636 J. Nat. Prod.

2000, 63, 636-642

Phytochemical Investigation of Aglaia rubiginosa

S. Weber, J. Puripattanavong, V. Brecht, and A. W. Frahm*

Department of Pharmaceutical Chemistry, Institute of Pharmacy, University of Freiburg, Hermann-Herder-Strasse 9,
79104 Freiburg, Germany

Received November 24, 1999

The phytochemical investigation of a methanolic leaf extract of Aglaia rubiginosa furnished 15 isoprenoid
constituents, eight of which represented new natural entities. Two androstane derivatives (1 and 2),
previously synthesized, and also obtained by microbiological transformations; an extraordinary 17-octanor-
cycloartane-ring-A-seco acid (3); four cycloartane-type triterpenes (4-7); and three unusual cholesterol
derivatives (8-10) were isolated, along with two known dammaranes (11 and 12), a stigmastandiol (13),
and β-sitosterol and its β-D-glucoside. Spectroscopic structure elucidation of the new natural products
(1-3, 6, 7, 8-10) is described.

With more than 100 species, Aglaia represents the of a five-membered ring ketone (C-17), and at δ 186.1,
largest genus in the family Meliaceae and constitutes an characteristic of a cross-coupled dienone-moiety (C-3), four
important part of the tropical forest in Indochina. The olefinic carbons at δ 169.6 (one quaternary carbon, C-5)
taxonomic species delimitation proved to be troublesome and at δ 155.1, 127.7, and 119.8 (three methine carbons,
and had to be revised several times. Based on the dehis- C-1, C-2, and C-4), together with one oxygenated methine
cence of the fruit and the flower characters, a taxonomic carbon at δ 68.0, identified as C-6 because it showed long-
monograph of the genus Aglaia recognizes two sections range coupling to H-4. The R-position of the C-6 hydroxyl
within this family, the section Aglaia (88 species) and the group was proven by positive NOE effects between
section Amoora (16 species).1 Some species are used in H-6βfH-8 and H-6βfH-19, respectively.
traditional medicine against different diseases, including Compared with that of 1, the 13C NMR spectrum of
cancer,2 heart problems,3 fever,3 inflammation,4 and cough.4 compound 2 showed broad similarities. Only the downfield-
So far only representatives of the section Aglaia have shifted signal at δ 68.0 assigned to C-6 was missing. The
been investigated phytochemically. Various triterpenes HREIMS exhibited a molecular ion peak at m/z 284.1775
(e.g., limonoids,5-7 cycloartanes,2,8,9 and tirucallanes10,11) against m/z 300.1725 for 1, with the former corresponding
and cyclopenta[b]benzofurans (flavaglines12) were isolated. to the supposed elemental formula C19H24O2 and leading
In particular, some of the benzofuran derivatives show to the identification of 2 as androsta-1,4-diene-3,17-dione,16
interesting phamacological properties: antileukemic,13 an- the non-hydroxylated parent compound of 1.
tiviral,14 and insecticidal15 activities have been found. As
part of our studies on the constituents of Aglaia species
from Thailand, we have examined the leaves of A. rubigi-
nosa (Hiern) Pannell, belonging to the section Amoora with
dehiscent fruits, which are dispersed by birds. It seemed
interesting from the phytochemical and taxonomic point
of view to establish whether the above-mentioned classes
of compounds discriminate the section Amoora from the
section Aglaia. This prompted us to direct our search
toward structurally differentiating compounds.

Results and Discussion

Repeated chromatographic separation of the methanolic The androstane derivatives 1 and 2 represent a very
leaf extract from A. rubiginosa resulted in the isolation of unusual type of steroid in plants with the characteristic
eight new steroid and triterpenoid compounds. The struc- ring A-cyclohexadienone structure, well-known from syn-
tures were elucidated by means of spectroscopic methods, thetic corticosteroid compounds. They were obtained previ-
such as EIMS, HREIMS, ESIMS, 1H NMR, 13C NMR, and ously from hyodeoxycholic acid17 by microbiological trans-
2D NMR techniques (gs-COSY, gs-HSQC, gs-HMQC, NOE- formation reactions.16 Their synthesis is described in the
SY). patent literature,18 but they have never been isolated from
The identification of 6R-hydroxyandrosta-1,4-diene-3,17- any plant until now.
dione (1) was carried out spectroscopically and finally A reduced number of 13C NMR signals for 18 carbons
confirmed by comparing the 1H spectral data with litera- excluded a normal triterpenoid structure for compound 3.
ture values16 and with those of a synthetic reference sample The IR spectrum exhibited absorptions in the hydroxyl
(Schering AG, Berlin). The 13C NMR data have been region (3400 cm-1) and for two carbonyl functions (1721
assigned for the first time by means of DEPT and gs- and 1683 cm-1) with a shoulder (1710 cm-1), pointing to
HMQC spectra. Seven indicative low-field signals out of the presence of a third carbonyl group. The 1H NMR
19 were assigned to two carbonyl carbons at δ 219.5, typical spectrum of 3 showed the presence of only two singlets for
methyl groups (δ 0.83, 1.14) and two 1H-doublets at δ 0.96
* To whom correspondence should be addressed. Tel.: +49-761-203-6335. and 1.68 (J ) 5.7 Hz) characteristic of geminal cyclopro-
Fax: +49-761-203-6351. E-mail: [email protected]. pane ring protons. On the basis of 2D NMR results
10.1021/np9905923 CCC: $19.00 © 2000 American Chemical Society and American Society of Pharmacognosy
Published on Web 04/06/2000
Phytochemical Investigation of Aglaia Journal of Natural Products, 2000, Vol. 63, No. 5 637

