Chiral discrimination of permethylated gluco-oligosaccharide toward amino acid ester salts

Article abstract of DOI:10.1016/S0040-4020(01)00390-8

The chiral discrimination ability of permethylated glucopyrano-oligosaccharides toward amino acid 2-propyl ester hydrochlorides was evaluated using FAB mass spectrometry. In the given permethylated homo-oligosaccharides, permethylated β-cello-oligosacchar

Full text of DOI:10.1016/S0040-4020(01)00390-8

TETRAHEDRON
Pergamon
Tetrahedron 57 ,2001) 4567±4578
Chiral discrimination of permethylated gluco-oligosaccharide
toward amino acid ester salts
Motohiro Shizuma,a,p Hiroshi Adachi,b Akinori Amemura,c Yoshio Takai,d Tokuji Takedaa
and Masami Sawadad,p
aOsaka Municipal Technical Research Institute, 1-6-50 Morinomiya, Joto-ku, Osaka 536-8553, Japan
bFaculty ofScience, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan
cFaculty ofEngineering, Fukuyama University, Sanzo 1 Gakuen-cho, Fukuyama, Hiroshima 729-0292, Japan
dMaterials Analysis Center, The Institute ofScienti®c and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki,
Osaka 567-0047, Japan
Received 14 March 2001;accepted 4 April 2001
AbstractÐThe chiral discrimination ability of permethylated glucopyrano-oligosaccharides toward amino acid 2-propyl ester hydro-
chlorides was evaluated using FAB mass spectrometry. In the given permethylated homo-oligosaccharides, permethylated b-cello-oligo-
saccharide series ,II) showed remarkably higher S-selectivity toward tryptophan ester salts ,Trp-O-iPr1) independent of the numbers ,n) of
the glucopyranose unit ,nˆ2±5). The hexamer and heptamer of the permethylated b-malto-oligosaccharide series ,I) showed the very similar
enantioselectivity to permethylated a- and b-cyclodextrin. q 2001 Elsevier Science Ltd. All rights reserved.
1. Introduction
isomalto-oligosaccharides ,b-MeImal,n), nˆ2±7, series
IV)] toward seven species of amino acid 2-propyl ester
hydrochlorides ,Scheme 2), which was evaluated using
FAB mass spectrometry. It was found that the b-MeCel,n)
values show remarkably higher enantioselectivity toward
Trp-O-iPr1. It is very interesting that the in¯uence of the
glucosyl bond style on the chiral discrimination ability is
very different among the series I, II, III and IV.
Carbohydrates play important roles in the molecular
recognition processes of living systems using the
complicated structures found in stereochemistry. Molecular
recognition in living systems involves chiral recognition,
which is a signi®cant and fundamental process. Carbo-
hydrates, polysaccharides and their derivatives have chiral
discrimination ability and this is applied to the chiral
stationary phase in liquid chromatography.1 Chiral discrimi-
nation of some linear oligosaccharides toward chiral
compounds has been reported.2 However, there are few
studies on the 1:1 enantioselective complexation of oligo-
saccharides and their derivatives with chiral compounds.3 In
this study we clarify the 1:1 enantioselective complexation
characteristics of oligosaccharide derivatives with chiral
organic ammoniums using FAB mass spectrometry.4
2. Results and discussion
2.1. Chiral discrimination
The chiral discrimination ability was evaluated by means of
a FAB mass spectrometry ,MS)/enantiomer labeled ,EL)
guest method ,Table 1).5 In this method, FAB mass spectra
of permethylated oligosaccharide ,S) and a 1:1 deuterium-
labeled/unlabeled enantiomeric mixture of the amino acid
ester salts ,GS±dx1 and GR1) in 3-nitrobenzyl alcohol ,NBA)
matrix were measured and the peak intensity of two
complex ions [,S1GR)1 and ,S1GS±dx)1] of the saccharide
with each enantiomer of amino acid ester salt was observed
in the spectra. The complex ion peaks differ about the m/z
,equal to x, where x is the number of deuteriums) by the
labeling with the deuterium. The chiral discrimination
ability of the saccharides is evaluated by the relative peak
intensity [I,S1GR)1/I,S1GS±dx)1ˆIR/IS±dx] values. The
details of this method have been already reported and
reviewed by others.6 In electrospray ionization ,ESI) mass
In the present article, we report on the chiral discrimination
ability of ®ve permethylated glucopyrano-oligosaccharide
series ,Scheme 1) [b-malto-oligosaccharides ,b-MeMal,n),
nˆ2±7, series I), b-cello-oligosaccharides ,b-MeCel,n),
nˆ2±5,
series
II),
a-laminari-oligosaccharides
,a-MeLam,n), nˆ2±5, series III-a), b-laminari-oligo-
saccharides ,b-MeLam ,n), nˆ2±5, series III-b), and
Keywords: amino acids and derivatives;carbohydrate;enantioselection;
molecular recognition.
Corresponding authors. Fax: 181-6-6963-8040;
e-mail: shizuma@omtri.city.osaka.jp
p
0040±4020/01/$ - see front matter q 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020,01)00390-8

4568
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
Scheme 1. Permethylated gluco-oligosaccharides ,RˆCH3).
spectrometry, various complex ions are easily and directly
detected from a given sample solution. However, the chiral
discrimination ability evaluated by ESIMS was decreased in
comparison with that by FABMS.7 For the above reason, the
FABMS method was adopted for the evaluation.
derivatives. The ability hardly depends on the numbers of
the glucopyranose units in series II, i.e. IR/IS±dx is roughly
constant at 0.6±0.5 for b-MeCel2±b-MeCel5. The
disaccharide MeCel2 in the series showed relatively higher
chiral discrimination ability. It is then suggested that the
binding site comprises the next glucopyranose unit to each
other. Note also that the permethylated b-d-glucopyranose
,monosaccharide) showed no chiral discrimination ability
toward all given amino acid ester hydrochlorides ,Table
1), supporting the above observation that two glucose
units are necessary for chiral discrimination.
