Preparation of new monomers aza-β3-aminoacids for solid-phase syntheses of aza-β3-peptides

Article abstract of DOI:10.1016/j.tetlet.2007.06.082

The preparation of new N^β-Fmoc-protected aza-β³-amino acids (aza-β³-aa) with proteinogenic side chains as well as their N^β-Fmoc, N^β-Cbz or N^β-Boc aza-β³-amino esters (from Pro, Asn, Asp, Glu, Gln) by successive nucleophilic substitutions will be described.

Full text of DOI:10.1016/j.tetlet.2007.06.082

Tetrahedron Letters 48 (2007) 5767–5770
Preparation of new monomers aza-b3-aminoacids for
solid-phase syntheses of aza-b3-peptides
*
`
Olivier Busnel and Michele Baudy-Floc’h
Groupe ‘Ciblage et Auto-Assemblages Fonctionnels’, UMR CNRS 6226, Institut de Chimie, Universite´ de Rennes I, 263 Av. du
Ge´ne´ral Leclerc, F-35042 Rennes Cedex, France
Received 4 May 2007; revised 3 June 2007; accepted 18 June 2007
Available online 22 June 2007
Abstract—The preparation of new Nb-Fmoc-protected aza-b3-amino acids (aza-b3-aa) with proteinogenic side chains as well as their
Nb-Fmoc, Nb-Cbz or Nb-Boc aza-b3-amino esters (from Pro, Asn, Asp, Glu, Gln) by successive nucleophilic substitutions will be
described.
Ó 2007 Elsevier Ltd. All rights reserved.
In recent years, there has been considerable interest in
the design and synthesis of non-natural oligomers that
form secondary structures and present enhanced meta-
bolic stability, bioavailability and biological absorp-
tion.1–7 In this class of peptidomimetics, peptides
consisting exclusively or including aza-b3-amino acid
have emerged as a promising new class of compounds
that form N–N turns and present potentially useful bio-
logical properties.8,9
We have previously reported two methods to prepare
aza-b3-amino acids, the first one consisting in a nucleo-
philic substitution of benzyl or t-Bu bromoacetate by
Na-substituted-Nb-protected hydrazines 1 (Scheme 1).11
Then, the required monomers 3 were obtained by depro-
tection of the carboxy protecting group of esters 2 in sat-
isfactory to good yields. Nevertheless, nucleophilic
substitution of bromo acetate by N-substituted Fmoc
hydrazine proceeds in low yield (36–50%). Therefore,
an alternative approach, in one step, to obtain aza-b3-
amino acid has been described, which relies on reductive
amination of glyoxylic acid and Nb-Fmoc protected-Na-
substituted hydrazine 1 (Scheme 2).12
For our ongoing projects on the synthesis of aza-b3-pep-
tides, we need to have ample access to the aza-b3-amino
acid building blocks with proteinogenic side chains and
Nb-Fmoc protection. With the exceptions of aza-b3-Lys,
aza-b3-Tyr, aza-b3-Arg only Fmoc-protected aza-b3-
amino acids with nonfunctionalized and achiral side
chains have been published.10 We described here, in de-
tail, the preparation of five new aza-b3-amino acid deriv-
atives, with the side chains of aspartic, glutamic, proline,
asparagine and glutamine.
N-Substituted Fmoc hydrazines 1 were prepared accord-
ing to literature procedures by reduction of Fmoc
hydrazones, derived from the reaction of Fmoc carbaz-
ate with either aldehyde or ketone.13,14 To introduce side
chains such as aspartic, glutamic, proline and aspara-
gine, the corresponding aldehydes were not available
R
O
R
O
2
2
BrCH CO PG
PG =Bn, H /Pd
1
2
2
1
2
PG
PG
N
N
1
2
PG NHNHR
N
H
OH
N
OPG
2
PG =
t
-Bu, CF CO H
H
3
2
K CO
2
3
3
2
1
Scheme 1. Synthesis of Nb-protected-aza-b3-amino acids.
Keywords: Aza-b3-amino acid; Fmoc protecting group; Peptidomimetics/nucleophilic substitution.
