Cytotoxic acylphenols from Myristica maingayi

Article abstract of DOI:10.1016/S0040-4020(00)00074-0

From the cytotoxic AcOEt extract of the fruits of Myristica maingayi, a Myristicaceae, five new acylphenols (promalabaricones B and C, maingayic acids B and C, and maingayone) were isolated, together with the known malabaricones A-C. The structures were determined from spectral analysis, including mass spectrometry and 2D NMR. The cytotoxicity of the new compounds and that of malabaricones was assessed against KB cells. A biosynthetic pathway for malabaricones, via the corresponding promalabaricones as precursors, is suggested. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4020(00)00074-0

TETRAHEDRON
Pergamon
Tetrahedron 56 (2000) 1707–1713
Cytotoxic Acylphenols from Myristica maingayi
a
a
b
a,*
Van Cuong Pham, Akino Jossang, Thierry S e´ venet and Bernard Bodo
a
Laboratoire de Chimie des Substances Naturelles-ESA 8041 CNRS—IFR 63 CNRS, Mus e´ um National d’Histoire Naturelle,
3, rue Buffon 75005 Paris, France
6
b
Institut de Chimie des Substances Naturelles, GDR 1206 CNRS-91198 Gif sur Yvette Cedex, France
Received 3 December 1999; accepted 28 January 2000
Abstract—From the cytotoxic AcOEt extract of the fruits of Myristica maingayi, a Myristicaceae, five new acylphenols (promalabaricones
B and C, maingayic acids B and C, and maingayone) were isolated, together with the known malabaricones A–C. The structures were
determined from spectral analysis, including mass spectrometry and 2D NMR. The cytotoxicity of the new compounds and that of
malabaricones was assessed against KB cells. A biosynthetic pathway for malabaricones, via the corresponding promalabaricones as
precursors, is suggested. ᭧ 2000 Elsevier Science Ltd. All rights reserved.
Introduction
Results and Discussion
Myristica maingayi is a tropical tree of the nutmeg family
The EtOAc extract of the dried fruits of M. maingayi was
chromatographed on silica gel to yield bioactive fractions
from which were separated, by a combination of chromato-
graphic procedures, five new compounds, 1–4 and 8, along
with malabaricones A–C (5–7).
(
Myristicaceae) occurring in Malaysia and Vietnam. The
family is known to produce lignans and acylphenols mainly
of the 2,6-dihydroxyphenyl type, such as malabaricones
A–C from M. malabarica and M. fragans. This aromatic
substitution type is less common than the 3,5-dihydroxy-
phenyl type which is widespread in Anacardiaceae,
1
–6
7
,8
Gingkoaceae and Proteaceae. In addition to their anti-
microbial properties, malabaricones have been recently
9
shown to have nematocide actitivity.
We have previously reported the isolation from the
Myristicaceae species Knema furfuracea, of cytotoxic and
antimicrobial acylphenols, knerachelins A and B, which
5
Promalabaricone B (1) was obtained as a microcrystalline
solid, without optical activity. Its HR FAB mass spectrum
are diarylpentanoids. In continuation of our search for cyto-
ϩ
5
,10
showed the [MϩH] ion at m/z 361.2024 leading to the
toxic compounds from Myristicaceae,
we have investi-
molecular formula C H O . The IR spectrum had strong
21
28
5
gated M. maingayi and observed for the AcOEt extract of the
absorptions for hydroxyls at 3418, methylenes at 2922 and
fruits a cytotoxic activity on human nasopharynx carcinoma
Ϫ1
Ϫ1
2852 and carbonyls at 1722 and 1640 cm .
KB cells (IC 23 mg ml ) and murine leukemia P 388 cells
(
5
0
Ϫ1
100% inhibition at 10 mg ml ).
1
1
1
1
13
The H NMR (Table 1), H– H and H– C COSY spectra
CD OD) depicted characteristic spin systems for a 1,4-
(
3
We now describe the bioguided isolation from M. maingayi
and the structure of two new 2-acylcyclohexane-1,3-diones,
named promalabaricones B (1) and C (2), of two structurally
related arylnonanoic acids, maingayic acids (3, 4) and of a
diarylnonanoid dimer termed maingayone (8), along with
known malabaricones A (5), B (6) and C (7). The bio-
activities are discussed and a biosynthetic relationship
between the isolated compounds is suggested.
disubstituted benzene ring and for ten methylenes, eight of
them forming together an octyl linear chain [–(CH ) –]:
2
8
four methylenes giving signals between dH 1.50 and
.00 ppm and four methylenes at dH 1.29 and 1.32 in a
3
broad singlet. A methinoxy proton at dH 4.27, vicinal to
the two remaining methylenes, formed with them a sym-
0
0
metrical AA BB X spin system (–CH –CH(OH)– CH –);
2
2
the chemical shift of these latter methylenes at d 2.63 and
H
2
.87 suggested they were vicinal to a carbonyl (Table 1).
Keywords: cytotoxicity; diarylnonanoids; promalabaricones B and C; main-
gayic acids B and C; maingayone; Myristicaceae.