(1H-1H COSY and NOESY), the signal groups at δ 2.68

and 2.74 (each 1H, d, J ) 16.6 Hz) were assigned to the
C-1 methylene group. The 13C NMR spectrum gave evi-
dence of three carbonyl signals, at δ 218.8 (five-membered
ring ketone), 209.7 (six-membered ring ketone), and 176.5
(carboxylic acid). The carboxylic acid structural element
was confirmed by the 1H NMR spectrum in DMSO-d6 with
a singlet at δ 11.8 integrating for one exchangeable proton.
The HREIMS showed the molecular ion peak at m/z
304.1685 and confirmed the proposed elemental formula
C18H24O4. These data allowed us to identify the skeleton
of 3 as a 17-octanor-cycloartane-ring-A-seco acid. To the
best of our knowledge this is the first report of a seco-
compound in the genus Aglaia. So far only very few
structurally related ring A seco-cycloartanes have been
isolated from other natural sources,19,20 with structures
substantially different from that of 3.
The number of publications on cycloartane-type triter-
penes in Aglaia species clearly indicates that this class of
compounds may be regarded as a distinctive chemotaxo-
nomic feature. Cycloartane-type triterpenes possess a
cyclopropane bridge between C-9 and C-10, and protons
attached to cyclopropyl rings characteristically appear as
a pair of doublets in the high-field 1H NMR region with a
geminal coupling constant (J ) 4.3 Hz). From the leaves
of A. rubiginosa we have isolated four such cycloartane
derivatives (4-7), which have not been isolated previously
from any Aglaia species. Two of them (6 and 7) proved to
be new compounds.
Based on its spectral characteristics and comparison with
published data,21 compound 4 was identified as cycloart-
23-ene-3β,25-diol, disregarding the (E)- or (Z)-configuration
of the double bond at C-23. When the 1H NMR spectrum
was recorded in benzene-d6, the narrow multiplet at δ 5.60
ppm observed in the CDCl3 spectrum was separated into
an AB-pattern with ∆δ ) 13.2 Hz and 3J(23/24-H) ) 16 Hz,
from which the (E)-configuration was unequivocally de-
duced. This contradicts the literature data for cycloart-
(23Z)-ene-3β,25-diol.21
In the 1H NMR spectrum of 5, when compared to that of
4, one of the three methyl singlets below 1 ppm was
missing, whereas a pair of doublets (1H each) appeared
downfield shifted to δ 3.50 and 3.75, respectively, the latter
overlapping with the H-3 multiplet. However, the coupling double bond (C-23 and C-24, δ 5.59, 2H, δ 125.6 and 139.3).
constant (J ) 10.5 Hz) confirmed a geminal relationship, The only structural feature capable of deshielding such
and the gs-HMQC spectrum indicated that the hydroxy- geminal methyl groups is an oxygen-bearing carbon atom.
methyl substituent is attached to C-4 either in the β- (C- Unfortunately, cycloart-23-ene-3β,25,28-triol (6) decom-
29) or in the R-position (C-28). A decision in favor of C-28 posed during the NMR experiments in CDCl3. The precise
followed from the observation of the high-field chemical interpretation of these corresponding spectra proved that
shift of C-29 from δ 14.0 in 4 to 10.1 in 5. From a detailed H2O had been eliminated from the side chain. The reaction
comparison of all spectral data with literature values,22 the might have been catalyzed by traces of hydrochloric acid
structure of 5 was deduced as cycloart-24-ene-3β,28-diol. from the CDCl3 solvent. Compound 6 has not yet been
The NMR experiments indicated that 6 has the same reisolated from the plant. The NMR data of 6 and of the
basic skeleton as 5, with hydroxyl groups at C-3 and C-28, resulting artifact 6a is given (see Table 1). These findings
but with a structurally different side chain at position 17. have been proven via compound 4, with an identical side
The side-chain moiety consisted of an OH-substituted chain. After dissolving 4 in the same impure CDCl3 solvent,
quaternary carbon (C-25, δ 70.8), two geminal methyl the regularly recorded 1H NMR spectra showed analogous
groups (C-26 and C-27, δ 1.30, 6H, δ 30.0 and 29.9), and a changes within 24 h at room temperature. The resulting
638 Journal of Natural Products, 2000, Vol. 63, No. 5 Weber et al.

