New diterpene glycosides of the fungus Acremonium striatisporum isolated from a Sea Cucumber

Article abstract of DOI:10.1021/np010503y

Three new diterpene glycosides, virescenosides O (1), P (2), and Q (3), have been isolated from a marine strain of Acremonium striatisporum KMM 4401 associated with the holothurian Eupentactafraudatrix. Their structures were determined on the basis of HRMALDIMS and NMR data as β-D-altropyranosido-19-isopimara-8(14),15-diene-7α,3β-diol (1), β-D-altropyranosido-19-7-oxoisopimara-8(9),15-diene-3β-ol (2), and β-D-mannopyranosido-19-isopimara-7,15-diene-3β-ol (3). The cytotoxic activity of the virescenosides was examined.

Full text of DOI:10.1021/np010503y

J . Nat. Prod. 2002, 65, 641-644
641
New Diter p en e Glycosid es of th e F u n gu s Acr em on iu m str ia tispor u m Isola ted
fr om a Sea Cu cu m ber
Shamil Sh. Afiyatullov, Anatoly I. Kalinovsky, Tatyana A. Kuznetsova,* Vladimir V. Isakov, Mikhail V. Pivkin,
Pavel S. Dmitrenok, and George B. Elyakov
Pacific Institute of Bioorganic Chemistry, Far East Branch of the Russian Academy of Sciences,
Vladivostok 22, Russian Federation
Received October 12, 2001
Three new diterpene glycosides, virescenosides O (1), P (2), and Q (3), have been isolated from a marine
strain of Acremonium striatisporum KMM 4401 associated with the holothurian Eupentacta fraudatrix.
Their structures were determined on the basis of HRMALDIMS and NMR data as â-D-altropyranosido-
1
9-isopimara-8(14),15-diene-7R,3â-diol (1), â-D-altropyranosido-19-7-oxoisopimara-8(9),15-diene-3â-ol (2),
and â-D-mannopyranosido-19-isopimara-7,15-diene-3â-ol (3). The cytotoxic activity of the virescenosides
was examined.
In our search for secondary metabolites from marine
fungi with cytotoxity and/or novel chemical structures, we
have previously isolated two new diterpene altrosides,
virescenosides M and N, from a marine strain of Acremo-
nium striatisporum originally separated from the holothu-
rian Eupentacta fraudatrix.1 Further investigation for
metabolites of this fungal strain has now led to the isolation
of three new cytotoxic glycosides, virescenosides O, P, and
Q (1, 2, and 3). We report herein the isolation and
structures of compounds 1-3 and their cytotoxic activity.
phase and normal-phase HPLC to yield individual glyco-
sides 1-3. The structures of new compounds 1-3 were
established by the interpretation of spectral data (NMR
and HRMALDIMS), as well as by comparison of their
spectra with those of related compounds.
The molecular formula of virescenoside O (1) was deter-
13
mined as C26
H
42
O
8
on the basis of HRMALDIMS and
C
1
13
NMR spectra. A close inspection of the H and C NMR
1
13
spectral data (Table 1) of 1 by DEPT and H- C 2D NMR
shift-correlated measurement (HMQC) revealed the pres-
ence of three quaternary methyls (δ 26.1, C-17; 23.9, C-18;
1
5.5, C-20); seven methylenes (δ 37.8, C-1; 28.8, C-2; 31.1,
1
C-6; 18.8, C-11; 34.4, C-12; 72.3, C-19; 63.5, C-6 ), including
two oxygen-bearing ones; seven oxygenated methines (δ
1
1
1
7
9.3, C-3; 72.6, C-7; 101.5, C-1 ; 71.7, C-2 ; 71.9, C-3 ; 67.0,
1
1
C-4 ; 77.1, C-5 ), including one methine linked to an
anomeric carbon; two tertiary (δ 47.8, C-5; 45.9, C-9) and
three C-C-bonded saturated quaternary carbons (δ 42.8,
C-4; 38.6, C-10; 37.5, C-13), and one monosubstituted
(148.9, 110.0) and one trisubstituted (140.6, 132.3) double
bond.