The FAB mass spectra of the permethylated penta-oligo-
saccharides in the present series with Trp-O-iPr1,Cl2) are
shown in Fig. 1. The diastereomeric 1:1 complex ions were
observed in the given saccharides and the amino acid ester
hydrochloride pair. In the case of series I, the chiral
discrimination was slightly detected toward Trp-O-iPr1
and Pro-O-iPr1 ,R-selectivity). In the case of series III,
the chiral discrimination ability was very small. In series
IV, the chiral discrimination was hardly observed except in
the case of MeImal2 toward Trp-O-iPr1 ,IR/IS±dxˆ0.65,
S-selectivity). Especially in the case of series II, the chiral
discrimination toward Trp-O-iPr1 was clearly detected
,Fig. 2). The chiral discrimination ability was larger than
the combination of the other series and the amino acid
2.2. Molecular dynamics of penta-oligosaccharides
The dynamic structures of the permethylated penta-oligo-
saccharides were simulated using the molecular dynamics
,MD) program. The most stable structure of the linear
saccharides is not easy to determine by molecular
simulation methods such as the molecular orbital ,MO)
and the molecular mechanics ,MM) calculations because

M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
4569
Scheme 2. Labeled/unlabeled amino acid 2-propyl ester hydrochloride pairs. ,R)-Enantiomer: R1ˆR2ˆCH,CH3)2; ,S)-enantiomer: R1ˆCH,CD3)2 [d6];
R2ˆCD,CD3)2, [d7].
Table 1. Chiral discrimination ability ,IR/IS±dx values) of permethylated gluco-oligosaccharides toward amino acid 2-propyl esters in FAB mass spectrometry
Permethylated oligosaccharides
I
Trp
Phe
Val
Met
Pro
Pgly
Tle
b-MeMal2
b-MeMal3
b-MeMal4
b-MeMal5
b-MeMal6
b-MeMal7
b-MeCel2
b-MeCel3
b-MeCel4
b-MeCel5
a-MeLam2
a-MeLam3
a-MeLam4
a-MeLam5
b-MeLam2
b-MeLam3
b-MeLam4
b-MeLam5
MeImal2
1.06
1.12
1.13
1.15
1.17
1.14
0.62
0.57
0.59
0.49
0.92
0.86
0.84
0.83
0.98
0.88
0.78
0.79
0.65
1.01
0.86
0.96
0.98
1.04
1.01
1.01
1.06
1.05
1.02
1.01
1.06
1.01
0.97
1.01
0.98
0.96
0.99
1.00
0.97
0.97
1.05
1.03
1.02
0.96
0.79
0.91
0.96
0.92
0.93
0.95
1.03
0.99
0.96
0.97
1.01
0.98
1.02
1.03
1.00
0.97
0.98
0.93
0.97
1.04
0.96
0.97
0.97
1.07
1.01
0.97
0.82
0.90
0.93
0.93
0.94
0.96
1.01
0.99
1.07
1.12
1.08
1.08
1.08
1.15
1.04
1.00
0.97
0.98
1.00
1.02
1.01
0.99
1.00
1.12
1.07
1.01
1.16
1.07
1.08
1.07
1.17
1.08
0.94
1.09
1.08
1.06
1.07
1.07
1.08
1.17
1.07
1.09
1.06
1.16
0.91
1.12
1.11
1.09
1.17
1.15
1.05
0.99
0.98
0.91
0.91
0.86
0.87
0.86
0.90
0.91
0.89
0.89
0.93
0.97
0.93
0.96
0.99
1.01
0.98
0.96
0.85
0.93
0.92
0.90
0.87
0.84
0.96
0.99
0.95
0.95
0.99
0.95
0.92
0.97
0.97
0.98
0.95
1.05
0.89
0.97
0.95
0.98
0.92
0.98
0.99
0.91
0.87
0.87
0.90
0.92
0.95
1.00
0.99
0.98
II
III-a
III-b
IV
MeImal3
MeImal4
MeImal5
MeImal6
MeImal7
b-MeGlc
18-crown-6
0.98
0.99
Ref.
of the ¯exibility of conformations like chains. So, the MD
was applied in order to evaluate the average structure of the
various conformations. The dynamic properties expected by
the MD simulation are summarized in Table 2. The
expected self-diffusion coef®cients, which are concerned
with the size and the shape of the molecules, for
b-MeCel5 and b-MeLam5 were larger than that for
b-MeMal5. For the averaged distance from C-1 of the
methoxy group in the reducing terminal to the C-4 of
the methoxy group in the irreducing terminal ,L), and the
averaged radius of gyration, b-MeCel5 was also larger than
b-MeMal5. The above expected properties determined by

4570
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
Figure 1. FAB mass spectra of permethylated penta-oligosaccharides with Trp-O-iPr1: ,a) b-MeMal5;,b) b-MeCel5;,c) a-MeLam5;,d) b-MeLam5;
,e) MeImal5.

M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
4571
Figure 2. Chiral discrimination ability ,IR/IS±dx values) of permethyl gluco-oligosaccharides toward Trp-O-iPr1. 18-crown-6, which is an achiral host, was
used as the standerd reference ,Ref.).
Table 2. Properties of permethylated penta-oligosaccharides calculated by molecular dynamics
a
Distance ,L) ,A)
Ê
Ê
Radius of gyration ,A)
Penta-oligosaccharides
Self-diffusion coef®cient
,cm2 s21 atom21
)
b-MeMal5
b-MeCel5
b-MeLam5
b-MeImal5
0.955£1026
0.254£1027
0.366£1027
0.177£1026
12.7
23.1
19.7
21.7
5.70
7.97
7.37
6.63
a
L: the averaged distance from C-1 of the methoxy group in the reducing terminal to C-4 of the methoxy group in the irreducing terminal.
the MD simulation corresponds with those of b-MeCel5
having the linear-type structure and b-MeMal5 having the
pseudo-ring-type structure ,Fig. 3). The structure of the free
,uncomplexed) permethylated oligosaccharides is not
necessarily the same as that of the complexes. However,
the free structure seems to be referable to the complex
structure, especially to the binding site.
