Stereo-regulated synthesis of peptides containing a β-trifluoromethyl- β-amino acid

Article abstract of DOI:10.1016/j.tet.2013.08.051

For the chemical conversions of a β-trifluoromethyl-β-amino acid ((S)-4,4,4-trifluoro-3-aminobutyric acid, 1), such as the N-terminus protection with benzyloxycarbonyl or tert-butoxycarbonyl group, the C-terminus protection with benzyl or tert-butyl group, and peptide elongation at the both termini, highly practical protocols were established. Through these conversions, the stereochemistry of 1 and/or its condensation counterpart was maintained. Because the protocols developed here are indispensable for the application of 1 in peptide engineering, they would expand the utility of 1 and its derivatives.

Full text of DOI:10.1016/j.tet.2013.08.051

Accepted Manuscript
Stereo-regulated synthesis of peptides containing a #-trifluoromethyl-#-amino acid
Joon-il Cho, Nao Nishizono, Nobutaka Iwahashi, Kazuhiko Saigo, Yasuhiro Ishida
PII:
S0040-4020(13)01338-0
10.1016/j.tet.2013.08.051
TET 24740
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 31 May 2013
Revised Date: 17 August 2013
Accepted Date: 19 August 2013
Please cite this article as: Cho J-i, Nishizono N, Iwahashi N, Saigo K, Ishida Y, Stereo-regulated
synthesis of peptides containing a #-trifluoromethyl-#-amino acid, Tetrahedron (2013), doi: 10.1016/
j.tet.2013.08.051.
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Stereo-regulated synthesis of peptides
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containing a β-trifluoromethyl-β-amino acid
Joon-il Cho, Nao Nishizono, Nobutaka Iwahashi, Kazuhiko Saigo, and Yasuhiro Ishida
RIKEN Center for Emergent Matter Science, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan
R
F3C
F3C
R
n
n
R' NH
NH
OR''
R' NH
NH
OR''
O
O
O
O
n = 0 or 1
n = 0 or 1
F3C
F3C
Cbz NH
OH
H2N
OBn
F3C
H2N
O
O
OH
F3C
F3C
H2N
O
Boc NH
OH
OtBu
O
O

ACCEPTED MANUSCRIPT
1
Tetrahedron
journal homepage: www.elsevier.com
Stereo-regulated synthesis of peptides containing a β-trifluoromethyl-β-amino acid
Joon-il Cho a, Nao Nishizono b, Nobutaka Iwahashi b, Kazuhiko Saigo c, Yasuhiro Ishida a
a RIKEN Center for Emergent Matter Science, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan
b Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-8656, Japan
c School of Environmental Science and Engineering, Kochi University of Technology, Miyanokuchi, Tosayamada-cho, Kami, Kochi 782-8502, Japan
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
For the chemical conversions of a β-trifluoromethyl-β-amino acid ((S)-4,4,4-trifluoro-3-
aminobutyric acid, 1), such as the N-terminus protection with benzyloxycarbonyl or tert-
butoxycarbonyl group, the C-terminus protection with benzyl or tert-butyl group, and peptide
elongation at the both termini, highly practical protocols were established. Through these
conversions, the stereochemistry of 1 and/or its condensation counterpart was maintained.
Because the protocols developed here are indispensable for the application of 1 in peptide
engineering, they would expand the utility of 1 and its derivatives.
Available online
Keywords:
amino acid
fluorinated amino acid
peptide synthesis
2009 Elsevier Ltd. All rights reserved.
1. Introduction
derived from other fluorinated amino acids, such as γ/δ-
fluorinated α-amino acids. To our wonder, however, studies on
The incorporation of fluorinated amino acids into peptides has
emerged as a new powerful tool for peptide engineering in the
the application of β-perfluoroalkyl-β-amino acids to peptide
engineering have been surprisingly rare. As far as we know,
even the synthesis of peptides containing β-perfluoroalkyl-β-
amino acids has been limited to a few examples.5 Furthermore,
only racemates of these fluorinated amino acids were used in
these early examples.
last decade.1
Owing to the peculiar characteristics of
perfluoroalkyl groups, including electron-withdrawing nature,
large dipole moment, hyper-hydrophobicity, and lipophobicity,2
the partial fluorination of peptides often gives rise to significant
alterations in the structures, functions, and properties of the
resultant peptides.1a,d In addition, 19F nucleus is highly NMR
sensitive, occurs in 100% natural abundance, and yet rarely exists
in biological systems, which allows us to monitor the
surrounding environment of the fluorinated positions by NMR.3
In particular, the incorporation of γ/δ-fluorinated α-amino acids
into peptides has been extensively explored.1
As another class of fluorinated amino acids, β-perfluoroalkyl-
β-amino acids, such as (S)-4,4,4-trifluoro-3-aminobutyric acid
(1), have attracted considerable recent attention.4 They are
expected to possess favorable properties as components of
fluorinated peptides, such as chemical stability, tolerance toward
racemization, and proper reactivity of the amino/carboxyl groups.
Moreover, the incorporation of β-perfluoroalkyl-β-amino acids
into peptides would bring significant influence on the structure
and properties of the peptides, in which the perfluoroalkyl groups
are directly connected to their back bones, unlike peptides
For a systematic study on the peptide synthesis using β-
perfluoroalkyl-β-amino acids, one of serious obstacles is their
hard accessibility, particularly in an enantiopure form. Although
several groups have reported the asymmetric synthesis of β-
perfluoroalkyl-β-amino acids such as 1, most of them are not
capable of producing enantiopure material in a practical scale.6–9
Contrary to them, we have recently reported a highly practical
method to obtain 1/its antipode in an enantiopure form, in which
the diastereoselective hydride reduction of a cyclic enamino-ester
is the key step.10 Owing to the advantages of this method,
including easy access to starting materials, scalability,
operational convenience, and high stereoselectivity, a multi-gram
quantity of enantiopure 1/its antipode can be easily produced,
which would allow us to explore a new mode of peptide
engineering based on 1. Here we report the first systematic study
on peptide formation using 1, especially focusing on the retention
of enantiomeric and diastereomeric purities through the
———
Corresponding author. Tel.: +81-48-462-1111 (ext. 6351); fax: +81-48-467-8214; e-mail: y-ishida@riken.jp (Y. Ishida)

ACCEPTED MANUSCRIPT
2
Tetrahedron
Route A (i)
Route A (i)
CbzCl (1.0 eq)
K2CO3 (10 eq)
H2O/1,4-dioxane
rt, 12 h
Boc2O (1.0 eq)
K2CO3 (10 eq)
H2O/1,4-dioxane
rt, 1 day
F3C
H2N
F3C
Cbz NH
F3C
F3C
H2N
OH
OH
Boc NH
OH
OH
56%
55%
O
O
O
O
1
4
1
6
Route B (i)
Route B (i)
Route B (iii)
Route B (iii)
SOCl2 (excess)
EtOH
rt, overnight
SOCl2 (excess)
EtOH
rt, overnight
LiOH (3.0 eq)
H2O/1,4-dioxane
rt, 1 h
LiOH (3.0 eq)
H2O/1,4-dioxane
rt, 1 h
quant.
OEt
91%
quant.
OEt
quant.
F3C
F3C
Cbz NH
F3C
F3C
Boc NH
96%
82%
HCl• H2N
OEt
HCl• H2N
OEt
Route B (ii)
O
2
O
O
Route B (ii)
Boc2O (2.0eq)
pyridine (1.2 eq)
DMAP (0.2 eq)
1,4-dioxane
rt, 1 day
O
CbzCl (3.5 eq)
pyridine (8 eq)
CH2Cl2
3
2
5
rt, 1 day
Route A: 56% (1 step) Route B: 87% (3 steps)
Route A: 55% (1 step) Route B: 82% (3 steps)
Scheme 1. N-Cbz protection of (S)-4,4,4-trifluoro-3-aminobutyric
acid (1).
Scheme 2. N-Boc protection of (S)-4,4,4-trifluoro-3-aminobutyric
acid (1).
protection/deprotection/coupling processes, as well as the
peculiar reactivity of the amino group in 1.
According to the reported procedure,7d 1 was quantitatively
converted into 2 by treatment with ethanol/thionyl chloride
(Scheme 1, Route B (i)). The following N-protection was carried
out under the conditions similar to those Soloshonok et al. have
reported, except for using CbzCl in place of phenylacetyl
chloride.9b Against to our expectation, however, the formation of
the target N-protected amino ester 3 was sluggish (13%), and
several kinds of unidentified byproducts were afforded.
Therefore, we surveyed reaction conditions and fortunately found
that the yield of 3 could be highly improved by using pyridine in
place of triethylamine (Scheme 1, Route B (ii), 96%). The ester
part of 3 was readily hydrolyzed by treatment with an aqueous
alkaline solution (Scheme 1, Route B (iii), 91%). Thus, the
target N-protected amino acid 4 was obtained in 87% overall
yield from 1 through three steps.
As described above, the choice of the base in the second step
of Route B brought an unexpected effect on the yield of 3. These
observations can be elucidated as follows, taking into account of
peculiar properties of the amino group in 2. Firstly, the basicity
of the amino group in 2 is comparable to or lower than that of
pyridine, so that pyridine was able to work as a proton captor in
this reaction system, contrary to the condensation of normal
aliphatic amines, where relatively strong base such as tertiary
amines are usually necessary to realize satisfactory yield.
Secondly, for the acylation of this amino group, specially
activated acylating reagents, such as ketenes and acyl
pyridiniums, are necessary to compensate the low nucleophilicity
of the amino group. In the acylation with acyl chlorides having
no hydrogen at the α-position of their carbonyl group, i.e. with
those incapable of forming ketenes, an N-heteroaromatic system
2. Results and discussion
2.1. N-Protection of 1.
As an indispensable step to incorporate 1 into peptides, we
started our work with the protection of the amino group. Taking
account of easiness in introduction and deprotection processes,
benzyloxycarbonyl (Cbz) and tert-butoxycarbonyl (Boc) groups
were chosen as the N-protecting groups.
The N-Cbz protection of 1 was conducted according to a
procedure generally used for various amino acids. For example,
1 was dissolved in an aqueous alkaline solution and then treated
with benzyl chloroformate (CbzCl) at rt for 12 h with extensive
stirring (Scheme 1, Route A). Although this protocol has been
well established and widely used for other amino acids, the target
N-protected amino acid 4 was obtained only in an unsatisfactory
yield (56%), where a considerable amount of 1 unreacted
remained. Even the optimization of reaction conditions (amount
of CbzCl, pH, etc.) could not improve the yield (data not shown).
In the Schotten-Baumann type reactions, i.e. the condensation
of amines with acyl chlorides in aqueous alkaline media, the
amines usually work as predominant nucleophiles to react with
acyl chlorides even in the presence of an excess amount of OH–,
owing to the difference in nucleophilicity. Unlike usual amines,
the amino group in
1 was considered to exhibit poor
nucleophilicity, comparable to or lower than that of OH–, so that
the hydrolysis of CbzCl was likely to proceed mainly. With this
consideration in mind, we next examined the N-protection in a
non-aqueous solution, which involved the following three steps
(Scheme 1, Route B): (i) the derivatization of 1 into the
corresponding amino ester hydrochloric acid salt 2 to enhance the
solubility in an aprotic medium, (ii) the N-protection of the
amino ester 2 under non-aqueous conditions, and (iii) the
hydrolysis of the ester moiety of the resultant N-Cbz amino ester
3. This synthetic route seemed to be so promising, because
Soloshonok and co-workers have reported an analogous reaction;
the condensation of 2 with 3,5-dinitrobenzoyl chloride proceeded
efficiently in dichloromethane by using triethylamine as a base.7d
with
certain
basicity,
such
as
pyridine,
4-
(dimethylamino)pyridine, or 1-methylimidazole, would be
suitable as a base to efficiently generate activated species. On
the other hand, in the reactions of acyl chlorides with a ketene-
forming ability, such as the previous report by Soloshonok et
al.,9b a relatively strong base can enhance the acylation.
We also performed the N-Boc protection of 1 by the
condensation with di-tert-butyl dicarbonate (Boc2O), wherein
two synthetic strategies (Scheme 2, Route A and Route B) were
again examined. In good agreement with the case of the N-Cbz

