G (2a ) a n d
Dim eth yl 24-Ep iqu a d r a n gu la r a te (3a ) to Dim eth yl Qu a -
d r a n gu la r a te H (4a ). To a stirred solution of 2a (10 mg) in
CHCl3 (2 mL), MnO2 (200 mg) was added, and the mixture
was stirred for 24 h at room temperature. The precipitate was
filtered off, and the filtrate was purified by preparative TLC
with MeOH-CHCl3 (1:9) to give 4a (2.7 mg, 26.8%). By the
same procedure, 3a (2.0 mg) also gave 4a (0.4 mg, 20.0%).
P r ep a r a tion of (R)- a n d (S)-MTP A Ester s of Dim eth yl
Qu a d r a n gu la r a te G (2a ) a n d Meth yl Qu a d r a n gu la r a te
L (8a ). To a solution of 2a (10 mg) in CHCl3 (0.5 mL) and
pyridine (0.5 mL), (R)-MTPA-Cl (100 µL) was added, and the
mixture was stirred overnight at room temperature. The
reaction mixture was then directly purified by preparative TLC
with MeOH-CHCl3 (1:19) to give (R)-MTPA ester 2c (15.4 mg;
70.4%). By the same procedure, the (S)-MTPA ester 2d (14.4
mg, 65.9%) and the (R)- and (S)-MTPA esters of 8a , 8b (3.7
mg, 47.8%), and 8c (3.5 mg, 38.9%) were prepared.
(R)-MTP A ester of d im eth yl qu a d r a n gu la r a te G (2c):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.59-7.30 (15H,
m, Ph-H × 3), 5.69 (1H, dd, J ) 12.5, 4.5 Hz, H-3), 5.27 (1H,
t, J ) 5.5 Hz, H-24), 5.10 (1H, br s, H-26), 4.87 (1H, br s, H-26),
4.78 (1H, br s, H-1), 3.58 (3H, s, MeO-21), 3.49 (3H, s, MeO-
28), 3.60, 3.47, 3.40 (each 3H, s, OMe × 3), 2.45 (1H, ddd, J )
13.0, 4.5, 4.0 Hz, H-2), 2.41 (1H, dd, J ) 12.0, 4.0 Hz, H-5),
2.19 (1H, td, J ) 10.0, 3.0 Hz, H-22), 2.00 (1H, br t, J ) 10.0
Hz, H-22), 1.90 (1H, ddd, J ) 13.0, 12.5, 3.5 Hz, H-2), 1.60
(1H, m, H-23), 1.50 (3H, s, H3-27), 1.48 (1H, m, H-23), 1.08
(3H, s, H3-29), 0.87 (3H, s, H3-18), 0.73 (1H, d, J ) 4.5 Hz,
H-19), 0.53 (1H, d, J ) 4.5 Hz, H-19), 0.52 (3H, s, H3-30);
HRFABMS m/z 1217.4624 (calcd for C62H71F9O13Na [M + Na]+,
1217.4649).
24-Ep iqu a d r a n gu la r ic a cid L (9): colorless amorphous
solid; [R]25 +76.2° (c 0.08, MeOH); IR νmax (KBr) 3450, 1700,
D
1470, 1380, 1050 cm-1; H NMR (pyridine-d5) δ 5.57 (1H, dd,
1
J ) 12.0, 4.5 Hz, H-3), 3.92 (1H, br s, H-1), 3.71 (1H, dd, J )
10.0, 2.0 Hz, H-24), 3.43 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.75
(1H, ddd, J ) 13.0, 9.0, 8.0 Hz, H-11), 2.50 (1H, ddd, J ) 12.5,
4.5, 4.0 Hz, H-2), 2.30 (1H, ddd, J ) 12.5, 12.0, 2.0 Hz, H-2),
1.74 (3H, s, H3-29), 1.55 (3H, s, H3-26), 1.52 (3H, s, H3-27),
1.05 (3H, s, H3-18), 1.01 (3H, d, J ) 6.5 Hz, H3-21), 0.98 (3H,
s, H3-30), 0.84 (1H, d, J ) 4.5 Hz, H-19), 0.56 (1H, d, J ) 4.5
Hz, H-19); HRFABMS m/z 529.3512 (calcd for C30H50O6Na [M
+ Na]+, 529.3505).
