Enantioselective fluorescence recognition of chiral amines by n-acyl-(s)-1-naphthylalanyl-(s)-phenylglycine and n-acyl-(s)-1-naphthylalanyl- (s)-1-naphthylalanine dipeptides bridged by a 1,2-phenylene or an ethylene spacer chain

Article abstract of DOI:10.3987/COM-13-12694

This study sought to support the development of enantioselective dipeptide fluorescence sensors by examining the ability of the N-acyl-(S)-1- naphthylalanyl-(S)-phenylglycine and N-acyl-(S)-1-naphthylalanyl-(S)-1- naphthylalanine dipeptides (bridged by the 1,2-phenylene or ethylene spacer chain) to discriminate between aliphatic amine-derived (S)- and (R)-enantiomers. The results indicated that both hydrogen-bonding interactions in the ground state and charge-transfer interactions in the excited state are involved in the fluorescence quenching of these bridged dipeptides by chiral amines to produce the upward curving Stern-Volmer plots. In addition, the numerical analysis of these plots led to the conclusion that the bridged naphthylalanylphenylglycine dipeptide shows the highest ability to differentiate the (S)-enantiomer from (R)-enantiomer when the emission of this dipeptide is quenched by chiral alaninols.

Full text of DOI:10.3987/COM-13-12694

HETEROCYCLES, Vol. 87, No. 5, 2013, pp. 1039 - 1058. © 2013 The Japan Institute of Heterocyclic Chemistry
Received, 6th March, 2013, Accepted, 4th April, 2013, Published online, 11th April, 2013
DOI: 10.3987/COM-13-12694
ENANTIOSELECTIVE FLUORESCENCE RECOGNITION OF CHIRAL
AMINES
BY
N-ACYL-(S)-1-NAPHTHYLALANYL-(S)-
PHENYLGLYCINE AND N-ACYL-(S)-1-NAPHTHYLALANYL-(S)-1-
NAPHTHYLALANINE DIPEPTIDES BRIDGED BY A 1,2-PHENYLENE
OR AN ETHYLENE SPACER CHAIN
Tomoe Ishikawa, Miho Sonobe, Akinori Hayakawa, Tetsutaro Igarashi,
and Tadamitsu Sakurai*
Department of Material and Life Chemistry, Faculty of Engineering, Kanagawa
University, Kanagawa-ku, Yokohama 221-8686, Japan
Abstract – This study sought to support the development of enantioselective
dipeptide fluorescence sensors by examining the ability of the N-acyl-(S)-1-
naphthylalanyl-(S)-phenylglycine
and
N-acyl-(S)-1-naphthylalanyl-(S)-1-
naphthylalanine dipeptides (bridged by the 1,2-phenylene or ethylene spacer
chain) to discriminate between aliphatic amine-derived (S)- and (R)-
enantiomers. The results indicated that both hydrogen-bonding interactions in
the ground state and charge-transfer interactions in the excited state are involved
in the fluorescence quenching of these bridged dipeptides by chiral amines
to produce the upward curving Stern–Volmer plots. In addition, the numerical
analysis of these plots led to the conclusion that the bridged
naphthylalanylphenylglycine dipeptide shows the highest ability to differentiate
the (S)-enantiomer from (R)-enantiomer when the emission of this dipeptide is
quenched by chiral alaninols.
INTRODUCTION
The intramolecular and intermolecular interactions of fluorescent host molecules (fluorophores) with
guest molecules in the ground and excited states are directly reflected in the emission intensities of the
former. Therefore, photophysical studies of these interactions have continued to contribute to the
1
development of various photofunctional host molecules. Recently, significant attention has been
directed toward the enantioselective recognition of chiral biomolecules by a wide variety of

1
b
fluorophores.
A survey of recent literature published showed that in order to enhance the ability of
these fluorophores to discriminate enantiomers of chiral guests, a number of molecules including 1,1´-
2
3
binaphthols and related derivatives, cyclohexane-based thioureas, various chromophore-substituted
4
5
6
macrocycles, 1,8-diacridyl naphthalene derivatives, !-amino acid-substituted calix[4]arenes, and
7
ingeniously-designed supramolecules have been developed as rigid and sophisticated chiral fluorophores.
However, only a few studies have been conducted into the enantioselective recognition of chiral amines
1
b,4e
(guests) by fluorophore-substituted peptides (hosts).
In a previous study we discovered that the fluorescence quenching of a (R,S)-N-benzoyl-1-
naphthylalanylphenylglycinamide derivative by chiral pyrrolidine-2-methanol (PMO) in 1,4-dioxane
8
proceeds via a singlet exciplex intermediate and proceeds enantioselectively. Despite the simple
acyclic structure of this dipeptide fluorophore, it showed a relatively large enantioselectivity in the
emission quenching by PMO (quenching constant ratio: KSV [(S)-PMO]/KSV [(R)-PMO] = 1.8). The
effects of the N-acyl substituent and peptide conformation on the stability of the exciplex are believed
to play major roles in discriminating the PMO-derived enantiomers. An inspection of the enantiomer
recognition mechanisms proposed so far reveals the importance of the charge-transfer and hydrogen-
bonding interactions and conformational rigidity in host–guest complexes as major factors controlling the
2
–7,9
enantioselectivity for chiral guest molecules.
Thus, it is expected that the enhancement of this
interaction and rigidity contributes to the increased difference in stability between the (R)- and (S)-
enantiomer-derived host–guest complexes.
The above considerations led us to assume that the connection between the two dipeptides previously
examined results in an enhanced conformational rigidity and creates a site of interaction with chiral
amines. To develop such a bridged dipeptide fluorophore and increase the enantioselectivity of this
dipeptide, we chose ethylene and 1,2-phenylene groups as bridging spacer chains and synthesized the
bridged dipeptides 1a, 1b, and 1c (Chart 1). For comparison, we synthesized 1d and 1e as reference
dipeptides containing no bridging spacer. In this paper, we report the bridging effects on the
enantioselective fluorescence quenching of (S)-1-naphthylalanine-derived dipeptides by the acyclic chiral
amines, N,N-dimethyl-1-phenylethylamine (PEA) and alaninol (ALO), and heterocyclic chiral amines,
PMO and 2-methylpiperazine (MPZ).
RESULTS AND DISCUSSION
(S)-3-(1-Naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester as a precursor of the bridged dipeptides 1a
and 1b and reference dipeptide 1d was prepared according to the following reaction pathway: the
benzyloxycarbonylation of the amino group in (S)-3-(1-naphthyl)alanine hydrochloride in
sodium hydroxide solution, the condensation reaction of N-benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine

