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animals survived much longer than those received 10 mg of
GTX-II (5). Therefore, acute toxicity might be milder in
order of 9a-hydroxy-GTX-II (7), 9b-hydroxy-GTX-II (11)
and dihydro GTX-II (12), 9b-hydroxy-dihydro-GTX-II (13)
and GTX-II (5). Based on these results, we can concluded
that the transformation of GTX-II to 9b-dihydro- and 9a-hy-
droxy-GTX-II resulted slight and marked reduction of acute
toxicity, respectively.
Experimental
All melting points (mps) are uncorrected. IR spectra were measured with
a Shimadzu IR-430 instrument. 1H- and 13C-NMR were measured on a
Unity-300 (Varian Co.) spectrometer in CDCl3 or pyridine-d5, using tetra-
Fig. 2. A Perspective View of the Molecule of 7
1
methylsilane (TMS) as an internal standard. The H- and 13C-NMR signals
of each derivatives were assigned DEPT, 1H–1H correlation spectroscopy
(H–H COSY), 13C–1H correlation spectroscopy (C–H COSY) and by com- H3), 1.78 (3H, s, C19-H3), 2.90 (1H, m, C7-H), 2.80 (1H, m, C2-H), 3.39 (1H,
parison with spectra of known derivatives. HR mass spectra were obtained m, C1-H), 3.89 (1H, t, Jϭ6.0 Hz, C3-H), 4.18 (1H, s, C14-H), 4.54 (1H, m,
with a Nihondenshi Co. mass spectrometer (JMS-700T). 13C-NMR data and C6-H), 5.39 and 5.62 (each 1H, d, Jϭ2.4 Hz, C20-H2).
assignment of GTX derivatives were shown in Table 1.
9b-Hydroxy-a-dihydro-GTX-II (13) A mixture of 9b-hydroxy-GTX-
9a-Hydroxy GTX-II (7) and 3b,5b,6b,9a,14b,15a-Hexahydroxy- II (11) (200 mg) and PtO2 (5 mg) in methanol (10 ml) was stirred at room
grayanotoxa-D1(10)-ene (8) Air was passed through a solution of iso-GTX- temperature for 5 h. under H2, and then filtered. The filtrate was concentrated
II (600 mg, 1.70 mmol) and Rose Bengal (2 mg) in methanol (10 ml) under under reduced pressure to leave white crystals. Which were crystallized
irradiation by UV light at 25 °C for 1.5 h. The mixture was treated with 5%- from hexane–ethyl acetate to give 9b-hydroxy-a-dihydro-GTX-II (13)
KOH soln. and active carbon, and the solvent was evaporated to give 360 mg (180 mg, 90%), mp 250—265 °C. EI-HR-MS m/z: 370.2357 (Calcd for
of products mixture, 7 and 8. The mixture was purified by silica gel (Wako- C20H34O6: 370.2355). 1H-NMR (pyridine-d5) d: 1.16 (3H, s, C18-H3), 1.47
gel C-300) column chromatography. Gradient elution with ethyl acetate from (3H, s, C17-H3), 1.66 (3H, s, C19-H3), 1.47 (3H, d, Jϭ15.9 Hz, C20-H3), 3.87
chloroform-ethyl acetate gave 7 (110 mg, 18%) and 8 (85 mg, 14%). The (1H, m, C3-H), 4.27 (1H, s, C14-H), 4.39 (1H, dd, Jϭ6.9, 9.0 Hz, C6-H).
crude 7 was recrystallized from hexane–ethyl acetate, mp 248—250 °C. EI-
X-Ray Crystallographic Analysis A crystal,9) 9a-hydroxy-GTX-II (7),
HR-MS m/z: 368.2191 (Calcd for C20H32O6: 368.2191). IR (Nujol) cmϪ1
:
used for X-ray crystallographic analysis, was obtained by slow evaporation
1
3362 (OH), 1634 (ϾCϭCH2), 1319, 1049, 1028, 927, 914; H-NMR (pyri- from an ethyl acetate solution at room temperature. X-ray diffraction data of
dine-d5) d: 1.10 (3H, s, C18-H3), 1.52 (3H, s, C19-H3), 1.61 (3H, s, C17-H3), the crystal were collected on a Rigaku AFC-5R diffractometer with graphite
2.30 (2H, m, C2-H2), 2.75 (1H, d, Jϭ15.3 Hz, C15-H), 2.91 (1H, dd, Jϭ2.7, monochromated CuKa radiation and a rotating anode generator. Of the 1739
15.6 Hz, C7-H), 3.16 (1H, dd, Jϭ6.0, 15.6 Hz, C7-H), 3.95 (1H, m, C3-H), reflections which were collected, 1716 were unique (Rintϭ0.000). The inten-
4.11 (1H, t, Jϭ9.6 Hz, C1-H), 4.71 (1H, m, C6-H), 5.13 (1H, d, Jϭ7.5 Hz, sities of three representative reflections were measured after every 150 re-
C14-H), 5.48 (2H, d, Jϭ6.6 Hz, C20-H2). 13C-NMR data of 7 was shown in
flections. The structure was solved by direct methods using the SHELX97
Table 1. The other product, 8, was recrystallized from hexane–ethyl acetate, program,10) and all the computations were carried out on the teXan crystallo-
mp 136—138 °C; IR (Nujol) cmϪ1: 3360 (OH), 1221, 1153, 1072, 1045,
graphic soft ware package.11) Final R-factors were Rϭ0.056, RWϭ0.111. The
997, 993. EI-HR-MS m/z: 368.2189 (Calcd for C20H32O6: 368.2191). 1H- atomic scattering factors used for non-hydrogen atoms were taken from the
NMR (pyridine-d5) d: 1.13 (3H, s, C18-H3), 1.48 (3H, s, C19-H3), 1.79 (3H, s, International Table12) and for hydrogen atoms from a reference.13)
C17-H3), 1.94 (3H, s, C20-H3), 2.97 (2H, dd, Jϭ3.3, 17.7 Hz, C2-H2), 3.00
(1H, m, C7-H), 3.44 (1H, dd, Jϭ7.2, 14.4 Hz, C7-H), 3.93 (1H, d, Jϭ4.5 Hz, References and Notes
C3-H), 4.81 (1H, dd, Jϭ7.2, 9.9 Hz, C6-H), 4.87 (1H, s, C14-H).
