Synthesis of model chromophores related to the gold fluorescent protein (GdFP)

Article abstract of DOI:10.1055/s-2007-965945

The two model chromophores 2 and 3 for the core 1 of the gold fluorescent protein (GdFP) were synthesized from commercially available 2-methyl-3- nitroaniline (4) in six synthetic steps and overall yields of 13% and 8%, respectively. The key step of the sequence is the chemoselective, reductive introduction of the amino group after assembly of the Z-configured 5-(indol-3-ylmethylene)imidazolin-4-one skeleton of the chromophore. Compound (Z)-2 was shown to undergo a light-initiated E/Z-isomerization, which allows access also to its E-isomer. Georg Thieme Verlag Stuttgart.

Full text of DOI:10.1055/s-2007-965945

PAPER
1103
Synthesis of Model Chromophores Related to the Gold Fluorescent Protein
(GdFP)
G
dFPChromoph
i
Lehrstuhl für Organische Chemie I, Technische Universität München, Lichtenbergstr. 4, 85747 Garching, Germany
Fax +49(89)28913315; E-mail: thorsten.bach@ch.tum.de
Received 19 January 2007
NH2
O
R
Abstract: The two model chromophores 2 and 3 for the core 1 of
the gold fluorescent protein (GdFP) were synthesized from com-
mercially available 2-methyl-3-nitroaniline (4) in six synthetic
steps and overall yields of 13% and 8%, respectively. The key step
of the sequence is the chemoselective, reductive introduction of the
amino group after assembly of the Z-configured 5-(indol-3-ylmeth-
ylene)imidazolin-4-one skeleton of the chromophore. Compound
(Z)-2 was shown to undergo a light-initiated E/Z-isomerization,
which allows access also to its E-isomer.
O
N
NH2
N
N
O
N
HN
N
HN
OH
H
1
2
3
R = Me
R = (CH2)6NHBoc
Key words: aldol reactions, bioorganic chemistry, heterocycles,
indoles, photochemistry
Figure 1 The structure of the chromophore 1 in GdFP as formed
from the amino acid triad threonine/4-aminotryptophan/glycine and
the structure of the model chromophores 2 and 3 synthesized in this
study
The green fluorescent protein (GFP) has revolutionized
almost all areas of the biological sciences.1 The visualiza-
tion of proteins and protein arrays by its fluorescence has
led to literally hundreds of applications in biology, bio-
chemistry and medicine.2 Biosynthetically, the in vivo
formation of the GFP chromophore from the amino acid
sequence serine/tyrosine/glycine is well understood3 and
model chromophores of GFP and related proteins have
been previously prepared by chemical synthesis.4
chromophore into a given protein via an amide linkage.
The syntheses of compounds 2 and 3 are described in this
paper.
While there is precedence for the Erlenmeyer azlactone
synthesis7,8 of indoles9 and the subsequent transformation
into the respective imidazolin-4-one,4,10 the appropriate
introduction of the amino group was an open question. We
found that it is beneficial to introduce the amino group re-
ductively at the very end of the reaction sequence
(Scheme 1). This route allowed us to start from commer-
cially available 2-methyl-3-nitroaniline (4) and to follow
the procedure by Bergman et al.11 for the preparation of al-
dehyde 6 via 4-nitroindole (5). Despite extensive optimi-
zation the formation of azlactone 7 proceeded only in
moderate yields. The best yield was obtained if acetylgly-
cine, NaOAc and Ac2O were stirred for an hour at 80 °C
prior to addition of aldehyde 6.