O
O
O
H
N
H
N
H
N
HN
HN
OBu-t
OBu-t
HN
HN
OBu-t
OBu-t
HN
HN
NHBu-t
O
O
Ph
Ph
O
O
Ph
Ph
O
O
O
O
O
O
O
O
Nap-1
Nap-1
NHBu-t
N
H
N
H
N
H
O
O
O
1-Nap = 1-naphthyl
1
a
1b
1c
Me
NMe2
OH
O
Me
HN
O
Nap-1
NHBu-t
O
Ph
O
N
H
HN
OBu-t
N
H
N
H
PMO
O
O
PEA
NH2
H
N
OH
1
d
1e
Me
N
H
Me
ALO
MPZ
Chart 1
with (S)-2-phenylglycine tert-butyl ester in 1,4-dioxane containing N-hydroxysuccinimide and water-
soluble carbodiimide, and the removal of the benzyloxycarbonyl group from N-acyl-(S)-3-(1-
naphthyl)alanyl-(S)-2-phenylglycine
dipeptide
by
catalytic
hydrogenation
in
methanol-
dimethylformamide. The acylation of the deprotected 3-(1-naphthyl)alanyl-2-phenylglycine dipeptide
with phthaloyl chloride, succinyl chloride, and benzoyl chloride in 1,4-dioxane containing pyridine led
to the successful formation of 1a, 1b, and 1d, respectively.
On the other hand, (S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide as a precursor of
the bridged dipeptide 1c and reference dipeptide 1e was prepared by a condensation reaction between (S)-
3
-(1-naphthyl)alanine tert-butylamide and N-(tert-butoxycarbonyl)-(S)-3-(1-naphthyl)alanine in 1,4-
dioxane containing water-soluble carbodiimide, followed by the deprotection of the resulting dipeptide in
acidic methanol solution. The former deprotected 3-(1-naphthyl)alaninamide was synthesized by the
amidation of N-benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine with tert-butylamine and the subsequent
catalytic hydrogenation of the resulting 1-naphthylalaninamide derivative. In addition, the latter N-
protected (S)-3-(1-naphthyl)alanine was obtained from the reaction of (S)-3-(1-naphthyl)alanine with tert-
butoxycarbonyl chloride in aqueous sodium hydroxide. The above precursor was acylated under weakly
basic conditions by using succinyl chloride and acetyl chloride to afford the bridged dipeptide 1c and
1
reference dipeptide 1e, respectively. The H NMR spectral analysis of 1a–c revealed that these bridged

dipeptides contain only a single diastereomer and that no epimerization at the chiral methine carbon takes
place during the protection, deprotection, and peptide bond-forming reactions described above.
3500
3000
2500
2000
1500
1000
–3
(
S)-PEA (mol dm )
0.00
0.01
0.02
0.03
0.04
0.05
500
0
3
00
350
400
450
500
550
600
Wavelength (nm)
–
4
–3
Figure 1. Fluorescence spectra of 1a (1.25 " 10 mol dm ) in argon-saturated 1,4-dioxane containing
various amounts of (S)-PEA at room temperature. The excitation wavelength was 295 nm.
4.5
4.0
3.5
3.0
2.5
2.0
1.5
1.0
1
b/(S)-PEA
1b/(R)-PEA
1d/(S)-PEA
a/(S)-PEA
d/(R)-PEA
1
1
1
a/(R)-PEA
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
–3
[
PEA] (mol dm )
–
4
–3
Figure 2. Stern–Volmer plots for the fluorescence quenching of 1a (1.25 " 10 mol dm ;ꢀꢁꢂ), 1b
–
4
–3
–4
–3
(
1.25 " 10 mol dm ;ꢃꢁꢄ), and 1d (2.5 " 10 mol dm ;ꢅꢁꢆ) by (S)-PEA (ꢀꢁꢃꢁꢅ) and (R)-PEA
ꢂꢁꢄꢁꢆ) in argon-saturated 1,4-dioxane. The excitation wavelength was 295 nm.
(
As typically shown in Figure 1, the 340-nm fluorescence of 1a was relatively efficiently quenched in the

presence of PEA in argon-saturated 1,4-dioxane with the appearance of a new fluorescence band at
approximately 460 nm. Because tertiary alkylamines are well known to form singlet exciplexes with
1
0
many naphthalene derivatives and also an isoemissive point was observed when the 460-nm emission
appeared, this new emission in the nonpolar solvent, 1,4-dioxane, can be attributed to singlet exciplex
fluorescence. Interestingly, the Stern–Volmer plots for the above fluorescence quenching are upward
2
curving, as shown in Figure 2, and can be approximated by the equation: I
where K and K are constants, and I and I
PEA, respectively), suggesting the contribution of both dynamic and static quenching processes.
0 1 2
/I = 1 + K [PEA] + K [PEA]
(
1
2
0
represent the fluorescence intensities of 1 with and without
5
a,5b,11
While a charge-transfer interaction between the excited-state 1a and ground-state PEA is responsible for
the former quenching, the latter quenching may be caused by an intermolecular hydrogen-bonding
interaction between the fluorophore and quencher in the ground state. In addition, we previously
revealed that tertiary alkylamine forms a hydrogen bond with the amide proton in N-acetyl-1-
1
2
naphthylalaninamide derivative to catalyze the hydrogen–deuterium exchange of this derivative.
Thus,
the hydrogen bond is considered to be formed between the PEA nitrogen and amide proton existing on
the amino side of the 1-naphthylalanine moiety. Almost the same upward curving Stern–Volmer plots
as those for 1a were obtained for the bridged dipeptide 1b and reference dipeptide 1d (Figure 2).
2
1.8
1.6
1.4
1.2
1
1
a
d
1
1b
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
PEA] (mol dm )
S
R
r r
Figure 3. Plots of the (I – 1)/(I – 1) ratios against PEA concentration for the fluorescence quenching
–
4
–3
–4
–3
–4
–3
of 1a (1.25 " 10 mol dm ), 1b (1.25 " 10 mol dm ), and 1d (2.5 " 10 mol dm ) by (S)-PEA and
(R)-PEA in argon-saturated 1,4-dioxane at room temperature. The excitation wavelength was 295 nm.
To estimate and compare the ability of 1a, 1b, and 1d to discriminate between the (S)- and (R)-