13
The J-modulated C NMR spectrum displayed only 15
signals, which suggested symmetrical moieties in the
molecule. In addition to the p-disubstituted benzene ring
*
Corresponding author. Tel.: ϩ33(0)140793129; fax: ϩ33(0)140793135;
e-mail: bodo@mnhn.fr
0
040–4020/00/$ - see front matter ᭧ 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(00)00074-0

1
708
V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
1
3
1
1
Table 1. C and H NMR data for promalabaricones B (1) and C (2) ( H:
with the cis (3.6 Hz) and trans (7.6 Hz) coupling constant
values determined for the methinoxy H-19. Promalabari-
cone B (1) is thus 3,5-dihydroxy-2-[9-(4-hydroxyphenyl)-
nonanoyl]-cyclohexen-1-one, according to the official
nomenclature.
1
3
4
00.13 MHz, C: 100.61 MHz CD
the official rule and is used here for simplicity)
3
OD; carbon numbering does not follow
16
1
2
C number
d
C
d
H
m (J (Hz))
d
C
d
H
m (J (Hz))
Promalabaricone C (2) was also obtained as a microcrystal-
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
206.3
41.5 2.96 t (7.5)
–
206.3
41.5
25.9
30.4
30.4
30.4
30.2
32.9
36.2
135.7
116.5
145.8
144.0
116.2
120.6
114.0
197.0
–
2.97 t (7.6)
line solid from MeOH, without optical activity. Its HR
ϩ
25.8 1.60
30.4 1.32
30.4 1.32
30.4 1.32
30.2 1.29
33.0 1.55
36.0 2.49 t (7.6)
134.9
130.2 6.97 m (8.5)
116.0 6.67 m (8.5)
156.2
116.0 6.67 m (8.5)
130.2 6.97 m (8.5)
114.1
197.1
m
m
m
m
m
m
1.60
1.33
1.33
1.33
1.30
1.55
2.47 t (7.7)
–
6.60 d (2.0)
m
m
m
m
m
m
CIMS showed the protonated molecular ion [MϩH] at
m/z 377, corresponding to the molecular formula
C H O , which differed from that of 1 only by an oxygen
21
28
6
1
13
atom more. The H and C NMR spectra were similar to
those of 1, showing the same substructures, especially the
n-octyl chain and the 1-acyl-4-hydroxycyclohexane-2,6-
dione ring, but differed in the aromatic part because a
0
1
2
3
4
5
6
7
8
3
,4-dihydroxyphenyl ring was observed instead of the
–
–
4-hydroxyphenyl one, as established from the observed
1
6.65 d (8.0)
6.47 dd (8.0, 2.0)
aromatic spin system. The final structure and the H and
13
C chemical shift assignments resulted from the HMBC
–
–
spectrum correlations (Table 1). Thus, promalabaricone
C (2) is 3,5-dihydroxy-2-[9-(3,4,-dihydroxyphenyl)non-
anoyl]-cyclohexen-1-one.
47.6 2.63 dd (16.6, 7.6) 47.8
.87 dd (16.6, 3.6)
64.1 4.27
47.6 2.63 dd (16.6, 7.6) 47.8
.87 dd 16.6, 3.6
2.65 dd (16.5, 7.7)
2.85 dd (16.5, 3.8)
4.28 m
2.65 dd (16.5, 7.7)
2.85 dd (16.5, 3.8)
2
1
2
9
0
m
(64.1)
2
2
1
197.1
197.0
–
and the n-octyl chain, were depicted signals for CH at d
2
C
C
C
4
1
1
7.6, one CHOH at d 64.1, one quaternary carbon at d
C
14.1, one CO at d 206.3 and two keto–enol carbons at d
C
97.1. In the HMBC spectrum, the carbonyl at d 206.3 was
C
correlated to the methylene at d 2.96, thus linking the octyl
H
C
3
protons at dH 6.67 vicinal to the phenolic hydroxyl and,
on the other hand, to the methylenes at 2.49 and 1.55,
thus ending the n-octyl chain. The carbons at dC 197.1
were correlated to the methinoxy proton at d 4.27 and to
The IR spectra of 3 and 4 showed strong absorptions at
H
Ϫ1
the methylenes displaying non-equivalent protons at
1705 cm , suggesting they were carboxylic acids, in agree-
13
2
–
.63 and 2.87, leading to the symmetrical substructure
ment with their C NMR spectra which displayed carbonyl
groups at d 177.8 and 178.1, respectively. The HR CIMS
indicated the molecular formulae C15 for 3 and
C H O for 4: the two compounds differed from each
15 22 4
CO–CH –CH(OH)–CH –CO–. Finally, the quaternary
C
2
2
carbon atom at d 114.1 was correlated to both the methyl-
H O
22 3
C
ene at dH 2.96 and those at dH 2.63 and 2.87: it was thus
linked to the three carbonyls, leading for compound 1 to a
1
other by an oxygen atom. Their H NMR spectra were simi-
lar in the aliphatic proton region, only showing spin systems
2
-acylcyclohexane-1,3-dione structure. However, this
11
cyclic b-triketonic structure is, as usual, not observed
and the presence of the quaternary carbon at dC 114.1
instead of a methine showed that the enolic tautomer (1)
characteristic of an Ar–(CH ) –CO– substructure, but
2 8
differed in the aromatic region: the NMR spectra indicated
they have a 4-hydroxyphenyl ring and a 3,4-dihydroxy-
phenyl ring, respectively. The substitution of the aromatic
rings and the final structures were confirmed by HMBC
experiments and mass spectral fragmentations.
