Efficient synthetic approach to fluorescent oxazole-4-carboxylate derivatives

Article abstract of DOI:10.1080/00397911.2012.706350

In our attempt to synthesize a halogenated analog of green fluorescent protein (GFP) chromophore, we discovered a simple and efficient synthetic strategy to the derivatives of oxazole-4-carboxylic acid substituted at positions 2 and 5. The method allows f

Full text of DOI:10.1080/00397911.2012.706350

This article was downloaded by: [Fordham University]
On: 17 February 2013, At: 12:00
Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK
Synthetic Communications: An International Journal
for Rapid Communication of Synthetic Organic
Chemistry
Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/lsyc20
An Efficient Synthetic Approach to Fluorescent
Oxazole-4-carboxylate Derivatives
Mikhail S. Baranov , Konstantin A. Lukyanov , Pavel E. Ivashkin & Ilia V. Yampolsky
Accepted author version posted online: 14 Feb 2013.
To cite this article: Mikhail S. Baranov , Konstantin A. Lukyanov , Pavel E. Ivashkin & Ilia V. Yampolsky (2013): An Efficient
Synthetic Approach to Fluorescent Oxazole-4-carboxylate Derivatives, Synthetic Communications: An International Journal for
Rapid Communication of Synthetic Organic Chemistry, DOI:10.1080/00397911.2012.706350
To link to this article: http://dx.doi.org/10.1080/00397911.2012.706350
Disclaimer: This is a version of an unedited manuscript that has been accepted for publication. As a service
to authors and researchers we are providing this version of the accepted manuscript (AM). Copyediting,
typesetting, and review of the resulting proof will be undertaken on this manuscript before final publication of
the Version of Record (VoR). During production and pre-press, errors may be discovered which could affect the
content, and all legal disclaimers that apply to the journal relate to this version also.
PLEASE SCROLL DOWN FOR ARTICLE
Full terms and conditions of use: http://www.tandfonline.com/page/terms-and-conditions
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to
anyone is expressly forbidden.
The publisher does not give any warranty express or implied or make any representation that the contents
will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should
be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims,
proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in
connection with or arising out of the use of this material.

ACCEPTED MANUSCRIPT
An efficient synthetic approach to fluorescent oxazole-4-carboxylate derivatives
Mikhail S. Baranov, Konstantin A. Lukyanov, Pavel E. Ivashkin, Ilia V. Yampolsky
Email: ivyamp@gmail.com
Abstract
Supplemental materials are available for this article. Go to the publisher’s online edition
of Synthetic Communications® to view the free supplemental file.
INTRODUCTION
Oxazoles represent an important class of heterocyclic compounds. [1,2] Oxazole moiety is
found among biologically active compounds. [3–12] Oxazole derivatives are used as
luminophores for scintillation [13] and as fluorescent labels. [14–17]
Known methods for synthesis of oxazole-4-carboxylates include metalation followed by
the action of electrophile, [18] palladium-catalysed coupling reactions [19] and cyclization
of brominated derivatives of dehydropeptides. [17, 20, 21]
RESULTS AND DISCUSSION
In the course of our systematic studies in the chemistry of arylideneimidazolones –
chromophores of GFP-like fluorescent proteins [22, 23] we attempted to introduce halogen
ACCEPTED MANUSCRIPT
1

ACCEPTED MANUSCRIPT
substituent at vinylic position of the GFP chromophore (A, Scheme 1). This halogenated
chromophore (A) would serve as a useful synthetic intermediate in the chemistry of
chromophores of fluorescent proteins.
Among several synthetic approaches to A we examined the one outlined in Scheme 1. It
is based on Erlenmeyer azlactone (1) formation from aldehyde and acylglycine with
subsequent opening of azlactone cycle with nucleophile, bromination of the
acylaminocinnamic acid 2 and finally, cyclization of vinylic bromides 3 upon the action
of base. However, instead of the desired brominated imidazolinone A the final cyclization
step produced only oxazoles 4. Due to little information value of NMR spectra in case of
highly substituted heterocycles we used X-ray analysis to confirm the structure of final
products 4 (Fig. 1).
This undesired oxazole formation can be viewed as a useful synthetic method for
preparation of a wide range of derivatives of 5-aryloxazole-4-carboxylic acids. The
method makes it possible to introduce different aryl substituents at the position 5, aryl or
alkyl substituents at position 2 of oxazole, and gives access not only to free carboxyl at
position 4, but also to a range of its amide derivatives. The advantages of the synthetic
strategy presented are: synthetic availability of a wide range of oxazolones 1, [23] much
better yields compared to an early report [21] and also avoiding palladium coupling and
metalation procedures.
ACCEPTED MANUSCRIPT
2

