Tetranorclerodanes and clerodane-type diterpene glycosides from Dicranopteris dichotoma

Article abstract of DOI:10.1021/np0603166

The acetone extract of Dicranopteris dichotoma afforded two new tetranorclerodanes, 18-hydroxyaylthonic acid (1) and 18-oxo-aylthonic acid (2), and four new clerodane-type diterpene glycosides, (6S,13S)-6-0-[6-O-acetyl- β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl]cleroda-3, 14-dien-13-ol (3), (65,13S)-6-0-[4-0-acetyl-β-D-glucopyranosyl-(1→4)- α-L-rhamnopyranosyl]cleroda-3,14-dien-13-ol (4), 6-O-[6-O-acetyl-β-D- glucopyranos-yl-(1→4)-α-L-rhamnopyranosyl]-(13E)-cleroda-3, 13-dien-15-ol (5), and 6-O-[β-D-glucopyranosyl]-(1→4)-α-L- rharnnopyranosyl-(13E)-cleroda-3,13-dien-15-ol (6), together with two known compounds, aylthonic acid (7) and (65,13S)-cleroda-3,14-diene-6,13-diol (8). The structures of these new compounds were established by a combination of 1D and 2D NMR techniques, MS, and acid hydrolysis. Compound 8 showed modest anti-HIV-1 activity.

Full text of DOI:10.1021/np0603166

J. Nat. Prod. 2007, 70, 265-268
265
Tetranorclerodanes and Clerodane-Type Diterpene Glycosides from Dicranopteris dichotoma
Xiao-Li Li,†, Liu-Meng Yang,§ Yu Zhao,†, Rui-Rui Wang,§ Gang Xu,† Yong-Tang Zheng,§ Lin Tu,†, Li-Yan Peng,†
Xiao Cheng,† and Qin-Shi Zhao*,†
State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences,
Kunming 650204, Yunnan, People’s Republic of China, Key Laboratory of Animal Models and Human Disease Mechanisms and Laboratory of
Molecular Immunopharmacology, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650223, Yunnan, People’s Republic of
China, and Graduate School of the Chinese Academy of Sciences, Beijing 100039, People’s Republic of China
ReceiVed July 3, 2006
The acetone extract of Dicranopteris dichotoma afforded two new tetranorclerodanes, 18-hydroxyaylthonic acid (1)
and 18-oxo-aylthonic acid (2), and four new clerodane-type diterpene glycosides, (6S,13S)-6-O-[6-O-acetyl-â-D-
glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]cleroda-3,14-dien-13-ol (3), (6S,13S)-6-O-[4-O-acetyl-â-D-glucopyranosyl-
(1f4)-R-L-rhamnopyranosyl]cleroda-3,14-dien-13-ol (4), 6-O-[6-O-acetyl-â-D-glucopyranos-yl-(1f4)-R-L-rhamnopyranosyl]-
(13E)-cleroda-3,13-dien-15-ol (5), and 6-O-[â-D-glucopyranosyl]-(1f4)-R-L-rhamnopyranosyl-(13E)-cleroda-3,13-dien-
15-ol (6), together with two known compounds, aylthonic acid (7) and (6S,13S)-cleroda-3,14-diene-6,13-diol (8). The
structures of these new compounds were established by a combination of 1D and 2D NMR techniques, MS, and acid
hydrolysis. Compound 8 showed modest anti-HIV-1 activity.
Dicranopteris species are very common ferns and grow as large
communities containing no other plant species. Previous studies
of Dicranopteris species have reported flavonoids,1,2 phenols,3
proanthocyanidins,4 and clerodane-type glycosides.2,5 Recent re-
search showed that some clerodane-type glycosides isolated from
Dicranopteris species accelerated the growth of the stems of lettuce
or inhibited the root growth.6 To find bioactive secondary metabo-
lites from this species, we chemically investigated the fronds of D.
dichotoma Bernb. and reported the isolation and structural elucida-
tion of two new highly oxygenated phenolic derivatives, dichoto-
mains A and B.6 Continuous screening of the fronds of D.