1
was predominant, inasmuch as the H NMR spectrum
recorded in DMSO-d showed a hydroxyl signal at dH
6
18.10, indicating a very strong intramolecular hydrogen
bonding. HMBC cross peaks of carbon at d 197.1 indicated
C
0
that enolization occurred into the ring, with formation of
symmetrical enols. Broadening of this signal evidenced
tautomeric equilibrium for the carbonyls in the ring.
On these bases, the structures of 9-(4 -hydroxyphenyl)-
0
0
nonanoic acid and 9-(3 ,4 -dihydroxyphenyl)-nonanoic
acid were assigned to 3 and 4, respectively, which were
named maingayic acids B and C.
As a result, signals for CO at 17 and 21 were identical, as
well as signals for the methylenes at 18 and 20, producing
symmetry of the cyclohexanedione ring.
Compounds 5, 6 and 7 were identified from their mass
spectral and NMR characteristics as malabaricones A, B
1,9
and C, respectively.
Carbon atoms at 1, 16, 17 and 21 of structure 1, are coplanar,
leading for carbons at 18, 19 and 20, to an average confor-
mation in which the substituents are eclipsed, in agreement
Confirmation of the structures of 1 and 2 arose from their
complete transformation by heating in pyridine at 110ЊC for

V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
1709
Scheme 1.
1
13
Table 2. NMR data for maingayone (8) ( H: 300.13 MHz, C: 75.47 MHz; CD
3
OD); a, b, c may be reversed within the same line
8
(substructure I)
8 (substructure II)
C number
d
C
d
H
m (J (Hz))
C number
d
C
d
H
m (J (Hz))
0
1
2
3
4
5
6
7
8
9
210.0
45.6
25.6
30.1
30.1
30.3
30.3
32.6
36.0
135.8
116.4
145.5
143.5
116.1
120.6
110.8
160.2
107.8
137.4
123.3
160.4
–
1
2
3
4
5
6
7
8
34.7
32.1
29.3
30.1
30.1
30.3
30.3
32.6
36.0
135.8
116.4
145.5
143.5
116.1
120.6
118.0
157.1
108.7
127.9
108.7
157.1
4.54
2.04, 2.41
1.50
1.30
1.30
dd (9.1, 6.8)
m
m
m
m
m
m
m
0
3.08
1.50
1.30
1.30
1.30
1.30
1.50a
2.42
–
t (7.2)
m
m
m
m
m
m
t (7.0)
0
0
0
0
1.30
1.30
0
0
1.50
2.40
0
a
9
t (7.0)
0
10
11
12
13
14
15
16
17
18
19
20
21
10
11
12
13
14
15
16
17
18
19
20
21
–
6.59
–
0
6.59
–
–
d (2.3)
d (2.3)
0
0
–
6.63
6.43
b
0
b
6.64c
d (8.0)
dd (8.0, 2.3)
d (8.0)
dd (8.0, 2.3)
0
c
6.45
–
0
–
–
6.28
6.79
6.28
–
0
–
0
6.30
7.57
–
d (8.5)
d (8.5)
d (8.0)
t (8.0)
d (8.0)
0
0
0
–
3
0 min, into malabaricones B and C. The 3,5-dihydroxy-
were observed between 2.07 and 2.23 ppm, and confirmed
cyclohexen-1-one ring underwent a dehydration and subse-
quent aromatisation, due to the enolic tautomerisation to
form the 2,6-dihydroxyphenyl ring of malabaricones
by its HR (ϩ)FAB MS indicating the molecular formula
1
1
1
13
1
C H O . The H NMR spectrum, H– H and C– H
58
68 17
COSY of 8 depicted four substituted benzene rings: two
0
(Scheme 1).
1,3,4-trisubstituted (a and a ), one 1,2,3,4-tetrasubstituted
(
0
b) and one 1,2,3-trisubstituted (b ) showing two ortho
Maigayone (8) was obtained as a colourless, not crystallized
protons at dH 6.30 and 7.57 (Jˆ8.5 Hz); in the aliphatic
region, signals for 16 methylenes were depicted (Table 2).
The NMR spectra were similar to those of malabaricone C,
with three main differences: (i) a tetrasubstituted benzene
powder, with a weak optical activity ([a] ˆϩ3Њ, MeOH).
D
Its IR spectrum indicated absorption bands of a conjugated
Ϫ1
carbonyl at 1702 cm together with hydroxyl bands at
Ϫ1
ϩ
0
3
385 cm and its FAB MS showed the [MϩH] ion at
ring (b ) instead of a trisubstituted one; (ii) only one
m/z 701 in agreement with the molecular formula
carbonyl at 210.0 ppm; and (iii) a methine signal at
4.54 ppm, strongly coupled with a non-equivalent methyl-
ene at 2.04 and 2.41 ppm and ending one polymethylene
chain (Fig. 1).
C H O . By acetylation with Ac O/Pyr, compound 8
42
52
9
2
gave an octa-acetyl derivative, as shown from its NMR
spectrum where eight acetyl groups forming sharp singlets
Figure 1. HMBC correlations of 8.