1) Miyajima S., Takei I., J. Agric. Chem. Jpn., 10, 1093—1103 (1934).
2) Kakisawa H., Kozima T., Yanai M., Nakanishi K., Tetrahedron, 21,
3091—3104 (1965).
3) Iwasa J., Kumazawa Z., Nakajima M., Agric. Biol. Chem., 25, 782—
801 (1961).
4) Hikino H., Ogura M., Ohta T., Takemoto T., Chem. Pharm. Bull., 18,
1071—1073 (1970).
5) Matsutani T., Seyama I., Narahashi T., Iwasa J., Pharmacol. Exp.
Ther., 217, 812—819 (1981).
6) Kaiya T., Sakakibara J., Nagoyashiritsudaigaku Kenkyuunenpo, 30,
1—34 (1982).
7) Katai M., Fujiwara M., Terai T., Meguri H., Chem. Pharm. Bull., 28,
3124—3126 (1980).
8) Terai T., Nippon Kagaku Kaishi, 1985, 1728—1733 (1985).
9) Crystal data: Mrϭ368.47, Monoclinic, aϭ13.326(3), bϭ18.431(3),
cϭ7.637(3) Å, Vϭ1875.7(7) Å3, Space group P212121, zϭ4, Dcϭ
1.305 g/cm3, m(CuKa)ϭ7.79 cmϪ1, l(CuKa)ϭ1.54178 Å, F(000)ϭ
800.00.
9b-Hydroxy-GTX-II-3,6,14,16-tetra-O-acetate (10) To a solution of
GTX-II-tetraacetate (9) (500 mg) in 10 ml dioxane and 1 ml water, SeO2
(40 mg) and 36%-H2O2 (1 ml) were added, and the solution was stirred at
room temperature for 1 week. The mixture was concentrated and extracted
with ethyl acetate. The combined ethyl acetate layer was dried over anhy-
drous Na2SO4 and evaporated to give oily products mixture. The products
mixture was purified by silica gel (Wakogel C-300) column chromatography.
Elution with hexane–ethyl acetate (4 : 3) and crystallization gave 160 mg
(31%) of 9b-hydroxy-GTX-II-tetraacetate (10), mp 211—212 °C. IR (Nujol)
cmϪ1: 3467 (OH), 1744 (CϭO), 1701 (CϭO), 1256 (–O–CO), 1034, 965.
1H-NMR (pyridine-d5) d: 1.07 (3H, s, C18-H3), 1.40 (3H, s, C19-H3), 2.00
(3H, s, C17-H3), 1.66, 1.99, 2.03, 2.40 (each 3H, s, –COCH3ϫ4), 2.10 (1H,
m, C7-H), 2.70 (C7-H), 2.30 (1H, m, C2-H), 2.90 (1H, m, C2-H), 3.18 (1H,
m, C13-H), 3.34 (1H, m, C1-H), 5.06 (1H, dd, Jϭ5.1, 7.5 Hz, C3-H), 5.15
(1H, dd, Jϭ3.9, 11.6 Hz, C6-H), 5.22 and 5.47 (each 1H, d, Jϭ2.1, C20-H2),
5.36 (1H, m, C14-H).
9b-Hydroxy-GTX-II (11) To
a solution of the tetraacetate (10)
(350 mg) in methanol (10 ml), 5%-KOH soln. (5 ml) was added, and the so-
10) Sheldric G. M., “Program for the Solution of Crystal Structures,” Uni-
versity of Gottingen, Germany, 1997.
lution was stirred at room temperature for 24 h. The mixture was neutralized
with dil. HCl and extracted with ethyl acetate. The combined ethyl acetate 11) teXan: Crystal Structure Analysis Package, Molecular Structure Cor-
layers were washed with water, and dried over anhydrous Na2SO4 and evapo-
poration (1979 and 1999).
rated to give crude 11 (205 mg, 85%). The product, 11, was recrystallized 12) “International Tables for X-ray Crystallography,” Vol. IV, The Kynoch
from ethyl acetate, mp 215—216 °C. IR (Nujol) cmϪ1: 3200 (OH), 1323,
Press, Birmingham, 1974.
1088, 1051, 982, 938. EI-HR-MS m/z: 368.2192 (Calcd for C20H32O6: 13) Stewart R. F., J. Chem. Phys., 42, 3175—3187 (1965).
1
368.2191). H-NMR (pyridine-d5) d: 1.22 (3H, s, C18-H3), 1.50 (3H, s, C17-