S
R
enantiomers of chiral PEA, the ratio of (I
r
– 1) for the (S)-enantiomer to (I
– 1), was used as a measure of this ability, where I = I /I. If the dynamic quenching is
predominant under given conditions, this ratio is estimated to be equal to the ratio of quenching constants,
r
– 1) for the (R)-enantiomer,
S
R
(I
r
– 1)/(I
r
r
0
S
R
S
S
R
R
K
SV /KSV , on the basis of the equations, I
r
= 1 + KSV [(S)-PEA] and I
r
= 1 + KSV [(R)-PEA]. An
S
R
inspection of the plots of (I
r
– 1)/(I
r
– 1) against [PEA], depicted in Figure 3, revealed that these ratios
can be regarded as approximately constant in any plots irrespective of the PEA concentration and give a
greater value for the 1a/PEA system (1.5) than for the 1d/PEA system (1.3). If the static quenching
takes place in competition with the dynamic quenching, the former result indicates these two quenching
processes to have an enantiodifferentiating ability comparable with each other. In addition, the latter
result that the ability to differentiate (S)-PEA and (R)-PEA is somewhat enhanced by bridging the two 1-
naphthylalanylphenylglycine dipeptides with the rigid 1,2-phenylene group led us to propose that owing
to the rigid bridging group, the charge-transfer and hydrogen-bonding interactions between the (R)-
enantiomer and naphthylalanine chromophore are subject to greater steric hindrance by the other
naphthylalanine. This interpretation is supported by the finding that the introduction of the flexible
ethylene spacer instead of the rigid 1,2-phenylene spacer lowers the enantiodiscriminating ability; in other
S
R
r r
words, the (I – 1)/(I – 1) ratio from 1.5 (1a) to 1.2 (1b) in the fluorescence quenching by chiral PEA.
O
O
O
O
O
2
h!
O
R
NH HN
O
R
PEA
O
R
NH HN
O
1b
Deactivation of Ar*
R
Ar*
Ar*
Ar*
(Ar/PEA)*
O
O
O
–
PEA
O
R
NH HN
O
R
O
R
NH HN
O
Deactivation of Ar*
R
(
Ar/PEA)*
Ar
Ar
Ar
O
H
N
R =
Ar =
OBu-t
Ph
Scheme 1
S
R
Interestingly, a contrasting bridging effect was observed between the (I
r
– 1)/(I
r
0
– 1) ratio and I /I ratio,
as can be seen from comparison of the Stern–Volmer plots for the fluorescence quenching of 1a, 1b, and
d by PEA (Figure 2). For example, the magnitude of the latter emission intensity ratio for 1d (1.8 at
1
–
3
[
(S)-PEA] = 0.05 mol dm ) was almost uninfluenced by bridging the naphthylalanylphenylglycine
dipeptide with the 1,2-phenylene group (1a, I /I = 1.8), whereas the ethylene bridge (1b) enhanced this
intensity ratio by a factor of approximately 2 (I /I = 4.1). If it is possible for one PEA molecule in the
0
0

0
ground state to deactivate the two naphthylalanine chromophores of 1 in the excited state, the I /I ratio is
expected to be increased by a factor of 2. Thus, the latter observation is consistent with the quenching
mechanism illustrated in Scheme 1, in which the deactivation of the exciplex formed between PEA and
one of the two naphthalene rings in the excited-state 1b is accompanied by the generation of the exciplex
derived from the other excited-state naphthalene ring and the chiral amine released by this
deactivation. The flexible ethylene bridge in 1b is considered to play a decisive role in the above
quenching mechanism.
2.0
1.8
1.6
1.4
1.2
1.0
0.8
MPZ
PMO
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
Chiral Amine] (mol dm )
S
R
Figure 4. Plots of the (I
r
– 1)/(I
r
– 1) ratios against chiral amine concentration for the fluorescence
–
4
–3
quenching of 1a (1.25 " 10 mol dm ) by (S)-PMO and (R)-PMO or (S)-MPZ and (R)-MPZ in argon-
saturated 1,4-dioxane at room temperature. The excitation wavelengths were 284 nm for PMO and 295
nm for MPZ.
On the other hand, from an analysis of the upward curving Stern–Volmer plots for the fluorescence
S
R
quenching of 1a by chiral PMO and MPZ, we estimated the (I
r r
– 1)/(I – 1) ratio at 1.0 and 1.2 for the
1
a/PMO and 1a/MPZ systems, respectively, as shown in Figure 4. These results confirmed that the
bridged dipeptide 1a has a lower enantiodiscriminating ability for these heterocyclic chiral amines than
for the acyclic chiral amine PEA. The less upward curving Stern-Volmer plots for the quenching of 1a
by chiral PMO in particular reflect that this secondary amine is a weaker base than the tertiary amine PEA
and hence contributes to the static quenching of 1a to a much lesser extent through the hydrogen-bonding
interaction (data not shown). Because MPZ forms a singlet exciplex with 1a to exhibit fluorescence at
around 405 nm, a charge-transfer interaction between the excited-state 1 and ground-state chiral amine
should also be involved in the emission quenching by chiral PMO. Thus, it is likely that there is a very

small difference in the stabilities of the hydrogen-bonding complexes and singlet exciplexes, formed
between 1a and each enantiomer of PMO having considerably less steric bulk than PEA. The negligible
effect of the 1,2-phenylene bridge on the ability to differentiate between (S)-PMO and (R)-PMO may be
1a
1b
Figure 5. Energy-minimized conformations of the bridged dipeptides 1a and 1b.
attributed to the minor steric effect of one naphthylalanine chromophore in 1a on the hydrogen-bonding
and charge-transfer interactions between the chiral PMO and other naphthylalanine chromophore. We
already demonstrated that the flexible ethylene bridge increased the fluorescence intensity ratio I
which can be utilized as a measure of the quenching efficiency of 1) for the reference dipeptide 1d by a
factor of approximately 2. The same bridging effect as noted in 1a was observed for the fluorescence
0
/I
(
–
3
quenching of 1b by PMO (I
0 0
/I = 1.9 for 1d and 3.2 for 1b at [(S)-PMO] = 0.05 mol dm ) and MPZ (I /I =
–
3
3
.1 for 1d and 7.2 for 1b at [(S)-MPZ] = 0.05 mol dm ). To confirm whether the two naphthalene rings
in 1a and 1b are able to interact with each other in their excited states, the conformational energies of
these two bridged dipeptides were minimized by MM2 and PM5 calculations (Figure 5). Clearly, the
two naphthalene rings in 1b carrying the flexible ethylene group are present in mutual proximity in the
ground state, and hence may easily interact with each other in the excited state, although the rigid 1,2-
phenylene group in 1a inhibits the close approach of these two naphthalene rings.
On the basis of the ability of 1a and 1b to discriminate the (S)- and (R)-enantiomers of PEA (tertiary
amine) and PMO (secondary amine), we predict that these two bridged dipeptides will display a poor
enantiodiscriminating ability for chiral ALO (primary amine) owing to the weaker charge-transfer

1.20
1.15
1.10
1.05
1.00
(
S)-ALO
R)-ALO
(
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
ALO] (mol dm )
–
4
–3
Figure 6. Stern–Volmer plots for the fluorescence quenching of 1a (1.25 " 10 mol dm ) by (S)-ALO
and (R)-ALO in argon-saturated 1,4-dioxane at room temperature. The excitation wavelength was 284
nm.
6
5
4
1b
3
2
1
1
a
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
ALO] (mol dm )
S
–
R
Figure 7. Plots of the (I
r
– 1)/(I
4
r
– 1) ratios against ALO concentration for the fluorescence quenching
–3
of 1a and 1b (1.25 " 10 mol dm ) by (S)-ALO and (R)-ALO in argon-saturated 1,4-dioxane at
room temperature. The excitation wavelengths were 284 nm.
interactions with this primary chiral amine than those with chiral PEA and PMO. Figure 6 illustrates
S
R
the Stern–Volmer plots for the fluorescence quenching of 1a by chiral ALO, and the (I
r r
– 1)/(I – 1)
ratio (used as a measure of the enantiodiscriminating ability of 1) is shown in Figure 7 as a function of the