dichotoma led to the discovery of two new tetranorclerodanes, 18-
hydroxyaylthonic acid (1) and 18-oxo-aylthonic acid (2), and four
new clerodane-type diterpene glycosides, (6S,13S)-6-O-[6-O-acetyl-
â-D-glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]cleroda-3,14-di-
ene-13-ol (3), (6S,13S)-6-O-[4-O-acetyl-â-D-glucopy-ranosyl-(1f4)-
R-L-rhamnopyranosyl]cleroda-3,14-dien-13-ol (4), 6-O-[6-O-acetyl-
â-D-glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]-(13E)-cleroda-
3,13-dien-15-ol (5), and 6-O-[â-D-glucopyranosyl-(1f4)-R-L-
rhamnopyranosyl]-(13E)-cleroda-3,13-dien-15-ol (6), together with
two known compounds, aylthonic acid (7)7 and (6S,13S)-cleroda-
3,14-diene-6,13-diol (8).6 In the present paper, we report the
structural characterization of compounds 1-8 and their anti-HIV-1
activities.
Results and Discussion
Compound 1 was obtained as colorless crystals. The molecular
formula of C16H26O4 was established on the basis of HRESIMS
[M - H]- (found 281.1755, calcd 281.1752). The 13C NMR
spectrum of 1 displayed the signals of one carboxylic acid group
(δC 174.4), two quaternary carbons (δC 45.0 and 40.5), two olefinic
carbons (δC 147.6 and 126.0), three methines (δC 74.9, 47.3, and
36.2), five methylenes (δC 66.3, 43.8, 37.2, 27.1, and 19.4), and
three methyls (δC 17.5, 17.0, and 16.4). Its IR spectrum showed
the absorption bands typical for the hydroxyl group (3483 cm-1
)
1
and carboxylic acid group (1700, 3251-2857 cm-1). The H and
13C NMR spectra of 1 were similar to those of aylthonic acid (7).7
The key difference was that the methyl group (Me-18) in 7 was
replaced by a hydroxymethyl group (δC 66.3) in 1, which was
confirmed by the HMBC correlations of H2-18 (δH 4.56, 4.41) with
C-3 (δC 126.0), C-4 (δC 147.6), and C-5 (δC 45.0).
The relative configuration of 1 was determined by a ROESY
experiment. The ROESY correlations of H-6/H-8 and H-10 and of
* Corresponding author. Tel: 86-871-5223058. Fax: 86-871-5215783.
E-mail: Qinshizhaosp@yahoo.com.
† Kunming Institute of Botany.
§ Kunming Institute of Zoology.
Graduate School of the Chinese Academy of Sciences.
10.1021/np0603166 CCC: $37.00
© 2007 American Chemical Society and American Society of Pharmacognosy
Published on Web 01/18/2007

266 Journal of Natural Products, 2007, Vol. 70, No. 2
Li et al.
identified a glucopyranosyl (1f4) rhamnosyl linkage. The C-6
location of the acetyl group in the glucopyranosyl unit was
confirmed by HMBC correlations of H-6′′ (δH 4.85 and 4.75) with
the carbonyl carbon (δC 170.9) of the acetyl group. Furthermore,
the sugar chain was linked to C-6 of the aglycone by HMBC
correlation of the anomeric proton (δH 5.35) of the rhamnopyranosyl
unit and C-6 (δC 86.5) of the aglycone. The rhamnopyranosyl and
glucopyranosyl units were in R- and â-configurations, respectively,
by the coupling constants of their anomeric protons (Table 1).5
The relative configuration was established by a ROESY experi-
ment. The ROESY correlations confirmed that H-6, H-8, and H-10
were â-oriented, while Me-17, Me-19, and Me-20 were R-oriented.
(6S,13S)-Cleroda-3,14-diene-6,13-diol (8) was also isolated from
this plant, and its specific rotation ([R]25 -6.7) coincided with
Figure 1. ROESY correlations of compound 1 and the aglycone
of 5.
D
the reported value.5 From a biogenetic perspective, compound 8
should be the aglycone of 3 because it was isolated from the same
genus. Thus, the structure of 3 was determined as (6S,13S)-6-O-
[6-O-acetyl-â-D-glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]cle-
roda-3,14-dien-13-ol.