1
710
V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
In the HMBC spectrum, carbons at d 135.8 ppm (C-10 and
derivatives (tyrosine or DOPA) by six acetate (malonate)
units, followed by reduction of the first three acetate units
and cyclisation of the last three acetate units into a triketonic
cyclohexane ring according to the phloroglucinol type
cyclisation; then the reduction of the para-carbonyl group
into an alcohol yields promalabaricones and further dehy-
dration of the ring hydroxyl leads to malabaricones (Fig 2).
This suggested mixed biosynthesis is similar to that now
well established for flavonoids, for which the cinnamoyl
moiety is elongated by only three acetate units, followed
by cyclisation. Acids 3 and 4 could result either from the
cleavage of promalabaricones at the triketonic methine
level, or from an unachieved promalabaricone biosynthesis.
C
0
0
1
2
9
0 on a and a rings) were correlated with protons at d
H
0
.40 and 2.42 (9 and 9 ) and carbons at 36.0 ppm (C-9 and
0
0
) with aromatic protons at 6.59, 6.43 and 6.45 ppm (a and
a -ring): two n-alkyl-dihydroxyphenyl groups were thus
defined, the aromatic a and a -rings ending two aliphatic
0
chains. The carbonyl at 210.0 ppm (C-1) was correlated
with the methylene at 3.08 ppm and with the aromatic
proton of b-ring at dH 6.30 (H-18), defining thus the
substructure I.
0
The methine proton at 4.54 ppm (H-1 ) was correlated in the
0
HMBC spectrum with the aromatic carbons at 118.0 (C-16 )
0
0
0
and 157.1 ppm (C17 and 21 ) of b -ring, allowing to define
substructure II (Table 2). It was also correlated with carbons
at 123.3 (C-20), 137.4 (C-19) and 160.4 (C-21) of b-ring,
indicating a diarylmethane structure and allowing the link-
age of substructures I and II as indicated in structure 8. This
was confirmed by the long range correlation between proton
When tested on human tumoral KB cells, all the compounds
showed significant cytotoxicity, malabaricone B being the
Ϫ1
more active with an IC50 of 3 mg ml (Table 3). Malabari-
cones were more toxic than the related promalabaricones,
and p-hydroxyphenyl compounds were generally more toxic
than the o,p-dihydroxyphenyl ones. These compounds were
responsible for the toxicity of the plant extract. Tubulin
could be the target for cytotoxicity of the active components
0
at 7.57 ppm (H-19) and the methine carbon 1 . The resulting
dimeric structure 8 for maingayone was in agreement with
mass spectral fragmentations. Compound 8 appears as a
dimeric product of malabaricone C (2) which could result
from the aldol condensation of one molecule 2 with another
molecule of malabaricone C, followed by the reduction of
the aldol hydroxyl.
Ϫ1
as an IC50 value of 10 mg ml was observed for the crude
extract. The mechanism of action of the Myristica phenols
may thus differ from that described for the 3,5-dihydroxy-
acylphenols which cleave DNA or inhibit DNA polymerase
8,12,13
b.
The biosynthesis of promalabaricones presumably results from
the elongation of a cinnamoyl type precursor, originating
from an amino acid such as phenyl alanine and its hydroxy-
All the isolated compounds had moderate or no activity
against Plasmodium falciparum, only malabaricone B
Figure 2. Hypothetical pathway for promalabaricones and malabaricones biosynthesis.

V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
1711
Table 3. Biological activity of acylphenols from M. maingayi: cytotoxicity
against KB cells, and inhibition of Plasmodium falciparum growth
malabaricones B (6, 1.76 g) and C (7, 11.42 g), respectively.
Fractions F18–F21 were combined (6.01 g) and further
chromatographed on silica gel with a AcOEt/MeOH
gradient starting from 9/1 to pure MeOH and 12 fractions
(f1–f12) were collected: fraction f9 yielded promalabari-
cone B (1, 70 mg) and fractions f4–f6 were combined
Compound
KB cells
IC50
mg ml
Plasmodium
falciparum
IC50 mg ml
(mM)
Ϫ1
Ϫ1
(
mM)
(
0.42 g) and submitted to repetitive column chromatography
Crude extract
23
–
to yield maingayic acid B (3, 42 mg).
Promalabaricone B (1)
Promalabaricone C (2)
Maingayic acid B (3)
Maingayic acid C (4)
Malabaricone A (5)
Malabaricone B (6)
Malabaricone C (7)
Maingayone (8)
6 (16)
9 (24)
30 (120)
8 (30)
50 (153)
3 (9)
Ͼ100 (Ͼ277)
16 (43)
45 (180)
56 (210)
32 (98)
Ͼ100 (Ͼ292)
20 (56)
Ͼ100 (Ͼ143)
Fraction F22 was chomatographed on silica gel with an
AcOEt/MeOH gradient starting from 9/1 to pure MeOH:
1
8 fractions (f1–f18) were collected. Fraction f11 yielded
4 (11)
18 (26)
maingayic acid C (4, 20 mg) and fraction f17 yielded
promalabaricone C (2, 120 mg). Fractions f3–5 (0.70 g)
were combined and chromatographed on a reversed phase
(
RP8) silica gel column eluted with a MeOH/H O gradient
2
showing a significative growth inhibition with an IC value
of 16 mg ml (Table 3).
from 1/1 to pure MeOH and yielded maingayone (8,
250 mg).
