Proteasome inhibitors from Neoboutonia melleri

Article abstract of DOI:10.1021/np200441h

Thirty new cycloartane derivatives (1-3, 5-12, 14-32) have been isolated from the leaves of Neoboutonia melleri. Their novelty stems from the loss of one of the C-4 methyl groups (1-3, 5-12, 14-25, and 32) and from the presence of an "extra" carbon atom i

Full text of DOI:10.1021/np200441h

Article
pubs.acs.org/jnp
Proteasome Inhibitors from Neoboutonia melleri
Christophe Long,† Josephine Beck,† Frederic Cantagrel,† Laurence Marcourt,† Laure Vendier,‡
́
́
́
Bruno David,† Fabien Plisson,† Fadila Derguini,† Isabelle Vandenberghe,§ Yannick Aussagues,†
Frederic Ausseil,† Catherine Lavaud,⊥ Francois Sautel,† and Georges Massiot*,†
̧
́
́
†USR CNRS-Pierre Fabre No. 3388 ETaC, Centre de Recherche et Dev
31035 Toulouse Cedex 01, France
‡UPR CNRS No. 8241, Laboratoire de Chimie de Coordination, 205 Route de Narbonne, 31077 Toulouse Cedex 04, France
§Institut de Recherche Pierre Fabre, Centre de Recherche en Oncologie Experimentale, Centre de Recherche et Dev
eloppement
Pierre Fabre, 3 Avenue Hubert Curien, 31035 Toulouse Cedex 01, France
⊥UMR CNRS-Universite
Reims Champagne-Ardenne No. 6229, Institut de Chimie Molec
́
eloppement Pierre Fabre, 3 Avenue Hubert Curien,
́
́
́
ulaire de Reims, Laboratoire de
Pharmacognosie, 51097 Reims, France
S
* Supporting Information
ABSTRACT: Thirty new cycloartane derivatives (1−3, 5−12,
14−32) have been isolated from the leaves of Neoboutonia
melleri. Their novelty stems from the loss of one of the C-4
methyl groups (1−3, 5−12, 14−25, and 32) and from the
presence of an “extra” carbon atom in the side chain (1−3, 5−12,
14−20, 26−29, and 30−32). Furthermore, compound 32
possesses a rare triterpene skeleton with the cyclopropane ring
fused onto C-1 and C-10, instead of C-9 and C-10. The struc-
tures were determined by spectrometric means, chemical cor-
relations, and X-ray crystallography of derivative 1c. The sub-
stitution pattern in ring A, with a cyclopropyl ring conjugated
with an α,β-unsaturated carbonyl moiety, confers to the molecule a particular reactivity, giving rise to a formal inversion of the
stereochemistry of the cyclopropane ring under UV irradiation. These compounds showed an interesting level of activity on the
proteasome pathway, thus motivating their evaluation as possible anticancer agents. The large number of isolated compounds
permitted a structure−activity relationship analysis, which showed that the presence of the two enone functions was a re-
quirement for the activity.
or the past decade, proteasome inhibitors have been con-
sidered as potentially useful drugs in the treatment of cancer,
Koutencha, “fish leaves” in the Bamum language. Neoboutonia is
an African tropical genus of the Euphorbiaceae, which, according
to the latest classifications, represents numerous varieties combined
into three species: N. melleri, N. diaguissensis Beille, and
N. macrocalyx Pax.5 N. melleri, a small tree (4−8 m) growing
in swamp areas from Cameroon/Angola to Sudan/
Mozambique, exhibits alternate, stipulate leaves, broadly ovate
or subcircular. It is locally considered as toxic and of interest
against diabetes. N. melleri was the subject of a single
phytochemical investigation,6 and articles on the genus are rare.7
Here we describe the isolation and structural characterization
of 30 new compounds, evaluation of their biological activity on
the proteasome pathway, and discussion of their structure−
activity relationship.
F
and this has culminated in the approval of bortezomib for the
treatment of multiple myeloma.1 Epoxomycin, lactacystin, and, the
most promising, salinosporamide A, presently in phase 1 clinical
trials, are examples of natural products that inhibit the proteasome
catalytic functions.2 In order to find new inhibitors that do not
particularly target proteolysis but the whole ubiquitin−proteasome
pathway, we have developed an assay based on a human colon
cancer cell line that stably expresses a 4-ubiquitin luciferase re-
porter protein (4Ub-Luc DLD-1).3 Under normal conditions, the
fusion protein (4Ub-Luc) is produced and, due to the ubiquitin
tag's presence, it is addressed to and degraded by the proteasome,
while inhibitors of the proteasome pathway lead to protein accumula-
tion and luminescence observation.4
The assay was applied to a collection of over 12 000 plant
extracts and 62 000 pure compounds including natural products
and derivatives. An extract from Neoboutonia melleri (Muell. Arg.)
Prain and two pure compounds isolated from it (1 and 2)
inhibited the proteasome pathway. The plant was collected in
Cameroon, in the late 1980s, following ethnopharmacological
observations. Its leaves were locally used to wrap fish, hence, its name
RESULTS AND DISCUSSION
The CH2Cl2 extract of N. melleri was dominated by two com-
pounds, which differed by the presence of an acetyl group and
■
Received: May 30, 2011
Published: December 14, 2011
© 2011 American Chemical Society and
American Society of Pharmacognosy
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Table 1. 13C NMR Data of Compounds 1-8
a
a
b
b
b
b
a
a
carbon
1
2
3
4
5
6
7
8
1
153.7, CH
128.3, CH
202.0, C
153.9, CH
128.2, CH
202.1, C
46.9, CH
42.6, CH
23.6, CH2
23.6, CH2
44.7, CH
26.1, C
155.5, CH
128.4, CH
202.4, C
47.6, CH
43.6, CH
24.3, CH2
24.3, CH2
45.2, CH
27.2, C
155.3, CH
128.5, CH
202.3, C
47.5, CH
43.1, CH
23.9, CH2
23.8, CH2
43.6, CH
27.4, C
33.5, CH
41.5, CH
213.2, C
50.5, CH
47.0, CH
26.5, CH2
26.2, CH2
48.6, CH
25.4, C
33.5, CH
41.5, CH
213.3, C
50.5, CH
47.0, CH
26.5, CH2
26.2, CH2
48.6, CH
25.4, C
154.4, CH
127.4, CH
202.5, C
43.8, CH
45.7, CH
65.6, CH
31.9, CH2
40.1, CH
25.3, C
154.6, CH
127.3, CH
202.7, C
43.8, CH
45.7, CH
65.6, CH
31.9, CH2
40.2, CH
25.4, C
2
3
4
46.9, CH
42.5, CH
23.4, CH2
23.5, CH2
44.3, CH
26.0, C
5
6
7
8
9
10
11
12
13
14
15
16
16a
16b
17
18
19
20
21
22
22a
22b
23
24
24a
25
26
26a
26b
27
28
30
31.9, C
31.9, C
32.9, C
33.3, C
30.3, C
30.3, C
29.6, C
29.7, C
27.4, CH2
32.1, CH2
47.4, C
27.5, CH2
32.1, CH2
47.4, C
28.1, CH2
32.9 CH2
46.9, C
28.4, CH2
32.8, CH2
44.8, C
27.4, CH2
33.3, CH2
46.8, C
27.4, CH2
33.5 CH2
46.7, C
27.1, CH2
32.3, CH2
46.3, C
27.2, CH2
32.3, CH2
46.1, C
46.0, C
45.8, C
48.4, C
52.4, C
48.0, C
47.9, C
46.7, C
46.7, C
46.0, CH2
76.0, CH
170.1, C
46.2, CH2
76.9, CH
170.1, C
21.8, CH3
50.1, CH
18.2, CH3
27.4, CH2
35.9, CH
11.7, CH3
74.8, CH
46.7, CH2
76.7, CH
171.3, C
41.7, CH2
82.5, CH
47.6, CH2
76.9, CH
171.3, C
22.1, CH3
51.4, CH
19.3, CH3
28.0, CH2
33.2, CH
13.2, CH3
78.5, CH
171.7, C
20.9, CH3
199.6, C
150.3, C
124.6, CH2
37.9, CH
66.4, CH2
47.8, CH2
77.5, CH
171.3, C
22.1, CH3
51.5,CH
19.5, CH3
28.0, CH2
36.3, CH
12.2, CH3
75.8, CH
46.8, CH2
76.2, CH
170.1, C
21.7, CH3
50.5, CH
19.0, CH3
32.6, CH2
32.4, CH
12.7, CH3
77.6, CH
170.6, C
20.7, CH3
198.6, C
149.2, C
123.8, CH2
37.1, CH
66.9, CH2
46.9, CH2
77.1, CH
170.2, C
21.8, CH3
50.4, CH
19.3, CH3
32.7, CH2
35.8, CH
11.5, CH3
74.8, CH
21.7, CH3
50.3, CH
17.8, CH3
27.0, CH2
32.4, CH
12.8, CH3
77.6, CH
170.6, C
22.1, CH3
51.3, CH
18.3, CH3
27.6, CH2
33.2, CH
13.3, CH3
78.3, CH
171.6, C
60.4, CH
21.3, CH3
26.5, CH2
38.2, CH
15.8, CH3
111.4, C
20.7, CH3
198.7, C
149.3, C
20.9, CH3
198.9, C
204.5, C
77.8, CH
153.3, C
205.6, C
204.4, C
148.0, C
149.2, C
149.1, C
147.9, C
123.7, CH2
37.2, CH
66.9, CH2
125.7, CH2
36.7, CH
67.0, CH2
125.6, CH2
34.9, CH
67.8, CH2
171.6, C
114.2, CH2
38.3, CH
69.0, CH2
126.5, CH2
37.6, CH
66.7, CH2
125.8, CH2
36.6, CH
67.0, CH2
21.1, CH3
17.3, CH3
11.3, CH3
20.0, CH3
16.3, CH3
10.8, CH3
19.6, CH3
16.5, CH3
10.8, CH3
19.6, CH3
17.8, CH3
11.3, CH3
19.3, CH3
17.2, CH3
11.2, CH3
20.6, CH3
17.2, CH3
11.2, CH3
20.7, CH3
16.3, CH3
10.8, CH3
20.3, CH3
16.5, CH3
10.8, CH3
20.3, CH3
a
b
In CDCl3. In CD3CN.
were named neoboutomellerone (1) and 22-de-O-acetylneobou-
tomellerone (2). Compound 1 showed a pseudomolecular ion
peak at m/z 591.3 [M + Na]+ in the positive ESI mode that
analyzed for C34H48O7Na. The UV spectrum exhibited two
maxima at 228 and 266 nm; the IR spectrum displayed strong OH
vibrations at 3430 cm−1 and carbonyl bands at 1735, 1665, and
1602 cm−1. Analysis of the 13C NMR spectrum (DEPTQ and
HSQC) permitted identification of seven methyl, eight methylene,
ten methine, and nine quaternary carbons, thus accounting for a
C34H47 partial formula and, therefore, a single OH group. Among
the deshielded signals, two were clearly assignable to ketocarbonyls
(δC 198.7 and 202.0), two ester carbonyls (δC 170.1 and 170.6),
and an oxymethylene carbon at δC 66.9. The esters were identified
as acetates on the basis of HMBC correlations between the CO
and methyls at δH 2.08 and 2.16. The composition determined by
HRMS was thus fully explained by NMR. A cyclohexenone moiety
with a quaternary carbon atom γ to the CO, a conjugated exo-
methylene, two acetates, two methyl groups on sp3 quaternary
carbons, and three methyl doublets were particular features of the
molecule.The observation of an isolated methylene with its most
upfield proton at δH 0.58, showing a small geminal coupling
constant (J = 4.3 Hz), suggested the presence of a cyclopropane
ring. Given the C30 composition of the basic skeleton (C34 minus
two acetyl moieties), the first structural hypothesis for 1 was that
of a diacetylated cycloartane derivative. On this basis, the two
methyl singlets were assigned to C-18 and C-30, and the HMBC
experiment permitted detection of the two quaternary carbons
C-13 and C-14 (shared correlations with the methyls, see Table 1
for chemical shifts) as well as C-8, C-17 (CH), C-12, and C-15
(CH2). The cyclohexenone moiety was located in ring A to
account for the observed long-range couplings between the
ketocarbonyl (C-3), H-1, H-4, H-5, and CH3-28; the most upfield
cyclopropane proton showed a correlation with H-1. Contrary to
most described cycloartanes,8 1 had a proton on C-4 and therefore
a single methyl group at this position. Analysis of the HMBC and
HSQC experiments permitted identification of all the carbon
atoms. The acetates were located on C-16 and C-22, and the
alcohol function was located on the methyl group that terminates
the chain. The location of the second enone moiety and the 30th
carbon atom was determined by the couplings observed between
the carbonyl at δC 198.7, the exo-methylene, and H-22, on one
hand, and between the exo-methylene protons and C-26 (or -27),
on the other. The extra carbon atom was thus placed at C-24, a
common alkylation site in the cycloartane series.9 The planar
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structure of neoboutomellerone (1) is thus 16,22-di-O-acetyl-26-
hydroxy-29-nor-24-methylcycloarta-1,24(24a)-diene-3,23-dione.
The second most abundant molecule was straightforwardly
identified as 22-de-O-acetylneoboutomellerone (2), H-22
appearing shielded at δH 4.74 instead of 5.57. Upon acetylation,
compounds 1 and 2 yielded the same 26-acetylneoboutomeller-
one (3), also isolated as a minor natural product, thus demon-
strating that the three compounds had the same configuration.
The latter was established through a ROESY experiment and by
the preparation of Mosher esters. The cyclopropyl hydrogen
atoms (H-19) were distinguished by the observation that the
exo hydrogen showed ROESY correlations with H-1 and H-11eq,
and the endo hydrogen with H-4, H-6, and H-8. These
observations showed that the A/B ring junction was trans
(H-5α) and that the C-4 methyl was equatorial (H-4β). In ring
D correlations between the angular methyl-18, H-8, and H-20
on one hand and between CH3-30, H-16, and H-17, on the
other, permitted assignment of a C/D trans ring junction with
OAc-16 and the side chain in a β-orientation. The configuration
of C-25 was determined by examination of the Mosher esters of
1 (1a and 1b), according to the method of Ivanchina.10 Ester
1a, prepared with S-α-methoxy-α-trifluoromethylphenylacetic
acid chloride, showed 0.12 ppm nonequivalence between the
diastereotopic H-26, while this value was 0.04 ppm for ester 1b
prepared with the R-acid chloride, thus implying a 25R con-
figuration. The configuration of C-22 was established similarly
on diol 2 by considering the chemical shifts of H-20; in this
case H-20 in the ester 2a, prepared from the S-acid chloride,
was found at higher field than in the ester 2b made with the
R-reagent (δH 2.57 vs 2.64). Finally, the configuration of C-20
was determined by the examination of the NMR spectra of
compound 4, a saponification product of 1, in which a cis- fused
five-membered ring is formed following hydrolysis of the
acetate at C-16, α-ketol rearrangement, and ring closure as a
hemiketal. In this compound, NOE correlations are observed
between H-20 and CH3-18, between H-17 and CH3-21, and
between CH3-30, H-16, and H-17, thus establishing a 20S
configuration. Compounds 1 and 2 could not be crystallized
for X-ray analysis. However, phenylcarbamate 1c was obtained
in crystalline form in the course of a chemical modification
program. The crystal structure determination confirmed these
assignments as depicted in the formulas (see Experimental Section
and Supporting Information for details).
were identified as 1,2-dihydroneoboutomellerone and 1,2-dihydro-
22-de-O-acetylneoboutomellerone.
