Linking of Alcohols with Vinyl Azides

Article abstract of DOI:10.1021/acs.orglett.6b00116

A protocol to link alcohols with vinyl azides has been established through fluoro- or bromo-alkoxylation of vinyl azides to provide α-alkoxy-β-haloalkyl azides. A series of primary and secondary alcohols including natural products and their derivatives such as sugars and steroids were successfully anchored with vinyl azides. The as-prepared cyanine dye linked testosterones were capable of rapid cell membrane imaging in real time.

Full text of DOI:10.1021/acs.orglett.6b00116

Letter
pubs.acs.org/OrgLett
Linking of Alcohols with Vinyl Azides
Yi-Feng Wang, Ming Hu, Hirohito Hayashi, Bengang Xing,* and Shunsuke Chiba*
Division of Chemistry and Biological Chemistry, School of Physical and Mathematical Sciences, Nanyang Technological University,
Singapore 637371, Singapore
S
* Supporting Information
ABSTRACT: A protocol to link alcohols with vinyl azides has
been established through fluoro- or bromo-alkoxylation of
vinyl azides to provide α-alkoxy-β-haloalkyl azides. A series of
primary and secondary alcohols including natural products and
their derivatives such as sugars and steroids were successfully
anchored with vinyl azides. The as-prepared cyanine dye linked
testosterones were capable of rapid cell membrane imaging in
real time.
Scheme 1. Installation of Azido Unit onto Alcohols
wing to their rich and diverse chemical reactivity, organic
azides are recognized as versatile synthons for the
O
preparation of nitrogen-containing compounds, in particular,
nitrogen heterocycles.1 The 1,3-dipole character of organic
azides is most commonly used to construct heterocyclic
frameworks through [3 + 2]-cycloaddition reactions with a
range of unsaturated bonds such as alkynes, alkenes, and
nitriles. Among them, the azide−alkyne cycloaddition2 to form
1,2,3-triazoles is actively applied for various areas such as
medicinal chemistry,3 chemical biology,4 and material sciences.5
For example, the azide−alkyne [3 + 2]-cycloaddition can be
used to label natural products, drugs, and biomolecules in vitro
and/or in vivo for bioimaging, drug delivery, or bioconjugation
purposes.6 For these opportunities, installation of the azido unit
at the appropriate position of the targeted materials is
necessarily indispensable. Hydroxyl groups (alcohols) are
prevalent in various molecules of biologically and medicinally
importance.7 Chemical conversion of alcohols into azides is
thus commonly implemented by substitution of the activated
hydroxyl group with the azido ion.8,9 Use of 2-azido-1,3-
dimethylimidazolinium salts9a or diarylphosphoryl azides with
DBU as a base9b−d enables direct azidation of alcohols (Scheme
1A).10 On the other hand, the protocol to install the azide unit
onto the alcohols would be an alternative strategy to use
alcohols for further applications.11 We conceived that such a
linking method could be achieved through electrophilic
halogenation of vinyl azides 1 in the presence of alcohols 2,
in which the initially formed α-haloiminodiazonium ions A12
could be trapped by external alcohols 2 (Scheme 1B).
Neighboring group participation of the installed halogen
atom (X) to the CN bond might result in stabilization of
iminodiazonium ions A through halonium ions B, preventing
formation of nitrilium ions by migration of substituent R.12a,b
Herein, we demonstrate this concept with the fluoro- or
bromo-alkoxylation of vinyl azides. Alcohols are uniquely
connected to haloalkyl azide moieties in the resulting
unprecedented aminal-type structure. Concise transformation
of the products, α-alkoxy-β-haloalkyl azides to halomethyl
nitrogen-heterocycles was demonstrated using thermal intra-
molecular [3 + 2]-azide-alkene/alkyne cycloaddition. More-
over, tagging of different types of fluorescent probes to
testosterone-linked azide was implemented by the click
reactions. The as-prepared fluorescent testosterone derivatives
having cyanine probes exhibited unique properties for rapid
imaging of cell membranes in real time.
