Highly efficient preparation of selectively isotope cluster-labeled long chain fatty acids via two consecutive Csp3-Csp3 cross-coupling reactions

Article abstract of DOI:10.1021/ol4036159

An efficient synthesis involving two copper-catalyzed alkyl-alkyl coupling reactions has been designed to easily access doubly isotope-labeled fatty acids. Such NMR- and IR-active compounds were obtained in excellent overall yields and will be further used for determining the conformation of an alkyl chain of lipidic biomolecules upon interaction with proteins.

Full text of DOI:10.1021/ol4036159

Letter
pubs.acs.org/OrgLett
Highly Efficient Preparation of Selectively Isotope Cluster-Labeled
3
3
Long Chain Fatty Acids via Two Consecutive Csp −Csp Cross-Coupling
Reactions
Sebastien Lethu,‡,§ Shigeru Matsuoka,*,‡,§ and Michio Murata*,‡,§
́
§
‡JST ERATO, Lipid Active Structure Project and Project Research Center for Fundamental Sciences, Osaka University, 1-1
Machikaneyama, Toyonaka, Osaka 560-0043, Japan
S
* Supporting Information
ABSTRACT: An efficient synthesis involving two copper-
catalyzed alkyl−alkyl coupling reactions has been designed to
easily access doubly isotope-labeled fatty acids. Such NMR-
and IR-active compounds were obtained in excellent overall
yields and will be further used for determining the
conformation of an alkyl chain of lipidic biomolecules upon
interaction with proteins.
In this study, by taking as an example [n, n+1-13C2, n, n
onformation of a saturated alkyl chain is one of the
+1-2H2] doubly labeled stearic acid (Figure 1), we aimed to
Cfundamental concepts in organic chemistry, where three
preferable states, anti (T), gauche+ (G+), and gauche− (G−),
are adopted.1
The conformation of flexible molecules is strongly correlated
with the macroscopic physical properties of hydrocarbon
assemblages containing long alkyl chains such as polymers,
Figure 1. [n, n+1-13C2, n, n+1-2H2] stearic acids.
detergent micelles, and biomembranes.2 Therefore, experimen-
tal methods to explore alkyl chain conformation have been
developed by means of vibration spectroscopy, neutron
diffraction, and NMR, combined with isotopically labeled
compounds.3 In these methodologies, 13C and/or 2H are
introduced at the methylene unit concerned to enable selective
observation by giving separable signals from large background
ones. Specifically, the 13CH2 or CD2 group provides
information about the chain mobility at the labeled methylene
or total information for two 13CH2−C or two CD2−C bonds
respectively.
establish a highly efficient route for the preparation of site-
selectively cluster-labeled linear hydrocarbon chains involving
3
3
two consecutive Csp −Csp cross-coupling reactions.
To establish the standardized labeling scheme, we designed
two common intermediates 10 and 11, which could be derived
from bromoacetic acid.5 In this study, we adopted [2-13C]-
bromoacetic acid twice to form a 13C−13C spin pair avoiding
the use of expensive 13C2 labeled precursors and constructed
the threo configuration for C2/C3 of the common intermediate
by the syn-selective deuteration of E-olefin (Scheme 1).
On the other hand, when investigating a dihedral angle of a
particular C−C bond, introduction of a single 13CH2 or CD2
group is not sufficient. For obtaining the segmental dihedral
angle in such dynamic molecules as fatty acids, it is a requisite
to determine the relative orientation between two neighboring
methylene groups by introducing multiple isotopes. The
minimum structure for defining an alkane chain conformation
is a butylene unit (C−CH2−CH2−C), where C2 and C3
should be CHDs with defined configurations, when aiming to
discriminate the G+ and G− orientations.
After activation as an acid chloride, the addition of ethanol
afforded ester 1, which was then immediately converted into
the corresponding phosphonate 2 by Arbuzov reaction with
triethylphosphite. Compound 2 was subsequently subjected to
Horner−Wadsworth−Emmons (HWE) olefination with
benzyloxyacetaldehyde, providing the monolabeled conjugated
ester 3, which was then reduced by diisobutylaluminium
hydride (DIBAL-H) to generate alcohol 4. Thus, a second
protecting group was required that would be sufficiently robust
for the conditions of our strategy while allowing an easy
separation of the two aldehydes after degradation by ozonolysis.