Table 1. 13C NMR Data for Cycloartane Triterpenes 4-7 (75

MHz, CDCl3, δ/ppm, mult)
position 4 5 6 6a 7
1 32.0 t 31.7 t 31.7 t 31.7 t 33.4 t
2 30.4 t 30.2 t 30.1 t 30.2 t 37.4 t
3 78.8 d 77.0 d 77.0 d 77.1 d 216.5 s
4 40.5 s 43.7 s 43.6 s 43.7 s 50.2 s
5 47.1 d 42.5 d 42.4 d 42.5 d 48.4 d
6 21.1 t 21.0 t 21.0 t 21.0 t 21.4 t
7 26.0 t 25.7 t 25.7 t 25.7 t 25.8 t
8 48.0 d 47.9 d 47.9 d 47.9 d 47.8 d
9 20.0 s 20.0 s 19.9 s 19.9 s 21.0 s
10 26.1 s 25.4 s 25.3 s 25.3 s 26.0 s
11 26.5 t 26.4 t 26.3 t 26.4 t 26.6 t
12 32.8 t 32.9 t 32.7 t 32.7 t 32.1 t
13 45.3 s 45.2 s 45.2 s 45.3 s 45.1 s
14 48.8 s 48.8 s 48.7 s 48.7 s 48.8 s
15 35.6 t 35.6 t 35.5 t 35.6 t 35.4 t
16 28.1 t 28.1 t 28.0 t 28.2 t 27.5 t
17 52.0 d 52.3 d 52.0 d 52.2 d 46.4 d
18 18.1 q 18.0 q 18.1 q 18.1 q 18.3 q
19 29.9 t 30.0 t 29.8 t 30.0 t 29.5 t
20 36.4 d 35.9 d 36.3 d 36.8 d 42.4 d
21 18.3 q 18.2 q 18.3 q 18.3 q 62.5 t
22 39.0 t 36.3 t 39.0 t 39.7 t 25.0 t
23 125.6 d 24.9 t 125.6 d 129.6 d 30.7 t
24 139.4 d 125.3 d 139.3 d 134.4 d 76.2 d
25 70.7 s 130.9 s 70.8 s 142.2 s 147.6 s
26 30.0 q 17.6 q 30.0 q 114.0 t 110.9 t
27 29.9 q 25.7 q 29.9 q 18.8 q 17.7 q
28 25.4 q 71.1 t 71.1 t 71.2 t 22.2 q
29 14.0 q 10.1 q 10.1 q 10.1q 20.7 q
30 19.3 q 19.3 q 19.3 q 19.3 q 19.4 q (δ 128.7), and three oxygenated methine carbons (δ 77.3,
74.1 and 72.5). These data were in good agreement with a
cholesterol-type triterpene structure with the chemical
product was identified as cycloarta-23,25-diene-3β,28-diol shifts of the olefinic carbons pointing to the ∆5-cholesterol
(4a), which, after a further 10 days under these conditions, series. In addition to the expected methyl (three doublets
rearranged its conjugated double-bond system to form the and two singlets) and methylene proton signals, the 1H
more stable cycloarta-22,24-diene-3β,28-diol (4b). This NMR spectrum of 8 showed resonances for one olefinic
reaction has not been observed previously in cycloartanes. proton at δ 5.68 (dd, H-6), one 1H doublet at δ 4.14 (H-
In contrast to the substances 4-6, compound 7 contained 4R), and a multiplet integrating for two protons at δ 3.59
a carbonyl group (δ 216.5) instead of a β-OH group at C-3. (H-3R, H-22) (Table 2). 1H-1H COSY and long-range 2D
Furthermore, the 1H NMR spectrum showed a pattern of NMR experiments (gs-HMQC) allowed us to locate three
a hydroxymethyl substituent instead of the methyl doublet, hydroxyl groups at C-3, C-4, and C-22, respectively. The
besides a new lowfield-shifted multiplet (δ 4.03) of another configuration at C-22 was deduced from the magnitude of
methine proton together with two broad singlets (δ 4.81 the observed β-effects on the chemical shift. According to
and 4.93), the characteristic pattern of an sp2 methylene the literature,24 8 is the (22R)-epimer, because of a strong
group. The 2D NMR experiments justified the location of β-effect on C-20. The β-positions of the hydroxyls at C-3
the hydroxyl groups at C-21 and C-24. The methylene and C-4 are based on the coupling constants of the attached
carbon was assigned to C-26. Duplication of most of the methine protons and proven by NOE experiments. A NOE
signals for the side chain and some of the signals of the was observed between H-3 and H-4, whereas no NOE was
basic skeleton in its 13C NMR spectrum indicated that 7 identified between H-3/H-4 and H3-19.
was a mixture of epimers. In fact, the doubling of the A more polar part of the methanolic extract yielded
resonances associated with C-24 and its adjacent carbons compound 9, with 13C NMR spectral data (Table 3) similar
was consistent with the presence of a mixture of C-24 to those of cholest-5-ene-3β,4β,22(R)-triol (8), but differing
epimers in the ratio of about 6:4. Therefore, compound 7 in the shift data for the olefinic carbons (δ 138.0, C-8 and
was assigned the structure 21,24(RS)-dihydroxycycloart- 119.2, C-7) and with a broad proton singlet at δ 5.36, both
25-en-3-one. indicating a double bond shift to position C-7/C-8. One of
According to NMR studies of 13C-enriched cycloartenol the five observed oxygenated carbons was quaternary,
biosynthesized from [1-13C]-, [2-13C]-, and [1,2-13C2]- bearing two methyl groups identified by two singlets at δ
acetate,23 and from our own results, some of the 13C NMR 1.25 (H3-26) and 1.26 (H3-27), respectively. Therefore, C-25
spectral assignments of cycloartenol derivatives in the must be oxygenated. A long-range coupling between H3-
literature require revision, namely, C-7, C-11, C-16, C-18, 21 and C-22 (δ 80.6) identified the second oxygenated
C-21, and C-28. Thus, 13C NMR data of all four cycloartanes carbon atom. The downfield shift of the C-22 (δ 80.6) and
(4-7) are presented in Table 1. C-25 (δ 79.6) signals in comparison to those of the 22,25-
In addition, compounds (8-10) were isolated with 27 dihydroxy derivative (δ 74 and δ 70, respectively) could be
carbon atoms and a steroid skeleton, representatives of a explained by a tetrahydrofuran ring structure in the C-17
class of compounds not yet reported from Aglaia species. side chain.25 This was confirmed by the HREIMS showing
The basic structure of steroid 8 was revealed by 13C NMR the molecular ion peak at m/z 432.3218 and leading to the
and DEPT experiments, with five methyl, nine methylene, elemental formula C27H44O4 with only four oxygen atoms
10 methine, and three quaternary carbons detected, inclu- instead of five for the respective open side-chain compound.
sive of one quaternary olefinic (δ 142.8), one tertiary olefinic The absolute configuration of C-22 was derived from NOE
Phytochemical Investigation of Aglaia Journal of Natural Products, 2000, Vol. 63, No. 5 639