Qu a d r a n gu la r ic a cid M (10): 1H NMR (pyridine-d5) δ 5.57
(1H, dd, J ) 12.0, 4.5 Hz, H-3), 5.27 (1H, br s, H-26), 4.97
(1H, br s, H-26), 4.36 (1H, t, J ) 6.0 Hz, H-24), 3.92 (1H, br s,
H-1), 3.43 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.76 (1H, ddd, J )
13.0, 9.0, 8.0 Hz, H-11), 2.50 (1H, ddd, J ) 12.5, 4.5, 4.0 Hz,
H-2), 2.30 (1H, ddd, J ) 12.5, 12.0, 2.0 Hz, H-2), 1.92 (3H, s,
H3-27), 1.74 (3H, s, H3-29), 1.06 (3H, s, H3-18), 0.99 (3H, s,
H3-30), 0.97 (3H, d, J ) 6.5 Hz, H3-21), 0.84 (1H, d, J ) 4.5
Hz, H-19), 0.55 (1H, d, J ) 4.5 Hz, H-19).
1
24-Ep iqu a d r a n gu la r ic a cid M (11): H NMR (pyridine-
d5) δ 5.57 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 5.22 (1H, br s, H-26),
4.97 (1H, br s, H-26), 4.36 (1H, t, J ) 6.0 Hz, H-24), 3.92 (1H,
br s, H-1), 3.43 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 2.76 (1H, ddd,
J ) 13.0, 9.0, 8.0 Hz, H-11), 2.50 (1H, ddd, J ) 12.5, 4.5, 4.0
Hz, H-2), 2.30 (1H, ddd, J ) 12.5, 12.0, 2.0 Hz, H-2), 1.93 (3H,
s, H3-27), 1.74 (3H, s, H3-29), 1.06 (3H, s, H3-18), 0.99 (3H, s,
H3-30), 0.97 (3H, d, J ) 6.5 Hz, H3-21), 0.84 (1H, d, J ) 4.5
Hz, H-19), 0.55 (1H, d, J ) 4.5 Hz, H-19).
(S)-MTP A ester of d im eth yl qu a d r a n gu la r a te G (2d ):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.59-7.30 (15H,
m, Ph-H × 3), 5.63 (1H, dd, J ) 12.5, 4.5 Hz, H-3), 5.30 (1H,
t, J ) 5.5 Hz, H-24), 4.99 (1H, br s, H-26), 4.92 (1H, br s, H-26),
4.77 (1H, br s, H-1), 3.57 (3H, s, MeO-21), 3.56 (3H, s, MeO-
28), 3.50, 3.45, 3.38 (each 3H, s, OMe × 3), 2.49 (1H, dd, J )
12.0, 4.0 Hz, H-5), 2.35 (1H, ddd, J ) 13.0, 4.5, 4.0 Hz, H-2),
2.15 (1H, td, J ) 10.0, 3.0 Hz, H-22), 2.00 (1H, br t, J ) 10.0
Hz, H-22), 1.77 (1H, ddd, J ) 13.0, 12.5, 3.5 Hz, H-2), 1.60
(3H, s, H3-27), 1.57 (1H, m, H-23), 1.40 (1H, m, H-23), 1.10
(3H, s, H3-29), 0.88 (3H, s, H3-18), 0.76 (1H, d, J ) 4.5 Hz,
7â-Hyd r oxy-23-d eoxojessic a cid (12): colorless crystals;
mp 219 °C; [R]25 +80.9° (c 0.07, MeOH); IR νmax (KBr) 3400,
D
1700, 1470, 1380 cm-1; H NMR (pyridine-d5) δ 5.60 (1H, dd,
1

64 J ournal of Natural Products, 2000, Vol. 63, No. 1
Banskota et al.
H-19), 0.72 (3H, s, H3-30), 0.52 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 1217.4657 (calcd for C62H71F9O13Na [M + Na]+,
1217.4649).
Hz, H-19), 0.81 (3H, s, H3-30), 0.63 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 1173.4744 (calcd for C61H71F9O11Na [M + Na]+,
1173.4750).
(R)-MTP A ester of m eth yl qu a d r a n gu la r a te L (8b):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.62-7.31 (15H,
m, Ph-H × 3), 5.71 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 4.89 (1H,
dd, J ) 9.0, 2.5 Hz, H-24), 4.81 (1H, br s, H-1), 3.50 (3H, s,
MeO-28), 3.62, 3.50, 3.41 (each 3H, s, OMe × 3), 2.46 (1H, ddd,
J ) 12.5, 4.5, 4.0 Hz, H-2), 2.42 (1H, dd, J ) 12.0, 4.5 Hz,
H-5), 1.92 (1H, dd, J ) 12.5, 12.0, 2.0 Hz, H-2), 1.50 (2H, m,
H2-23), 1.19 (3H, s, H3-29), 1.10 (3H, s, H3-26), 1.10 (3H, s,
H3-27), 0.79 (3H, s, H3-18), 0.76 (1H, d, J ) 4.5 Hz, H-19), 0.68
(3H, d, J ) 6.5 Hz, H3-21), 0.55 (1H, d, J ) 4.5 Hz, H-19), 0.53
(3H, s, H3-30); HRFABMS m/z 1191.4825 (calcd for C61H73F9O12-
Na [M + Na]+, 1191.4856).
