Stereochemical determination of acyclic structures based on carbon- proton spin-coupling constants. A method of configuration analysis for natural products

Article abstract of DOI:10.1021/jo981810k

A method for elucidating the relative configuration of acyclic organic compounds was developed on the basis of carbon-proton spin-coupling constants (^2,3J(C,H)) and interproton spin-coupling constants (³J(H,H)). This method is based

Full text of DOI:10.1021/jo981810k

866
J . Org. Chem. 1999, 64, 866-876
Ster eoch em ica l Deter m in a tion of Acyclic Str u ctu r es Ba sed on
Ca r bon -P r oton Sp in -Cou p lin g Con sta n ts. A Meth od of
Con figu r a tion An a lysis for Na tu r a l P r od u cts
Nobuaki Matsumori,†,§ Daisuke Kaneno,† Michio Murata,*,† Hideshi Nakamura,‡ and
Kazuo Tachibana†
Department of Chemistry, School of Science, The University of Tokyo,
Hongo, Bunkyo-ku, Tokyo 113-0033, J apan, and Division of Chemistry, Graduate School of Science,
Hokkaido University, Sapporo 060-0810, J apan
Received September 4, 1998
A method for elucidating the relative configuration of acyclic organic compounds was developed on
the basis of carbon-proton spin-coupling constants (2,3J C,H) and interproton spin-coupling constants
(3J H,H). This method is based on the theory that, in acyclic systems, the conformation of adjacent
asymmetric centers is represented by staggered rotamers, and their relative stereochemistry can
2,3
3
be determined using
J
and J H,H, because the combined use of these J values enables the
C,H
identification of the predominant staggered rotamer(s) out of the six possible derived from threo
and erythro configurations. Detailed conformational analysis for model compounds 1-4 revealed
that this method is useful in most cases for assignment of the configuration of acyclic structures
occurring in natural products, in which stereogenic methine carbons are often substituted with a
methyl or a hydroxy (alkoxy) group. This J -based configuration analysis was applied to the
stereochemical elucidation of carboxylic acid 5 derived from zooxanthellatoxin and proven to be a
practical method even for natural products with complicated structures.
In tr od u ction
prymnesins,8 and zooxanthellatoxins.9 These compounds
are very difficult to crystalize and, to make matters
worse, their acyclic parts are laborious to synthesize
because of the number of chiral centers they contain.
Knowing the configuration of natural products has
become crucial because it provides essential information
for both total synthesis and molecular mode of actions,
which are now regarded as the most challenging fields
in organic and bioorganic chemistry. NMR-based meth-
ods have been devised for this purpose; e.g., NOE-based
methods in combination with molecular mechanics cal-
culations have been proposed for flexible molecules,
particularly for macrocyclic compounds, as reported for
macrolides10 and other compounds.11 However, even with
new NOE-based techniques, it is still very difficult to
assign the stereochemistry of highly flexible carbon
chains because the presence of multiple conformers, in
which minor populations often make disproportionately
large contributions to NOE intensity, occasionally leads
to contradictory distance constraints.
Natural products possessing complicated structures
and potent biological activities have been isolated from
marine sources and microorganisms in the last two
decades, and their structures have been successfully
elucidated mainly by modern NMR techniques. Among
these compounds, acyclic or macrocyclic structural moi-
eties appear frequently. Their configurations, however,
have been determined chiefly by X-ray or laborious
synthetic work as seen for palytoxin,1 amphotericin B,2
calyculins,3 fumonisins,4 and swinholides.5 Assignments
of configuration of acyclic structures are extremely dif-
ficult even with current NMR-based methods. This may
be one reason that a fairly large number of acyclic
moieties in important natural products remain stereo-
chemically unassigned; e.g., aflastatin,6 amphidinols,7
* To whom correspondence should be addressed. E-mail address:
murata@chem.s.u-tokyo.ac.jp. FAX/telephone: +81-3-5800-6898.
† The University of Tokyo.
‡ Hokkaido University, Present address of H.N.: Faculty of Agri-
culture, Nagoya University.
§ Present address: Department of Biotechnology, Division of Agri-
culture and Life Sciences, The University of Tokyo, 1-1 Yayoi, Bunkyo-
ku, Tokyo 113-8657, J apan.
(7) (a) Satake, M.; Murata, M.; Yasumoto, T.; Fujita, T.; Naoki, H.
J . Am. Chem. Soc. 1991, 113, 9859-9861. (b) Paul, G. K.; Matsumori,
N.; Murata, M.; Tachibana, K. Tetrahedron Lett. 1995, 36, 6279-6282.
(8) Igarashi, T.; Satake, M.; Yasumoto, T. J . Am. Chem. Soc. 1996,
118, 479-480.
(1) (a) Uemura, D.; Ueda, K.; Hirata, Y.; Katayama, C.; Tanaka, J .
Tetrahedron Lett. 1980, 21, 4861-4864. (b) Cha, J . K.; Christ, W. J .;
Finan, J . M.; Fujioka, H.; Kishi, Y.; Klein, L. L.; Ko, S. S.; Leder, J .;
McWhorter, J r. W. W.; Pfaff, K.-P.; Yonaga, M.; Uemura, D.; Hirata,
Y. J . Am. Chem. Soc. 1982, 104, 7369-7371.
(2) (a) Mechlinski, W.; Schaffner, C. P.; Ganis, P.; Avitabile, G.
Tetrahedron Lett. 1970, 3873-3876. (b) Ganis, P.; Avitabile, G.;
Mechlinski, W.; Shaffner, C. P. J . Am. Chem. Soc. 1971, 93, 4560-
4564.
(3) Kato, Y.; Fusetani, N.; Matsunaga, S.; Hashimoto, K.; Fujita,
S.; Furuya, T. J . Am. Chem. Soc. 1986, 108, 2780-2781.
(4) Harmange, J .-C.; Boyle, C. D.; Kishi, Y. Tetrahedron Lett. 1994,
35, 6819-6822.
(5) Kitagawa, I.; Kobayashi, M.; Katori, T.; Yamashita, M.; Tanaka,
J .; Doi, M.; Ishida, T. J . Am. Chem. Soc. 1990, 112, 3710-3712.
(6) Sakuda, S.; Ono, M.; Furihata, K.; Nakayama, J .; Suzuki, A.;
Isogai, A. J . Am. Chem. Soc. 1996, 118, 7855-7856.
(9) Nakamura, H.; Asari, T.; Murai, A.; Kan, Y.; Kondo, T.; Yoshida,
K.; Ohizumi, Y. J . Am. Chem. Soc. 1995, 117, 550-551.
(10) (a) Kobayashi, M.; Aoki, S.; Kitagawa, I.; Kobayashi, Y.;
Nemoto, N.; Fujikawa, A. Symposium papers of 38th Symposium on
the Chemistry of Natural Products; Organizing Committee of the
Symposium: Sendai, 1996; pp 61-66. (b) Kobayashi, M.; Aoki, S.;
Kitagawa, I. Tetrahedron Lett. 1994, 35, 1243-1246. (c) Fujita, K.;
Fujiwara, M.; Yamasaki, C.; Matsuura, T.; Furihata, K.; Seto, H.
Symposium papers of 38th Symposium on the Chemistry of Natural
Products; Organizing Committee of the Symposium: Sendai, 1996; pp
379-384. (d) Hayakawa, Y.; Kim, J .-W.; Adachi, H.; Shin-ya, K.; Fujita,
K.; Seto, H. J . Am. Chem. Soc. 1998, 120, 3524-3525.
(11) (a) Falk, M.; Spierenburg, P. F.; Walter, J . A. J . Comput. Chem.
1996, 17, 409-417. (b) Ko¨ck, M.; J unker, J . J . Org. Chem. 1997, 62,
8614-8615.