The search for new fluorescent protein probes is – among
others – driven (a) by the desired variation of the fluores-
cence wavelength at similar excitation wavelengths and
(b) by the need for additional information to be collected
by the fluorescent probe, e.g., about the polarity of its bio-
logical environment. In this respect the recent discovery
of a gold fluorescent protein (GdFP) by Budisa et al. is re-
markable.5 The protein was generated by selective pres-
sure incorporation6 of the non-canonical amino acid 4-
aminotryptophan. It contains the chromophore
1
The concomitant N-acetylation of indole in the azlactone
synthesis could be suppressed if Ca(OAc)2 instead of
NaOAc was used as a base.9e Yields were lower, however,
and we consequently decided to employ the NaOAc
method9b with the N-deacetylation being likely to occur
upon subsequent treatment with amines. Azlactone 7 was
obtained as a single diastereoisomer, to which the Z-con-
figuration was assigned based on analogy to previous
work.9,12 Further evidence for this assignment was ob-
tained in the course of the conversion of azlactone 7 into
products 8 and 9 (vide infra). For our purpose methyl-
amine and N-tert-butoxycarbonyl(Boc) protected 1,6-
diaminohexane13 were used as amine nucleophiles. Since
the amines not only displace the oxygen atom in the het-
erocycle but should also deacetylate the indole fragment
they were used in excess (3.5 equiv). Yields for imidazo-
(Figure 1), which is formed from the protein sequence
threonine/4-aminotryptophan/glycine. The protein exhib-
its in aqueous solution an absorption maximum at 466 nm
(20 °C, pH 8) and an emission maximum at 574 nm5 with
a remarkably high Stokes shift of ca. 4000 cm–1. This
property invites a closer inspection of the chromophore
and its photophysical behavior. The 5-(alkylidene)imida-
zolin-4-one 2 was considered as a reasonable model to
elucidate the origin of the Stokes shift and to further study
the unrestricted chromophore under varying conditions.
In addition, model compound 3 was chosen as a synthetic
target because it would allow the incorporation of the
SYNTHESIS 2007, No. 7, pp 1103–1106
x
x
.
x
x
.2
0
0
7
Advanced online publication: 20.02.2007
DOI: 10.1055/s-2007-965945; Art ID: Z01507SS
© Georg Thieme Verlag Stuttgart · New York

1104
B. Prüger, T. Bach
PAPER
NO2
(Figure 3). The compound exhibits a hypsochromic ex-
tinction maximum relative to GdFP (Tris-buffer: 20 mM
tris(hydroxymethyl)aminomethane HCl, pH 8) with the
hypsochromic shift depending on the solvent. Absorption
maxima were determined to be at 417 nm in MeCN, at
430 nm in H2O and at 442 nm in EtOH as the solvent. The
peak width at half-height of the absorption band is about
4700 cm–1. It is slightly larger than the peak width at half-
height of the GdFP band but independent of the solvent.
Similarly, the extinction coefficient e is identical in EtOH
and MeCN (ca. 10000 L mol–1 cm–1). The fluorescence
properties of model compound 2 are currently being stud-
ied.
NO2
1. HC(OEt)3 [TsOH]
2. (COOEt)2, KOEt
1. DMF, POCl3
2. NaOH
N
H
88%
66%
NH2
4
5
O
OH
O
O
N
N
H
O
NO2
O
NO2
NaOAc (Ac2O)
140 °C, 30 min
H
N
N
43%
H
O
6
7
O
R
N
NO2
H2 [Lindlar]
(MeOH, EtOAc)
r.t., 4 h
H2N−R, K2CO3
(EtOH)
80 °C, 16 h
N
2
3
N
H
55%
35%
97%
95%
R = Me
8
9
R = (CH2)6NHBoc
Scheme 1 Synthesis of model compounds 2 and 3 starting from
commercially available nitroaniline 4
Figure 3 Comparison between the normalized absorption spectra of
GdFP and of the model chromophore 2 in various solvents
lin-4-ones 8 and 9 were expectedly in the moderate
range.4,10 The reaction proceeds stepwise, i.e., via nucleo-
philic amine-induced azlactone ring opening and subse-
quent ring closure to the imidazolinone. Using N-
methylamine as amine, ring-opened intermediate 10
(Figure 2) could be isolated if the reaction was stopped af-
ter four hours. The assignment of the two distinct amide
NH protons was possible and NOE experiments estab-
lished the spatial proximity of the NHAc group and the
proton at C-2 of the indole.