5
0
Ϫ1
Determination of cytotoxic activity
Experimental
General experimental procedures
The cytotoxicity assays were carried out in 96-well
microtiter plates in triplicate against human nasopharynx
3
Ϫ1
carcinoma KB cell lines ꢀ3 × 10 cells ml † using a modi-
Melting points were determined on a Reichert microscope
apparatus and are uncorrected. Optical rotations were
measured on a Perkin–Elmer 141 polarimeter at room
temperature. IR spectra were recorded in KBr disks on a
Nicolet Impact 400D spectrophotometer, and UV spectra
with an Uvikon 930 Kontron spectrophotometer. Mass
spectra were obtained either on a Nermag R 10-10 in the
CI or EI mode or on a ZAB2-SEQ (VG analytical) for the
FAB MS.
14
fication of the published method. KB cells were maintained
in Dulbecco’s D-MEM medium, supplemented with 10%
Ϫ1
fetal calf serum, NaHCO (3.7 g l ), l -glutamine (2 mM),
3
Ϫ1
Ϫ1
penicillin G (100 units ml ), streptomycin (100 mg ml )
and gentamycin (10 mg ml ). After 72 h incubation at 37ЊC
Ϫ1
in air/CO (95/5) with or without test compounds, cell
2
growth was estimated by colorimetric measurement of
stained living cells by neutral red. Optical density was deter-
mined at 540 nm on a Titertek Multiscan photometer. The
IC value was defined as the concentration of sample neces-
1
13
50
H and C spectra were recorded on a Bruker AC 300
sary to inhibit the cell growth to 50% of the control.
spectrometer operating at 300.13 and 75.43 MHz, respec-
1
tively, or on an Avance 400 Bruker (400.13: H and
1
3
1
1
00.61 MHz: C). H chemical shifts were referenced
Determination of antimalarial activity
relative to CHD OD at 3.31 ppm or CHCl at 7.24 ppm
2
3
13
and C chemical shifts to central peak of CD OD at 49.0
3
The assays on Plasmodium falciparum were performed
on the chloroquine resistant-strain FcB1/Colombia.
P. falciparum was maintained in vitro on human type O
or CDCl at 77.0 ppm. For the HMBC spectra, the delay was
3
ϩ
optimized for a coupling constant of 7 Hz.
erythrocytes in RPMI 1640 culture medium supplemented
Plant material
with 27.5 mM NaHCO , 25 mM HEPES, 11 mM glucose
3
ϩ
and 7.5% heat treated human O serum in an atmosphere
The fruits of Myristica maingayi Hk, f. were collected at
Gua Musang (Malaysia) in August 1997. A voucher
specimen (no. KL 4525) is on deposit in the Herbarium of
the Department of Chemistry, University of Malaysia
of 3% CO , 6% O , 91% N at 37ЊC and pH 7.4. The experi-
2
2
2
ments were carried out in 96-well plates according to the
15
semi-automated microdilution technique. Stock solutions
of acylphenols were prepared in DMSO. Different concen-
trations were tested on parasite cultures (2% parasitaemia,
(Kuala-Lumpur).
2
% haematocrite) for 24 h, at 37ЊC prior to the addition of
3
Extraction and isolation
0.5 mCi per well of [ H]hypoxanthine. DMSO concen-
tration never exceeded 0.05%, a concentration which
has no effect upon parasite growth. After further incubation
for 24 h, the cells were harvested onto filter paper. The
dried disks were counted with a scintillation spectrometer.
The growth inhibition was determined by comparison of
the incorporated radioactivity of treated culture to control
culture without drug. The concentrations causing 50%
of growth inhibition (IC ) were obtained from the drug
The powder of dried fruits (1.3 kg) was extracted with
EtOAc at room temperature. The residue (83 g) obtained
after evaporation of the solvent was chomatographed on a
silica gel column with a CH Cl /MeOH gradient starting
2
2
from 95/5 to pure MeOH and yielding 36 fractions
(
F1–F36). Fractions F1–F5 were combined (2.34 g) and
further chromatographed on silica gel with a cyclohexane/
AcOEt gradient starting from 95/5 to pure AcOEt and 28
fractions were collected: fraction 20 yielded malabaricone
A (5, 0.2 g). Fractions F9 and F12 corresponded to pure
50
concentration–response curves and the results expressed
as the mean determined from several independent experi-
ments.

1
712
V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
Promalabaricone B (1). C H O ˆ360. Colorless micro-
lmax (log e): 205 (4.2), 221 (3.9), 272 (3.6), 340 (3.1). IR
(KBr) nmax (cm ): 3262, 2930, 2914, 2854, 1632, 1603,
2
1
28
5
2
2
Ϫ1
crystals mp: 140–142ЊC (MeOH); [a] ˆ0Њ (c 0.5, MeOH).
D
UV (MeOH) l (log e): 203 (4.1), 226 (4.0), 274 (4.1). IR
1510, 1459, 1347, 1242, 1045, 965, 874, 722. CI MS
max
Ϫ1
ϩ
ϩ
(
KBr) nmax (cm ): 3418, 2922, 2852, 1722 (sh), 1640,
(NH ) m/z (%): 344 ([MϩNH ] , 20), 327 ([MϩH] ,
3
4
13
1
582, 1517, 1447, 1406, 1239, 1064, 824, 783, 611, 511.