Compound 4 was obtained as a pale yellow powder. Its molecular
1
formula C34H56O12 was determined by analysis of H, 13C, and
DEPT NMR data and verified by HRESIMS (found 655.3700, calcd
655.3693), which revealed 11 unsaturation degrees. The resem-
blance of the NMR spectra (Tables 1 and 2) to those of 3 suggested
that 4 was a related clerodane-type diterpene glycoside. The acetyl
group in compound 4 was determined to be at C-4′′ (δC 72.3) of
the glucosyl moiety from the HMBC correlation of H-4′′ (δH 5.66)
to the carbonyl carbon (δC 170.5). Sugar analysis of 4 was carried
out using the same method as for 3. Therefore, compound 4 was
identified as (6S,13S)-6-O-[4-O-acetyl-â-D-glucopyranosyl-(1f4)-
R-L-rhamnopyranosyl]cleroda-3,14-dien-13-ol.
Figure 2. HMBC correlations of compounds 1 and 3.
H-10/H-11â indicated that H-6, H-8, and H-10 were â-oriented and,
accordingly, the hydroxyl group at C-6 was R-oriented. The ROESY
correlation of Me-19/Me-20 and H-1R and of Me-20/Me-17
suggested that Me-17, Me-19, and Me-20 possessed R-orientations
(Figure 1). The structure of 1 was 18-hydroxyaylthonic acid.
Compound 2, colorless crystals, had the molecular formula
C16H24O4 on the basis of the HRESIMS (found 279.1596 and calcd
279.1596). The similarity of the 1H and 13C NMR spectra of 1 and
2 (Tables 1 and 2) suggested that 2 was a related tetranorclerodane
derivative. The only difference between the two compounds was
that the hydroxymethylene moiety (δC 66.3, C-18) in 1 was replaced
by an aldehydic function (δC 198.2) in 2. This was confirmed by
the HMBC correlations between the aldehydic proton (δH 9.42)
and C-3 (δC 157.8), C-4 (δC 151.4), and C-5 (δC 45.2) and supported
by the downfield shift of C-3 and C-4 from δC 123.0 and 147.6 in
1 to δC 157.8 and 151.4 in 2. The relative configuration was
established via a ROESY experiment. Similar ROESY correlations
to those in 1 confirmed that H-6, H-8, and H-10 were â-oriented,
while Me-17, M-19, and Me-20 were R-oriented. Thus, compound
2 was identified as 18-oxo-aylthonic acid.
Compound 5 showed a molecular ion at m/z 655 [M - H]-, and
the molecular formula C34H56O12 was established by HRESIMS
1
(found 655.3703, calcd 655.3693). The H and 13C NMR spectra
of 5 (Tables 1 and 2) indicated the presence of a diterpene, two
hexoses, and an acetyl group. The 1H and 13C NMR spectra of the
aglycone of 5 were similar to those of 3, except for the resonances
of the side chain (Tables 1 and 2). HMBC correlations from Me-
16 (δH 1.76) to C-14 (δC 126.6) and from H2-15 (δH 4.46) to C-13
(δC 138.0) confirmed that the side chain had a C-13-C-14 double
bond and C-15 hydroxymethyl moiety. The acetyl group was located
at C-6′′ of the glucosyl moiety by HMBC correlations of H2 (δH
4.85 and 4.74) with the carbonyl carbon (δC 170.9) of the acetyl
group. Acid hydrolysis of 5 by the same method used for 3 led to
decomposition of the aglycone, but gave D-glucose and L-rhamnose.
The two monosaccharides were identified by comparison of their
Rf and specific rotation with those of authentic samples.
The ROESY correlations of Me-16/H2-15 established the 13E
configuration of the C-13-C-14 double bond. The â-orientation
of H-6, H-8, and H-10 and the R-orientation of Me-17, M-19, and
Me-20 were also confirmed by ROESY correlations (Figure 1).
Thus, the structure of 5 was defined as 6-O-[6-O-acetyl-â-D-
glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]-(13E)-cleroda-3,13-
dien-15-ol.