For this reason, we chose the tert-butyldiphenylsilyl (TBDPS)
group that is stable under various conditions6 and more
lipophilic than benzyl. After protection of the hydroxyl group of
Accordingly, a standardized cluster-labeling scheme for a
saturated alkyl chain can be separated into two stages: a
versatile preparation of the butylene common intermediate that
4
2
can cover all combinations of 13C and H labeling patterns,
and the practical incorporation of the cluster-labeled common
intermediate into an alkyl chain in high yield retaining the
configuration of C2/C3.
Received: December 13, 2013
© XXXX American Chemical Society
A
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Scheme 1. Synthesis of the Common Synthons 10 and 11
Table 1. Optimization of Alkyl−Alkyl Coupling
a
entry
starting material
CuCl2 (mol %)
acetylene (mol %)
R
RMgBr (equiv)
yield (%)
XX
1
2
3
4
10
10
11
11
2
5
5
5
10
20
20
20
C6H13
C6H13
C6H13
C14H29
1.5
2
50
80
96
98
12
12
12
13
2
2
a
Isolated yield.
4 as a corresponding TBDPS ether, the resulting olefin 5 was
ozonolyzed to labeled aldehyde 6. Using the previously
synthesized phosphonate 2, a second HWE reaction provided
ester 7 in a separable mixture E/Z 98:2 with the two positions
of the conjugated olefin labeled with 13C. From olefin (E)-7,
deuteration of the double bond was carried out with
Wilkinson’s catalyst to give labeled ester 8 in almost
quantitative yield with full deuterium incorporation.7,8 The
diastereomeric purity of this reaction was found to be
satisfactory in favor of the cis/threo compound.8 A second
reduction with DIBAL-H afforded hydroxyl 9. Finally, two
desired common synthons were obtained either as a
brominated compound, 10, or as a tosylated one, 11, in overall
yields, after 10 steps, of 48% and 47% respectively.
tosylated 11 proved to be a more suitable precursor than 10 as
the coupling product was obtained under the same conditions
in almost quantitative yield with both short (C6H13) and long
(C14H29) alkyl chains (entries 3 and 4). As depicted in Scheme
2, two more steps from 13, i.e. desilylation by tetrabutylalu-
Scheme 2. Final Steps for 2,3-Labeled Stearic Acid 15
minium fluoride (TBAF) and Jones’ oxidation, led to the
preparation of 15, one of the labeled stearic acids, with a 66%
overall yield from the common synthon 11.
With those two labeled intermediates in hand, we envisaged
that further extension of an acyl chain should be achieved
through a direct cross-coupling between the alkyl electrophile
10 or 11 and an alkyl metal. Recently, remarkable advances
While the tail part was successfully installed, we found that
the coupling of the fatty acyl head was more challenging.
Because of the propensity of carboxylic acids to bind metals,11
direct coupling is delicate12 and the acid moiety is often
protected as an ester. Various methodologies involving alkyl-
metal bearing functional groups, such as Suzuki−Miyaura9,13
and Negishi9,14 couplings, have been elaborated. However, no
reaction occurred between 10 (or 11) and the appropriate ester
as a coupling partner.13b,c,14a This forced us to adopt an
alternative strategy where a protected hydroxyl is to be
converted to the expected fatty acids by a routine deprotection
and oxidation sequence. In order to accomplish such a coupling
reaction, the previously used methodology was employed using
freshly prepared Grignard reagents bearing a protected
hydroxyl group. To our knowledge, this coupling reaction
involving Grignard reagents with heteroatoms has not been
3
3
have been brought about to accelerate applications of Csp −Csp
cross-coupling reactions for organic synthesis. Alkyl electro-
philes containing β-hydrogen atoms were originally thought to
be unsuitable. Nevertheless, several groups have developed
3
3
powerful Csp −Csp Suzuki−Miyaura, Negishi, or Kumada−
Corriu couplings.9 Among these, we adopted a method based
on a copper-catalyzed Kumada−Corriu coupling developed by
Kambe and co-workers,10 to first link the tail of the stearic acid.
