Extension of a cascade reaction: Microwave-assisted synthesis of the GFP chromophore derivatives

Article abstract of DOI:10.1002/jhet.315

(Chemical Equation Presented) A facile microwave-assisted synthesis of imidazol-5(4H)-one derivatives is accomplished via reactions of 4-arylmethylene-2-phenyloxazol-5(4H)-ones with urea (or ammonium acetate) in ethylene glycol. The cascade reaction is simple to perform and occurs under mild conditions with broad scope of applicability.

Full text of DOI:10.1002/jhet.315

354
Extension of a Cascade Reaction: Microwave-Assisted Synthesis
of the GFP Chromophore Derivatives
Vol 47
Runhong Jia,a Shu Yan,b Bo Jiang,b,c Feng Shi,b and Shu-Jiang Tub*
aLianyungang Teacher’s College, Lianyungang, Jiangsu 222006, People’s Republic of China
bCollege of Chemistry and Chemical Engineering, Xuzhou Normal University, Xuzhou,
Jiangsu 221116, People’s Republic of China
cCollege of Chemistry, Chemical Engineering and Materials Science, Suzhou University, Suzhou,
People’s Republic of China
*E-mail: jiarunhong@126.com
Received July 19, 2009
DOI 10.1002/jhet.315
Published online 2 March 2010 in Wiley InterScience (www.interscience.wiley.com).
A facile microwave-assisted synthesis of imidazol-5(4H)-one derivatives is accomplished via reac-
tions of 4-arylmethylene-2-phenyloxazol-5(4H)-ones with urea (or ammonium acetate) in ethylene gly-
col. The cascade reaction is simple to perform and occurs under mild conditions with broad scope of
applicability.
J. Heterocyclic Chem., 47, 354 (2010).
INTRODUCTION
Green fluorescent protein (GFP) including imidazoli-
dinone skeleton (Fig. 1) [1], is an autofluorescent pro-
tein of 238 amino acid residues that is derived from the
Pacific Northwest jellyfish, Aequorea victoria [2]. The
GFP chromophore exhibits promising applications in
molecular and cell biology due to its intrinsic visible
fluorescence, which is easily detectable by fluorescence
spectroscopy [3–5]. Therefore, much interest has been
geared toward the engineering of novel color variants
[6,7] of the GFP in light of its wide applicability in the
life sciences.
Figure 2. Compounds 3.
tone [12–14]. All these methods [12–17] have one or
more limitations such as inaccessibility of precursors,
narrow substrate scope, and operational complexity.
Recently, a microwave-improved synthesis of imidazo-
lones using graphite as support has been reported [15].
In the light of current studies, the development of a
practically simple, economical, and high-yielding route
to a wide variety of imidazol-5(4H)-one derivative (Fig.
2) is strongly desired. Herein, we like to report a cas-
cade reaction of 4-arylmethylene-2-phenyloxazol-5(4H)-
ones 1 (Compounds 1 were conveniently prepared fol-
lowing a literature procedure: [18]) with urea 2 (or am-
monium acetate) for synthesis of imidazol-5(4H)-ones
under microwave irradiation (MWI) at 150ꢀC in ethyl-
ene glycol (Scheme 1).
In addition, the imidazolone core represents an attrac-
tive pharmacophore that displays extensively pharmaco-
logical and medicinal activities [8,9]. In particular, imi-
dazol-5(4H)-one and its derivatives have possessed an
unique role in drug discovery and crop protection
[10,11], serving as combinatorial chemistry groups. In
general routes, the imidazolone ring is formed by con-
densing glycine ester of acetimidic or phenylacetimidic
acid in the solvents, such as benzene, dioxane, and ace-
RESULTS AND DISCUSSION
To choose the most appropriate solvents, the MW-
assisted reaction (Scheme 2) of 4-(4-chlorobenzylidene)-
2-phenyloxazol-5(4H)-one (1c, 1 mmol) and urea (2,
1.5 mmol) was examined using glacial acetic acid
(HOAc), ethylene glycol, ethanol (EtOH), N,N-dimethyl
Figure 1. GFP chromophore.