0
ALO concentration. The small I /I ratios (for example, 1.12 for 1a, 1.25 for 1b, and 1.17 for 1d at [(S)-
–
3
ALO] = 0.05 mol dm ) reflect the inefficiency of the emission quenching by chiral ALO and are
consistent with the weaker charge-transfer interactions of 1 with chiral ALO that we predicted. In
S
R
r r
addition, the unexpected observation of the relatively large (I – 1)/(I – 1) ratio of 3.0 for the 1a/ALO
and 1b/ALO systems demonstrates that these systems display the highest enantiodiscriminating ability
among the 1/chiral amine systems examined (Figure 7). This observation also reveals that the ability
to discriminate (S)-ALO and (R)-ALO is independent of the flexibility or rigidity of the spacer chain
connecting the two naphthylalanylphenylglycine dipeptides. On the other hand, the pronouncedly
curved Stern–Volmer plots (depicted in Figure 6) indicate an increased contribution of the static
S
R
quenching to the total excited-state deactivation of 1 by chiral ALO. If we consider that the (I
r r
– 1)/(I
–
1) ratio of 1.3 for the reference dipeptide 1d is not so large and the Stern–Volmer plots for this
dipeptide is only weakly upward curving, the hydrogen-bonding interaction (that is responsible for the
observed static quenching) between 1a or 1b and a chiral ALO molecule should occur in the
two naphthylalanine chromophores bridged by the 1,2-phenylene or ethylene spacer chain. It is likely
that in addition to the amino hydrogen in ALO, the hydroxy hydrogen is also involved in the above
S
R
interaction, as schematically illustrated in Figure 8. A large difference in the (I
r
– 1)/(I
r
– 1)
ratio between the 1a/ALO and 1d/ALO systems strongly suggests that a chiral ALO molecule (which is
hydrogen bonded to the amide carbonyl oxygen atoms in the naphthylalanine chromophores) plays a
major role in differentiating between (S)-ALO and (R)-ALO; hence, the former enantiomer forms a more
stabilized hydrogen-bonded complex than the latter enantiomer. Unfortunately, it was difficult
to estimate the equilibrium constant for the formation of these hydrogen-bonded complexes because of
very minor changes in the ultraviolet (UV) absorption spectra of 1a and 1b (measured in the presence of
chiral ALO), in the wavelength range where the absorption of this amine is negligible.
Ar
R
O
HN
H
N
R =
O
O
O
OBu-t
O
O
H
H
Ph
N
H
Me
HN
Ar
R
Ar =
Figure 8. Schematic illustration for the hydrogen-bonding interaction between the amide carbonyl
oxygen atoms in 1a and the hydroxy and amino hydrogen atoms in (S)-ALO.

On the basis of the considerations described above, we predicted that the ability of the bridged dipeptide 1
to discriminate (S)-ALO from (R)-ALO is enhanced with increasing number of the 1-naphthylalanine
S
R
r r
residue in this dipeptide. To verify whether our prediction is correct, the (I – 1)/(I – 1) ratios for the
bridged (S)-1-naphthylalanyl-(S)-1-naphthylalanine dipeptide 1c and its reference dipeptide 1e were
evaluated from the analysis of the Stern–Volmer plots for the fluorescence quenching of these dipeptides
by chiral ALO and compared with that for 1b. As shown in Figure 9, the Stern–Volmer plots obtained
1.3
1.2
1.1
1.0
(S)-ALO
(R)-ALO
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
ALO] (mol dm )
–
4
–3
Figure 9. Stern–Volmer plots for the fluorescence quenching of 1c (1.25 " 10 mol dm ) by (S)-ALO
and (R)-ALO in argon-saturated 1,4-dioxane at room temperature. The excitation wavelength was 284
nm.
were much more weakly upward curved as compared with those for 1b. This finding indicates a
decreased contribution of the static quenching of 1c by ALO and compatibility with the minor
participation of the hydrogen bonds (between an ALO molecule and the amide carbonyl oxygen atoms of
the naphthylalanine chromophores) depicted in Figure 8. The presence of the four bulky naphthyl-
alanine chromophores in 1c is assumed to inhibit the formation of such hydrogen bonds, as can be
inferred from the energy-minimized conformation of this bridged dipeptide (Figure 10). This
assumption is supported by the fact that the ability of the bridged naphthylalanylnaphthylalanine
S
R
dipeptide 1c to discriminate the ALO enantiomers [(I
r
– 1)/(I
– 1) = 2.0, see Figure 11]. In addition, a comparison
– 1) ratio for 1c (2.0) with that for 1b (3.0) confirms that contrary to our prediction, an
increase in the number of the naphthylalanine chromophore (2 # 4) lowers the enantiodiscriminating
r
– 1) = 2.0, see Figure 11] is comparable
S
R
r r
with that of the reference dipeptide 1e [(I – 1)/(I
S
R
r r
of the (I – 1)/(I

1c
Figure 10. Energy-minimized conformations of the bridged dipeptide 1c.
6
5
4
3
1
e
2
1
1c
0
0.01
0.02
0.03
0.04
0.05
0.06
–
3
[
ALO] (mol dm )
S
R
Figure 11. Plots of the (I
r
– 1)/(I
r
– 1) ratios against ALO concentration for the fluorescence
–
4
–3
–4
–3
quenching of 1c (1.25 " 10 mol dm ) and 1e (2.5 " 10 mol dm ) by (S)-ALO and (R)-ALO in argon-
saturated 1,4-dioxane at room temperature. The excitation wavelengths were 284 nm.
ability. Therefore, these considerations led to the conclusion that the bridged dipeptide 1 bearing the
two naphthylalanine chromophores displays the highest ability to differentiate (S)-ALO from (R)-ALO.
EXPERIMENTAL