HR-MS and NMR demonstrated that the pair of new com-
pounds 7 and 8 contained an extra oxygen atom compared to
1
their respective parent compounds 1 and 2. The H NMR
spectra showed a broad singlet (W1/2 = 10 Hz) at δH 4.20 in the
new compounds, corresponding to a CH at δC 65.6. In the
COSY experiment, this signal displayed couplings to upfield
CH and CH2, but the HMBC failed to yield any useful cor-
relations. Assuming that the basic skeleton of the neoboutomel-
lerones was conserved, C-6 was left as the unique position for
the hydroxylation. In agreement, the neighboring C-5 and C-7
were downfield shifted (4 and 9 ppm, respectively), while C-8
exhibited a γ shielding effect (3 ppm). This substitution also
induced a strong deshielding of C-19 (Δδ +6, δ effect) and,
most importantly, of the endo proton of the cyclopropane
(ΔδH +0.63) moiety, which could be explained by the presence
of a 6β-OH group in 7 and 8. A ROESY experiment led to the
same conclusion with the observation of interactions between H-6
and CH3-28, and between H-4 and H-19 endo, thus supporting
the configurations of C-4, C-5, and C-6. Consequently, the
structures of 7 and 8 were respectively determined to be 6β-
hydroxyneoboutomellerone and 6β-hydroxy-22-de-O-acetylneo-
boutomellerone. Along with compound 7, another isomer (9)
was isolated, in which either CH3-18 or CH3-30 was replaced by a
hydroxymethylene function. No HMBC correlations were
observed between this CH2 and the backbone atoms, but the
remaining methyl singlet displayed four long-range CH
correlations, including one with CH2-15 (identified by scalar
coupling with H-16). The OH was therefore placed at C-18, and
this accounted for the chemical shift of the remaining CH3 at δH
1.00 (CH3-18 at δH >1.15, CH3-30 at δH <1). Compound 9 was
thus assigned the structure of 18-hydroxyneoboutomellerone.
Compound 10 appeared to belong to the same series as 1,
displaying NMR signals for two O-acetyl groups, the γ,δ-
cyclopropyl-α,β-unsaturated carbonyl, and the full and
unmodified side chain. The HRMS indicated a C34H46O8
composition corresponding to the formula of compound 7
plus one degree of unsaturation. Analysis of the 1H NMR data
was facilitated by the absence of signal overlap, with the
sequence CH3-28, H-4, H-5, H-6, H-7, H-8 giving rise to well-
separated signals, in which H-6 and H-7 appeared as a doublet
These molecules show a high degree of novelty, and their
highly functionalized side chain is unique. Furthermore, there
are only a few examples of nor-29-cycloart-1-en-3-ones.11 With
these two major compounds and a valuable biological activity at
hand, it was decided to enlarge the panel of molecules in order
to evaluate the structural requirements for activity. A deriva-
tization program was thus undertaken,12 as well as a systematic
search for related natural products contained in this particular
plant and in a few plants well known as sources of cycloartanes
(Cimicifuga simplex, Musa sapientum, and Mangifera indica).
Two minor compounds, 5 and 6, respectively the 1,2-dihydro
derivatives of 1 and 2, were first isolated. Their HR-MS compo-
sitions were in agreement with the expected formulas (C34H50O7
1
and C32H48O6), and their H NMR spectra did not exhibit a
deshielded AX system for H-1 and H-2. Both molecules showed
a strong shielding of C-28 (δC 11.2), as a result of its equatorial
position, β to a carbonyl group. All other 13C NMR signals
except those of rings A and B were similar to those of 1−3 (see
Table 1). On the basis of these data, the structures of 5 and 6
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of doublets at δH 3.15 and 3.00. These two protons were
attached to carbon atoms resonating at δC 52.8 and 55.0, re-
spectively, suggesting that they were incorporated in an epoxide
ring. In this particular compound, H-5 was distinguished as an
isolated broad doublet with J = 11.3 Hz (δH 2.45), thus proving
its axial position and the trans-fusion of the A/B rings. The cyclo-
propane endo proton showed an Overhauser interaction with H-8
but not with H-6/7, allowing assignment of a β-configuration for
the epoxide moiety. Shielding of the cyclopropane exo-proton,
possibly as a result of interaction with the epoxide, is noteworthy.
Compound 10 is thus 6β,7β-oxidoneoboutomellerone.
The last compound in the “regular” series, 11, had a
C32H48O7 molecular formula, as established by HR-MS, indica-
tive of a hydroxylated analogue of 2. Absence of the AB system
for H-1 and H-2 in the enone, as well as significant differences
in the UV spectrum, suggested a 1,4-Michael addition of water
to the enone functionality. An HMBC correlation between the
exo-cyclopropane H-19 and the hydroxymethine allowed the
hydroxy group to be placed at C-1. H-1 appeared as a broad
triplet with J = 3 Hz (δH 3.80), characteristic of an equatorial
proton, and, consequently, the hydroxy group was deduced to
be axial. This was confirmed by the observation of an NOE
interaction between H-1 and the exo-cyclopropane proton.
Compound 11 was thus identified as 1,2-dihydro-1α-hydroxy-
22-de-O-acetylneoboutomellerone.
with C-7 being almost 7 ppm downfield (Table 2). In
acetonitrile-d3, the H NMR spectrum displayed several well-
1
separated signals, including H-17, H2-12, H-15, and endo H-19,
while H-5 was masked by the broad acetonitrile residual signal,
and exo H-19 partially overlapped with H3-30. A ROESY
experiment showed a correlation between H-5 and the endo
H-19, suggesting that these atoms were cofacial, thus offering
two possibilities: H-5β in a “normal” series or H-5α with H2-19
α-oriented. The presence of a cis-fused decalin moiety was
deduced by the observation of an NOE correlation between
H-1 and exo H-19, a feature not observed in any product in the
trans series. The decision between the two hypotheses came in
a serendipitous manner with the observation of a slow conver-
sion of 1 into 14 upon storage in acetonitrile or by irradiation
under a UV lamp, which could only be explained by opening of
the cyclopropane (cleavage of the C-9/C-10 bond activated by the
enone system) and ring closure to yield the thermodynamically
more favorable cis isomer.13 The configuration of all stereogenic
centers was assumed to be as in compound 1, and the proposed
structure for 14 was 9,10-di-epi-25ξ-neoboutomellerone. Com-
pound 15 is isomeric to 2 and was assigned the structure 9,10-
di-epi-25ξ-22-de-O-acetylneoboutomellerone owing to the strong
similarity between the 13C NMR spectra of 15 and 2.
At this point, all isolated compounds could be considered as
being derived from 1 and 2 and thus were assigned the same
configuration as evidenced by the series of related 13C NMR
chemical shifts presented in Tables 1 and 2. Among the minor
compounds, several isomers were isolated, including 12, in which
the 1H and 13C NMR spectra were almost superimposable with
the corresponding spectra of 1. The compounds were, however,
quite different, with 12 being slightly less polar on reversed-
phase HPLC. Since minor differences were observed for the
signals of CH-25 and CH2-26, 12 was proposed to be the C-25
epimer of 1, i.e., 25-epi-neoboutomellerone. This hypothesis
was supported by the Dess-Martin oxidation of 1 into aldehyde
13, which was found to be unstable, rapidly converting into
isomers at C-25. Compound 14, another isomer of 1, gave a
13C NMR spectrum remarkably different from that of 1, despite
the presence of all the functionalities, i.e., the two enones, the
cyclopropane, and the esters (see Table 2). The most significant
chemical shift differences were found for the signals of ring B,
The next set of new compounds (16−20) shared the same side
chain as before but without oxidation at C-26. As for 1 and 2,
the difference between 16 and 17 was an acetyl group at C-22.
The absence of a deshielded signal for H-26 and the presence
of two diastereotopic methyl doublets for H3-26 and H3-27
suggested that these compounds were 26-deoxyneoboutomel-
lerone (16) and 22-de-O-acetyl-26-deoxyneoboutomellerone (17),
respectively. Compound 18 belonged to the 22-OAc series, but
its molecular weight was 16 amu higher than observed for 16
(m/z 591.3287 for [M + Na]+). The 1H NMR spectrum of 18
showed similarity to that of 7, including a broad deshielded
singlet for H-6 and characteristic deshielding for endo H-19
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Table 2. 13C NMR Data of Compounds 9−16
a
b
b
a
b
a
a
b
carbon
9
10
11
12
13
14
15
16
1
153.5, CH
128.3, CH
202.0, C
154.2, CH
128.4, CH
202.1, C
74.0, CH
49.3, CH2
212.2, C
50.5, CH
39.5, CH
26.1, CH2
26.2, CH2
48.7, CH
26.0, C
153.7, CH
128.3, CH
202.0, C
155.5, CH
128.4, CH
202.4, C
155.4, CH
126.5, CH
200.6, C
155.7, CH
126.3, CH
200.7, C
47.8, CH
39.4, CH
20.6, CH2
31.2, CH2
40.7, CH
33.9, C
155.6, CH
128.4, CH
202.5, C
2
3
4
47.0, CH
43.0, CH
23.5, CH2
24.0, CH2
45.0, CH
25.8, C
45.6, CH
41.5, CH
52.8, CH
55.0, CH
38.9, CH
27.2, C
46.9, CH
42.5, CH
23.4, CH2
23.5, CH2
44.3, CH
26.0, C
47.6, CH
43.6, CH
24.3, CH2
24.3, CH2
45.2, CH
27.2, C
47.7, CH
39.4, CH
20.6, CH2
31.2, CH2
40.7, CH
33.7, C
47.6, CH
43.6, CH
24.3, CH2
24.3, CH2
45.2, CH
27.7, C
5
6
7
8
9
10
11
12
13
14
15
16
16a
16b
17
18
19
20
21
22
22a
22b
23
24
24a
25
26
27
28
30
32.0, C
31.8, C
34.0, C
31.9, C
33.4, C
26.6, C
26.6, C
32.9, C
27.5, CH2
27.7, CH2
47.5, C
27.4, CH2
32.1, CH2
47.1, C
26.5, CH2
33.3, CH2
46.6, C
27.4, CH2
32.1, CH2
47.4, C
28.1, CH2
33.0, CH2
46.9, C
29.8, CH2
32.1, CH2
47.8, C
29.9, CH2
32.1, CH2
47.8, C
28.1, CH2
32.9, CH2
46.8, C
51.0, C
47.3, C
47.9, C
46.0, C
48.3, C
46.1, C
46.0, C
48.4, C
46.2, CH2
76.6, CH
169.4, C
44.7, CH2
76.3, CH
171.3, C
22.1, CH3
49.7, CH
15.2, CH3
22.1, CH2
33.2, CH
13.7, CH3
78.6, CH
171.7, C
20.9, CH3
199.6, C
47.9, CH2
77.5, CH
171.3, C
22.1, CH3
51.5, CH
19.6, CH3
28.1, CH2
36.3, CH
12.2, CH3
75.8, CH
46.0, CH2
75.9, CH
170.1, C
46.7, CH2
76.8, CH
171.2, C
22.1, CH3
51.2, CH
18.3, CH3
27.6, CH2
33.8, CH
13.4, CH3
78.2, CH
171.6, C
43.8, CH2
76.0, CH
170.1, C
44.0, CH2
76.9, CH
170.1, C
21.8, CH3
49.3, CH
15.3, CH3
31.4, CH2
35.9, CH
12.1, CH3
75.0, CH
46.7, CH2
76.6, CH
171.3, C
22.1, CH3
51.3, CH
18.2, CH3
27.7, CH2
33.0, CH
13.2, CH3
78.5, CH
171.7, C
21.6, CH3
49.6, CH
64.9, CH2
28.5, CH2
32.8, CH
13.4, CH3
77.4, CH
170.5, C
21.7, CH3
50.3, CH
17.8, CH3
27.0, CH2
32.2, CH
12.7, CH3
77.8, CH
170.7, C
21.7, CH3
49.5, CH
15.1, CH3
31.3, CH2
32.5, CH
13.2, CH3
77.7, CH
170.6, C
20.7, CH3
198.5, C
20.7, CH3
198.6, C
20.9, CH3
198.3, C
20.6, CH3
198.8, C
149.3, C
20.9, CH3
199.5, C
205.6, C
204.6, C
149.3, C
150.3, C
149.1, C
149.2, C
145.8, C
148.0, C
154.3, C
123.6, CH2
37.4, CH
66.8, CH2
16.4, CH3
10.8, CH3
21.7, CH3
124.7, CH2
38.0, CH
66.4, CH2
17.2, CH3
11.3, CH3
19.6, CH3
126.5, CH2
37.6, CH
66.7, CH2
17.2, CH3
11.0, CH3
20.7, CH3
124.3, CH2
37.9, CH
66.8, CH2
15.5, CH3
10.8, CH3
19.6, CH3
128.6, CH2
48.9, CH
201.7, CH
13.8, CH3
11.3, CH3
20.0, CH3
123.7, CH2
37.2, CH
66.9, CH2
16.3, CH3
12.2, CH3
18.6, CH3
125.6, CH2
36.7, CH
67.0, CH2
16.4, CH3
12.2, CH3
18.6, CH3
122.3, CH2
29.7, CH
21.6, CH3
22.4, CH3
11.3, CH3
20.0, CH3
a
b
In CDCl3. In CD3CN.
(δH 1.81). This feature, as well as the overall concordance between
the spectra of 7 and 18, indicated that 18 was 6β-hydroxy-26-
deoxyneoboutomellerone. Compound 19, like the preceding
compounds, was characterized by four methyl doublets and
therefore lacked oxidation at the terminus of the side chain.
This compound had a molecular weight of 512, corresponding
to a C32H48O5 composition. This confirmed the absence of an
oxygen atom and suggested that 19 was a dihydro analogue of a
derivative of 2. Since the characteristic signals for the ring A
double bond were missing, it was deduced that 19 was 1,2-
dihydro-22-de-O-acetyl-26-deoxyneoboutomellerone. Com-
pound 20 was an isomer of 16 (C34H48O6), and, given the
previous analyses, it was readily identified as the cyclopropyl
isomer of 16, i.e., 9,10-di-epi-26-deoxyneoboutomellerone.
Compounds 21−23 differed from the previous compounds
by the absence of the C-24 exo-methylene group. Examination
of the NMR spectra showed that C-24a was replaced by a
proton (δH 6.15 0.2) and that the chain was terminated by a
3-methyl-2-butenoyl moiety. In these three compounds, the
tetracyclic cycloartane core was intact, and, as for compounds 1
and 2, 21 and 22 differed only by the presence of a C-22-O-acetyl
moiety. Compound 21 was thus 24a-nor-24,25-didehydro-26-
deoxyneoboutomellerone, and 22 was 22-de-O-acetyl-24a-nor-
24,25-didehydro-26-deoxyneoboutomellerone. While 21 and
22 were shown by 13C NMR to belong to the normal A/B trans
series, 23 displayed a markedly different spectrum, character-
istic of the cis α-series. Consequently, its structure was deduced
to be 9,10-di-epi-24a-nor-24,25-didehydro-26-deoxyneobouto-
mellerone (see Table 3).
Compounds 24 and 25 had molecular weights of 412 and
428, respectively, corresponding to molecular formulas C26H36O4
and C26H36O5. Although six carbon atoms were missing, the 1H
NMR spectra of both compounds showed strong resemblance
with those of the neoboutomellerones and displayed signals for
an acetate, an α,β-unsaturated carbonyl, a cyclopropane, and
the angular methyl groups. Initially, it was proposed that these
compounds were lacking C-23 to C-27 of the “regular” series
side chain. A formyl proton doublet at δH 9.58 (J = 1.8 Hz)
coupling with a doublet of doublets of quartets (δH 2.86)
suggested that truncation had happened between C-22 and C-23.