C
V
2010 HeteroCorporation

March 2010
Extension of a Cascade Reaction: Microwave-Assisted Synthesis of the
GFP Chromophore Derivatives
355
Scheme 1
Table 1
Solvent optimization for the synthesis of 3c under MWI.
Entry
Solvent
Temp (ꢀC)
Time (min)
Yield (%)
1
2
3
4
5
HOAc
Glycol
EtOH
DMF
150
150
150
150
150
5
5
5
5
5
trace
93
63
79
formamide (DMF), water as the solvent (2.0 mL) at
150ꢀC, respectively. All the reactions were carried out
at the maximum power of 300 W. As shown in Table 1,
we could see the reaction in glycol gave the best results
(Table 1, entry 2).
Water
32
In a further study, aromatic amine 4 was employed
instead of urea 2 in this case. The reactions proceeded
smoothly too. However, the desired products 3 were not
detected. Instead, a series of open-chain products 5 were
obtained in high yields (Scheme 4). The reason may be
attributed to the nucleophilicity of amine. When R is a
phenyl group, the resulting N-phenylformamide deriva-
tive decreases the nucleophilicity of amine, which is dif-
ficult to form closed-ring products 3. The synthesis of
these compounds has reported in our previous study
(Compounds 5 were conveniently prepared following a
literature procedure: [19]).
To further optimize reaction conditions, the same
reaction was performed in ethylene glycol and 300 W at
the temperatures ranging from 110 to 180ꢀC in incre-
ments of 10ꢀC each time. The yield of product 3c was
improved from 23 to 93%, when the temperature was
raised from 110 to 150ꢀC (Table 2, entries 1–5). How-
ever, no significant increase in the yield of product 3c
was observed as the reaction temperature was raised
from 150 to 180ꢀC (Table 2, entries 5–8). Therefore,
150ꢀC phenyloxazol-5(4H)-ones were performed, which
leads to the corresponding 4-arylmethylene-2-phenyl-
1H-imidazol-5(4H)-ones with good yields.
Under the optimal conditions [glycol, 150ꢀC, 300 W
(maximum power)], reactions of different 4-arylmethy-
lene-2-phenyloxazol-5(4H)-ones were performed, which
leads to the corresponding 4-arylmethylene-2-phenyl-
1H-imidazol-5(4H)-ones with good yields. The elec-
tronic effect of the aryl group in 4-arylmethylene-2-phe-
nyloxazol-5(4H)-ones was investigated. As shown in
Table 3, both electron-withdrawing (such as nitro) and
electron-donating (such as alkoxy) groups readily pro-
vided compounds 3 in good yields. Moreover, the heter-
ocyclic phenyloxazol-5(4H)-ones such as 2-phenyl-4-
((thiophen-2-yl)methylene) oxazol-5(4H)-one (Table 3,
entry 9) still displayed a high reactivity under this stand-
ard condition.
In summary, we demonstrated a simple method, using
readily available starting materials and simple experi-
mental procedures, for the efficient synthesis of imida-
zol-5(4H)-one derivative and related compounds. Partic-
ularly, valuable features of this cascade reaction
included operational simplicity, high yields, increased
safety for small-scale high-speed synthesis, and broader
substrate scope.
EXPERIMENTAL
All reactions were performed in a monomodal EmrysTM
Creator from Personal Chemistry, Uppsala, Sweden. Melting
points were determined in open capillaries and are uncorrected.
IR spectra were recorded on a FTIR-tensor 27 spectrometer in
KBr. 1H NMR spectra were measured on a DPX 400 MHz
spectrometer using TMS as an internal standard and DMSO-d6
as solvent. Elemental analysis was determined by using a Per-
kin-Elmer 240c elemental analysis instrument.