10.1021/jo981810k CCC: $18.00 © 1999 American Chemical Society
Published on Web 01/20/1999

Stereochemical Determination of Acyclic Structures
J . Org. Chem., Vol. 64, No. 3, 1999 867
F igu r e 1. Dihedral angle dependence of spin-coupling con-
3
3
stants, J H,H,
2J C,H, and J C,H. a: Vicinal 1H-1H coupling
constants, 3J H,H, when the dihedral angles are 180° (anti) and
60° (gauche). b: Geminal 13C-1H coupling constants, J C,H
,
2
when the dihedral angles between 13C-attached oxygen and
the proton are 180° (anti) and 60° (gauche). c: Vicinal 13C-
1H coupling constants, 3J C,H, when the dihedral angles are 180°
(anti) and 60° (gauche). The figures in parentheses represent
the values of 1,2-dioxygenated systems (see Table 1 for details).
F igu r e 2. Asymmetric centers in acyclic or macrocyclic
structures occurring in natural products mostly consist of
methine carbons substituted with a hydroxy (alkoxy) or methyl
group. a: C1-C10 and C83-92 parts of palytoxin. b: Terminal
chains of maitotoxin (C4-C14 and C135-C142). c: Acyclic
parts of zooxanthellatoxin A (C44-C59 and C77-C84). Num-
bering in the structures denotes carbon numbers of entire
structures.
The spin-coupling constant is a useful NMR parameter
for conformational studies of biomolecules as seen in its
application to polypeptides or polynucleotides. In par-
ticular, proton-proton vicinal coupling constants (3J H,H
)
are most frequently used for this purpose because of their
dependence on dihedral angles. These coupling constants,
including those due to carbon-proton coupling (2J C,H and
3J C,H), possess several advantages over NOE for acyclic
systems. In systems with conformational changes, the
coupling constants are observed as a weighted average
of those due to each conformer, which greatly facilitates
the determination of a population ratio among conform-
ers. For configuration assignments, using interproton J
values (3J H,H) alone is inadequate because two H/H-
a Karplus-type equation (Figure 1)14 and thus can be
utilized theoretically for stereochemical analysis as is the
3
3
case with J H,H. The value of J C,H has been used for
conformational studies of proteins and oligonucleotides.15
In addition, geminal carbon-proton coupling constants
(2J C,H) also provide conformational information;16 when
an oxygen functionality on a carbon atom is gauche to
gauche rotamers cannot be distinguished as shown in
2
its geminal proton, J C,H becomes large, and when it is
2,3
Figures 1 and 3. Additional information from
J
may
C,H
anti, the value becomes small (Figure 1). However, to our
knowledge, comprehensive configuration analysis of com-
plicated natural products based on carbon-proton cou-
pling constants (2,3J C,H) has not been reported except
those from our group,13 probably due to difficulty in
dramatically expand the utility of the coupling constants
in configuration analysis. Thanks to the progress in two-
dimensional NMR techniques and hardware innovation
in the past decade,12 2,3J C,H values in complicated natural
products have become measurable.
In this study we report the development of a method
called J -based configuration analysis for assigning the
relative configuration of natural products, in particular,
their acyclic subunits.13
determining 2,3J C,H values accurately. Therefore, we first
2,3
set up a basic strategy for correlating
J
values with
C,H
the configuration of acyclic asymmetric carbons.
(14) (a) Lemieux, R. U.; Nagabhushan, T. L.; Paul, B. Can. J . Chem.
1972, 50, 773-776. (b) Schwarcz, J . A.; Perlin, A. S. Can. J . Chem.
1972, 50, 3667-3676. (c) Wasylishen, R.; Schaefer, T. Can. J . Chem.
1973, 51, 1, 961-973. For a review, see: Hansen, P. E. Prog. NMR
Resu lts
Rota tion a l Isom er s a n d Str u ctu r a l An a lysis. Vici-
nal carbon-proton spin-coupling constants (3J C,H) follow
Spectrosc. 1981, 14, 175-296.
2,3
(15) Applications of
J
to conformational analyses and/or ste-
C,H
reospecific assignments of methylene protons have been reported for
proteins (a-c), carbohydrates (d, e), and polynucleotides (f, g): (a)
Wang, A. C.; Bax, A. J . Am. Chem. Soc. 1996, 118, 2483-2494. (b)
Kessler, H.; Griesinger, C.; Wagner, K. J . Am. Chem. Soc. 1987, 109,
6927-6933. (c) Hansen, P. E. Biochemistry 1991, 30, 10457-10466.
(d) Podlasek, C. A.; Wu, J . Stripe, W. A.; Bondo, P. B.; Serianni, A. S.
J . Am. Chem. Soc. 1995, 117, 8635-8644. (e) Duker, J .; Serianni, A.
S. Carbohydr. Res. 1993, 249, 281-303. (f) Hines, J . V.; Landry, S.
M.; Varani, G.; Tinoco, I., J r. J . Am. Chem. Soc. 1994, 116, 5823-
5831. (g) Marino, J . P.; Schwalbe, H.; Glaser, S. J .; Griesinger, C. J .
Am. Chem. Soc. 1996, 118, 4388-4395.
(16) (a) Schwarcz, J . A.; Cyr, N.; Perlin, A. S. Can. J . Chem. 1975,
53, 1872-1875. (b) Parella, T.; Sanchez-Ferrando, F.; Virgili, A. Magn.
Reson. Chem. 1994, 32, 657-664. (c) Podlasek, C. A.; Wu, J .; Stripe,
W. A.; Bondo, P. B.; Serianni, A. S. J . Am. Chem. Soc. 1995, 117, 8635-
8644.
(12) For a review, see: Eberstadt, M.; Gemmechker, G.; Mierke, D.
G.; Kessler, H. Angew. Chem., Int. Ed. Engl. 1995, 34, 1671-1695.
(13) Applications of this methodology to the configuration assign-
ment of natural products have been published in the following
papers: (a) Matsumori, N.; Murata, M.; Tachibana, K. Tetrahedron
1995, 51, 12229-12238. (b) Matsumori, N.; Nonomura, T.; Sasaki, M.;
Murata, M.; Tachibana, K.; Satake, M.; Yasumoto, T. Tetrahedron Lett.
1996, 37, 1269-1272. (c) Sasaki, M.; Matsumori, N.; Maruyama, T.;
Nonomura, T.; Murata, M.; Tachibana, K.; Yasumoto, T. Angew. Chem.,
Int. Ed. Engl. 1996, 35, 1672-1675. (d) Nonomura, T.; Sasaki, M.;
Matsumori, N.; Murata, M.; Tachibana, K.; Yasumoto, T. Angew.
Chem., Int. Ed. Engl. 1996, 35, 1675-1678. (e) Sakai, R.; Kamiya, H.;
Murata, M.; Shimamoto, K. J . Am. Chem. Soc. 1997, 119, 4112-4116.
(f) Murata, M.; Matsumori, N.; Tachibana, K. Nippon Kagaku Kaishi
1997, 749-757.

868 J . Org. Chem., Vol. 64, No. 3, 1999
Matsumori et al.
proton signals coupling with carbons of interest must be
separated from each other to enable a first-order analysis
of 1H-1H coupling. To satisfy this, the number of bonds
separating asymmetric centers should be one, two, or at
most three, since propylene (CH2CH2CH2) or longer
carbon chains usually give rise to overlapping proton
signals. Thus, in this study, stereochemical correlations
between 1,2-methines and between 1,3-methines bearing
hydroxy and/or methyl groups are the main focus. Even
with all these conditions, this method would greatly
assist in the determination of the configuration of natural
products, because the acyclic asymmetric carbons occur-
ring in these compounds, probably more than 80%, fall
under these categories.