Upon irradiation at 419 nm (irradiation source: RPR
4190 Å) product (Z)-2 underwent geometrical isomeriza-
tion into the corresponding E-isomer (Scheme 2). Under
our conditions (DMSO-d6 as solvent, 35 °C) a photosta-
tionary state was reached after 30 minutes. The Z/E ratio
was determined by NMR as 60:40. There was no indica-
tion for other photochemical processes.
O
hν (λ = 419 nm)
30 min
N
N
N
O
NH2
NH2
H
N
δ (NHMe):
δ (NHAc):
δ (H-C2):
7.73 ppm (q)
7.91 ppm (s)
9.16 ppm (s)
O
N
(DMSO-d6)
NO2
O
N
Z/E = 60:40
H
N
N
2
H
H
H
10
N
(Z)-2
(E)-2
H
Figure 2 Chemical shift data for the relevant protons in compound
10 and observed NOE contact
Scheme 2 E/Z-Isomerization of compound (Z)-2
In conclusion, the model chromophores 2 and 3 were ob-
tained by conventional condensation and functional group
transformation chemistry. The azlactone formation and
nitro group reduction were closely studied. Optimized
conditions are reported for these transformations. Further
photophysical and biophysical data of the chromophores
will be reported in due course.
The reduction of the nitro group was eventually conducted
with hydrogen in the presence of Lindlar catalyst (5%
Pd/CaCO3/Pb). Other attempts to produce 4-aminoindoles
2 and 3 from the nitroindoles 8 and 9 met with little suc-
cess. Reduction experiments with hydrogen and other cat-
alysts or with SnCl2 in EtOH and EtOAc led to partial or
complete degradation.
Product 2 was obtained as intensely red colored solid, and
3 as equally intense oil. They are well soluble in organic
All reactions involving water-sensitive chemicals were carried out
in flame-dried glassware with magnetic stirring under argon. All
solvents and UV spectra of compound 2 were recorded solvents, EtOAc, CH2Cl2 and MeOH for column chromatography
Synthesis 2007, No. 7, 1103–1106 © Thieme Stuttgart · New York

PAPER
GdFP Chromophore
1105
were distilled prior to use. Acetylglycine was prepared from glycine
according to literature precedence.14 N-(tert-Butoxycarbonyl)-1,6-
diaminohexane was obtained from 1,6-diaminohexane.13 All other
chemicals were commercially available and were used without fur-
ther purification.
MS (EI, 70 eV): m/z (%) = 284 (55, [M+]), 238 (86), 232 (11), 154
(13), 130 (18), 56 (100), 44 (11).
HRMS (EI): m/z calcd for C14H12N4O3: 284.0909; found: 284.0910.
UV/Vis (MeCN): lmax (log e) = 377 nm (4.35).
TLC: Merck glass sheets (0.25 mm silica gel 60, F254), eluent given
in brackets. Detection by UV or coloration with cerium ammonium
molybdate (CAM). NMR: Bruker AV-250, AV-360, AV-500. H
Isolation of 2-Acetylamino-3-(indol-3-yl)-N-methylacrylamide
(10)
Upon premature (4 h) addition of H2O to the above-mentioned re-
action mixture (preparation of 8), a bright yellow solid precipitated,
which was not soluble neither in the aqueous nor in the organic lay-
er. The precipitate was filtered, washed with H2O and CH2Cl2, and
dried in vacuo; mp >250 °C (dec.).
1H NMR (250 MHz): d = 2.00 (s, 3 H), 2.67 (d, J = 4.5 Hz, 3 H),
7.32 (t, J = 8.2 Hz, 1 H), 7.42 (s, 1 H), 7.73 (q, J = 4.5 Hz, 1 H), 7.82
(d, J = 8.2 Hz, 1 H), 7.84 (d, J = 8.2 Hz, 1 H), 7.91 (s, 1 H), 9.16 (s,
1 H), 12.28 (br s, 1 H).
13C NMR (62.9 MHz): d = 23.0 (q), 26.1 (q), 107.8 (s), 117.6 (d),
117.8 (s), 118.3 (d), 120.8 (d), 122.3 (d), 126.2 (s), 130.7 (d), 138.1
(s), 142.1 (s), 165.3 (s), 169.4 (s).