Compound 3 was obtained as a pale yellow powder, and its
molecular formula was indicated as C34H56O12 by HRESIMS (found
655.3691, calcd 655.3693), corresponding to seven unsaturation
1
degrees. The H and 13C NMR spectra of 3 showed resonances
characteristic of a diterpene, two hexoses, and an acetyl group
(Tables 1 and 2). Assignment of each glycosidic proton system
1
was achieved by analysis of H-1H COSY and HMQC-TOCSY
Compound 6 was formulated as C32H54O11 from HRESIMS
experiments. The diterpene possessed a secondary methyl, three
teriary methyls, four olefinic carbons, an oxygenated methine, and
an oxygenated quaternary carbon, which suggested that it was
similar to (6S,13S)-cleroda-3,14-diene-6,13-diol (8),5 a common
aglycone of the clerodane-type glycosides of Dicranopteris.2,5 The
sugar moieties comprised a rhamnopyranosyl and a glucopyranosyl
unit by similarity of their spectroscopic data with literature data.2,5,8
Acid hydrolysis of 3 could not afford aglycone, which was
decomposed under acid condition, but gave D-glucose and L-
rhamnose. The two monosaccharides were identified by comparison
of their Rf and specific rotation with those of authentic samples.9,10
HMBC correlation between the anomeric proton (δH 5.19) of the
glucopyranosyl unit and C-4 (δC 85.2) of the rhamnosyl unit
1
(found 613.3578, calcd 613.3587). The H and 13C NMR spectra
of 6 (Tables 1 and 2) were co-incident with 5 except for the absence
of an acetyl group in 6. Sugar analysis of 6 was also carried out as
for 5. The ROESY correlations of Me-16 (δH 1.77) with H2-15
(δH 4.48) suggested that the double bond between C-13 and C-14
was also E-configured. Accordingly, compound 6 was determined
to be 6-O-[â-D-glucopyranosyl-(1f4)-R-L-rhamnopyranosyl]-(13E)-
cleroda-3,13-dien-15-ol.
Compounds 1-8 were tested for cytotoxicity against C8166 cells
(CC50), and anti-HIV-1 activity was evaluated by the inhibition
assay for the cytopathic effects of HIV-1 (EC50), using AZT as a
positive control. Compound 8 exerted modest cytotoxic activity

Tetranorclerodanes from Dicranopteris dichotoma
Journal of Natural Products, 2007, Vol. 70, No. 2 267
Table 1. 1H NMR Assignments of Compounds 1-6a
no.
1
2
3
4
5
6
1 R
1 â
2 R
2 â
3
1.77 (m)
2.10 (m)
2.16 (m)
2.30 (m)
5.66 (br s)
3.98 (t, 4.3)
1.82 (2H, m)
1.55 (m)
1.87 (m)
2.11 (m)
2.31 (m)
6.64 (d, 3.2)
3.71 (dd, 11.1, 4.5)
1.72 (2H, m)
1.50 (m)
1.61 (m)
1.92 (2H, m)
1.48 (2H, overlapped)
1.48 (m)
1.52 (m)
1.93 (2H, m)
1.50 (m)
1.56 (m)
1.95 (2H, m)
1.89 (2H, m)
5.13 (m)
3.39 (m)
1.60 (overlapped)
1.51 (overlapped)
2.15 (m)
5.11 (s)
3.39 (m)
1.72 (2H, m)
5.18 (overlapped)
3.39 (m)
2.15 (m)
1.57 (m)
1.44 (m)
1.23 (m)
1.23 (m)
1.29 (m)
1.85 (2H, m)
5.13 (s)
6
3.40 (m)
2.15 (m)