2.3. Comparison between the b-MeMal series and the
permethylated cyclodextrin series
Because the b-MeMal5 was expected to be a pseudo-ring
structure by the MD simulation, it is reasonably assumed
that b-MeMal6 and b-MeMal7 have similar pseudo-ring
structures. On the other hand, the complex structure of
Figure 3. Averaged molecular structure of permethylated penta-oligosaccharides expected by molecular simulation: ,a) b-MeCel5;,b) b-MeMal5.

4572
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
Figure 4. Comparison of the IR/IS±dx values of b-MeMal,n) ,nˆ6 and 7) and those of permethylated cyclodextrin [MeCD,n)] ,nˆ6 and 7) in terms of chiral
discrimination toward amino acid 2-propyl ester hydrochlorides: ,a) MeMal6;,b) MeMal7;,c) MeCD6;and ,d) MeCD7.
cyclodextrinÐwhich has a cyclic structure consisting of
a,1!4)-linked glucopyranose unitsÐwith tryptophan has
been studied in detail by Lipkowitz et al.8 In that study, the
indole moiety of the tryptophan locates in the hole of
a-cyclodextrin, and the functional groups in the tryptophan
such as the amino group and the carboxylic group interact
with the oxygens of the carbohydrate. The complex of the
a-cyclodextrin with ,R)-tryptophan was assumed to be
more stable than the complex with ,S)-tryptophan. The
chiral discrimination of permethylated cyclodextrins
,a-MeCD and b-MeCD) toward some amino acid ester
salts has been previously reported.4 Also, the MeCD series
did not show any remarkable chiral discrimination ability as
in b-MeMal,n). However, the chiral discrimination of the
MeCD series is characteristically in good agreement with
the chiral discrimination of series I oligosaccharides except
in the case toward Met-O-iPr1 ,Fig. 4). The good agreement
suggests that the binding site of series I structures is the
same as that of cyclodextrins although the structure cannot
be con®rmed yet using the spectrometric method. The
Figure 5. 1H-NMR spectra of b-MeCel2 adding NH4SCN in acetone-d6 at 298 K. In the case of [NH4SCN]/[b-MeCel2]ˆ0, [b-MeCel2]ˆ0.00865 M.

M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
4573
complex structure of b-MeMal,n) molecules is not easily
clari®ed because of the small spectral change induced by the
complexation compared with cyclodextrins.
2.4. Expected complex structure of series II molecules
with Trp-O-iPr1
In order to clarify the binding site of the cationic moiety
,NH31) of Trp-O-iPr1 in the complex with b-MeCel2, the
1H-NMR-induced shifts of b-MeCel2 that occur when
adding NH41,SCN2) were followed in acetone-d6 at 258C
,Fig. 5).9 For facile observation of the induced shifts, an
ammonium ion was used in order to omit the redundant
interaction such as the p-effect of the indole moiety of
Trp-O-iPr1 etc. Acetone-d6 was selected as the solvent
because of the solvent effect which enhances the electro-
static interaction with the cation and the solubility of
ammonium thiocyanate. The speci®c induced shifts of the
pyranose ring protons were observed. Especially, the B-3
proton showed larger down-®eld shifts in its signal
,0.16 ppm). Therefore, ammonium would bind mainly to
the oxygen of the OMe-3 group.10 The 1:1 association
constant was evaluated at ca. 1 M21 from the induced
shifts.
Figure 7. Fluorescence spectra of ,S)-Trp-O-iPr1 and ,S)-Trp-,S)-Trp-O-
iPr1 in H2O at 298 K. The exciting wavelength was 287 nm: ,a) ,S)-Trp-O-
iPr1Cl2, [A1Cl2]ˆ3.53£1025 M;,b) , S)-Trp-O-iPr1Cl21b-MeCel2, [S]/
[A1Cl2]ˆ50 ,mol/mol);,c)
,
S)-Trp-,S)-Trp-O-iPr1, [A1Cl2]ˆ3.87£
1025 M;,d) , S)-Trp-,S)-Trp-O-iPr11b-MeCel2, [S]/[A1Cl2]ˆ100 ,mol/
Further, for clarifying the interaction between b-MeCel2
mol).
1
and the indole moiety of Trp-O-iPr1, H-NMR spectra of
b-MeCel2 were measured while adding ,S)-Trp-O-iPr1 in
D2O at 258C ,Fig. 6). Here, D2O was selected as the solvent
in order to enhance the electrophobic interaction such as the
CH±p interaction: generally, the CH±p interaction, for
example, between b-MeCel2 and the indole moiety of
Trp-O-iPr1 in the complex,would be enhanced in the gas
phase. Most of the peaks showed slight down-®eld shifts or
no shift. One of the OMe groups showed a small up-®eld
shift ,0.02 ppm) by the p-electron effect of the indole
moiety.11 The indole moiety must be located near the
methoxy group. The methoxy group could not be assigned.
of the same amino acid units, the ¯uorescence intensity of
the dipeptide ester salt was smaller than that of the mono-
amino acid ester salt because the exciting energy of the
indole moieties was quenched by the p±p intramolecular
interaction between the indole moietes.12 By adding an
excess of MeCel2 to ,S)-Trp-O-Pr1 ,Cl2) solution, the
¯uorescence intensity was reduced by about 25% on the
anisotropy effect. By adding the same amount of MeCel2
to ,S)-Trp-,S)-Trp-O-iPr1 ,Cl2), the intensity was reduced
by about 14%. This reduction of the ¯uorescence intensity
suggests that the quenching became weaker during the inter-
molecular interaction between the indole moieties and
b-MeCel2, which inhibits the intramolecular stacking of
the indole moieties in ,S)-Trp-,S)-Trp-O-iPr1 ,Cl2). In the
case of b-MeCD, which has the larger association ability
toward electrophobic compounds in water,13 the
¯uorescence intensity was enhanced by about 15% as the
larger inhibition effect on the quenching in the dipeptide. In
adding b-MeMal2 instead of b-MeCel2, the reduction of
¯uorescence intensity of the ,S)-Trp-O-iPr1 ,Cl2) solution
was at the same level as that of the ,S)-Trp-,S)-Trp-O-iPr1
,Cl2).