General methods
UV absorption and fluorescence spectra were recorded at room temperature on a Shimadzu UV-3150
spectrophotometer and Hitachi F-4500 spectrofluorometer, respectively.
A cell with a 10-
1
13
mm pathlength was used.
H and C NMR spectra were recorded on a JEOL JNM-ECA500
spectrometer. Chemical shifts were determined using tetramethylsilane as an internal standard.
Electrospray ionization time-of-flight (ESI-TOF) mass spectra were recorded using a JEOL JMS-T100LC
AccuTOF mass spectrometer. Matrix-assisted laser desorption/ionization (MALDI)-TOF mass spectra
were recorded on a Shimadzu/Kratos TOF mass spectrometer. Elemental analyses were performed on a
Perkin-Elmer PE2400 series II CHNS/O analyzer. MM2 and PM5 calculations were accomplished
using CAChe 5.0 for Windows available from Fujitsu Ltd (2002). Spectrophotometric grade 1,4-
dioxane was used without further purification. All other chemicals were obtained from commercial
sources at the highest grade available.
N-Benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine
–
3
(
S)-3-(1-Naphthyl)alanine hydrochloride (2.01 g, 8.0 mmol) was dissolved in 2 mol dm aqueous
sodium hydroxide (8.0 mL, 16 mmol) and the resulting solution was cooled on ice. Benzyloxycarbonyl
–
3
chloride (2.05 g, 12 mmol) and 2 mol dm aqueous sodium hydroxide (4.0 mL, 8.0 mmol) were
successively added to this solution. The reaction mixture was stirred for 1 h at room temperature and its
–
3
pH was adjusted to 2 with 2 mol dm hydrochloric acid. The resulting precipitates were collected by
suction filtration and recrystallized from aqueous EtOH to afford the title compound as a colorless crystal
1
(
2.71 g, 97%). H NMR (500 MHz, DMSO-d
6
) ! = 3.22 (1H, dd, J = 10.3, 14.3 Hz), 3.64 (1H, dd, J =
4
7
7
7
5
1
.0, 14.3 Hz), 4.31 (1H, ddd, J = 4.0, 8.6, 10.3 Hz), 4.91 (1H, d, J = 12.6 Hz), 4.95 (1H, d, J = 12.6 Hz),
.23 (2H, d, J = 6.9 Hz), 7.29 (1H, dd, J = 6.9, 7.4 Hz), 7.33 (2H, dd, J = 7.4, 7.4 Hz), 7.40–7.44 (2H, m),
.53 (1H, dd, J = 6.9, 8.0 Hz), 7.60 (1H, dd, J = 6.9, 8.6 Hz), 7.78 (1H, d, J = 8.6 Hz), 7.82 (1H, d, J =
1
3
6
.4 Hz), 7.94 (1H, d, J = 8.0 Hz), 8.12 (1H, d, J = 8.6 Hz); C NMR (125 MHz, DMSO-d ) ! = 33.9,
4.8, 65.2, 123.1, 125.3, 125.6, 126.3, 127.2, 127.4 (2C), 127.5, 127.7, 128.3 (2C), 128.8, 131.3, 133.4,
33.7, 136.9, 156.0, 173.4.
N-Benzyloxycarbonyl-(S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester
N-Benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine (2.01 g, 5.7 mmol) and N-hydroxysuccinimide (0.79 g,
6
.9 mmol) were dissolved in 1,4-dioxane (20 mL). N-Ethyl-N´-(3-dimethylaminopropyl)carbodiimide
hydrochloride (1.65 g, 8.6 mmol) was added to this dioxane solution with stirring at 0 °C. The reaction
mixture was stirred for 2 h at room temperature and (S)-2-phenylglycine tert-butyl ester (1.42 g, 6.9
mmol) was added. The resulting mixture was stirred for an additional 1 h at room temperature. After

the evaporation of the solvent, the residual solid was dissolved in EtOAc (30 mL). The EtOAc solution
was washed with 10% aqueous citric acid and water and dried over magnesium sulfate. The removal of
the solvent gave a crystalline solid, which was purified by column chromatography on silica gel (70–230
1
mesh, EtOAc/hexane = 2/1 v/v) to afford the title compound (2.32 g, 75%). H NMR (500 MHz,
DMSO-d
6
) ! = 1.36 (9H, s), 3.21 (1H, dd, J = 10.9, 14.3 Hz), 3.54 (1H, dd, J = 4.0, 14.3 Hz), 4.60 (1H,
ddd, J = 4.0, 9.2, 10.9 Hz), 4.87 (1H, d, J = 12.6 Hz), 4.91 (1H, d, J = 12.6 Hz), 5.31 (1H, d, J = 6.9 Hz),
7.18 (2H, d, J = 6.3 Hz), 7.30 (2H, dd, J = 6.3, 9.2 Hz), 7.31 (1H, dd, J = 9.2, 9.2 Hz), 7.36–7.43 (6H, m),
7.48 (1H, d, J = 6.9 Hz), 7.52 (1H, dd, J = 6.9, 7.4 Hz), 7.56 (1H, dd, J = 6.9, 8.0 Hz), 7.64 (1H, d, J =
9.2 Hz), 7.81 (1H, d, J = 8.0 Hz), 7.93 (1H, d, J = 7.4 Hz), 8.24 (1H, d, J = 8.6 Hz), 8.83 (1H, d, J = 6.9
1
3
Hz); C NMR (125 MHz, DMSO-d
6
) ! = 27.5 (3C), 34.8, 54.8, 57.2, 65.2, 81.2, 123.8, 125.3, 125.5,
26.0, 127.0, 127.3 (2C), 127.5, 127.6, 127.7 (2C), 128.2, 128.3 (2C), 128.5, 128.6 (2C), 131.6, 133.3,
33.5, 136.4, 136.9, 155.8, 169.3, 171.5.
1
1
(S)-3-(1-Naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester
To a solution of N-benzyloxycarbonyl-(S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester (1.08
g, 2.0 mmol) in argon-saturated MeOH/DMF (3/1 v/v, 8 mL), 10% palladium on carbon (0.10 g) was
added. The mixture was stirred for 48 h under hydrogen atmosphere at room temperature. After the
catalyst had been removed by filtration, the filtrate was evaporated to dryness under reduced
pressure. The resulting crystalline solid was subjected to column chromatography on silica gel (70–230
1
mesh, EtOAc/hexane = 2/1 v/v) to afford the title compound (0.73 g, 90%). H NMR (500 MHz,
DMSO-d
6
) ! = 1.35 (9H, s), 1.85 (2H, s), 2.92 (1H, dd, J = 8.6, 13.7 Hz), 3.56 (1H, dd, J = 4.6, 13.7 Hz),
.62 (1H, dd, J = 4.6, 8.6 Hz), 5.30 (1H, d, J = 6.9 Hz), 7.35–7.42 (7H, m), 7.51 (1H, dd, J = 6.9, 8.0 Hz),
.54 (1H, dd, J = 6.9, 8.0 Hz), 7.79 (1H, d, J = 6.9 Hz), 7.92 (1H, d, J = 8.0 Hz), 8.16 (1H, d, J = 8.6 Hz),
3
7
8
1
1
1
3
6
.67 (1H, d, J = 6.9 Hz); C NMR (125 MHz, DMSO-d ) ! = 27.5 (3C), 38.1, 55.4, 56.6, 81.4, 123.8,
25.4, 125.5, 125.9, 126.8, 127.2 (2C), 127.6, 128.1, 128.5, 128.6 (2C), 131.7, 133.5, 134.8, 137.0, 168.5,
74.3.
N,N´-(1,2-Phenylenedicarbonyl)-bis[(S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester]
1a)
(
To a solution of (S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester (0.67 g, 1.7 mmol) and
pyridine (0.11 g, 1.3 mmol) in 1,4-dioxane (10 mL), phthaloyl chloride (0.14 g, 0.66 mmol) was slowly
added with stirring at 0 °C. The reaction mixture was stirred for 72 h at room temperature. After the
solvent had been removed in vacuo, the residual solid was dissolved in EtOAc (20 mL). The EtOAc
solution was washed with water and dried over magnesium sulfate. The removal of EtOAc afforded a