1
and 13C NMR spectra were recorded in DMSO-d6 at ambient tem-
perature, unless stated otherwise. Chemical shifts are reported rela-
tive to tetramethylsilane as internal standard. Apparent multiplets
which occur as a result of the accidental equality of coupling con-
stants of magnetically nonequivalent protons are marked as virtual
(virt.). The multiplicities of the 13C NMR signals were determined
by DEPT experiments. IR: PerkinElmer 1600 FT-IR. MS: Finnigan
MAT 8200 (EI). UV/Vis: PerkinElmer Lambda 9, Lambda 35.
Flash chromatography was performed on silica gel 60 (Merck, 230–
400 mesh) (ca. 50 g for 1 g of material to be separated) with the in-
dicated eluent.
4-(N-Acetyl-4-nitroindol-3-ylmethylene)-2-methyloxazolin-5-
one (7)
3-[N-(tert-Butoxycarbonyl)-6-aminohexyl]-2-methyl-5-(4-ni-
troindol-3-ylmethylene)imidazolin-4-one (9)
A suspension of acetylglycine14 (358 mg, 3.06 mmol) and NaOAc
(233 mg, 2.84 mmol) in Ac2O (2.50 mL, 2.72 g, 26.6 mmol) was
stirred for 1 h at 80 °C. 3-Formyl-4-nitroindole (6; 500 mg,
2.63 mmol) was added and the mixture was stirred at 140 °C for
30 min. The dark red mixture was cooled overnight in the refriger-
ator (4 °C). The brownish precipitate was filtered, washed with H2O
and dried in vacuo at 50 °C. Purification by flash chromatography
(CH2Cl2–MeOH, 99:1) yielded 306 mg (43%) of azlactone 7 as yel-
low needles; mp 228–235 °C (dec.); Rf = 0.84 (CH2Cl2–
MeOH, 95:5).
Following the procedure for imidazolinone 8, azlactone 7 (407 mg,
1.30 mmol) was reacted with N-(tert-butoxycarbonyl)-1,6-
diaminohexane13 (986 mg, 4.55 mmol) and K2CO3 (400 mg,
2.89 mmol) in EtOH (12 mL). After purification by flash chroma-
tography (EtOAc), 9 (211 mg, 35%) was obtained as a yellow solid;
mp 125–144 °C; Rf = 0.39 (CH2Cl2–MeOH, 95:5).
IR (KBr): 3294 (w), 3159 (m), 2972 (m), 2930 (s), 2858 (m), 1681
(s), 1631 (s), 1558 (m), 1517 (s), 1355 (s), 1324 (s), 1170 (m), 785
(m), 736 (m), 611 cm–1 (m).
1H NMR (360 MHz): d = 1.23–1.31 (m, 4 H), 1.36 (br s, 11 H),
1.48–1.59 (m, 2 H), 2.36 (s, 3 H), 2.89 (q, J = 6.4 Hz, 2 H), 3.54 (t,
J = 7.2 Hz, 2 H), 6.69–6.78 (m, 1 H), 7.38 (virt. t, J = 7.9 Hz, 1 H),
7.45 (s, 1 H), 7.91 (dd, J = 7.9, 0.9 Hz, 1 H), 7.93 (dd,
J = 7.9, 0.9 Hz, 1 H), 8.93 (s, 1 H), 12.65 (s, 1 H).
IR (KBr): 1799 (s), 1722 (s), 1650 (s), 1567 (m), 1519 (s), 1341 (s),
965 (m), 794 (m), 747 (m), 666 cm–1 (m).