ACCEPTED MANUSCRIPT
3
F3C
Boc NH
F3C
K2CO3 (1.0 eq)
BnBr (1.1 eq)
F3C
F3C
Cbz NH
isobutene
H2SO4 (cat.)
Cbz NH
OH
OtBu
OtBu
OH
Boc NH
OBn
OBn
DMF, rt, 1 day
62%
CH2Cl2, rt, 4 days
87%
O
O
O
O
O
6
9
4
7
F3C
H2N
F3C
H2N
H2 (1 atm)
Pd/C (cat.)
CF3CO2H (excess)
CH2Cl2, rt, 3 h
quant.
CF CO H•
3
2
MeOH/CH2Cl2, rt, 12 h
quant.
O
10
8
Scheme 3. O-tBu protection of (S)-4,4,4-trifluoro-3-aminobutyric
acid (1).
Scheme 4. O-Bn protection of (S)-4,4,4-trifluoro-3-aminobutyric
acid (1).
t
protection, the Schotten-Baumann type reaction was found to be
unsuitable for the N-Boc protection of 1 (Scheme 2, Route A,
55%). Meanwhile, in the alternative route (Scheme 2, Route B),
there was some room for optimization. In the second step (N-
acylation), the protocol widely used for O-Boc protection was
employed, taking account of the moderate nucleophilicity of the
amino group in 1 comparable to the hydroxyl group in usual
aliphatic alcohols. For example, the amino ester 2 was treated at
rt with Boc2O (2.0 eq) and pyridine (1.2 eq) in the presence of a
catalytic amount of 4-(dimethylamino)pyridine (DMAP; 0.2 eq)
as an activator of Boc2O. Although the yield of target N-Boc
amino ester 5 was again unsatisfactory when the reaction was
conducted in dichloromethane (60%, Table S1, entry 1), reaction
efficiency was notably enhanced when 1,4-dioxane was used in
place of dichloromethane (82%, Table S1, entry 2). Since the
increment of reagents brought little effect on the yield of 5 (84%,
Table S1, entry 3), we chose the conditions of entry 2 as optimal
ones (Scheme 2, Route B (ii)). The hydrolysis of 5 proceeded
smoothly to afford the target N-Boc protected amino acid 6 in
quantitative yield (Scheme 2, Rout B (iii)). Overall, the N-Boc
protection of 1 was achieved in 82% yield through three steps.
In the transformations shown in Schemes 1 and 2, particularly
in the steps under basic/alkaline conditions, the racemization of 1
and its derivatives were anticipated. Fortunately, however, any
detectable racemization did not take place in these
transformations, which was confirmed by 19F NMR spectroscopy
by the aid of chiral NMR shift reagents (Figures S1 and S2).
the target Bu ester 7 was obtained in good yield (87%). In
addition, the N-protecting group of 7 was smoothly cleaved by
t
hydrogenolysis to afford the target amino acid Bu ester 8 in
quantitative yield.
The O-Bn protection of the carboxyl group in 1 was also
achieved by employing a general procedure (Scheme 4). The N-
Boc amino acid 6 was treated with benzyl bromide and K2CO3 in
N,N-dimethylformamide to afford the Bn ester 9 in acceptable
yield (62%). The cleavage of the N-protecting group was
efficiently promoted by trifluoroacetic acid (TFA) to give TFA
salt of the target amino acid Bn ester (10) in quantitative yield.
As in the case of the N-protection, any detectable racemization
of 1 and its derivatives again did not take place through the
transformations in Schemes 3 and 4, which was confirmed by 19F
NMR spectroscopy by the aid of chiral NMR shift reagents
(Figures S3 and S4).
2.3. Incorporation of 1 into peptides: elongation at the N-
terminus.
With these N- and O-protected derivatives of 1 in hand, we
next attempted the synthesis of peptides incorporating 1.
Particularly, in the peptide elongation at the N-terminus of 1, the
low nucleophilicity of the amino group in 1 was an anticipated
obstacle, as in the case of the N-protection of 1. In addition, the
activation method of the acid component should be carefully
chosen, in order to avoid the racemization of the stereogenic
centers in the components. Moreover, reaction conditions
applicable to solid-state synthesis are more desirable from
practical viewpoints.
2.2. O-Protection of 1.
Firstly, we examined the peptide-bond formation at the N-
terminus of 1. As an acid component, N-benzyloxycarbonyl-L-β-
homoalanine (11) was chosen, because (i) β-amino acids such as
11 can be obtained by the simple homologation of natural amino
acids, of which chemistry has been well explored by Seebach et
al.,11 and (ii) the α-carbon of the carboxyl group in 11 is
unsubstituted and is not stereogenic, which is favorable from the
viewpoints of the easiness in the activation of the carboxyl group
It has already been reported that 1 can be readily converted
into amino esters by using thionyl chloride and the corresponding
alcohols.7d
Despite its simplicity and reliability, this
esterification method requires an excess amount of alcohols and
is applicable only to the reaction of primary or secondary
alcohols with sufficient volatility and availability in a large scale.
Among esters inaccessible with this method, tert-butyl (tBu) and
benzyl (Bn) esters of 1 are of special synthetic utility, in terms of
orthogonality to other protecting groups in deprotection
processes. Therefore, we attempted the synthesis of these two
esters, which would expand the scope of the peptide engineering
of 1.
and
avoidance
of
complexity
arising
from
racemization/epimerization.
The condensation of 2 with 11 is summarized in Table 1. As
might be anticipated from the low nucleophilicity of the amino
group in 2, usual peptide-forming methods were found to be
unsuitable for the present condensation; the usage of 3-(3-
In general, the O-tBu protection of amino acids is conducted
by a three-stage synthesis: the N-Cbz protection, the O-tBu
protection, and the cleavage of the N-protecting group. We
dimethylaminopropyl)-1-ethylcarbodiimide
hydrochloride
t
(EDC•HCl) and 1-hydroxybenzotriazole (HOBt), as condensing
and activating reagents, respectively, afforded the target
dipeptide 11/2 in unsatisfactory yield (Table 1, entry 1, 41%).
Contrary to this, when a guanidinium-type condensing reagent
(O-(benzotriazol-1-yl)-N,N,N’,N’-tetramethyluronium
applied the same strategy for the synthesis of Bu ester of 1
t
(Scheme 3). The introduction of Bu group to 4 was conducted
according to a general procedure widely employed for the O-
protection of carboxylic acids. For example, 4 was dissolved in
dichloromethane and treated with isobutene in the presence of a
catalytic amount of concentrated sulfuric acid. As we expected,
hexafluorophosphate [HBTU] or O-(7-azabenzotriazol-1-yl)-
N,N,N’,N’-tetramethyluronium hexafluorophosphate [HATU])