1
1
The correlations observed in the H- H COSY and
HMQC spectra of 1 and double resonance experiments
indicated the presence of the following isolated spin
systems: -CH
CHOH- (C-5-C-7), -CHdCH
C-19). Furthermore, an additional spin system involved
one anomeric proton, four oxymethine ones, and protons
2
-CH
2
-CHOH- (C-1-C-3), >CH-CH
2
-
2
(C-15-C-16), -CH -O-
2
(
1
1
1
1
of a hydroxymethyl group (C-1 -C-6 ). The H- H COSY
spectrum of 1 contained a cross-peak attributed to long-
range coupling between the olefinic proton at δ 5.57 (H-
1
4) and a carbinyl proton at δ 4.39 (H-7). In addition, H-14
was allylically coupled to a signal at δ 2.40 (H-9), which
was further correlated with two signals at δ 1.60 (H-11R)
and 1.45 (H-11â). This information together with the data
1
13
obtained from the H- C HMBC spectrum (Table 1)
indicated that 1 was a diterpene monoside with a tricyclic
aglycon structure.
Resu lts a n d Discu ssion
A direct comparison of 1H and 13C NMR spectra of 1 with
1
The fungus was cultured for 21 days on rice medium
specially modified by us.1 The CHCl
-MeOH (2:1, v/v)
extract of the culture of A. striatisporum was fractionated
by Si gel column chromatography followed by reversed-
those of the glycosides M and N , and virescenosides A-C,
obtained earlier from the terrestrial fungus Acremonium
3
luzulae2 suggests that virescenoside O has the structure
-5
1
3
of an isopimaradienic altroside. The C NMR spectrum of
contained signals for a monosubstituted double bond at
δ 148.9 (d) and 110.0 (t). The proton signals of a typical
1
*
Corresponding author. Fax: 7-42320314050. E-mail: piboc@stl.ru.
1
0.1021/np010503y CCC: $22.00
© 2002 American Chemical Society and American Society of Pharmacognosy
Published on Web 04/02/2002

6
42 J ournal of Natural Products, 2002, Vol. 65, No. 5
Afiyatullov et al.
Ta ble 1. 1H and 13C NMR Data of Virescenoside O (1) in C5D5N (J , Hz)
1
1
atom
1
δC
δH
H- H COSY
HMBC (C)
NOESY (H)
37.8 t
1R: 1.23 m
â: 1.73 m
2R: 2.02 m
â: 1.95 m
1â, 2R,â
1R, 2R,â
1R,â, 2â, 3
1R,â, 2R, 3
2R,â
3, 9
11â, 20
1
2
28.8 t
2
19a, 20
1R, 5, 18
3
4
5
6
79.3 d
42.8 s
47.8 d
31.1 t
3.67 dd (4.8, 11.0)
2.16 dd (2.3, 12.8)
6R,â
3, 9, 18
18
19b, 20
14
6R: 2.23dt (2.3, 13.0)
5, 6â, 7
5, 6R, 7
6R,â, 14
6
â: 1.91 td (3.4, 12.8)
7
8
9
72.6 d
140.6 s
45.9 d
38.6 s
18.8 t
4.39 t (3.4)
2.40 m
11R,â
1R, 5
1
1
0
1
11R: 1.60 m
1â: 1.45 m
9, 11â, 12R,â
9, 11R, 12R,â
11R,â
1
1
1
1
1
1
2
3
4
5
6
34.4 t
37.5 s
132.3 d
148.9 d
110.0 t
1.45 m
5.57 d (1.9)
5.81 dd (10.7, 17.5)
16a: 4.94 dd (1.5, 10.7)
7, 9
7, 17
17
16a,b
15, 16b
15, 16a
13
17
1
6b: 5.03 dd (1.5, 17.5)
1
1
1
7
8
9
26.1 q
23.9 q
72.3 t
1.09 s
1.57 s
19a: 4.24 d (10.0)
12, 13, 14, 15
3, 4, 5, 19
14, 15, 16b
3, 5, 6R, 19a,b
1
19b
21
1 , 2â,18, 20
1
9b: 4.49 d (10.0)
6â,18, 20
2
1
2
3
4
5
6
0
15.5 q
101.5 d
71.7 d
71.9 d
67.0 d
77.1 d
63.5 t
0.88 s
1, 5, 9, 10
1â, 2â, 11â, 19a,b
19a,b
1
5.54 d (1.7)
4.63 dd (1.7, 4.9)
4.76 dd (3.3, 4.9)
4.82 dd (3.3, 8.1)
4.57 m
1
1
1
1
1
1
1
1
1
1
1 , 3
1
2 , 4
1
3 , 5
1
4 , 6 a,b
1
1
1
41
6 a: 4.40 dd (5.3, 11.4)
5 , 6 b
1
1
1
6
b: 4.51 dd (3.7, 11.4)