*
Corresponding author. Fax: +33 223236933; e-mail: Michele.Baudy-Floch@univ-rennes1.fr
0040-4039/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2007.06.082

5768
O. Busnel, M. Baudy-Floc’h / Tetrahedron Letters 48 (2007) 5767–5770
Fmoc, PG2 = Me) or glutamic methyl ester 2 (n = 2,
R
N
O
OHCCO H
R = (CH2)2CO2t-Bu, PG1 = Fmoc, PG2 = Me). Then
saponification of the corresponding C-terminal methyl
ester, catalyzed by CaCl2 to suppress Fmoc cleavage
under basic conditions,18,19 leads to the corresponding
N-Fmoc protected aza-b3-aspartic 3 (n = 1, R = (CH2)2-
CO2t-Bu) in an overall yield of 40% or glutamic acid 3
(n = 2, R = (CH2)2CO2t-Bu) in an overall yield of 20%
(Scheme 4).
2
Fmoc
FmocNHNHR
N
OH
H
NaBH CN,MeOH
3
1
HCl (2N)
3
Scheme 2. Synthesis of Fmoc-aza-b3-amino acid.
1) RCHO
X
Asparagine or glutamine side chains could be intro-
duced by aminolysis of ester group. First of all, Boc-
aza-b3-Gly-OMe 8 (n = 1) was made in 70% yield by cat-
alytic hydrogenation of the condensation product from
glyoxylic methyl ester obtained in situ by oxidation of
dimethyl L-tartrate. Analogue 8 (n = 2) was obtained
by nucleophilic substitution of methyl bromopropionate
in 40% yield. Introduction of asparagine and glutamine
chains was achieved by aminolysis of the ester group of
8.20 Then nucleophilic substitution of benzyl bromoace-
tate by hydrazine 9 leads, respectively, to Boc-aza-b3-
2) NaBH CN,MeOH
3
FmocNHNH
FmocNHNH(CH ) CO t-Bu
2 n 2
2
1 R=(CH ) CO t-Bu
2
2 n
Br(CH ) CO t-Bu
2 n 2
10-20%
DIPEA
Scheme 3. Synthesis of Nb-substituted Fmoc hydrazines.
or did not lead to hydrazones (Scheme 3). In the litera-
ture the introduction of aspartic acid was achieved by
reduction of the condensation product from glyoxylic
acid and 9-fluorenylmethyl carbazate. The acylhydraz-
one was then reduced with 5 fold excess of cyanoboro-
hydride and the resulting acid was then protected as
t-butyl ester using t-butyl trichloroacetimidate in an
overall yield of 20%.15 Nucleophilic substitution of the
corresponding bromo acetate or propionate with Fmoc
carbazate would be an alternative to introduce side
chains. Unfortunately, this reaction proceeds in some-
what lower yield (20%) than the corresponding Boc or
Cbz carbazates (80%).16
Asn-OBn
2
(n = 1, R = CH2CONH2, PG1 = Boc,
PG2 = Bn) and Boc-aza-b3-Gln-OBn 2 (n = 2, R =
CH2CONH2, PG1 = Boc, PG2 = Bn). After deprotec-
tion of hydrazine function with HClg, the Nb-Fmoc
derivatives
2
(R = CH2CONH2,
PG1 = Fmoc,
PG2 = Bn) and 2 (R = (CH2)2CONH2, PG1 = Fmoc,
PG2 = Bn) are preferably obtained by reaction of
Fmoc-Cl and NEt3 (Scheme 5).