Screening of a variety of halonium ion sources in the
reactions of vinyl azide 1a with cinnamyl alcohol (2a) revealed
that 1-(chloromethyl)-4-fluoro-1,4-diazoniabicyclo[2.2.2]-
octane bistetrafluoroborate (Selectfluor)13,14 and 2,4,4,6-tetra-
bromocyclohexa-2,5-dienone (TBCO)15 worked well to give
the desired α-alkoxy-β-haloalkyl azides 3aa (for fluoride) and
4aa (for bromide) in 80% and 82% yields, respectively (Scheme
2, conditions A and B; see the Supporting Information for
optimization of the reaction conditions). Having these
Received: January 13, 2016
© XXXX American Chemical Society
A
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a,b
Scheme 2. Substrate Scope on Alcohols
a
Unless otherwise noted, the reactions were carried out by treatment of alcohols 2 and vinyl azide 1a (1.2−2.2 equiv) with Selectfluor (1.2−2.2
equiv) in CH3CN (conditions A) or TBCO (1.2−2.2 equiv) in CH2Cl2 (conditions B) at 0 °C to room temperature under a N2 atmosphere.
b
c
1
Isolated yields and diastereoselectivity (dr) determined by H and 19F NMR measurements were recorded above. Isolated yields of the linking
d
e
product of the secondary hydroxyl group. The reaction was conducted in CH3CN−CH2Cl2 (5:1). The reaction was conducted in CH2Cl2−THF
(1:1).
protocols in hand, the generality of alcohols 2 to link with vinyl
azide 1a was next examined (Scheme 2; see the Supporting
Information for the substituent compatibility on vinyl azides
1b−i for these linking protocols). Both alkenyl (2b with aryl-
substituted alkene and 2c with terminal alkene) and alkynyl
(2d−f with both terminal and internal alkynes) alcohols reacted
smoothly, providing the corresponding β-haloalkyl azides 3 and
4 in good yields. Other functional groups such as N-tosylindolyl
(for 2g), phenylsulfonyl (for 2h), diethyl phosphoryl (for 2i),
and pinacolboryl (for 2j) moieties were all compatible under
the present reaction conditions. In the reactions of diol 2k
having primary and secondary hydroxyl groups, the primary
one was preferentially linked (for 3ak and 4ak).16 For linking of
3,5-dimethylphenol (2l), the bromination protocol with TBCO
only worked to give 4al, while the reaction was sluggish due to
overbromination. We next investigated tagging natural products
and their derivatives having hydroxyl group(s) in their
structure.17 Glucose derivative 2m18 and rac-menthol (2n)
were readily employed for the present linking protocol. It was
found that nopol (2o) and nerol (2p) having reactive alkenes
(strained and/or electron rich) are not applicable for the
linking protocol with Selectfluor, while that with TBCO
delivered the corresponding azides 4ao and 4ap in good yields.
Steroids such as androsterone (2q) and testosterone (2r) could
be anchored well with both protocols, while the bromination
method was only amenable for linking of cholesterol (2s) and
betulin (2t) having an electron-rich alkene. In the reaction of
betulin (2t), its primary hydroxyl group was selectively linked,
while the secondary one next to the quaternary carbon was kept
intact due to steric hindrance.
Intramolecular 1,3-dipolar azide−alkene cycloaddition1 of
alkenyl alcohol-linked azides under thermal reaction conditions
afforded unique nitrogen heterocycles, 2-(halomethyl)-2,5-
dihydrooxazoles (for 5aa and 6aa) and 2-(fluoromethyl)-5,6-
dihydro-2H-1,3-oxazine 7ab, in good yields (Scheme 3).
Similarly, intramolecular 1,3-dipolar azide−alkyne cycloaddition
Scheme 3. Intramolecular Azide−Alkene/Alkyne
Cycloaddition
a
Two-step yield from vinyl azide 1a (see the Supporting Information).