100), 137 (20), 91 (9). C NMR (75.47 MHz, CD OD):
3
ϩ
CI MS (NH ), m/z (%): 361 ([MϩH] , 100), 343
24.4 (C-3), 29.1 (C-7), 29.3 (C-4), 29.3 (C-5), 29.3 (C-6),
31.4 (C-8), 35.9 (C-9), 44.7 (C-2), 108.2 (C-18), 108.2
(C-20), 110.0 (C-16), 125.5 (C-13), 128.1 (C-12), 128.1
(C-14), 128.3 (C-11), 128.3 (C-15), 135.9 (C-19), 142.9
3
ϩ
(
[MϪH OϩH] , 94), 284 (10), 268 (12), 233 (15). HR
2
ϩ
FAB MS, m/z: 361.2024 ([MϩH] ), calcd C H O ˆ
2
1
29
5
⅐ϩ
3
61.2015. EI MS m/z (%): 360 (M , 5), 342 (23), 232
1
1
(20), 204 (25), 137 (95), 107 (100).
H
NMR
(C-10), 161.3 (C-17), 161.3 (C-21), 208.5 (C-1). H NMR
(
300.13 MHz, DMSO-d ): 1.25 (8H, br.s, CH -4 to 7),
(300.13 MHz, CD OD): 1.33 (8H, br.s, CH -4 to 7), 1.61
6
2
3
2
1
.48 (4H, br.s, CH -3 and 8), 2.42 (2H, t, Jˆ7.5 Hz,
(2H, p, Jˆ7.5 Hz, CH -8), 1.71 (2H, p, Jˆ7.5 Hz, CH -3),
2
2
2
CH -9), 2.93 (2H, t, Jˆ7.4 Hz, CH -2), 2.81 (2H, m)
2.60 (2H, t, Jˆ7.5 Hz, CH -9), 3.15 (2H, t, Jˆ7.5 Hz,
2
2
2
and 2.51 (2H, m) CH -18 and 20), 4.14 (1H, br.s,
CH -2), 6.40 (2H, d, Jˆ8.3 Hz, H-18, 20), 7.17 (3H, m,
2
2
H-19), 6.64 (2H, m, Jˆ8.6 Hz, H-12, 14), 6.94 (2H, m,
Jˆ8.6 Hz, H-11, 15), 9.08 (1H, br.s, OH-13), 18.10 (1H,
br.s, OH-19).
H-11, 13, 15), 7.26 (3H, m, H-12, 14, 19).
Malabaricone B (6). C H O ˆ342. Colorless crystals
21
26
4
1
(MeOH), mp: 133–135ЊC (lit. : mp 81–82ЊC). UV
Promalabaricone C (2). C H O ˆ376. Mp: 162–164ЊC
(MeOH) lmax (log e): 203 (4.1), 224 (4.2), 270 (3.9), 341
(3.3). IR (KBr) nmax (cm ): 3572, 3348, 2921, 2848, 1637,
2
1
28
6
2
2
Ϫ1
(
(
(
MeOH); [a] ˆ0Њ (c 0.5, MeOH). UV (MeOH) l
D
max
max
log e): 204 (4.3), 222 (2.4), 272 (3.9). IR (KBr) n
1588, 1517, 1460, 1394, 1341, 1255, 1222, 1193, 1045, 959,
Ϫ1
ϩ
cm ): 3420, 2930, 2854, 1729, 1637, 1585, 1400, 1262,
830, 808, 782, 722. CI MS (NH ) m/z (%): 360 ([MϩNH ] ,
3
4
ϩ
ϩ
13
1
038, 815. CI MS (NH ), m/z (%): 377 ([MϩH] , 29), 359
10), 343 ([MϩH] , 100), 137 (10), 107 (10). C NMR
3
ϩ
⅐ϩ
[
(
(
MϪH OϩH] , 100). EI MS m/z (%): 376 (M , 5), 358
(75.47 MHz, CD OD): 24.4 (C-3), 29.0 (C-5), 29.0 (C-6),
2
3
20), 248 (25), 221 (21), 149 (100), 137 (85), 123 (70), 110
80).
29.2 (C-7), 29.5 (C-4), 31.6 (C-8), 35.0 (C-9), 44.7 (C-2),
107.8 (C-18), 107.8 (C-20), 110.1 (C-16), 115.0 (C-12),
1
15.0 (C-14), 129.3 (C-11), 129.3 (C-15), 134.6 (C-10),
Maingayic acid B (3). C H O ˆ250. Colorless crystals
135.7 (C-19), 153.6 (C-13), 161.6 (C-17), 161.6 (C-21),
208.6 (C-1), H NMR (300.13 MHz, CD OD): 1.29 (8H,
1
5
22
3
1
(
MeOH), mp: 106–107ЊC. UV (MeOH) lmax (log e): 202
3
Ϫ1
(
3.6), 224 (3.6), 278 (2.9). IR (KBr) nmax (cm ): 3400,
br.s, CH -4 to 7), 1.51 (2H, p, Jˆ7.4 Hz, CH -8), 1.64
2
2
2
1
925, 2856, 1778 (sh.), 1702, 1620, 1602, 1520, 1473,
(2H, p, Jˆ7.4 Hz, CH -3), 2.46 (2H, t, Jˆ7.4 Hz, CH -9),
2
2
251, 1239, 1219, 859, 818, 725. CI MS m/z (%): 268
3.10 (2H, t, Jˆ7.4 Hz, CH -2), 6.32 (2H, d, Jˆ8.2 Hz, H-18,
2
ϩ
⅐ϩ
(
[MϩNH ] , 95), 250 (M , 57), 233 (100), 205 (10), 107
20). HRMS, m/z: 250.1589 ([M] ), calcd C H O ˆ
20), 6.69 (2H, m, Jˆ8.4 Hz, H-12, 14), 6.96 (2H, m,
4
⅐ϩ
(
Jˆ8.4 Hz, H-11, 15), 7.14 (1H, t, Jˆ8.2 Hz, H-19).