As shown in Table 1, the initial attempt gave a promising
50% yield (entry 1). To improve the yield, we optimized the
reaction conditions by increasing the amount of the catalyst,
additive, and organomagnesium reagent. After such adjust-
ments, the yield rose to a good 80% (entry 2) with brominated
10. Nevertheless, in contrast to the report by Kambe,10
B
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described to date. A survey over several possible protecting
groups15 revealed three candidates to be promising: tetrahy-
dropyran (THP), p-methoxybenzyl (PMB), and benzyl (Bn) as
shown in Table 2. After two steps, THP afforded the desired
After synthesizing the two stearic acids 15 and 23 containing
the isotope labels near both of the acyl termini, we had all the
tools in hand to produce fatty acids where the labeled moiety
would be located in the central portion. However, the order of
the two couplings had to be reexamined. Due to the volatility of
the intermediates if the tail of the fatty acid was coupled first,
we had to carry out the troublesome separation of the desired
compound from the byproducts after the second coupling with
protected-hydroxyl Grignard reagents. Thus, it quickly came to
light that the head of the fatty acid had to be coupled first.
On this basis, pure products 25a−c were obtained in
satisfactory yield after coupling the protected-hydroxyl chain
and deprotection of TBDPS (Scheme 4). Tosylation and the
3
3
Table 2. Survey of Different Protecting Groups for Csp −Csp
a
Coupling
b
entry
PG
compound
yield (% after 2 steps)
Scheme 4. Synthesis of “Middle”-Labeled Stearic Acids 29a−
c
1
2
3
THP
PMB
Bn
16
43
93
96
17
25b
a
Reaction conditions: 11 (0.5 mmol), CuCl2 (5 mol %), methyl-
phenylacetylene (20 mol %), Grignard reagent (1 mmol), THF (0.5
b
mL), 0 °C to rt, 1 h. Isolated yield.
compound 16 in only 43% yield, much lower than our
expectations. The more robust PMB and Bn groups revealed
excellent yields in the two-step sequence. Because Bn exhibited
a better result, we chose it as the protecting group for the rest
of our study.
The synthesis of 16,17-labeled stearic acid 23 was realized by
the same strategy with the appropriate chain length (Scheme
3
3
3). As expected, the Csp −Csp coupling and subsequent
Scheme 3. Synthesis of 16,17-Labeled Stearic Acid 23
subsequent second coupling afforded the desired protected
stearyl alcohols 27a−c in an efficient way. Finally, debenzyla-
tion furnished alcohols 28a−c, which were oxidized to
complete the synthesis of the fatty acids 29a−c in overall
yields of, respectively, 68%, 68%, and 65% from 11.
In summary, from [2-13C]-bromoacetic acid and via a
versatile common precursor, we established a simple and
efficient route for accessing a range of stearic acids bearing
variable labeled positions with a 13CHD-13CHD moiety in their
3
aliphatic chain. This methodology involves two powerful Csp −
3
Csp couplings that could also be applied for the synthesis of
other labeled fatty acids or more complex molecules possessing
an alkyl chain linker. Their application in the investigation on
protein−lipid interactions is now in progress by means of solid
NMR and will be reported in due course.
desilylation afforded the labeled monoprotected diol 19 in
90% yield after two steps. Activation as tosylate 20 and
subsequent nucleophilic reduction afforded the protected
alcohol 21. Deprotection of the Bn group to give 16,17-labeled
stearyl alcohol 22 was found to be more efficient and much
faster with a Lewis acid such as boron trichloride than with an
oxidant such as dichlorodicyanoquinone (DDQ) or by classical
catalytic hydrogenation with Pd/C. Finally, Jones’ oxidation led
to the desired stearic acid 23 in 65% overall yield after six steps
from 11.
ASSOCIATED CONTENT
* Supporting Information
■
S
Experimental procedures, full spectroscopic data, and copies of
1H and 13C NMR spectra for labeled stearic acids and their
intermediates are provided. This material is available free of
charge via the Internet at http://pubs.acs.org.
C
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AUTHOR INFORMATION
Corresponding Authors
■
*E-mail: matsuokas11@chem.sci.osaka-u.ac.jp.
*E-mail: murata@chem.sci.osaka-u.ac.jp.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
This work was supported in part by JSPS KAKENHI Grant
Numbers 24681045, 24651244, and 25242073.
■
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■
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