Table 2. 1H NMR Data for Cholesterol Derivatives 8-10 (300 In the adopted conformation of the cyclic C-17 side chain,
MHz, CDCl3, δ/ppm, TMS, J Hz, multa) H-17 is spatially close to H2-23, with dihedral angles
position 8 9b 10 between H-22 and H-20 of about 25° and between H-22
1 1.05/1.83 1.40 (m) 1.08/1.83 and H-23a+b of about 25° and 120°, respectively. The
2.45 (dd, 3.3, 14.1) corresponding coupling constants of 7, 7, and 4 Hz are
2 1.64/1.88 4.49 (br s) 1.6-1.8 found in the splitting pattern of H-22. Independent evi-
3 3.59 (m) 3.79 (br s) 3.57 (m) dence for the (22R)-configuration is derived from the 13C
4 4.14 (d, 3.4) 4.21 (br s) 3.81 (t, 2.6)
5 1.50 1.33 NMR chemical shift of C-23 (δ 25.7), which is shielded
6 5.68 (dd, 1.6, 4.8) 1.90/2.85 1.78/2.36 compared to the (22S)-epimer (around 30 ppm).25 The
7 2.08 (2H) 5.36 (br s) 5.24 (m) hydroxylation pattern of ring A was again settled by
8 1.54 1H-1H COSY and gs-HMQC experiments. The NOESY
9 0.89 1.74 1.62
11 1.46 (2H) 1.59 (2H) 1.48 (2H) spectrum revealed indirectly the β-position of all three
12 1.17/2.01 1.26/2.08 1.20/2.03 hydroxyl groups in ring A. The structure of compound 9
14 0.98 1.85 1.74 was established accordingly as (22R,25)-epoxy-cholest-7-
15 1.11/1.60 1.4-1.7 (2H) 1.4-1.6 (2H) ene-2β,3β,4β-triol.
16 1.38/1.70 1.44/1.90 1.44/1.78
17 1.12 1.25 1.16 The 1H and 13C NMR spectra of 10 were similar to those
18 0.72 (s) 0.58 (s) 0.54 (s) of compound 9. The only difference was the number of
19 1.20 (s) 1.59 (s) 1.01 (s) downfield-shifted methine protons with three (instead of
20 1.68 1.90 1.81
21 0.94 (d, 6.4) 1.00 (d, 6.7) 0.91 (d, 6.7) four) multiplets observed in the region δ 3.5-4.5 in the 1H
22 3.59 (m) 4.09 (ddd, 4, 7, 7) 4.04 (ddd, 3.5, 7, 7) NMR spectrum and only four, instead of five, carbon
23 1.20/1.32 1.68 (2H) 1.68 (2H) resonances between δ 72 and 81 in the 13C NMR spectrum.
24 1.16/1.40 1.65 (2H) 1.66 (2H) Accordingly, one secondary hydroxyl group was absent in
25 1.55
26 0.92 (d, 6.4) 1.25 (s) 1.22 (s) 10. An unusual 4-hydroxyl substitution was supported
27 0.91 (d, 6.6) 1.26 (s) 1.23 (s) unambiguously by 1H-1H COSY relations between H-4 (δ
a All 1H NMR shifts given without multiplicities consist of 3.81)fH-3 and H-4fH-5 and by strong long-range cross-
nonresolved overlapping multiplets; their correct values and peaks between H-4fC-2, H-4fC-6, and H-4fC-10, respec-
assignments were determined by 2D hetero-techniques. tively. Further comparison with the spectral data of
b Spectrum recorded in pyridine-d .
5 compound 9 revealed 10 to contain the identical cyclized
C-17 side chain. Taking into account all available data,
Table 3. 13C NMR Data for Cholesterol Derivatives 8-10 (75
MHz, CDCl3 δ/ppm, TMS, mult)
compound 10 proved to be the new (22R,25)-epoxy-cholest-
7-ene-3β,4β-diol. It is interesting to note that 9 and 10
position 8 9a 10 display the same side chain as some ecdysteroids,26 the
1 36.9 t 45.1 t 37.3 t well-known insect molting hormones with an (R)-configured
2 25.4 t 73.0 d 25.6 t C-22.