ACCEPTED MANUSCRIPT
4
Tetrahedron
Table 2. N-Terminal elongation of the amino esters derived from 1
(2 and 2’) with various β- or α-amino acids (11–15).
Table 1. N-Terminal elongation of the amino ester derived from 1
(2) with N-benzyloxycarbonyl-L-β-homoalanine (11).
F3C
yielda)
entry
product
CF3
reagent
Cbz NH
OH
+
HCl• H2N
OEt
O
O
1
Cbz NH
Boc NH
NH
OEt
OMe
OEt
HBTU (1.0 eq)
11
2
69%
O
11
O
O
O
O
O
/
/
/
/
/
2
F3C
NH
reagent
base (5.0 eq)
CF3
Cbz NH
OEt
2b)
3b)
HBTU (1.0 eq)
HATU (1.0 eq)
9%
CH2Cl2, rt
O
O
NH
11/2
65%
O
12
2'
yielda)
entry
reagent
base
Cbz
HN
CF3
1b)
2b)
3b)
4c)
5c)
EDC•HCl (1.0 eq), HOBt (1.0 eq)
HBTU (1.0 eq)
iPr2NEt
iPr2NEt
iPr2NEt
Et3N
41%
69%
61%
64%
86%
4
5
NH
HBTU (1.0 eq)
62%
O
13
HATU (1.0 eq)
2
(COCl)2 (6.0 eq)
(COCl)2 (6.0 eq)
Pyridine
Cbz
HN
CF3
HBTU (1.0 eq)
HBTU (1.0 eq)
a) Isolated yield. b) 2 and a base were successively added to a
mixture of 11 and condensing reagent(s). c) 11 was treated with
oxalyl chloride and then reacted with 2 in the presence of a base.
NH
OEt
48%
71%
O
14
2
BnO
Boc
HN
CF3
6b),c)
7b)
was used, the yield of 11/2 was notably improved (entries 2 and
3, 69 and 61%, respectively), in good agreement with the relative
reactivity of intermediates generated from these condensing
reagents.12 In every case, the obtained dipeptides exhibited a set
of resonances in 1H and 19F NMR spectra, indicating that the
racemization/epimerization of the stereogenic carbons did not
occur so that the target dipeptide 11/2 was obtained in a
stereopure form (Figure S5).
NH
OMe
DEPBT (2.0 eq) 73%
O
15
2'
a) Isolated yield. b) In place of ethyl ester 2, methy ester 2' was
used. c) Product was obtained as a mixture of diastereoisomers.
As an alternative approach for the same condensation, the
reaction of 2 with an acyl chloride derived from 11 was
examined, taking account of the cost of reagents, atom economy,
convenience in isolation, and scalability. According to the
reported procedure, 11 was converted into the acyl chloride by
treatment with oxalyl chloride. The acyl chloride, obtained as a
crude form by removal of volatiles from the reaction mixture,
was then mixed with 2 in dichloromethane in the presence of a
base to afford the target dipeptide 11/2 in good to excellent yields
(Table 1, entries 4 and 5). In good agreement with the case of the
N-protection of 1, the yield of 11/2 was significantly influenced
by the choice of base. When triethylamine was used as a base,
11/2 was obtained in 64% yield (entry 4), which is comparable to
those achieved by guanidinium-type condensing reagents (HATU
and HBTU). In this case, the condensation was likely to proceed
via the generation of a ketene from the acyl chloride. On the
other hand, when pyridine was used in place of triethylamine, the
yield of 11/2 was improved to 86% (entry 5), most likely owing
to the activity of the in-situ formed acyl pyridinium. In addition
to such good yield, another benefit of this condensation method
is the convenience in isolation. Because the reaction mixture
contained only hydrophilic and volatile materials except for the
target dipeptide, pure 11/2 could be isolated by simple washing
of the reaction mixture with aqueous acidic and alkaline
solutions. Again, the preservation of the stereochemistry of the
stereogenic carbons was verified by 19F NMR spectroscopy (data
not shown).
For the condensation with an N-protected β-amino acid with
conformationally
butoxycarbonylamino)cyclohexane-1-carboxylic
constrained
structure
[(1R,2R)-2-(tert-
acid, 12],
HBTU was not efficient as the condensing reagent (entry 2),
while HATU afforded the target dipeptide 12/2 in satisfactory
yield (entry 3), indicating the effect of steric hindrance around
the carbonyl group in 12. The procedure using HBTU was
applicable to the condensation with N-protected α-amino acids
[N-benzyloxycarbonyl-L-alanine (13) and N-benzyloxycarbonyl-
L-valine (14)] to afford the target dipeptides in acceptable yields
(entries 4 and 5). In all cases of entries 2–5, the resultant
1
dipeptides showed a set of signals in H and 19F NMR (for 19F
NMR, see Figure S7, b–d), suggesting the retention of
stereochemistry of stereogenic carbons through the condensation,
although the stereogenic carbon in 12–14 are located at the α-
position of the carbonyl group and are generally prone to
racemize. Among these dipeptides, 13/2 was chosen as a
representative to conduct more unambiguous study on
stereopurity, where an authentic sample of its diastereoisomer
was independently prepared, as in the case of 11/2 (Figure S5).
Comparison of the diastereoisomers in 19F NMR proved that any
detectable racemization/epimerization took place through the
formation of 13/2 (Figure S6). Only an exception was the
condensation with the N-protected α-amino acid bearing a polar
functionality [O-benzyl-N-(tert-butoxycarbonyl)-L-serine (15)],
where HBTU caused significant stereopurity loss of the target
dipeptide 15/2 (entry 6).
However, by using 3-
Heartened by these results, we next applied these optimized
conditions to the condensation with various β- or α-amino acid
derivatives (12–15), in order to prove the general utility of the β-
perfluoroalkyl β-amino acid 1 in peptide engineering (Table 2).
(diethoxyphosphoryloxy)-1,2,3-benzotriazin-4(3H)-one
(DEPBT)13 in place of HBTU, 15/2 could be obtained in
satisfactory yield (entry 7) in a stereopure form (Figure S7, e).

ACCEPTED MANUSCRIPT
5
and the preservation of the stereopurity was unambiguously
Table 3. O-Terminal elongation of the N-protected amino acids
derived from 1 (4 and 6) with various β- or α-amino esters (16–20).
confirmed. In addition, the reactivity of the amino group in 1
toward various acylating reagents was clearly elucidated in terms
of its basicity and nucleophilicity. As far as we are aware of, this
is the first stereo-regulated synthesis of peptides containing a β-
yielda)
entry
product
F3C
perfluoroalkyl-β-amino
acid.
By
applying
the
protection/deprotection/condensation methods established here,
various peptides containing 1 could be constructed, which should
contribute to the development of novel peptides containing
fluoroalkyl-β-amino acids.
1
82%
Cbz NH
NH
O
OEt
O
O
4 16
/
/
/
/
F3C
4. Experimental section
4.1. General
2
3
4
5
83%
84%
80%
85%
Boc NH
NH
17
OMe
O
6
4
1H, 13C, and 19F NMR spectra were recorded on a JEOL model
JNM-ECA500 spectrometer operating at 500.16, 125.77, and
F3C
OEt
1
470.62 MHz for H, 13C, and 19F NMR, respectively. Chemical
Cbz NH
NH
18
O
shifts were determined with respect to an internal ((CH3)4Si) and
O
1
an external (C6F6) reference for H and 19F NMR, respectively.
Matrix-assisted laser desorption ionization time-of-flight mass
(MALDI-TOF-MS) spectrometry was performed on an Applied
Biosystems model Voyager-DE™ STR or a Bruker model MDS
SCIEX 4800 MALDI TOF Analyzer using 2,5-dihydroxybenzoic
acid (DHB) or α-cyano-4-hydroxycinnamic acid (CHCA) as a
matrix. Electrospray-ionization TOF-MS (ESI-TOF-MS) spectra
were recorded on a JEOL model JMS-T100LC AccuTOF
spectrometer on a positive mode. Specific rotation was measured
with a JASCO model P-2200 polarimeter.
F3C
OMe
O
Boc NH
NH
19
O
6
OBn
F3C
OMe
O
Boc NH
NH
6/20
O
Materials
(S)-4,4,4-Trifluoro-3-aminobutyric acid (1),10 ethyl (S)-4,4,4-
a) Isolated yield.
trifluoro-3-aminobutyrate hydrochloride (2),7d methyl (S)-4,4,4-
trifluoro-3-aminobutyrate
benzyloxycarbonyl-L-β-homoalanine
aminobutyrate hydrochloride
butoxycarbonylamino)cyclohexane-1-carboxylic acid (12),15 and
hydrochloride
(11),14
(16),14
(2’),7d
ethyl
(1R,2R)-2-(tert-
N-
(S)-3-
2.4. Incorporation of 1 into peptides: elongation at the O-
terminus.
methyl
(1R,2R)-2-aminocyclohexane-1-carboxylate
To prove the general utility of the β-perfluoroalkyl β-amino
acid 1 in peptide engineering, the peptide-bond formation at its
O-terminus was also performed, although this reaction was of
little synthetic challenge (Table 3). The condensation of N-
protected derivatives of 1 (4 and 6) with various β- or α-amino
acid esters [L-β-homoalanine ethyl ester hydrochloride (16),
hydrochloride (17)15 were prepared according to the methods in
literatures. Dichloromethane was distilled twice successively
from P2O5 and CaH2 and stored over activated molecular sieves.
Other chemicals were used as received.
4.3. Synthesis of N- and O-protected derivatives of 1
4.3.1. Synthesis of 4 (Scheme 1, Route A)
methyl
(1R,2R)-2-aminocyclohexane-1-carboxylate
hydrochloride (17), L-alanine ethyl ester hydrochloride (18), L-
valine methyl ester hydrochloride (19), and O-benzyl-L-serine
methyl ester hydrochloride (20)] was conducted by applying a
common method; the N-protected amino acid (4 or 6) dissolved
in dichloromethane was treated with EDC•HCl (1.0 eq), HOBt
(1.0 eq), triethylamine (5.0 eq), and the amino ester (16–20, 1.0
eq) to afford the target dipeptide in satisfactory yield (entries 1–
To a solution of 1 (235 mg, 1.50 mmol) in water/1,4-dioxane
(1:2, v/v, 20 mL) were successively added K2CO3 (2.07 g, 15
mmol) and benzyl chloroformate (210 ꢀL, 1.50 mmol) at rt.
After being stirred at rt for 12 h, the resulting mixture was treated
with 1 M aqueous hydrochloric acid until the pH of the mixture
became ca. 1 and extracted with ethyl acetate (3 × 30 mL).
Organic layers combined were washed with brine (100 mL),
dried over anhydrous Na2SO4, and concentrated under reduced
1
5). In all cases, a set of signals were observed in H and 19F
NMR (for 19F NMR, see Figure S10), suggesting the retention of
stereochemistry during the condensation.
To confirm the
pressure to dryness to afford 4 as a white solid (245 mg, 0.84
mmol, 56%); mp 145–147 ºC.
27
D
2.1 (c 0.5, MeOH). 1H
retention of stereopurity more unambiguously, 4/16 and 4/18
were chosen as representatives of dipeptides prepared from an N-
protected β- and α-amino acids, respectively. Thus, authentic
samples of their diastereoisomers were independently prepared,
and the comparison of their 19F NMR spectra clearly excluded the
possibility of racemization/epimerization (Figures S8 and S9).
[
α
]
NMR (500 MHz, CD3OD, 25 °C): δ 7.31 (m, 5H), 5.12 (d, 1H, J
= 12.3 Hz), 5.10 (d, 1H, J = 12.3 Hz), 4.71 (m, 1H), 2.78 (dd, 1H,
J1 = 16.6 Hz, J2 = 4.0 Hz), 2.61 (dd, 1H, J1 = 16.3 Hz, J2 = 9.9
Hz) ppm. 13C NMR (125 MHz, CD3OD, 25 °C): δ 172.3, 158.2,
137.9, 129.5, 129.0, 128.7, 126.6 (q, J = 279 Hz), 67.9, 51.4 (q, J
= 31 Hz), 34.0 ppm. 19F NMR (470 MHz, CD3OD, 25 °C): δ –
77.9 (d, J = 8.1 Hz) ppm. IR (ATR): 3324, 3040, 2986, 2636,
1703, 1536, 1455, 1420, 1368, 1341, 1292, 1246, 1191, 1173,
1126, 1057, 1031, 1005, 971, 935, 864, 779, 736, 695, 645, 575
cm–1. MALDI-TOF-MS: [M + Na]+ calcd. 314.06, found 313.98.
3. Conclusions
The protection, deprotection, and condensation of a β-
perfluoroalkyl-β-amino acid ((S)-4,4,4-trifluoro-3-aminobutyric
acid, 1) were systematically studied. For every process, synthetic
routes and the reaction conditions were thoroughly optimized,