Since the 13C NMR spectrum showed that the tetracyclic core
was intact, 24 was proposed to be 23,24,24a,25,26,27-hexa-nor-
neoboutomelleron-22-al. This was confirmed by the high-yield
oxidation of 2 into 24 by MnO2. It is worth noting that 1 was
recovered unchanged under the same conditions. The 13C
NMR spectra of 24 and 25 could barely be distinguished except
for the formyl carbonyl carbon, which was replaced by a signal
at δC 182.0. It was deduced that 25 was the corresponding acid
38
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
Table 3. 13C NMR Data of Compounds 17−24
a
a
a
a
a
a
a
a
carbon
17
18
19
20
21
22
23
24
1
155.7, CH
128.3, CH
202.5, C
47.6, CH
43.6, CH
24.3, CH2
24.3, CH2
45.3, CH
27.2, C
156.2, CH
127.5, CH
203.4, C
33.5, CH2
41.5, CH2
213.3, C
50.5, CH
47.0, CH
26.5, CH2
26.2, CH2
48.6, CH
27.4, C
157.3, CH
126.5, CH
201.0, C
155.7, CH
128.4, CH
202.6, C
47.6, CH
43.5, CH
24.2, CH2
24.2, CH2
44.9, CH
27.0, C
155.7, CH
128.4, CH
202.5, C
47.6, CH
43.5, CH
24.3, CH2
24.2, CH2
45.0, CH
27.1, C
157.3, CH
126.5, CH
201.0, C
48.5, CH
40.4, CH
21.3, CH2
31.9, CH2
41.4, CH
27.6, C
155.4, CH
128.5, CH
202.3, C
47.6, CH
43.3, CH
24.1, CH2
24.0, CH2
44.3, CH
27.3, C
2
3
4
44.9, CH
46.6, CH
65.6, CH
32.7, CH2
41.1, CH
26.5, C
48.5, CH
40.4, CH
21.3, CH2
31.9, CH2
41.5, CH
27.7, C
5
6
7
8
9
10
11
12
13
14
15
16
16a
16b
17
18
19
20
21
22
22a
22b
23
24
24a
25
26
27
28
30
33.0, C
30.8, C
33.5, C
34.9, C
32.9, C
32.8, C
31.5, C
33.0, C
28.1, CH2
33.1, CH2
46.7, C
27.7, CH2
33.1, CH2
47.1, C
27.4, CH2
33.5, CH2
46.7, C
30.4, CH2
32.9, CH2
47.0, C
28.0, CH2
32.7, CH2
46.7, C
28.1, CH2
32.9, CH2
46.2, C
30.4, CH2
32.7
28.0, CH2
32.6, CH2
46.5, C
46.8, C
48.3, C
47.7, C
47.9, C
48.8, C
48.7, C
48.2, C
48.8, C
48.4, C
46.9, CH2
77.3, CH
171.3, C
22.1, CH3
51.4, CH
18.5, CH3
27.7, CH2
36.2, CH
12.3, CH3
75.8, CH
47.5, CH2
76.8, CH
171.3, C
47.8, CH2
77.5, CH
171.3, C
22.1, CH3
51.6, CH
19.5, CH3
28.0, CH2
36.2, CH
12.2, CH3
75.7, CH
44.6, CH2
76.5, CH
171.3, C
46.1, CH2
76.1, CH
171.3, C
21.8, CH3
51.1, CH
18.1, CH3
27.4, CH2
32.7, CH
13.2, CH3
81.5, CH
171.7, C
21.2, CH3
196.9, C
120.5, CH
46.3, CH2
76.5, CH
171.3, C
21.8, CH3
51.2, CH
18.3, CH3
27.4, CH2
34.4, CH
12.2, CH3
79.0, CH
44.1, CH2
76.1, CH
171.3, C
21.8, CH3
50.2, CH
15.4, CH3
31.6, CH2
32.8, CH
13.5, CH3
81.6, CH
171.6, C
21.2, CH3
196.9, C
120.5, CH
45.3, CH2
75.1, CH
170.9, C
21.3, CH3
50.7, CH
18.4, CH3
26.9, CH2
45.1, CH
13.4, CH3
205.2, CH
22.1, CH3
51.5, CH
19.4, CH3
33.0, CH2
33.0, CH
13.1, CH3
78.5, CH
171.7, C
22.1, CH3
50.6, CH
15.4, CH3
31.6, CH2
33.0, CH
13.6, CH3
78.6, CH
171.7, C
20.9, CH3
199.4, C
20.9, CH3
199.6, C
205.4, C
205.4, C
202.5, C
153.1, C
154.3, C
153.1, C
154.3, C
120.3, CH
124.3, CH2
29.5, CH
22.0, CH3
22.6, CH3
11.3, CH3
20.1, CH3
122.3, CH2
29.7, CH
22.4, CH3
21.7, CH3
11.0, CH3
20.7, CH3
124.3, CH2
29.5, CH
22.0, CH3
22.6, CH3
11.3, CH3
20.8, CH3
122.3, CH2
29.7, CH
21.6, CH3
22.4, CH3
12.6, CH3
18.9, CH3
159.8, C
159.9, C
159.7, C
28.0, CH3
21.0, CH3
11.3, CH3
19.8, CH3
28.1, CH3
21.3, CH3
11.3, CH3
19.9, CH3
28.0, CH3
21.0, CH3
12.6, CH3
18.8, CH3
11.3, CH3
19.4, CH3
a
In CD3CN.
of 24, i.e., 23,24,24a,25,26,27-hexa-nor-neoboutomelleron-22-
oic acid.
Six more compounds (26−31) were isolated and, although
devoid of any biological activity (vide infra), are interesting
from a biosynthetic standpoint. They all shared common func-
tionalities in rings A and B: a hydroxy group at C-3 and a
quaternary C-4 bearing a hydroxycarbonyl or ester function.
These compounds could be considered as the precursors for
the aforementioned compounds, undergoing oxidation of a C-4
methyl group of a cycloartane, oxidation of C-3, followed by
decarboxylation and introduction of the Δ1,2 double bond,
paralleling the well-known biosynthesis of the steroids. All six
compounds had a 3S configuration (3β-OH) based on the large
vicinal coupling between H-3 and H-2α (ca. 11 Hz) and a 4S
configuration to account for the strong shielding of the axial
C-28 methyl group (δC ca. 10). In the 1H NMR spectrum of 27,
H-5 was observed as a doublet of doublets with J = 12.2 and
3.4 Hz and was therefore α-axially oriented. Configurations of the
other ring junctions, and of C-16, C-20, and C-22, were most
likely the same as for the major compounds given the simi-
larities in chemical shift values. Compounds 26 and 27 were
isomers with a C33H50O7 molecular formula, and comparison of
their NMR spectra showed that they differed in the location of
the side chain hydroxy group: C-22 for 27 and C-25 for 26.
However, it was not possible to define the configuration of
C-25 in compound 26 due to a paucity of material. Compound
29 was not oxidized at C-16 nor C-22, as shown by the pre-
sence of two methylene resonances at δC 29.3 and 46.5. The C-22
methylene group gave rise to two well-separated doublets of
doublets, with 15.6 Hz geminal couplings and vicinal couplings
39
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
of 3.1 and 10.1 Hz, the smaller constant being close to that
observed in all the HO-22 derivatives. The mass spectrum of
compound 28 indicated a molecular formula of C31H46O5,
corresponding to the loss of HOAc with regard to 26 and 27.
Accordingly, no signal was observed for an acetate group in
the 1H NMR spectrum. A noteworthy feature of the 13C NMR
spectrum was the presence of a quaternary carbon at δC 108,
which showed HMBC couplings with two exo-methylene protons,
sp2 C-24a, and an oxymethylene corresponding to C-26. These
characteristics were best accounted for by a ketal derived from a
precursor possessing a C-23 carbonyl group and hydroxy
groups at C-16 and C-26. No NOE correlations between
protons of the spiroketal rings were evident, and we could not
assign configurations to C-23 and C-25. Compounds 30 and 31
shared with 29 the absence of oxidation of C-16 and C-22, but
analysis of the NMR spectra suggested that these highly polar
compounds were glycosides. In both compounds, the sugar
was tentatively identified as an α-L-rhamnose. The difference
between the two compounds was in the side chain, with 31 having
the same side chain as 16 and 17, and 30 having the truncated
chain of 21 and 22. Therefore, the respective structures of these
compounds were 16-acetyl-3β,26-dihydroxy-24-methyl-25ξ-cy-
cloart-24(24a)-en-23-on-29-oic acid (26), 16-acetyl-3β,22β-
dihydroxy-24-methylcycloart-24(24a)-en-23-on-29-oic acid (27),
3β,16β,22β-trihydroxy-24-methyl-(16,23:23,26)-diepoxycy-
cloart-24(24a)-en-29-oic acid (28), 3β-hydroxy-24-methylcy-
cloart-24(24a)-en-23-on-29-oic acid (29), 3β-hydroxy-29-O-
(α-L-rhamnopyranosyl)cycloart-24-en-23-on-29-oic acid (30),
and 3β-hydroxy-29-O-(α-L-rhamnopyranosyl)-24-methylcy-
cloart-24(24a)-en-23-on-29-oic acid (31).
bond was evident, with H-11 being a broad doublet coupling
with H2-12. This assignment was supported by an HMBC
correlation to C-18. A broad hydroxymethine singlet [δH 4.16
(δC 73.2)] showed HMBC couplings with the methine, the
methylene, and the quaternary carbon of the cyclopropane
moiety. The COSY experiment showed a weak but definite
coupling between this proton and the cyclopropane proton at
δH 1.88. Consequently, the hydroxy group was placed at C-2,
and the cyclopropane bridge between C-1 and C-10. A NOESY
experiment showed a correlation between H-2 and the exo-
cyclopropane proton, suggesting that this proton and the
cyclopropane moiety were β-cofocially oriented, and therefore
the 2-OH group must be α-oriented. This molecule is another
member of a rare family of triterpenes that are linked to the
parent compounds through acidic rearrangements, for example,
simplexol and O-methylcinicimol, arising from cimigenol.15
In our case, the putative epoxide 33, an oxidation product of
1, should be the intermediate between 32 and the regular
cycloartanes, and the rearrangement should be favored by an
anti relationship between the cyclopropane and the oxirane,
hence giving further support to the 2α-OH configuration.
On the basis of the above evidence, the structure of 32 was
proposed to be 16,22-diacetyl-2,26-dihydroxy-29-nor-24-
methyl-19(9→1)-abeo-cycloart-9(11),24(24a)-diene-3,23-
dione.
Structure−Activity Relationship Analysis. Twenty-four
of the isolated compounds the two non-natural derivatives, 4
and 13, were evaluated in the cellular proteasome assay (Table 5).
Cimiracemosides F and G,16 cycloeucalone,17 28-nor-cycloeuc-
alone, mangiferonic, and mangiferolic acids,18 isolated from
Cimicifuga simplex, Musa sapientum, and Mangifera indica, were
also tested, but none were found to be active. Table 5 lists the
induction factors (IFs) observed for the cycloartanes. The IF is
defined as the increase in luciferase signal measured in the
4UB-Luc-DLD-1 cells, after 7 h exposure to the test compound,
as compared to untreated cells.3b Epoxomycin was used as re-
ference with an IF of 100 at 0.5 μM. Due to the cytotoxicity of
the compounds and/or the accumulation of proteasome sub-
strates (known to produce cytotoxic effects), it was not possible
to observe luminescence at the highest concentrations. It also
explains why a maximum on the induction factor (IF) versus
concentration plots is observed (bell-shaped curve). The two
major compounds (1, 2) from the plant were also the most
active, with IF values of 48 and 45 at 1 μM, respectively, and
the activity decreased with the acylation level at the C-26
hydroxy group, with 1, 2 > 3. The integrity of the side chain
seemed to be another requirement since compounds such as 4
or 24 showed no activity. Simple modifications, such as the
removal of the exo-methylene in this side chain (21 and 22) or
the alteration of CH3-18 into a hydroxymethlene (9), also
reduced the biological activity. However, a C-6 hydroxy group
(7, 8), an epoxy moiety on C-6/C-7 (10), a configurational
change at C-25 (12), a cyclopropyl isomerism (14, 15), or
substitutions at C-26 (13, 16, 17) represent structural modi-
fications that do not significantly affect biological activity.
Compound 32 was an isomer of 7, with its 1H NMR spectrum
displaying all the signals corresponding to rings C and D and of
the side chain. However, the enoyl AB system in 7 was missing
and the usual upfield doublet for H-19 was replaced by a
doublet of doublets at δH 0.41 (J = 9.9, 4.7 Hz) and 0.94 [broad
triplet (J = 4.7 Hz)], with both signals coupled to a broadened
doublet of doublets at δH 1.88 (J = 9.9, 4.7 Hz). The values of
the vicinal coupling constants in the cyclopropane ring indicated
that the most upfield proton was endo, since it is known that in
such systems Jtrans > Jcis.14 A trisubstituted C-9/C-11 double
40
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Journal of Natural Products
Article
Table 4. 13C NMR Data of Compounds 25−32
a
a
c
c
c
c
c
a
carbon
25
26
27
28
29
30
31
32
1
153.7, CH
128.3, CH
202.0, C
31.4, CH2
29.4, CH2
75.4, CH
54.5, C
32.8, CH2
30.5, CH2
76.3 CH
56.0, C
32.8, CH2
30.5, CH3
76.3, CH
55.9, C
32.8, CH2
30.5, CH2
76.3, CH
55.8, C
32.7, CH2
30.4, CH2
75.9, CH
56.5, C
32.7, CH2
30.4, CH2
75.9, CH
56.5, C
23.6, CH
73.2, CH
213.2, C
2
3
4
46.8, CH
42.3, CH
23.4, CH2
23.3, CH2
43.4, CH
40.4, CH
47.6, CH
26.6, CH2
29.5, CH2
44.2, CH
140.4, C
5
44.0, CH
22.8, CH2
25.6, CH2
47.7, CH
19.7, C
45.7, CH
24.1, CH2
26.9, CH2
49.7, CH
20.8, C
45.5, CH
23.9, CH2
27.1, CH2
49.1, CH
20.8, C
45.6, CH
24.1, CH2
26.7, CH2
49.3, CH
21.1, C
46.0, CH
23.9, CH2
26.5, CH2
49.3, CH
21.2, C
46.0, CH
23.9, CH2
26.5, CH2
49.3, CH
21.2, C
6
7
8
c
9
n.d.