3 72.5 d 72.7 d 72.6 d
4 77.3 d 76.0 d 73.2 d The first compound isolated from an Aglaia species was
5 142.8 s 45.9 d 44.4 d aglaiol,3 a tetracyclic triterpene with a dammarane skel-
6 128.7 d 27.2 t 26.0 t eton. We have now isolated two more dammaranes from
7 32.1 t 119.2 d 117.9 d A. rubiginosa, which were identified as the known (20S,24S)-
8 31.9 d 138.0 s 139.0 s dihydroxydammar-25-en-3-one (11) and (20S,25)-dihydroxy-
9 50.2 d 52.8 d 50.7 d
10 36.1 s 34.4 s 34.2 s dammar-23-en-3-one (isofouquierone) (12).27,28 Further-
11 20.6 t 21.3 t 21.0 t more, three stigmasterol derivatives were identified as the
12 39.7 t 39.8 t 39.4 t ubiquitous β-sitosterol, its glucoside, and stigmast-5-ene-
13 42.7 s 44.2 s 43.9 s 3β,7R-diol (13).29 The NMR data and the spectral properties
14 56.5 d 54.9 d 54.6 d of these compounds were consistent with those in the
15 24.4 t 23.6 t 23.1 t
16 27.4 t 27.6 t 27.3 t literature.27-29
17 53.2 d 54.3 d 53.7 d The accumulation of cycloartanes, dammaranes, choles-
18 11.9 q 12.1 q 11.8 q terol derivatives, and even the biogenetically more ad-
19 21.0 q 18.5 q 15.2 q vanced androstanes in one single species, gives rise to the
20 42.3 d 39.2 d 38.5 d
21 12.4 q 13.0 q 12.4 q supposition that the biogenesis of these different structural
22 74.1 d 80.6 d 80.2 d types is closely connected. The androstanes 1 and 2
23 27.7 t 25.7 t 24.8 t represent a novel group of steroids from plants. Related
24 36.0 t 39.1 t 38.9 t pregnane derivatives without the characteristic A-ring
25 28.2 d 79.6 s 79.7 s dienone have been isolated from species in the Meliaceae
26 22.5 q 28.2 q 28.0 q
27 22.9 q 29.0 q 28.6 q
(Melia30 and Trichilia31 species), the Simaroubaceae,32 and
the Burseraceae.33 Due to their rarity, these steroids may
a Recorded in pyridine-d5. be regarded as significant chemotaxonomic evidence sup-
experiments in combination with the analysis of the porting the proposed link between these three families that
coupling constants of the H-22 signal group, based on the all belong to the order Rutales. We did not isolate any of
configurational and conformational arrangement of the the insecticidal flavaglines or related cyclopenta[b]benzo-
CH3-18-C-13-CH-17-CH-20-CH3-21 structural frag- furans from A. rubiginosa. The absence of flavonoids, which
ment, in which H-17 is antiperiplanar to H-20 at the (S)- represent important biosynthetic precursors of the flava-
configured C-20. Inspecting the Dreiding model of com- glines,5 corroborates these results.
pound 9, we discovered the crucial NOE effects between On the other hand the hydroxylated cholesterol deriva-
H-17 and H2-23 as well as between H-20 and H-22, together tives show a structural relationship to the ecdysteroids,26,34
with the missing NO enhancements between H-22 and H3- and the A-ring dienone structure of the androstanes is also
21 are compatible only with the (R)-configuration of C-22. present in the insecticidal petuniasterones,35 pointing to
640 Journal of Natural Products, 2000, Vol. 63, No. 5 Weber et al.