We envisaged that ammonia, obtained by urea (or
ammonium acetate), attacks on carbonyl group in 4-aryl-
methylene-2-phenyloxazol-5(4H)-ones 1 as a nucleo-
phile. Bond formation should lead directly to intermedi-
ate 7 via ring opening, which then poised to attack adja-
cent reactive carbonyl group. Finally, an intramolecular
condensation occurred and 4-arylmethylene-2-phenyl-
1H-imidazol-5(4H)-ones 3 were obtained (Scheme 3).
Table 2
Temperature optimization for the synthesis of 3c under MWI.
Entry
Temp (ꢀC)
Time (min)
Yield (%)
1
2
3
4
5
6
7
8
110
120
130
140
150
160
170
180
5
5
5
5
5
5
5
5
23
58
75
82
93
71
58
41
Scheme 2
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

356
R. Jia, S. Yan, B. Jiang, F. Shi, and S.-J. Tu
Vol 47
Table 3
Physical and analytical data of compounds 3.
Entry
Compd.
Ar
Time (min)
Yield (%)
Mp (ꢀC)
1
2
3
4
5
6
7
8
9
3a
3b
3c
3d
3e
3f
3g
3h
3i
C6H5
4-FC6H4
4-ClC6H4
2,4-Cl2C6H3
3,4-Cl2C6H3
4-CH3C6H4
2-CH3OC6H4
4-CH3OC6H4
Thien-2-yl
5 (3) [15]
5
87 (85) [15]
84
>300 (272–273) [15]
286–287
5 (5) [15]
6 (4) [15]
6
93 (91) [15]
83 (97) [15]
85
>300 (289–290) [15]
268–269 (273–274)[15]
277–278
5 (4) [15]
6
95 (70) [15]
82
278–279 (288–289) [15]
278–279 (254–255) [15]
>300 (289–290) [15]
>300 (291–292) [15]
5 (3) [15]
5 (2) [15]
87 (84) [15]
86 (80) [15]
1H NMR (400 MHz, DMSO) (d, ppm): 12.14 (s, 1H, NH),
8.41 (t, J ¼ 8.0 Hz, 2H, ArH), 8.18 (d, J ¼ 8.0 Hz, 2H, ArH),
7.68–7.59 (m, 3H, ArH), 7.35 (t, J ¼ 8.8 Hz, 2H, ArH), 7.06
(s, 1H, CH).
Anal calcd. For C16H11FN2O, C, 72.17; H, 4.16; N, 10.52;
found C, 72.24; H, 4.14; N, 10.59%.
Sample experimental
4-Arylmethylene-2-phenyl-1H-imidazol-5(4H)-one (3). In a
10 mL EmrysTM reaction vial, 4-arylmethylene-2-phenyloxa-
zol-5(4H)-ones (1 mmol) with urea (1.5 mmol), ammonium ac-
etate (1.5 mmol), in ethylene glycol (2.0 mL) were mixed and
then capped. The mixture was irradiated by microwave at 300
W and 150ꢀC for a given time. The automatic mode stirring
helped the mixing and uniform heating of the reactants. Upon
completion, monitored by TLC, the reaction mixture was
cooled to room temperature and then poured into cold water,
filtered to give the crude products, which were further purified
by recrystallization from 95% EtOH. The reaction time and
the yields are listed in Table 3. The analytical data of new
products are as following:
4-(4-Chlorobenzylidene)-2-phenyl-1H-imidazol-5(4H)-one
(3c). IR (KBr): 3153, 3124, 3066, 2987, 1703, 1641, 1541,
1456, 1261, 1180, 1092, 923, 788, 691 cmꢁ1
.
1H NMR (400 MHz, DMSO) (d, ppm): 12.18 (s, 1H, NH),
8.36 (d, J ¼ 8.4 Hz, 2H, ArH), 8.19 (d, J ¼ 8.4 Hz, 2H,
ArH), 7.67–7.56 (m, 5H, ArH), 7.05 (s, 1H, CH).
Anal calcd. For C16H11ClN2O, C, 67.97; H, 3.92; N, 9.91;
found C, 67.88; H, 3.89; N, 9.99%.