10
11
12
13
14
15
16
16a
16b
17
18
19
20
21
22
22a
22b
23
24
24a
25
26
27
28
29
30
1′
32.0, C
27.4, CH2
32.0, CH2
45.2, C
25.1, C
26.6, C
26.8, C
26.4, C
26.3, C
26.3, C
30.0, C
26.0, CH2
32.3, CH2
45.9, C
27.1, CH2
33.8, CH2
46.9, C
27.0, CH2
34.1, CH2
45.7, C
27.4, CH2
34.0, CH2
46.6, C
27.4, CH2
33.9, CH2
46.6, C
27.4, CH2
33.9, CH2
46.6, C
116.1, CH
36.2, CH2
45.1, C
47.5, C
47.1, C
48.3, C
47.4, C
50.1, C
50.1, C
50.1, C
45.2, C
44.0, CH2
75.0, CH
171.8, C
21.1, CH3
52.0, CH
18.0, CH3
26.5, CH2
38.7, CH
17.5, CH3
182.0, C
45.7, CH2
75.1, CH
171.0, C
21.2, CH3
54.7, CH
18.5, CH3
30.2, CH2
27.3, CH
19.2, CH3
44.8, CH2
48.1, CH2
78.1, CH
172.3, C
21.9, CH3
51.8, CH
19.5, CH3
31.3, CH2
36.2, CH
12.1, CH3
76.0, CH
44.7, CH2
74.0, CH
36.5, CH2
29.3, CH2
36.5, CH2
29.3, CH2
36.5, CH2
29.3, CH2
45.0, CH2
76.2, CH
170.1, C
21.7, CH3
49.1, CH
15.1, CH3
15.8, CH2
32.4, CH
12.8, CH3
77.6, CH
170.6, C
20.7, CH3
198.7, C
57.7, CH
20.9, CH3
31.3, CH2
26.8, CH
21.0, CH3
43.8, CH2
53.8, CH
18.5, CH3
30.7, CH2
35.2, CH
19.8, CH3
46.5, CH2
53.8, CH
18.6, CH3
30.8, CH2
34.9, CH
19.7, CH3
52.7, CH2
53.8, CH
18.6, CH3
30.8, CH2
35.1, CH
19.7, CH3
46.4, CH2
203.1, C
205.1, C
108.0, C
205.0, C
204.2, C
204.9, C
151.8, C
154.2, C
159.2, C
157.1, C
125.3, CH
157.1, C
149.3, C
123.8, CH2
36.6, CH
67.6, CH2
16.1, CH3
9.1, CH3
180.8, C
20.0, CH3
123.2, CH2
29.9, CH
22.6, CH3
21.9, CH3
10.2, CH3
181.7, C
106.2, CH2
38.7, CH
72.7, CH2
14.3, CH3
10.1, CH3
181.4, C
122.6, CH2
29.1, CH
22.3, CH3
22.4, CH3
10.0, CH3
181.2, C
19.7, CH3
122.6, CH2
29.1, CH
22.3, CH3
22.4, CH3
9.9, CH3
176.2, C
123.7, CH2
37.2, CH
66.9, CH2
16.3, CH3
10.8, CH3
157.0, C
27.7, CH3
20.9, CH3
9.9, CH3
176.2, C
10.8, CH3
19.0, CH3
20.7, CH3
20.0, CH3
19.7, CH3
94.9, CH
71.2, CH
72.4, CH
73.3, CH
72.4, CH
17.9, CH3
19.8, CH3
94.9, CH
71.2, CH
72.4, CH
73.3, CH
72.4, CH
17.9, CH3
19.4, CH3
2′
3′
4′
5′
6′
a
c
c
In CDCl3 In methanol-d4. n.d., not detected.
Moreover, minor changes in ring A were tolerated; the 1,2-
dihydro derivatives (5, 6) and 1-hydroxy derivative (11) showed
some activity, possibly due to reoxidation in vivo. Compounds
lacking the A-ring enone function (26−28, 30−32) or the
C-16-ester moiety (4, 28) were also found to be weakly
active. Acetylation of the C-22-hydroxy group results in
minor decreases of activity among comparable structures
(1/2, 5/6, 7/8, 16/17, and 21/22). Finally, enone function-
alities associated with the side chain and in ring A appear to
be crucial to maintain the biological activity, thus leaving the
C-16-ester group and the C-22-hydroxy function as possible
points of modification.
species such as N. melleri. Cursory investigation of the
terpenes from the twigs did not show the presence of
cycloartanes but rather daphnanes, as observed in the only
published investigation on similar material.6 Despite the
ubiquity of these triterpenes, the oxidation level of the
molecules discussed here is most peculiar, with the rare
combination of an α,β-unsaturated carbonyl with a cyclo-
propane giving rise to photochemical reactivity. The two
Michael acceptors in ring A and in the side chain add further
biological properties to the neoboutomellerones. In fact, the
two moieties may act as reversible or irreversible inhibitors
across the ubiquitin−proteasome pathway. From a medicinal
point of view, the novelty of the isolated compounds and
their interesting pharmacological properties motivate the
antitumor activity evaluation of the most abundant
molecules 1 and 2.19
CONCLUSION
■
From a phytochemical standpoint, it is not surprising to find
cycloartanes as major secondary metabolites in a Euphorbiaceae
41
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
reference RBL-273 in the Pierre Fabre Botanical Conservatory in
Cambounet-sur-le-Sor (France). The plant was collected again by one
of us (B.D.) at the same location and identified by comparison of
the vouchers at the National Herbarium of the Museum National
d’Histoire Naturelle in Paris. HPLC of the extracts showed similar
profiles.
Table 5. Induction Factors for Cycloartanes
induction factor (4Ub Luc DLD-1 cell) at dose (μM)
compound
10
5
1
0.5
1
4.4
2.9
37
28
48
48
45
12
2.1
1.4
1.1
2
Extraction and Isolation. The dried leaves (1 kg) of N. melleri
were powdered and extracted at room temperature with CH2Cl2 (15 L)
overnight. After filtration, the extract was concentrated under reduced
pressure. The residue (45 g) was dissolved in CH2Cl2 (1 L) and stirred
for 1 h with 100 g of activated vegetable charcoal. After filtration and
concentration under reduced pressure, the extract (32 g) was parti-
tioned between MeOH and cyclohexane. The MeOH fraction (15 g)
was subjected to silica gel CC (600 g, 57 × 420 mm) using
cyclohexane−EtOAc (50:50) to give 50 fractions of 150 mL. All the
fractions were analyzed by TLC on silica gel using the solvent mixture
CH2Cl2−MeOH (97:3) and pooled according to TLC into four
fractions (F1−F4). Fraction F1 (6.0 g) was purified by semipreparative
HPLC RP-18 chromatography, eluting with a linear gradient (80 to
100% B), to give after repeated HPLC purification 17 (84 mg), 19 (53 mg),
23 (6 mg), 3 (5 mg), 22 (25 mg), 21 (18 mg), 16 (76 mg), 24 (7 mg),
20 (3 mg), and 18 (1 mg). Fractions F2 (5.5 g) and F3 (1.5 g) were
purified by semipreparative RP-18 chromatography, eluting with a
linear gradient (45−100% B). After repeated HPLC purification of F2,
compounds 1 (2.5 g, 0.25%), 6 (180 mg, 0.018%), and 5 (107 mg,
0.0107%) were obtained. Fraction F3, after repeated HPLC
purification, afforded the second most abundant compound, 2 (600 mg),
10 (11 mg), and 15 (63 mg). Fraction F4 (1.0 g) was purified by
semipreparative RP-18 chromatography, to give after repeated HPLC
purification with a linear gradient (90−100% B and 50−100% B) the
most polar compounds, 26 (3 mg), 30 (5 mg), 27 (7 mg), 29 (23 mg),
28 (17 mg), 11 (12 mg), 25 (3 mg), 31 (2 mg), 7 (34 mg), 8 (18 mg),
14 (80 mg), 12 (5 mg), 32 (1 mg), and 9 (1 mg).
3
4
2.6
1.0
0.9
5
48
77
45
10
31
35
12
72
11
71
70
84
0.9
1.2
5.1
1.1
1.2
2.8
1.6
1.5
1
6
7
1.2
8
9
1.1
10
11
12
13
14
15
16
17
21
22
24
26
27
28
30
31
32
15
74
60
31
24
1.3
1.0
2
1.4
0.9
9.1
0.9
7.2
2.4
1.1
1.1
1.1
1.0
1.2
1.2
1.0
1.1
1.2
19
20
44
53
74
65
34
5.2
3.0
1.3
1.3
1.3
1.3
1.0
1.0
1.9
2.8
1.8
1.3
1.1
1.1
15
Neoboutomellerone (1): white fluffy solid, [α]20 −55 (c 0.16,
D
CHCl3); UV (MeOH) λmax (log ε) 224 (3.77), 266 (3.83) nm; IR
(film) νmax 3430, 1735, 1665, 1602, 1230 cm−1; 1H NMR (CDCl3, 500
MHz) δ 6.85 (1H, d, J = 10.1 Hz, H-1), 6.13 (1H, s, H-24a), 6.02 (1H,
d, J = 10.1 Hz, H-2), 5.89 (1H, br s, H-24a), 5.57 (1H, d, J = 2.1 Hz,
H-22), 5.09 (1H, dt, J = 4.6, 7.6 Hz, H-16), 3.59 (2H, d, J = 5.8 Hz, 2
H-26), 2.88 (1H, sext, J = 6.5 Hz, H-25), 2.60 (1H, dqd, J = 11.0, 7.0,
2.1 Hz, H-20), 2.31 (1H, dd, J = 14.0, 7.9 Hz, H-15a), 2.28 (1H, dd,
J = 11.1, 7.5 Hz, H-17), 2.19 (1H, dq, J = 12.7, 7.9 Hz, H-4), 2.16 (3H, s,
OAc-22), 2.08 (3H, s, OAc-16), 1.75−1.65 (3H, m, H-6, 2 H-12), 1.54
(1H, m, H-11), 1.47 (1H, m, H-7), 1.32 (1H, dd, J = 13.7, 4.6 Hz, H-
15b), 1.20 (1H, d, J = 4.3 Hz, H-19a), 1.18 (3H, s, H-18), 1.12 (3H, d,
J = 7.0 Hz, H-27), 1.10 (3H, d, J = 6.7 Hz, H-28), 0.95 (3H, s, H-30),
0.90 (3H, d, J = 6.7 Hz, H-21), 0.58 (1H, d, J = 4.3 Hz, H-19b); 13C
NMR, see Table 1; HR-ESIMS m/z 591.3297 (calcd for C34H48O7Na
591.3292).
EXPERIMENTAL SECTION
■
General Experimental Procedures. Optical rotations were
determined on a Perkin-Elmer 341 automatic polarimeter. UV spectra
were obtained in MeOH using a UV MC2 Safas spectrophotometer.
An FT-IR Bruker Tensor 27 spectrophotometer was used for scanning
IR spectroscopy. The NMR spectra were recorded on a Bruker Avance
II spectrometer equipped with a 13C cryoprobe at 500 MHz for H
1
and 125 MHz for 13C; 2D experiments were performed using standard
Bruker programs. The ESIMS and MS/MS were performed using a
Bruker Esquire-LC ion trap mass spectrometer; the samples were
introduced by infusion in a solution of MeOH. HR-ESIMS were
obtained on a Bruker MicrOTOF. TLC was carried out on precoated
silica gel 60F254 (Merck) with CH2Cl2−MeOH (97:3), and spots were
visualized by heating after spraying with 3% H2SO4 + 1% vanillin. CC
was carried out on a prepacked Kieselgel cartridge (40−60 μm) with
cyclohexane−EtOAc (50:50). Analytical HPLC was performed on a
Merck-Hitachi apparatus equipped with an L-7200 automated sample
injector, an L-7100 pump, a L-7450 diode array detector, a D-7000
interface, and Lachrom HSM or EZChrom software. A prepacked C18
reversed-phase column (Lichrospher 100 RP-18, 4 × 125 mm, 5 μm)
was used for analytical HPLC with a binary gradient elution (solvent
A: H2O; solvent B: MeCN) and a flow rate of 1 mL·min−1. Semipre-
parative HPLC was performed on an apparatus equipped with a VWR
International LaPrep pump P110, a VWR LaPrep P314 Dual λ
absorbance detector, and EZChrom software. A 50 × 250 mm column
(NW 50, Merck) with LiChroprep RP-18 (15−25 μm) and a pre-
packed C18 reversed-phase column (Hibar-Lichrospher 100 RP-18,
25 × 250 mm, 5 μm) were used for semipreparative HPLC with a
binary gradient elution (solvent A: H2O; solvent B: MeCN) and a flow
rate of 100 or 30 mL.min−1, and the chromatogram was monitored at
210 and 270 nm.
Preparation of Compound 1c. To a stirred solution of 1 (53 mg,
0.093 mmol) in 1 mL of CH2Cl2 were added DMAP (6 mg, 0.047
mmol, 0.5 equiv), phenyl isocyanate (11 μL, 0.102 mmol, 1.1 equiv),
and Et3N (11 μL, 0.102 mmol, 1.1 equiv). The mixture was stirred at
room temperature, and after 22 h, it was diluted with EtOAc and the
organic solution was successively washed with 4% aqueous HCl,
NaHCO3, and brine. The organic solution was dried over MgSO4 and
concentrated under vacuum. The residue was purified by chromatog-
raphy on silica gel (elution: cyclohexane−EtOAc, 1:0 to 0:1) to give 1c
1
as a white solid (30 mg, 47%): H NMR (500 MHz, CD3CN) δ 7.68
(1H, br s, H-26b), 7.41 (2H, br d, J = 8.3 Hz, H-26d, H-26 h), 7.29
(2H, dd, J = 7.6, 8.3 Hz, H-26 g, H-26e), 7.03 (1H, tt, J = 7.6, 1.2 Hz,
H-26f), 6.94 (1H, d, J = 10.1 Hz, H-1), 6.15 (1H, s, H-24a), 6.02 (1H,
d, J = 0.6 Hz, H-24a), 5.90 (1H, d, J = 10.1 Hz, H-2), 5.54 (1H, d,
J = 2.1 Hz, H-22), 5.09 (1H, dt, J = 4.4, 7.7 Hz, H-16), 4.13 (1H, dd,
J = 10.7, 7.0 Hz, H-26a), 4.06 (1H, dd, J = 10.7, 6.4 Hz, H-26b), 3.07
(1H, sext, J = 7.0 Hz, H-25), 2.61 (1H, ddq, J = 2.3, 10.9, 7.0 Hz,
H-20), 2.30 (1H, dd, J = 11.0, 7.3 Hz, H-17), 2.10 (3H, s, OAc-22),
2.03 (3H, s, OAc-16), 1.55 (1H, ddd, J = 6.6, 9.0, 14.8 Hz, H-11b),
1.37 (1H, dd, J = 13.9, 4.1 Hz, H-15b), 1.24 (1H, d, J = 4.6 Hz,
H-19a), 1.18 (3H, s, H-18), 1.11 (3H, d, J = 7.0 Hz, H-27), 1.02
Plant Material. The original plant sample was collected by Rene
́
Belle
́
(Pierre Fabre Research Institute) in Malantouen (Cameroon) on
June 21, 1988, and an herbarium specimen was deposited under
42
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
(3H, d, J = 6.7 Hz, H-28), 0.95 (3H, s, H-30), 0.85 (3H, d, J = 7.0 Hz,
H-21), 0.57 (1H, d, J = 4.6 Hz, H-19b); 13C NMR (125 MHz,
CD3CN) δ 202.4 (C-3), 198.8 (C-23), 171.8 (C-22a), 171.3 (C-16),
155.5 (C-1), 149.2 (C-24), 139.9 (C-26c), 129.9 (C-26e, 26 g), 128.4
(C-2), 125.7 (C-24a), 123.9 (C-26f), 119.5 (C-26d, 26 h), 78.5 (C-22),
76.7 (C-16), 68.5 (C-26), 51.3 (C-17), 48.3 (C-14), 47.6 (C-4), 46.8
(C-13), 46.7 (C-15), 45.2 (C-8), 43.6 (C-5), 35.0 (C-25), 33.4 (C-20),
32.9 (C-12), 28.1 (C-11), 27.6 (C-19), 27.2 (C-9), 24.3 (C-6, 7), 22.1
(C-16b), 21.0 (C-22b), 20.0 (C-30), 18.3 (C-18), 17.1 (C-27), 13.4
(C-21), 11.3 (C-28); ESIMS m/z: 688.24 [M + H]+.
2-(S)-MTPA ester (2b): 1H NMR (methanol-d4, 500 MHz) δ 4.40
(2H, m, H-26), 2.65 (1H, m, H-20); ESIMS m/z 981 [M + Na]+.