5 , 6 a
ABX system of a vinyl group at δ 5.81 (1H, dd, 10.7, 17.5,
Hz), 5.03 (1H, dd, 1.5, 17.5, Hz), and 4.94 (1H, dd, 1.5, 10.7,
Hz) in the spectrum of 1 indicated the C-15, C-16 position
of this double bond.6 Furthermore, the position of the
C-17 methyl group (δ 1.09, s) and of the exo-vinyl group at
between H-2â and H -20 as well as between H-3R and H-5R
3
indicated a trans ring fusion between rings A and B. The
NOESY spectrum exhibited the cross-peaks H-1R/H-9R and
H-5R/H-9R, indicating these protons to be on the same side
of the molecule. NOEs were also observed between H-3R
-9
1
13
C-13 were argued by H- C HMBC and NOE measure-
ments (Table 1). The stereochemistry at C-13 in 1 was
assigned to be the same as sandaracopimaradienic deriva-
tives on the basis of the similarity of C-15-C-17 chemical
shifts for these compounds.1
and H
consistent with a ∆
3
-18 and between H-7â and H-14. All these data are
8
(14),15
-sandaracopimaradienic skeleton
with an equatorial hydroxyl group at C-3 and an axial
hydroxymethyl and hydroxyl groups at C-4 and C-7 for the
0-13
1
aglycon part of 1. The strong NOEs from H-1 to H-19a
Localization of the trisubstituted double bond (δ 140.6,
s, 132.3, d) at the C-8, C-14 position was evident from the
indicated that the sugar moiety was linked at C-19.
A comparison of the 13C NMR spectrum of 1 with the
data published for R- and â-D-altropyranoses as well as a
good coincidence of carbon signals due to the glycosidic
moiety with those of virescenosides A, M, and N together
1
1
H- H COSY and HBMC correlations. A direct comparison
1
3
of C NMR shifts of 1 with the values published for 7R-
hydroxysandaracopimar-8(14),15-dienic derivatives1
3-15
con-
1
1
1
firmed this deduction. The small coupling constants of the
H-7 signal at δ 4.39 (1H, t, 3.4) indicated that 1 contained
an allylic secondary alcohol function with an axial config-
with magnitudes of H- H spin coupling constants in H
NMR spectra of 11
,5,16-18
elucidated the presence of a â-D-
altropyranoside unit of C1 form in 1. Acid hydrolysis of
virescenoside O gave D-altropyranose and 1,6-anhydro-â-
D-altropyranose, which were identified by NMR spectra and
optical rotation. On the basis of all the data above, the
structure of virescenoside O was established as â-D-
altropyranosido-19-sandaracopimara-8(14),15-diene-3â,7R-
diol.
uration. Furthermore, in comparison with the spectra of
sandaracopimaric acid1
0,12
the shift of the C-9 signal from
δ 50.7 to 45.9 may be explained by the γ-effect of an axial
hydroxyl at C-7. The COSY and HMBC data allowed the
assignment of the signal at δ 79.3 (C-3) to a secondary
3
hydroxyl-bearing carbon, adjacent to a quaternary sp C
carbon. The relative stereochemistry of the proton at C-3
was defined on the basis of the H-H coupling constants
In HRMALDIMS virescenoside P (2) gave a quasimo-
lecular ion at m/z 503.2630 [M + Na]. These data, coupled
1
1
1
3
(
J ) 4.8, 11.0 Hz) observed between H-3 and H-2R,â and
with C NMR spectral data (DEPT), established the
molecular formula of 2 as C26 . The general features
assigned as axial.