To establish J -based configuration analysis, we cat-
egorized dihedral interactions into several cases. Our first
assumption was that, in the systems in Figure 3, con-
formations can be represented by three staggered rota-
mers. This seems to be quite reasonable for acyclic carbon
chains with hydroxy and methyl substitutions, which,
unlike bulky substituents, usually cause no significant
deviations from the anti or gauche orientation.21,22 During
our investigations of over 30 diastereorelationships oc-
curring in 10 natural and synthetic compounds,23 no
single example of deviation over 10° has been found so
far (if their dihedral angles deviate from the staggered
3
rotamer by over 10°,22 J H,H becomes an atypical value
and unconditional assignment of stereochemistry cannot
be achieved, particularly when conformational change is
involved). The occurrence of these unusual conformers
can be judged by accurate measurements of coupling
constants. 3J H,H can be precisely determined (0.1-0.2 Hz)
2,3
F igu r e 3. Dependence of 3J H,H and
J
on dihedral angles
C,H
between vicinal methine carbons in 2,3-disubstituted butane
systems. Frequently occurring pairs of substituents in natural
products, Me/OR and OR/OR, are presented (Me/Me is omitted
since it rarely occurs in natural products). R stands for H,
using decoupling difference spectra or E.COSY-type
2,3
experiments,24 but measurements of
J
with an
C,H
3
alkyl, or/and acyl groups. When J H,H is small () gauche), the
accuracy better than 1 Hz are not usually attainable
owing to the low signal-to-noise ratio and limited digital
resolution in 2D methodologies. In the following experi-
ments, therefore, values of 3J H,H, which were used for not
only stereochemical determination but also conforma-
tional analysis, were determined with an accuracy of 0.1
four rotamers A-1, A-2, B-1, and B-2 can be clearly identified
2,3
3
using
J
values; either 3J (H-2,C4)/3J (C1,H-3), J (Cx,H-3)/
C,H
2J (C3,H-2) or J (C2,H-3)/2J (C3,H-2) can be used to identify the
four rotamers. H/H-anti rotamers A-3 and B-3 can be distin-
guished on the basis of NOEs.25
2
2
3
For structure elucidation of natural products, stereo-
genic carbons substituted with a hydroxy, an alkoxy,17
or a methyl group are particularly important. In other
words, most asymmetric centers existing in acyclic
subunits are methine carbons bearing a hydroxy or a
methyl as seen in the side chains of palytoxin, maito-
toxin,18 and zooxanthellatoxins (Figure 2).9 To determine
the configuration of multiple asymmetric carbons, the
diastereomeric relationship between two neighboring
carbons must be assigned; as shown in Figure 2, 1,2-,
1,3-, and 1,4-asymmetric systems are commonly found
in natural products. For accurate measurements of 2,3J C,H
by 2D NMR methods such as HETLOC19 and HMBC,20
Hz, while J C,H and J C,H were obtained with single digit
accuracy.
Con figu r a tion Assign m en t for System s w ith On e
Dom in a n t Con for m er . 1,2-Meth in e System s. To as-
sign the stereochemical relationship for a pair of vicinal
asymmetric carbons, we have to choose a single con-
former with a correct configuration from the staggered
rotamers possible in threo and erythro diastereomers
(21) Eliel, E. L.; Allinger, N. L.; Angyal, S. J .; Morrison, G. A.
Conformational Analysis; J ohn Wiley & Sons. Inc.: New York, 1965;
pp 13-21.
(22) Dependence of 3J H,H on dihedral angles was examined for model
compounds 1-4 on the basis of a modified Karplus equation28 and force
field calculations. The C1-C2-C3-C4 dihedral angles in the stable
conformers for 1-3 deviated from 180° by less than 10°; 1 was most
stable at a dihedral angle C1/C2/C3/C4 of 183° and had the weighted
(17) In configuration studies of natural products, the stereochemical
relationship between asymmetric carbons residing in cyclic ether or
lactone structures and those in a side chain is often important as seen
for C15-C14 of maitotoxin (Figure 2).13c,d To such systems, the J -based
method was successfully applied. Although applications to acyloxy
substitutions are not plentiful, preliminary investigations on tri-O-
acetyl model compounds 1 and 2 suggested that this method can be
applied for acetoxy-substituted chains.
3
3
average J H-2,H-3 of 0.7 Hz. In the range of 166-190°, J H-2,H-3 in 1
does not significantly (less than 1 Hz) deviate from the calculated value.
Compounds 2 and 3 were also most stable in the C1/C4-anti conforma-
tions at 180° (8.7 Hz) and at 175° (2.7 Hz), respectively. Within the
ranges of 157-203° and 167-185°, 2 and 3 showed no significant
3
aberrations in J H-2,H-3 values (<1 Hz), respectively.
(23) We have examined the diastereomeric relationship of asym-
metric carbons residing in acyclic portions for maitotoxin,13b-d amphi-
dinol,7 okadaic acid,13a zooxanthellatoxin, and several synthetic com-
pounds. Force field calculations (CFF91 force field on DISCOVER/
INSIGHT II) carried out for okadaic acid and maitotoxin revealed that
deviations of all 1H/1H-dihedral angles in acyclic structures from 60°
or 180° were less than 10° as is the case with the model compounds.22
(24) Bru¨schweiler, R.; Madsen, J . C.; Griesinger, C.; Sørensen, O.
W.; Ernst R. R. J . Magn. Reson. 1987, 73, 380-385.
(18) Murata, M.; Naoki, H.; Matsunaga, S.; Satake, M.; Yasumoto,
T. J . Am. Chem. Soc. 1994, 116, 7098-7107.
(19) (a) Otting, G.; Wu¨thrich, K. Quart. 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.
(20) (a) Zhu, G.; Bax, A. J . Magn. Reson. 1993, 104A, 353-357. (b)
Zhu, G.; Live, D.; Bax, A. J . Am. Chem. Soc. 1994, 116, 8370-8371.

Stereochemical Determination of Acyclic Structures
J . Org. Chem., Vol. 64, No. 3, 1999 869
(Figure 3). Among those, four conformers, A-1, A-2, B-1,
3
2
3
and B-2, can be identified using J H,H, J C,H, and J C,H
,
while the two rotamers A-3 and B-3 with an H/H-anti
orientation cannot be distinguished. For these anti
conformers, NOE experiments should be a practical way
to assign their configuration. In acyclic organic com-
pounds with methyl or hydroxy substituents, these H/H-
anti conformers usually assume a C/C-anti orientation
(or an extended form), in which no NOE (or ROE) should
be observed between H-1 and H-4 (B-3 in Figure 3). In
the case of the H/H-anti and C/C-gauche conformation
(A-3), if present, H-1 and H-4 should come within the
range of NOE.25 Using these criteria, all six conformers
could be discriminated, with their relative configuration
(threo or erythro) determined accordingly.
1,3-Meth in e System s. To determine the relative
configuration of two methine carbons separated by a
methylene group, the pair of diastereotopic methylene
protons must be assigned stereospecifically. This task is
not always easy using NOE, particularly for large
molecules with masses above 1000 Da, in which spin
diffusion between methylene protons tends to obscure
NOE-based assignments. Using coupling constants, as-
signments can be made for large molecules as reported
for maitotoxin.13b-d The method for assigning diaste-
reotopic methylene protons with respect to the adjacent
methine is similar to that for vicinal methines described
above. In these structural units, six conformers are
possible when a pair of protons on a methylene is
stereochemically labeled according to their chemical
shifts. All these conformers can be unambiguously identi-
2,3
fied using 3J H,H and
J
, as depicted in Figure 4. With
C,H
2,3
F igu r e 4. Dependence of 3J H,H and
J
on dihedral angles
C,H
one methylene-methine relationship in hand, the same
examination of another relationship leads to diastereo-
meric assignment of the 1,3-methine groups via the
stereospecifically labeled methylene protons. If 1H NMR
signals of relevant protons are separated, this method
can be applied to 1,4-methine systems separated by an
ethylene group.
between methine and methylene in 2-substituted butane
systems. R stands for H, alkyl, or/and acyl groups. Six sets of
J values are derived from two different stereospecific assign-
ments for methylene protons on C3, which are labeled accord-
ing to their chemical shifts; Hh is observed at higher field and
Hl is at lower field. With J H,H alone, none of the six rotamers
3
3
h
can be distinguished. However, with use of either J C1,H-3
/
3
3
3
2
2
h
l
h
l
l
J C1,H-3 , J CX,H-3 / J CX,H-3 , or J C2,H-3 / J C2,H-3 , they are clearly
Con figu r a tion Assign m en t for System s w ith Al-
ter n a tin g Con for m er s. In acyclic systems, two or three
rotamers frequently coexist. The contributions from
minor rotamers that comprise less than 1 Hz are negli-
identified, which leads to the stereospecific assignment of the
methylene protons on C3 with respect to C2.