26-Acetylneoboutomellerone (3): yellow gum; [α]20D −61 (c 0.05,
CHCl3); UV (MeOH) λmax (log ε) 222 (3.78), 266 (3.74) nm; IR
(film) νmax 3299, 2933, 2874, 1736, 1699, 1596, 1372, 1230, 1022 cm−1;
1H NMR (CD3CN, 500 MHz) δ 6.94 (1H, d, J = 10.1 Hz, H-1),
6.10 (1H, s, H-24a), 5.95 (1H, d, J = 0.6 Hz, H-24a), 5.88 (1H, d,
J = 10.1 Hz, H-2), 5.52 (1H, d, J = 2.1 Hz, H-22), 5.09 (1H, dt, J = 4.6,
7.6 Hz, H-16), 4.04 (1H, dd, J = 7, 11 Hz, H-26a), 3.99 (1H, dd,
J = 6.5, 11 Hz, H-26b), 3.00 (1H, sext, J = 6.9 Hz, H-25), 2.57 (1H,
ddq, J = 11.0, 2.1, 7.0 Hz, H-20), 2.28 (1H, dd, J = 11.0, 7.6 Hz, H-17),
2.08 (3H, s, OAc-22), 2.05 (3H, s, OAc-16), 1.96 (3H, s, OAc-26),
1.56 (1H, ddd, J = 6.3, 8.5, 15 Hz, H-11b), 1.45 (1H, m, H-7a), 1.37
(1H, dd J = 14.5, 4.5 Hz, H-15b), 1.24 (1H, d, J = 4.6 Hz, H-19a), 1.19
(1H, m, H-7b), 1.18 (3H, s, H-18), 1.07 (3H, d, J = 7.0 Hz, H-27),
1.02 (3H, d, J = 6.7 Hz, H-28), 0.95 (3H, s, H-30), 0.94 (1H, m,
H-6b), 0.84 (3H, d, J = 7.0 Hz, H-21), 0.57 (1H, d, J = 4.6 Hz,
H-19b); 13C NMR, see Table 1; HR-ESIMS m/z 633.3399 (calcd for
C36H50O8Na 633.3398).
Preparation of Compound 3. To a stirred solution of compound 2
(50 mg, 0.088 mmol) in 4 mL of CH2Cl2 at 0 °C were added pyridine
(300 μL, 3.7 mmol) and acetyl chloride (60 μL, 0.88 mmol, 10 equiv).
The mixture was stirred overnight, MeOH (0.5 mL) was added, and
the mixture was concentrated under vacuum. The crude material was
purified by chromatography on silica gel (elution: from cyclohexane−
EtOAc, 7:3 to 6:4) to yield 3 (52 mg, 75%), identical by all means with
the natural product.
X-ray Crystallographic Analysis of Compound 1c. Data were
collected at low temperature (180 K) on a Bruker Kappa Apex II
diffractometer using graphite-monochromated Mo Kα radiation (λ =
0.71073 Å) and equipped with an Oxford Cryosystems Cryostream
cooler device. The structure was solved by direct methods using
SHELXS-8620 and refined by means of least-squares procedures on F2
with the aid of the program SHELXL9720 included in the software
package WinGX version 1.63.21 The atomic scattering factors were
taken from International Tables for X-ray Crystallography.22 All hy-
drogen atoms were placed geometrically and refined by using a riding
model, except for the atoms H30A and H30B, which were located by
Fourier differences. All non-hydrogen atoms were anisotropically
refined, and in the last cycles of refinement, a weighting scheme was
used, where weights were calculated from the following formula: w =
2
1/[σ2(Fo ) + (aP)2 + bP] where P = (Fo2 + 2Fc2)/3. Drawing was per-
formed with the program ORTEP32 with 30% probability displace-
ment ellipsoids for non-hydrogen atoms.23
Crystal Data for 1c: orthorhombic, C41H53NO8, space group
P212121 with a = 11.5565(7) Å, b = 14.6892(10) Å, c = 22.0210(15) Å,
V = 3738.2(4) Å3, Z = 4, Dcalcd 1.222 g/cm3, m = 0.084 mm−1, F(000) =
1480. Crystal size: 0.35 × 0.2 × 0.08 mm3. Independent reflections:
6975 with Rint = 0.0333. The final agreement factors are R1 = 0.0394
and wR2 = 0.0972 [I > 2σ(I)]. Crystallographic data are deposited at
the Cambridge Crystallographic Data center (deposition no. CCDC
826141).
Preparation of 22,23,26-Trihydroxy-29-nor-24-methyl-16,22-ep-
oxycycloarta-1,24(24a)-diene-3,23-dione (4). To a stirred solution
of compound 1 (20 mg, 0.038 mmol) in CH3CN (300 μL) was added
NaOH (570 μL, 0.437 mmol, c = 1 mol/L, 15 equiv). The mixture was
stirred for 22 h at room temperature, then diluted with EtOAc, and
filtered over Celite. After concentration under vacuum, the product
was purified by chromatography on silica gel (elution: cyclohexane−
1
EtOAc, 1:1) to obtain 4 as a translucent film (3 mg, 17%): H NMR
22-De-O-acetylneoboutomellerone (2): [α]20D −55 (c 0.16, CHCl3);
UV (MeOH) λmax (log ε) 228 (3.71), 266 (3.86) nm; IR (film)
(CD3CN, 500 MHz) δ 6.96 (1H, d, J = 10.1 Hz, H-1), 5.91 (1H, d,
J = 10.1 Hz, H-2), 5.24 (1H, s, H-24a), 5.11 (1H, s, H-24a), 4.66 (1H,
dt, J = 7.2, 8.5 Hz, H-16), 3.97 (1H, d, J = 5.6 Hz, H-23), 3.79 (1H, d,
J = 5.6 Hz, OH-23), 3.48 (1H, s, OH-22), 3.47 (1H, m, H-26a), 3.40
(1H, ddd, J = 9.6, 8.7, 4.6 Hz, H-26b), 3.23 (1H, t, J = 4.7 Hz, OH-26),
2.63 (1H, sext, J = 7.0 Hz, H-25), 2.47 (1H, quint, J = 6.8 Hz, H-20), 1.31
(1H, d, J = 4.6 Hz, H-19a), 1.27 (1H, m, H-7b), 1.13 (3H, s, H-18),
1.03 (3H, d, J = 6.7 Hz, H-28), 1.00 (3H, d, J = 7.0 Hz, H-27), 0.95
(3H, d, J = 6.7 Hz, H-21), 0.92 (3H, s, H-30), 0.50 (1H, d, J = 4.6 Hz,
H-19b); 13C NMR, see Table 1; ESIMS m/z 507.3 [M + Na]+, 991.6
[2M + Na]+.
1
νmax 3453, 1725, 1661, 1601, 1245, 1043 cm−1; H NMR (CDCl3,
500 MHz) δ 6.85 (1H, d, J = 10.1 Hz, H-1), 6.18 (1H, s,
H-24a), 6.02 (1H, s, H-24a), 5.99 (1H, d, J = 10.1 Hz, H-2), 5.35 (1H,
dt, J = 4.6, 7.5 Hz, H-16), 4.74 (1H, dd, J = 5.5, 1.5 Hz, H-22), 3.65
(2H, d, J = 6.1 Hz, H-26), 3.52 (1H, d, J = 5.8 Hz, OH-22), 2.96 (1H,
sext, J = 6.7 Hz, H-25), 2.52 (1H, dd, J = 11.0, 7.3 Hz, H-17), 2.46
(1H, m, H-20), 2.35 (1H, dd, J = 14.0, 7.9 Hz, H-15a), 2.19 (1H, dq,
J = 12.5, 6.7 Hz, H-4), 2.10 (3H, s, OAc-16), 2.05 (1H, m, H-11a),
1.99 (1H, dt, J = 5.2, 13.4 Hz, H-5), 1.97 (1H, dd, J = 10.5, 6.9 Hz,
H-8a), 1.65 (1H, m, H-12a), 1.34 (1H, dd, J = 14.0, 4.4 Hz, H-15b),
1.18 (1H, d, J = 4.5 Hz, H-19a), 1.18 (3H, s, H-18), 1.14 (3H, d,
J = 7.0 Hz, H-27), 1.10 (3H, d, J = 7.0 Hz, H-28), 0.99 (3H, s, H-30),
0.88 (1H, m, H-6b), 0.66 (3H, d, J = 7.0 Hz, H-21), 0.57 (1H, d,
J = 4.5 Hz, H-19b); 13C NMR, see Table 1; HR-ESIMS m/z 549.3179
(calcd for C32H46O6Na 549.3187).
1,2-Dihydroneoboutomellerone (5): yellow gum; [α]20D +36 (c 0.08,
CHCl3); UV (MeOH) λmax (log ε) 216 (3.78), 290 (2.63) nm; IR
(film) νmax 3513, 2965, 2871, 1734, 1705, 1452, 1376, 1231 cm−1; 1H
NMR (CD3CN, 500 MHz) δ 6.05 (1H, s, H-24a), 5.90 (1H, d, J = 0.9
Hz, H-24a), 5.53 (1H, d, J = 2.1 Hz, H-22), 5.08 (1H, dt, J = 4.3, 7.6
Hz, H-16), 3.53 (1H, dt, J = 10.6, 5.8 Hz, H-26a), 3.39 (1H, dt,
J = 10.6, 6.1 Hz, H-26b), 2.77 (1H, sext, J = 6.7 Hz, H-25), 2.68 (1H,
t, J = 5.6 Hz, OH-26), 2.60 (1H, ddq, J = 10.7, 1.8, 6.7 Hz, H-20), 2.43
(1H, td, J = 13.6, 6.4 Hz, H-2a), 2.20 (1H, dd, J = 14.0, 7.9 Hz,
H-15a), 2.09 (3H, s, OAc-22), 2.11 (1H, m, H-11a), 2.03 (3H, s, OAc-16),
1.82 (1H, m, H-1a), 1.60−1.55 (2H, m, H-1b, H-5), 1.36 (1H, dd, J =
14.5, 4.1 Hz, H-15b), 1.23 (3H, s, H-18), 1.10 (1H, m, H-7b), 1.03
(3H, d, J = 7.0 Hz, H-27), 0.97 (3H, s, H-30), 0.91 (3H, d, J = 6.7 Hz,
H-28), 0.84 (3H, d, J = 7.0 Hz, H-21), 0.75 (1H, dq, J = 2.4, 12.6 Hz,
H-6b), 0.65 (1H, br d, J = 4.0 Hz, H-19a), 0.47 (1H, d, J = 4.0 Hz,
H-19b); 13C NMR, see Table 1; HR-ESIMS m/z 593.3451 (calcd for
C34H50O7Na 593.3449).
Preparation of Mosher Ester Derivatives 1a, 1b, 2a, and 2b. Com-
pounds 1 and 2 (10 mg) in dry pyridine (2.0 mL) were tran-
sferred into an HPLC vial (2 mL) capped with a septum and were
treated with 30 μL (175 μmol) of (S)-(+)-α-methoxy-α-trifluorome-
thylphenylacetyl chloride (MTPA chloride). The mixtures were stirred
at room temperature for 2 h. After concentration under reduced
pressure, they were purified by semipreparative HPLC RP-18 chroma-
tography, eluting with a linear gradient (55−100% B) to afford 1a and
2a. Derivatives 1b and 2b were prepared in a similar fashion by using
(R)-(−)-α-methoxy-α-trifluoromethylphenylacetyl chloride.
1-(R)-MTPA ester (1a): 1H NMR (CD3CN, 500 MHz) δ 4.39 (1H,
dd, J = 6.2, 10.9 Hz, H-26a), 4.27 (1H, dd, J = 5.7, 10.9 Hz, H-26b);
ESIMS m/z 807 [M + Na]+.
1,2-Dihydro-22-de-O-acetylneoboutomellerone (6): yellow gum;
[α]20D +11 (c 0.12, CHCl3); UV (MeOH) λmax (log ε) 214 (3.74), 262
(3.23) nm; IR (film) νmax 3435, 2932, 2873, 1731, 1705, 1700, 1452,
1-(S)-MTPA ester (1b): 1H NMR (CD3CN, 500 MHz) δ 4.34 (2H,
AB part of an ABX system, H-26); ESIMS m/z 807 [M + Na]+.
2-(R)-MTPA ester (2a): 1H NMR (methanol-d4, 500 MHz) δ 4.44
(1H, dd, J = 6.0, 10.9 Hz, H-26a), 4.35 (1H, dd, J = 5.1, 10.9 Hz, H-
26b), 2.59 (1H, m, H-20); ESIMS m/z 981 [M + Na]+.
1
1376, 1237, 1022, 997 cm−1; H NMR (CD3CN, 500 MHz) δ 6.12
(1H, s, H-24a), 5.99 (1H, d, J = 0.9 Hz, H-24a), 5.20 (1H, dt, J = 4.4,
7.6 Hz, H-16), 4.72 (1H, dd, J = 6.1, 2.1 Hz, H-22), 3.56 (1H, dt, J =
10.5, 5.8 Hz, H-26a), 3.54 (1H, d, J = 6.1 Hz, OH-22), 3.41 (1H, dt,
43
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J = 10.5, 5.8 Hz, H-26b), 2.83 (1H, sext, J = 6.7 Hz, H-25), 2.67 (1H,
t, J = 5.8 Hz, OH-26), 2.07 (1H, m, H-11a), 2.03 (3H, s, OAc-16),
1.82 (1H, m, H-1a), 1.37 (1H, ddd, J = 14.2, 4.4, 0.9 Hz, H-15b), 1.22
(3H, s, H-18), 1.12 (1H, dq, J = 2.7, 12.8 Hz, H-7b), 1.05 (3H, d,
J = 7.3 Hz, H-27), 0.97 (3H, d, J = 0.6 Hz, H-30), 0.91 (3H, d, J = 6.7
Hz, H-28), 0.75 (1H, dq, J = 2.4, 12.5 Hz, H-6b), 0.65 (1H, m,
H-19a), 0.64 (3H, d, J = 6.4 Hz, H-21), 0.47 (1H, d, J = 4.0 Hz,
H-19b); 13C NMR, see Table 1; HR-ESIMS m/z 551.3342 (calcd for
C32H48O6Na 551.3343).
(2H, m, H-11a, H-19a), 1.40 (1H, ddd, J = 15.9, 3.1, 1.6 Hz,
H-11b), 1.23 (3H, d, J = 7.0 Hz, H-28), 1.20 (3H, s, H-18), 1.03 (3H,
d, J = 7.0 Hz, H-27), 0.94 (3H, s, H-30), 0.86 (3H, d, J = 7.0 Hz,
H-21), 0.03 (1H, d, J = 4.0 Hz, H-19b); 13C NMR, see Table 2; HR-
ESIMS m/z 605.3091 (calcd for C34H46O8Na 605.3085).
1,2-Dihydro-1α-hydroxy-22-de-O-acetylneoboutomellerone
(11): yellow gum; [α]20D +30 (c 0.12, CHCl3); UV (MeOH) λmax (log ε)
216 (3.920) nm; IR (film) νmax 3401, 2934, 2876, 1707, 1669, 1376,
1237, 1021 cm−1; 1H NMR (CD3CN, 500 MHz) δ 6.12 (1H, s,
H-24a), 5.99 (1H, d, J = 0.9 Hz, H-24a), 5.21 (1H, dt, J = 4.3, 7.5 Hz,
H-16), 4.72 (1H, dd, J = 5.3, 1.7 Hz, H-22), 3.80 (1H, t, J = 3.1 Hz, H-
1), 3.55 (2H, dd, J = 10.9, 6.1 Hz, H-26a + OH-22), 3.42 (1H, dd,
J = 10.9, 7 Hz, H-26b), 2.83 (1H, sext, J = 6.4 Hz, H-25), 2.80 (1H, br
s, OH-1), 2.69 (1H, br s, OH-26), 2.64 (1H, ddd, J = 14.0, 3.7, 0.5 Hz,
H-2a), 2.46 (1H, ddq, J = 11.0, 2.1, 6.7 Hz, H-20), 2.42 (1H, dd,
J = 11.0, 7.0 Hz, H-17), 2.14 (1H, m, H-5), 2.03 (3H, s, OAc-16), 1.74
(1H, m, H-6a), 1.21 (3H, s, H-18), 1.05 (3H, d, J = 7.0 Hz, H-27),
1.01 (3H, s, H-30), 0.92 (3H, d, J = 6.4 Hz, H-28), 0.80 (1H, dq,
J = 2.4, 12.6 Hz, H-6b), 0.73 (1H, d, J = 4.3 Hz, H-19a), 0.64 (3H, d,
J = 6.6 Hz, H-21), 0.48 (1H, d, J = 4.3 Hz, H-19b), 13C NMR, see
Table 2; HR-ESIMS m/z 567.3296 (calcd for C32H48O7Na 567.3292).