crystalline solid, which was purified by column chromatography on silica gel (70–230 mesh,
1
EtOAc/hexane = 2/1 v/v) to yield the title compound as a colorless crystal (0.45 g, 72%). H NMR (500
MHz, DMSO-d
6
) ! = 1.31 (18H, s), 3.34 (2H, dd, J = 9.2, 14.3 Hz), 3.74 (2H, dd, J = 4.6, 14.3 Hz), 4.90
(2H, ddd, J = 4.6, 9.2, 8.6 Hz), 5.28 (2H, d, J = 6.9 Hz), 7.13 (2H, dd, J = 3.4, 6.0 Hz), 7.24 (4H, d, J =
4
.0 Hz), 7.26 (2H, dd, J = 5.1, 5.1 Hz), 7.35 (4H, dd, J = 4.0, 5.1 Hz), 7.38 (2H, dd, J = 3.4, 5.7 Hz), 7.40
(2H, d, J = 8.0 Hz), 7.44 (2H, d, J = 6.0 Hz), 7.48–7.51 (4H, m), 7.76 (2H, d, J = 8.0 Hz), 7.89 (2H, d, J =
1
3
7
.4 Hz), 8.18 (2H, d, J = 6.9 Hz), 8.40 (2H, d, J = 6.9 Hz), 8.52 (2H, d, J = 8.6 Hz); C NMR (125 MHz,
) ! = 27.5 (6C), 53.4 (2C), 57.3 (2C), 66.3 (2C), 81.0 (2C), 123.6 (2C), 125.3 (2C), 125.5 (2C),
26.1 (2C), 127.0 (2C), 127.3 (2C), 127.5 (2C), 127.7 (4C), 127.9 (2C), 128.4 (4C), 128.6 (2C), 129.4
DMSO-d
6
1
(2C), 131.6 (2C), 133.4 (2C), 134.1 (2C), 135.6 (2C), 136.5 (2C), 167.7 (2C), 169.0 (2C), 170.6 (2C);
+
ESI-TOF-MS: m/z calcd for C58
H
58
N
4
NaO
8
[M + Na] , 961.413; Found: 961.415. Anal. Calcd for
58 58 4 8
C H N O : C, 74.18; H, 6.23; N, 5.97; Found: C, 74.18; H, 6.02; N, 5.94.
N,N´-Ethylenedicarbonyl-bis[(S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester] (1b)
Following the above protocol, (S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester (0.32 g, 0.79
mmol), pyridine (0.056 g, 0.71 mmol), and succinyl chloride (0.055 g, 0.36 mmol) in 1,4-dioxane (4 mL)
1
yielded the title compound as a colorless crystal (0.22 g, 70%). H NMR (500 MHz, DMSO-d
6
) ! =
1
.34 (18H, s), 1.97–2.02 (2H, m), 2.09–2.15 (2H, m), 3.16 (2H, dd, J = 9.2, 14.3 Hz), 3.52 (2H, dd, J =
.6, 14.3 Hz), 4.80 (2H, ddd, J = 4.6, 8.6, 9.2 Hz), 5.27 (2H, d, J = 6.9 Hz), 7.32–7.41 (14H, m), 7.50 (2H,
4
dd, J = 6.9, 8.6 Hz), 7.54 (2H, dd, J = 6.9, 8.0 Hz), 7.77 (2H, d, J = 8.6 Hz), 7.90 (2H, d, J = 8.0 Hz), 8.16
1
3
(
2H, d, J = 8.6 Hz), 8.22 (2H, d, J = 8.6 Hz), 8.79 (2H, d, J = 6.9 Hz); C NMR (125 MHz, DMSO-d
27.5 (6C), 30.5 (2C), 34.8 (2C), 52.7 (2C), 57.1 (2C), 81.2 (2C), 123.9 (2C), 125.2 (2C), 125.5 (2C),
25.9 (2C), 127.0 (2C), 127.4 (2C), 127.7 (4C), 128.1 (2C), 128.4 (2C), 128.5 (4C), 131.7 (2C), 133.3
6
) !
=
1
(2C), 133.5 (2C), 136.4 (2C), 169.2 (2C), 171.1 (2C), 171.2 (2C); ESI-TOF-MS: m/z calcd for
+
C
54
H
58
N
4
NaO
8
[M + Na] , 913.413; Found, 913.415. Anal. Calcd for C54
H
58
N
4
O
8
: C, 72.79; H, 6.56; N,
6
.29; Found: C, 72.80; H, 6.28; N, 6.16.
N-Benzoyl-(S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester (1d)
Following the above protocol, (S)-3-(1-naphthyl)alanyl-(S)-2-phenylglycine tert-butyl ester (0.25 g, 0.61
mmol), pyridine (0.050 g, 0.67 mmol), and benzoyl chloride (0.090 g, 0.67 mmol) in 1,4-dioxane (3 mL)
1
yielded the title compound as a colorless crystal (0.30 g, 97%). H NMR (500 MHz, DMSO-d
6
) ! =
1
.36 (9H, s), 3.45 (1H, dd, J = 10.9, 14.3 Hz), 3.66 (1H, dd, J = 4.0, 14.3 Hz), 5.05 (1H, ddd, J = 4.0, 8.6,
0.9 Hz), 5.33 (1H, d, J = 6.9 Hz), 7.37–7.45 (8H, m), 7.49 (1H, dd, J = 7.4, 8.0 Hz), 7.53 (1H, d, J = 8.0
1
Hz), 7.59 (2H, dd, J = 6.9, 8.6 Hz), 7.72 (2H, d, J = 8.6 Hz), 7.77 (1H, d, J = 8.0 Hz), 7.90 (1H, d, J = 7.4