3
2,3
Ta ble 1. J H,H a n d
J
Va lu es (Hz) for An ti a n d
C,H
Ga u ch e Or ien ta tion s in Acyclic System s
gible (in most cases, 1 Hz is around 15% of the difference
2,3
in
J
between anti and gauche conformers). In other
C,H
3J H,H 2J C,H
anti gauche gauchea
3J C,H
words, if one conformer comprises over 85% of the
population, configuration analysis can be safely carried
antib
anti gauche
oxygenation large small
large
small large small
out only for the dominant conformer. In the following
2,3
sections, the correlation of
J
/3J H,H in erythro and
C,H
none
mono
di
9-12 2-4
8-11 1-4
-c
-
6-8
1-3
1-3
-5 to -7d 0 to -2d 6-8
7-10 0e-3 -4 to -6f 2-0f
5-7g 1-3g
3
(25) If J H,H shows an anti-typical value, NOE analysis should be
reliable since no significant conformational change takes place. In the
A-3 system, H-1 and H-4 should come close enough to give rise to a
prominent NOE, since the carbon chains extending from C1 and C4
tend to adopt a C/C-anti conformation as depicted below. During our
studies,13 however, we have not encountered the predominant occur-
rence of A-3; usually, A-3 coexists with A-1, where their configuration
can be assigned by coupling constants (Figure 5). For the B-3 case, in
addition to the absence of NOE (or ROE) between H-1 and H-4, NOE
(ROE) between H-1 and a proton on Y or between H-4 and a proton on
X should be observed when Y or X is a methyl or hydroxyl group;
NOESY experiments in DMSO-d6 sometimes show prominent NOEs
from OH protons in these systems.
a,b
Oxygen functions on relevant carbons are gauche and anti
to their vicinal protons, respectively (see Figure 1b).
do not show dihedral dependence. J C,H values are those between
c 2
J
values
C,H
d
2
an oxygenated carbon and a proton on the neighboring carbon.
e The value (0) is observed for the system where both protons are
f 2
anti to oxygen atoms such as A-1 in Figure 3.
those between an oxygenated carbon and a proton on the neigh-
values are those between a
proton on an oxygenated carbon and a carbon at the vicinal
position via another oxygenated carbon (see Figure 1c).
J
values are
C,H
g 3
boring oxygenated carbon.
J
C,H
threo configurations is examined for 1,2-methine and 1,3-
methine systems with alternating conformers.
3
1,2-Meth in e System s. When J H,H attains an inter-
mediate value between anti and gauche (see Table 1),
two major rotamers with H/H-anti and gauche orienta-
tions should be considered. These conformational changes

870 J . Org. Chem., Vol. 64, No. 3, 1999
Matsumori et al.
2,3
F igu r e 5. Dependence of 3J H,H and
J
on dihedral angles between vicinal methine carbons in alternating butane systems. R
C,H
3
stands for H, alkyl, or/and acyl groups. When J H,H is a medium value as a mixture of anti and gauche, the four pairs of alternating
rotamers A-2/A-3, A-1/A-3, B-2/B-3, and B-1/B-3 can be clearly identified using either J C4,H-2/3J C1,H-3, 3J CX,H-3/2J C3,H-2, or J C2,H-3
/
3
2
2J C3,H-2. H/H-gauche pairs, A-1/A-2 and B-1/B-2, cannot be identified with their configurations, but these pairs rarely occur in
acyclic structures of natural products since they are thought to be thermodynamically disfavored pairs. See Table 1 for small,
medium, and large volumes.
are often observed in natural products.13 In such a case,
four out of six alternating pairs can be unambiguously
ral products when rotational changes occur as C-1/C-3
or D-1/D-3 in Figure 6 (these conformers are observed in
5), while the other pairs of conformers are rarely present
because thermodynamically unlikely rotamers (C-2 and
D-2) with double gauche interactions between C1/C4 and
between X/C4 are involved. Similar treatments to those
for the alternating methine-methine systems are pos-
sible for the elucidation of the relative configuration
between 1,3-methines through stereospecific assignment
of diastereotopic methylene protons (Figure 6).
2,3
3
identified using
J
and J H,H as shown in Figure 5,
C,H
where all four H/H-anti/gauche pairs of rotamers A-2/
A-3, A-1/A-3, B-2/B-3, and B-1/B-3 give rise to different
combinations of J values. When both alternating rota-
mers have an H/H-gauche orientation (A-1/A-2 or B-1/
B-2 in Figure 5), their configuration cannot be assigned
3
2,3
using J H,H and
J
alone. In these conformers, how-
C,H
ever, all the substituents on C2 and C3 are gathered in
one side, thus making them thermodynamically dis-
favored. As far as we know, the occurrence of these pairs
of rotamers is extremely rare in natural products.
Another case in which the J -based method fails to
determine configuration is that of three rotamers having
comparable populations, which leads to an averaging of
all J values.26 In this case, if a minor rotamer comprises
less than 10% of the population, its contribution can be
disregarded and the configuration can be assigned as
described above (an example of this case will be discussed
later).
2
3
Mea su r em en ts of J C,H a n d J C,H Va lu es. Several
practical methods have been proposed for the measure-
ment of 2,3J C,H 12 Among those, we adopted two methods,
.
hetero half-filtered TOCSY (HETLOC)19 and phase-
1
sensitive HMBC (PS-HMBC).20 HETLOC gives a H-1H
2,3
2D spectrum, where
J
is represented by the disloca-
C,H
1
tion, along F2, of a doublet cross-peaks split by J C,H
(Figure 7). With this method, accurate
2,3
J
values can
C,H
be determined even for a sample at millimolar concentra-
tion regardless of the size of the J coupling. Another
advantage of this method is that signs of J values can
be obtained (Figure 7). However, HETLOC has two
drawbacks: all carbons including those residing between
a relevant carbon and its long-range coupling proton have
to be protonated, and the sensitivity is dependent on the
3
1,3-Meth in e System s. When at least one of J H,H
values for methylene protons has an intermediate value,
the presence of multiple rotamers should be considered.
This conformational change is occasionally seen in natu-
3
magnitude of each J H,H through which TOCSY transfers
(26) The presence of three rotamers with comparable populations
magnetization. In such a case, PS-HMBC20 is used, which
3
can be recognized by the J H,H value according to Table 1. In such a
3
1
case, J H,H becomes a marginal value between gauche and medium
is designed to reproduce a H signal shape by setting a
because of the coexistence of two gauche and one anti rotamer.
refocus time before acquisition and by the application of
13C-decoupling during acquisition. In an original sequence
of PS-HMBC reported by Bax et al.20 a 1D reference
2,3
Additionally, none of the
J
values should take the typical gauche
C,H
values since, unlike a mixure of two conformers, anti interactions
2,3
should be involved in all 3J H,H and
J
(Figure 5). A minor conformer
C,H
with 10% or less population does not affect the average J value
spectrum is necessary to obtain the absolute value of
significantly, and thus its presence does not usually result in
a
2,3
deviation in J values.
J
, although this is not always possible for natural
C,H

Stereochemical Determination of Acyclic Structures
J . Org. Chem., Vol. 64, No. 3, 1999 871
2,3
F igu r e 6. Dependence of 3J H,H and
J
on dihedral angles between methine and methylene in alternating butane systems. R
C,H
stands for H, alkyl, or/and acyl groups. Methylene protons are labeled according to their chemical shifts; Hh (H-3h) and Hl (H-3l)
are protons with higher and lower-field signals, respectively. When either J H-2,H-3 or J H-2,H-3 takes a medium value, the
3
3
h
l
alternating rotamers should be considered. Six pairs of the rotamers with the different prochiral assignments for methylene
3
3
3
3
2
2
h
h
l
h
l
l
protons can be clearly distinguished using either J C1,H-3 / J C1,H-3 , J CX,H-3 / J CX,H-3 , or J C2,H-3 / J C2,H-3
.
