Conversion of azides into diazo compounds in water

Article abstract of DOI:10.1021/ja407822b

Diazo compounds are in widespread use in synthetic organic chemistry but have untapped potential in chemical biology. We report on the design and optimization of a phosphinoester that mediates the efficient conversion of azides into diazo compounds in phosphate buffer at neutral pH and room temperature. High yields are maintained in the presence of common nucleophilic or electrophilic functional groups, and reaction progress can be monitored by colorimetry. As azido groups are easy to install and maintain in biopolymers or their ligands, this new mode of azide reactivity could have substantial utility in chemical biology.

Full text of DOI:10.1021/ja407822b

Communication
pubs.acs.org/JACS
Conversion of Azides into Diazo Compounds in Water
†
,†,∥
Ho-Hsuan Chou and Ronald T. Raines*
†
∥
Department of Chemistry and Department of Biochemistry, University of WisconsinMadison, Madison, Wisconsin 53706,
United States
*
S Supporting Information
ditions and that tolerates the sensitive functionalities in
biological molecules could have high utility.
ABSTRACT: Diazo compounds are in widespread use in
synthetic organic chemistry but have untapped potential in
chemical biology. We report on the design and
optimization of a phosphinoester that mediates the
efficient conversion of azides into diazo compounds in
phosphate buffer at neutral pH and room temperature.
High yields are maintained in the presence of common
nucleophilic or electrophilic functional groups, and
reaction progress can be monitored by colorimetry. As
azido groups are easy to install and maintain in
biopolymers or their ligands, this new mode of azide
reactivity could have substantial utility in chemical biology.
8f,10
According to our putative mechanism (Scheme 1),
azide
deimidogenation begins with nucleophilic attack of the
Scheme 1
iazo compounds are among the most versatile inter-
D
mediates in organic synthesis. Because of their inherent
dipolar nature, diazo compounds can readily participate in 1,3-
dipolar cycloaddition reactions with a wide range of
1
dipolarophiles. Moreover, C-protonation gives rise to2
diazonium ions, which are highly reactive alkylating agents.
Thermal or photochemical generation of carbenes, along with
transition metal-mediated carbenoid formation, facilitates
addition to double bonds and insertion into C−H, O−H,
3
and N−H bonds. This broad reactivity makes diazo
compounds attractive for applications in chemical biology,
4
,5
having special promise in the labeling of proteins and as
tunable reactants in 1,3-dipolar cycloaddition reactions with
phosphine on the azide to form a phosphazide. Then, the
reaction takes one of two possible pathways, depending on the
ability of the pendant acyl group to trap the phosphazide
1
,6
cycloalkynes.
Although the first diazo compound was synthesized in the
9th century, there are still few methods for their construction:
intermediate prior to N (g) extrusion. We hypothesized that
7
2
1
this ability correlates with the pK of the conjugate acid of the
a
diazo transfer to an activated C−H acceptor, diazotization of an
amine, decomposition or oxidation of a hydrazone, rearrange-
ment of an N-acyl-N-nitrosoamine, and fragmentation of a
leaving group, and that more reactive acyl groups would favor
diazo compound formation and less reactive groups would
favor amides.
To test our hypothesis, we surveyed the reactivity of a wide
variety of potential phosphinoesters with α-azido-N-benzylace-
tamide. Reaction mixtures had equimolar reactants, which were
incubated at room temperature for 4 h, quenched with
5
c,7,8
triazene.
The harsh conditions required to access diazo
compounds can provoke undesirable reactivity with common
functional groups, and the need for organic solvents hampers
the generation of diazo compounds in many molecules of
biological interest.
saturated aqueous NaHCO , and then stirred for another 8 h.
3
Recently, we reported that an azide can be converted into the
We found that the diazo compound/amide product ratio does
corresponding diazo compound by the generation and
indeed correlate with the pK of the conjugate acid of the
a
8
f
subsequent decomposition of an acyl triazene. The
phosphinoester reagent that mediates this deimidogenation is,
however, nearly insoluble in water and unstable to hydrolysis,
leaving group (Table 1). When the pK
intermediate phosphazide underwent rapid N
rather than acyl transfer, yielding exclusively the Staudinger
a
was ≥9.2, the
2
(g) extrusion
ligation product. As the pK was lowered from 9.0 to 7.6, the
and the preferred solvent is THF/H O (20:3). As azido groups
a
2
9
have become a privileged functionality in chemical biology, a
stable, water-soluble reagent that converts azides into their
corresponding diazo compounds under physiological con-
Received: July 29, 2013
Published: September 23, 2013
©
2013 American Chemical Society
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Journal of the American Chemical Society
Communication
Table 1. Phosphinoester Reactivity and Stability
a
product ratio
diazo compound
% decomposition at pH
XH
pKa
amide
4.0
7.0
9.0
12.0
methanol
15.5
10.6
10.0
9.4
9.2
9.0
8.7
8.5
7.6
7.1
6.9
6.0
5.5
0
0
100
100
100
100
100
67
27
33
17
3
ethanethiol
4-fluorophenol
0
benzylmercaptan
0
3
3
3
3
2
4
2
-(dimethylamino)phenol
-chlorophenol
0
33
63
67
83
97
97
98
97
,5-difluorophenol
-hydroxypyridine
,2,2-trifluoroethanethiol
-nitrophenol
<5
<5
<5
<5
<5
12
<5
<5
<5
<5
<5
<5
40
<5
5
12
13
10
<5
54
<5
21
62
35