1
3
Hz), 8.34 (1H, d, J = 8.6 Hz), 8.69 (1H, d, J = 8.6 Hz), 8.94 (1H, d, J = 6.9 Hz); C NMR (125 MHz,
DMSO-d ) ! = 27.5 (3C), 34.4, 53.4, 57.2, 81.2, 123.9, 125.3, 125.6, 126.1, 127.1, 127.3 (2C), 127.5,
27.8 (2C), 128.2 (3C), 128.5, 128.6 (2C), 131.3, 131.6, 133.3, 133.8, 133.9, 136.4, 166.3, 169.4, 171.4.
Anal. Calcd for C H N O : C, 75.57; H, 6.34; N, 5.51; Found: C, 75.18; H, 6.21; N, 5.42.
6
1
3
2
32
2
4
N-Benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine tert-butylamide
N-Benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine (1.39 g, 4.0 mmol) and N-hydroxysuccinimide (0.69 g,
6
.0 mmol) were dissolved in 1,4-dioxane (20 mL). To this dioxane solution, a solution of N-ethyl-N´-
(3-dimethylaminopropyl)carbodiimide hydrochloride (1.15 g, 6.0 mmol) in DMF (5 mL) was added
dropwise with stirring at 0 °C. The reaction mixture was stirred for 2 h at room temperature and tert-
butylamine (0.58 g, 8.0 mmol) was added. After the resulting solution had been stirred for an additional
3
h at the same temperature, it was poured into water (100 mL). The resulting precipitate was collected
by suction filtration and recrystallized from aqueous EtOH to afford the title compound as a colorless
1
crystal (1.38 g, 86%). H NMR (500 MHz, DMSO-d
6
) ! = 1.20 (9H, s), 3.20 (1H, dd, J = 8.5, 13.7 Hz),
3
7
=
.41 (1H, dd, J = 5.7, 13.7 Hz), 4.36 (1H, ddd, J = 5.7, 7.4, 8.5 Hz), 4.94 (2H, s), 7.22 (2H, d, J = 6.8 Hz),
.27–7.33 (3H, m), 7.36–7.43 (2H, m), 7.36–7.43 (2H, m), 7.51 (1H, dd, J = 6.8, 8.0 Hz), 7.56 (1H, dd, J
1
3
7.4, 8.0 Hz), 7.80 (1H, dd, J = 5.7, 6.8 Hz), 7.92 (1H, d, J = 8.0 Hz), 8.20 (1H, d, J = 8.5 Hz); C NMR
) ! = 28.3 (3C), 35.2, 50.2, 55.7, 65.1, 122.8 (2C), 123.9, 125.2, 125.4, 125.9, 126.9,
27.3 (2C), 127.5, 127.6, 128.4, 131.7, 133.3, 133.7, 137.0, 155.6, 170.4.
(125 MHz, DMSO-d
6
1
(S)-3-(1-Naphthyl)alanine tert-butylamide
To a solution of N-benzyloxycarbonyl-(S)-3-(1-naphthyl)alanine tert-butylamide (0.49 g, 1.2 mmol) in
argon-saturated MeOH (20 mL), 10% palladium on carbon (0.10 g) was added. The mixture was stirred
for 24 h under a hydrogen atmosphere at room temperature. After the catalyst had been removed by
filtration, the filtrate was evaporated to dryness under reduced pressure. The crude product was purified
3
by column chromatography on silica gel (70–230 mesh, CHCl /MeOH = 15/1 v/v) to yield the title
1
compound (0.29 g, 89%). H NMR (500 MHz, DMSO-d
6
) ! = 1.23 (9H, s), 1.78 (2H, s), 2.92 (1H, dd, J
=
8.5, 13.7 Hz), 3.43 (1H, dd, J = 5.1, 8.5 Hz), 3.55 (1H, dd, J = 5.1, 13.7 Hz), 7.39–7.56 (5H, m), 7.78
1
3
(
1H, d, J = 6.8 Hz), 7.90 (1H, d, J = 8.0 Hz), 8.19 (1H, d, J = 8.5 Hz); C NMR (125 MHz, DMSO-d
6
) !
=
28.4 (3C), 39.0, 49.6, 56.1, 123.9, 125.3, 125.5, 125.9, 126.8, 127.5, 128.5, 131.7, 133.5, 135.1, 173.8.
N-(tert-Butoxycarbonyl)-(S)-3-(1-naphthyl)alanine
S)-3-(1-Naphthyl)alanine hydrochloride (1.07 g, 4.3 mmol) was dissolved in 2 mol dm aqueous
sodium hydroxide (4.2 mL, 8.4 mmol) and the resulting solution was cooled on ice. tert-
–
3
(

–
3
Butoxycarbonyl chloride (1.12 g, 5.1 mmol) and 2 mol dm aqueous sodium hydroxide (2.1 mL, 4.2
mmol) were successively added to the above solution. The reaction mixture was stirred for 1 h at
–
3
room temperature and its pH was adjusted to 2 with 2 mol dm hydrochloric acid. The oily product was
extracted several times with EtOAc and the combined EtOAc extracts were washed with water and dried
over magnesium sulfate. The removal of the solvent yielded the title compound as an oily liquid (1.28 g,
1
9
5%). H NMR (500 MHz, DMSO-d
6
) ! = 1.28 (9H, s), 3.20 (1H, dd, J = 10.3, 13.7 Hz), 3.58 (1H, dd,
J = 3.4, 13.7 Hz), 4.21 (1H, ddd, J = 3.4, 8.0, 10.3 Hz), 7.23 (1H, d, J = 8.0 Hz), 7.41 (1H, dd, J = 6.9, 6.9
Hz), 7.43 (1H, d, J = 6.9 Hz), 7.52 (1H, dd, J = 7.4, 8.0 Hz), 7.58 (1H, dd, J = 7.4, 8.5 Hz), 7.79 (1H, d, J
1
3
=
6.9 Hz), 7.92 (1H, d, J = 8.0 Hz), 8.11 (1H, d, J = 8.5 Hz); C NMR (500 MHz, DMSO-d ) ! = 28.1
6
(3C), 33.9, 54.5, 78.1, 123.2, 125.4, 125.6, 126.3, 127.1, 127.5, 128.8, 131.4, 133.5, 134.0, 155.5, 173.8.
N-(tert-Butoxycarbonyl)-(S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide
N-(tert-Butoxycarbonyl)-(S)-3-(1-naphthyl)alanine (1.12 g, 4.2 mmol) and N-hydroxysuccinimide (0.72 g,
6
.2 mmol) were dissolved in 1,4-dioxane (20 mL). To this dioxane solution, a solution of N-ethyl-N’-
3-dimethylaminopropyl)carbodiimide (0.72 g, 6.2 mmol) in DMF (5 mL) was added with stirring at 0
C. The reaction mixture was stirred for 2h at room temperature and (S)-3-(1-naphthyl)alanine tert-
(
°
butylamide (1.57 g, 5.0 mmol) was added to this mixture. After the resulting solution had been stirred
for an additional 3 h at the same temperature, it was poured into water (100 mL). The resulting
crystalline solid was collected by suction filtration and subjected to column chromatography on silica gel
(70–230 mesh, EtOAc/hexane = 2/1 v/v) to yield the title compound as a colorless crystal (1.98 g, 84%).
1
H NMR (500 MHz, DMSO-d
6
) ! = 1.15 (9H, s), 1.26 (9H, s), 3.06 (1H, dd, J = 10.3, 14.3 Hz), 3.27–
.40 (3H, m), 4.20 (1H, ddd, J = 7.4, 9.7, 14.3 Hz), 4.65 (1H, dd, J = 7.4, 14.3 Hz), 7.22–7.24 (2H, m),
.33 (1H, dd, J = 7.4, 8.0 Hz), 7.41 (1H, d, J = 7.4 Hz), 7.41 (1H, d, J = 7.4 Hz), 7.52–7.59 (5H, m),
3
7
7
1
3
.76–7.81 (2H, m), 7.88–7.93 (3H, m), 8.07 (1H, d, J = 8.5 Hz), 8.31 (1H, d, J = 8.5 Hz); C NMR (125
) ! = 28.0 (3C), 28.2 (3C), 30.7, 34.6, 35.6, 50.2, 53.6, 78.4, 123.4, 124.2, 125.2, 125.3,
25.5 (2C), 126.0, 126.1, 126.9, 127.0, 127.2, 127.8, 128.4, 128.6, 131.5, 131.9, 133.3 (2C), 133.5, 133.8,
55.1, 169.7, 171.1.
MHz, DMSO-d
6
1
1
(S)-3-(1-Naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide
To a solution of N-(tert-butoxycarbonyl)-(S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-
–
3
butylamide (0.41 g, 0.72 mmol) in MeOH (20 mL), 4 mol dm hydrochloric acid (2 mL) was added with
stirring at room temperature. The reaction mixture was allowed to stand for 3 h at this temperature.
After the pH of the mixture had been adjusted to 10 with sodium carbonate, the product was extracted
several times with EtOAc. The combined EtOAc extracts were washed with saturated aqueous