1H NMR (360 MHz): d = 2.40 (s, 3 H), 2.79 (s, 3 H), 7.41 (s, 1 H),
7.63 (virt. t, J = 8.2 Hz, 1 H), 8.08 (dd, J = 8.2, 0.9 Hz, 1 H), 8.80
(dd, J = 8.2, 0.9 Hz, 1 H), 8.98 (s, 1 H).
13C NMR (90.6 MHz): d = 15.4 (q), 24.1 (q), 112.2 (s), 120.1 (s),
121.0 (d), 121.7 (d), 122.3 (d), 125.5 (d), 131.4 (s), 135.8 (d), 136.8
(s), 142.7 (s), 166.3 (s), 166.7 (s), 169.8 (s).
13C NMR (90.6 MHz): d = 15.3 (q), 25.9 (t), 28.2 (q), 29.3 (t), 39.7
(t), 77.2 (s), 109.1 (s), 117.8 (s), 118.7 (d), 119.1 (d), 119.6 (d),
121.5 (d), 134.8 (s), 136.5 (d), 138.7 (s), 142.3 (s), 160.8 (s), 169.2
(s).
MS (EI, 70 eV): m/z (%) = 313 (39, [M+]), 271 (40), 225 (6), 201
MS (EI, 70 eV): m/z (%) = 469 (1, [M+]), 439 (1), 413 (1), 368 (1),
323 (1), 266 (1), 241 (2), 224 (3), 197 (4), 162 (5), 132 (3), 116 (5),
112 (4), 98 (4), 86 (3), 56 (33), 44 (34), 41 (100), 39 (31).
(13), 184 (13), 154 (14), 145 (14), 44 (19), 43 (100).
HRMS (EI): m/z calcd for C15H11N3O5: 313.0699; found: 313.0701.
2,3-Dimethyl-5-(4-nitroindol-3-ylmethylene)imidazolin-4-one
(8)
HRMS (EI): m/z calcd for C24H31N5O5: 469.2325; found: 469.2370.
UV/Vis (MeCN): lmax (log e) = 377 nm (4.46).
K2CO3 (517 mg, 3.74 mmol) was added to a suspension of azlac-
tone 7 (531 mg, 1.70 mmol) and methylamine (40% in H2O,
0.51 mL, 459 mg, 5.94 mmol) in EtOH (17 mL). The mixture was
stirred for 16 h at 80 °C. After addition of H2O (25 mL), the dark
red solution was extracted with CH2Cl2 (5 × 25 mL). The combined
organic layers were washed with brine (75 mL), dried (MgSO4) and
filtered. The product was concentrated in vacuo and purified by
flash chromatography (CH2Cl2–MeOH, 95:5). Compound 8 was
obtained as a dark red solid; yield: 265 mg (55%); mp 240–248 °C
(dec.); Rf = 0.18 (CH2Cl2–MeOH, 95:5).
5-(4-Aminoindol-3-ylmethylene)-2,3-dimethylimidazolin-4-one
[(Z)-2]
Compound 8 (150 mg, 528 mmol) was dissolved in MeOH (2.5 mL)
and EtOAc (2.5 mL). Lindlar catalyst (5% Pd/CaCO3/Pb,
26.4 mmol Pd; 56.0 mg) was added and the mixture was stirred vig-
orously for 4 h under a H2 atmosphere (1 atm). The mixture was fil-
tered over Celite and washed thoroughly with MeOH (200 mL).
The solvent was removed in vacuo and the residue was purified by
silica gel chromatography (EtOAc). Compound 2 was obtained as a
red solid; yield: 131 mg (97%); mp 240 °C (dec.); Rf = 0.34
(EtOAc).
IR (KBr): 3155 (m) 1677 (s), 1625 (s), 1560 (m), 1351 (s), 1326 (s),
1138 (s), 775 (m), 735 (m), 558 cm–1 (m).
1H NMR (360 MHz): d = 2.34 (s, 3 H), 3.09 (s, 3 H), 7.37 (virt. t,
J = 8.0 Hz, 1 H), 7.44 (s, 1 H), 7.91 (dd, J = 8.0, 0.9 Hz, 1 H), 8.80
(dd, J = 8.0, 0.9 Hz, 1 H), 8.93 (s, 1 H), 12.65 (s, 1 H).