ACCEPTED MANUSCRIPT
We characterized optical spectra of the chromophores obtained (Fig. 2 and Table 1). As
expected, all chromophores possessed absorption maxima in UV region. At the same
time, their emission peaks were in violet-blue region of the visible spectrum due to a
large Stockes shift. The brightest fluorescence (quantum yield 0.82) was observed for 4b.
Chromophore 4a with an ionizable hydroxyl group showed expected red shift of
absorption and emission spectra upon deprotonation in basic conditions. Other
chromophores (including 4d possessing a carboxylic group) did not change absorption
maxima in the pH range 2-12. Compared to 4b, 4a had considerably lower quantum yield
(0.25), which further drastically decreased to 0.01 in the anionic state. Introduction of
phenyl substituent in 4c (compared to methyl in 4b) resulted in a significantly red-shifted
absorption, but practically did not change emission maximum.
We believe that 5-aryl-4-carboxyoxazole is a promising core for creation of new
fluorescent dyes. This core can ensure high quantum yield of fluorescence and emission
in the visible spectrum in spite of low size of the molecule. As spectral properties of these
compounds are very sensitive to introduced substituents we expect that red shifted
fluorophores can be obtained with this core structure.
EXPERIMENTAL
NMR spectra were recorded on Bruker Avance III 700 spectrometer. UV-vis spectra were
recorded with a Varian Cary 100 spectrophotometer. Fluorescence excitation and
emission spectra were recorded with a Varian Cary Eclipse fluorescence
spectrophotometer.
ACCEPTED MANUSCRIPT
3

ACCEPTED MANUSCRIPT
Oxazolones 1a-c were prepared by literature procedures. [23]
GENERAL PROCEDURE FOR PREPARATION OF ACYLAMINOCINNAMIC
ACIDS 2
2a: To a suspension of 1a (2.45 g, 10.0 mmol) in dry acetonitrile (50 mL)
cyclohexylamine (1.49 g, 15.0 mmol) was added. The mixture was stirred for 30 minutes,
while white precipitate was formed. Then the reaction mixture was cooled to 0oC and
filtered, precipitate was washed by small amount of acetonitrile and diethyl ether. Yield
2.65 g (77%). Mp 224-227oC
1H NMR (DMSO-d6) δ 9.33 (br. s, 1H), 7.69 (d, 1H, J=7.67Hz), 7.56 (d, 2H, J=8.11Hz),
7.14 (d, 2H, J=8.11Hz), 6.90 (s, 1H), 3.61 (m, 1H), 2.27 (s, 3H), 1.98 (s, 3H), 1.72-1.68
(m, 4H), 1.59 (m, 1H), 1.27 (m, 4H), 1.12 (m, 1H).
13C NMR (DMSO-d6) δ 20.76 (CH3), 22.79 (CH3), 24.78 (2xCH2), 25.20 (CH2), 32.18
(2xCH2), 48.19 (CH), 121.76 (2xCH), 125.55 (CH), 130.26 (2xCH), 130.60, 131.94,
150.09, 164.08, 168.96, 169.13.
HRMS (ESI) calcd for C19H24N2O4 344.1736, found 344.1742.
GENERAL PROCEDURE FOR BROMINATED ACYLAMINOCINNAMIC
ACIDS 3
ACCEPTED MANUSCRIPT
4