40 8
H O
1
1
13
The H NMR spectrum of 1 showed two signals corre-
of the UV and H and C NMR spectra of 2 (Table 2 and
sponding to an AB system coupling at δ 4.24 and 4.49 (each
the Experimental Section) closely resembled those of
virescenoside M with the exception of proton and carbon
1
1
H, each d, 10.0 Hz), which were consistent with the
presence of CH
O- group linked to a quaternary sp3
carbon. The position and stereochemistry of the methyl
1.57, s) and hydroxymethyl (72.3, t) groups at C-4 and
2
C
signals belonging to the ring A.
1
1
Correlations observed in the H- H COSY and HMQC
spectra and double resonance experiments on 2 indicated
the presence of an isolated spin system corresponding to
(
methyl group (0.88, s) at C-10 were established on the basis
of NOEs and HBMC data. Furthermore, NOE correlations
the sequence -CH
2
-CH -CHOH- (C-1-C-3). Thus, this
2

Diterpene Glycosides of the Fungus Acremonium
J ournal of Natural Products, 2002, Vol. 65, No. 5 643
Ta ble 2. 1H and 13C NMR Data of Virescenosides P (2) and Q (3) in C5D5N (J , Hz)
2
3
atom
1
δC
δH
δC
δH
34.6 t
1R: 1.23 m
â: 1.73 m
2R: 1.91 m
â: 2.02 m
38.3 t
1R: 1.15 m
1
1â: 1.78 m
2
28.1 t
28.6 t
2R: 1.88 m
2
2â: 2.03 m
3
4
5
6
77.5 d
42.7 s
50.0 d
37.1 t
3.47 dd (4.5, 11.4)
79.0 d
42.6 s
51.3 d
24.3 t
3.57 dd (4.0, 11.9)
1.75 dd (3.3, 14.5)
6R: 2.89 dd (3.3, 18.0)
â: 3.24 dd (14.5, 18.0)
1.28 m
6R: 2.04 m
6â: 2.30 m
5.33 m
6
7
8
9
1
1
.
199.5 s
128.4 s
164.2 s
39.6 s
122.0 d
135.4 s
52.1 d
35.3 s
20.4 t
1.60 m
0
1
23.0 t
11R: 2.07 m
1â: 2.07 m
12R: 1.50 m
2â: 1.21 m
11R: 1.47 m
11â: 1.30 m
12R: 1.32 m
12â: 1.45 m
1
1
2
33.9 t
36.2 t
1
1
1
3
4
34.4 s
33.7 t
36.9 s
46.1 t
14R: 2.56 brd (17.6)
14R: 2.02 brd (15.0)
14â: 1.95 brd (15.0)
5.88 dd (10.7, 17.5)
16a: 4.96 dd (1.5, 10.7)
16b: 5.03 dd (1.5, 17.5)
0.90 s
1
4â: 2.12 brd (17.6)
1
1
5
6
145.9 d
111.4 t
5.74 dd (10.8, 17.4)
16a: 4.90 dd (1.5, 17.4)
150.4 d
109.4 t
1
6b: 4.97 dd (1.5, 10.7)
1
1
1
7
8
9
27.7 q
23.0 q
71.9 t
0.94s
1.35 s
19a: 4.16 d (10.4)
21.4 q
24.1 q
71.3 t
1.42 s
19a: 4.22 d (10.2)
19b: 4.54 d (10.2)
0.95 s
1
9b: 4.70 d (10.4)
2
1
2
3
4
5
6
0
17.5 q
101.2 d
71.9 d
72.5 d
66.7 d
76.5 d
63.5 t
1.21 s
15.6 q
102.5 d
71.7 d
75.6 d
68.9 d
78.8 d
62.8 t
1
5.45 d (1.4)
4.56 dd (1.4, 4.4)
4.74 dd (3.2, 4.4)
4.78 dd (3.2, 8.5)
4.49 m
4.90 d (0.9)
1
1
1
1
1
4.54 dd (0.9, 3.3)
4.13 dd (3.3, 9.2)
4.58 t (9.2)
3.88 ddd (2.6, 5.4, 9.2,)
1
1
6 a: 4.36 dd (6.0, 12.0)
6 a: 4.38 dd (5.4, 11.5)
1
1
6
b: 4.47 dd (3.5, 12.0)
6 b: 4.55 dd (2.6, 11.5)
metabolite has the same structure as virescenoside M
except that it possesses one less hydroxyl group. The
magnitudes of the vicinal coupling constants (4.5, 11.4 Hz)
between H-3 (δ 3.47) and H-2R,â (δ 2.02, 1.91) revealed an
equatorial configuration of the alcohol function at C-3.