13C NMR (90.6 MHz): d = 15.2 (q), 26.1 (q), 109.0 (s), 117.8 (s),
118.6 (d), 119.0 (d), 119.4 (d), 121.4 (d), 134.9 (s), 136.4 (d), 138.6
(s), 142.2 (s), 161.2 (s), 169.1 (s).
IR (KBr): 3420 (w), 3344 (m), 3160 (m), 1668 (m), 1627 (s), 1499
(s), 1426 (s), 1329 (s), 780 (m), 612 (m), 553 cm–1 (m).
1H NMR (360 MHz): d = 2.32 (s, 3 H), 3.09 (s, 3 H), 5.09 (s, 2 H),
6.43 (d, J = 7.6 Hz, 1 H), 6.77 (d, J = 7.6 Hz, 1 H), 6.89 (virt. t,
J = 7.6 Hz, 1 H), 7.56 (s, 1 H), 8.49 (d, J = 1.8 Hz, 1 H), 11.69 (s, 1
H).
Synthesis 2007, No. 7, 1103–1106 © Thieme Stuttgart · New York

1106
B. Prüger, T. Bach
PAPER
13C NMR (90.6 MHz): d = 15.2 (q), 26.1 (q), 102.2 (d), 107.6 (d),
111.4 (s), 115.2 (s), 121.9 (d), 123.2 (d), 131.7 (d), 133.3 (s), 137.5
(s), 142.6 (s), 158.7 (s), 168.9 (s).
MS (EI, 70 eV): m/z (%) = 254 (72, [M+]), 232 (9), 199 (38), 182
(5), 154 (15), 73 (6), 56 (100), 44 (23).
(2) Examples: (a) Ehrhardt, E. Curr. Opin. Plant Biol. 2003, 6,
622. (b) Chudakov, D. M.; Lukyanov, K. A. Biochemistry
(Moscow) 2003, 68, 952. (c) Michaelson, D.; Philips, M.
Methods Enzymol. 2000, 406, 296. (d)Prasher, D. C. Trends
Genet. 1995, 11, 320.
(3) (a) Shimomura, O. FEBS Lett. 1979, 104, 220. (b) Cody, C.
W.; Prasher, D. C.; Westler, W. M.; Prendergast, F. G.;
Ward, W. W. Biochemistry 1993, 32, 1212. (c) Reid, B. G.;
Flynn, G. C. Biochemistry 1997, 36, 6786.
(4) Examples: (a) Kojima, S.; Ohkawa, H.; Hirano, T.; Maki, S.;
Niwa, H.; Ohashi, M.; Inouye, S.; Tsuji, F. I. Tetrahedron
Lett. 1998, 39, 5239. (b) He, X.; Bell, A. F.; Tonge, P. J.
Org. Lett. 2002, 4, 1523. (c) Yampolsky, I. V.; Remington,
S. J.; Martynov, V. I.; Potapov, V. K.; Lukyanov, S.;
Lukyanov, K. A. Biochemistry 2005, 44, 5788. (d) Hager,
B.; Schwarzinger, B.; Falk, H. Monatsh. Chem. 2006, 137,
163.
HRMS (EI): m/z calcd for C14H14N4O: 254.1168; found: 254.1172.
UV/Vis (MeCN): lmax (log e) = 419 nm (3.99).
UV/Vis (EtOH): lmax (log e) = 438 nm (4.00).
E/Z-Isomerization of 5-(4-Aminoindol-3-ylmethylene)-2,3-di-
methylimidazolin-4-one
(Z)-2 was dissolved in DMSO-d6 in a NMR tube and irradiated for
30 min at 419 nm (light source: RPR 4190 Å, 35 °C). An equilibri-
um was established between the two isomers, with an E/Z ratio of
40:60.