Extraction and Isolation. Air-dried leaves of A. rubigi-

nosa (10 kg) were ground and exhaustively extracted with
methanol. The methanolic extract (550 g) was evaporated to
dryness under reduced pressure. A part of the residue (94 g)
was preadsorbed on Kieselgur (Seitz-Filter-Werke, Bad
Kreuznach, Germany) and successively eluted with n-hexane,
ethyl acetate, and methanol, respectively. The EtOAc fraction
(37 g) and the MeOH fraction (10 g) were roughly separated
by column chromatography on Sephadex LH-20 with MeOH
(600 fractions of 10 mL and 200 fractions of 15 mL, respec-
tively) followed by repeated chromatography on Si gel with
different eluents. The n-hexane residue (47 g) was directly
chromatographed on Si gel using a cyclohexane-EtOAc gradi-
ent (1000 fractions of 15 mL). Lobar column chromatography
(UV detection, 254 nm) was used to finally purify the com-
pounds.
n-Hexane Extract. Fractions 166-300 of the chromato-
graphic separation of the n-hexane extract directly yielded 1
g of β-sitosterol. Si gel chromatography of fractions 301-485
(5 g) with cyclohexane-EtOAc (2:1) yielded 4 (100 mg) and
that of the following fractions 486-670 (4 g) with cyclohexane-
EtOAc (1:1), compound 10 (10 mg). Fractions 671-900 (11.5
g) were separated by Si gel rechromatography with cyclohex-
ane-EtOAc (2:1), which led to the isolation of 5 (20 mg)
(fractions 533-750), 11 (20 mg) (fractions 751-920), and 13
(30 mg) (fractions 980-1119). Fractions 921-979 (1 g) were
further purified by repeated Lobar column chromatography
(2% MeOH in CH2Cl2) to yield finally 10 mg of 2 as an oily
substance and 10 mg of 12.
EtOAc Extract. Fractions 76-117 (16 g) of the chromato-
graphic separation of the EtOAc extract on Sephadex LH-20
were further separated by Si gel chromatography (cyclohex-
ane-EtOAc gradient). Fractions 417-500 (930 mg) were
eluted with cyclohexane-EtOAc (1:1) and afforded 80 mg of
8, while fractions 1037-1056 contained β-sitosterol-β-D-glu-
coside (20 mg). Fractions 566-1025 (6 g), were further purified
by repeated Lobar column chromatography (3% MeOH in
CH2Cl2), which led to the isolation of 7 (15 mg) (fractions 29-
50), an additional amount of 13 (8 mg) (fractions 51-61), and
6 (5 mg) (fractions 62-88), respectively. Lobar column chro-
matography of a portion (5 g) of fractions 118-148 (7.6 g) from
different mechanisms and active principles with a signifi- the EtOAc extract, using CH2Cl2-EtOAc (4:1), afforded 100
cance in the chemical plant-insect interactions. mg of 3 and 20 mg of 1, respectively.
It is hypothesized that these significant deviations MeOH Extract. The MeOH extract was chromatographed
observed in the secondary metabolism might represent on a Sephadex LH-20 column with MeOH to yield 30 mg of 9
important differences between the section Amoora and the (fractions 54-70) as an amorphous powder.
section Aglaia. Further phytochemical investigations may 6R-Hydroxyandrosta-1,4-diene-3,17-dione (1): mp 240-