Fluorescence spectra of aqueous solutions of ,S)-Trp-O-Pr1
,Cl2) and ,S)-Trp-,S)-Trp-O-iPr1 ,Cl2) were measured at
258C ,Fig. 7). Further, after adding MeCel2 to each solution,
¯uorescence spectra were measured. Under equal quantities
From the above results, the mechanism of chiral
discrimination found in the action of permethylated cello-
oligosaccharides toward Trp-O-iPr1 can be accounted for
by three interaction concepts described as follows.14
1. The ammonium cation moiety in Trp-O-iPr1 binds to the
methoxy oxygen ,OMe-3) of one glucopyranose moiety
,electrostatic interaction).
2. The indole moiety in Trp-O-iPr1 binds to the methoxy
group of the next glucopyranose moiety ,CH±p inter-
action).15
3. The other substituents of the asymmetric carbon in Trp-
O-iPr1 are the 2-propyl ester group and the hydrogen.
Figure 6. Methyl protons in 1H-NMR spectra of MeCel2 in D2O at 298K:
,a) b-MeCel2 only;,b)
b-MeCel21Trp-O-iPr1,Cl2), [Trp-O-iPr1]/
[b-MeCel2]ˆca. 15. One methyl proton ,p) showed high-®eld shift
,0.02 ppm).

4574
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
In the situation where the 2-propyl ester group locates near
part of the permethylated cello-oligosaccharide, the
complex becomes unstable with respect to repulsion ,steric
effects). In the situation where the latter hydrogen locates
near part of the permethylated cello-oligosaccharide,
the complex becomes more stable. The difference in the
stability of these diastereomeric complexes re¯ects the
chiral discrimination toward Trp-O-iPr1. In order to clarify
the binding site in more detail, further investigations must
be undertaken by means of X-ray crystallography because
of the small interactions between the given permethylated
oligosaccharides and amino acid ester salts.
4.2. Series I
1
4.2.1. b-MeMal2. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.64 ,d, 1H, J1,2ˆ3.9 Hz, H1a-B), 4.15
3
3
,d, 1H, J1,2ˆ7.7 Hz, H1b-A), 3.83 ,dd, 1H, J3,4ˆ8.8 Hz,
3J4,5ˆ9.7 Hz, H4-A), 3.64 ,s, 3H, ZCH3), 3.56 ,s, 3H,
ZCH3), 3.55 ,s, 3H, ZCH3), 3.54 ,s, 6H, ZCH3), 3.53 ,s,
3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.33 ,s, 3H, ZCH3), 3.60±
3.37 ,m, 16H, pyranose ring protons), 3.42 ,dd,
3
3
1H, J2,3ˆ9.5 Hz, J3,4ˆ9.0 Hz, H3-B), 3.41 ,dd, 1H,
3J2,3ˆ9.2 Hz, J3,4ˆ8.8 Hz, H3-A), 3.26 ,dd, 1H, J3,4ˆ
3
3
3
3
8.8 Hz, J4,5ˆ9.9 Hz, H4-B), 3.19 ,dd, 1H, J1,2ˆ3.9 Hz,
3J2,3ˆ9.7 Hz, H4-B), 3.06 ,dd, 1H, J1,2ˆ7.8 Hz,
3
3J2,3ˆ9.2 Hz, H4-A) from TMS. FT-IR ,neat, cm21) 1186,
1146, 1030 ,ether).
3. Summary
1
4.2.2. b-MeMal3. Colorless syrup; H-NMR ,300 MHz,
3
We clari®ed that permethylated oligosaccharides consisting
of only a glucopyranose ring unit show various levels of
chiral discrimination toward the present amino acid ester
salts depending on the number of the units and the glycosyl
link style such as a,1!4), b,1!4), b,1!3), and a,1!6).
For example, the present permethylated cello-oligo-
saccharides of the b,1!4) bond form showed different
chiral discriminations toward tryptophan derivatives from
the present permethylated malto-oligosaccharides of the
a,1!4) bond form. The reasons are that the structures
and the binding sites of the oligosaccharides are very
different from each other as is presumed using NMR, FL
spectral and MD calculation methods. It was assumed that
series I structures have the same binding behavior as that
found with the permethylated cyclodextrins, and that series
II molecules have a binding site which consists of two
adjacent glucopyranose moieties in a linear-type structure.
CDCl3) d ,ppm) 5.64 ,d, 1H, J1,2ˆ3.7 Hz, H1a), 5.54 ,d,
3
3
1H, J1,2ˆ3.9 Hz, H1a), 4.16 ,d, 1H, J1,2ˆ7.7 Hz, H1b),
3.62 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.56 ,s, 3H,
ZCH3), 3.55 ,s, 6H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.53 ,s,
6H, ZCH3), 3.51 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.35
,s, 3H, ZCH3), 3.32 ,s, 3H, ZCH3), 3.60±3.06 ,m, 18H,
pyranose ring protons) from TMS. FT-IR ,neat, cm21) 1186,
1148, 1102, 1040 ,ether). FABMS ,NBA matrix) m/z 697
,M1K)1.
1
4.2.3. b-MeMal4. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.65 ,d, 1H, J1,2ˆ3.8 Hz, H1a), 5.56 ,d,
3
3
1H, J1,2ˆ3.9 Hz, H1a), 5.54 ,d, 1H, J1,2ˆ3.7 Hz, H1a),
3
4.17 ,d, 1H, J1,2ˆ7.7 Hz, H1b), 3.61 ,s, 3H, ZCH3), 3.58
,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.56 ,s, 6H, ZCH3),
3.55 ,s, 3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.53 ,s, 3H,
ZCH3), 3.52 ,s, 3H, ZCH3), 3.51 ,s, 3H, ZCH3), 3.49
,s, 3H, ZCH3), 3.36 ,s, 3H, ZCH3), 3.35 ,s, 3H, ZCH3),
3.30 ,s, 3H, ZCH3), 3.26 ,s, 3H, ZCH3), 3.60±3.02 ,m,
In monosaccharides, there are many epimers such as
glucose, galactose, mannose, etc. The oligosaccharides
suitably consisting of epimers must show a strongly
different chiral or molecular discrimination ability.