IC ,H/IC ,H ) sin2(π 2,3J C ,H∆)/sin2(π 2,3J C ,H∆)
a
b
a
b
where IC ,H and IC ,H are the intensities of cross-peaks due
a
b
to Ca-H and Cb-H couplings, respectively, and ∆ is a
time interval in the pulse sequence.20
To minimize the effect of intramolecular hydrogen
bonding, we used a mixed solvent of pyridine-d5 and CD3-
3
2,3
OD (1:1) for measurement of J H,H and
J
. However,
C,H
the coupling constants of model compounds 1 and 2
measured in the mixed solvent were not very different
from those in a CDCl3 solution,27 in which hydrogen
bonds should make a greater contribution to the distribu-
tion of rotational conformation, implying that they may
not play a major role in these systems. We have mea-
2,3
sured
J
values for several highly oxygenated com-
C,H
pounds13b-f in the mixed solvent or methanol. Successful
assignments of their configurations based on these data
2,3
indicate that significant deviation from typical
J
C,H
F igu r e 7. Partial HETLOC spectrum of carboxylic acid 5 from
zooxanthellatoxin. An enlarged spectrum for H-3′ at δ 4.027
on F2 versus H-4′ at δ 3.25 on F1 is shown. Dislocation along
values (Table 1) hardly occurs even in polyhydroxy
structural moieties, which means that intramolecular
hydrogen bonding causes no significant conformational
aberration (less than 1 Hz) from the staggered rotamers
in these polar solvents.
1
F2 of the doublet cross-peak split along F1 with J C4′,H-4′
2
corresponds to J C4′,H-3′ (-1 Hz). Upper left way of the
dislocation indicates that the sign of the J is minus.
1
products due to extensive overlap of H signals. In this
study 2,3J C,H values were not directly measured but were
obtained from the ratio of their cross-peak magnitudes
with respect to a common proton. In this PS-HMBC
experiments, the absolute values are obtained by the
following equation using 2,3J C,H derived from other meth-
ods (e.g., HETLOC) as reference:13a
(27) Coupling constants of 1 and 2 in CDCl3 are as follows. 1:
2
2
3
3J H-2,H-3 ) 3.4 Hz; J C2,H-3 ) 0 Hz, J C3,H-2 ) -2 Hz; 2: J H-2,H-3
)
6.2 Hz, 2J C2,H-3 ) -4 Hz, 2J C3,H-2 ) -3 Hz. These values indicate that
their C2-C3 bond rotations in CDCl3 are similar to those in C5D5N-
CD3OD (1:1) (see Table 3); the difference by 0.8 Hz in J H-2,H-3 of 2 is
3
thought to be caused by hydrogen bonding between 2-OH and 3-OH,
which resulted in the increased population of an OH/OH-gauche
rotamer in CDCl3.

872 J . Org. Chem., Vol. 64, No. 3, 1999
Matsumori et al.
3
Ta ble 2. Ca lcu la ted J H,H for An ti a n d Ga u ch e Or ien ta tion s in F lu ctu a tin g System s of Meth in e-Meth ylen e a n d
Meth in e-Meth in e in Su bstitu ted Hexa n es
3
a
interproton dihedral angle (clockwise) and J H-3,H-4 (Hz)
substitution
(C2/C5 dihedral angle)
corresponding
rotamers
near 60°
near 180°
near 300°
3-Meb (C2/C5 60°)
(C2/C5 180°)
C-3/D-3
C-1/D-1
C-2/D-2
C-3/D-3
C-1/D-1
C-2/D-2
A-2,3,1
B-2,3,1
A-2,3,1
B-2,3,1
64° 2.5 (2.1)h
177° 11.6 (12.1)
173° 11.3 (12.0)
304° 4.0 (3.4)
297° 2.6 (2.4)
(C2/C5 300°)
53° 4.4 (3.9)*1
62° 1.8 (1.4)
3-OHc (C2/C5 60°)
(C2/C5 180°)
173° 10.3 (10.9)
184° 10.2 (10.7)
302° 3.7 (3.6)
294° 2.7 (2.5)
302° 1.0 (0.7)
290° 1.7 (1.5)
297° 3.2 (3.0)
298° 1.7 (1.5)
(C2/C5 300°)
63° 1.8 (1.4)
65° 3.5 (3.3)*2
70° 1.7 (1.5)
67° 1.3 (1.1)
70° 2.5 (2.1)
3,4-threo OH/OHd
3,4-erythro OH/OHe
3,4-threo Me/OHf
3,4-erythro Me/OHg
176° 8.1 (8.5)
179° 8.4 (8.8)
182° 9.5 (10.1)
181° 9.6 (10.2)
a
*The values with an asterisk do not fit to the criteria in Table 1, but the rotamers with *1 and *2, corresponding to C-2 (D-2) and A-2,
respectively, rarely occur in natural products because of their thermodynamically disfavored conformations. The calculations were carried
out for: b3(R)-methylhexane, c3(S)-hexanol, d(3R,4R)-hexanediol, emeso-3,4-hexanediol, f(3R,4R)-4-methyl-3-hexanol, and g (3R,4S)-4-methyl-
d-g
h
3-hexanol.
These configurations correspond to those of model compounds 1, 2, 3, and 4, respectively. The values in parentheses are
those at each local energy minimum (see Experimental Section for details).
3
2,3
Ta ble 3. J H,H a n d
J
(Hz) w ith Resp ect to th e C2-C3
C,H
Bon d in Mod el Com p ou n d s 1-4a
compound H-2/H-3 C2/H-3 C3/H-2 C1/H-3 C4/H-2 Me/H-3
1
2
3
4
2.9 (2.5)b
7.0 (7.8)
3.0 (2.5)
5.7 (6.2)
0c
-4
-2
-2
-1
-3
-1
-5
2
1
3
2
3
-
-
6
3
3d
4
4
a
The values were determined in C5D5N-CD3OD (1:1) for 1 and
b
2 or in C5D5N-CD3OD (1:2) for 3 and 4. Coupling constants in
c 2,3
parentheses were measured at -33 °C.
J
values were
C,H
d
rounded to single digits because of the digital resolution. Since
the accurate value could not be determined for 3 due to overlapping
3
satellite signals of H2-1, J C1,H-3 was measured with a deacetyl
derivative of 3, of which all 3J H,H values were almost identical with
those of 3, suggesting the close similarity of their conformations.
J -Ba sed Con figu r a t ion An a lysis Usin g Mod el
Com p ou n d s. Model compounds 1-4 were designed and
synthesized to examine the validity of the J -based
method in the configuration analysis of natural products
(substituted hexanes in Table 2, although ideal for
determining the interaction between substituents, were
5
3
2,3
Typ ica l Va lu es of J H,H a n d
w ith Oxygen F u n ction a lities. Electron-withdrawing
substituents such as a hydroxy group affect the magni-
J
for System s
C,H
1
unsuitable due to overlapping signals in H NMR). With
2,3
tude of 3J H,H and
J
. During our configuration analy-
C,H
3
vicinal dihydroxy models 1 and 2, their J H,H values were
ses for acyclic structures appearing in natural products,13
typical of H-2/H-3 gauche and anti, respectively (Table
3). For the threo isomer 1, there are three cases in which
3J H-2,H-3 assumes a gauche value; i.e., a single rotamer
with A-1, A-2 (Figure 3), or alternating A-1/A-2 (Figure
5). Among these, the A-1 rotamer is best fitted for the
3
2,3
we categorized J H,H and
J
values as small, medium,
C,H
or large to fit them into the criteria in Figures 3-6. We
reexamined the categories with respect to the number
of oxygen functionalities and defined the allowances for
3
3
2
small (gauche J H,H and J C,H; anti H/OH for J C,H) and
2
3
observed values of J C2,H-3 and J C4,H-2 (Table 3), which
3
3
2
large (anti J H,H and J C,H; gauche H/OH for J C,H), as
shown in Table 1. The values between small and large
are regarded as medium in Figures 5 and 6. To validate
these criteria, J values were calculated for simple models
using both MM2 and a Karplus-type equation. Haasnoot
et al. proposed a modified Karplus-type equation, in
which the orientation of substituents and their electro-
negativity are taken into account.28 However, since this
equation was based on data from relatively rigid struc-
indicate anti C2-OH/H-3 and gauche C4/H-2, respec-
2
3
tively; in A-2, both J C2,H-3 and J C4,H-2 should be large
and, in the alternating pair of A-1/A-2, both should be
medium. These results indicated the dominant conforma-
tion of 1 to be A-1 in Figure 3. In the other vicinal diol
system, erythro isomer 2 predominantly takes an all-anti
orientation such as B-3 in Figure 3, in which all the
2,3
relevant
J
showed gauche values (Table 3). In these
C,H
threo and erythro dihydroxy systems, an extended (C1/
C4-anti) conformer was predominant.