ACCEPTED MANUSCRIPT
6
Tetrahedron
4.3.2. Synthesis of 3 (Scheme 1, Route B)
To a 1,4-dioxane solution (2 mL) of 2 (111 mg, 0.50 mmol)
were successively added pyridine (100 ꢀL, 0.60 mmol), 4-
(dimethylamino)pyridine (12 mg, 0.10 mmol), and di-tert-butyl
dicarbonate (219 mg, 1.00 mmol) at rt. The mixture was stirred
at the temperature for 1 day. Precipitates generated in the
resultant mixture were filtered off, and the filtrate was
concentrated under reduced pressure to dryness. The resultant
residue was diluted with ethyl acetate (3 mL) and washed with
0.5 M aqueous solution of citric acid (2 × 3 mL), a saturated
aqueous solution of NaHCO3 (3 mL), and water (3 mL). The
organic layer was dried over anhydrous Na2SO4 and concentrated
To a dichloromethane solution (10 mL) of 2 (336 mg, 1.50
mmol) and pyridine (1.0 mL, 12 mmol) was dropwise added
benzyl chloroformate (0.75 mL, 5.3 mmol) at 0 °C. After being
allowed to warm to rt and then stirred at the temperature for 1
day, the reaction mixture was concentrated under reduced
pressure to dryness, diluted with ethyl acetate (10 mL), and
successively washed with 1 M aqueous hydrochloric acid (10
mL), a saturated aqueous solution of NaHCO3 (10 mL) and brine
(10 mL). The organic layer was dried over anhydrous Na2SO4
and concentrated to dryness under reduced pressure to afford 3 as
27
under reduced pressure to dryness to afford 5 as a white solid
a white solid (464 mg, 1.40 mmol, 96%); mp 84–86 ºC. [α]
D
27
D
1
(117 mg, 0.41 mmol, 82%); mp 69–71 ºC.
[
α
]
29.2 (c 0.5,
15.1 (c 0.5, CHCl3). H NMR (500 MHz, CDCl3, 25 °C): δ 7.36
(m, 5H), 5.51 (d, 1H, J = 9.2 Hz), 5.14 (s, 2H), 4.76 (m, 1H),
4.16 (q, 2H, J = 7.2 Hz), 2.77 (dd, 1H, J1 = 16.5 Hz, J2 = 4.9 Hz),
2.62 (dd, 1H, J1 = 16.0 Hz, J2 = 7.5 Hz), 1.24 (t, 3H, J = 7.2 Hz)
ppm. 13C NMR (125 MHz, CD3OD, 55 °C): δ 170.5, 158.2,
137.9, 129.5, 129.1, 128.8, 126.4 (q, J = 281 Hz), 70.7, 68.1,
62.3, 51.5 (q, J = 32 Hz), 34.5, 14.4 ppm. 19F NMR (470 MHz,
CDCl3, 25 °C): δ –79.5 (d, J = 7.2 Hz) ppm. IR (ATR): 3307,
3064, 2997, 2907, 1732, 1702, 1669, 1541, 1457, 1420, 1387,
1365, 1349, 1302, 1284, 1253, 1230, 1182, 1122, 1056, 1024,
969, 912, 884, 838, 779, 747, 735, 698, 659, 623, 577, 530 cm–1.
MALDI-TOF-MS: [M + Na]+ calcd. 342.09, found 342.00.
CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ 5.22 (d, 1H, J =
8.0 Hz), 4.70 (brs, 1H), 4.19 (q, 2H, J = 7.5 Hz), 2.75 (dd, 1H, J1
= 16.1 Hz, J2 = 5.2 Hz), 2.58 (dd, 1H, J1 = 15.8 Hz, J2 = 7.8 Hz),
1.45 (s, 9H), 1.27 (t, 3H, J = 7.5 Hz) ppm. 13C NMR (125 MHz,
CDCl3, 25 °C): δ 169.2, 154.6, 124.7 (q, J = 279 Hz), 80.7, 61.3,
49.4 (q, J = 31 Hz), 33.7, 28.1, 14.0 ppm. 19F NMR (470 MHz,
CDCl3, 25 °C): δ –79.6 (d, J = 7.2 Hz) ppm. IR (ATR): 3331,
2987, 2941, 2909, 2878, 1740, 1694, 1662, 1532, 1461, 1435,
1386, 1371, 1344, 1298, 1249, 1177, 1154, 1123, 1069, 1053,
1027, 919, 876, 850, 787, 753, 723, 700, 654, 635, 587, 527 cm–
1. MALDI-TOF-MS: [M + Na]+ calcd. 308.11, found 308.05.
4.3.6. Synthesis of 6 (Scheme 2, Route B)
4.3.3. Synthesis of 4 (Scheme 1, Route B)
The N-Boc protected derivative of 1 (6) was obtained
quantitatively from 5 in the same procedure as that for the
synthesis of 4 from 3. Physical properties of this sample were
essentially identical to those of 6 obtained by Scheme 2, Route
A.
To a 1,4-dioxane solution (3 mL) of 3 (306 mg, 1.00 mmol)
was added an aqueous solution (3 mL) of LiOH•H2O (126 mg,
3.00 mmol) at 0 °C. The mixture was allowed to warm to rt and
stirred at the temperature for 1 h. The resultant mixture was
treated with 3 M aqueous hydrochloric acid (1.5 mL) and
extracted with ethyl acetate (3 × 3 mL). Organic layers
combined were dried over anhydrous Na2SO4 and concentrated
under reduced pressure to dryness. The resultant residue was
dissolved in a mixture of saturated aqueous solution of NaHCO3
(5 mL) and water (5 mL). The aqueous solution was washed
with chloroform (3 × 5 mL), treated with 3 M aqueous
hydrochloric acid until the pH of the mixture became ca. 3, and
extracted with ethyl acetate (3 × 5 mL). Organic layers
combined were washed with brine (15 mL), dried over anhydrous
Na2SO4 and concentrated to dryness under reduced pressure to
afford 4 as a white solid (266 mg, 0.91 mmol, 91%). Physical
properties of this sample thus obtained were essentially identical
to those of 4 obtained by Scheme 1, Route A.
4.3.7. Synthesis of 7
To a mixture of 4 (2.92 g, 10.0 mmol) and concentrated
sulfuric acid (0.1 mL) in dichloromethane (50 mL) was added
isobutene at 0 °C with stirring until the total volume of the
solution became ca. 80 mL. The mixture was allowed to warm to
rt and stirred at the temperature for 4 days. The resultant mixture
was treated with triethylamine (0.26 mL) and concentrated under
reduced pressure. The resultant residue was diluted with ethyl
acetate (20 mL) and successively washed with a saturated
aqueous solution of NaHCO3 (2 × 10 mL) and water (2 × 10 mL).
The organic layer was dried over anhydrous Na2SO4 and
concentrated under reduced pressure. The resultant residue was
subjected to silica gel column chromatography eluted with
hexane/ethyl acetate (1:1, v/v) to afford 7 as a white solid (3.02
4.3.4. Synthesis of 6 (Scheme 2, Route A)
g, 8.7 mmol, 87%); mp 65–67 ºC. [α]
27 15.8 (c 0.5, CHCl3). 1H
To a solution of 1 (235 mg, 1.50 mmol) in water/1,4-dioxane
(1:2, v/v, 20 mL) were successively added K2CO3 (2.07 g, 15
mmol) and di-tert-butyl dicarbonate (330 mg, 1.50 mmol) at 0
°C. After being stirred at rt for 1 day, the resultant mixture was
treated with 1 M aqueous hydrochloric acid until the pH of the
mixture became ca. 2 and extracted with dichloromethane (3 ×
30 mL). Organic layers combined were washed with brine (100
mL), dried over anhydrous Na2SO4, and concentrated under
D
NMR (500 MHz, CDCl3, 25 °C): δ 7.35 (m, 5H), 5.54 (d, 1H, J =
9.7 Hz), 5.16 (d, 1H, J = 12.3 Hz), 5.12 (d, 1H, J = 12.3 Hz),
4.71 (m, 1H), 2.69 (dd, 1H, J1 = 16.0 Hz, J2 = 4.6 Hz), 2.58 (dd,
1H, J1 = 16.1 Hz, J2 = 8.0 Hz), 1.42 (s, 9H) ppm. 13C NMR (125
MHz, CDCl3, 25 °C): δ 168.2, 155.5, 135.8, 128.5, 128.3, 128.2,
124.7 (q, J = 282 Hz), 82.2, 67.4, 50.1 (q, J = 31 Hz), 34.6, 27.8
ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –79.5 (d, J = 7.7
Hz) ppm. IR (ATR): 3302, 3075, 2987, 2933, 2898, 1731, 1702,
1673, 1551, 1472, 1458, 1423, 1393, 1370, 1343, 1291, 1248,
1179, 1156, 1127, 1069, 1050, 1029, 1003, 957, 916, 868, 840,
777, 753, 700, 658, 628, 603, 579 cm–1. MALDI-TOF-MS: [M +
Na]+ calcd. 370.12, found 370.00.
reduced pressure to dryness to afford 6 as a white solid (210 mg,
26
0.80 mmol, 55%); mp 109–111 ºC. [
α
]
10.5 (c 0.5, MeOH).
D
1H NMR (500 MHz, CD3OD, 25 °C): δ 4.64 (m, 1H), 2.73 (dd,
1H, J1 = 16.5 Hz, J2 = 4.0 Hz), 2.56 (dd, 1H, J1 = 16.3 Hz, J2 =
10.4 Hz), 1.46 (s, 9H) ppm. 13C NMR (125 MHz, CD3OD, 55
°C): δ 172.3, 157.3, 126.6 (q, J = 282 Hz), 81.1, 50.9 (q, J = 31
Hz), 34.3, 28.6 ppm. 19F NMR (470 MHz, CD3OD, 25 °C): δ –
77.9 (d, J = 8.1 Hz) ppm. IR (ATR): 3354, 2989, 2941, 1692,
1524, 1463, 1434, 1422, 1395, 1371, 1298, 1253, 1228, 1154,
1121, 1087, 1050, 1025, 965, 926, 859, 783, 765, 644, 623, 526
cm–1. MALDI-TOF-MS: [M + Na]+ calcd. 280.08, found 280.00.
4.3.8. Synthesis of 8
To
a
solution of
7
(416 mg, 1.20 mmol) in
methanol/dichloromethane (1:1, v/v, 10 mL) was added 5% Pd/C
(416 mg), and the resultant mixture was degassed and purged
with hydrogen. After being stirred at rt for 12 h, the catalyst was
filtered off through a celite pad, and the solid was washed with a
mixture of dichloromethane/methanol (10:1, v/v). The filtrate
4.3.5. Synthesis of 5 (Scheme 2, Route B)