ACCEPTED MANUSCRIPT
3a: To a solution of 2a (1.58 g, 5.0 mmol) in dry dichloromethane (50 mL) NaHCO3
(1.68 g, 20.0 mmol) was added. Then a solution of bromine (1.0g, 6.3 mmol) in
dichloromethane (10 mL) was added with stirring. After the addition was complete the
mixture was stirred for additional 30 minutes, filtered, diluted with 100 mL of
dichloromethane, washed with 5% Na2SO3 (50 mL), water (2x50mL), brine (50 mL),
dried over Na2SO4 and concentrated in vacuo. The resulting white solid was washed with
small amount of diethyl ether giving 1.85 g (88%) of 3a containing 4a as a trace impurity.
Mp 221-224oC.
H NMR (DMSO-d6) δ 9.38 (br. s, 1H), 7.66 (d, 1H, J=7.89Hz), 7.33 (d, 2H, J=8.55Hz),
7.08 (d, 2H, J=8.33Hz), 3.26 (m, 1H), 2.26 (s, 3H), 1.99 (s, 3H), 1.49 (m, 2H), 1.42 (m,
1H), 1.30 (m, 2H), 1.06 (m, 2H), 0.94 (m, 1H), 0.77 (m, 2H).
13C NMR (DMSO-d6) δ 20.72 (CH3), 22.39 (CH3), 24.27 (2xCH2), 25.03 (CH2), 32.18
(2xCH2), 47.50 (CH), 113.05, 121.34 (2xCH), 130.37 (2xCH), 134.66, 135.59, 150.47,
161.32, 167.91, 168.86.
HRMS (ESI) calcd for C19H23BrN2O4 422.0841, found 422.0848.
GENERAL PROCEDURE FOR PREPARATION OF OXAZOLES 4
4a: A solution of 3a (0.847 g, 2.0mmol) in wet DMF (30 mL DMF and 5 mL water) was
refluxed with K2CO3 (0.69 g, 5.0 mmol) for 1-2 minutes. The solvent was removed in
vacuo, the mixture was dissolved in 100 mL DCM, acidified with 5% HCl, washed with
ACCEPTED MANUSCRIPT
5

ACCEPTED MANUSCRIPT
water (2x50 mL), brine and dried over Na2SO4. The solution was concentrated in vacuo
to give a white solid, which was purified by column chromatography:
(CHCl3:EtOH=20:1). Yield 440 mg, (73%). Mp 202-204oC.
1H NMR (DMSO-d6) δ 9.90 (s, 1H), 8.06 (d, 2H, J=8.33Hz), 7.75 (d, 1H, J=7.89Hz),
6.85 (d, 2H, J=8.55Hz), 3.74 (m, 1H), 2.47 (s, 3H), 1.78 (m, 2H), 1.71 (m, 2H), 1.58 (m,
1H), 1.38-1.27 (m, 4H), 1.13 (m, 1H).
13C NMR (DMSO-d6) δ 13.19 (CH3), 24.76 (2xCH2), 25.05 (CH2), 32.15 (2x CH2), 47.56
(CH), 115.06 (2xCH), 118.10, 127.43, 129.27 (2xCH), 151.25, 157.84, 158.61, 160.01.
HRMS (ESI) calcd for C17H20N2O3 300.1474, found 300.1468.
Please see the Supplementary Information, available online, for complete experimental
and spectral details.
ACKNOWLEDGMENT
We gratefully acknowledge grants by Molecular and Cell Biology program of the
Russian Academy of Sciences, by Ministry of Education and Science of the Russian
Federation (16.740.11.0367 and 16.512.11.2139) and by Russian Foundation for Basic
Research (11-04-01293-а).
REFERENCES
ACCEPTED MANUSCRIPT
6

