Absolute stereochemistry of amphidinolide Q

Article abstract of DOI:10.1021/ol8013123

(Chemical Equation Presented) The absolute configurations at five chiral centers in amphidinolide Q (1), a cytotoxic 12-membered macrolide isolated from a marine dinoflagellate Amphidinium sp., were elucidated to be 4R, 7R, 9S, 11R, and 13R on the basis of NMR analyses and a modified Mosher's method,

Full text of DOI:10.1021/ol8013123

ORGANIC
LETTERS
2008
Vol. 10, No. 17
3709-3711
Absolute Stereochemistry of
Amphidinolide Q
Yohei Takahashi,† Takaaki Kubota,† Eri Fukushi,§ Jun Kawabata,§ and
Jun’ichi Kobayashi*,†
Graduate School of Pharmaceutical Sciences, Hokkaido UniVersity,
Sapporo 060-0812, Japan, and Graduate School of Agriculture,
Hokkaido UniVersity, Sapporo 060-0859, Japan
jkobay@pharm.hokudai.ac.jp
Received June 10, 2008
ABSTRACT
The absolute configurations at five chiral centers in amphidinolide Q (1), a cytotoxic 12-membered macrolide isolated from a marine dinoflagellate
Amphidinium sp., were elucidated to be 4R, 7R, 9S, 11R, and 13R on the basis of NMR analyses and a modified Mosher’s method.
Amphidinolides are a series of cytotoxic macrolides pos-
sessing unique structural features isolated from laboratory
cultured marine dinoflagellates Amphidinium sp.1 Amphi-
dinolide Q (1) isolated from a dinoflagellate Amphidinium
sp. (Y-5 strain) is a cytotoxic 12-membered macrolide having
C1 branches at vicinal carbons (C-13 and C-14) and an R,ꢀ-
unsaturated ester moiety.2 The gross structure of 1 has been
elucidated by 2D NMR, whereas the stereochemistry remains
unsolved due to lack of sample.2 Quite recently, we could
reisolate amphidinolide Q (1) from the same strain so that
this sample was used to elucidate the stereochemistry of 1.
Here, we describe the elucidation of the absolute configura-
tions at five chiral centers in 1 on the basis of the NMR
analysis and a modified Mosher’s method.
(JBCA) method3 and NOESY correlations. The geminal,
vicinal, and long-range J(C,H) coupling constants were
obtained by the resolution-enhanced 1H spectrum, the HET-
LOC,4 and J-IMPEACH-MBC5 spectra in C6D6. A small
3
2
value for J(H-4/H-5a) and a large value for J(H-5b/C-4)
indicated that H-4/H-5a and 4-OH/H-5b were both gauche
3
(Figure 1). A middle value for J(H-4/H-5b) suggested that
The relative stereochemistry for amphidinolide Q (1) was
elucidated on the basis of the J-based configuration analysis
† Graduate School of Pharmaceutical Sciences.
§ Graduate School of Agriculture.
Figure 1. Rotation models for the C-4/C-5 bond of amphidinolide
(1) (a) Kobayashi, J.; Kubota, T. J. Nat. Prod. 2007, 70, 451–460. (b)
Kobayashi, J. J. Antibiot. 2008, 61, 271–284. (c) Takahashi, Y.; Kubota,
T.; Kobayashi, J. J. Antibiot. 2007, 60, 376–379. (d) Oguchi, K.; Tsuda,
M.; Iwamoto, R.; Okamoto, Y.; Kobayashi, J.; Fukushi, E.; Kawabata, J.;
Ozawa, T.; Masuda, A.; Kitaya, Y.; Omasa, K. J. Org. Chem. 2008, 73,
1567–1570.
Q (1).
the C-4/C-5 bond existed as two conformers, while NOESY
correlations for H-4/H-5a, H-4/H-5b, and H-5a/H3-17 in C6D6
indicated that the C-4/C-5 bond existed mainly as a
(2) Kobayashi, J.; Takahashi, M.; Ishibashi, M. Tetrahedron Lett. 1996,
37, 1449–1450.
10.1021/ol8013123 CCC: $40.75
Published on Web 08/05/2008
2008 American Chemical Society