24H, pyranose ring protons) from TMS. FT-IR ,neat, cm21
)
1188, 1151, 1102, 1042 ,ether). FABMS ,NBA matrix) m/z
1001 ,M1K)1.
1
4.2.4. b-MeMal5. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.64 ,d, 1H, J1,2ˆ3.7 Hz, H1a), 5.57 ,d,
3
3
4. Experimental
4.1. Materials
1H, J1,2ˆ3.9 Hz, H1a), 5.56 ,d, 1H, J1,2ˆ3.0 Hz, H1a),
5.53 ,d, 1H, 3J1,2ˆ3.9 Hz, H1a), 4.17 ,d, 1H, 3J1,2ˆ7.7 Hz,
H1b), 3.61 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.56 ,s, 3H,
ZCH3), 3.55 ,s, 6H, ZCH3), 3.55 ,s, 3H, ZCH3), 3.54 ,s,
3H, ZCH3), 3.53 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.51
,s, 3H, ZCH3), 3.50 ,s, 3H, ZCH3), 3.46 ,s, 3H, ZCH3),
3.31 ,s, 3H, ZCH3), 3.30 ,s, 3H, ZCH3), 3.27 ,s, 3H,
ZCH3), 3.60±3.02 ,m, 30H, pyranose ring protons) from
TMS. FT-IR ,neat, cm21) 1151, 1102, 1026 ,ether).
FABMS ,NBA matrix) m/z 1105 ,M1K)1.
Permethylated oligosaccharides: the present permethylated
oligosaccharides were prepared from free oligosaccharides
,hydroxyl type) by the Hakomori method.16 The commercial
products were used in the cases of malto-oligosaccharides
,Hayashihara Biochemical Laboratories, Inc.), cello-oligo-
saccharides ,Sigma), and isomalto-oligosaccharides
,Seikagaku Corporation). Laminari-oligosaccharides were
synthesized by an enzyme reaction.17 The prepared
permethylated oligosaccharides were puri®ed using
medium-pressure liquid chromatography ,silica gel 60,
AcOEt/n-hexane/MeOHˆ5:5:1 by volume) to the anomers,
respectively. Isomalto-oligosaccharides could not be
puri®ed to the anomers. Note that the anomeric mixtures
were used in these experiments.
1
4.2.5. b-MeMal6. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.65 ,d, 1H, J1,2ˆ3.7 Hz, H1a), 5.58 ,d,
3
3
1H, J1,2ˆ3.5 Hz, H1a), 5.57 ,d, 1H, J1,2ˆ4.0 Hz, H1a),
5.56 ,d, 1H, 3J1,2ˆ3.9 Hz, H1a), 5.53 ,d, 1H, 3J1,2ˆ3.7 Hz,
H1a), 4.17 ,d, 1H, 3J1,2ˆ7.7 Hz, H1b), 3.61 ,s, 3H, ZCH3),
3.59 ,s, 3H, ZCH3), 3.58 ,s, 3H, ZCH3), 3.57 ,s, 6H,
ZCH3), 3.56 ,s, 3H, ZCH3), 3.56 ,s, 3H, ZCH3), 3.55 ,s,
3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.53 ,s, 3H, ZCH3), 3.52
,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.51 ,s, 3H, ZCH3),
3.50 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.40 ,s, 3H,
Physical and spectral properties of the permethylated gluco-
oligosaccharides are presented in the following sections.

M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
4575
ZCH3), 3.39 ,s, 3H, ZCH3), 3.36 ,s, 3H, ZCH3), 3.34 ,s,
3H, ZCH3), 3.31 ,s, 3H, ZCH3), 3.60±3.02 ,m, 36H,
nose ring protons), 3.62 ,s, 3H, ZCH3), 3.58 ,s, 3H, ZCH3),
3.55 ,s, 3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.52 ,s, 3H,
ZCH3), 3.40 ,s, 3H, ZCH3), 3.39 ,s, 3H, ZCH3) from
TMS. FT-IR ,KBr disk, cm21) 1068 ,ether). FABMS
,NBA matrix) m/z 1105 ,M1K)1.
pyranose ring protons) from TMS. FT-IR ,neat, cm21
1188, 1156, 1102, 1038 ,ether). FABMS ,NBA matrix)
)
m/z 1309 ,M1K)1.
1
4.2.6. b-MeMal7. Colorless syrup; H-NMR ,300 MHz,
4.4. Series III-a
3
CDCl3) d ,ppm) 5.65 ,d, 1H, J1,2ˆ3.7 Hz, H1a), 5.58±
3
1
4.4.1. a-MeLam2. Colorless syrup; H-NMR ,300 MHz,
5.57 ,d, 4H, J1,2ˆ3.5 Hz, H1a), 5.53 ,d, 1H,
3
3J1,2ˆ4.0 Hz, H1a), 4.17 ,d, 1H, J1,2ˆ7.9 Hz, H1b), 3.61
3
CDCl3) d ,ppm) 4.84 ,d, 1H, J1,2ˆ3.5 Hz, H1), 4.69
3
3
,d, 1H, J1,2ˆ7.7 Hz, H1), 4.08 ,t, 1H, J2,3ˆ3J3,4ˆ9.0 Hz,
H3), 3.60±3.13 ,m, 9H, pyranose ring protons), 3.62 ,s, 3H,
ZCH3), 3.59 ,s, 3H, ZCH3), 3.53 ,s, 3H, ZCH3), 3.52
,s, 3H, ZCH3), 3.48 ,s, 3H, ZCH3), 3.42 ,s, 3H, ZCH3),
3.41 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3) 3.29 ,dd, 1H,
,s, 3H, ZCH3), 3.59 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3),
3.57 ,s, 6H, ZCH3), 3.56 ,s, 3H, ZCH3), 3.55 ,s, 3H,
ZCH3), 3.54 ,s, 3H, ZCH3), 3.53 ,s, 3H, ZCH3), 3.53 ,s,
3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.51
,s, 3H, ZCH3), 3.51 ,s, 3H, ZCH3), 3.50 ,s, 3H, ZCH3),
3.40 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.39 ,s, 3H,
ZCH3), 3.39 ,s, 3H, ZCH3), 3.35 ,s, 3H, ZCH3), 3.33
,s, 3H, ZCH3), 3.31 ,s, 3H, ZCH3), 3.60±3.02 ,m, 42H,
pyranose ring protons) from TMS. FT-IR ,neat, cm21) 1188,
1156, 1102, 1024 ,ether). FABMS ,NBA matrix) m/z 1467
,M1K)1.