(Z)-2 (60%)
(5) Bae, J. H.; Rubini, M.; Jung, G.; Wiegand, G.; Seifert, M. H.
J.; Azim, M. K.; Kim, J.-S.; Zumbusch, A.; Holak, T. A.;
Moroder, L.; Huber, R.; Budisa, N. J. Mol. Biol. 2003, 328,
1071.
(6) (a) Budisa, N.; Rubini, M.; Bae, J. H.; Weyher, E.; Wenger,
W.; Golbik, R.; Huber, R.; Moroder, L. Angew. Chem. Int.
Ed. 2002, 41, 4066. (b) Budisa, N. ChemBioChem 2004, 5,
1176. (c) Budisa, N.; Pal, P. P. Biol. Chem. 2004, 385, 893.
(7) (a) Erlenmeyer, E. Jr. Liebigs Ann. Chem. 1893, 275, 1.
(b) Erlenmeyer, E. Jr.; Früstück, E. Liebigs Ann. Chem.
1895, 284, 36.
1H NMR (360 MHz): d = 2.32 (s, 3 H), 3.09 (s, 3 H), 5.09 (s, 2 H),
6.43 (d, J = 7.6 Hz, 1 H), 6.77 (d, J = 7.6 Hz, 1 H), 6.89 (virt. t,
J = 7.6 Hz, 1 H), 7.56 (s, 1 H), 8.49 (d, J = 1.8 Hz, 1 H), 11.69 (s, 1
H).
(E)-2 (40%)
1H NMR (360 MHz): d = 2.25 (s, 3 H), 3.14 (s, 3 H), 5.12 (s, 2 H),
6.46 (d, J = 7.6 Hz, 1 H), 6.79 (d, J = 7.6 Hz, 1 H), 6.91 (virt. t,
J = 7.6 Hz, 1 H), 7.96 (s, 1 H), 9.38 (d, J = 2.8 Hz, 1 H), 11.79 (s, 1
H).
(8) Reviews: (a) Rosen, T. In Comprehensive Organic
Synthesis, Vol. 2; Trost, B.; Fleming, I.; Heacthcock, C. H.,
Eds.; Pergamon: Oxford, 1991, 395–408. (b) Mukerjee, A.
K. Heterocycles 1987, 26, 1077. (c) Mukerjee, A. K.;
Kumar, P. Heterocycles 1981, 16, 1995. (d) Filler, R. Adv.
Heterocycl. Chem. 1965, 4, 75. (e) Cornforth, J. W. In
Heterocyclic Compounds, Vol. 5; Elderfield, R. C., Ed.;
Wiley: New York, 1957, 298–417. (f) Baltazzi, E. Q. Rev.
Chem. Soc. 1955, 9, 150. (g) Cornforth, J. W. In Chemistry
of Penicillin; Clarke, H. T.; Johnson, J. R.; Robinson, R.,
Eds.; Princeton University Press: Princeton, N. J., 1948,
688–848. (h) Carter, H. E. Org. React. 1947, 3, 198.
(9) (a) Ojima, I.; Kato, K.; Nakahashi, K.; Fuchikami, T.; Fujita,
M. J. Org. Chem. 1989, 54, 4511. (b) Oba, M.; Ueno, R.;
Fukuoka, M.; Kainosho, M.; Nishiyama, K. J. Chem. Soc.,
Perkin Trans. 1 1995, 1603. (c) Donati, D.; Garzon-
Aburbeh, A.; Natalini, B.; Marchioro, C.; Pellicciari, R.
Tetrahedron 1996, 52, 9901. (d) Mesaik, M. A.; Rahat, S.;
Khan, K. M.; Zia-Ullah; Choudhary, M. I.; Murad, S.;
Ismail, Z.; Atta-ur-Rahman; Ahmad, A. Bioorg. Med.
Chem. 2004, 12, 2049. (e) Guella, G.; N’Diaye, I.; Fofana,
M.; Marcini, I. Tetrahedron 2006, 62, 1165.