46
13
98
16
,4,6-trifluorophenol
3
N-hydroxysuccinimide
2
pentafluorophenol
3
a
1
Determined by H NMR spectroscopy: δ 4.73 (1H, diazo compound) and 3.95 (2H, amide).
a
product ratio began to increase. When the pK was ≤7.1, the
Scheme 2. Synthetic Route to Phosphinoester 1
a
reaction produced the diazo compound, almost exclusively.
In addition to promoting diazo-compound formation, a
useful reagent must exhibit high chemical stability. We were
concerned about this attribute because esters formed with
alcohols having pK ≤7.1 can be unstable to hydrolysis.
a
Accordingly, we assessed stability by stirring a reagent in
phosphate buffer containing 40% (v/v) THF (to enhance
1
solubility) for 19 h and evaluating decomposition by H NMR
spectroscopy. Hydrolysis was not observed at pH 7.0 (Table 1),
except for the N-hydroxysuccinimide (NHS) ester, which was
8f
the object of our previous study. Most of the phosphinoesters
were likewise stable at pH 9.0. On the basis of these reactivity/
stability screens, ease-of-synthesis, and chromogenicity, the 4-
nitrophenyl ester was chosen as ideal.
a
Conditions: (a) Mg, aryl bromide, THF, 95%; (b) BH
8%; (c) NaOMe, methy acrylate, MeOH, 71%; (d) 7 N KOH,
−SMe , THF,
2
3
9
MeOH, 98%; (e) MeOH/toulene reflux, 97%; (f) 4-nitrophenol,
EDC, CH Cl , 72%.
2
2
Next, we sought a reagent that was soluble in water. We
reported previously that N,N-dimethylamino groups imparted
water solubility to phosphinothioesters and enabled the
The ability of phosphinoester 1 to convert azides into their
corresponding diazo compounds was assessed first in
phosphate buffer (Table 2). α-Azido-N-benzylacetamide was
treated with phosphinoester 1 in buffers of various pH for 24 h,
11
traceless Staudinger ligation in water. N,N-Dimethylamino
groups, however, led to rapid decomposition of the
phosphinoesters used herein. After screening other functional
11
groups that confer water solubility on similar phosphines, we
settled on methoxyethoxymethyl (MEM) as a preferred
Table 2. Effect of pH and Organic Cosolvent on Diazo
Compound Formation by Phosphinoester 1
1
2
group.
The synthesis of an optimized reagent for the conversion of
azides into diazo compounds in water began from 3-
bromobenzyl alcohol that was MEM-protected, converted
into the corresponding Grignard compound, and added to
diethyl phosphite to give the bis-aryl phosphine oxide (Scheme
2
). Reduction of the phosphine oxide and subsequent
a
b
c
b
entry solvent
yield (%)
entry
solvent
yield (%)
protection with borane dimethylsulfide gave the borane-
13
1
2
3
4
pH 5.0
69
88
91
71
5
6
7
8
9
50% v/v MeOH
80
74
75
74
78
protected phosphine. Direct deprotonation of the protected
phosphine and subsequent conjugate addition with methyl
acrylate followed by hydrolysis afforded the phosphine-
pH 6.0
pH 7.0
pH 8.0
50% v/v CH CN
3
50% v/v DMF
1% v/v DMSO
20% v/v E.G.
8
f,14
carboxylic acid.
Removing the borane protecting group
15
with a mild methanol reflux, and installing the 4-nitro-
phenylester using standard coupling conditions provided
phosphinoester 1 in 6 steps and 45% overall yield.
a
b
c
10 mM sodium phosphate buffer. Isolated yield. In 10 mM sodium
phosphate buffer, pH 7.0. E.G. = ethylene glycol.
1
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Journal of the American Chemical Society
Communication
and the ensuing diazo compound was isolated by chromatog-
raphy on silica gel. The reaction was found to be efficient under
the conditions screened, with the optimal yield being achieved
at neutral pH without the addition of an organic cosolvent
In conclusion, we have developed a phosphinoester that
mediates the efficient conversion of azides into diazo
compounds in phosphate buffer at neutral pH. This conversion
is tolerant to the functional groups relevant to chemical biology.
No other method for generating diazo compounds has these
(
entry 3). This result bodes well for biological applications, as
the optimal conversion was attained near physiological
conditions. Under those conditions, reaction progress can be
attributes. Azido groups, which can be introduced with a simple
9
S 2 reaction, have found widespread use in chemical biology.
N
16
monitored by quantifying the yellow 4-nitrophenolate anion.
Accordingly, a reagent capable of converting azides into their
smaller and even more versatile diazo congeners in water could
have substantial utility.
Then, we probed functional group compatibility using
derivatives of azido-glycine. The initial test was to probe the
tolerance of azide deimidogenation by phosphinoester 1 to
strong nucleophiles, such as alcohols, amines, and thiols, all of
which are capable of undergoing acyl transfer reactions with 4-
ASSOCIATED CONTENT
Supporting Information
■
*
S
17
nitrophenyl esters. We found that acyl transfer was not
Experimental procedures and spectral data for novel com-
pounds. This material is available free of charge via the Internet
at http://pubs.acs.org.
competitive with deimidogenation in phosphate buffer at pH
7.0 (Table 3).
Table 3. Chemoselectivity of Diazo-Compound Formation
in Water by Phosphinoester 1
AUTHOR INFORMATION
Corresponding Author
rtraines@wisc.edu
■
a
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We are grateful to Dr. N. A. McGrath, Dr. E. Myers, and C. H.
Eller for contributive discussions and assistance. This work was
supported by grant R01 GM044783 (NIH). This study made
use of the National Magnetic Resonance Facility at Madison,
which is supported by grants P41 RR002301 and P41
GM066326 (NIH).
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