3
tures such as cyclic compounds, the derived J H,H values
must be adjusted for flexible acyclic systems. Values of
3J H,H for 3-substituted and 3,4-disubstituted hexane
(Table 2) were calculated as a weighted average, in which
fluctuation from a staggered angle was taken into ac-
count. The range of J values in Table 1 appears to be
reasonable when compared with calculated values (Table
2).
In vicinal-methylhydroxy systems, threo model 3 re-
3
2,3
vealed a typical J H-2,H-3 value for gauche and a
J
C,H
value for anti C3-OH/H-2 and Me/H-3, again confirming
that 3 takes mainly the A-1 configuration (Table 3). In
contrast, erythro hydroxy-methyl model 4 showed the
medium value of 3J H-2,H-3 (5.7 Hz), which suggested the
presence of two or more alternating rotamers. The values
3
3
3
of J H-2,H-3, J H-2,C4, and J C1,H-3 suggested that a pair
(28) Haasnoot, C. A. G.; De Keeuw, F. A. A. M.; Altona, C.
Tetrahedron 1980, 36, 2783-2792.
of B-2/B-3 rotamers is predominant (Table 3). However,

Stereochemical Determination of Acyclic Structures
J . Org. Chem., Vol. 64, No. 3, 1999 873
3
2,3
3
2,3
Ta ble 4. J H,H a n d
J
(Hz) w ith Resp ect to th e C3-C4
Ta ble 5. J H,H a n d
J
(Hz) for th e C3′-C7′ P a r t of 5
C,H
C,H
Bon d in Mod el Com p ou n d s 1-4a
fr om Zooxa n th ella toxin a
H-3/
H-4R
H-3/
H-4â
C3/
H-4R
C3/
H-4â
C2/
H-4R
C2/
H-4â
coupled
nuclei
coupled
nuclei
coupled
nuclei
compound
3J H,H
2J C,H
3J C,H
1
2
3
4
5.5
8.9
5.4
8.8
8.2
3.0
8.3
3.9
-4
-6
-4
-7
-6
-3
-7
-3
3
3
-
3
4
2
-
2
H-3′/H-4′
H-4′/H-5′
H-5′/H-6′hb
H-5′/H-6′lb
H-6′h/H-7′
H-6′l/H-7′
4.5
3.0
7.0
7.0
7.5
7.0
C3′/H-4′
C4′/H-3′
C4′/H-5′
C5′/H-4′
C5′/H-6′h
C5′/H-6′l
-1
-1
1
0
-6
-4
C2′/H-4′
2
5
C26′/H-6′h
a
The values were determined in the same conditions as those
in Table 3.
The values were determined in CD3OD. H-6′h and H-6′l are
a
b
those at δ 1.42 and 1.62, respectively.
2
3J Me,H-3 and J C3,H-2 do not accurately correspond to these
rotamers, which implies that the third rotamer B-1 exists
with a comparable population. For configuration analysis,
the assumption of B-2/B-3 rotamers may lead to the
correct answer, but in these cases with three rotamers
of comparable populations, this method should be care-
fully applied. The value of 3J H-2,H-3 of 4 at -33 °C (shown
in parentheses in Table 3) revealed that the population
of the B-1 rotamer decreased, which may allow us to
regard 4 as consisting of alternating B-2/B-3 rotamers.
The other models 1-3 also gave rise to typical values at
low temperatures, which may facilitate the J -based
analysis.29
Force field calculations (MM2* force field, MacroModel)
further supported these results. As expected, for models
1-3, in which the J data indicated C1/C4-anti conformers
to be predominant, the calculation also showed that their
lowest energy appeared at this orientation.22 Conversely,
the calculations revealed that the Me/C4-anti conformer
is most stable in 4, which parallels the results from the
J -based analysis.
extensive spectroscopic analysis of the intact toxins and
the degradation products of periodate oxidation or LiOH
hydrolysis.9 The carboxylic acid 5 obtained from the
hydrolysates was further subjected to stereochemical
elucidation by spectroscopic and synthetic approaches.31
In this study the relative configuration for the C3′-C7′
part was investigated by the J -based configuration
analysis.
Using HETLOC and PS-HMBC, J values were suc-
cessfully determined (Table 5). The rotational conformer
with respect to the C3′-C4′ bond was treated as an
alternating system because of the medium value (4.5 Hz)
2
of 3J H-3′,H-4′. The J values of -1 Hz for both J C3′,H-4′ and
2J C4′,H-3′ can be regarded as medium values (Table 1).
Because the data suggested that H-3′/4′-OH and H-4′/3′-
OH alternated anti and gauche interaction, only the pair
of rotamers shown in Figure 8a, corresponding to A-1 and
A-3 (Figure 5), satisfied all these requirements. Thus, the
diastereomeric relation between C3′ and C4′ was deter-
mined to be 3′R* and 4′R*. With respect to the C4′-C5′
2
For examination of the rotational conformer around a
methine-methylene bond, the C3-C4 bonds in 1-4 are
good models. In all four systems, the extended conformers
are dominant because the protons that are anti to H-3
bond, the values of 2J C4′,H-5′ and J C5′,H-4′ were determined
to be 1 and 0 Hz, respectively, indicating the predominant
anti orientation for 4′-OH/H-5′ and 5′-OH/H-4′ (Figure
8b). Because the specific rotamer can be identified by two
anti substituents, the major rotamer around this bond
was determined to be A-1 in Figure 3, hence showing the
relative stereochemistry to be 4′R* and 5′S*.
3
give rise to large J H,H values of 8.2-8.9 Hz (Table 4). In
3
contrast, the gauche protons show values of J H,H that
vary from 3.0 to 5.5 Hz, the latter of which is too large
for gauche interaction, suggesting the presence of a minor
rotamer with a C2/phenyl-gauche orientation. In model
For the stereochemical relationship between C5′ and
C7′, assignments of diastereotopic methylene protons
were first attempted. With respect to the C5′-C6′ bond,
3
3
1, the values of J C2,H-4R and J C2,H-4â (accurate values
are 2.6 and 3.7 Hz, respectively) reveal that a C2/phenyl-
gauche conformer is present, where H-4â is anti to C2
(Table 4). On the other hand, a pair of 3J H,H values typical
of anti/gauche interactions is observed for 2. This is also
3
h
a medium value of 7 Hz was observed for both
J H-5′,H-6′
3
l
and J H-5′,H-6′ , which suggested the presence of two
alternating rotamers between anti and gauche orienta-
2
h
tions. On the other hand, the large J C5′,H-6′ (-6 Hz) fell
3
3
supported by J C2,H-4R and J C2,H-4â (accurate values are
2.5 and 1.9 Hz, respectively), showing their gauche
orientation. These observations clearly demonstrated
within the range of typical values (Figures 6 and 8c),
indicating the gauche orientation of 5′-OH/H-6′h in both
rotamers. A single pair (D-1/D-3) of rotamers in Figure
6 meets these requirements. Regarding the C6′-C7′
bond, the presence of alternating rotamers was again
implied by medium values of 3J H,H for H-6′h/H-7′ (7.5 Hz)
and H-6′l/H-7′ (7.0 Hz). These data showed that H-6′h/
H-7′ was gauche and H-6′l/H-7′ was anti in one rotamer,
whereas their orientations were exchanged in the other.
2,3
3
that based on
J
and J H,H, two diastereotopic protons
C,H
can be assigned stereospecifically.