1.57 (m)
1.45 (m)
1.26 (m)
1.30 (m)
1.39 (m)
1.87(2H, m)
7 R
7 â
8
2.40 (m)
2.05 (m)
2.52 (d, 13.6)
2.62 (d, 13.6)
2.36 (m)
1.82 (m)
2.51 (2H, m)
2.10 (m)
1.29 (m)
1.52 (m)
1.60 (m)
1.60 (m)
1.52 (m)
6.18 (m)
5.17 (m)
5.57 (m)
10
1.31 (m)
11 R
11 â
12 R
12 â
14
15 R
15 â
1.58 (overlapped)
1.51 (overlapped)
1.58 (m)
1.55 (m)
6.18 (m)
5.74 (m)
4.46 (2H, m)
5.75 (m)
4.48 (2H, m)
5.24 (overlapped)
5.57
(dd, 6.5, 17.3)
1.51 (3H, s)
0.79
16
17
1.51 (3H, s)
0.74
1.76 (3H, s)
0.72
1.77 (3H, s)
0.73
1.00
0.98
(3H, d, 6.7)
4.41 (d., 12.7)
4.56 (d., 12.7)
1.80 (3H, s)
0.79 (3H, s)
(3H, d, 6.7)
9.42 (s)
(3H, d, 6.3)
1.74 (3H, overlapped)
(3H,overlapped)
1.74 (3H, s)
(3H, d, 6.6)
1.74 (3H, s)
(3H, d, 6.7)
1.74 (3H, s)
18
19
20
Rha
1
2
3
4
5
6
1.21 (3H, s)
0.73 (3H, s)
1.07 (3H, s)
0.74 (3H, s)
1.07 (3H, s)
0.74 (3H, overlapped)
1.04 (3H, s)
0.44 (3H, s)
1.08 (3H, s)
0.67 (3H, s)
5.35 (br s)
4.50 (m)
4.54 (m)
4.38 (t, 8.3)
4.20 (m)
1.74
5.33 (br s)
4.49 (m)
4.55 (m)
4.44 (t, 8.3)
4.20 (m)
1.74
5.35 (br s)
4.48 (m)
4.52 (m)
4.40 (t, 8.2)
4.21 (m)
1.72
5.34 (br s)
4.50 (m)
4.54 (m)
4.45 (t,8.3)
4.24 (m)
1.69
(3H, overlapped)
(3H, overlapped)
(3H, overlapped)
(3H, d, 6.3)
Glc
1
2
3
4
5.19 (d, 7.3)
4.12 (t, 7.3)
4.16 (m)
4.00 (t, 7.3)
3.86 (m)
5.28 (d, 7.3)
4.29 (m)
4.15 (m)
5.66 (m)
3.84 (m)
4.15 (m)
4.06 (m)
5.20 (d, 7.2)
4.16 (m)
4.17 (m)
4.00 (t, 7.2)
3.85 (m)
4.85 (d, 11.3)
4.75 (m)
5.26 (d, 7.2)
4.16 (t, 7.2)
4.25 (m)
4.29 (t, 7.2)
3.80 (m)
5
6
4.85 (d, 11.3)
4.74 (m)
4.47 (m)
4.38 (m)
Ac
CH3
1.95 (3H, s)
1.98 (3H, s)
1.93 (3H, s)
a Spectra were recorded in C5D5N; chemical shifts (δ) are in ppm and J in Hz.
Extraction and Isolation. The dry fronds of D. dichotoma (32 kg)
against C8166 cells with CC50 > 653.04 µM and showed anti-
were powdered and extracted with acetone (3 × 30 L) for 24 h at room
temperature. The acetone extract was concentrated in vacuo to give a
crude extract (2.3 kg), which was then suspended in H2O and subjected
to column chromatography over DM 130 eluting with H2O and 95%
EtOH, respectively. The fraction eluting by 95% EtOH was concentrated
in vacuo to give a residue (1.5 kg), which was then subjected to column
chromatography over silica gel (200-300 mesh) eluting with CHCl3-
MeOH (from 1:0 to 0:1) to afford fractions A-H. Fraction B was
subjected to column chromatography over silica gel eluting with
CHCl3-MeOH (100:1) to yield 1 (30 mg), 2 (10 mg), 7 (20 mg), and
8 (47 mg). Fraction D was subjected to RP-18 and silica gel column
chromatography, repeatedly, to afford 6 (100 mg). Further purification
on Sephadex LH-20, RP-18, and semipreparation HPLC (Agilent 1100
HPLC system; Zorbax SB-C18, 250 × 9.4 mm; UV detector) yielded
3 (25 mg), 4 (7.8 mg), and 5 (6 mg).