6β-Hydroxyneoboutomellerone (7): yellow gum; [α]20D −43 (c 0.13,
CHCl3); UV (MeOH) λmax (log ε) 212 (3.84), 220 (3.84), 266 (3.87)
nm; IR (film) νmax 3409, 2937, 2876, 1733, 1662, 1454, 1374, 1232,
1
1022 cm−1; H NMR (CDCl3, 500 MHz) δ 6.76 (1H, d, J = 10 Hz,
H-1), 6.14 (1H, s, H-24a), 5.96 (1H, d, J = 10 Hz, H-2), 5.90 (1H, s,
H-24a), 5.59 (1H, d, J = 2.1 Hz, H-22), 5.10 (1H, dt, J = 4.4, 7.7 Hz,
H-16), 4.20 (1H, br s, H-6), 3.60 (2H, d, J = 5.8 Hz, H-26), 2.89 (1H,
sext, J = 6.4 Hz, H-25), 2.35 (1H, dd, J = 14.0, 7.9 Hz, H-15a), 2.20
(1H, m, H-11a), 2.16 (3H, s, OAc-22), 2.08 (3H, s, OAc-16), 1.99
(1H, m, H-5), 1.86 (1H, d, J = 3.7 Hz, H-19a), 1.76 (2H, m, 2 H-12),
1.56 (1H, m, H-7a), 1.48 (1H, dt, J = 1.7, 13.3 Hz, H-7b), 1.25 (3H, s,
H-18), 1.24 (3H, d, J = 7.0 Hz, H-28), 1.13 (3H, d, J = 7.0 Hz, H-27),
1.0 (3H, s, H-30), 0.91 (3H, d, J = 6.7 Hz, H-21), 0.72 (1H, d, J = 3.7
Hz, H-19b); 13C NMR, see Table 1; HR-ESIMS m/z 607.3243 (calcd
for C34H48O8Na 607.3241).
25-Epi-neoboutomellerone (12): yellow gum; [α]20 +9 (c 0.03,
D
CHCl3); UV (MeOH) λmax (log ε) 210 (3.54), 264 (3.54) nm; IR
(film) νmax 3386, 2927, 1732, 1667, 1570, 1375, 1233, 1022 cm−1; 1H
NMR (CDCl3, 500 MHz) δ 6.82 (1H, d, J = 9.8 Hz, H-1), 6.11 (1H, s,
H-24a), 5.98 (1H, d, J = 9.8 Hz, H-2), 5.83 (1H, d, J = 0.9 Hz, H-24a),
5.52 (1H, d, J = 2.1 Hz, H-22), 5.07 (1H, dt, J = 4.4, 7.7 Hz, H-16),
3.56 (2H, t, J = 6.0 Hz, 2 H-26), 2.96 (1H, sext, J = 6.7 Hz, H-25), 2.59
(1H, m, H-20), 2.28 (1H, dd, J = 7.4, 14.4 Hz, H-17), 2.25 (1H, m,
H-15a), 2.18 (1H, m, H-4), 2.14 (3H, s, OAc-22), 2.06 (3H, s, OAc-
16), 1.45 (1H, m, H-7), 1.30 (1H, dd, J = 14.2, 4.7 Hz, H-15b), 1.18
(1H, d, J = 4.6 Hz, H-19a),1.16 (3H, s, H-18), 1.09 (3H, d, J = 7.0 Hz,
H-27), 1.08 (3H, d, J = 6.7 Hz, H-28), 0.93 (3H, s, H-30), 0.89 (3H, d,
J = 7.0 Hz, H-21), 0.55 (1H, d, J = 4.6 Hz, H-19); 13C NMR, see Table
2; HR-ESIMS m/z 591.3291 (calcd for C34H48O7Na, 591.3292).
Preparation of 25ξ-Neoboutomelleron-26-al (13). To a stirred
solution of 1 (100 mg, 0.18 mmol) in CH2Cl2 (3.5 mL) was added
350 μL of pyridine (2 mL/mmol), and the mixture was cooled to 0 °C.
The Dess-Martin reagent (1.76 mL, 0.528 mmol, c = 3 mol/L, 3
equiv) was added dropwise, and the mixture was allowed to warm.
After 3 h, the mixture was diluted with EtOAc, and a saturated solution
of Na2S2O3 was added. The aqueous layer was extracted three times
with EtOAc (10 mL), and the combined organic layers were washed
successively with NaHCO3 (10 mL) and brine. The organic solution
was dried over MgSO4 and concentrated under vacuum. The residue
was purified by chromatography on silica gel (cyclohexane−EtOAc,
7:3 to 6:4) to give 13 (66 mg, 66%). 1H NMR (500 MHz, CD3CN) δ
9.52 (1H, d, J = 0.6 Hz, H-26), 6.94 (1H, d, J = 10.1 Hz, H-1), 6.31
(1H, s, H-24a), 6.06 (1H, s, H-24), 5.90 (1H, d, J = 10.1 Hz, H-2),
5.58 (1H, d, J = 2.1 Hz, H-22), 5.10 (1H, dt, J = 4.4, 7.7 Hz, H-16),
3.45 (1H, q, J = 7.2 Hz, H-25), 2.64 (1H, m, H-20), 2.31 (1H, dd,
J = 11.0, 7.6 Hz, H-17), 2.09 (3H, s, OAc-22), 2.04 (3H, s, OAc-16),
1.38 (1H, dd, J = 13.7, 4.0 Hz, H-15b), 1.25 (1H, d, J = 4.4 Hz,
H-19a), 1.18 - 1.22 (7H, m, H-7, -27, -18), 1.03 (3H, d, J = 6.7 Hz,
H-28), 0.96 (3H, s, H-30), 0.94 (1H, qd, J = 12.8, 3.7 Hz, H-6b), 0.85
(3H, d, J = 7.0 Hz, H-21), 0.58 (1H, d, J = 4.4 Hz, H-19b);); 13C
NMR, see Table 2; ESI-MS m/z 567.3 [M + H]+.
6β-Hydroxy-22-de-O-acetylneoboutomellerone (8): yellow gum;
[α]20D −8 (c 0.14, CHCl3); UV (MeOH) λmax (log ε) 210 (3.94), 266
(3.78) nm; IR (film) νmax 3435, 2934, 2876, 1729, 1666, 1453, 1375,
1236, 1023 cm−1; 1H NMR (CDCl3, 500 MHz)) δ 6.76 (1H, d, J = 10
Hz, H-1), 6.18 (1H, s, H-24a), 6.03 (1H, s, H-24a), 5.94 (1H, d, J = 10
Hz, H-2), 5.35 (1H, dt, J = 4.4, 7.1 Hz, H-16), 4.74 (1H, s, H-22), 4.19
(1H, br s, H-6), 3.64 (2H, d, J = 6.1 Hz, H-26), 3.59 (1H, d, J = 5.5
Hz, OH-22), 2.96 (1H, sext, J = 6.4 Hz, H-25), 2.58 (1H, m, H-4),
2.52 (1H, m, H-17), 2.45 (1H, m, H-20), 2.37 (1H, dd, J = 13.7, 7.9
Hz, H-15a), 2.30 (1H, dd, J = 12.7, 4.4 Hz, H-8), 2.22 (1H, m, H-11a),
2.09 (3H, s, OAc-16), 1.99 (1H, dd, J = 12.8, 2.4 Hz, H-5), 1.84 (1H,
d, J = 3.1 Hz, H-19a), 1.75 (2H, m, 2 H-12), 1.58 (1H, dt, J = 4.4, 12.9
Hz, H-7a), 1.48 (1H, t, J = 13.1 Hz, H-7b), 1.40−1.20 (2H, m, H-15b,
H-11b), 1.24 (3H, d, J = 7.3 Hz, H-28), 1.23 (3H, br s, H-18), 1.14
(3H, d, J = 7.0 Hz, H-27), 1.04 (3H, s, H-30), 0.70 (1H, d, J = 3.1 Hz,
H-19b), 0.67 (3H, d, J = 6.4 Hz, H-21); 13C NMR, see Table 1; HR-
ESIMS m/z 565.3133 (calcd for C32H46O7 Na 565.3136).
18-Hydroxyneoboutomellerone (9): yellow gum; [α]20D −16 (c 0.03,
1
CHCl3); UV (MeOH) λmax (log ε) 214 (3.14), 266 (2.96) nm; H
NMR (CDCl3, 500 MHz) δ 6.80 (1H, d, J = 10 Hz, H-1), 6.07 (1H, s,
H-24a), 5.98 (1H, d, J = 10 Hz, H-2), 5.86 (1H, br d, H-24), 5.57 (1H,
d, J = 2.1 Hz, H-22), 5.18 (1H, dt, J = 4.3, 7.6 Hz, H-16), 4.06 (1H, m,
H-18a), 3.94 (1H, d, J = 12.2 Hz, H-18b), 3.58 (2H, m, 2 H-26), 2.86
(1H, sext, J = 7.0 Hz, H-25), 2.72 (1H, m, H-20), 2.36 (1H, dd,
J = 11.3, 7.6 Hz, H-17), 2.28 (1H, dd, J = 14.3, 7.9 Hz, H-15a), 2.19
(1H, m, H-4), 2.14 (3H, s, OAc-22), 2.09 (3H, s, OAc-16), 2.03 (2H, m,
H-5, H-8), 1.85 (1H, m, H-12a), 1.70 (1H, m, H-6a), 1.63 (1H, m, H-
12b), 1.60 (1H, m, H-15b), 1.35 (2H, m, H-7a, H-11b), 1.19 (1H, d,
J = 4.3 Hz, H-19a), 1.16 (1H, m, H-7b), 1.11 (3H, d, J = 7.3 Hz,
H-27), 1.09 (3H, d, J = 6.9 Hz, H-28), 1.00 (3H, s, H-30), 0.93 (3H, d,
J = 6.7 Hz, H-21), 0.90 (1H, m, H-6b), 0.60 (1H, d, J = 4.3 Hz,
H-19b); 13C NMR, see Table 2; HR-ESIMS m/z 607.3216 (calcd for
C34H48O8Na 607.3241).
6β,7β-Oxidoneoboutomellerone (10): yellow gum; [α]20 −86
9,10-Di-epi-25ξ-neoboutomellerone (14): yellow gum; [α]20 −48
D
D
(c 0.16, CHCl3); UV (MeOH) λmax (log ε) 224 (3.92), 262 (3.93) nm;
IR (film) νmax 3411, 2966, 2878, 1735, 1669, 1375, 1233, 1022 cm−1;
1H NMR (CD3CN, 500 MHz) δ 6.94 (1H, d, J = 10 Hz, H-1), 6.06
(1H, s, H-24a), 5.91 (1H, d, J = 10.0 Hz, H-2), 5.90 (1H, br s, H-24a),
5.54 (1H, d, J = 2.1 Hz, H-22), 5.16 (1H, dt, J = 4.7, 7.7 Hz, H-16),
3.53 (1H, dd, J = 10.4, 6.4 Hz, H-26a), 3.39 (1H, dd, J = 10.4, 6.6 Hz,
H-26b), 3.15 (1H, dd, J = 4.3, 1.5 Hz, H-6), 3.00 (1H, dd, J = 4.3, 1.8
Hz, H-7), 2.83 (1H, br d, J = 1.2 Hz, H-8), 2.77 (1H, sext, J = 6.7 Hz,
H-25), 2.70 (1H, br s, OH-26), 2.63 (1H, ddq, J = 10.8, 2.1, 7.0 Hz,
H-20), 2.55 (1H, dq, J = 12.5, 7.0 Hz, H-4), 2.45 (1H, br d, J = 11.3 Hz,
H-5), 2.33 (1H, dd, J = 13.4, 7.9 Hz, H-15a), 2.28 (1H, dd, J = 10.8,
7.8 Hz, H-17), 2.08 (3H, s, OAc-22), 2.06 (3H, s, OAc-16), 2.04
(c 0.13, CHCl3); UV (MeOH) λmax (log ε) 210 (3.73), 270 (3.93) nm; IR
(film) νmax 3417, 2937, 2875, 1734, 1666, 1373, 1233, 1022 cm−1; 1H
NMR (CDCl3, 500 MHz) δ 6.39 (1H, d, J = 10.4 Hz, H-1), 6.13 (1H,
s, H-24a), 5.99 (1H, d, J = 10.4 Hz, H-2), 5.88 (1H, d, J = 0.6 Hz, H-
24a), 5.58 (1H, d, J = 2.1 Hz, H-22), 5.15 (1H, dt, J = 5.0, 7.7 Hz, H-
16), 3.60 (2H, t, J = 5.5 Hz, 2 H-26), 2.88 (1H, sext, J = 6.4 Hz, H-25),
2.62 (1H, m, H-20), 2.28 (1H, dd, J = 11.9, 7.3 Hz, H-17), 2.26 (1H,
m, H-15a), 2.20 (1H, m, H-4), 2.16 (3H, s, OAc-22), 2.10 (3H, s,
OAc-16), 1.96 (1H, m, H-8), 1.85 (1H, dt, J = 5.0, 13.0 Hz, H-12a),
1.6 (1H, dd, J = 13.0, 4.1 Hz, H-12a), 1.28 (1H, d, J = 5.0 Hz, H-19a),
1.14 (3H, s, H-18), 1.12 (3H, d, J = 6.1 Hz, H-28), 1.10 (3H, s,
H-27), 0.93 (3H, br s, H-30), 0.91 (3H, d, J = 7.0 Hz, H-21), 0.88
44
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(1H, d, J = 5.0 Hz, H-19b); 13C NMR, see Table 2; HR-ESIMS m/z
591.3295 (calcd for C34H48O7Na, 591.3292).
δ 6.03 (1H, s, H-24a), 5.94 (1H, d, J = 1.1 Hz, H-24a), 5.20 (1H, td,
J = 4.2, 7.7 Hz, H-16), 4.70 (1H, d, J = 1.4 Hz, H-22), 2.86 (1H, dsept,
J = 0.9, 7.0 Hz, H-25), 2.03 (3H, s, OAc-16), 1.86−1.78 (2H, m, H-1,
H-12), 1.22 (3H, s, H-18), 1.10 (3H, d, J = 6.7 Hz, H-27), 1.02 (3H, d,
J = 6.7 Hz, H-26), 0.97 (1H, d, J = 0.6 Hz, H-30), 0.91 (3H, d, J =
6.8 Hz, H-28), 0.76 (1H, dq, J = 2.3, 12.5 Hz, H-6b), 0.64 (1H, m,
H-19a), 0.64 (3H, d, J = 6.1 Hz, H-21), 0.47 (1H, d, J = 4.0 Hz,
H-19b); 13C NMR, see Table 3; HR-ESIMS m/z 535.3393 (calcd for
C32H48O5Na, 535.3394).