3
3J1,2ˆ3.5 Hz, J2,3ˆ9.5 Hz, H2), 2.99±2.91 ,m, 1H,
pyranose ring proton) from TMS. FT-IR ,neat, cm21
1102, 1051 ,ether). FABMS ,NBA matrix) m/z 493
)
,M1K)1.
1
4.4.2. a-MeLam3. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 4.84 ,d, 1H, J1,2ˆ3.5 Hz, H1), 4.69
3
3
,d, 1H, J1,2ˆ7.9 Hz, H1), 4.69 ,d, 1H, J1,2ˆ8.0 Hz, H1),
4.3. Series II
3
4.08 ,t, 1H, J2,3ˆ3J3,4ˆ9.0 Hz, H3), 3.77±3.65 ,m, 2H,
pyranose ring protons), 3.63 ,s, 3H, ZCH3), 3.61 ,s, 3H,
ZCH3), 3.58 ,s, 3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.52
,s, 3H, ZCH3), 3.51 ,s, 3H, ZCH3), 3.48 ,s, 3H, ZCH3),
3.41 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.63±2.96 ,m,
15H, pyranose ring proton) from TMS. FT-IR ,neat,
cm21) 1100, 1053 ,ether). FABMS ,NBA matrix) m/z 697
,M1K)1.
4.3.1. b-MeCel2. Colorless crystal;mp ˆ75±768C;
1H-NMR ,300 MHz, CDCl3) d ,ppm) 4.31 ,d, 1H,
3
3J1,2ˆ7.9 Hz, H1,b)), 4.15 ,d, 1H, J1,2ˆ7.7 Hz, H1,b)),
3.71±3.11,m, 10H, pyranose ring protons), 3.62 ,s, 3H,
ZCH3), 3.58 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.55 ,s,
3H, ZCH3), 3.53 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.48
,s, 3H, ZCH3), 3.62 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3),
3
3.39 ,s, 3H, ZCH3), 3.02 ,t, 1H, J1,2ˆ3J2,3ˆ8.1 Hz, H2),
1
4.4.3. a-MeLam4. Colorless syrup; H-NMR ,300 MHz,
3
2.93 ,t, 3H, 1H, J1,2ˆ3J2,3ˆ8.1 Hz, H2) from TMS. FT-IR
3
,KBr disk, cm21) 1132, 1100, 1067, 1044 ,ether). FABMS
CDCl3) d ,ppm) 4.84 ,d, 1H, J1,2ˆ3.4 Hz, H1), 4.69 ,d,
3
3
,NBA matrix) m/z 493 ,M1K)1.
1H, J1,2ˆ7.5 Hz, H1), 4.68 ,d, 1H, J1,2ˆ7.9 Hz, H1),
3
3
4.68 ,d, 1H, J1,2ˆ8.1 Hz, H1), 4.08 ,t, 1H, J2,3ˆ3J3,4ˆ
9.2 Hz, H3), 3.63±3.40 ,m, 2H, pyranose ring protons),
3.63 ,s, 3H, ZCH3), 3.61 ,s, 3H, ZCH3), 3.58 ,s, 3H,
ZCH3), 3.54 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.51 ,s,
3H, ZCH3), 3.48 ,s, 3H, ZCH3), 3.41 ,s, 3H, ZCH3), 3.41
,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.32±2.93 ,m, 20H,
pyranose ring proton) from TMS. FT-IR ,neat, cm21) 1127,
1053, 1053 ,ether). FABMS ,NBA matrix) m/z 901
,M1K)1.
4.3.2. b-MeCel3. Colorless crystal;mp ˆ106±1088C;
1H-NMR ,300 MHz, CDCl3) d ,ppm) 4.32 ,d, 2H,
3
3J1,2ˆ7.9 Hz, H1,b)), 4.15 ,d, 1H, J1,2ˆ7.7 Hz, H1,b)),
3.79±3.10 ,m, 15H, pyranose ring protons), 3.62 ,s, 3H,
ZCH3), 3.58 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.55 ,s,
3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.40
,s, 3H, ZCH3), 3.62 ,s, 3H, ZCH3), 3.39 ,s, 3H, ZCH3),
3
3.38 ,s, 3H, ZCH3), 3.02 ,t, 1H, J1,2ˆ3J2,3ˆ7.9 Hz, H2),
3
2.97 ,t, 1H, J1,2ˆ3J2,3ˆ8.1 Hz, H2), 2.93 ,t, 3H, 1H,
1
4.4.4. a-MeLam5. Colorless syrup; H-NMR ,300 MHz,
3J1,2ˆ3J2,3ˆ6.7 Hz, H2) from TMS. FT-IR ,KBr disk,
cm21) 1142, 1124, 1090, 1042 ,ether). FABMS ,NBA
matrix) m/z 697 ,M1K)1.
3
CDCl3) d ,ppm) 4.84 ,d, 1H, J1,2ˆ3.3 Hz, H1), 4.70
3
3
,d, 2H, J1,2ˆ7.7 Hz, H1), 4.69 ,d, 2H, J1,2ˆ7.7 Hz, H1),
3
4.08 ,t, 1H, J2,3ˆ3J3,4ˆ9.2 Hz, H3), 3.78±2.89 ,m, 29H,
pyranose ring protons), 3.63 ,s, 3H, ZCH3), 3.60 ,s, 3H,
ZCH3), 3.58 ,s, 3H, ZCH3), 3.55 ,s, 3H, ZCH3), 3.52
,s, 3H, ZCH3), 3.48 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3)
from TMS. FT-IR ,neat, cm21) 1100, 1055 ,ether). FABMS
,NBA matrix) m/z 1105 ,M1K)1.
