Notes
J ournal of Natural Products, 2001, Vol. 64, No. 2 221
Ta ble 1. NMR Data of Compound 3 (CD3OD)
MTP A Ester s of th e Com p ou n d 3a . Compound 3a (2 mg)
was esterified with (-)-R-methoxy-R-(trifluoromethyl)phenyl-
acetyl chloride (5 µL) in dry pyridine (0.2 mL) for 2 h at room
1
position
δH (mult., J in Hz)
δC
HMBC ( H to C)
1
1
2
3
3
4
5
6
7
7
8
9
0
1
2
3
4
5
6
7
8
9
a
b
3.80 (dd, 6.6-9.5)
3.90 (dd, 6.6-9.5)
1.82 (m)
76.4
2, 16
2, 16
temperature to give, after removal of the solvent, the 1,18-di-
1
(
-)-MTPA ester: H NMR (CDCl
3
) δ 4.08 (1H, dd, J ) 10.3
36.8
37.0
and 6.6 Hz, H-1a) and 4.23 (1H, dd, J ) 10.3 and 6.0 Hz, H-1b),
.83 (1H, m, H-2), 1.14 (1H, m, H-3a), 1.29 (1H, overlapped,
H-3b), 1.35 (1H, overlapped, H-6), 1.68 (1H, m, H-10), 4.19-
1H, dd, J ) 10.3 and 5.88 Hz, H-18a), 4.27 (1H, dd, J ) 10.3
and 5.88 Hz, H-18b), 1.12 (2H, m, H-13), 1.50 (1H, m, H-14),
a
b
1.19 (m)
1
a
1.46
a
1.35-1.40
27.6
41.1
36.6
41.1
(
1.32
1.45
17
5, 7, 17
a
0
0
.90 (3H, d, J ) 6.6 Hz, H-16), 0.84 (3H, d, J ) 6.6 Hz, H-15),
.81 (3H, d, J ) 6.6 Hz, H-17), 0.84 (3H, d, J ) 6.6 Hz, H-19).
The 1,18-di-(+)-MTPA ester of 3a was prepared using (+)-
a
b
1.14 (m)
1.34 (m)
17
a
1.35-1.40
27.6
34.7
41.7
34.7
27.6
43.0
31.3
25.3
19.5
22.4
73.9
25.3
a,b
R-methoxy-R-(trifluoromethyl)phenylacetyl chloride (5 µL) in
an identical manner. H NMR (CDCl ) values were identical
3
1.32
1.68
1.40
7a, 7b, 11
9, 11
9
1
1
1
1
1
1
1
1
1
1
1
a,b
with those reported for the (-)-MTPA diester except for the
signal of the C-1 protons: δ 4.15 (2H, d, J ) 6.3 Hz, H-1).
Cell Cu ltu r es a n d Biologica l Activity. WEHI 164 cells
(murine fibrosarcoma cell line) were maintained in adhesion
on Petri dishes with Dulbecco’s Modified Eagle’s Medium
a
1.35-1.40
1.21 (m)
1.57 (m)
11, 12, 14, 15, 19
15, 19
13, 14, 19
1, 2
5, 6, 7
9, 10, 11
13, 14, 15
0.91 (3H, d, 6.3)
0.98 (3H, d, 6.3)
0.89 (3H, d, 6.3)
3.94 (3H, d, 6.0)
0.91 (3H, d, 6.3)
(DMEM) supplemented with 10% (v/v) heat-inactivated fetal
bovine serum (FBS), 25 mM HEPES, penicillin (100 U/mL),
and streptomycin (100 µg/mL).
All reagents for cell culture were from Biowhittaker. MTT
[3-(4,5-dimethylthiazol-2-yl)-2,5-phenyl-2H-tetrazolium bro-
mide] and 6-mercaptopurine were from Sigma.
a
Signals overlapped by other resonances. b Values with the
same superscript may be interchanged.
3
WEHI 164 (3.5 × 10 ) cells were plated on 96-well plates in
Com p ou n d 5: colorless amorphous solid; IR (KBr) νmax
2
50 µL and allowed to adhere at 37 °C in 5% CO /95% air for 2
-
1 1
1
(
0
3
110, 1240 cm ; H NMR δ 4.00 (2H, t, J ) 6 Hz, H-1), 1.68
h. Thereafter, 50 µL of a 1:4 v/v serial dilution of the test
compounds 1-5 were added, and then the cells were incubated
for 96 h. In some experiments 6-mercaptopurine (6-MP) was
added as standard compound for antiproliferative activity. The
cells’ viability was assessed through an MTT conversion assay
as previously described.3 The viability of each cell line in
response to treatment with compounds 1-5 and 6-MP was
calculated as % dead cells ) 100 - (OD treated/OD control) ×
100. The results are expressed as IC50 (the concentration that
inhibited the cell growth by 50%). Statistical analysis was
made by paired two-tailed Student’s t-test: The level of
statistically significant difference was defined as P < 0.01.