ence of a â-D-mannopyranoside unit in 3. On the basis of
all the above data, the structure of virescenoside Q was
established as â-D-mannopyranosido-19-isopimara-7,15-
diene-3-â-ol.
It was shown that virescenosides O, P, and Q exhibited
cytotoxic action against tumor cells of Ehrlich carcinoma
The relative stereochemistry of 2 was defined by analysis
of NMR chemical shifts and coupling constant values and
by NOESY correlations (Table 2 and the Experimental
Section). The CD spectrum of 2a obtained upon acid
hydrolysis of 2 showed the characteristic Cotton effects at
(
IC50 ) 20-100 µM) in vitro. Virescenoside P showed
cytotoxic effects on developing eggs of the sea urchin
Strongylocentrotus intermedius (MIC50 ) 5.0 µM).
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. 1H and 13C NMR
3
24 (positive), 258 (negative), and 210 (positive) nm, which
were in good agreement with those for methyl 7-oxo-13-
epi-pimara-8,15-dien-18-oate.9 These data led us to the
conclusion that the aglycon 2a belonged to the normal 5R-
pimarane series, and the structure of virescenoside P was
established as â-D-altropyranosido-19-7-oxoisopimara-8,15-
diene-3â-ol.
spectra were recorded in both CDCl and pyridine on a Bruker
3
DPX-300 spectrometer operating at 300 and 75.4 MHz with
TMS as internal standard. HRMALDIMS analyses were
carried out with a Bruker Biflex time-of-flight mass spectrom-
eter equipped with a UV-nitrogen laser (337 nm). CD spectra
were obtained with a J ASCO model J -500. UV spectra were
recorded on a Specord UV-vis spectrometer in MeOH. GLCMS
analyses were done on a Hewlett-Packard HP6890 GG system,
with an HP-5MS capillary column (30.0 m × 250 µm × 0.25
µm) at 210 °C. Helium was used as the carrier gas, and the
ionizing voltage was 70 eV. Optical rotations were measured
by a Perkin-Elmer 141 polarimeter.
The molecular formula of virescenoside Q (3) was estab-
42 7
H O C
on the basis of HRMALDIMS and 13
lished as C26
NMR spectra. The carbon signals of the aglycon part of 3
Table 2) were very similar to those of virescenol B19 except
(
for the chemical shifts of C-3, C-18, and C-19, and accord-
ingly it is proposed that 3 has the same aglycon structure
as virescenoside B.
Cu ltiva tion of A. str ia tispor u m . The cultivation of the
fungus was performed as previously reported.1
Acid hydrolysis of virescenoside Q gave 3a , which was
identical to isovirescenol B by NMR spectra and optical
rotation. Besides 3a , acid hydrolysis of 3 gave D-mannose,
which was identified by optical rotation and by GLCMS
as the corresponding aldononitrile peracetate. These data
Extr a ction a n d Isola tion . At the end of the incubation
period, the mycelium and medium were homogenized and
thrice extracted then with a mixture of CHCl -MeOH (2:1,
3
v/v, ca. 2 L). After evaporation of the solvent, the residual
material (4 g) was passed over normal-phase silica, which was
1
1
1
1
and the magnitudes of H-1 -H-6 and C-1 -H-1 (158 Hz)
3
eluted first with CHCl (500 mL) followed by a step gradient
spin-coupling constants in the NMR spectra of 31
6-18
and
from 5% to 20% MeOH in CHCl (total volume 2 L). Fractions
3
NOE data (see Experimental Section) elucidated the pres-
of 10 mL were collected and combined by TLC examination.