1
5
22
3
1
3
2
3
3
50.1569. C NMR (75.47 MHz, CD OD): 26.0 (C-3),
3
0.2 (C-4), 30.2 (C-5), 30.3 (C-6), 30.4 (C-7), 32.9 (C-8),
5.0 (C-2), 36.0 (C-9), 116.0 (C-12), 116.0 (C-14), 130.2
Malabaricone C (7). C H O ˆ358. Colorless crystals
21
26
5
1
(MeOH), mp: 127–128ЊC; (lit. : mp 123–124ЊC). UV
(MeOH) lmax (log e): 206 (4.5), 222 (4.3), 272 (4.1), 341
C-11), 130.2 (C-15), 134.8 (C-10), 156.1 (C-13), 177.8
1
Ϫ1
(3.4). IR (KBr) nmax (cm ): 3467, 3296, 2934, 2855, 1683
3
CH -4 to 7), 1.56 (4H, m, CH -3, 8), 2.26 (2H, t,
( e´ p.), 1637, 1604, 1532, 1460, 1393, 1367, 1255, 1124,
2
2
Jˆ7.5 Hz, CH -2), 2.47 (2H, t, Jˆ7.6 Hz, CH -9), 6.68
1045, 955, 801, 742, 538. CI MS (NH ) m/z (%): 376
2
2
3
ϩ
ϩ
(
2H, m, Jˆ8.3 Hz, H-12, 14), 6.95 (2H, m, Jˆ8.3 Hz,
([MϩNH ] , 12), 359 ([MϩH] , 100), 137 (10), 123 (8).
C NMR (75.47 MHz, CD OD): 24.3 (C-3), 28.9 (C-7),
4
13
H-11, 15).
3
2
9.2 (C-4), 29.2 (C-5), 29.2 (C-6), 31.4 (C-8), 35.0 (C-9),
Maingayic acid C (4). C H O ˆ266. Colorless crystals
44.6 (C-2), 107.5 (C-18), 107.5 (C-20), 110.1 (C-16), 114.9
(C-14), 115.2 (C-11), 120.0 (C-15), 135.3 (C-10), 135.7
(C-19), 141.8 (C-13), 143.8 (C-12), 161.8 (C-17), 161.8
(C-21), 208.7 (C-1).
1
5
22
4
(
MeOH), mp: 133–135ЊC. UV (MeOH) l
(log e): 204
max
Ϫ1
(
4.1), 220 (3.8), 278 (3.5). IR (KBr) nmax (cm ): 3433,
2
8
929, 2856, 1713, 1611, 1529, 1389, 1207, 1117, 871,
ϩ
12. CI MS (NH ), m/z (%): 284 ([MϩNH ] , 85), 266
3
4
⅐ϩ
1
(
M , 50), 249 (100), 233 (10), 205 (5), 151 (5), 123 (15).
HRMS, m/z: 266.1499 ([M] ), calcd C H O ˆ266.1518.
H NMR (300.13 MHz, CD OD): 3.09 (2H, t, Jˆ7.4 Hz,
3
⅐ϩ
CH -2), 1.62 (2H, p, Jˆ7.4 Hz, CH -3), 1.30 (2H, m,
1
5
22
4
2
2
13
C NMR (75.47 MHz, CD OD): 26.2 (C-3), 30.2 (C-4),
CH -4), 1.26 (6H, br.s, CH -5 to 7), 1.49 (2H, p,
2 2
3
3
3
0.2 (C-5), 30.4 (C-6), 30.4 (C-7), 32.9 (C-8), 35.2 (C-2),
Jˆ7.4 Hz, CH -8), 2.39 (2H, t, Jˆ7.4 Hz, CH -9), 6.62
2
2
6.2 (C-9), 116.2 (C-14), 116.5 (C-11), 120.6 (C-15), 135.7
(1H, d, Jˆ2.0 Hz, H-11), 6.67 (1H, d, Jˆ8.0 Hz, H-14),
6.50 (1H, dd, Jˆ8.0, 2.0 Hz, H-15), 6.31 (2H, d,
Jˆ8.1 Hz, H-18, 20), 7.12 (1H, t, Jˆ8.1 Hz, H-19).