9,10-Di-epi-22-de-O-acetyl-25ξ-neoboutomellerone (15): yellow
gum; [α]20 −53 (c 0.13, CHCl3); UV (MeOH) λmax (log ε)
D
210 (4.12), 270 (4.20) nm; IR (film) νmax 3421, 2937, 2877, 1732,
1
1666, 1609, 1453, 1376, 1240, 1024 cm−1; H NMR (CDCl3, 500
MHz) δ 6.39 (1H, d, J = 10.1 Hz, H-1), 6.19 (1H, s, H-24a), 6.02 (1H,
s, H-24a), 5.99 (1H, d, J = 10.1 Hz, H-2), 5.39 (1H, dt, J = 5.0, 7.6 Hz,
H-16), 4.73 (1H, br s, H-22), 3.64 (1H, d, J = 5.8 Hz, 2 H-26), 3.54
(1H, br s, OH-22), 2.96 (1H, sext, J = 6.7 Hz, H-25), 2.53 (1H, dd,
J = 11.0, 7.6 Hz, H-17), 2.48 (1H, m, H-20), 2.31 (1H, dd, J = 13.3, 7.8
Hz, H-15), 2.11 (3H, s, OAc-16), 1.84 (1H, dt, J = 4.7,, 13.4 Hz,
H-12), 1.60 (1H, ddd, J = 13.4, 4.0, 1.5 Hz, H-12b), 1.33 (1H, m,
H-15), 1.28 (1H, d, J = 4.9 Hz, H-19a), 1.13 (6H, d, J = 7.0 Hz, H-27,
H-28); 1.12 (3H, s, H-18), 0.97 (3H, s, H-30), 0.88 (1H, d, J = 4.9 Hz,
H-19b), 0.67 (3H, d, J = 6.4 Hz, H-21); 13C NMR, see Table 2; HR-
ESIMS m/z 549.3190 (calcd for C32H46O6Na, 549.3187).
9,10-Di-epi-26-deoxyneoboutomellerone (20): yellow gum;
[α]20 −98 (c 0.11, CHCl3); UV (MeOH) λmax (log ε) 212 (4.08),
D
270 (4.21) nm; IR (film) νmax 3451, 2936, 2873, 2876, 1734, 1667,
1
1609, 1452, 1374, 1238 cm−1; H NMR (CD3CN, 500 MHz) δ 6.50
(1H, d, J = 10.2 Hz, H-1), 5.95 (1H, s, H-24a), 5.88 (1H, d, J = 10.2
Hz, H-2), 5.82 (1H, d, J = 1.2 Hz, H-24a), 5.52 (1H, d, J = 2.0 Hz,
H-22), 5.14 (1H, dt, J = 4.5, 7.7 Hz, H-16), 2.81 (1H, dsept, J = 1.0,
7.0 Hz, H-25), 2.57 (1H, ddq, J = 10.9, 2.1, 6.9 Hz, H-20), 2.29 (1H,
dd, J = 10.8, 7.7 Hz, H-17), 2.22 (2H, m, H-4, H-8a), 2.09 (3H, s,
OAc-22), 2.04 (3H, s, OAc-16), 2.00 (1H, m, H-7), 1.83 (1H, dt,
J = 4.4, 12.5, H-12a), 1.63 (1H, ddd, J = 2.0, 5.0, 13.0 Hz, H-12b), 1.42
(1H, ddd, J = 2.2, 5.0, 14.0, H-11b), 1.36 (1H, dd, J = 4.7, 14.0 Hz,
H-15b) 1.30 (1H, d, J = 4.7 Hz, H-19a), 1.13 (3H, s, H-18), 1.07 (3H,
d, J = 7.0 Hz, H-26), 1.04 (3H, d, J = 6.6 Hz, H-28), 1.00 (3H, d,
J = 6.9 Hz, H-27), 0.93 (3H, s, H-30), 0.92 (1H, m, H-19), 0.86 (3H,
d, J = 6.9 Hz, H-21); 13C NMR, see Table 3; HR-ESIMS m/z
575.3338 (calcd for C34H48O6Na, 575.3343).
26-Deoxyneoboutomellerone (16): yellow gum; [α]20 −102
D
(c 0.08, CHCl3); UV (MeOH) λmax (log ε) 220 (3.95), 264 (3.77) nm;
IR (film) νm1ax 3437, 2962, 2931, 2873, 1734, 1669, 1599, 1455, 1407,
1237 cm−1; H NMR (CD3CN, 500 MHz) δ 6.94 (1H, d, J = 10 Hz,
H-1), 5.94 (1H, s, H-24a), 5.90 (1H, d, J = 10.0 Hz, H-2), 5.83 (1H, d,
J = 1.2 Hz, H-24a), 5.51 (1H, d, J = 2.4 Hz, H-22), 5.09 (1H, dt,
J = 4.3, 7.6 Hz, H-16), 2.81 (1H, dsept, J = 0.9, 7.0 Hz, H-25), 2.56
(1H, ddq, J = 11.0, 2.1, 7.0 Hz, H-20), 2.29 (1H, dd, J = 11.0,, 7.3 Hz,
H-17), 2.18 (2H, m, H-4, H-15a), 2.09 (3H, s, OAc-22), 2.02 (3H, s,
OAc-16), 1.56 (1H, ddd, J = 7.0, 9.0, 15.0 Hz, H-11b), 1.44 (1H, ddt,
J = 7.0, 13.5, 4.0 Hz, H-7a), 1.36 (1H, ddd, J = 14.0, 4.3, 0.9 Hz,
H-15b), 1.24 (1H, d, J = 4.6 Hz, H-19a), 1.18 (3H, s, H-18), 1.07 (3H,
d, J = 6.7 Hz, H-26), 1.02 (3H, d, J = 7.0 Hz, H-28), 1.00 (3H, d,
J = 7.0 Hz, H-27), 0.95 (3H, s, H-30), 0.85 (3H, d, J = 7.0 Hz, H-21),
0.58 (1H, d, J = 4.6 Hz, H-19b); 13C NMR, see Table 2; HR-ESIMS
m/z 575.3341 (calcd for C34H48O6Na, 575.3343).
24a-Nor-24,25-didehydro-26-deoxyneoboutomellerone (21):
yellow gum; [α]20 −73 (c 0.03, CHCl3); UV (MeOH) λmax (log ε)
D
242 (3.96) nm; IR (film) νmax 3433, 2932, 2931, 2874, 1735, 1671,
1615, 1378, 1234 cm−1; 1H NMR (CD3CN, 500 MHz) δ 6.94 (1H, d,
J = 10.1 Hz, H-1), 6.14 (1H, sept, J = 1.3 Hz, H-24), 5.90 (1H, d,
J = 10.1 Hz, H-2), 5.10 (1H, dt, J = 4.6, 7.8 Hz, H-16), 4.85 (1H, d,
J = 1.8 Hz, H-22), 2.59 (1H, ddq, J = 10.9, 1.5, 7.0 Hz, H-20), 2.12
(3H, d, J = 1.2 Hz, H-27), 2.09 (3H, s, OAc-22), 2.08 (3H, s, OAc-16),
1.98 (3H, m, H-26), 1.26 (1H, d, J = 4.3 Hz, H-19a), 1.20 (3H, s,
H-18), 1.02 (3H, d, J = 6.7 Hz, H-28), 0.93 (3H, d, J = 0.9 Hz, H-30),
0.85 (3H, d, J = 7.0 Hz, H-21), 0.56 (1H, d, J = 4.3 Hz, H-19b); 13C
NMR, see Table 1; HR-ESIMS m/z 561.3182 (calcd for C33H46NaO6,
561.3187).
22-De-O-acetyl-26-deoxyneoboutomellerone (17): yellow gum;
[α]20 −91 (c 0.12, CHCl3); UV (MeOH) λmax (log ε) 220 (3.87),
D
266 (3.86) nm; IR (film) νmax 3457, 2933, 2931, 2873, 1734, 1669,
1456, 1375, 1237 cm−1; 1H NMR (CD3CN, 500 MHz) δ 6.93 (1H, d,
J = 10.0 Hz, H-1), 6.02 (1H, s, H-24a), 5.93 (1H, d, J = 1.2 Hz,
H-24a), 5.89 (1H, d, J = 10.0 Hz, H-2), 5.20 (1H, dt, J = 4.4, 7.6 Hz,
H-16), 4.71 (1H, d, J = 5.8 Hz, H-22), 3.54 (1H, d, J = 5.8 Hz, OH-
22), 2.85 (1H, dsept, J = 0.9, 6.9 Hz, H-25), 2.42 (2H, m, H-17,
H-20), 2.22 (1H, dd, J = 13.9, 7.8 Hz, H-15a), 2.17 (1H, dq, J = 12.7,
6.9 Hz, H-4), 2.02 (3H, s, OAc-16), 1.54 (1H, ddd, J = 7.0, 8.5, 15.0
Hz, H-11), 1.45 (1H, ddt, J = 7.0, 14.0, 4.0 Hz, H-7a), 1.37 (1H, ddd,
J = 13.9, 4.4, 1.0 Hz, H-15), 1.23 (1H, d, J = 4.3 Hz, H-19a), 1.17 (3H,
s, H-18), 1.09 (3H, d, J = 7.0 Hz, H-26), 1.03 (3H, d, J = 6.7 Hz,
H-28), 1.02 (3H, d, J = 6.7 Hz, H-27), 0.96 (3H, d, J = 0.6 Hz, H-30),
0.64 (3H, d, J = 6.4 Hz, H-21), 0.57 (1H, d, J = 4.3 Hz, H-19b); 13C
NMR, see Table 3; HR-ESIMS m/z 533.3234 (calcd for C32H46O5Na,
533.3237).
22-De-O-acetyl-24a-nor-24,25-didehydro-26-deoxyneobouto-
mellerone (22): yellow gum; [α]20 −80 (c 0.04, CHCl3); UV
D
(MeOH) λmax (log ε) 242 (3.95) nm; IR (film) νmax 3464, 2933, 2874,
1
1732, 1671, 1618, 1449, 1378, 1237 cm−1; H NMR (CD3CN, 500
MHz) δ 6.94 (1H, d, J = 10 Hz, H-1), 6.17 (1H, sept, J = 1.3 Hz,
H-24), 5.89 (1H, d, J = 10.0 Hz, H-2), 5.19 (1H, dt, J = 4.7, 7.9 Hz,
H-16), 3.99 (1H, dd, J = 4.9, 1.2 Hz, H-22), 3.54 (1H, d, J = 5.2 Hz,
OH-22), 2.47 (1H, ddq, J = 11.3, 1.8, 7.0 Hz, H-20), 2.35 (1H, dd,
J = 11.0, 7.6 Hz, H-17), 2.18 (1H, m, H-4), 2.17 (3H, d, J = 1.2 Hz,
H-27), 2.05 (3H, s, OAc-16), 1.94 (3H, m, H-26), 1.56 (1H, ddd,
J = 7.5, 8.5, 14.0 Hz, H-11), 1.47 (1H, m, H-7), 1.39 (1H, dd, J = 13.6,
5.0 Hz, H-15), 1.25 (1H, d, J = 4.6 Hz, H-19a), 1.20 (3H, s, H-18),
1.02 (3H, d, J = 6.7 Hz, H-28), 0.95 (3H, d, J = 0.6 Hz, H-30), 0.65
(3H, d, J = 6.7 Hz, H-21), 0.56 (1H, d, J = 4.6 Hz, H-19b); 13C NMR,
see Table 3; HR-ESIMS m/z 519.3076 (calcd for C31H44O5Na,
519.3081).
6β-Hydroxy-26-deoxyneoboutomellerone (18): yellow gum;
[α]20 −61 (c 0.12, CHCl3); UV (MeOH) λmax (log ε) 220 (3.83),
D
266 (3.74) nm; IR (film) νmax 3414, 2965, 2939, 2876, 1737, 1668,
1600, 1376, 1232 cm−1; 1H NMR (CD3CN, 500 MHz) δ 6.84 (1H, d,
J = 9.9 Hz, H-1), 5.95 (1H, s, H-24a), 5.84 (1H, d, J = 1.2 Hz, H-24a),
5.82 (1H, d, J = 9.9 Hz, H-2), 5.53 (1H, d, J = 2.1 Hz, H-22), 5.10
(1H, dt, J = 4.3, 7.6 Hz, H-16), 4.05 (1H, br s, H-6), 2.81 (1H, dsept,
J = 1.0, 6.7 Hz, H-25), 2.61 (1H, d, J = 3.4 Hz, OH-6), 2.57 (1H, ddq,
J = 2.2, 10.0, 6.9 Hz, H-20), 2.48 (1H, dq, J = 13.1, 6.7 Hz, H-4), 2.32
(1H, dd, J = 11.0, 7.6 Hz, H-17), 2.29 (1H, dd, J = 12.8, 4.9 Hz, H-8a),
2.22 (1H, dd, J = 13.7, 7.9 Hz, H-15a), 2.21 (1H, m, H-11), 2.10 (3H,
s, OAc-22), 2.03 (3H, s, OAc-16), 1.81 (1H, d, J = 3.1 Hz, H-19a),
1.75 (2H, m, 2 H-12), 1.52 (1H, dt, J = 13.4, 4.7 Hz, H-7), 1.24 (3H, s,
H-18), 1.14 (3H, d, J = 6.7 Hz, H-28), 1.08 (3H, d, J = 6.7 Hz, H-26),
1.01 (3H, d, J = 7.0 Hz, H-27), 1.00 (3H, s, H-30), 0.86 (3H, d, J =
7.0 Hz, H-21), 0.67 (1H, d, J = 3.4 Hz, H-19b); 13C NMR, see Table
3; HR-ESIMS m/z 591.3287 (calcd for C34H48O7Na, 591.3292).
1,2-Dihydro-22-de-O-acetyl-26-deoxyneoboutomellerone (19): yel-
low gum; [α]20D +19 (c 0.09, CHCl3); IR (film) νmax 3401, 2930, 2871,
9,10-Di-epi-24a-nor-24,25-didehydro-26-deoxyneoboutomeller-
one (23): yellow gum; [α]20D −55 (c 0.07, CHCl3); UV (MeOH) λmax
(log ε) 210 (3.77), 244 (3.98) nm; IR (film) νmax 3409, 2936, 2874,
1
1734, 1669, 1612, 1447, 1374, 1235 cm−1; H NMR (CD3CN, 500
MHz) δ 6.50 (1H, d, J = 10.1 Hz, H-1), 6.15 (1H, sept, J = 1.2 Hz,
H-24), 5.88 (1H, d, J = 10.1 Hz, H-2), 5.16 (1H, dt, J = 4.8, 7.7 Hz,
H-16), 4.86 (1H, d, J = 1.7 Hz, H-22), 2.62 (1H, ddq, J = 11.0, 1.5, 6.9
Hz, H-20), 2.12 (3H, d, J = 1.1 Hz, H-27), 2.10 (6H, 2 s, OAc-16,
OAc-22), 1.44−1.35 (2H, m, H-11, H-15), 1.30 (1H, d, J = 4.9 Hz,
H-19a), 1.15 (3H, s, H-18), 1.05 (3H, d, J = 6.8 Hz, H-28), 0.93
(1H, m, H-19b), 0.91 (3H, d, J = 1.0 Hz, H-30), 0.86 (3H, d, J = 7.0
Hz, H-21); 13C NMR, see Table 3; HR-ESIMS m/z 561.3187 (calcd
for C33H46O6Na, 561.3187).
23,24,24a,25,26,27-Hexa-nor-neoboutomelleron-22-al (24): yellow
1
1734, 1707, 1453, 1377, 1240 cm−1; H NMR (CD3CN, 500 MHz)
gum; [α]20 −74 (c 0.11, CHCl3); UV (MeOH) λmax (log ε) 210
D
45
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
(3.51), 266 (3.83) nm; IR (film) νmax 3640, 2938, 2874, 1732, 1669,
H-3), 2.87 (1H, dsept, J = 1.0, 6.8 Hz, H-25), 2.72 (1H, dd, J = 15.6,
3.1 Hz, H-22a), 2.48 (1H, dd, J = 15.6, 10.1 Hz, H-22b), 1.08 (3H, s,
H-28), 1.04 (3H, s, H-18), 1.02 (3H, d, J = 6.7 Hz, H-27), 1.01 (3H, d,
J = 7.0 Hz, H-26), 0.95 (3H, s, H-30), 0.84 (3H, d, J = 6.4 Hz, H-21),
0.62 (1H, d, J = 4.2 Hz, H-19a), 0.41 (1H, d, J = 4.2 Hz, H-19b); 13C
NMR, see Table 4; HR-ESIMS m/z 507.3441 (calcd for C31H48O4Na,
507.3445).