2H, m, H-2), 1.45 (2H, m, H-3) 1.37 (H-4), 1.36 (2H, m, H-5),
.98 (3H, t, J ) 6.7 Hz, H-6); 13C NMR (CD
3.7 (C-2), 26.8 (C-3), 24.5 (C-4), 20.5 (C-5), and 13.5 (C-6);
OD) δ 66.0 (C-1),
3
negative FABMS m/z 181; HRFABMS m/z 181.0520 [M -
-
6 13 4
Na(K)] , C H O S requires 181.0535.
Solvolysis of Com p ou n d s 1-3. Compounds 1 (1.5 mg), 2
1.5 mg), and 3 (5 mg) were desulfated using the same
(
procedure. Each compound was dissolved in a dioxane-
pyridine mixture (1:1, 3 mL) and heated at 130 °C (3 h). H
2
O
(5 mL) was added to the cooled solution before extraction with
CHCl
3
(3 × 4 mL). The organic phase was evaporated in vacuo
to give the respective alcohols 1a (0.8 mg), 2a (1 mg), and 3a
Ack n ow led gm en t. This work is the result of a research
supported by “MURST- PRIN Chimica dei Composti Organici
di Interesse Biologico”, Italy. We wish to thank Prof. Angelo
Tursi (Istituto di Zoologia ed Anatomia Comparata, Universita`
di Bari, Italy) for identifying the organism. Mass, IR, and NMR
experiments were performed at the “Centro di Ricerca Inter-
dipartimentale di Analisi Strumentale”, Universit a` di Napoli
(
(
(
(
4 mg).
Com p ou n d 1a : EIMS m/z 256; H NMR (CD
2H, t, J ) 6 Hz, H-1), 0.94 (3H, t, J ) 6.7 Hz, H-17).
1
3
OD) δ 3.63
1
Com p ou n d 2a : EIMS m/z 270; H NMR (CD
3
OD) δ 3.63
2H, t, J ) 6 Hz, H-1), 0.94 (3H, t, J ) 6.7 Hz, H-18).
1
Com p ou n d 3a : EIMS m/z 300; H NMR (CDCl
1H, dd, J ) 6.6 and 10.3 Hz, H-1a), 3.50 (1H, dd, J ) 5.9 and
0.3 Hz, H-1b), 1.61 (1H, overlapped, H-2), 1.09 (1H, over-
3
) δ 3.42
“Federico II”.
1
lapped, H-3a), 1.37 (1H, overlapped, H-3b), 1.28 (2H, over-
lapped, H-4), 1.29 (1H, overlapped, H-5a), 1.09 (1H, over-
lapped, H-5b), 1.39 (1H, m, H-6), 1.31 (2H, overlapped, H-9 or
H-11), 1.25 (2H, overlapped, H-9 or H-11), 1.46 (1H, m, H-10),
Refer en ces a n d Notes
(
(
1) Davidson, B. S. Chem. Rev. 1993, 93, 1771-1791.
2) Aiello, A.; Fattorusso, E.; Menna, M.; Carnuccio, R.; D’Acquisto, F.
Tetrahedron 1997, 53, 5877-5882.
1
.16 (2H, m, H-13), 1.54 (1H, m, H-14), 0.87 (3H, d, J ) 6.6
(3) Aiello, A.; Fattorusso, E.; Menna, M.; Carnuccio, R.; Iuvone, T.
Tetrahedron 1997, 53, 11489-11492.
4) Breitmaier, E.; Woelter, W. In Carbon-13 NMR Spectroscopy, 3rd ed.;
Hz, H-15), 0.91 (3H, d, J ) 6.6 Hz, H-16), 0.84 (3H, d, J ) 6.6
Hz, H-17), 3.54 (2H, d, J ) 5.15 Hz, H-18) and 0.87 (3H, d, J
)
3
(
VCH: Weinheim, 1987.
1
3
6.6 Hz, H-19); C NMR (CDCl
3
) δ 68.5 (C-1), 35.7 (C-2),
(5) Ohtani, I.; Kusumi, T.; Kasham, Y.; Kakisawa, H. J . Am. Chem. Soc.
1991, 113, 4092-4096.
3.2 (C-3), 24.3 (C-4), 37.3 (C-5), 32.5 (C-6), 37.3 (C-7), 24.3
(
6) De Riccardis, F.; Minale, L.; Riccio, R.; Giovannitti, B.; Iorizzi, M.;
(C-8), 31.2 (C-9), 40.6 (C-10), 31.2 (C-11), 24.3 (C-12), 39.1
(C-13), 27.8 (C-14), 22.6 (C-15), 16.4 (C-16), 19.5 (C-17), 65.7
(C-18), 22.6 (C-19).
Debitus, C. Gazz. Chim. Ital. 1993, 123, 79-86.
NP000438C