4.3.3. b-MeCel4. Colorless crystal;mp ˆ116±1188C;
1H-NMR ,300 MHz, CDCl3) d ,ppm) 4.33 ,d, 1H,
3
3J1,2ˆ7.5 Hz, H1,b)), 4.32 ,d, 2H, J1,2ˆ7.9 Hz, H1,b)),
4.15 ,d, 1H, 3J1,2ˆ7.7 Hz, H1,b)), 3.92±2.80 ,m, 24H, pyra-
nose ring protons), 3.62 ,s, 3H, ZCH3), 3.58 ,s, 3H, ZCH3),
3.55 ,s, 3H, ZCH3), 3.54 ,s, 3H, ZCH3), 3.52 ,s, 3H,
ZCH3), 3.41 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.39 ,s,
3H, ZCH3) from TMS. FT-IR ,KBr disk, cm21) 1126, 1090,
,ether). FABMS ,NBA matrix) m/z 901 ,M1K)1.
4.5. Series III-b
1
4.5.1. b-MeLam2. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 4.68 ,d, 1H, J1,2ˆ7.7 Hz, H1), 4.13
3
3
4.3.4. b-MeCel5. Colorless crystal;mp ˆ134±1358C;
1H-NMR ,300 MHz, CDCl3) d ,ppm) 4.33 ,d, 2H,
,d, 1H, J1,2ˆ7.9 Hz, H1), 3.76 ,t, 1H, J2,3ˆ3J3,4ˆ8.8 Hz,
H3), 3.67±3.14 ,m, 9H, pyranose ring protons), 3.63 ,s, 3H,
ZCH3), 3.59 ,s, 3H, ZCH3), 3.57 ,s, 3H, ZCH3), 3.52
,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3), 3.07 ,dd, 1H,
3
3J1,2ˆ7.7 Hz, H1,b)), 4.32 ,d, 2H, J1,2ˆ7.7 Hz, H1,b)),
4.15 ,d, 1H, 3J1,2ˆ7.7 Hz, H1,b)), 3.79±2.92 ,m, 30H, pyra-

4576
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
3
3J2,3ˆ3J3,4ˆ8.4 Hz, H3), 2.93 ,m, 1H, pyranose ring proton)
from TMS. FT-IR ,neat, cm21) 1107, 1059 ,ether). FABMS
,NBA matrix) m/z 493 ,M1K)1.
proton H1a), 4.96 ,d, 4H, J1,2ˆ3.0 Hz, H1a), 4.12
3
,d, J1,2ˆ7.8 Hz, reducing terminal proton H1b) from
TMS, a/b ratioˆ1/0.23;FT-IR ,KBr disk, cm 21) 1159,
1102, 1041 ,ethers).
1
4.5.2. b-MeLam3. Colorless syrup; H-NMR ,300 MHz,
3
4.6.5. MeImal6. Colorless solid; 1H-NMR ,300 MHz,
CDCl3) d ,ppm) 4.69 ,d, 1H, J1,2ˆ7.7 Hz, H1), 4.69 ,d,
3
3
3
1H, J1,2ˆ8.1 Hz, H1), 4.14 ,d, 1H, J1,2ˆ8.1 Hz, H1),
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.2 Hz, reducing terminal
3.77 ,t, 1H, J2,3ˆ3J3,4ˆ8.8 Hz, H3), 3.74 ,t, 1H, J2,3ˆ
3J3,4ˆ7.3 Hz, H3), 3.67±3.15 ,m, 13H, pyranose ring
protons), 3.63 ,s, 3H, ZCH3), 3.61 ,s, 3H, ZCH3), 3.59 ,s,
3H, ZCH3), 3.55 ,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.40
,s, 3H, ZCH3), 3.07 ,t, 1H, 3J2,,3ˆ3J3,4ˆ8.8 Hz, H3), 3.02 ,t,
3
3
3
proton H1a), 4.96 ,d, 5H, J1,2ˆ3.0 Hz, H1a), 4.13
3
,d, J1,2ˆ7.8 Hz, reducing terminal proton H1b) from
TMS, a/b ratioˆ1/0.24;FT-IR ,KBr disk, cm 21) 1174,
1016.
3
1H, J2,,3ˆ3J3,4ˆ8.8 Hz, H3), 2.93 ,m, 1H, pyranose ring
4.6.6. MeImal7. Colorless solid; 1H-NMR ,300 MHz,
proton) from TMS. FT-IR ,neat, cm21) 1105, 1059
3
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.2 Hz, reducing terminal
,ether). FABMS ,NBA matrix) m/z 697 ,M1K)1.
3
proton H1a), 4.96 ,d, 6H, J1,2ˆ3.0 Hz, H1a), 4.13
3
,d, J1,2ˆ7.5 Hz, reducing terminal proton H1b) from
1
4.5.3. b-MeLam4. Colorless syrup; H-NMR ,300 MHz,
TMS, a/b ratioˆ1/0.22;FT-IR ,KBr disk, cm 21) 1108,
3
CDCl3) d ,ppm) 4.70 ,d, 1H, J1,2ˆ7.7 Hz, H1), 4.69
1096, 1044, 1028.
3
3
,d, 2H, J1,2ˆ7.9 Hz, H1), 4.13 ,d, 1H, J1,2ˆ7.9 Hz, H1),
3.75 ,t, 1H, J2,3ˆ3J3,4ˆ8.8 Hz, H3), 3.65±3.43 ,m, 9H,
3
pyranose ring protons), 3.63 ,s, 3H, ZCH3), 3.61 ,s, 3H,
ZCH3), 3.60 ,s, 3H, ZCH3), 3.59 ,s, 3H, ZCH3), 3.55
,s, 3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.40 ,s, 3H, ZCH3),
3.40 ,s, 3H, ZCH3), 3.36±2.90 ,m, 1H, pyranose ring
proton) from TMS. FT-IR ,neat, cm21) 1107, 1057
,ether). FABMS ,NBA matrix) m/z 901 ,M1K)1.