(S)-MTP A ester of m eth yl qu a d r a n gu la r a te L (8c):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.57-7.31 (15H,
m, Ph-H × 3), 5.65 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 4.91 (1H,
dd, J ) 9.0, 2.5 Hz, H-24), 4.80 (1H, br s, H-1), 3.56 (3H, s,
MeO-28), 3.53, 3.50, 3.39 (each 3H, s, OMe × 3), 2.42 (1H, dd,
J ) 12.0, 4.5 Hz, H-5), 2.36 (1H, ddd, J ) 12.5, 4.5, 4.0 Hz,
H-2), 1.95 (1H, td, J ) 11.0, 3.5 Hz, H-11), 1.80 (1H, ddd, J )
12.5, 12.0, 3.5 Hz, H-2), 1.60 (2H, m, H2-23), 1.19 (3H, s, H3-
29), 1.11 (3H, s, H3-26), 1.11 (3H, s, H3-27), 0.84 (3H, s, H3-
18), 0.77 (1H, d, J ) 4.5 Hz, H-19), 0.75 (3H, d, J ) 6.5 Hz,
H3-21), 0.74 (3H, s, H3-30), 0.55 (1H, d, J ) 4.5 Hz, H-19);
HRFABMS m/z 1191.4840 (calcd for C61H73F9O12Na [M + Na]+,
1191.4856).
Cyt ot oxic Assa y. Cellular viability in the presence and
absence of experimental agents were determined using the
standard 3-(4,5-dimethylthiazol-2-yl)-2,5-dimethyltetrazolium
bromide (Sigma, St. Louis, MO) assays, as described previ-
ously.29
Su p p or tin g In for m a tion Ava ila ble: 1H and 13C NMR data of
1a -4a , 6a -8a , 12a , 13a , 2b, and 12b. This material is available free
of charge via the Internet at http://pubs.acs.org.
Refer en ces a n d Notes
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Deh yd r a tion of MTP A Ester s 8b a n d 8c w ith P OCl3.
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(R)-MTP A ester of m eth yl qu a d r a n gu la r a te M (10b):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.68-7.36 (15H,
m, Ph-H × 3), 5.76 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 5.38 (1H,
dd, J ) 10.0, 2.0 Hz, H-24), 5.02 (1H, br s, H-26), 4.94 (1H, br
s, H-26), 4.86 (1H, br s, H-1), 3.55 (3H, s, MeO-28), 3.68, 3.53,
3.47 (each 3H, s, OMe × 3), 2.53 (1H, ddd, J ) 12.5, 4.5, 4.0
Hz, H-2), 2.49 (1H, dd, J ) 12.0, 4.5 Hz, H-5), 1.99 (1H, dd, J
) 12.5, 12.0, 2.0 Hz, H-2), 1.74 (1H, m, H-23), 1.71 (3H, s, H3-
27), 1.50 (1H, s, H-23), 1.17 (3H, s, H3-29), 0.84 (3H, s, H3-18),
0.79 (1H, d, J ) 4.5 Hz, H-19), 0.77 (3H, d, J ) 6.5 Hz, H3-21),
0.61 (1H, d, J ) 4.5 Hz, H-19), 0.56 (3H, s, H3-30); HRFABMS
m/z 1173.4735 (calcd for C61H71F9O11Na [M + Na]+, 1173.4750).
(S)-MTP A ester of m eth yl qu a d r a n gu la r a te M (10c):
colorless amorphous solid; 1H NMR (CDCl3) δ 7.60-7.38 (15H,
m, Ph-H × 3), 5.71 (1H, dd, J ) 12.0, 4.5 Hz, H-3), 5.34 (1H,
dd, J ) 10.0, 2.0 Hz, H-24), 4.94 (1H, br s, H-26), 4.90 (1H, br
s, H-26), 4.86 (1H, br s, H-1), 3.63 (3H, s, MeO-28), 3.60, 3.56,
3.45 (each 3H, s, OMe × 3), 2.56 (1H, dd, J ) 12.0, 4.5 Hz,
H-5), 2.43 (1H, ddd, J ) 12.5, 4.5, 4.0 Hz, H-2), 1.83 (1H, dd,
J ) 12.5, 12.0, 2.0 Hz, H-2), 1.82 (1H, m, H-23), 1.61 (3H, s,
H3-27), 1.56 (1H, m, H-23), 1.17 (3H, s, H3-29), 0.88 (3H, s,
H3-18), 0.86 (3H, d, J ) 6.5 Hz, H3-21), 0.83 (1H, d, J ) 4.5
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NP990336Q