5-(4-Aminoindol-3-ylmethylene)-3-[N-(tert-butoxycarbonyl)-6-
aminohexyl]-2-methylimidazolin-4-one (3)
Following the procedure for chromophore 2, imidazolinone 9
(130 mg, 277 mmol) and Lindlar catalyst (5% Pd/CaCO3/Pb,
13.9 mmol Pd; 29.5 mg) in MeOH (1.5 mL) and EtOAc (1.5 mL)
were stirred vigorously for 4 h under a H2 atmosphere. After work-
up and purification by flash chromatography (EtOAc), chro-
mophore 3 (117 mg, 95%) was obtained as a dark red oil; Rf = 0.54
(EtOAc–MeOH, 90:10).
1H NMR (360 MHz): d = 1.18–1.29 (m, 4 H), 1.37 (s, 11 H), 1.47–
1.60 (m, 2 H), 2.34 (s, 3 H), 2.89 (q, J = 6.5 Hz, 2 H), 3.54 (t,
J = 7.2 Hz, 2 H), 5.11 (br s, 1 H), 6.44 (dd, J = 7.7, 0.8 Hz, 1 H),
6.74 (s, 2 H), 6.78 (dd, J = 7.7, 0.8 Hz, 1 H), 6.89 (virt. t, J = 7.7 Hz,
1 H), 7.57 (s, 1 H), 8.49 (d, J = 2.7 Hz, 1 H), 11.72 (d, J = 2.7 Hz, 1
H).
13C NMR (90.6 MHz): d = 15.2 (q), 25.9 (t), 26.1 (t), 28.2 (q), 28.8
(t), 29.1 (t), 29.4 (t), 39.7 (t), 77.2 (s), 102.2 (d), 107.7 (d), 111.5 (s),
115.3 (s), 122.1 (d), 123.2 (d), 131.8 (d), 133.1 (s), 137.6 (s), 142.6
(s), 155.6 (s), 158.2 (s), 168.9 (s).
HRMS (EI): m/z calcd for C24H33N5O3: 439.2583; found: 439.2576.
UV/Vis (MeCN): lmax (log e) = 421 nm (4.20).
(10) (a) Erlenmeyer, E. Jr.; Wittenberg, F. Liebigs Ann. Chem.
1904, 337, 294. (b) Narang, K. S.; Ray, J. N. J. Chem. Soc.
1931, 976. (c) Williams, D. L.; Ronzio, A. R. J. Am. Chem.
Soc. 1946, 68, 647. (d) Pfleger, R.; Markert, G. Chem. Ber.
1957, 90, 1494.
Acknowledgment
(11) (a) Bergman, J.; Sand, P. Org. Synth. 1987, 65, 146.
(b) Melhado, L. L.; Brodsky, J. L. J. Org. Chem. 1988, 53,
3852. (c) Bergman, J.; Sand, P. Tetrahedron 1990, 46, 6085.
(12) Rao, Y. S.; Filler, R. Synthesis 1975, 749.
We thank Dr. N. Budisa, Prof. M. E. Michel-Beyerle, and Prof. A.
Skerra for helpful discussions. Financial support by the Deutsche
Forschungsgemeinschaft (SFB 533) is gratefully acknowledged.
(13) (a) Byk, G.; Gilon, C. J. Org. Chem. 1992, 57, 5687.
(b) Muller, D.; Zeltser, I.; Bitan, G.; Gilon, C. J. Org. Chem.
1997, 62, 411. (c) Pons, J.-F.; Fauchière, J.-L.; Lamaty, F.;
Molla, A.; Lazaro, R. Eur. J. Org. Chem. 1998, 853.
(14) Herbst, R. M.; Shemin, D. Org. Synth. Coll. Vol. II; Wiley:
New York, 1943, 11–12.
References
(1) Reviews: (a) Zimmer, M. Chem. Rev. 2002, 102, 759.
(b) Tsien, R. Y. Annu. Rev. Biochem. 1998, 67, 509.
Synthesis 2007, No. 7, 1103–1106 © Thieme Stuttgart · New York