conformer with gauche relationships for C-3/H-5a and H-4/
H-5b. The relative stereochemistry of the C-7 to C-11 moiety
in 1 was deduced from analyses of the HETLOC, J-
IMPEACH-MBC, and NOESY spectra in C6D6. The middle
values for J(H-7/H-8b) and J(H-9/H-10a) indicated that
C-7/C-8 and C-9/C-10 bonds adapted two conformers, while
NOESY cross-peaks of H-7/H-9, H-8a/H-10a, H-9/H-11, and
H-10b/H3-19 as well as small J-values for H-7/H-8a, H-8b/
C-6, H-9/H-10b, and H-10a/C-19 allowed us to assign the
major confomer for C-7/C-9 and C-9/C-10 bonds as shown
in Figure 2a and 2c, respectively. NOESY correlations for
10a), 3J(H-11/H-10b), and 2J(H-10b/C-11) revealed that C-8/
C-9 and C-10/C-11 bonds existed as a single conformer as
shown in Figure 2b and 2d, respectively.
Relative configurations at C-4 and C-7 were deduced from
NOESY correlations for H-2/H-8a, H3-17/H-5a, H-4/H2-5,
and H-5a/H-7 for 1 as shown in Figure 3.
3
3
Figure 3. Selected NOESY correlations and relative stereochemistry
for the C-2∼C-11 portion of amphidinolide Q (1).
The relationship of C-11 and C-13 could not be deduced
from the JBCA method due to the overlapping of signals
for H-12a and H-12b in C6D6, whereas these signals were
3
slightly separated in CDCl3. A small value for J(H-11/H-
3
12a) and a large value for J(H-11/H-12b) suggested a
gauche relationship of H-11/H-12a and an anti relationship
for H-11/H-12b, respectively. In addition, an anti relationship
of H-12a/11-O deduced from a small J-value of H-12a/C-
11 and NOESY correlations for H-11/H-12a and H-11/H-
13 revealed that the C-11/C-12 bond existed as a single
conformer as shown in Figure 4a. On the other hand, NOESY
Figure 2. Rotation models for (a) C-7/C-8, (b) C-8/C-9, (c) C-9/
C-10, and (d) C-10/C-11 bonds of amphidinolide Q (1).
H-7/H-9, H-8a/H-10a, H-9/H-11, and H-10b/H-11 and values
for J(H-9/H-8a), J(H-9/H-8b), J(H-8a/C-19), J(H-11/H-
Figure 4. Rotation models for (a) C-11/C-12 and (b) C-12/C-13
bonds of amphidinolide Q (1).
3
3
3
3
(3) Matsumori, N.; Kaneno, D.; Murata, M.; Nakamura, H.; Tachibana,
K. J. Org. Chem. 1999, 64, 866–876.
cross-peaks of H-11/H-13, H-12a/H-21a, and H-12b/H3-20
(4) (a) Otting, G.; Wu¨thrich, K. Q. ReV. Biophys. 1990, 23, 39–96. (b)
Wollborn, U.; Leibfritz, D. J. Magn. Reson. 1992, 98, 142–146. (c) Kurz,
M.; Schmieder, P.; Kessler, H. Angew. Chem., Int. Ed. Engl. 1991, 30, 1329–
1331.
3
as well as the large value for J(H-13/H-12a) and middle
values for 3J(H-13/H-12b) and 3J(H-12a/C-20) indicated that
the C-12/C-13 bond existed as a pair of two conformers as
shown in Figure 4b.
(5) Williamson, R. T.; Marquez, B. L.; Gerwick, W. H.; Martin, G. E.;
Krishnamurthy, V. V. Magn. Reson. Chem. 2001, 39, 127–132.
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Org. Lett., Vol. 10, No. 17, 2008

The molecular mechanics calculations of 1 were carried
out by using Sybyl 6.5 (MMFF94 force-field).6 One of the
most stable confomers explained well the NOESY cor-
reations observed for 1 (Figure 5). Thus, the relative
stereochemistry of amphidinolide Q (1) was elucidated as
shown.
Figure 6. ∆δ values [∆δ (in ppm) ) δS - δR] obtained for the
(S)- and (R)-MTPA esters (2a and 2b, respectively) of amphi-
dinolide Q (1).
mixture of two linear methyl esters generated by dehydration
of C-4 and C-5 and epimerization of C-7. The mixture was
treated with (R)- and (S)- MTPACl, and the resulting bis-
(S)- and (R)-MTPA esters were obtained by HPLC purifica-
tion. ∆δ values for 3a and 3b suggested the absolute
configuration at C-11 to be R (Figure 7). Thus, the absolute
configurations at the five chiral centers in 1 were concluded
to be 4R, 7R, 9S, 11R, and 13R, respectively.
Figure 5. Selected NOESY correlations and a stable conformer
for macrolactone ring of amphidinolide Q (1) obtained from the
molecular mechanics calculations (hydrogen atoms of methyl groups
were omitted).
To elucidate the absolute configurations of amphidinolide
Q (1), a modified Mosher’s method7 was applied as follows.
Treatment of 1 with (R)-(-)- and (S)-(+)-2-methoxy-2-
trifluoro-2-phenylacetyl chloride (MTPACl) gave the (S)- and
(R)-MTPA esters (2a and 2b, respectively) of 1. ∆δ values
Figure 7. ∆δ values [∆δ (in ppm) ) δS - δR] obtained for the
(S)- and (R)-MTPA esters (3a and 3b, respectively) of the linear
methyl ester derivatives of amphidinolide Q (1).
1
(∆δ ) δS - δR) obtained from H NMR data of 2a and 2b
are shown in Figure 6. The ∆δ values for H2-5 and H3-18
are positive, while negative ∆δ values are observed for H-2,
H2-10, H-13, H3-16, H3-17, H3-19, H3-20, and H2-21.8 These
results indicated that the absolute configuration at C-4
was R.
The relative stereochemistry of 1 was elucidated as
described above, while the absolute configuration at C-11
was elucidated directly by application of a modified Mosher
method to the linear methyl ester derivatives of amphidino-
lide Q (1). Treatment of 1 with K2CO3 in MeOH yielded a
Acknowledgment. The authors thank Ms. S. Oka, Center
for Instrumental Analysis, Hokkaido University, for mea-
surements of ESIMS. This work was partly supported by
grants from a Takeda Science Foundation, a Mochida
Memorial Foundation for Medical and Pharmaceutical
Research, and a Grant-in-Aid for Scientific Research from
the Ministry of Education, Culture, Sports, Science and
Technology of Japan.
Supporting Information Available: Experimental pro-
cedures and one- and two-dimensional NMR spectra for 1,
2a, 2b, 3a, and 3b. This material is available free of charge
via the Internet at http://pubs.acs.org.
(6) Halgren, T. A. J. Am. Chem. Soc. 1992, 114, 7827–7643.
(7) Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J. Am. Chem.
Soc. 1991, 113, 4092–4095.
(8) A positive ∆δ value for H-11 seems to be due to the anisotropic
effect of the ester carbonyl group at C-1.
OL8013123
Org. Lett., Vol. 10, No. 17, 2008
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