ACCEPTED MANUSCRIPT
7
and washing were combined and concentrated under reduced
successively washed with 0.5 M aqueous hydrochloric acid (3 ×
pressure to dryness to afford 8 as a white solid (254 mg, quant);
3 mL), a saturated aqueous solution of NaHCO3 (3 × 3 mL), and
brine (3 × 3 mL). The organic layer was dried over anhydrous
Na2SO4 and concentrated under reduced pressure. The resultant
residue was subjected to silica gel column chromatography eluted
with hexane/ethyl acetate (2:1, v/v) to afford 11/2 as a white solid
(139 mg, 0.34 mmol, 69%). When EDC•HCl/HOBt or HATU
27
mp 132–134 ºC.
[
α
]
–19.9 (c 0.2, MeOH). 1H NMR (500
D
MHz, CDCl3, 25 °C): δ 8.71 (brs, 2H), 4.37 (brs, 1H), 2.93 (m,
2H), 1.47 (s, 9H) ppm. 13C NMR (125 MHz, CDCl3, 25 °C): δ
167.5, 123.7 (q, J = 281 Hz), 83.1, 50.1 (q, J = 32 Hz), 32.9, 28.0
ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –77.4 (brs) ppm. IR
(ATR): 3447, 3387, 2955, 2853, 2691, 2528, 1727, 1587, 1528,
1459, 1427, 1387, 1372, 1328, 1285, 1237, 1196, 1137, 1097,
1070, 946, 899, 849, 804, 761, 729, 652, 564, 541 cm–1. ESI-
TOF-MS: [M + Na]+ calcd. 236.09, found 236.07.
was used in place of HBTU, 11/2 was obtained in 41% or 61%
27
D
yield, respectively; mp 177–179 ºC. [
α
]
14.0 (c 0.08, CHCl3).
1H NMR (500 MHz, CDCl3, 25 °C): δ 7.34 (m, 5H), 6.64 (brs,
1H), 5.39 (brs, 1H), 5.09 (s, 2H), 5.03 (m, 1H), 4.15 (q, 2H, J =
7.0 Hz), 4.07 (m, 1H), 2.71 (dd, 1H, J1 = 16.1 Hz, J2 = 5.2 Hz),
2.59 (dd, 1H, J1 = 16.3 Hz, J2 = 7.2 Hz), 2.48 (m, 2H), 1.26 (d,
3H, J = 5.7 Hz), 1.25 (t, 3H, J = 6.9 Hz) ppm. 13C NMR (125
MHz, CDCl3, 55 °C): δ 170.4, 169.2, 155.9, 136.6, 128.4, 128.0,
127.9, 124.6 (q, J = 282 Hz), 66.6, 61.4, 47.5 (q, J = 32 Hz),
44.7, 42.5, 33.2, 20.3, 13.9 ppm. 19F NMR (470 MHz, CDCl3, 25
°C): δ –78.9 (d, J = 7.7 Hz) ppm. IR (ATR): 3316, 3301, 3067,
2980, 2927, 2853, 1728, 1681, 1660, 1535, 1456, 1427, 1371,
1341, 1309, 1282, 1263, 1203, 1176, 1120, 1065, 1025, 978, 948,
909, 881, 841, 779, 749, 723, 695, 662, 578, 552 cm–1. MALDI-
TOF-MS: [M + Na]+ calcd. 427.15, found 427.02.
4.3.9. Synthesis of 9
To an N,N-dimethylformamide solution (100 mL) of 6 (3.85 g,
15.0 mmol) were successively added K2CO3 (2.07 g, 15.0 mmol)
and benzyl bromide (2.0 mL, 16.5 mmol) at rt. The mixture was
stirred at the temperature for 1 day, treated with water (20 mL),
and extracted with ethyl acetate (3 × 30 mL). Organic layers
combined were successively washed with water (30 mL) and
brine (30 mL), dried over anhydrous Na2SO4 and concentrated
under reduced pressure. The resultant residue was subjected to
silica gel column chromatography eluted with hexane/ethyl
acetate (2:1, v/v) to afford 9 as a white solid (3.26 g, 9.4 mmol,
27 21.6 (c 0.5, CHCl3). 1H NMR (500 MHz,
4.4.2. Synthesis of 11/2 (via the conversion of 11
into acyl chloride)
62%); mp 65 ºC. [α]
D
CDCl3, 25 °C): δ 7.36 (m, 5H), 5.21 (d, 1H, J = 8.5 Hz), 5.15 (s,
2H), 4.73 (brs, 1H), 2.79 (dd, 1H, J1 = 16.1 Hz, J2 = 4.6 Hz), 2.65
(dd, 1H, J1 = 15.7 Hz, J2 = 7.7 Hz), 1.45 (s, 9H) ppm. 13C NMR
(125 MHz, CDCl3, 25 °C): δ 169.0, 154.6, 135.1, 128.6, 128.5,
128.4, 124.7 (q, J = 282 Hz), 80.8, 67.2, 49.4 (q, J = 32 Hz),
33.7, 28.1 ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –79.6 (d,
J = 7.2 Hz) ppm. IR (ATR): 3341, 3037, 3016, 2979, 2939,
1730, 1697, 1664, 1525, 1460, 1440, 1389, 1373, 1336, 1287,
1248, 1164, 1147, 1121, 1049, 1024, 979, 908, 853, 780, 751,
697, 657, 632, 580 cm–1. MALDI-TOF-MS: [M + Na]+ calcd.
370.12, found 370.03.
To a dichloromethane solution (5 mL) of 11 (142 mg, 0.60
mmol) was added oxalyl chloride (0.26 mL, 3.0 mmol). The
mixture was allowed to warm to rt, stirred at the temperature for
3 h, and concentrated under reduced pressure to dryness to give a
residue containing the acyl chloride derived from 11. To the
resultant residue were successively added dichloromethane (3
mL), a dichloromethane solution (2 mL) of 2 (111 mg, 0.50
mmol), and pyridine (0.20 mL, 2.5 mmol) at rt. After being
stirred at the temperature for 12 h, the resultant mixture was
concentrated under reduced pressure, diluted with ethyl acetate
(10 mL), and washed with 0.5 M aqueous hydrochloric acid (3 ×
3 mL), a saturated aqueous solution of NaHCO3 (3 × 3 mL), and
brine (3 × 3 mL). The organic layer was dried over anhydrous
Na2SO4 and concentrated under reduced pressure. The resultant
residue was subjected to silica gel column chromatography eluted
with hexane/ethyl acetate (1:1, v/v) to afford 11/2 as a white solid
(175 mg, 0.43 mmol, 86%). When triethylamine was used in
place of pyridine, 11/2 was obtained in 64% yield. Physical
properties of the product samples were identical to those of 11/2
obtained by the method using a condensing reagent [Section
4.4.1. Synthesis of 11/2 (by using a condensing reagent)].
4.3.10. Synthesis of 10
To a dichloromethane solution (10 mL) of 9 (694 mg, 2.00
mmol) was added trifluoroacetic acid (TFA, 10 mL) at rt. After
being stirred at rt for 3 h, the resultant mixture was concentrated
under reduced pressure to dryness to afford 10 as a white solid
(724 mg, quant.); mp 106–109 ºC. [α]
27 7.7 (c 0.2, MeOH). 1H
D
NMR (500 MHz, CD3OD, 25 °C): δ 7.38 (m, 5H), 5.24 (d, 1H, J
= 12.3 Hz), 5.23 (d, 1H, J = 12.3 Hz), 4.53 (m, 1H), 3.14 (dd, 1H,
J1 = 17.8 Hz, J2 = 4.0 Hz), 2.89 (dd, 1H, J1 = 17.8 Hz, J2 = 9.2
Hz) ppm. 13C NMR (125 MHz, CD3OD, 25 °C): δ 169.8, 162.3
(q, J = 36.0 Hz), 136.7, 129.7, 129.6, 129.5, 125.1 (q, J = 279
Hz), 117.8 (q, J = 289 Hz), 68.7, 50.6 (q, J = 32.8 Hz), 32.4 ppm.
19F NMR (470 MHz, CD3OD, 25 °C): δ –76.4 (d, J = 6.3 Hz), –
76.5 ppm. IR (ATR): 3072, 2924, 2853, 2813, 2674, 1750, 1625,
1551, 1500, 1439, 1423, 1390, 1330, 1293, 1255, 1220, 1185,
1165, 1136, 1105, 1075, 1011, 978, 947, 910, 888, 839, 804, 741,
724, 696, 657, 573, 548 cm–1. MALDI-TOF-MS: [M –
CF3COOH + Na]+ calcd. 270.07, found 269.96.
4.4.3. Synthesis of 12/2’
The dipeptide 12/2’ was obtained as a white solid from 12 and
2’ in a procedure similar to the synthesis of 11/2, where HATU
27
was used as a condensing regent (65%); decomp. >220 ºC. [α]
D
–3.6 (c 0.3, CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ 6.76
(d, 1H, J = 8.6 Hz), 5.07 (m, 1H), 4.57 (brs, 1H), 3.72 (s, 3H),
3.52 (m, 1H), 2.74 (dd, 1H, J1 = 16.0 Hz, J2 = 5.2 Hz), 2.64 (dd,
1H, J1 = 16.1 Hz, J2 = 7.5 Hz), 2.31 (brs, 1H), 2.00 (m, 1H), 1.75
(m, 1H), 1.41 (s, 9H), 1.28 (m, 4H) ppm. 13C NMR (125 MHz,
CDCl3/CD3OD (2:1, v/v), 55 °C): δ 175.5, 170.1, 156.5, 125.6 (q,
J = 281 Hz), 79.9, 52.4, 51.6, 51.2, 48.0 (q, J = 32 Hz), 33.8,
33.4, 30.0, 28.5, 25.2, 25.1 ppm. 19F NMR (470 MHz, CDCl3, 25
°C): δ –78.6 (d, J = 7.6 Hz) ppm. IR (ATR): 3332, 3309, 3216,
3015, 2989, 2937, 2857, 1741, 1678, 1666, 1533, 1440, 1417,
1391, 1356, 1320, 1296, 1278, 1253, 1228, 1175, 1141, 1120,
1074, 1054, 1011, 993, 930, 906, 895, 868, 780, 747, 661, 639,
596, 560 cm–1. MALDI-TOF-MS: [M + Na]+ calcd. 419.18,
found 413.31.
4.4. Synthesis of dipeptides composed of 1
4.4.1. Synthesis of 11/2 (by using a condensing
reagent)
To a dichloromethane solution (3 mL) of 11 (119 mg, 0.50
mmol) was added HBTU (190 mg, 0.50 mmol) at 0 °C. After
being stirred at the temperature for 30 min, the mixture was
allowed to warm to rt, and treated with a dichloromethane
i
solution (5 mL) of 2 (111 mg, 0.50 mmol), followed by Pr2NEt
(0.43 mL, 2.5 mmol). After being stirred at the temperature for
12 h, the resultant mixture was concentrated under reduced
pressure to dryness, diluted with ethyl acetate (10 mL), and
4.4.4. Synthesis of 13/2