The synthesis of peptides containing asparagine or glu-
tamine with unprotected side-chains is known to be
fraught with different types of problems. Reaction of
the activated a-carboxyl group of asparagine or gluta-
mine with b-carboxamide function leads to nitrile and
succinimide derivatives. This is often the cause of incom-
plete or low coupling with the amino acid and of by-
product formation.21 The trityl group seems to be a
good protecting group for carboxamide functions of
asparagine and glutamine as tritylated carboxamide
could have a cleavage rate similar to the usual t-butyl-
type protecting group for peptides. The carboxamide
Therefore, to overcome this problem Cbz-aza-b3-aspar-
tic OMe 2 (n = 1, R = CH2CO2t-Bu, PG1 = Cbz,
PG2 = Me) and Cbz-aza-b3-glutamic OMe 2 (n = 2,
R = (CH2)2CO2t-Bu, PG1 = Cbz, PG2 = Me) were first
prepared by two successive nucleophilic substitutions.16
After hydrogenolysis of 2 in the presence of Pd/C to
remove the benzyl group,17 treatment of 6 with FmocCl
in the presence of NaHCO3 afforded N-Fmoc protected
aza-b3-aspartic 2 (n = 1, R = CH2CO2t-Bu, PG1 =
t-BuO C
2
O
O
BrCH CO Me
Br(CH ) CO t-Bu
2
2
2
2 n
H
n
CbzNHNH
Cbz
N
2
Cbz
N
N
DIPEA
n=1: 90%
DIPEA
n =1 : 80%
N
Ot-Bu
OMe
n
H
H
4
5
2 R=(CH2)nCO2t-Bu, n=1 or 2,
PG1=Cbz, PG2=Me
t-BuO C
2
t-BuO C
O
n
2
H , Pd/C
O
2
FmocCl
NaHCO
n
N
N
FmocHN
OMe
95%
3
H N
2
OMe
n=1 : 65%
2 R=(CH ) CO
PG =Fmoc, PG =Me
t
-Bu, n=1 or 2,
2 n
2
6
1
2
t-BuO C
2
NaOH, CaCl
n =1 : 85%
O
2
n
N
FmocHN
OH
t-Bu, n=1 or 2
3 R=(CH ) CO
2 n
2
Scheme 4. Synthesis of Fmoc-aza-b3-Asp-OH (n = 1) and Fmoc-aza-b3-Glu-OH (n = 2).

O. Busnel, M. Baudy-Floc’h / Tetrahedron Letters 48 (2007) 5767–5770
5769
n=1
(CHOHCO Me)
2
2
O
n
O
n
NH /MeOH
3
H
H
H IO ,70%
5
4
BocNHNH
Boc
N
Boc
O
N
2
n=2
N
OMe
N
NH
2
n=1 : 85%
CO Me
2
H
H
Br
7
9
8
DIPEA, 40%
O
BrCH CO Bn
a)HClg
O
2
2
H N
O
2
H N
n
2
n
Fmoc
N
b)NEt , FmocCl
Boc
N
K CO
2
3
3
N
H
OBn
N
OBn
n =1 : 55%
H
n =1 : 54%
2 R= (CH ) CONH , n=1 or 2,
2 n
2
2 R= (CH ) CONH , n=1 or 2,
2 n
2
1
2
1
2
PG =Fmoc, PG =Bn
PG = Boc, PG =Bn
O
O
Trt-OH
AcOH, Ac O
H , Pd/C
O
O
TrtHN
2
TrtHN
n
n
Fmoc
N
Fmoc
N
2
n=1 : 99%
N
OH
N
H
OBn
H SO
2
4
H
n=1 : 55%
2 R= (CH ) CONHTrt, n=1 or 2,
PG = Fmoc, PG =Bn
Scheme 5. Synthesis of Fmoc-aza-b3-Asn-OH (n = 1) Fmoc-aza-b3-Gln-OH (n = 2).
3 R= (CH ) CONHTrt, n=1 or 2
2 n
2 n
1
2
functions of asparagine and glutamine analogues were
tritylated by an acid-catalyzed reaction with triphenyl-
methanol and acetic anhydride in glacial acetic acid at
with carboxybenzyloxy chloride. Treatment of the
orthogonally protected hydrazine with sodium hydride
in DMF followed by reaction with 1,3-dibromopropane
afforded the protected five-membered cyclic hydrazine
12.22 After deprotection of the Boc group using trifluoro
acetic acid or HClg, nucleophilic substitution of tert-bu-
60 °C. Catalytic hydrogenation of ester
2 (R =
(CH2)2CONHTrt, PG1 = Fmoc, PG2 = Bn) finally gives
Fmoc-aza-b3-Asn(Trt)-OH 3 (n = 1, R = CH2CON-
HTrt) in 30% overall yield and Fmoc-aza-b3-Gln(Trt)-
OH 3 (n = 2, R = (CH2)2CONHTrt) in 10% overall
yield (Scheme 5).
tyl bromoacetate affords Cbz-aza-b3-Pro-OtBu
2
(R = (CH2)3, PG1 = Cbz, PG2 = t-Bu). After reductive
deprotection of the Cbz group using 10% Pd/C as a cat-
alyst in methanol, the Nb-Fmoc derivative 2,
R = (CH2)3, PG1 = Fmoc, PG2 = t-Bu, is obtained by
nucleophilic substitution of Fmoc-Cl by 13. Finally
deprotection of the Boc group using HClg, leads to pro-
line analogue Fmoc-aza-b3-Pro-OH 3, R = (CH2)3,
PG1 = Fmoc, in 35% overall yield (Scheme 6).