1
1603, 1455, 1375, 1238 cm−1; H NMR (CD3CN, 500 MHz) δ 9.58
(1H, d, J = 1.8 Hz, H-22), 6.95 (1H, d, J = 10.0 Hz, H-1), 5.90 (1H, d,
J = 10.0 Hz, H-2), 5.24 (1H, dt, J = 5.5, 8.2 Hz, H-16), 2.86 (1H, ddq,
J = 11.0, 2.0, 7.3 Hz, H-20), 2.39 (1H, dd, J = 11.0, 8.2 Hz, H-17), 2.16
(1H, m, H-4), 1.89 (3H, s, OAc-16), 1.47 (1H, m, H-7), 1.33 (1H, ddq,
J = 13.4, 5.6, 1.1 Hz, H-15), 1.27 (1H, d, J = 4.3 Hz, H-19a), 1.17 (3H, s,
H-18), 1.09 (3H, d, J = 7.3 Hz, H-21), 1.02 (3H, d, J = 6.7 Hz, H-28),
0.96 (3H, d, J = 0.9 Hz, H-30), 0.55 (1H, d, J = 4.3 Hz, H-19b); 13C NMR,
see Table 3; HR-ESIMS m/z 435.2503 (calcd for C26H36O4Na, 435.2506).
23,24,24a,25,26,27-Hexa-nor-neoboutomelleron-22-oic acid
3β-Hydroxy-29-O-(α-L-rhamnopyranosyl)cycloart-24-en-23-on-
29-oic acid (30): yellow gum; [α]20 −22 (c 0.17, CHCl3); UV
D
(MeOH) λmax (log ε) 238 (4.27), 300 (3.09) nm; IR (film) νmax 3291,
2932, 2867, 1723, 1677, 1607, 1445, 1378, 1311 cm−1; 1H NMR
(methanol-d4, 500 MHz) δ 6.16 (1H, sept, J = 1.2 Hz, H-24), 5.99
(1H, d, J = 1.8 Hz, H-1′), 4.03 (1H, dd, J = 11.3, 4.3 Hz, H-3), 3.86
(1H, dd, J = 3.4, 1.8 Hz, H-2’), 3.69 (1H, dd, J = 9.5, 3.4 Hz, H-3′),
3.61 (1H, dq, J = 9.5, 6.2 Hz, H-5′), 3.44 (1H, t, J = 9.5 Hz, H-4’), 2.52
(1H, dd, J = 15.0, 3.1 Hz, H-22a), 2.11 (3H, d, J = 0.9 Hz, H-27), 1.99
(1H, dd, J = 12.1, 3.8 Hz, H-5), 1.90 (3H, d, J = 0.9 Hz, H-26), 1.22
(3H, d, J = 6.1 Hz, H-6’), 1.13 (3H, s, H-28), 1.04 (3H, s, H-18), 0.94
(3H, s, H-30), 0.86 (3H, d, J = 6.4 Hz, H-21), 0.64 (1H, d, J = 4.1 Hz,
H-19a), 0.44 (1H, d, J = 4.1 Hz, H-19b); 13C NMR, see Table 4; HR-
ESIMS m/z 639.3856 (calcd for C36H56O8Na, 639.3867).
(25): yellow gum; [α]20 −57 (c 0.06, CHCl3); UV (MeOH) λmax
D
(log ε) 208 (3.52), 266 (3.74) nm; IR (film) νmax 3406, 2930, 2875,
1
1725, 1666, 1456, 1376, 1247 cm−1; H NMR (CDCl3, 500 MHz) δ
6.86 (1H, d, J = 10.0 Hz, H-1), 6.01 (1H, d, J = 10.0 Hz, H-2), 5.46
(1H, br q, J = 7.3 Hz, H-16), 2.73 (1H, m, H-20), 2.42 (1H, dd,
J = 10.2, 9.1 Hz, H-17), 2.20 (1H, dq, J = 12.0, 6.9 Hz, H-4), 2.00 (3H,
s, OAc-16), 1.75 (1H, m, H-12), 1.35 (1H, m, H-15), 1.23 (1H, d,
J = 4.5 Hz, H-19a), 1.21 (3H, d, J = 7.0 Hz, H-21), 1.16 (3H, s, H-18),
1.10 (3H, d, J = 6.8 Hz, H-28), 0.96 (3H, s, H-30), 0.55 (1H, d, J = 4.5
Hz, H-19b); 13C NMR, see Table 4; HR-ESIMS m/z 451.2428 (calcd
for C26H36O5Na, 451.2455).
3β-Hydroxy-29-O-(α-L-rhamnopyranosyl)-24-methylcycloart-24-
(24a)-en-23-on-29-oic acid (31): yellow gum; [α]20 −16 (c 0.13,
D
16-Acetyl-3β,26-dihydroxy-24-methyl-25ξ-cycloart-24(24a)-en-
23-on-29-oic acid (26): yellow gum; [α]20D −7 (c 0.04, CHCl3); UV
(MeOH) λmax (log ε) 220 (3.57) nm; IR (film) νmax 3310, 2933, 2871,
CHCl3); UV (MeOH) λmax (log ε) 220 (3.95) nm; IR (film) νmax
3291, 2935, 2865, 1725, 1659, 1462, 1376, 1248, 1147, 1105 cm−1; 1H
NMR (methanol-d4, 500 MHz) δ 6.05 (1H, s, H-24a), 5.99 (1H, d,
J = 1.8 Hz, H-1′), 5.76 (1H, d, J = 1.2 Hz, H-24a), 4.03 (1H, dd,
J = 4.3, 11 Hz, H-3), 3.86 (1H, dd, J = 3.5, 1.8 Hz, H-2’), 3.70 (1H, dd,
J = 9.5, 3.5 Hz, H-3′), 3.61 (1H, dq, J = 9.5, 6.2 Hz, H-5′), 3.45 (1H, t,
J = 9.5 Hz, H-4’), 2.87 (1H, sept, J = 6.7 Hz, H-25), 2.72 (1H, dd,
J = 15.6, 2.7 Hz, H-22a), 2.48 (1H, dd, J = 15.6, 9.8 Hz, H-22b), 1.22
(3H, d, J = 6.2 Hz, H-6’), 1.12 (3H, s, H-28), 1.04 (3H, s, H-18), 1.02
(3H, d, J = 7.0 Hz, H-27), 1.01 (3H, d, J = 7.0 Hz, H-26), 0.95 (3H, s,
H-30), 0.84 (3H, d, J = 6.4 Hz, H-21), 0.64 (1H, d, J = 4.1 Hz, H-19a),
0.44 (1H, d, J = 4.1 Hz, H-19b); 13C NMR, see Table 4; HR-ESIMS
m/z 653.4014 (calcd for C37H58O8Na, 653.4024).
1
1732, 1669, 1555, 1378, 1247, 1028 cm−1; H NMR (CDCl3, 500
MHz) δ 6.03 (1H, s, H-24a), 5.75 (1H, s, H-24a), 5.26 (1H, dt,
J = 4.9, 7.8 Hz, H-16), 4.07 (1H,br d, J = 7.9 Hz, H-3), 3.55 (1H, dd,
J = 10.6, 5.2 Hz, H-26a), 3.49 (1H, dd, J = 10.6, 7.0 Hz, H-26b), 2.95
(1H, sext, J = 6.5 Hz, H-25), 2.52 (3H, m, H-20, 2 H-22), 2.11 (1H,
dd, J = 13.7, 8.2 Hz, H-15), 1.95 (3H, s, OAc-16), 1.89 (1H, dd,
J = 9.9, 8.1 Hz, H-17), 1.80 (1H, m, H-2), 1.17 (3H, s, H-18), 1.13
(3H, br.s, H-28), 1.05 (3H, d, J = 7.0 Hz, H-27), 0.91 (3H, s, H-30),
0.88 (3H, d, J = 5.8 Hz, H-21), 0.62 (1H, d, J = 4.0 Hz, H-19a), 0.40
(1H, d, J = 4.0 Hz, H-19b); 13C NMR, see Table 4; HRESIMS m/z
581.3443 (calcd for C33H50O7Na, 581.3449).
16,22-Diacetyl-2,26-dihydroxy-29-nor-24-methyl-19(9→1)-abeo-
cycloart-9(11),24(24a)-dien-3,23-dione (32): yellow gum; [α]20D +77
(c 0.13, CHCl3); UV (MeOH) λmax (log ε) 210 (3.97), 268 (3.17) nm;
IR (film) νmax 3394, 2926, 2854, 1734, 1694, 1566, 1376, 1229, 1021 cm−1;
1H NMR (CDCl3, 500 MHz) δ 6.12 (1H, s, H-24a), 5.87 (1H, d,
J = 0.6 Hz, H-24a), 5.56 (1H, d, J = 2.1 Hz, H-22), 5.20 (1H, d, J = 6.1
Hz, H-11), 5.09 (1H, dt, J = 4.4, 7.6 Hz, H-16), 4.16 (1H, d, J = 2.1
Hz, H-2), 3.60 (3H, m, OH-2, 2 H-26), 2.88 (1H, sext, J = 6.5 Hz, H-
25), 2.56 (1H, m, H-20), 2.36 (1H, dd, J = 14.0, 7.9 Hz,
H-15a), 2.27 (1H, dd, J = 11.1, 7.5 Hz, H-17), 2.15 (3H, s, OAc-
22), 2.07 (3H, s, OAc-16), 1.88 (1H, dd, J = 9.9, 4.7 Hz, H-1), 1.37
(1H, dd, J = 14.2, 4.1 Hz, H-15b), 1.11 (3H, d, J = 7.3 Hz, H-27), 1.05
(3H, d, J = 6.4 Hz, H-28), 0.94 (1H, bt, J = 4.7 Hz, H-19a), 0.88 (3H,
d, J = 7.0 Hz, H-21), 0.81 (3H, s, H-30), 0.79 (3H, s, H-18), 0.41 (1H,
dd, J = 9.9, 4.7 Hz, H-19b); 13C NMR, see Table 4; HR-ESIMS m/z
607.3245 (calcd for C34H48O8Na, 607.3241).
16-Acetyl-3β,22β-dihydroxy-24-methylcycloart-24(24a)-en-23-
on-29-oic acid (27): yellow gum; [α]20 0 (c 0.03, CHCl3); UV
D
(MeOH) λmax (log ε) 216 (3.58) nm; IR (film) νmax 3302, 2929, 2864,
1732, 1671, 1542, 1454, 1385 cm−1; 1H NMR (methanol-d4, 500
MHz) δ 5.99 (1H, s, H-24a), 5.89 (1H, d, J = 1.2 Hz, H-24a), 5.27
(1H, dt, J = 4.0, 7.3 Hz, H-16), 4.71 (1H, d, J = 1.5 Hz, H-22), 4.03
(1H, dd, J = 11.3, 4.6 Hz, H-3), 2.87 (1H, dsept, J = 1, 6.7 Hz, H-25),
2.24 (1H, dd, J = 14.0, 7.9 Hz, H-15a), 2.06 (3H, s, OAc-16), 1.94 (1H,
dd, J = 12.2, 3.4 Hz, H-5), 1.72 (1H, m, H-2), 1.19 (3H, s, H-18), 1.11
(3H, d, J = 7.0 Hz, H-27), 1.07 (3H, s, H-28), 1.03 (3H, d, J = 7.0 Hz,
H-26), 0.99 (3H, s, H-30), 0.69 (3H, d, J = 6.1 Hz, H-21), 0.63 (1H, d,
J = 4.1 Hz, H-19a), 0.42 (1H, d, J = 4.1 Hz, H-19b); 13C NMR, see
Table 4; HR-ESIMS m/z 581.3455 (calcd for C33H50O7Na, 581.3449).
3β,16β,22β-Trihydroxy-24-methyl-(16,23:23,26)-diepoxycycloart-
24(24a)-en-29-oic acid (28): yellow gum; [α]20 −38 (c 0.05,
D
CHCl3); UV (MeOH) λmax (log ε) 208 (3.41), 270 (2.74) nm; IR
(film) νmax 3300, 2929, 2864, 1650, 1541, 1385 cm−1; 1H NMR
(methanol-d4, 500 MHz) δ 5.24 (1H, d, J = 3.1 Hz, H-24a), 4.92 (1H,
d, J = 3.1 Hz, H-24a), 4.27 (1H, dt, J = 8.6, 7.1 Hz, H-16), 4.03 (1H,
dd, J = 11.3, 4.6 Hz, H-3), 3.94 (1H, t, J = 7.8 Hz, H-27a), 3.30 (1H, t,
J = 7.8 Hz, H-27b), 2.75 (1H, m, H-25), 2.07 (1H, ddd, J = 15.3, 10.5,
5.3 Hz, H-7a), 1.95 (1H, dd, J = 12.4, 4.1 Hz, H-5), 1.90 (1H, m,
H-20), 1.84 (1H, dd, J = 12.8, 7.9 Hz, H-15a), 1.78 (1H, dd, J = 13.7,
2.7 Hz, H-22a), 1.74 (1H, m, H-2a), 1.49 (1H, t, J = 13.7 Hz, H-22b),
1.41 (1H, br dd, J = 12.5, 6.7 Hz, H-15b), 1.18 (3H, s, H-18), 1.08
(3H, s, H-28), 1.07 (3H, d, J = 6.8 Hz, H-26), 0.93 (3H, d, J = 7.0 Hz,
H-21), 0.92 (3H, s, H-30), 0.68 (1H, d, J = 4.0 Hz, H-19a), 0.41
(1H, d, J = 4.0 Hz, H-19b); 13C NMR, see Table 4; HR-ESIMS m/z
521.3230 (calcd for C31H46O5Na, 521.3237).
Proteasome Assay. Experimental procedures for the assay are
detailed in ref 3.
ASSOCIATED CONTENT
* Supporting Information
■
S
This material is available free of charge via the Internet at
http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Author
*Tel: +33-5-34506517. E-mail: georges.massiot.externe@
pierre-fabre.com.
■
3β-Hydroxy-24-methylcycloart-24(24a)-en-23-on-29-oic acid
(29): yellow gum; [α]20 −34 (c 0.06, CHCl3); UV (MeOH) λmax
D
ACKNOWLEDGMENTS
The authors are grateful to R. Belle
collection, to P. Arimondo, C. Bailly, and N. Guilbaud for their
■
(log ε) 220 (3.78) nm; IR (film) νmax 3380, 2930, 2869, 1678, 1552,
1406, 1374 cm−1; 1H NMR (methanol-d4, 500 MHz) δ 6.05 (1H, s, H-
24a), 5.75 (1H, d, J = 0.9 Hz, H-24a), 4.02 (1H, dd, J = 11.4, 4.4 Hz,
́
for historical plant
46
dx.doi.org/10.1021/np200441h | J. Nat. Prod. 2012, 75, 34−47

Journal of Natural Products
Article
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V. G.; Nageswara Rao, K.; Radhika, P. Phytochemistry 1999, 50, 1229−
1236.
interest and attentive support during the research program, to
A. Samson and P. Vergnes for driving the screening process, to
C. Gau, C. Menendez, N. Molinier, and M. J. Serrano for their
highly skilled contribution to the isolation of the com-
pounds, and to N. Chansard for pharmacological evaluation.
(19) Beck, J.; Guminski, Y.; Long, C.; Marcourt, L.; Derguini, F.;
́
Plisson, F.; Grondin, A.; Vandenberghe, I.; Vispe, S.; Brel, V.;
Aussagues, Y.; Ausseil, F.; Arimondo, P. B.; Massiot, G.; Sautel, F.;
Cantagrel, F. Bioorg. Med. Chem., in press.
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