6
44 J ournal of Natural Products, 2002, Vol. 65, No. 5
Afiyatullov et al.
Fractions containing the desired compounds were further
purified by reversed-phase HPLC on a Silasorb-ODS column
NH
2
OH‚HCl (1 mg) and pyridine (0.5 mL) at 100 °C for 1 h. A
2
solution obtained was heated with Ac O (0.5 mL) at 100 °C
(
10 µm, 9.6 × 200 mm, 220 nm) eluting with a step gradient
from 52% to 75% MeOH in H O and then by normal-phase
HPLC on a Zorbax SIL column (5 µm, 4.6 × 150 mm) using
CO (70:30) as eluent to yield 1 (6 mg), 2 (4.5
for 1 h and concentrated in vacuo to dryness. The aldononitrile
peracetate was analyzed by means of GLC and GLCMS.
2
2
0
Isop im a r a -7,15-d ien e-3â,19-d iol (3a ): [R] +98° (c 0.16,
D
1
13
EtOAc-(CH
mg), and 3 (4.2 mg).
3
)
2
3
CHCl ); H and C NMR spectra and optical rotation data
obtained for 3a were in agreement with published data
7
,19
for
â-D-Altr op yr a n osid o-19-sa n d a r a cop im a r a -8(14),15-d i-
isovirescenol B.
2
0
en e-3â,7r-d iol (1): colorless amorphous solid; [R]
0
HRMALDIMS m/z 505.2790 (calcd for C26
D
-44° (c
N), see Table 1;
42 8
O Na, 505.2772).
Acid ic Hyd r olysis of Vir escen osid e O (1). Acidic hy-
drolysis of compound 1 (12 mg) was performed as described
above for 2. The residue obtained after evaporation of the
1
13
.5, MeOH); H and C NMR spectra (C
5
D
5
H
â-D-Altr op yr a n osid o-19-7-oxoisop im a r a -8,15-d ien e-3â-
water layer was purified on a Zorbax NH
2
column (5 µm, 4.6
2
0
ol (2): colorless amorphous solid; [R]
D
+31° (c 0.2, MeOH);
×
150 mm) eluting with 90% AcCN to yield 1.7 mg of
1
13
UV (MeOH) λmax (log ꢀ) 248 (3.7) nm; H and C NMR spectra
N), see Table 2; HMBC correlation (H/C) H-16a,b/C-13,
1
,6-anhydro-â-D-altropyranose (altrosan) and 1.2 mg of D-
20 13
(
C
5
D
5
altrose, [R]
D
2
+32.8° (c 0.6, H O). The C NMR spectrum
C-15; Me-17/C-12, C-13, C-14, C-15; Me-18/C-3, C-4, C-5,
C-19; Me-20/C-1, C-5, C-9, C-10; NOESY correlations (H/H)
1
1
obtained for the monosaccharide was in agreement with
20
published data for D-altrose. The acetylation of altrosan with
R/3,5, 1â/11â, 2â/19b,20, 3/5,18, 5/18, 6â/19b,20, 11R/15,
20
Ac
CHCl
published data.
2
O and pyridine afforded the triacetate, [R]
D
-166° (c 0.3,
1
2R/15, 14R/16b,17, 14â/17, 15/17, 16b/17, 18/19a, 19a,b/20,1 ;
1
3
). Its
H NMR spectrum was in agreement with
HRMALDIMS m/z 503.2630 (calcd for C26
H
40
O
8
Na, 503.2616).
2
â-D-Man n opyr an osido-19-isopim ar a-7,15-dien e-3â-ol (3):
2
0
1
colorless amorphous solid; [R]
D
-20° (c 0.45, MeOH); H and
Ack n ow led gm en t. We thank Dr. N. Prokof’eva for biotest-
ing of obtained virescenosides. This work was supported by
Russian Foundation of Base Research grants N 00-15-97397
and N 00-04-48034.