1
(
C-10), 144.0 (C-13), 146.0 (C-12), 178.1 (C-1). H NMR
(300.13 MHz, CD OD): 1.30 (8H, br.s, CH -4 to 7), 1.53
3
2
(
2
6
2H, t, Jˆ6.5 Hz, CH -3), 1.58 (2H, p, Jˆ7.1 Hz, CH -8),
2
2
.27 (2H, t, Jˆ7.2 Hz, CH -2), 2.43 (2H, t, Jˆ7.4 Hz, CH -9),
Maingayione (8). C H O ˆ700. Amorphous solid.
2
2
42 52
9
2
2
.46 (1H, dd, Jˆ8.0, 2.0 Hz, H-15), 6.58 (1H, d, Jˆ2.2 Hz,
[a] ˆϩ3Њ (c 0.5, MeOH). UV (MeOH) l
(log e): 206
max
Ϫ1
D
H-11), 6.64 (1H, d, Jˆ8.0 Hz, H-14).
(4.8), 277 (4.2), 354 (3.4). IR (KBr) n
(cm ): 3385,
max
2
929, 2856, 1702, 1611, 1523, 1468, 1441, 1362, 1283,
ϩ
Malabaricone A (5). (C H O )ˆ326. Colorless crystals
1242, 1117, 994, 815, 789. MS FAB m/z (%): 701
([MϩH] , 5), 629 (3), 591 (45), 441 (9) 473 (16), 343
2
1
26
3
1
ϩ
(MeOH), mp: 80–82ЊC (lit. : mp 81–82ЊC). UV (MeOH)

V. C. Pham et al. / Tetrahedron 56 (2000) 1707–1713
1713
(
25), 123 (100). CI MS (NH ), m/z (%): 591 (1), 495 (1), 416
2. Orabi, K. Y.; Mossa, J. S.; El-Feraly, F. S. J. Nat. Prod. 1991,
54, 856–859.
3
(2), 399 (6), 387 (18), 359 (100), 343 (23).
3
. Cooray, N. F.; Jansz, E. R.; Wimalasena, S.; Wijesekera, T. P.;
ϩ
Maingayione octa-acetate. SM FAB m/z (%): 1059
Nair, B. M. Phytochemistry 1987, 26, 3369–3371.
4. Kumar, N. S.; Herath, H. M. T. B.; Karunaratne, V. Phyto-
chemistry 1988, 27, 465–468.
5. Zahir, A.; Jossang, A.; Bodo, B. J. Nat. Prod. 1993, 56, 1634–
1637.
6. Pinto, M. M. M.; Kijjoa, A.; Mondranondra, I. O.; Guti e´ rrez,
A. B.; Herz, W. Phytochemistry 1990, 29, 1985–1988.
7. Kozubek, A. Chem. Rev. 1999, 99, 1–25.
8. Lytollis, W.; Scannell, R. T.; An, H.; Murty, V. S.; Reddy, K. S.;
Barr, J. R.; Hecht, S. M. J. Am. Chem. Soc. 1995, 117, 12683–
ϩ
(
[MϩNa] , 100); HR MS obs. 1059.4275, calcd for
1
C H O Na: 1059.4354. H NMR (300.13 MHz, CDCl ):
58
68 17
3
0
0
1
2
8
.23 (16H, br.s, 8 CH ), 1.55 (8H, m, 4 CH : 3, 8, 3 and 8 ),
2
2
.07 (3H, s), 2.13 (6H, s), 2.20 (3H, s) and 2.23 (12H, s):
0
0
-CO–CH , 2.55 (8H, m, 4 CH : 2, 9, 2 and 9 ), 4.21 (1H, t,
3
2
0
0
0
Jˆ7.5 Hz, H-1 ), 6.88 (2H, d, Jˆ8.0 Hz, H-18 and 20 ),
0
6
.90–7.10 (6H, m, arom. a and a -ring), 7.10 (1H, d,
0
Jˆ8.7 Hz, H-18), 7.20 (1H, t, Jˆ8.0 Hz, H-19 ), 7.50 (1H,
d, Jˆ8.7 Hz, H-19).
1
9
2690.
. Hosoi, S.; Kiuchi, F.; Nakamura, N.; Imasho, M.; Ahad Ali, M.;
Acknowledgements
Sasaki, Y.; Tanaka, E.; Tsumamoto, Y.; Kondo, K.; Tsuda, U.
Chem. Pharm. Bull. 1999, 47, 37–43.
We thank Dr J. P. Brouard for the mass spectra, Dr A. Blond
for the 400 MHz NMR spectra, Dr Ph. Grellier for the
antimalarial bioassays and Dr C. Wiart for collecting the
plant material. The French Minist e` re de la Coop e´ ration is
gratefully acknowledged for a scholarship to one of us
10. Jossang, A.; Jossang, P.; Hadi, H. A.; S e´ venet, T.; Bodo, B.
J. Org. Chem. 1991, 56, 6527–6530.
11. Rubinov, D. B.; Rubinova, I. L.; Akhrem, A. A. Chem. Rev.
1999, 99, 1047–1065.
12. Singh, U. S.; Scannell, R. T.; An, H.; Carter, B. J.; Hecht, S. M.
J. Am. Chem. Soc. 1995, 117, 12691–12699.
(V.C.P.).
1
4
3. Deng, J. Z.; Starck, S. R.; Hecht, S. M. J. Nat. Prod. 1999, 62,
77–480.
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