ACCEPTED MANUSCRIPT
8
Tetrahedron
The dipeptide 13/2 was obtained as a white solid from 13
concentrated under reduced pressure. The resultant residue was
subjected to silica gel column chromatography eluted with
and 2 in a procedure similar to the synthesis of 11/2, where
HBTU was used as a condensing regent (62%); mp 114–116 ºC.
hexane/ethyl acetate (2:1, v/v) to afford 4/16 as a white solid
27
[
α
]
28 8.0 (c 0.5, CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ
(166 mg, 0.41 mmol, 82%); mp 165–169 ºC. [
α
]
3.3 (c 0.2,
D
D
7.35 (m, 5H), 6.98 (d, 1H, J = 7.5 Hz), 5.24 (m, 1H), 5.12 (s,
2H), 5.00 (m, 1H), 4.26 (m, 1H), 4.15 (q, 2H, J = 7.3 Hz), 2.73
(m, 1H), 2.62 (dd, 1H, J1 = 16.1 Hz, J2 = 7.4 Hz), 1.40 (d, 3H, J =
6.9 Hz), 1.25 (t, 3H, J = 7.2 Hz) ppm. 13C NMR (125 MHz,
CDCl3, 25 °C): δ 172.7, 168.9, 156.0, 136.0, 128.4, 128.1, 127.8,
124.5 (q, J = 281 Hz), 66.8, 61.3, 50.3, 47.5 (q, J = 32 Hz), 33.2,
18.3, 13.8 ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –79.0 (d,
J = 7.2 Hz) ppm. IR (ATR): 3338, 3092, 3035, 2979, 2934,
1723, 1696, 1682, 1529, 1432, 1369, 1347, 1297, 1264, 1245,
1200, 1175, 1142, 1127, 1117, 1068, 1025, 963, 952, 892, 876,
846, 785, 739, 718, 694, 660, 627, 575, 525 cm–1. MALDI-TOF-
MS: [M + Na]+ calcd. 413.13, found 413.03.
CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ 7.34 (m, 5H),
6.37 (d, 1H, J = 7.5 Hz), 6.28 (d, 1H, J = 8.6 Hz), 5.14 (s, 2H),
4.66 (m, 1H), 4.32 (m, 1H), 4.15 (m, 2H), 2.77 (m, 1H), 2.62 (dd,
1H, J1 = 15.2 Hz, J2 = 4.9 Hz), 2.47 (m, 3H), 1.27 (t, 3H, J = 7.2
Hz), 1.20 (d, 3H, J = 6.9 Hz) ppm. 13C NMR (125 MHz, CDCl3,
55 °C): δ 171.6, 167.5, 155.7, 136.1, 128.5, 128.2, 128.0, 124.9
(q, J = 283 Hz), 67.4, 60.7, 50.7 (q, J = 32 Hz), 42.4, 34.7, 29.7,
19.7, 14.1 ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –78.7 (d,
J = 7.2 Hz) ppm. IR (ATR): 3298, 3068, 3036, 2988, 2973,
2929, 1723, 1694, 1635, 1543, 1498, 1458, 1439, 1377, 1356,
1308, 1248, 1176, 1149, 1121, 1049, 1032, 936, 912, 875, 854,
824, 774, 752, 697, 670, 649, 610, 580, 564, 521 cm–1. MALDI-
TOF-MS: [M + Na]+ calcd. 427.15, found 427.02.
4.4.5. Synthesis of 14/2
4.4.8. Synthesis of 6/17
The dipeptide 14/2 was obtained as a white solid from 14 and
2 in a procedure similar to the synthesis of 11/2, where HBTU
The dipeptide 6/17 was obtained as a white solid from 6 and
27
was used as a condensing reagent (48%); mp 155–156 ºC. [
α
]
17 in the same procedure for the synthesis of 4/16 (83%);
D
23.4 (c 0.5, CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ 7.35
(m, 5H), 6.83 (d, 1H, J = 9.7 Hz), 5.33 (d, 1H, J = 6.9 Hz), 5.11
(s, 2H), 5.02 (m, 1H), 4.16 (q, 2H, J = 7.5 Hz), 4.02 (t, 1H, J =
7.5 Hz), 2.73 (dd, 1H, J1 = 15.5 Hz, J2 = 4.0 Hz), 2.66 (dd, 1H, J1
= 16.0 Hz, J2 = 6.3 Hz), 2.12 (m, 1H), 1.25 (t, 3H, J = 6.9 Hz),
0.97 (d, 3H, J = 6.3 Hz), 0.93 (d, 3H, J = 6.9 Hz) ppm. 13C NMR
(125 MHz, CDCl3, 25 °C): δ 171.3, 169.1, 156.4, 136.1, 128.5,
128.2, 127.9, 124.5 (q, J = 281 Hz), 67.0, 61.5, 60.3, 47.4 (q, J =
32 Hz), 33.1, 31.2, 18.8, 17.7, 13.9 ppm. 19F NMR (470 MHz,
CDCl3, 25 °C): δ –78.7 (d, J = 7.7 Hz) ppm. IR (ATR): 3282,
3074, 2974, 2960, 2911, 2874, 1734, 1692, 1666, 1531, 1469,
1456, 1388, 1372, 1340, 1293, 1246, 1229, 1184, 1122, 1072,
1038, 1028, 967, 926, 902, 873, 843, 788, 748, 695, 658, 594,
576 cm–1. MALDI-TOF-MS: [M + Na]+ calcd. 441.16, found
441.30.
decomp. > 200 ºC. [
α
]
10.7 (c 0.3, MeOH). 1H NMR (500
27
D
MHz, CDCl3, 25 °C): δ 5.96 (d, 1H, J = 5.8 Hz), 5.67 (d, 1H, J =
5.8 Hz), 4.56 (m, 1H), 3.99 (m, 1H), 2.60 (dd, 1H, J1 = 15.5 Hz,
J2 = 5.2 Hz), 2.39 (dd, 1H, J1 = 14.4 Hz, J2 = 5.2 Hz), 2.39 (td,
1H, J1 = 12.0 Hz, J2 = 4.0 Hz), 2.03 (m, 1H), 1.95 (m, 1H), 1.74
(m, 2H), 1.62 (m, 1H), 1.45 (s, 9H), 1.19 (m, 2H) ppm. 13C
NMR (125 MHz, CD3OD, 55 °C): δ 176.0, 169.6, 157.1, 126.8
(q, J = 282 Hz), 81.1, 52.3, 51.2, 51.1 (q, J = 31 Hz), 50.3, 35.8,
33.1, 29.9, 28.6, 25.7, 25.5 ppm. 19F NMR (470 MHz, CDCl3, 25
°C): δ –78.9 (d, J = 5.4 Hz) ppm. IR (ATR): 3327, 3263, 3062,
2998, 2982, 2966, 2941, 2862, 1735, 1698, 1646, 1542, 1438,
1367, 1308, 1276, 1251, 1208, 1157, 1122, 1051, 1015, 973, 917,
876, 861, 758, 692, 655, 628, 570, 554 cm–1. MALDI-TOF-MS:
[M + Na]+ calcd. 419.18, found 419.33.
4.4.9. Synthesis of 4/18
4.4.6. Synthesis of 15/2
The dipeptide 4/18 was obtained as a white solid from 4 and
The dipeptide 15/2 was obtained as a transparent oil from 15
and 2 in a procedure similar to the synthesis of 11/2, where
18 in the same procedure for the synthesis of 4/16 (84%); mp
27
188–191 ºC. [
α
]
–29.9 (c 0.5, MeOH). 1H NMR (500 MHz,
D
DEPBT13 was used as a condensing reagent (73%); [
α
]
25 23.4 (c
CD3OD, 25 °C): δ 7.33 (m, 5H), 5.10 (q, 2H, J = 7.5 Hz), 4.76
(m, 1H), 4.35 (q, 2H, J = 7.3 Hz), 4.15 (q, 2H, J = 7.1 Hz), 2.69
(dd, 1H, J1 = 14.9 Hz, J2 = 4.0 Hz), 2.55 (dd, 1H, J1 = 15.2 Hz, J2
= 10.0 Hz), 1.32 (d, 3H, J = 6.9 Hz), 1.25 (t, 3H, J = 7.2 Hz)
ppm. 13C NMR (125 MHz, CD3OD, 25 °C): δ 174.0, 170.2,
158.0, 138.0, 129.4, 129.1, 128.8, 126.6 (q, J = 280 Hz), 68.0,
62.4, 60.3, 51.7 (q, J = 32 Hz), 49.7, 35.3, 17.5, 14.4 ppm. 19F
NMR (470 MHz, CDCl3, 25 °C): δ –78.7 (d, J = 7.7 Hz) ppm. IR
(ATR): 3295, 3068, 3035, 2982, 2929, 2875, 2855, 1735, 1698,
1643, 1542, 1451, 1437, 1377, 1358, 1324, 1305, 1277, 1247,
1228, 1172, 1151, 1119, 1078, 1049, 1030, 979, 937, 915, 878,
864, 827, 755, 738, 696, 639, 606, 580, 565 cm–1. MALDI-TOF-
MS: [M + Na]+ calcd. 413.13, found 413.26.
D
0.5, CHCl3). 1H NMR (500 MHz, CDCl3, 25 °C): δ 7.32 (m,
5H), 7.22 (d, 1H, J = 5.8 Hz), 5.38 (brs, 1H), 5.07 (m, 1H), 4.56
(d, 1H, J = 11.5 Hz), 4.53 (d, 1H, J = 11.5 Hz), 4.31 (brs, 1H),
3.89 (dd, 1H, J1 = 9.2 Hz, J2 = 4.0 Hz), 3.66 (s, 3H), 3.55 (m,
1H), 2.74 (dd, 1H, J1 = 16.1 Hz, J2 = 5.2 Hz), 2.61 (dd, 1H, J1 =
16.3 Hz, J2 = 7.8 Hz), 1.45 (s, 9H) ppm. 13C NMR (125 MHz,
CDCl3, 25 °C): δ 170.5, 169.3, 155.4, 137.1, 128.4, 127.9, 127.8,
124.4 (q, J = 282 Hz), 80.3, 73.5, 69.5, 53.7, 52.2, 47.5 (q, J = 32
Hz), 33.1, 28.1 ppm. 19F NMR (470 MHz, CDCl3, 25 °C): δ –
79.0 (d, J = 3.6 Hz) ppm. IR (ATR): 3304, 3032, 2978, 2956,
2930, 2871, 1744, 1712, 1672, 1523, 1496, 1455, 1391, 1366,
1285, 1238, 1168, 1126, 1023, 997, 960, 913, 860, 781, 737, 698,
662, 611, 566 cm–1. MALDI-TOF-MS: [M + Na]+ calcd. 471.17,
found 471.36.
4.4.10. Synthesis of 6/19
The dipeptide 6/19 was obtained as a white solid from 6 and
4.4.7. Synthesis of 4/16
19 in the same procedure for the synthesis of 4/16 (80%); mp
27
D
To a dichloromethane solution (2 mL) of 4 (146 mg, 0.50
mmol) were successively added EDC•HCl (96 mg, 0.50 mmol),
HOBt (68 mg, 0.50 mmol), a dichloromethane solution (3 mL) of
16 (84 mg, 0.50 mmol), and triethylamine (0.27 mL, 2.5 mmol)
at rt. After being stirred at the temperature for 12 h, the resultant
mixture was concentrated to dryness under reduced pressure,
diluted with ethyl acetate (15 mL), and successively washed with
0.5 M aqueous hydrochloric acid (3 × 20 mL), a saturated
aqueous solution of NaHCO3 (3 × 20 mL), and brine (3 × 20
mL). The organic layer was dried over anhydrous Na2SO4 and
179–181 ºC. [
α
]
40.9 (c 0.5, CHCl3). 1H NMR (500 MHz,
CDCl3, 25 °C): δ 6.19 (d, 1H, J = 8.6 Hz), 5.94 (d, 1H, J = 9.2
Hz), 4.61 (m, 1H), 4.56 (m, 1H), 3.75 (s, 3H), 2.69 (dd, 1H, J1 =
15.5 Hz, J2 = 4.6 Hz), 2.55 (dd, 1H, J1 = 15.0 Hz, J2 = 5.1 Hz),
2.17 (m, 1H), 1.44 (s, 9H), 0.94 (d, 3H, J = 6.9 Hz), 0.91 (d, 3H,
J = 6.9 Hz) ppm. 13C NMR (125 MHz, CDCl3, 25 °C): δ 172.8,
168.7, 154.9, 124.9 (q, J = 283 Hz), 80.5, 57.2, 52.3, 49.9 (q, J =
31 Hz), 34.4, 31.2, 28.2, 18.8, 17.6 ppm. 19F NMR (470 MHz,
CDCl3, 25 °C): δ –78.8 (d, J = 7.2 Hz) ppm. IR (ATR): 3324,
3014, 2980, 2953, 2933, 2878, 1741, 1692, 1652, 1530, 1468,