HIV-1 activity with EC50 ) 59.43 µM and a selectivity index (CC50/
EC50) more than 10.99.
Experimental Section
General Experimental Procedures. Both 1D and 2D NMR
experiments were performed on a Bruker AM-400 or a DRX-500
spectrometer. Chemical shifts (δ) are expressed in ppm with reference
to the solvent signals. MS was recorded on a VG Auto Spec-3000 or
a Finnigan MAT 90 intrument. IR spectra were recorded on a Bio-Rad
FTS-135 spectrometer for KBr pellets. UV data were obtained on a
UV 210A spectrometer. Optical rotations were measured with a Horiba
SEPA-300 polarimeter or a Perkin-Elmer model 241 polarimeter.
Column chromatography was performed either on silica gel (200-300
mesh, Qingdao Marine Chemical, China), silica gel H (10-40 µm,
Qingdao Marine Chemical, China), or MCI gel CHP20P (75-150 µm,
Mitsubishi Chemical Corporation, Tokyo, Japan). Semipreparative
HPLC was performed on a Hewlett-Packard instrument (column:
Zorbax SB-C18, 250 × 9.4 mm; UV detector). Fractions were
monitored by TLC, and spots were visualized by heating silica gel plates
sprayed with 10% H2SO4 in EtOH.
Compound 1: colorless crystals from MeOH; mp 207-208 °C;
[R]18 -51.9 (c 1.0, MeOH); UV (MeOH) λmax (log ꢀ) 202 nm (2.1);
D
IR (KBr) νmax 3483, 3251, 2960, 2857, 1700, 1252 cm-1; H and 13C
1
NMR data, see Tables 1 and 2; negative ESIMS m/z 281 [M - H]-;
HRESIMS m/z 281.1755 (calcd for C16H26O4 281.1752).
Compound 2: colorless crystals from MeOH; mp 161-162 °C;
[ga]17D -43.3 (c 0.8, MeOH); UV (MeOH) λmax (log ꢀ) 202 nm (2.5);
Plant Material. The fronds of D. dichotoma were collected from
Xiaowei Mountain, Nanxi, Hekou County, Yunnan Province, P. R.
China, in March 2005. A specimen (CZ 007) was deposited in the
Herbarium of the Department of Taxonomy, Kunming Institute of
Botany, Chinese Academy of Sciences, and was identified by one of
the authors (X.C.).
1
IR (KBr) νmax 3435, 2963, 2889,1709, 1228, 991 cm-1; H and 13C
NMR data, see Tables 1 and 2; negative ESIMS m/z 279 [M - H]-;
HRESIMS m/z 279.1596 (calcd for C16H24O4 279.1596).
Compound 3: pale yellow powder; [R]18 -50.5 (c 3.2, MeOH);
D
UV (MeOH) λmax (log ꢀ) 202 nm (2.4); IR (KBr) νmax 3424, 2960,

268 Journal of Natural Products, 2007, Vol. 70, No. 2
Table 2. 13C NMR Assignments of 1-6a
Li et al.
Compound 5: pale yellow powder; [R]18 -47.8 (c 0.6, MeOH);
D
UV (MeOH) λmax (log ꢀ) 202 nm (2.0); IR (KBr) νmax 3426, 2961,
2927, 2876,1741, 1633, 1450, 1384, 1054 cm-1; 1H and 13C NMR data,
see Tables 1 and 2; negative ESIMS m/z 655 [M - H]-; HRESIMS
m/z 655.3703 (calcd for C34H56O12 655.3693).
no.