Proline is the amino acid with the highest propensity to
occur in turn structures in proteins, because of the
constraints of pyrrolidine ring formation. tert-But-
oxycarbonylhydrazine 7 was converted to 1-tert-butoxy-
carbonyl-2-carboxybenzyloxyhydrazine 11 by treatment
Boc
Cbz
1) NaH
2) Br-(CH ) -Br
N
N
ZCl
2 3
NH
BocHN
2
NHCbz
BocHN
89%
82%
7
10
11
Br-CH -CO
tBu
2
1) HCl
2) NEt
2
g
3
Cbz
Cbz
K CO
2
3
N
CO t-Bu
2
N
N
NH
98%
Toluene
85%
2
12
1
2
R= (CH ) , PG =Cbz, PG =
t
-Bu
2 3
Fmoc
H Pd/C
2
FmocCl
CO tBu
2
N
HN
N
CO
t
2
Bu
N
NaHCO aq
3
99%
79%
2
13
1
2
R= (CH ) , PG =Fmoc, PG =
t-Bu
2 3
Fmoc
HCl
g
CO H
N
N
2
74%
3 R= -(CH ) -
2 3
Scheme 6. Synthesis of Nb-Fmoc-aza-b3-Pro-OH.

5770
O. Busnel, M. Baudy-Floc’h / Tetrahedron Letters 48 (2007) 5767–5770
(CDCl3)d ppm: 169.92 CO2Me, 168.80 CO2tBu, 155.21
NHCO, 136.16 Car, 128.29 128.14 127.97 CHar, 81.61
C(CH3), 66.62 OCH2, 57.71 NCH2, 57.04 NCH2, 51.61
OCH3, 27.87 CH3. HRMS (ESI) m/z calculated for
C17H24N2O6Na [M+Na]+: 375.1532; found, 375.1526
(2 ppm).
In conclusion, we described herein the preparation of
five new Nb-Fmoc-aza-b3-amino acids with four hetero-
atomic side chains by successive nucleophilic substitu-
tions. The solid-phase synthesis of oligomers or hybrid
peptides incorporating these analogues will be published
later.
17. General procedure for catalytic hydrogenolysis of 6 (n = 1).
Compound 6 (3.21 g, 7.5 mmol) was dissolved in methanol
(30 mL) with 10% Pd/C (250 mg) for 12 h. The palladium
was removed by filtration over Celite and the resulting
solution was concentrated under vacuum to afford 7 as a
yellow oil (1.46 g, 95%). 1H NMR (CDCl3) d ppm: 1.49 (s,
9H, CH3), 3.70 (s, 2H, CH2), 3.76 (s, 3H, CH3), 3.82 (s,
2H, CH2), 5.01 (s, 2H, NH2). 13C NMR (CDCl3) d ppm:
170.78 CO2Me, 169.53 CO2tBu, 81.46 C(CH3), 60.00
NCH2, 59.22 NCH2, 51.51 OCH3, 27.94 CH3. HRMS
(ESI) m/z calculated for C9H20N2O4 [M+H]+: 219.1345;
found, 219.1359 (6 ppm).
References and notes
1. Gante, J. Angew. Chem., Int. Ed. Engl. 1994, 33, 1699–
1720.
2. Gellman, S. H. Acc. Chem. Res. 1998, 31, 173–180.
3. Seebach, D.; Schaeffer, L.; Gessier, F.; Bindscha¨dler, P.;
Ja¨ger, C.; Josien, D.; Kopp, S.; Lelais, G.; Mahajan, Y.
R.; Micuch, P.; Sebesta, R.; Schweizer, B. W. Helv. Chim.
Acta 2003, 86, 1852–1861.
18. General procedure for hydrolysis of 2 (2.19 g, 5 mmol) in
methanol (77 mL) was added a solution of NaOH (0.24 g,
1.2 equiv) in aqueous CaCl2 0.8 M (9.77 g in 33 mL of
water). The mixture was stirred at room temperature for
7 h. Methanol was removed by evaporation and the
resulting solution was acidified with HCl 2 N and
extracted twice with methylene chloride (40 mL). The
organic layers were combined, dried over Na2SO4 and
concentrated. The crude oil was purified by chromatog-
raphy on silica gel (ethyl acetate/methylene chloride 1:1
and ether/methanol 1:1) to give 3 as a moss (1.81 g, 85%).