Ap p lica tion to Ca r boxylic Acid Moiety Obta in ed
fr om Zooxa n th ella toxin . Zooxanthellatoxin-A (ZT-A)
and ZT-B were isolated as potent vasoconstrictive sub-
stances from a symbiotic marine dinoflagellate Symbio-
dinium sp.9,30 The structures of ZTs were determined by
3
h
In addition, a medium value for J C26′,H-6′ (5 Hz) sug-
gested the C26′/H-6′h orientation to be anti in one
rotamer and gauche in the other (Figure 8d). Thus, C6′-
C7′ was thought to undergo conformational change
corresponding to C-1/C-3 in Figure 6. Assembling these
two diastereomeric relationships led to the elucidation
of C5′/C7′ to be 5′S* and 7′S*. Consequently, the relative
(29) As judged from their relaxation times, molecular motions and
internal rotations of 1-4 seemed to be much faster than those of
natural products with masses around 1000 Da, for which this J -based
method was originally designed. Measurements at low temperature,
which slow these motions to the level of larger molecules, resulted in
decreasing the population of the minor conformers in 1-4. The values
at low temperature in parentheses (Table 3) may reproduce the
corresponding stucture units in larger molecules more precisely.
(30) (a) Nakamura, H.; Asari, T.; Fujimaki, K.; Maruyama, K.;
Murai, A.; Ohizumi, Y.; Kan, Y. Tetrahedron Lett. 1995, 36, 7255-
7258. (b) Nakamura, H.; Asari, T.; Murai, A.; Kondo, T.; Yoshida, K.;
Ohizumi, Y. J . Org. Chem. 1993, 58, 313-314.
(31) Nakamura, H.; Maruyama, K.; Murai, A. Abstracts of Papers,
72th Annual Meeting of J apan Society of Chemistry, Tokyo; J apan
Society of Chemistry: Tokyo, 1997; 3G4 29 (p 1048). Structural studies
by synthesis will be published elsewhere.

874 J . Org. Chem., Vol. 64, No. 3, 1999
Matsumori et al.
F igu r e 8. Configuration analysis of carboxylic acid 5. a: Alternating rotamers with respect to the C3′-C4′ bond. b: The
predominant rotamer with respect to the C4′-C5′ bond. c and d: Pairs of alternating rotamers with respect to the C5′-C6′ and
C6′-C7′ bonds, respectively.
2,3
3
F igu r e 9. Configuration assignment for vicinal methine systems on the basis of
J
and J H,H, and partly NOEs.
C,H
configuration for the C3′-C7′ portion of 5 was determined
as 3′R*, 4′R*, 5′S*, and 7′S*. This assignment has been
confirmed by the synthesis of the ozonolysis product
corresponding to C1′-C8′.31
Figure 9 is a schematic diagram showing the assign-
ment of the diastereomeric relationship of vicinal methine
systems. In the first step, the six possible rotamers or
the alternating pairs arising from threo and erythro
stereoisomers are classified into three cases according to
3
Discu ssion
their size of J H,H values. For vicinal dioxygenated
systems, the J value becomes smaller as shown in Table
Besides the compounds described above, this J -based
analysis has been used for the stereochemical determi-
nation of several natural products,13 which resulted in
the elucidation of 40 stereogenic centers. These successful
applications demonstrate that this method is reliable for
acyclic structures with hydroxyl (alkoxyl) and methyl
groups. Its applicability to the other substituents, such
as halogen, nitrogen, carbonyl, and bulky functionalities,
is currently being investigated and will be published in
due course.
3
1. When J H,H shows a typical anti orientation, NOE
2,3
experiments are necessary, because
J
does not
C,H
provide any further information on configuration. In
3
other cases with J H,H values less than 8 Hz (7 Hz), 2,3J C,H
values allow us to assign configurations and conforma-
tions according to Figures 3, 5, and 9.
There are three exceptions in which the 2,3J C,H analysis
fails to determine configuration as described earlier: one
is the case in which the two alternating conformations

Stereochemical Determination of Acyclic Structures
J . Org. Chem., Vol. 64, No. 3, 1999 875
F igu r e 10. Configuration assignment for 1,3-methine systems.
have an H/H-gauche orientation as A-1/A-2 or B-1/B-2
in Figure 5, and the others are those in which three
staggered rotamers coexist with comparable populations
or where a rotational conformer deviates from the stag-
gered one. These three cases, to our knowledge, rarely
occur in acyclic structures of natural products.
we reduced the number of synthetic diastereomers needed
to 2 from 128 possible candidates using this J -based
method.13c
NOE analysis is an excellent method for conformation
and configuration assignments of cyclic compounds as
seen in numerous examples in structure studies of
natural products. Conversely, the J -based method works
better for acyclic systems than for cyclic compounds, in
which deviations from the staggered rotamers frequently
occur. Therefore, the complementary use of both the
J -based method and NOE analysis greatly facilitates the
stereochemical determination of complicated organic
compounds.
A schematic diagram of the configuration assignment
for 1,3-methine systems is given in Figure 10. Prochiral
protons on the methylene are first designated as Hh and
Hl according to their 1H chemical shifts, and the rota-
tional conformation is determined for one CH-CH2 bond
3
2,3
using J H,H and
J
(see Figures 4, 6, and 10). In the
C,H
next step, the rotamer for the other CH2-CH bond is
examined in the same way. The four possible cases, 1,3-
syn with â-Hh/R-Hl, 1,3-anti with â-Hh/R-Hl, 1,3-anti with
R-Hh/â-Hl, and 1,3-syn with R-Hh/â-Hl, can be discrimi-
nated, thus leading to the elucidation of the 1,3-methine
diastereomeric relationship in combination with the
stereospecific assignment of the methylene protons.
Exp er im en ta l Section
Gen er a l P r oced u r e. Unless otherwise noted, all reagents
and NMR solvents were obtained from standard commercial
sources and used without further purification. Column chro-
matography was carried out on silica gel (E. Merck, 70-230
mesh) with use of hexanes-EtOAc as a mobile phase.
This J -based configuration analysis is applicable to all
organic compounds bearing asymmetric centers, which
include not only natural products but synthetic molecules
such as intermediates for natural product syntheses. As
2,3
Meth od s for Mea su r in g
J
. Geminal and vicinal
C,H
carbon-proton coupling constants (2,3J C,H) were measured
using two methods. The 2D hetero half-filtered TOCSY (HET-
LOC) experiment, which was originally proposed by Otting and
Wu¨thrich,19 is one of the most powerful methods for measuring
reported previously,13a the amount of a sample necessary
2,3
J
. In this study the pulse sequence reported by Leibfritz
C,H
2,3
to determine
J
using the natural abundance of the
C,H
was used, which consisted of a BIRD-half-filter-TOCSY
succession.19b Another method used in this study was phase-
sensitive HMBC reported by Zhu and Bax.20 All the spectra
were measured with a J EOL A-500 spectrometer (500 MHz)
13C isotope is about 10 µmol when a modern NMR
instrument of 400-600 MHz is used; this amount is
comparable to that used in most structure elucidations.
Compared to conventional approaches consisting of chemi-
cal degradation and synthesis of all possible diastereo-
mers, our method greatly diminishes labor and time. As
demonstrated in the structure studies on maitotoxin,13b-d,32
(32) The relative configuration of side chains of maitotoxin has been
independently elucidated by Harvard group, who reached the identical
conclusion: Zheng, W.; DeMattei, J . A.; Wu, J .-P.; Duan, J . J .-W.; Cook,
L. R.; Oinuma, H.; Kishi, Y. J . Am. Chem. Soc. 1996, 118, 7946-7968.

876 J . Org. Chem., Vol. 64, No. 3, 1999
Matsumori et al.
in the phase-sensitive mode proposed by States.33 3J H,H values
were extracted from 1D 1H NMR and 2D E.COSY spectra.34
For the HETLOC spectra of model compounds 1-4, the MLEV-
17 spin-lock periods including a trim pulse (2.5 ms) were set
by SiO2 column chromatography. After reduction of each
isomer to the alcohol and then acetylation of the resultant
alcohol, the olefin was dihydroxylated with osmium tetraoxide
to yield the corresponding erythro- and threo-dihydroxy com-
pounds from the Z- and E-isomers, respectively. The hydroxy-
methyl models (3 and 4) were prepared from the phenylallyl
alcohol obtained as an intermediate for the synthesis of 1 and
2 by epoxidation with 3-chloroperbenzoic acid and reductive
cleavage with MeLi in the presence of CuI to give 1,3-diol with
excellent regioselectivity. After regioselective acetylation, both
diastereomers were subjected to the NMR measurements.