1
2
3
4
5
6
1
2
3
4
5
6
7
8
19.4 t
27.2 t
18.7 t
29.0 t
18.1 t
27.0 t
18.1 t
27.0 t
17.9 t
27.0 t
17.9 t
27.0 t
126.0 d
147.6 s
45.0 s
74.9 d
37.2 t
36.2 d
40.5 s
47.3 d
43.8 t
157.8 d
151.4 s
45.2 s
74.1 d
36.7 t
35.6 d
40.4 s
46.7 d
43.6 t
123.1 d
143.6 s
44.2 s
86.5 d
35..3 t
34.4 d
38.1 s
46.0 d
32.7 t
123.0 d
143.6 s
44.2 s
86.4 d
35.3 t
34.4 d
38.1 s
46.1 d
32.7 t
122.9 d
143.6 s
44.1 s
86.3 d
35.2 t
34.3 d
38.4 s
45.9 d
37.2 t
123.0 d
143.6 s
44.0 s
86.2 d
35.2 t
34.3 d
38.6 s
45.9 d
37.2 t
Compound 6: pale yellow powder; [R]18 -62.1 (c 0.7, MeOH);
D
UV (MeOH) λmax (log ꢀ) 203 nm (2.7); IR (KBr) νmax 3419, 2961,
2934, 2877, 1636, 1450, 1384, 1058 cm-1; 1H and 13C NMR data, see
Tables 1 and 2; negative ESIMS m/z 613 [M - H]-; HRESIMS m/z
613.3578 (calcd for C32H54O11 613.3587).
9
Acid Hydrolysis of 3, 4, 5, and 6. A solution of 3 (10 mg) in 2 M
HCl (3 mL) was heated in a water bath at 70 °C for 6 h. After cooling,
the reaction mixture was neutralized with NaHCO3 and extracted with
CHCl3. Through TLC comparison with an authentic sample using
CHCl3-MeOH (8:2) as a developing system, D-glucose and L-rhamnose
were detected in the water layer (Rf ) 0.16 and 0.43, respectively).
The aqueous solution was further concentrated to dryness and subjected
to silica gel chromatography eluting with CHCl3-MeOH (9:1) to give
10
11
12
13
14
15
16
17
18
19
20
Rha
1
2
3
4
5
6
Glc
1
2
3
174.4 s
174.3 s
35.9 t
35.9 t
25.6 t
25.6 t
72.6 s
147.2 d
111.5 t
28.6 q
16.0 q
23.0 q
16.1 q
18.4 q
72.6 s
147.3 d
111.4 t
28.7 q
16.0 q
22.9 q
16.4 q
18.3 q
138.0 s
126.6 d
58.7 t
23.8 q
15.9 q
23.0 q
16.3 q
18.0 q
140.0 s
126.6 d
58.7 t
23.4 q
15.9 q
22.9 q
16.3 q
18.0 q
16.4 q
66.3 t
17.0 q
17.5 q
16.3 q
198.2 d
16.8 q
17.0 q
D-glucose (2 mg), [R]18 +40 (c 0.2, MeOH), and L-rhamnose (1.3
D
mg), [R]18D +10 (c 0.15, MeOH). The aglycone of 3 was not obtained,
because the TLC of the CHCl3 part indicated that there were at least
four products, and was not subjected to further isolation and identifica-
tion due to the limited amount. Acid hydrolysis of 4, 5, and 6 by the
same method used for 3 led to decomposition of the aglycones, but
gave D-glucose and L-rhamnose. The two monosaccharides were
identified by comparison of their Rf and specific rotation with those of
authentic samples.
103.4 d
72.3 d
72.8 d
85.2 d
68.3 d
18.2 q
103.3 d
72.7 d
72.8 d
84.9 d
68.3 d
18.2 q
103.4 d
72.2 d
72.8 d
85.2 d
68.3 d
18.1 q
103.3 d
72.2 d
72.8 d
85.2 d
68.4 d
18.2 q
Anti-HIV-1 Assay. Cytotoxicity against C8166 cells (CC50) was
assessed using the MTT method, and anti-HIV-1 activity was evaluated
by the inhibition assay for the cytopathic effects of HIV-1 (EC50).11
106.6 d
76.4 d
78.4 d
71.6 d
75.3 d
64.6 t
106.5 d
76.0 d
76.6 d
72.3 d
76.0 d
62.2 t
106.7 d
76.4 d
78.4 d
71.6 d
75.4 d
64.6 t
106.9 d
76.6 d
78.6 d
71.5 d
78.6 d
62.7 t
4
5
6
Ac
CH3
CO
Acknowledgment. The work was supported by grants to Q.-S.Z.
from project P-06-4 of State Key Laboratory of Phytochemistry and
Plant Resources in West China, Kunming Institute of Botany, Chinese
Academy of Sciences, and to Y.-T.Z. from Key Scientific and
Technological Projects of China (2004BA719A14) and Yunnan Prov-
ince (2004NG12).