help to establish a final chemotaxonomic classification. 242 °C; [R]20D +96.6° (c 0.11, MeOH); UV (MeOH) λmax () 240
(11 506) nm; CD (MeCN) λ (θ) 298 (7484), 258 (-2753), 236
Experimental Section (13 309) nm; IR (KBr) νmax 3480 (OH), 1738 (17-CdO), 1655
General Experimental Procedures. Melting points were (3-CdO), 1613 (CdC) cm-1; 1H NMR (CDCl3) δ 6.99 (1H, d, J
determined on a Mel-Temp II apparatus (Laboratory Devices, ) 10.0 Hz, H-1), 6.22 (1H, t, J ) 1.8 Hz, H-4), 6.48 (1H, dd, J
) 1.9, 10.3 Hz, H-2), 4.49 (1H, ddd, J ) 1.7, 5.5, 11.7 Hz, H-6),
Holliston, MA) and are uncorrected. Optical rotations were
2.45 (1H, dd, J ) 9.0, 19.0 Hz, H-16b), 2.33 (1H, ddd, J ) 3.7,
measured on a Perkin-Elmer 241 polarimeter. IR spectra were
5.5, 12.0 Hz, H-7b), 2.07 (1H, dd, J ) 10.0, 19.0 Hz, H-16a),
obtained on a Perkin-Elmer 1605 FT-IR spectrometer. 1H NMR 1.96 (1H, m, H-15b), 1.85 (2H, m, H-8, H-12b), 1.83 (1H, m,
and 13C NMR spectra were recorded on a Varian Unity 300 H-11b), 1.65 (1H, m, H-11a), 1.59 (1H, m, H-15a), 1.29 (1H,
(1H NMR at 299.95 MHz, 13C NMR at 75.4 MHz) using TMS m, H-14), 1.26 (1H, m, H-12a), 1.21 (3H, s, H-19), 1.10 (1H,
as internal standard. With a Finnigan MAT 312 apparatus, m, H-7a), 1.07 (1H, m, H-9), 0.91 (3H, s, H-18); 13C NMR
mass spectra were obtained at 70 eV. Column chromatography (CDCl3) δ 219.5 (s, C-17), 186.1 (s, C-3), 169.6 (s, C-5), 155.1
was performed on Sephadex LH-20 (Pharmacia, Freiburg, (d, C-1), 127.7 (d, C-2), 119.8 (d, C-4), 68.0 (d, C-6), 52.1 (d,
Germany); Si 60 (63-200 µm) (Merck, Darmstadt, Germany), C-9), 50.2 (d, C-14), 47.7 (s, C-13), 43.5 (s, C-10), 41.1 (t, C-7),
and Lobar A (240 × 10 mm), B (310 × 25 mm), and C (440 × 35.6 (t, C-16), 33.6 (d, C-8), 31.1 (t, C-12), 22.0 (t, C-11), 21.9
37 mm) Lichroprep Si 60 (40-63 µm) (Merck), respectively. (t, C-15), 19.1 (q, C-19), 13.8 (q, C-18); EIMS (70 eV) m/z 300
TLC spots (Si gel 60 F254, Merck) were detected with a UV254 [M]+ (12), 282 [M - H2O]+ (4), 255 (17), 228 (10), 171 (10),
lamp and by 40% H2SO4 followed by heating at 120 °C for 5 159 (11), 147 (17), 138 (55), 121 (31), 107 (72), 91 (64), 79 (92),
min. 67 (47), 55 (81), 41 (100); HREIMS m/z 300.1725 (calcd for
Plant Material. The leaves of A. rubiginosa (Hiern) Pannell C19H24O3, 300.1726).
(Meliaceae) were collected from a peat swamp forest in Androsta-1,4-diene-3,17-dione (2): oil, [R]20D +66.7° (c
Narathiwat Province (Thailand) in May 1994. A voucher 0.14, MeOH), UV (MeOH) λmax () 240 (7652) nm [242-243
specimen (no. ES-94051) is deposited at the herbarium of the nm];36 CD (MeOH) λ (θ) 331 (-1607), 295 (5564), 260 (-4694),
Department of Pharmaceutical Botany, Faculty of Pharma- 230 (10 444) nm; 1H and 13C NMR data were coincident with
ceutical Sciences, Chulalongkorn University, Bangkok, Thai- those from the literature;37 HREIMS m/z 284.1775 (calcd for
land. C19H24O2, 284.1776).
Phytochemical Investigation of Aglaia Journal of Natural Products, 2000, Vol. 63, No. 5 641