sodium chloride and dried over magnesium sulfate. The removal of the solvent in vacuo yielded the title
1
compound as a colorless crystal (0.30 g, 90%). H NMR (500 MHz, DMSO-d
6
) ! = 1.14 (9H, s), 1.84
(2H, s), 2.63 (1H, dd, J = 9.1, 14.8 Hz), 3.32–3.34 (3H, m), 3.45 (1H, dd, J = 4.0, 9.1 Hz), 4.67 (1H, dd, J
=
7.4, 14.8 Hz), 7.23 (1H, d, J = 7.0 Hz), 7.33–7.41 (3H, m), 7.49–7.59 (5H, m), 7.76–7.80 (2H, m),
1
3
7
.89–7.91 (2H, m), 8.06 (1H, d, J = 8.0 Hz), 8.21 (1H, d, J = 8.5 Hz), 8.31 (1H, d, J = 8.5 Hz); C NMR
(
125 MHz, DMSO-d ) ! = 28.3 (3C), 35.9, 37.8, 50.2, 55.6, 59.7, 123.6, 124.3, 125.1, 125.3, 125.4, 125.5,
6
1
1
25.9 (2C), 126.8, 127.0, 127.3, 127.9, 128.4, 128.5, 131.5, 132.0, 133.3, 133.4, 133.6, 134.7, 169.9,
73.8.
N,N´-Ethylenedicarbonyl-bis[(S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide]
1c)
(
To a solution of (S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide (0.81 g, 1.7 mmol)
and pyridine (0.096 g, 1.2 mmol) in 1,4-dioxane (15 mL), succinyl chloride (0.094 g, 0.61 mmol) was
slowly added with stirring at 0 °C. The reaction mixture was stirred for 24 h at room temperature.
After the mixture had been concentrated to dryness under reduced pressure, the residual solid was
dissolved in EtOAc (25 mL). The EtOAc solution was washed with 10% aqueous citric acid and water
and dried over magnesium sulfate. The removal of the solvent afforded a crystalline solid, which was
purified by column chromatography on silica gel (70–230 mesh, EtOAc/hexane = 2/1 v/v) to yield the
1
title compound as a colorless crystal (0.36 g, 21%). H NMR (500 MHz, DMSO-d
6
) ! = 1.14 (18H, s),
2
3
8
7
7
8
.17 (1H, d, J = 9.1 Hz), 2.32 (1H, d, J = 9.1 Hz), 2.64 (1H, d, J = 14.8 Hz), 2.74 (1H, d, J = 14.8 Hz),
.06 (2H, dd, J = 9.1, 13.7 Hz), 3.29–3.34 (4H, m), 3.50 (2H, dd, J = 5.7, 13.7 Hz), 4.54 (4H, ddd, J = 7.4,
.0, 14.8 Hz), 7.10 (2H, br s), 7.20 (2H, d, J = 6.8 Hz), 7.27 (2H, dd, J = 7.4, 8.0 Hz), 7.33–7.42 (4H, m),
.44–7.47 (4H, m), 7.51 (2H, dd, J = 6.8, 7.4 Hz), 7.56 (2H, dd, J = 7.4, 8.0 Hz), 7.73 (2H, d, J = 8.5 Hz),
.79 (2H, d, J = 8.0 Hz), 7.85–7.88 (2H, m), 7.91 (2H, d, J = 8.0 Hz), 8.05–8.07 (2H, m), 8.15 (1H, d, J =
1
3
.0 Hz), 8.15 (1H, d, J = 8.0 Hz), 8.27 (2H, d, J = 8.0 Hz), 8.35 (2H, d, J = 6.8 Hz); C NMR (125 MHz,
) ! = 28.3 (6C), 34.5 (2C), 34.9 (2C), 42.8 (2C), 50.2 (3C), 54.0 (3C), 123.5 (2C), 123.9 (2C),
25.2 (4C), 125.4 (2C), 125.5 (2C), 126.0 (2C), 126.1 (2C), 127.0 (4C), 127.2 (2C), 127.5 (2C), 128.5
DMSO-d
6
1
(2C), 128.6 (2C), 131.5 (2C), 131.7 (2C), 133.3 (2C), 133.4 (2C), 133.5 (2C), 133.8 (2C), 169.8 (2C),
+
1
70.6 (2C), 171.2, 172.1; MALDI-TOF-MS: m/z calcd for C64
H
68
N
6
NaO
6
[M + Na] , 1017.26; Found,
1
017.11.
N-Acetyl-(S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide (1e)
Following the above protocol, (S)-3-(1-naphthyl)alanyl-(S)-3-(1-naphthyl)alanine tert-butylamide (0.28 g,
0
.60 mmol), pyridine (0.057 g, 0.72 mmol), and acetyl chloride (0.056 g, 0.71 mmol) in 1,4-dioxane (20

1
mL) afforded the title compound as a colorless crystal (0.30 g, 98%). H NMR (500 MHz, DMSO-d
1.15 (9H, s), 1.76 (3H, s), 3.12 (1H, dd, J = 8.5, 13.7 Hz), 3.25–3.34 (2H, m), 3.46 (1H, dd, J = 6.3,
3.7 Hz), 4.58 (2H, dd, J = 7.4, 8.0 Hz), 7.21–7.24 (2H, m), 7.31 (1H, dd, J = 7.4, 7.4 Hz), 7.36 (1H, d, J
6.3 Hz), 7.41 (1H, dd, J = 7.4, 8.0 Hz), 7.49–7.60 (4H, m), 7.75 (1H, d, J = 8.0 Hz), 7.80 (1H, d, J = 8.0
6
) !
=
1
=
1
3
Hz), 7.89–7.94 (2H, m), 8.09–8.12 (2H, m), 8.22–8.27 (2H, m); C NMR (125 MHz, DMSO-d
6
) ! = 22.4,
2
1
8.3 (3C), 34.5, 35.0, 50.2, 53.6, 53.9, 123.6, 123.9, 125.2, 125.3, 125.5 (2C), 126.0 (2C), 126.9, 127.0,
27.1, 127.6, 128.5, 128.6, 131.5, 131.8, 133.3, 133.4, 133.6, 133.7, 169.3, 169.7, 170.8; MALDI-TOF-
+
MS: m/z calcd for C32
H
35
N
3
NaO
3
[M + Na] , 509.64; Found, 509.81.
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