1
3
C NMR spectra (C D N), see Table 2; HMBC correlation (H/
5 5
C) H-15/C-13, C-17; H-16a,b/C-13; Me-17/C-12, C-13, C-14,
C-15; Me-18/C-3, C-4, C-5, C-19; Me-20/C-1, C-5, C-9, C-10;
H-1 /C-2 ; NOESY correlation (H/H) 1R/3,9, 1â/11R, 20, 2â/
1
2
1
1
1
9a,b,20, 3/5,18, 5/9, 6R/18, 6â/19b,20, 7/14â, 9/12R, 11â/17,-
0, 12R/14R,15, 14R/15,16b, 14â/ 17, 15/17, 16b/17, 18/19a,b,
Refer en ces a n d Notes
1
1
1
1
1
1
9a,b/20,1 , 19b/6â, 1 /3 ,5 , 3 /5 ; HRMALDIMS m/z 489.2809
(1) Afiyatullov, Sh. Sh.; Kuznetsova, T. A.; Isakov, V. V.; Pivkin, M. V.;
Prokof’eva, N. G.; Elyakov, G. B. J . Nat. Prod. 2000, 63, 848-850.
2) Cagnolli-Bellavita, N.; Ceccherelli, P.; Ribaldi, M.; Polonsky, J .;
(
calcd for C26
42 7
H O Na, 489.2823).
(
Acid ic Hyd r olysis of Vir escen osid e P (2). A solution of
Baskevitch, Z. Gazz. Chim. Ital. 1969, 99, 1354-1363.
(3) Cagnolli-Bellavita, N.; Ceccherelli, P.; Mariani, R.; Polonsky, J .;
compound 2 (5 mg) in 0.1 M TFA (1 mL) was heated in a
stoppered reaction vial for 30 min. The water layer was
Baskevitch, Z. Eur. J . Biochem. 1970, 15, 356-359.
4) Cagnolli-Bellavita, N.; Ceccherelli, P.; Ribaldi, M.; Polonsky, J .;
(
extracted with CHCl
of the extract was chromatographed on a Si gel column (0.8 ×
cm), eluting first with hexane and finally with a solvent
3
. The residue obtained after evaporation
Baskevitch-Varon, Z.; Varenne, J . J . Chem. Soc., Perkin Trans. 1
1
977, 351-354.
6
(5) Bellavita, N.; Bernassau, J . M.; Ceccherelli, P.; Raju, M. S.; Wenkert,
E. J . Am. Chem. Soc. 1980, 102, 17-20.
system of hexane-ethyl acetate (60:40), to yield 1.8 mg of 2a .
(
(
6) Wenkert, E.; Beak, P. J . Am. Chem. Soc. 1961, 83, 998-1000.
7) Polonsky, J .; Baskevitch, Z.; Cagnolli-Bellavita, N.; Ceccherelli, P.
Bull. Soc. Chim. Fr. 1970, 5, 1912-1918.
7
-Oxoisop im a r a -8,15-d ien e-3â,19â-d iol (2a ): colorless
20
amorphous solid; [R]
D
+44° (c 0.15, MeOH); UV (MeOH) λmax
6
-
(
∆
3
log ꢀ) 253 (4.04) nm; CD (3.10 × 10 M, MeOH) ∆ꢀ324 +0.96,
(8) Borkosky, S.; Bardon, A.; Catalan, C. A. N.; Diaz, J . G.; Herz, W.
1
Phytochemistry 1995, 40, 1477-1479.
9) De Pascual, J . T.; Barrero, A. F.; Muriel, L.; San Feliciano, A.; Grande,
ꢀ
258 -0.15, and ∆ꢀ210 +1.64; H NMR (300 MHz, J , CDCl ) δ
3
(
.49 (1H, dd, 4.8, 11.4, H-3), 1.73 (1H, dd, 3.5, 14.4, H-5), 2.57
M. Phytochemistry 1980, 19, 1153-1156.
(10) (10) Wenkert, E.; Buckwalter, B. L. J . Am. Chem. Soc. 1972, 94,
4367-4369.