ACCEPTED MANUSCRIPT
9
V. A.; Ohkura, H.; Yasumoto, M. J. Fluorine Chem. 2006, 127,
930.
1435, 1393, 1367, 1310, 1250, 1207, 1180, 1152, 1122, 1055,
1025, 975, 915, 881, 853, 781, 645, 622, 563, 550 cm–1.
MALDI-TOF-MS: [M + Na]+ calcd. 393.16, found 393.27.
8. (a) Lebouvier, N.; Laroche, C.; Huguenot, F.; Brigaud, T.
Tetrahedron Lett. 2002, 43, 2827; (b) Funabiki, K.; Nagamori, M.;
Matsui, M. J. Fluorine Chem. 2004, 125, 1347; (c) Fustero, S.;
Jiménez, D.; Sanz-Cervera, J. F.; Sánchez-Roselló, M.; Esteban,
E.; Simón-Fuentes, A. Org. Lett. 2005, 7, 3433; (d) Santos, M. D.;
Crousse, B.; Bonnet-Delpon, D. Synlett 2008, 399.
9. (a) Galushko, S. V.; Shishkina, I. P.; Soloshonok, V. A. J.
Chromatogr. 1992, 592, 345; (b) Soloshonok, V. A.; Kirilenko, A.
G.; Fokina, N. A.; Shishkina, I. P.; Galushko, S. V.; Kukhar, V.
P.; Švedas, V. K.; Kozlova, E. V. Tetrahedron: Asymmetry 1994,
5, 1119; (c) Soloshonok, V. A.; Kirilenko, A. G.; Fokina, N. A.;
Kukhar, V. P.; Galushko, S. V.; Švedas, V. K.; Resnati, G.
Tetrahedron: Asymmetry 1994, 5, 1225; (d) Michaut, V.; Metz, F.;
Paris, J.-M.; Plaquevent, J.-C. J. Fluorine Chem. 2007, 128, 889.
10. Ishida, Y.; Iwahashi, N.; Nishizono, N.; Saigo, K. Tetrahedron
Lett. 2009, 50, 1889.
4.4.11. Synthesis of 6/20
The dipeptide 6/20 was obtained as a white solid from 6 and
20 in the same procedure for the synthesis of 4/16 (85%); mp
27
D
146–148 ºC. [
α
]
53.7 (c 0.5, CHCl3). 1H NMR (500 MHz,
CDCl3, 25 °C): δ 7.32 (m, 5H), 6.45 (d, 1H, J = 6.9 Hz), 5.91 (d,
1H, J = 8.6 Hz), 4.73 (dt, 1H, J1 = 10.9 Hz, J2 = 2.9 Hz), 4.63 (m,
1H), 4.53 (d, 1H, J = 12.6 Hz), 4.48 (d, 1H, J = 12.0 Hz), 3.91
(dd, 1H, J1 = 9.8 Hz, J2 = 2.9 Hz), 3.75 (s, 3H), 3.67 (dd, 1H, J1 =
9.7 Hz, J2 = 2.9 Hz), 2.69 (dd, 1H, J1 = 15.5 Hz, J2 = 4.6 Hz),
2.53 (dd, 1H, J1 = 14.9 Hz, J2 = 4.6 Hz), 1.45 (s, 9H) ppm. 13C
NMR (125 MHz, CDCl3, 55 °C): δ 170.4, 168.4, 154.8, 137.4,
128.3, 127.8, 127.6, 125.0 (q, J = 283 Hz), 80.4, 73.3, 69.5, 52.8,
52.3, 49.9, 34.2, 28.1 ppm. 19F NMR (470 MHz, CDCl3, 25 °C):
δ –78.8 (d, J = 6.8 Hz) ppm. IR (ATR): 3315, 2987, 2942, 2871,
2860, 1740, 1698, 1650, 1532, 1441, 1367, 1340, 1308, 1249,
1232, 1177, 1152, 1120, 1101, 1056, 1028, 984, 920, 882, 853,
782, 736, 697, 672, 635, 612, 580 cm–1. MALDI-TOF-MS: [M +
Na]+ calcd. 471.17, found 471.34.
11. (a) Seebach, D.; Matthews, J. L. Chem. Commun. 1997, 21, 2015;
Seebach, D.; Beck, A. K.; Bierbaum, D. J. Chem. Biodivers. 2004,
1, 1111.
12. Valeur, E.; Bradley, M. Chem. Soc. Rev. 2009, 38, 606.
13. Li, H.; Jiang, X.; Ye, Y.; Fan, C.; Romoff, T.; Goodman, M. Org.
Lett. 1999, 1, 91.
14. Arvidsson, P. I.; Frackenpohl, J.; Seebach, D. Helv. Chim. Acta
2003, 86, 1522.
15. Nohira, H.; Ehara, K.; Miyashita, A. Bull. Chem. Soc. Jpn. 1970,
43, 2230.
Acknowledgments
A part of this research is financially supported by the Grant-
in-Aid for Young Scientists (B) 21750137 (MEXT, Japan).
Supplementary material
Optimization of reaction conditions for the synthesis of 5 and
19F NMR spectra of the derivatives of 1. Supplementary data
associated with this article can be found in the online version, at
http://dx.doi.org/XXXXXXX.
References and notes
1. (a) Yoder, N. C.; Kumar, K. Chem. Soc. Rev. 2002, 31, 335; (b)
Marsh, E. N. G.; Buer, B. C.; Ramamoorthy, A. Mol. BioSyst.
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21, 453; (d) Salwiczek, M.; Nyakatura, E. K.; Gerling, U. I. M.;
Ye, S.; Koksch, B. Chem. Soc. Rev. 2012, 41, 2135.
2. (a) Müller, K.; Faeh, C.; Diederich, F. Science 2007, 317, 1881;
(b) O’Hagan, D. Chem. Soc. Rev. 2008, 37, 308.
3. Cobb, S.; Murphy, C. J. Fluorine Chem. 2009, 130, 132.
4. (a) Aceña, J. L.; Simón-Fuentes, A.; Fustero, S. Curr. Org. Chem.
2010, 14, 928; (b) Qiu, X. L.; Qing, F. L. Eur. J. Org. Chem. 2011,
3261.
5. (a) Su, T.; Naughton, M. A.; Smyth, M. S.; Rose, J. W.; Arfsten,
A. E.; McCowan, J. R.; Jakubowski, J. A.; Wyss, V. L.;
Ruterbories, K. J.; Sall, D. J.; Scarborough, R. M. J. Med. Chem.
1997, 40, 4308; (b) Volonterio, A. Bellosta, S.; Bravo, P.;
Canavesi, M.; Corradi, E.; Meille, S. V.; Monetti, M.; Moussier,
N.; Zanda, M. Eur. J. Org. Chem. 2002, 428; (c) Formicola, L.;
Maréchal, X.; Basse, N.; Bouvier-Durand, M.; Bonnet-Delpon, D.;
Milcent, T.; Reboud-Ravaux, M.; Ongeri, S. Bioorg. Med. Chem.
Lett. 2009, 19, 83.
6. (a) Fustero, S.; Salavert, E.; Pina, B.; Ramírez de Arellano, C.;
Asensio, A. Tetrahedron 2001, 57, 6475; (b) Fustero, S.; Pina, B.;
Salavert, E.; Navarro, A.; Ramírez de Arellano, M. C.; Fuentes, A.
S. J. Org. Chem. 2002, 67, 4667; (c) Fustero, S.; Sánchez-Roselló,
M.; Sanz-Cervera, J. F.; Aceña, J. L.; del Pozo, C.; Fernández, B.;
Bartolomé, A.; Asensio, A. Org. Lett. 2006, 8, 4633. (d) Fustero,
S.; del Pozo, C.; Catalán, S.; Alemán, J.; Parra, A.; Marcos, V.;
Ruano, J. L. G. Org. Lett. 2009, 11, 641.
7. (a) Soloshonok, V. A.; Kirilenko, A. G.; Kukhar, V. P.; Resnati,
G. Tetrahedron Lett. 1993, 34, 3621; (b) Soloshonok, V. A.;
Kirilenko, A. G.; Galushko, S. V.; Kukhar, S. V. Tetrahedron Lett.
1994, 35, 5063; (c) Soloshonok, V. A.; Ono, T.; Soloshonok, I. V.
J. Org. Chem. 1997, 62, 7538; (d) Soloshonok, V. A.; Ohkura, H.;
Yasumoto, M. J. Fluorine Chem. 2006, 127, 924; (e) Soloshonok,