B
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reactions of 3ae and 3af were also performed to give biyclic
triazoles 8ae and 9af, respectively, in good yields.
that the resulting triazole-linked aminal-type moiety was fairly
stable under physiologically relevant serum conditions (pH 7.4)
and in aqueous phosphate buffer solution (pH 6.6−3.0) (see
the Supporting Information). Moreover, MTT assay showed
that there was little cell viability impaired during the process of
cell staining, indicating less toxicity and suitable biocompati-
bility of the triazole-linked aminal-type moiety in living cell
applications (see the Supporting Information).
Human embryonic kidney 293 (HEK293) cells were then
incubated with 10−12 (2.0 μM). The capability of these dye-
linked testosterones 10−12 as a membrane staining probe was
examined by confocal microscopy. As shown in Figure 1 and
Figure S2, while fluorescence of dansyl probe 10 was found
inside the cells rather than on the membrane surface probably
due to the rapid internalization of dansyl probe 10 into the cells
(Figure 1a), the strong fluorescence signals of 11 and 12 were
found on the cellular membrane (Figures 1c,d). These
observations are most likely attributed to the difference of
the charge properties between dansyl-linked testosterone 10
and cyanine-linked ones 11 and 12. The fluorescence signal
overlap of dansyl probe 10 and organelle specific dyes (Figure
1b with BODIPY TR ceramide, one commonly used Golgi
apparatus tracker) indicated more prominent localization in the
Golgi apparatus than other major organelles (e.g., mitochondria
and lysosomes) (Figure S4). Further investigation is ongoing to
account for such interesting differences between dansyl probe
10 and cyanine-linked probes 11 and 12.
We next explored the potential applicability of the present
linking protocol in bioimaging. As indispensable important cell
components, steroids have been found to maintain the cell
membrane’s integrity and fluidity as well as to secure the
membrane-associated biological activities in living cells. It has
been reported that several dye-labeled steroid analogues can be
used for specific membrane staining.19,20
Thus, testosterone-linked azide 3ar was chosen, and the click
reactions of 3ar and alkynes were tethered to fluorescent
probes such as N-propargyldansylamide, and azadibenzocy-
clooctyne−cyanine dyes (DBCO−Cy3 and DBCO−Cy5) were
implemented to prepare 10−12 (Figure 1). It is noteworthy
Further studies to develop modular and selective linking
protocols of other essential functional groups, such as amines
and thiols with vinyl azides, are currently underway in our
laboratory. We envision that such a unique protocol would
enable promising applicability for medicinal and biological
studies.
ASSOCIATED CONTENT
■
S
* Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.6b00116.
Experimental procedures and characterization of new
compounds (PDF)
X-ray crystallographic data for compound 5aa (CIF)
AUTHOR INFORMATION
■
Corresponding Authors
*E-mail: bengang@ntu.edu.sg.
*E-mail: shunsuke@ntu.edu.sg.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was supported by funding from Nanyang
Technological University and the Singapore Ministry of
Education (Academic Research Fund Tier 2: MOE2013-T2-
1-060 for S.C.). We thank Dr. Yongxin Li and Dr. Rakesh
Ganguly (Division of Chemistry and Biological Chemistry,
Nanyang Technological University) for assistance with X-ray
crystallographic analysis. We also acknowledge Mr. Yichen
Ding and Prof. Liang Yang at the Singapore Centre on
Environmental Life Sciences Engineering (SCELSE), Nanyang
Figure 1. Synthesis of dye-linked testosterones 10−12 and
fluorescence imaging of live HEK293 cells by incubation of 10−12
at 37 °C, respectively. Cells incubation with (a) dansyl 10 (2 μM) for
5 min, λex = 405 nm; (b) fluorescence signals overlap of dansyl 10 with
BODIPY TR ceramide for the Golgi apparatus staining; (c) Cy-3 11
(2.0 μM) for 2 min, λex = 514 nm; and (d) Cy-5 12 (2.0 μM) for 2
min, λex = 633 nm. The cell nucleus staining (c and d) by Hoechst
33258 (λex = 405 nm) was used as control. Scale bar in all images = 20
μm.
C
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further conversion to amides or nitrogen heterocycles; see: (a) Zhang,
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- Asian J. 2014, 9, 2458.
Technological University, for the assistance in setting up the
cell-imaging facility.
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D
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