4-(2,4-Dichlorobenzylidene)-2-phenyl-1H-imidazol-5(4H)-
one (3d). IR (KBr): 3159, 3129, 3062, 2986, 1708, 1638,
4-Benzylidene-2-phenyl-1H-imidazol-5(4H)-one (3a). IR (KBr):
3124, 3067, 2989, 1698, 1639, 1539, 1496, 1452, 1419, 1322,
1536, 1455, 1413, 1361, 1249, 1181, 1098, 915, 694 cmꢁ1
.
1266, 1187, 1031, 922, 775, 687 cmꢁ1
.
1H NMR (400 MHz, DMSO) (d, ppm): 12.36 (s, 1H, NH),
9.06 (s, 1H, ArH), 8.15 (d, J ¼ 7.2 Hz, 2H, ArH), 7.70–
7.61(m, 4H, ArH), 7.51 (dd, J1 ¼ 8.4 Hz, J2 ¼ 2.4 Hz, 1H,
ArH), 7.18 (s, 1H, CH).
1H NMR (400 MHz, DMSO) (d, ppm): 12.12 (s, 1H, NH),
8.32 (d, J ¼ 7.2 Hz, 2H, ArH), 8.18 (d, J ¼ 7.6 Hz, 2H,
ArH), 7.67–7.69 (m, 3H, ArH), 7.52–7.44 (m, 3H, ArH), 7.04
(s, 1H, CH).
HRMS (ESI): m/z [MþH]þ calcd for C16H12N2O: 249.1023;
Anal calcd. For C16H10Cl2N2O, C, 60.59; H, 3.18; N, 8.83;
found C, 60.65; H, 3.14; N, 8.89%.
found: 249.1023.
Anal calcd. For C16H12N2O, C, 77.40; H, 4.87; N, 11.28;
found C, 77.50; H, 4.79; N, 11.32%.
4-(3,4-Dichlorobenzylidene)-2-phenyl-1H-imidazol-5(4H)-
one (3e). IR (KBr): 3127, 3068, 2986, 1711, 1646, 1547,
1455, 1416, 1355, 1249, 1125, 908, 784, 685 cmꢁ1
.
4-(4-Fluorobenzylidene)-2-phenyl-1H-imidazol-5(4H)-one
(3b). IR (KBr): 3159, 3128, 3071, 2991, 1707, 1645, 1595,
1H NMR (400 MHz, DMSO) (d, ppm): 12.23 (s, 1H, NH),
8.60 (s, 1H, ArH), 8.33 (dd, J1 ¼ 8.4 Hz, J2 ¼ 1.2 Hz, 1H,
ArH), 8.18 (d, J ¼ 7.2 Hz, 2H, ArH), 7.76 (d, J ¼ 8.4 Hz,
1H, ArH), 7.68–7.61 (m, 3H, ArH), 7.04 (s, 1H, CH).
1500, 1457, 1235, 1157, 921, 842, 784, 691 cmꢁ1
.
Scheme 3
Scheme 4
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

March 2010
Extension of a Cascade Reaction: Microwave-Assisted Synthesis of the
GFP Chromophore Derivatives
357
Anal calcd. For C16H10Cl2N2O, C, 60.59; H, 3.18; N, 8.83;
found C, 60.51; H, 3.25; N, 8.78%.
Acknowledgments. The authors are grateful for financial sup-
port from the National Natural Science Foundation of China (No.
20672090), Natural Science Foundation of the Jiangsu Province
(No. BK2006033), and the Qing Lan Project of Jiangsu Province
(No. 08QLT001).
4-(4-Methylbenzylidene)-2-phenyl-1H-imidazol-5(4H)-one
(3f). IR (KBr): 3152, 3122, 3062, 2984, 1701, 1638, 1598,
1496, 1456, 1256, 1180, 1028, 922, 816, 789, 690 cmꢁ1
.