ACCEPTED MANUSCRIPT
[1]
Taylor, E. C.; Wipf. P. Oxazoles: Synthesis, Reactions, and Spectroscopy; John
Wiley & Sons: Hoboken, 2003.
[2]
Turchi, I. J. In Oxazoles; John Wiley & Sons: New York, 1986; and references
therein.
[3]
Searle, P. A.; Molinski, F. F. J. Am. Chem. Soc. 1995, 117, 8126.
Roesener, J. A.; Scheuer, P. J. J. Am. Chem. Soc. 1986, 108, 846.
Paris, J. M.; Barriere, J. C.; Smith, C.; Bost, P. E. Recent Progress in the Chemical
[4]
[5]
Synthesis of Antibiotics; Lukacs, G., Ohno, M., Eds.; Springer-Verlag: Berlin, 1990, 183.
[6]
[7]
Matsumaga, S.; Fusetani, N. Tetrahedron Lett. 1991, 32, 5605.
Ichiba, T.; Yoshida, W. Y.; Scheuer, P. J.; Higa, T.; Gravalos, D. G. J. Am. Chem.
Soc. 1991, 113, 3173.
[8]
Shin-ya, K.; Wierzba, K.; Matsuo, K.; Ohtani, T.; Yamada, Y.; Furihata, K.;
Hayakawa, Y.; Seto, H. J. Am. Chem. Soc. 2001, 123, 1262.
[9]
Searle, P. A.; Molinski, T. F. J. Am. Chem. Soc. 1995, 117, 8126.
[10] Rosener, J. A.; Scheuer, P. J. J. Am. Chem. Soc. 1986, 108, 846.
[11] Dalisay, D. S.; Rogers, E. W.; Edison, A. S.; Molinski, T. F. J.Nat. Prod. 2009, 72,
732.
[12] Hagelueken, G.; Albrecht, S. C.; Steinmetz, H.; Jansen, R.; Heinz,D. W.; Kalesse,
M.; Schubert, W.-D. Angew. Chem., Int. Ed. 2008, 48, 595.
[13] Dyer, A. In An Introduction to Liquid Scintillation Counting; Heyden: London,
UK, 1974, 12.
[14] Pavlopoulos, T. G.; Hammond, P. R. J. Am. Chem. Soc. 1974, 96, 6568.
ACCEPTED MANUSCRIPT
7

ACCEPTED MANUSCRIPT
[15] Ionescu, S.; Popovici, D.; Balaban, A. T.; Hillebrand, M. Spectrochim. Acta, Part
A 2005, 62, 252.
[16] Lukavenko, O. N.; Eltsov, S. V.; Grigorovich, A. V.; Mchedlov-Petrossyan, N. O.
J. Mol. Liq. 2009, 145, 167.
[17] Ferreira, P. M. T.; Castanheira, E.M. S.; Monteiro, L. S.; Pereira, G.; Vilaça, H.
Tetrahedron 2010, 66, 8672.
[18] Williams, D. R.; Fu, L. Org. Lett. 2010, 12, 808.
[19] Hodgetts, K. J.; Kershaw, M. T. Org. Lett. 2002, 4, 2905.
[20] Shin, C.; Abe, C.; Yonezawa, Y. Chem. Lett. 2004, 33, 664.
[21] Pfleger, R.; Strandtmann, M. Chem. Ber. 1957, 1455.
[22] Ivashkin, P. E.; Lukyanov, K.A.; Lukyanov, S.; Yampolsky, I. V. J.Org. Chem.
2011, 76, 2782.
[23] Ivashkin, P. E.; Yampolsky, I.V.; Lukyanov, K. A. Russ. J. Bioorg. Chem. 2009,
35, 652.
ACCEPTED MANUSCRIPT
8

ACCEPTED MANUSCRIPT
Table 1. Spectral properties of 4a-d.
Chromophor Water
EtOH
e
Absorbanc Extinction Emissio Fluorescenc Absorbanc Extinction
e max, nm coefficien n max,
t, M-1cm-1 nm
e quantum
yield
0.25
e max, nm coefficien
t, M-1cm-1
4a
290
322
291
333
309
19000
24000
18000
12000
24000
400
460
395
393
411
300
340
298
317
315
26000
31000
27000
25000
17000
4a anion a
0.01
4b
4c
4d
0.82
0.15
0.65
a In the presence of 10 mM NaOH.
ACCEPTED MANUSCRIPT
9

ACCEPTED MANUSCRIPT
Scheme 1. Synthesis of oxazoles (4) via Erlenmeyer azlactones. Attempt to synthesize
halogenated GFP chromophore derivative (A)
ACCEPTED MANUSCRIPT
10

ACCEPTED MANUSCRIPT
Figure 1. Structure of 4b by X-ray crystallography.
ACCEPTED MANUSCRIPT
11

ACCEPTED MANUSCRIPT
Figure 2. Absorbance and fluorescence spectra of oxazoles 4a-d in water.
ACCEPTED MANUSCRIPT
12