4-P h en yl-th r eo-2,3-d ih yd r oxyet h yl Acet a t e (1). The
preparation of this compound is detailed in the Supporting
2
3
for 30 ms to measure J C,H and 60 ms to measure J C,H. The ∆
in the half-filter was set at 3.45 ms and 4 ms, which was
optimized for CH/CH2 and CH3, respectively. The nondiagonal
version of HETLOC19b was measured for 3 and 4, in which
some of cross-peaks overlapped on diagonal peaks. The HET-
LOC spectra of 1-4 were measured for 9 h at 27 °C with a
data size of 4K(F2) × 128(F1) points for the spectral width of
3000 Hz by 3000 Hz. The phase-sensitive (PS)-HMBC spectra20
were recorded for 9 h with the delay (∆) set at 40 ms with a
data size of 2K(F2) × 128(F1) points for the spectral width of
3000 Hz (1H) and 25000 Hz (13C). For HETLOC and PS-HMBC,
2-fold zero-filling was conducted for both dimensions to give
the digital resolution in F2 of 0.38 for HETLOC and 0.75 Hz
for PS-HMBC.
1
Information. H NMR (500 MHz, CD3OD-C5D5N, 1:1; For δH
of 1-4, CHD2OD was taken as the reference at δ 3.30) δ 7.26-
7.30 (m, 2H), 7.17-7.20 (m, 2H), 7.10 (m, 1H), 4.33 (dd, 1H),
4.27 (dd, 1H), 3.93 (ddd, 1H), 3.85 (dd, 1H), 3.01 (dd, 1H), 2.89
(dd, 1H), 1.88 (s, 3H). 13C NMR (125 MHz, C5D5N-CD3OD,
1:1; For δC of 1-4, CD3OD was taken as the reference at δ
49.0) 172.0, 140.7, 130.5, 129.2, 127.0, 73.7, 71.7, 67.4, 40.8,
21.0. HRMS (FAB): calcd for C12H16O4Na 247.0946, obsd
247.0938.
The carboxylic acid 5 was obtained from zooxanthellatoxins
by basic hydrolysis with LiOH.9 The sample (8 mg) was
dissolved in 0.5 mL of CD3OD and subjected to NMR measure-
3
ments. J H,H values were determined from 1D 1H NMR and
2D E.COSY,34 the latter of which was carried out with a data
4-P h en yl-er yth r o-2,3-d ih yd r oxyeth yl Aceta te (2). The
preparation of this compound is detailed in the Supporting
Information. 1H NMR (500 MHz, CD3OD-C5D5N, 1:1) δ 7.28-
7.30 (m, 2H), 7.16-7.19 (m, 2H), 7.10 (m, 1H), 4.52 (dd, 1H),
4.32 (dd, 1H), 3.92 (ddd, 1H), 3.84 (dd, 1H), 3.19 (dd, 1H), 2.76
(dd, 1H), 2.15 (s, 3H). 13C NMR (125 MHz, C5D5N-CD3OD,
1:1) 172.2, 140.9, 130.8, 129.1, 126.9, 74.3, 73.5, 67.6, 41.0,
21.0. HRMS (FAB): calcd for C12H16O4Na 247.0946, obsd
247.0963.
4-P h en yl-th r eo-3-d ih yd r oxy-2-m eth yleth yl Aceta te (3).
The preparation of this compound is detailed in the Supporting
Information. 1H NMR (500 MHz, CD3OD-C5D5N, 2:1) δ 7.20-
7.24 (m, 4H), 7.13 (m, 1H), 4.08 (dd, 1H), 3.97 (dd, 1H), 3.92
(ddd, 1H), 2.77 (dd, 1H), 2.72 (dd, 1H), 1.92 (s, 3H), 1.83 (m,
1H), 0.97 (s, 3H). 13C NMR (125 MHz, CD3OD-C5D5N, 2:1)
172.2, 140.9, 130.3, 129.3, 127.0, 72.9, 68.1, 42.2, 38.2, 20.9,
10.7. HRMS (FAB): calcd for C13H18O3Na 245.1154, obsd
245.1169.
4-P h en yl-er yth r o-3-d ih yd r oxy-2-m eth yleth yl Aceta te
(4). The preparation of this compound is detailed in the
Supporting Information. 1H NMR (500 MHz, CD3OD-C5D5N,
2:1) δ 7.20-7.25 (m, 4H), 7.13 (m, 1H), 4.28 (dd, 1H), 4.04 (dd,
1H), 3.72 (ddd, 1H), 2.86 (dd, 1H), 2.64 (dd, 1H), 1.97 (s,
3H),1.00 (d, 3H). 13C NMR (125 MHz, CD3OD-C5D5N, 2:1)
172.4, 141.0, 130.6, 129.2, 127.0, 75.0, 67.3, 41.9, 39.4, 21.0,
14.7. HRMS (FAB): calcd for C13H18O3Na 245.1154, obsd
245.1140 as (M + Na)+.
size of 4K (F2) × 512 (F1) points for a spectral width of 3000
Hz, and a squared sine-bell window function shifted by -2π/7
2,3
was applied to both interferograms.
J
values were mea-
C,H
sured by HETLOC experiments in the following conditions;
2
BIRD delay, 470 ms; spin-lock period, 30 ms for J C,H and 60
3
ms for J C,H with each 2.5 ms trim pulses; ∆, 3.45 ms, data
points, 2K (F2) × 128(F1); spectral width, 3000 Hz by 3000 Hz;
2-fold zerofilling to both dimensions. Unless otherwise noted,
a mixed solvent of pyridine-methanol (1:1) was used for NMR
measurements to minimize the effect of intramolecular hy-
drogen bonding.
3
Rota tion a l Con for m er s a n d J H,H. The conformations of
rotamers in Table 2 and footnote 22 for model compounds 1-4
were calculated using MM2* force field on MacroModel. The
populations of these rotamers were estimated from a calculated
potential curve. On the basis of these populations, the weighted
3
average values of J H,H in Table 2 were calculated using the
modified Karplus equation.28 The averaged values for gauche
60°, anti 180°, and another gauche 300° were obtained from
conformers with the dihedral angles between 0° and 120°,
those between 120° and 240°, and those between 240° and
360°, respectively. For calculations of the conformations of
model compounds 1-4 in footnote 22, a potential curve with
respect to the C2-C3 bond was obtained under non-hydrogen-
bonding conditions since NMR data of 1-4 were collected in
highly polar solvents. In these molecular mechanics calcula-
tions, the C1-C2 and C3-C4 bonds of 1-4 were supposed to
have an extended conformation. The same conditions were set
for the calculations of subsituted hexanes in Table 2.
Ack n ow led gm en t. This study was supported by a
Grant-in-Aid from the Ministry of Education, Science,
Sports and Cultures of J apan, and by the “Research for
the Future” program of the J apan Society for the
Promotion of Science (J SPS-RFTF96I00301). We are
grateful to Drs. T. Nonomura and M. Sasaki in our
laboratory for discussions, and Messrs. H. Kumaki, H.
Utsumi, and T. Hinomoto (J EOL Co. LTD) for their
assistance in NMR measurements.
Syn th esis of Mod el Com p ou n d s. Model compounds were
designed to reproduce rotational conformers for vicinal dihy-
droxy and vicinal hydroxy-methyl systems occurring in natural
products. Syntheses of model compounds 1 and 2 for models
of 1,2-dihydroxy system were started with the coupling of
phenylacetaldehyde with methyl (tripheylphosphoranylidene)-
acetate in methanol to furnish methyl 4-phenyl-2-butenoate
with both E- and Z-geometry, which could be readily separated
Su p p or tin g In for m a tion Ava ila ble: Selected spectra
used for configuration assignments of 5; the synthetic details
of the model compounds 1-4. This material is available free
of charge via the Internet at http://pubs.acs.org.
(33) States, D. J .; Haberkorn, R. A.; Ruben, D. J . J . Magn. Reson.
1982, 48, 286-292.
(34) (a) Griesinger, C.; Sørensen, O. W.; Ernst, R. R. J . Am. Chem.
Soc. 1985, 107, 6394-6396. P.E. COSY (Mueller, L. J . Magn. Reson.
1987, 72, 191-196) could be used in lieu of E. COSY.
J O981810K