Ring A seco-17-octanor-5,17-dioxo-cycloartane-1-car- (100), 361 (16), 329 (16), 302 (17), 289 [M - side chain
boxylic acid (5a,6-methano-3a,9b-dimethyl-3,7-dioxo-cy- C8H17O]+ (9) 285 (21), 274 [M - CH3 - side chain C8H17O]+
clopentano[a]naphthalene-6-yl-ethanoic acid) (3): mp (26), 271 [M - H2O - side chain C8H17O]+ (20), 243 (13), 229
198 °C; [R]20D +60.8° (c 0.11, MeOH); UV (MeOH) λmax () 280 (14), 173 (13), 147 (16), 131 (13), 105 (18), 83 (27), 69 (15), 55
(69) nm; CD (MeOH) λ (θ) 302 (2048), 271 (-2475), 214 (9168) (19); HREIMS m/z 418.3454 (calcd for C27H46O3, 418.3447).
nm; IR (KBr) νmax 3400 (OH), 2934, 1721 (CdO), 1710 (sh, Cd (22R,25)-Epoxycholest-7-ene-2β,3β,4β-triol (9): mp >250
O), 1683 (CdO), 1457, 1377, 1113, 968 cm-1; 1H NMR (CDCl3) °C (dec); [R]20D +23.1° (c 0.1, MeOH); IR (KBr) νmax 3386 (OH),
δ 2.70 (2H, d, J ) 7.6, H-1a, H-1b), 2.40 (2H, m, H-6b, H-16b), 3306 (OH), 2964 (OH), 1443 and 1379 (CdC), 1142 and 1096
2.35 (1H, m, H-8), 2.32 (1H, m, H-6), 2.23 (1H, dt, J ) 9.0, (C-O-C) cm-1; 1H NMR (pyridine-d5) and 13C NMR (pyridine-
19.5, H-16a), 1.96-1.97 (2H, m, H-7b, H-11b), 1.73-1.82 (2H, d5), see Tables 2 and 3; EIMS (70 eV) m/z 432 [M]+ (20), 414
m, H-15a, H-15b), 1.80 (1H, m, H-7a), 1.68 (1H, d, J ) 5.7, [M - H2O]+ (9), 334 (6), 185 (6), 171 (7), 152 (14), 145 (13),
H-19b), 1.65 (1H, m, H-12b), 1.56 (1H, m, H-11a), 1.49 (1H, 131 (14), 105 (27), 100 (47), 99 (100), 81 (100); HREIMS m/z
m, H-12a), 1.14 (3H, s, H-30), 0.96 (1H, d, J ) 5.7, H-19a), 432.3218 (calcd for C27H44O4, 432.3239).
0.83 (3H, s, H-18); 13C NMR (CDCl3) δ 218.8 (s, C-17), 209.7 (22R,25)-Epoxycholest-7-ene-3β,4β-diol (10): mp 230 °C;
(s, C-5), 176.5 (s, COOH), 53.5 (s, C-13), 44.3 (s, C-14), 38.4 (s, [R]20D +23.5° (c 0.1, MeOH); IR (KBr) νmax 3422 (OH), 2964
C-10), 38.3 (d, C-8), 34.0 (s, C-9), 33.8 (t, C-1), 33.6 (t, C-6), (OH), 1455 and 1375 (CdC), 1146 and 1047 (C-O-C) cm-1;
33.6 (t, C-16), 30.9 (t, C-15), 27.1 (t, C-11), 24.8 (t, C-12), 24.0 1
H NMR (CDCl3) and 13C NMR (CDCl3), see Tables 2 and 3;
(t, C-19), 19.7 (q, C-30), 19.4 (t, C-7), 17.8 (q, C-18); EIMS (70 EIMS (70 eV) m/z 416 [M]+ (3), 398 [M - H2O]+ (2), 185 (3),
eV) m/z 304 [M]+ (18), 286 [M - H2O]+ (71), 271 [M - H2O - 171 (3), 152 (9), 145 (7), 131 (10), 105 (14), 100 (20), 99 (100),
CH3]+ (42), 258 (15), 243 (16), 229 (12), 201 (9), 187 (7), 173 81 (100); ESIMS m/z 439 [M + Na]+ (100) {M+ ) 416}.
(8), 161 (10), 133 (9), 105 (31), 91 (68), 79 (72), 55 (100), 41
(89); HREIMS m/z 304.1685 (calcd for C18H24O4, 304.1675).
Acknowledgment. This investigation was financially sup-
Cycloart-23-ene-3β,25,28-triol (6): Physical data for 6
ported by “Fonds der Chemischen Industrie”. We wish to thank
could not be obtained because of decomposition during NMR
analysis (H2O-elimination, which led to 6a); 1H NMR (CDCl3) also Schering AG (Berlin, Germany) for the generous gift of
δ 5.59 (2H, m, H-23, H-24), 3.74 (1H, dd, J ) 5.0, 10.7, H-3), the androstane derivative 1 as synthetic reference sample, and
3.71 (1H, d, J ) 10.5, H-28b), 3.50 (1H, d, J ) 10.5, H-28a), we gratefully acknowledge Dr. E. Saifah (Department of
2.16 (1H, m, H-22b), 1.98 (1H, m, H-11b), 1.90 (1H, m, H-16b), Pharmaceutical Botany, Faculty of Pharmaceutical Sciences,
1.70 (1H, m, H-22a), 1.74 (1H, m, H-2b), 1.6 (3-H, m, H-2a, Chulalongkorn University, Bangkok, Thailand) for collecting
H-12a, H-12b), 1.56 (1H, m, H-17), 1.5 (1H, m, H-1b), 1.48 (1H, and identifying the plant material, as well as Dr. J. Wörth
m, H-8), 1.44 (1H, m, H-20), 1.44 (1H, m, H-5), 1.40 (1H, m, (Department of Organic Chemistry, University of Freiburg,
H-6b), 1.36 (1H, m, H-7b), 1.30 (6H, s, H-26, H-27), 1.29 (1H, Germany) for the mass spectra.
m, H-16a), 1.28 (2H, m, H-15a, H-15b), 1.22 (1H, m, H-1a),
1.12 (1H, m, H-11a), 1.06 (1H, m, H-7a), 0.96 (3H, s, H-18), References and Notes
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