ACCEPTED MANUSCRIPT
Supplementary Material
Stereo-regulated synthesis of peptides containing a
β-trifluoromethyl-β-amino acid
Joon-il Cho a, Nao Nishizono b, Nobutaka Iwahashi b, Kazuhiko Saigo c and Yasuhiro Ishida a*
a RIKEN Center for Emergent Matter Science, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan
b Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of
Tokyo, Hongo, Bunkyo-ku, Tokyo 113-8656, Japan
c School of Environmental Science and Engineering, Kochi University of Technology, Miyanokuchi,
Tosayamada-cho, Kami, Kochi 782-8502, Japan
Table of Contents
(i) Optimization of reaction conditions for the synthesis of 5........................................................... S2
(ii) Determination of the enantiomeric purity of 4 and 6....................................................................S3
(iii) Determination of the enantiomeric purity of 8 and 10..................................................................S4
(iv) Determination of the diastereomeric purity of 11/2 and 13/2.......................................................S5
(v) 19F NMR spectra of 11/2, 12/2’, 13/2, 14/2, and 15/2’ ................................................................ S6
(vi) Determination of the diastereomeric purity of 4/16 and 4/18.......................................................S7
(vii) 19F NMR spectra of 4/16, 6/17, 4/18, 6/19, and 6/20....................................................................S8
S1

ACCEPTED MANUSCRIPT
(i) Optimization of reaction conditions for the synthesis of 5
Table S1. N-Boc protection of the amino ester derived from 1 (2).
F3C
F3C
reagent, base
HCl• H2N
OEt
Boc NH
OEt
solvent
rt, 1 day
O
O
2
5
yielda)
entry reagent
base
solvent
1
2
3
Boc2O (2.0 eq), DMAP (0.2 eq) Pyridine (1.2 eq) CH2Cl2
60%
82%
84%
Boc2O (2.0 eq), DMAP (0.2 eq) Pyridine (1.2 eq) 1,4-Dioxane
Boc2O (3.5 eq), DMAP (0.2 eq) Pyridine (8.0 eq) 1,4-Dioxane
a) Isolated yield.
S2

ACCEPTED MANUSCRIPT
(ii) Determination of the enantiomeric purity of 4 and 6
(a)
(b)
(c)
–78.4 –78.5 –78.6 –78.7 –78.8 –78.9 –79.0 –79.1 –79.2 –79.3 –79.4 –79.5
19F (ppm)
Figure S1. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 4 (10 mM), (b) the antipode of 4 (10 mM),
and (c) a mixture of 4 (5 mM) and its antipode (5 mM), where (2S,5S)-2,5-diphenylpyrrolidine (25 mM)
was added as a chiral NMR shift reagent.
(a)
(b)
(c)
–78.5 –78.6 –78.7 –78.8 –78.9 –79.0 –79.1 –79.2 –79.3 –79.4 –79.5 –79.6
19F (ppm)
Figure S2. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 6 (10 mM), (b) the antipode of 6 (10 mM),
and (c) a mixture of 6 (5 mM) and its antipode (5 mM), where (2S,5S)-2,5-diphenylpyrrolidine (25 mM)
was added as a chiral NMR shift reagent.
S3

ACCEPTED MANUSCRIPT
(iii) Determination of the enantiomeric purity of 8 and 10
(a)
(b)
(c)
–76.7 –76.8 –76.9 –77.0 –77.1 –77.2 –77.3 –77.4 –77.5 –77.6 –77.7
19F (ppm)
Figure S3. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 8 (10 mM), (b) the antipode of 8 (10 mM),
and (c) a mixture of 8 (5 mM) and its antipode (5 mM), where dimethyl L-(+)-tartrate (1.0 M) was added
as a chiral NMR shift reagent.
(a)
(b)
(c)
–79.1 –79.2 –79.3 –79.4 –79.5 –79.6 –79.7 –79.8 –79.9 –80.0 –80.1
19F (ppm)
Figure S4. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 10 (10 mM), (b) the antipode of 10 (10
mM), and (c) a mixture of 10 (5 mM) and its antipode (5 mM), where (S)-(–)-1,1'-bi-2-naphthol (250
mM) was added as a chiral NMR shift reagent.
S4

ACCEPTED MANUSCRIPT
(iv) Determination of the diastereomeric purity of 11/2 and 13/2
(a)
(b)
–78.5
–78.6
–78.7
–78.8
–78.9
–79.0
–79.1
–79.2
19F (ppm)
Figure S5. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 11/2 (10 mM) and (b) an authentic mixture
of 11/2 and its diastereoisomer (10 mM in total, prepared from racemic
N-benzyloxycarbonyl-β-homoalanine and 2).
(a)
(b)
–78.6
–78.7
–78.8
–78.9
–79.0
–79.1
–79.2
–79.3
19F (ppm)
Figure S6. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 13/2 (10 mM) and (b) an authentic mixture
of 13/2 and its diastereoisomer (10 mM in total, prepared from racemic N-benzyloxycarbonyl-alanine
and 2).
S5

ACCEPTED MANUSCRIPT
(v) 19F NMR spectra of 11/2, 12/2’, 13/2, 14/2, and 15/2’
(a)
(b)
(c)
(d)
(e)
–78.2 –78.3 –78.4 –78.5 –78.6 –78.7 –78.8 –78.9 –79.0 –79.1 –79.2 –79.3
19F (ppm)
Figure S7. Partial 19F NMR spectra in CDCl3 at 25 °C of dipeptides (10 mM); (a) 11/2, (b) 12/2’, (c) 13/2,
(d) 14/2, and (e) 15/2’.
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ACCEPTED MANUSCRIPT
(vi) Determination of the diastereomeric purity of 4/16 and 4/18
(a)
(b)
–78.5
–78.6
–78.7
–78.8
–78.9
–79.0
–79.1
19F (ppm)
Figure S8. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 4/16 (10 mM) and (b) an authentic mixture
of 4/16 and its diastereoisomer (10 mM in total, prepared from 16 and racemic
3-benzyloxylcarbonylamino-4,4,4,-trifluorobutylic acid).
(a)
(b)
–78.5
–78.6
–78.7
–78.8
–78.9
–79.0
–79.1
19F (ppm)
Figure S9. Partial 19F NMR spectra in CDCl3 at 25 °C of (a) 4/18 (10 mM) and (b) an authentic mixture
of 4/18 and its diastereoisomer (10 mM in total, prepared from 18 and racemic
3-benzyloxylcarbonylamino-4,4,4,-trifluorobutylic acid).
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ACCEPTED MANUSCRIPT
(vii) 19F NMR spectra of 4/16, 6/17, 4/18, 6/19, and 6/20
(a)
(b)
(c)
(d)
(e)
–78.2 –78.3 –78.4 –78.5 –78.6 –78.7 –78.8 –78.9 –79.0 –79.1 –79.2 –79.3
19F (ppm)
Figure S10. Partial 19F NMR spectra in CDCl3 at 25 °C of dipeptides (10 mM); (a) 4/16, (b) 6/17, (d)
4/18, (d) 6/19, and (e) 6/20.
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