1H NMR (400 MHz, DMSO) (d, ppm): 12.09 (s, 1H, NH),
8.22 (d, J ¼ 7.8 Hz, 2H, ArH), 8.17 (d, J ¼ 8.0 Hz, 2H,
ArH), 7.67–7.59 (m, 3H, ArH), 7.32 (d, J ¼ 8.0 Hz, 2H,
ArH), 7.01 (s, 1H, CH), 2.38 (s, 3H, CH3).
Anal calcd. For C17H14N2O, C, 77.84; H, 5.38; N, 10.68;
found C, 77.89; H, 5.31; N, 10.79%.
REFERENCES AND NOTES
4-(2-Methoxybenzylidene)-2-phenyl-1H-imidazol-5(4H)-one
(3g). IR (KBr): 3147, 3071, 2985, 2839, 1691, 1633, 1455,
[1] Bourotte, M.; Schmitt, M.; Follenius-Wund, A.; Pigault, C.;
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Naenna, T.; Prachayasittikul, V. J Comput Chem 2007, 28, 1275.
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1249, 1168, 1026, 923, 755 cmꢁ1
.
1H NMR (400 MHz, DMSO) (d, ppm): 12.11 (s, 1H, NH),
8.92 (d, J ¼ 8.0 Hz, 1H, ArH), 8.17 (d, J ¼ 8.0 Hz, 2H,
ArH), 7.65–7.58 (m, 3H, ArH), 7.46–7.41 (m, 2H, ArH), 7.13–
7.10 (d, J ¼ 7.6 Hz, 1H, ArH), 7.09 (s, 1H, CH), 3.92 (s, 3H,
OCH3).
Anal calcd. For C17H14N2O2, C, 73.37; H, 5.07; N, 10.07;
found C, 73.42; H, 5.11; N, 10.00%.
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USA 1994, 91, 12501.
4-(4-Methoxybenzylidene)-2-phenyl-1H-imidazol-5(4H)-one
(3h). IR (KBr): 3121, 3065, 2969, 1699, 1596, 1456, 1304,
1264, 1172, 1028, 922, 827, 690 cmꢁ1
.
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1H NMR (400 MHz, DMSO) (d, ppm): 12.03 (s, 1H, NH),
8.31 (d, J ¼ 8.4 Hz, 2H, ArH), 8.16 (d, J ¼ 7.8 Hz, 2H,
ArH), 7.65–7.58 (m, 3H, ArH), 7.08 (d, J ¼ 8.0 Hz, 2H,
ArH), 7.02 (s, 1H, CH), 3.84 (s, 3H, OCH3).
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Anal calcd. For C17H14N2O2, C, 73.37; H, 5.07; N, 10.07;
found C, 73.30; H, 5.17; N, 9.98%.
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1984, 40, 2395.
2-Phenyl-4-((thiophen-2-yl)methylene)-1H-imidazol-5(4H)-
one (3i). IR (KBr): 3116, 3061, 2985, 1698, 1634, 1457, 1419,
[13] Cornforth, J. W.; Huang, H. T. J Chem Soc 1948, 1960.
[14] Lehr, H.; Karlan, S.; Goldberg, M. W. J Am Chem Soc
1953, 3640.
1320, 1256, 1200, 1121, 922, 891, 788, 692 cmꢁ1
.
1H NMR (400 MHz, DMSO) (d, ppm): 12.06 (s, 1H, NH),
8.16 (dd, J1 ¼ 8.0 Hz, J2 ¼ 1.2 Hz, 2H, ArH), 7.92 (d, J ¼
5.2 Hz, 1H, ArH), 7.74 (d, J ¼ 2.7 Hz, 1H, ArH), 7.64–7.60
(m, 3H, ArH), 7.39 (s, 1H, CH), 7.21–7.18 (m, 1H, ArH).
HRMS (ESI): m/z [MþH]þ calcd for C14H10N2OS:
255.0587; found: 255.0580.
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Anal calcd. For C14H10N2OS, C, 66.12; H, 3.96; N, 11.02;
found C, 66.19; H, 3.90; N, 11.11%.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet