Synthesis of fluorescent D-amino acids with 4-acetamidobiphenyl and 4-N,N-dimethylamino-1,8-naphthalimido containing side chains

Article abstract of DOI:10.1016/j.tetlet.2015.06.059

We report the synthesis and fluorescence properties of two aromatic D-amino acids. The key step for the synthesis of (R)-2-amino-3-(4′-acetamido-[1,1′-biphenyl]-4-yl)propanoic acid was a Suzuki cross-coupling reaction between the pinacol diester of N-acetylphenylboronic acid and Fmoc-D-p-bromo-phenylalanine. The second amino acid, (R)-2-amino-4-(4′-N,N-dimethylamino-1,8-naphthalimido)butanoic acid [(R)-2-amino-4-DMNA-butanoic acid], was synthesized in four steps from 4-bromo-1,8-naphthalic anhydride. The amino acids were prepared in a form that could be readily incorporated into a peptide sequence by solid phase peptide synthesis using the Fmoc-strategy. Evaluation of the fluorescence properties of the biphenyl amino acid showed a maximum emission at 384 nm (excitation at 295 nm) while the naphthalimido amino acid showed a maximum emission at 545 nm (excitation at 450 nm).

Full text of DOI:10.1016/j.tetlet.2015.06.059

Tetrahedron Letters 56 (2015) 4780–4783
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Synthesis of fluorescent D-amino acids with 4-acetamidobiphenyl
and 4-N,N-dimethylamino-1,8-naphthalimido containing side chains
⇑
⇑
Jyotirmoy Maity, Dmytro Honcharenko , Roger Strömberg
Department of Biosciences and Nutrition, Karolinska Institute, Novum, Huddinge SE-141 83, Sweden
a r t i c l e i n f o
a b s t r a c t
Article history:
We report the synthesis and fluorescence properties of two aromatic D-amino acids. The key step for
thesynthesisof(R)-2-amino-3-(4 -acetamido-[1,1 -biphenyl]-4-yl)propanoicacidwasaSuzukicross-coupling
Received 24 February 2015
Revised 28 May 2015
Accepted 19 June 2015
Available online 26 June 2015
0
0
reaction between the pinacol diester of N-acetylphenylboronic acid and Fmoc-
D
-p-bromo-phenylalanine.
0
The second amino acid, (R)-2-amino-4-(4 -N,N-dimethylamino-1,8-naphthalimido)butanoic acid [(R)-2-
amino-4-DMNA-butanoic acid], was synthesized in four steps from 4-bromo-1,8-naphthalic anhydride.
The amino acids were prepared in a form that could be readily incorporated into a peptide sequence by solid
phase peptide synthesis using the Fmoc-strategy. Evaluation of the fluorescence properties of the biphenyl
amino acid showed a maximum emission at 384 nm (excitation at 295 nm) while the naphthalimido amino
acid showed a maximum emission at 545 nm (excitation at 450 nm).
Keywords:
Aromatic amino acids
Fluorescent amino acids
Biphenyl amino acid
Naphthalimido amino acid
Ó 2015 Elsevier Ltd. All rights reserved.
In recent years, fluorescence spectroscopy techniques have
been applied as powerful tools for studying the structure and func-
tion of proteins. Unnatural aromatic amino acids with fluorescence
properties are widely used for this purpose and have been
3-(6-acetylnaphthalen-2-ylamino)-2-aminopropionic acid (Anap).14
The assortment of fluorescent amino acids with diverse properties
has enabled researchers to examine the biological processes of
protein binding, trafficking and localization and also protein
1
15
deployed as a fluorescent labels at the N-terminus of proteins,
conformational studies involving them.
2
utilized for FRET analysis of protein conformation change,
To access libraries of unnatural aromatic amino acids with differ-
ent fluorescent properties and/or biological activities, there is still a
demand for the synthesis of new aromatic amino acids. Cost effec-
3
incorporated into proteins as alternative chromophores, placed
in dihydrofolate reductase to study its conformational change
4
upon inhibitor binding, included in a model peptide system villin
tive, enantioselective and efficient synthetic protocols have been
5
16–19
headpiece HP35 fragment to quench various fluorophores, used as
developed for the synthesis of coumarin amino acids,
5-substi-
6
a substrate to establish a sensitive assay procedure for
acid oxidase activity in mammalian tissues, and incorporated into
the molecular chaperone GroEL for its in vivo visualization.
D
-amino
tuted tryptophans, Fmoc-L-diaminopropyl-(7-nitrobenz-2-oxa-
6
20
21
1,3-diazol-4-yl)-OH, benz-X-asparagine derivatives, bis-alanine
derivatives bearing (oligo)thiophene and benzoxazole units as the
7
2
2,23
Modified peptides with unnatural fluorescent amino acids have
heteroaromatic bridge,
alanine-substituted benzoacridone
derivatives, and amino acid derivatives of 4-ethoxymethylene-
2-[1]-naphthyl-5(4H)-oxazolone. However, among the syntheti-
8
24
been used to screen and quantify EGF receptor-binding peptides
and in the study of dynamic protein interactions.9 An unnatural
fluorescent amino acid Lys(BODIPYFL) was incorporated into the
mouse muscle nicotinic acetylcholine receptor (nAChR) b-subunit
25
cally accessible amino acids with fluorescence properties, very
few are
D-amino acids. As an example, the aromatic amino acid
0
0
M2 domain 19 position (b19 ) and studied for cell-based single
D-tryptophan has been synthesized and evaluated for its biological
1
0
25
molecule detection. Efficient methods have even been developed
to genetically encode unnatural amino acids for incorporation
into proteins to enhance their properties. Among these, there
significance showing an enhancement of MC receptor potency
2
6
when substituted in place of L-Trp.
The broad range of applications of unnatural aromatic amino
acids in biochemistry encouraged us to explore the synthesis of
are quite
a few unnatural aromatic amino acids, such as
the para-substituted derivatives of phenyl alanine,1
1–13
2-
amino-3-(8-hydroxyquinolin-5-yl)propanoic acid (HqAla), and
two novel aromatic D-amino acids. Herein we describe the synthe-
1
2
0
0
sis of (R)-2-amino-3-(4 -acetamido-[1,1 -biphenyl]-4-yl)propanoic
0
acid (1) and (R)-2-amino-4-(4 -N,N-dimethylamino-1,8-naphthal-
imido)butanoic acid [(R)-2-amino-4-DMNA-butanoic acid] (2) in
their Fmoc-protected form, compounds 6 and 11, respectively,
making them suitable for incorporation into a peptide sequence
⇑
Corresponding authors.
E-mail addresses: dmytro.honcharenko@ki.se (D. Honcharenko), roger.
stromberg@ki.se (R. Strömberg).
http://dx.doi.org/10.1016/j.tetlet.2015.06.059
0
040-4039/Ó 2015 Elsevier Ltd. All rights reserved.

J. Maity et al. / Tetrahedron Letters 56 (2015) 4780–4783
4781
NHFmoc
O
B
O
B
COOH
5
Br
(i)
(ii)
O
O
H N
H COCHN
2
3
3
4
NHFmoc
NH2
COOH
(iii)
COOH
6
1
H
3
COCHN
H COCHN
3
2 2 2 3 3
Scheme 1. Reagents and conditions: (i) Ac (5 mol %), THF–ethylene glycol (10:1), Na CO , 66 °C, 3 h, 81%; (iii) 33% aq NH , 55 °C, 18 h, quant.
O, rt, 1 h, quant;29 (ii) 5, PdCl
Table 1
Standardization of reaction conditions for the Suzuki coupling reaction of 4 and 5
Entry
Solvent system
Catalyst (5 mol %)
Time (h)
Base
NaH
Temp (°C)
Yield of 6 (%)
1
2
3
4
5
6
7
THF–H
THF–H
THF
2
O (10:1)
O (10:1)
10% Pd/C
10% Pd/C
16
16
4
4
4
2
PO
4
/Na
2
CO
3
(2:1)
22
22
66
66
66
66
66
—
—
2
Na
3
Na
3
Na
2
PO
PO
CO
4
a
Pd(PPh
Pd(PPh
3
)
)
4
4
ꢂ40
a
THF
THF
3
4
3
ꢂ40
a
PdCl
PdCl
PdCl
2
2
2
K
K
2
CO
CO
3
ꢂ50
b
THF–ethylene glycol (10:1)
THF–ethylene glycol (10:1)
3
3
2
3
72
b
Na
2
CO
3
81
a
Product formation as judged by visualization by TLC under UV (254 nm).
isolated yield.
b
using the solid phase peptide synthesis (SPPS) Fmoc-strategy.27,28
The fluorescence properties of these unnatural aromatic amino
acids in their unprotected form were also evaluated.
compound 8 precipitated from the reaction mixture to afford suf-
ficiently pure product that was used in the next step without
2
purification. Anhydride 8 was reacted with N -(tert-butoxycar-
Our strategy for the synthesis of biphenyl aromatic amino acid 6
began with acetylation of 4-(4,4,5,5-tetramethyl-1,3,2-diox-
aborolan-2-yl)aniline (3) at the para-amino position to produce
compound 4 (Scheme 1). Subsequently, the biphenyl moiety was
constructed by a Suzuki cross-coupling reaction between the
bonyl)-2,4-diaminobutanoic acid (Boc-
D
-Dab-OH, 9) to afford
0
2
(R)-4-(4 -N,N-dimethylamino-1,8-naphthalimido)-N -(tert-butoxy-
29
40
carbonyl)-2-aminobutanoic acid (10). The Boc-protecting group
was then removed using trifluoroacetic acid in dichloromethane
to give the unprotected amino acid 2, which was used for the
characterization of its fluorescent properties. To obtain a suitable
building block for solid phase peptide synthesis using the Fmoc-
pinacol diester of 4-N-acetamidophenylboronic acid (4) and
a
Fmoc-
2
(
D
-p-bromo-phenylalanine (5) to afford N -Fmoc-(R)-
0
0
a
-amino-3-(4 -acetamido-[1,1 -biphenyl]-4-yl)propanoic acid (6)
Scheme 1).
Various reaction conditions (Table 1) for the synthesis of the
desired biphenyl compound were investigated using palladium
strategy, compound 2 was reprotected at the N position by
treatment with Fmoc-hydroxysuccinimide ester (Fmoc-OSu)41
affording compound 11 (Scheme 2).
Studies of the fluorescent properties of aromatic amino acids 1
and 2 were performed in a solvent mixture of 50% aqueous metha-
3
0,31
32–34
35–37
on carbon (Pd/C),
PdCl
2
and Pd(PPh
)
3 4
which have
ꢁ
4
ꢁ4
all been extensively used as catalysts for coupling reactions.
Negligible amounts of the desired product were obtained using
nol at concentrations of 7.7 ꢀ 10
M
and 2.9 ꢀ 10 M,
1
0% Pd/C in THF–water, and most of the starting material was
recovered (entries 1 and 2, Table 1). Different inorganic bases such
as Na PO , Na CO and K CO as well as several solvent mixtures
O
O
O
O
O
O
BocHN
COOH
O
3
4
2
3
2
3
(i)
(ii)
were examined. Decent conversions to the desired product were
obtained when the reaction was performed in refluxing THF with
NH2
O
N
9
3 4 2
Pd(PPh ) (entries 3 and 4, Table 1) and substitution of PdCl
Br
N
7
8
resulted in a higher yield (entry 5, Table 1). Changing the solvent
to THF–ethylene glycol (10:1) resulted in even higher yields of
the cross-coupling product (81%) after isolation by chromatogra-
HOOC
NHBoc
10
N
FmocHN
COOH
O
H N
COOH
O
38
2
phy (entries 6 and 7, Table 1). The acetamidobiphenyl amino acid
1
13
6
was characterized by H and C NMR spectroscopy as well as by
HRMS.
To carry out fluorescence spectroscopic characterization of par-
O
N
O
N
(iv)
(iii)
ent amino acid the Fmoc-group was removed from compound 6 by
treatment with ammonia to afford the unprotected biphenyl
amino acid 1 (Scheme 1).
0
11
N
2
N
The second aromatic amino acid synthesized was (R)-4-(4 -N,N-
dimethylamino-1,8-naphthalimido)-2-aminobutanoic acid (2)
Scheme 2. Reagents and conditions: (i) isoamylalcohol, 3-(dimethylamino)propi-
onitrile, 132 °C, 16 h, quant; (ii) 9, aq NaHCO , dioxane, reflux, 1 h, 66%; (iii)
3
(Scheme 2). 4-Bromo-1,8-naphthalic anhydride (7) was converted
39
into 8 by heating at reflux with 3-dimethylamino propionate
trifluoroacetic acid, DCM, 0 °C, 2 h; (iv) Fmoc-OSu, water–dioxane (1:1), NaHCO
3
,
3
9
in isoamyl alcohol.
Upon cooling to room temperature,
0 °C, 2 h, 71% (over two steps).

4
782
J. Maity et al. / Tetrahedron Letters 56 (2015) 4780–4783
was in the same range as other reported naphthalimido
9
,43,44
fluorophores.
A small red shift in the emission maximum
was observed for 2 when fluorescence measurements were per-
formed in pure water (maximum kem at 548 nm) compared to
the emission in less polar methanol (maximum kem at 533 nm)
(
see ESI, Fig. S14).
To demonstrate the incorporation of one of the fluorescent
amino acids into a peptide-like structure, compound 12 (Fig. 3)
was synthesized using SPPS and a recently reported triamino acid
4
5
(
see ESI, Scheme S1).
We have synthesized two unnatural fluorescent
with -Fmoc protection (and -Boc protection in one case). The
D
-amino acids
a
a
synthesis of acetamidobiphenyl amino acid 6 was achieved in
two steps staring from the pinacol ester of p-amino-phenyl boronic
acid (3) and the fluorescent amino acid 11 was obtained from
cyclic anhydride 7 in four steps. Both of these amino acids possess
different fluorophores that were found to be highly fluorescent
with properties that were distinctive and complementary in
respect to both excitation and emission wavelengths. The
acetamidobiphenyl amino acid 1 showed a maximum fluorescence
intensity at a shorter wavelength (384 nm) than the naphthalimido
amino acid (2) which had a maximum at a longer wavelength
Figure 1. Fluorescence emission spectrum of (R)-2-amino-3-(p-acetamido-
biphenyl)propanoic acid (1) in aqueous methanol (1:1) showing a maximum at
3
84 nm (excitation wavelength was 295 nm).
(
543 nm). These aromatic amino acids with significant fluores-
cence properties should be valuable for incorporation into peptide
sequences for studies of biological significance. Because they are
D
-amino acids they should be particularly valuable for incorpora-
tion into - or -peptides which are resistant to enzymatic
degradation.
D
D,L
Acknowledgment
We gratefully acknowledge financial support from AlphaBeta
and the Swedish Research Council.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.tetlet.2015.06.
059.
Figure 2. Fluorescence emission spectrum of (R)-2-amino-4-DMNA-butanoic acid
2) in aqueous methanol (1:1) showing maximum at 543 nm (excitation
wavelength was 450 nm).
(
a
References and notes
1
2
3
4
5
.
.
.
.
.
Watanabe, T.; Miyata, Y.; Abe, R.; Muranaka, N.; Hohsaka, T. ChemBioChem
2008, 9, 1235–1242.
Kajihara, D.; Abe, R.; Iijima, I.; Komiyama, C.; Sisido, M.; Hohsaka, T. Nat.
Methods 2006, 3, 923–929.
Budisa, N.; Alefelder, S.; Bae, J. H.; Golbik, R.; Minks, C.; Huber, R.; Moroder, L.
Protein Sci. 2001, 10, 1281–1292.
Chen, S.; Fahmi, N. E.; Wang, L.; Bhattacharya, C.; Benkovic, S. J.; Hecht, S. M. J.
Am. Chem. Soc. 2013, 135, 12924–12927.
respectively. Both compounds showed high levels of fluorescence
and the relative fluorescence intensities of the aromatic amino
acids were measured at various excitation wavelengths (see ESI,
Figs. S12 and S13) to determine the maximum emission wave-
length (kem) and maximum excitation wavelength (kex). The
acetamidobiphenyl amino acid 1 showed a maximum kem at
Goldberg, J. M.; Speight, L. C.; Fegley, M. W.; Peterson, E. J. J. Am. Chem. Soc.
2012, 134, 6088–6091.
3
84 nm with a maximum kex at 295 nm (Fig. 1). This result was
4
2
6. Hamase, K.; Nagayasu, R.; Morikawa, A.; Konno, R.; Zaitsu, K. J. Chromatogr. A
006, 1106, 159–164.
comparable with the fluorescent properties of 4-aminobiphenyl.
2
The aromatic amino acid 2 displayed a maximum kem at 543 nm
with a maximum kex at 450 nm (Fig. 2). The emission for 2
7
8
9
.
.
.
Charbon, G.; Wang, J.; Brustad, E.; Schultz, P. G.; Horwich, A. L.; Jacobs-Wagner,
C.; Chapman, E. Bioorg. Med. Chem. Lett. 2011, 21, 6067–6070.
Kitamatsu, M.; Yamamoto, T.; Futami, M.; Sisido, M. Bioorg. Med. Chem. Lett.
2010, 20, 5976–5978.
Loving, G.; Imperiali, B. J. Am. Chem. Soc. 2008, 130, 13630–13638.
N
10. Pantoja, R.; Rodriguez, E. A.; Dibas, M.; Doughtrty, D. A.; Lester, H. A. Biophys. J.
009, 96, 226–237.
1. Liu, W.; Brock, A.; Chen, S.; Chen, S.; Schultz, P. G. Nat. Methods 2007, 3, 239–
44.
2
O
1
2
O
OH
N
1
1
2. Niu, W.; Guo, J. Mol. BioSyst. 2013, 9, 2961–2970.
3. Wang, F.; Robbins, S.; Guo, J.; Shen, W.; Schultz, P. G. PloS One 2010, 5, e9354.
O
O
O
O
NH2
HO
N
H
N
H
14. Chatterjee, A.; Guo, J.; Lee, H. S.; Schultz, P. G. J. Am. Chem. Soc. 2013, 135,
12540–12543.
15. Krueger, A.; Imperiali, B. ChemBioChem 2013, 14, 788–799.
HN
N
H
1
1
6. Xu, X.; Hu, X.; Wang, J. Beilstein J. Org. Chem. 2013, 9, 254–259.
7. Koopmans, T.; van Haren, M.; van Ufford, L. Q.; Beekman, J. M.; Martin, N. I.
Bioorg. Med. Chem. 2013, 21, 553–559.
12
O
NH
2
Figure 3. Chemical structure of the peptide-like compound 12 containing
fluorescent (R)-2-amino-4-DMNA-butanoic acid unit.
a
18. Kele, P.; Sui, G.; Huo, Q.; Leblanc, R. Tetrahedron: Asymmetry 2000, 11, 4959–
4963.

J. Maity et al. / Tetrahedron Letters 56 (2015) 4780–4783
4783
0 0 0
40. Procedure for the synthesis of (R)-2-amino-4-(4 -N,N-dimethylamino-1 ,8 -
2
1
2
2
9. Wang, J.; Xie, J.; Schultz, P. G. J. Am. Chem. Soc. 2006, 128, 8738–8739.
0. Dugau, I.; Mazarguil, H. Tetrahedron Lett. 2000, 41, 6063–6066.
1. Esteves, C. I. C.; Silva, A. M. F.; Raposo, M. M.; Costa, S. P. G. Tetrahedron 2009,
naphthalimido)-N -(tert-butoxycarbonyl)butanoic acid (10): Compound
8
(0.266 g, 1.1 mmol) was kept under a nitrogen atmosphere in a three neck
round bottom flask fitted with water condenser. Dioxane (25 mL) was added
through a septum whereupon the mixture was heated at reflux and stirred
6
5, 9373–9377.
2. Costa, S. P. G.; Batista, R. M. F.; Raposo, M. M. M. Tetrahedron 2008, 64, 9733–
737.
3. Costa, S. P. G.; Oliveira, E.; Lodeiro, C.; Roposo, M. M. M. Tetrahedron Lett. 2008,
9, 5258–5261.
2
2
9
vigorously. Boc-
D-Dab-OH (9, 0.218 g, 1.0 mmol) was dissolved in an aqueous
solution (5 mL) containing NaHCO
3
(0.42 g, 5 mmol) and added into the
4
reaction mixture. The reaction mixture was heated at reflux for 1 h. TLC
showed complete consumption of the amino acid, the mixture was
concentrated under reduced pressure and water (60 mL) was added. The
2
2
4. Taki, M.; Sisido, M. US 2010/0152417 A1.
5. Koczan, G.; Csik, G.; Csampai, A.; Balog, E.; Bosze, S.; Sohar, P.; Hudecz, F.
Tetrahedron 2001, 57, 4589–4598.
water phase was washed with Et
pH 3 in an ice-bath whereupon a yellow precipitate formed. The aqueous phase
2
O (2 ꢀ 60 mL) and acidified with 2 N HCl till
2
6. Holder, J. R.; Bauzo, R. M.; Xiang, Z. M.; Haskell-Luevano, C. J. Med. Chem. 2002,
4
5, 3073–3081.
was extracted with DCM (3 ꢀ 80 mL) and the combined organic layers were
2
2
2
7. Rink, H. Tetrahedron Lett. 1987, 28, 3787–3790.
2 4
dried over Na SO , filtered and concentrated under reduced pressure. The
8. https://www.anaspec.com/html/peptide_notes.html.
9. Ting, R.; Adam, M. J.; Ruth, T. J.; Perrin, D. J. Am. Chem. Soc. 2005, 127, 13094–
crude product was purified by column chromatography (40–50% ethyl acetate
in hexane containing 0.1% AcOH) to afford compound 10 (0.27 g, 66%). R = 0.2
ꢁ40.6 (c 0.032, MeOH); H NMR
): d = 8.49 (d, J = 7.2 Hz, 1H), 8.39 (d, J = 7.6 Hz, 1H), 8.36 (d,
J = 8.4 Hz, 1H), 7.58 (t, J = 8.0 Hz, 1H), 7.02 (d, J = 8.4 Hz, 1H), 5.52 (d, J = 7.6 Hz,
1H), 4.27–4.16 (m, 3H), 3.04 (s, 6H), 2.21–2.11 (m, 2H), 1.35 (s, 9H) ppm. 13
NMR (100.6 MHz, CDCl ): d = 174.6, 165.1, 164.6, 157.5, 156.0, 133.4, 131.9,
f
2
2
1
1
3095.
(50% ethyl acetate–hexane, 1% AcOH), [
(400 MHz, CDCl
a]
D
3
0. Maegawa, T.; Kitamura, Y.; Sako, S.; Udzu, T.; Sakurai, A.; Tanaka, A.;
Kobayashi, Y.; Endo, K.; Bora, U.; Kurita, T.; Kozaki, A.; Monguchi, Y.; Sajiki,
H. Chem. Eur. J. 2007, 13, 5937–5943.
1. Simeone, J. P.; Sowa, J. R., Jr. Tetrahedron 2007, 63, 12646–12654.
2. Guo, F.; Zhou, R.; Jiang, Z.; Wang, W.; Fu, H.; Zheng, X.; Chen, H.; Li, R. Catal.
Commun. 2015, 66, 87–90.
3. Guan, J. T.; Song, X. M.; Zhang, Z. Y.; Wei; Ben, M. W.; Dai, Z. Q. Appl. Organomet.
Chem. 2015, 29, 87–89.
4. Bora, U.; Mondal, M. Green Chem. 2012, 14, 1873–1876.
5. Silva, N. O.; Abreu, A. S.; Ferreira, P. M. T.; Monteiro, L. S.; Queiroz, M. R. P. Eur. J.
Org. Chem. 2002, 2524–2528.
3
C
3
3
3
131.7, 130.5, 125.2, 125.0, 122.7, 114.2, 113.4, 80.4, 52.0, 44.9, 36.8, 30.8,
ꢁ
28.5 ppm. HRMS (ESI-TOF) (m/z): calcd for C23
found 440.1827.
H
26
N
3
O
6
[MꢁH] 440.1827,
3
0
0
0
41. Procedure for the synthesis of (R)-2-amino-4-(4 -N,N-dimethylamino-1 ,8 -
2
3
3
naphthalimido)-N -(9-fluorenylmethoxycarbonyl)butanoic
acid
(11):
Compound 10 (0.12 g, 0.27 mmol) was dissolved in DCM (3 mL). Cold
trifluoroacetic acid (3 mL) was added over 5 min and the reaction mixture
was stirred for 2 h. The reaction mixture was concentrated to dryness under
reduced pressure and excess TFA removed by coevaporation with chloroform
(3 ꢀ 10 mL). The crude product was dried under vacuum overnight then
3
3
6. Ito, S.; Ueda, M.; Sekiguchi, R.; Kawakami, J. Tetrahedron 2013, 69, 4259–4269.
7. Christakakou, M.; Schön, M.; Schnürch, M.; Mihovilovic, M. D. Synlett 2013,
2
411–2418.
3
8. Procedure for synthesis of (R)-2-amino-(9-fluorenylmethoxycarbonyl)amino-
3
dissolved in water (4 mL) containing NaHCO (0.114 g, 1.35 mmol). Dioxane
0
0
3
-(4 -acetamido-[1,1 -biphenyl]-4-yl)propanoic acid (6): Compound
64 mg, 0.25 mmol) was dissolved in mixture of tetrahydrofuran and
ethylene glycol (10:1, 15 mL) at rt. N-(9-Fluorenylmethoxycarbonyl)-4-
bromo- -phenylalanine (5, 96 mg, 0.21 mmol) was added and the mixture
was kept stirring. PdCl (10 mol %, 4 mg) was added and nitrogen was bubbled
through the reaction mixture for 15 min. Na CO (66 g, 0.62 mmol) was then
4
(10 mL) was added, followed by Fmoc-OSu (0.101 g, 0.3 mmol). The reaction
mixture was stirred at 0 °C for 2 h. When the TLC showed complete
consumption of the starting material the reaction mixture was diluted with
(
a
D
water (20 mL). The aqueous layer was washed with Et
acidified with 6 N HCl to adjust the pH to 6. The product was extracted with
4
SO ,
2
O (2 ꢀ 20 mL) and
2
2
3
DCM (3 ꢀ 25 mL) and the combined organic layers were dried over Na
2
added into the reaction mixture whereupon the flask was filled with nitrogen
and heated at 66 °C for 3 h. When the TLC showed completion of the reaction
water (20 mL) was added into the reaction mixture which was then extracted
with ethyl acetate (3 ꢀ 30 mL). The combined organic layers were washed with
filtered and concentrated under reduced pressure. The crude compound was
purified by column chromatography (0.5–1% MeOH in DCM containing 0.1%
AcOH) to afford compound 11 (0.116 g, 71%). R
f
= 0.6 (2.5% MeOH–DCM, 1%
): d = 8.51 (d,
22
1
AcOH), [
a
]
D
ꢁ8.0 (c 0.05, MeOH); H NMR (400 MHz, CDCl
3
brine-water (1:1, 2 ꢀ 25 mL), dried over Na
2
SO
4
and concentrated to dryness
J = 6.8 Hz, 1H), 8.41 (d, J = 8.4 Hz, 1H), 8.35 (d, J = 8.4 Hz, 1H), 7.67 (d, J = 7.2 Hz,
2H), 7.59–7.52 (m, 3H), 7.31 (t, J = 7.2 Hz, 2H), 7.23 (t, J = 7.2 Hz, 2H), 7.00 (d,
J = 8.0 Hz, 1H), 5.96 (d, J = 8.0 Hz, 1H), 4.35–4.31 (m, 1H), 4.29–4.19 (m, 4H),
under reduced pressure. The crude product was purified by column
chromatography (2–3% MeOH in DCM containing 0.1% AcOH) to afford
2
7
compound 6 (87 mg, 81%). R
f
= 0.25 (5% MeOH–DCM, 1% AcOH), [
O), H NMR (400 MHz, CD OD): d = 7.67 (d, J = 6.8 Hz, 2H), 7.52–7.44
m, 6H), 7.37 (d, J = 7.6 Hz, 3H), 7.28–7.12 (m, 5H), 4.36 (dd, J = 7.6, 4.8 Hz, 1H),
a
]
D
ꢁ10.0
4.13 (t, J = 7.2 Hz, 1H), 3.03 (s, 6H), 2.34–2.25 (m, 1H), 2.19–2.11 (m, 1H) ppm.
1
13
(
c 0.05, H
2
3
3
C NMR (100.6 MHz, CDCl ): d = 174.0, 165.2, 164.8, 157.7, 156.2, 144.0, 141.4,
(
133.6, 132.1, 131.8, 130.6, 127.8, 127.3, 125.4, 125.1, 125.0, 122.6, 120.1, 113.9,
4
4
.23–4.19 (m, 1H), 4.12–4.08 (m, 1H), 4.05–4.01 (m, 1H), 3.16 (dd, J = 14.0,
113.3, 67.4, 52.2, 47.2, 44.9, 36.8, 31.1 ppm. HRMS (ESI-TOF) (m/z): calcd for
13
ꢁ
.8 Hz, 1H), 2.82 (dd, J = 14.0, 9.6 Hz, 1H), 2.04 (s, 3H) ppm.
C
NMR
C
33
H
28
N
3
O
6
[MꢁH] 562.1984, found 562.1984.
(
100.6 MHz, CD
3
OD): d = 171.7, 158.4, 145.3, 142.6, 140.3, 139.2, 137.9,
42. Bridges, J. W.; Creaven, P. J.; Williams, R. T. Biochem. J. 1965, 96, 872–878.
43. Grabtchev, I.; Meallier, T.; Kontantinova, T.; Popova, M. Dyes Pigments 1995, 28,
41–46.
1
4
5
37.7, 130.9, 128.8, 128.2, 128.0, 127.7, 126.4, 126.3, 121.5, 120.9, 68.0, 56.9,
8.4, 38.4, 23.9 ppm. HRMS (ESI-TOF) (m/z): calcd for C32
19.1925, found 519.1924.
H
27
2
N O
5
[MꢁH]ꢁ
44. Grabtchev, I.; Philipova, Tz.; Meallier, P.; Guittonneau, S. Dyes Pigments 1996,
31, 31–34.
3
9. Koll a´ r, J.; Hrdlovic, P.; Chmela, S.; Sarakha, M.; Guyot, G. J. Photochem. Photobiol.
A 2005, 170, 151–159.
45. Maity, J.; Honcharenko, D.; Strömberg, R. PLoS One 2015, 10, e0124046.