20.8 q
170.9 s
21.1 q
170.5 s
20.8 q
170.9 s
a Spectra were recorded in C5D5N; chemical shifts (δ) are in ppm.
References and Notes
Table 3. Cytotoxicity and Anti-HIV-1 Activity of Compounds
1-8
(1) Kishimoto, Y. Yakugaku Zasshi 1955, 75, 1437-1439.
(2) Diraviam, P. R.; Visuvasam, S. M.; Alexis, J. B.; Subarayan, G.;
Toshiyuki, U.; Masako, S.; Akinobu, T.; Hiroyuki, F.; Nobutoshi,
T. Chem. Pharm. Bull. 1995, 43, 1800-1803.
cytotoxicity,
CC50 (µM)
anti-HIV-1 activity,
selectivity index,
CC50/EC50
compound
EC50 (µM)
(3) Kuraishi, T.; Mitadera, Y.; Murakami, T.; Tanaka, N.; Saiki, Y.; Chen,
C. M. Yakugaku Zasshi 1983, 103, 679-682.
1
2
3
4
708.52
713.37
120.93
304.51
135.7
301.51
325.30
110.29
145.40
76.31
>2.34
>2.19
(4) Yoshiki, K.; Masafumi, M.; Gen-ichiro, N.; Itsuo, N. Chem. Pharm.
Bull. 1990, 38, 856-860.
1.10
>2.09
1.78
(5) Tadashi, A.; Tadashi, O.; Yoshikazu, H.; Takayuki, S.; Mihoko, U.;
Shinji, O. Phytochemistry 1997, 46, 839-844.
5
6
7
8
AZT
325.33
750.81
653.04
5746.1
85.06
197.01
59.43
>3.82
(6) Li, X. L.; Cheng, X.; Yang, L. M.; Wang, R. R.; Zheng, Y. T.; Xiao,
W. L.; Zhao, Y.; Xu, G.; Lu, Y.; Chang, Y.; Zheng, Q. T.; Zhao, Q.
S.; Sun, H. D. Org. Lett. 2006, 8, 1937-1940.
>3.81
>10.99
390 406.06
0.0147
(7) Pinto, A. C.; Epifanio, R. A.; Zocher, D. H. T. Biochem. Syst. Ecol.
2004, 32, 597-561.
(8) Wada, H.; Nakamura, H.; Toyoshima, Y.; Hakamatsuka, T.; Tanaka,
N.; Cambie, R. C.; Braggins, J. E. Aust. J. Chem. 1998, 51, 171-
173.
2924, 2877,1731, 1642, 1451, 1384, 1053 cm-1; 1H and 13C NMR data,
see Tables 1 and 2; negative ESIMS m/z 655 [M - H]-; HRESIMS
m/z 655.3691 (calcd for C34H56O12 655.3693).
(9) Hua, Y.; Liu, H. Y.; Ni, W.; Chen, C. X.; Lu, Y.; Wang, C.; Zheng,
Q. T. J. Nat. Prod. 2003, 66, 898-900.
(10) Pawar, R. S.; Bhutani, K. K. J. Nat. Prod. 2004, 67, 668-671.
(11) Wang, J. H.; Tam, S. C.; Huang, H.; Ouyang, D. Y.; Wang, Y. Y.;
Zheng, Y. T. Biochem. Biophys. Res. Commun. 2004, 317, 965-971.
Compound 4: pale yellow powder; [R]18 -16.2 (c 0.6, MeOH);
D
UV (MeOH) λmax (log ꢀ) 202 nm (2.2); IR (KBr) νmax 3441, 2959,
2922, 2852,1728, 1634, 1451, 1384, 1059 cm-1; 1H and 13C NMR data,
see Tables 1 and 2; negative ESIMS m/z 655 [M - H]-; HRESIMS
m/z 655.3700 (calcd for C34H56O12 655.3693).
NP0603166