1H NMR (CDCl3) d ppm: 1.51 (s, 9H, CH3), 3.63 (s, 2H,
CH2), 3.71 (s, 2H, CH2), 4.23 (t, 1H, J = 6.3 Hz, CH), 4.56
(d, 2H, J = 6.3 Hz, CH2), 7.29–7.80 (m, 8H, CHar). 13C
NMR (CDCl3) d ppm: 171.80 CO2H, 169.46 CO, 157.19
COFmoc, 143.43, 141.32 Car, 127.89, 127.19, 125.07, 120.08
CHar, 82.99 C(CH3), 67.68 CH2Fmoc, 59.25, 58.60 NCH2,
47.05 CHFmoc, 28.10 CH3. HRMS (ESI) m/z calculated for
C23H25N2O6Na2 [MÀH+2Na]+: 471.1508; found,
471.1509 (0 ppm).
4. Nielsen, P. E.; Egholm, M.; Berg, R. H.; Buchardt, O.
Science 1991, 254, 1497–1500.
5. Kirshenbaum, K.; Zukermann, R. N.; Dill, K. A. Curr.
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633–636.
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Gall, P.; Baudy-Floc’h, M.; Le Grel, P. J. Org. Chem.
2001, 66, 4923–4929.
´
9. Baudy-Floc’h, M.; Muller, S.; Busnel, O. 23 Decembre
2004, CNRS WO 2004/111086.
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S.; Bondon, A.; Muller, S.; Baudy-Floc’h, M. J. Org.
Chem. 2005, 26, 10701.
11. Cheguillaume, A.; Doubli-Bounoua, I.; Baudy-Floc’h, M.;
Le Grel, P. Synlett 2000, 331–334.
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2005, 46, 7073.
19. Pascal, R.; Sola, R. Tetrahedron Lett. 1998, 39, 5031–5034.
20. General procedure for aminolysis 9 (n = 2). The ester 9
(10.5 g, 43 mmol) was dissolved into a solution of meth-
anol with ammoniac 6 N (20 mL). The solution was stirred
at room temperature for 3 days and concentrated under
vacuum. The crude oil was precipitated in ether (10 mL) to
afford 10 as a white powder (8.92 g, 85%): mp = 105 °C.
1H NMR (CDCl3) d ppm: 1.48 (s, 9H, CH3), 2.41 (t, 2H,
J = 5.9 Hz, CH2), 3.14 (t, 2H, J = 5.9 Hz, CH2), 5.60 (br,
1H, NH), 6.24 (s, 1H, NH amide), 7.06 (s, 1H, NH amide).
13C NMR (CDCl3) d ppm: 173.93, 156.07, 79.79, 47.04,
32.83, 27.32. HRMS (ESI) m/z calculated for
C8H17N3O3K [M+K]+: 242.0907; found, 242.0909
(1 ppm).
13. Dutta, A. S.; Morley, J. S. J. Chem. Soc., Perkin Trans. 1
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878.
16. General procedure for nucleophilic substitution of Cbz-aza-
b3-Gly-OtBu 5 (n = 1). To a solution of 5 (n = 1) (5.12 g,
18.3 mmol) in toluene (40 mL), were added DIPEA
(2.39 g, 1 equiv) and methyl-bromoacetate (5.6 g, 2 equiv).
The mixture was stirred at 75 °C for 4 days, cooled at
room temperature and filtrated. The filtrate was concen-
trated under vacuum and the resulting crude oil was
purified by chromatography on silica gel (ethyl acetate/
methylene chloride 7:93) to give 6 as a yellow oil (5.80 g,
90%). 1H NMR (CDCl3) d ppm: 1.47 (s, 9H, CH3), 3.72 (s,
5H, CH2+CH3), 3.82 (s, 2H, CH2), 5.16 (s, 2H, CH2), 7.10
(br, 1H, NH), 7.29–7.40 (m, 5H, CHar). 13C NMR
21. Sieber, P.; Riniker, B. Tetrahedron Lett. 1991, 32(6), 739.
22. Toya, T.; Yamaguchi, K.; Endo, Y. Bioorg. Med. Chem.
2002, 10, 953–962.