(
(
1H,dd, 3.5, 17.5, H-6R), 2.33 (1H, dd, 14.4, 17.5, H-6â), 2.36
1H, brd, 17.7, H-14R), 2.57 (1H, brd, 17.7, H-14â), 5.67 (1H.
(
11) San Feliciano, A.; Medarde, M.; Lopez, J . L.; del Corral, J . M. M.;
Puebla, P.; Barrero, A. F. Phytochemistry 1988, 27, 2241-2248.
12) Bardyshev, I. I.; Degtyarenko, A. S.; Pekhk, T. L.; Yankovskaya, G.
S. Zh. Org. Khim. 1981, 17, 2568-2573.
dd, 10.8, 17.4, H-15), 4.83 (1H, dd, 1.4, 17.4, H-16a), 4.97 (1H,
dd, 1.4, 10,8, H-16b), 3.43 (1H, d, 11.2, H-19a), 4.29 (1H, d,
1
(
1.2, H-19b), 1.02 (3H, s, H
3
-17), 1.24 (3H, s, H
3
-18), 1.06 (3H,
1
3
(13) De Kimpe, N.; Schamp, N.; van Puyvelde, L.; Dube S.; Chagnon-Dube
M.; Borremans, F.; Anteunia M. J . O.; Declercg J .-P.; Germain, G.;
van Meerssche, M. J . Org. Chem. 1982, 47, 3628-3630.
s, H
3
-20); C NMR (75.4 MHz, CDCl ) δ 33.9 (t, C-1), 27.7 (t,
3
C-2), 79.9 (d, C-3), 42.4 (s, C-4), 49.6 (s, C-5), 34.9 (t, C-6), 199.0
s, C-7), 129.1 (s, C-8), 164.5 (s, C-9), 39.2 (s, C-10), 23.2 (t,
(
(
(
(
14) Rao, Ch. B.; Suseela, K.; Raju, G. B. S. Ind. J . Chem. 1984, 23B, 177-
C-11), 33.7 (t, C-12), 34.5 (s, C-13), 33.4 (t, C-14), 145.1 (d,
C-15), 111.8 (t, C-16), 28.2 (q, C-17), 21.9 (q, C-18), 63.9 (t,
C-19), 18.4 (q, C-20); EIMS m/z 318 [M] (6), 300 (3), 170 (10),
179.
15) Touche, E. M. G.; Lopez, E. G.; Reyes, A. P.; Sanchez, H.; Honecker,
F.; Achenbach, H. Phytochemistry 1997, 45, 387-390.
16) King-Morris, M. J .; Serianni, A. S. J . Am. Chem. Soc. 1987, 109,
+
1
48 (100), 82 (99).
Acid ic Hyd r olysis of Vir escen osid e Q (3). A solution of
3
501-3508.
(17) Perlin, A. S.; Casu, B.; Koch, H. J . Can. J . Chem. 1970, 48, 2596-
606.
2
compound 3 (5 mg) in 0.1 M TFA (1 mL) was heated in a
boiling water bath for 1 h. The lipid part of the hydrolysate
was purified as described above to yield 1.2 mg of 3a . The
residue obtained after evaporation of the water layer was
purified on a Separon SGX NH
mm) eluting with 90% AcCN to yield 0.8 mg of D-mannose,
(
18) Podlasek, C. A.; Wu, J .; Stripe, W. A.; Bondo, P. B.; Serianni, A. S. J .
Am. Chem. Soc. 1995, 117, 8635-8644.
(19) Polonsky, J .; Baskevitch, Z.; Cagnoli-Bellavita, N.; Ceccherelli, P.;
Buckwalter, B. L.; Wenkert, E. J . Am. Chem. Soc. 1972, 94, 4369-
4
370.
2
column (7 µm, 3 × 150
(
20) Bock, K.; Beck, M. Acta Chem. Scand. 1980, B34, 389.
2
0
[
R]
D
+14.1° (c 0.4, H
2
O). Monosaccharide was treated with
NP010503Y