4.7. Further materials
Amino acid 2-propyl ester hydrochlorides: the present
amino acid 2-propyl ester hydrochlorides were prepared
by the esteri®cation of the free amino acids with 2-propanol
under HCl catalyst.18 ,S)-Enantiomers were reacted with
deuterium-labeled 2-propanol ,99 at.% D, CDN isotopes).
,R)-Enantiomers were reacted with unlabeled 2-propanol
,Wako).
1
4.5.4. b-MeLam5. Colorless syrup; H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 4.70 ,d, 1H, J1,2ˆ7.7 Hz, H1), 4.69 ,d,
3
3
2H, J1,2ˆ7.7 Hz, H1), 4.13 ,d, 1H, J1,2ˆ7.9 Hz, H1),
3.75 ,m, 4H, pyranose ring protons), 3.62±3.40 ,m, 1H,
pyranose ring protons), 3.63 ,s, 3H, ZCH3), 3.61 ,s, 3H,
ZCH3), 3.60 ,s, 3H, ZCH3), 3.59 ,s, 3H, ZCH3), 3.56 ,s,
3H, ZCH3), 3.52 ,s, 3H, ZCH3), 3.50 ,s, 3H, ZCH3), 3.48
,s, 3H, ZCH3), 3.41 ,s, 3H, ZCH3), 3.31±2.90 ,m, 16H,
pyranose ring proton) from TMS. FT-IR ,neat, cm21) 1098,
1055 ,ether). FABMS ,NBA matrix) m/z 1105 ,M1K)1.
7S)-Trp-7S)-Trp-O-iPr17Cl2): equimolar Boc-,S)-Trp and
,S)-Trp-O-iPr1 were dissolved in DMF containing 1.1
equimolar triethylamine and HOBt and the mixture
condensed with equimolar 1-,3-dimethylaminopropyl)-3-
ethylcarbodiimide hydrochloride ,EDC).19 After cleavage
of the protective group with TFA, the counter anion was
exchanged with Cl2 using an anion-exchange resin
,Dowex AG1-X2, Cl2 form). The product was
recrystallized in MeOH/AcOEt to yield 54% product.
1H-NMR ,300 MHz, DMSO) d ,ppm) 11.03 ,s, 1H, NH),
7.72 ,d, 1H, 3Jˆ7.5 Hz, ZCZCHyCH), 7.52 ,d, 1H,
3Jˆ6.8 Hz, ZCZCHyCH), 7.38 ,d, 1H, 3Jˆ7.8 Hz,
ZCZCHyCH), 7.35 ,d, 1H, 3Jˆ6.9 Hz, CZCHyCH),
7.25±6.98 ,m, 6H, aromatic protons), 4.84 ,m, 1H,
CHZNH21), 4.60 ,m, 1H, CHZNH31), 4.03 ,m, 1H,
ZCH,CH3)2), 3.20±3.10 ,m, 4H, methylene), 1.15 ,d, 3H,
3Jˆ6.1 Hz, ZCH,CH3)2), 1.01 ,d, 3H, 3Jˆ6.6 Hz,
ZCH,CH3)2) from TMS.
4.6. Series IV: anomeric mixture
4.6.1. MeImal2. Colorless solid; 1H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.1 Hz, reducing terminal
3
proton H1a), 4.96 ,d, 1H, J1,2ˆ3.1 Hz, H1a), 4.13
3
,d, J1,2ˆ7.8 Hz, reducing terminal proton H1b) from
TMS, a/b ratioˆ1/0.28;FT-IR ,KBr disk, cm 21) 1108,
1068 ,ethers).
4.6.2. MeImal3. Colorless solid; 1H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.0 Hz, reducing terminal
3
proton H1a), 4.97 ,d, 2H, J1,2ˆ3.0 Hz, H1a), 4.13
3
4.8. General
,d, J1,2ˆ7.5 Hz, reducing terminal proton H1b) from
TMS, a/b ratioˆ1/0.23;FT-IR ,KBr disk, cm 21) 1140,
1102, 1040 ,ethers).
FAB mass spectra were measured with a JEOL SX-102
mass spectrometer and the data were analyzed with a
JMA-DA 6000 data processing system. NMR spectra were
measured with a JEOL AL-300 spectrometer. FL spectra
were measured with a Shimadzu RF-5300PC ¯uorescence
photometer.
4.6.3. MeImal4. Colorless solid; 1H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.1 Hz, reducing terminal
3
proton H1a), 4.97 ,d, 3H, J1,2ˆ3.1 Hz, H1a), 4.14
3
,d, J1,2ˆ7.5 Hz, reducing terminal proton H1b) from
TMS, a/b ratioˆ1/0.25;FT-IR ,KBr disk, cm 21) 1102,
1042 ,ethers).
Synthesized compounds were assigned using other
instrumental analyses ,FT-IR: Shimadzu FT-IR 8100
spectrometer;elemental analysis: Perkin±Elmer 2400;
melting points: Yanaco-MP apparatus).
4.6.4. MeImal5. Colorless solid; 1H-NMR ,300 MHz,
3
CDCl3) d ,ppm) 5.03 ,d, J1,2ˆ3.1 Hz, reducing terminal

M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
4577
4.9. FABMS/EL guest method
,Aldrich, 99.5 at.% D) was used as a solvent without
puri®cation. TMS was used as internal standard. The KS
values for the complexation equilibrium between
b-MeCel2 and ammonium thiocyanate at 258C ,298 K)
were determined by the following procedure. To an NMR
tube containing a 0.6 mL acetone-d6 solution of b-MeCel2
,8.65 mM) was added dropwise ,0, 5, 10, 15, 20, 25, and
30 mL) an acetone-d6 solution of ammonium thiocyanate
FAB mass spectra ,positive mode) were measured operating
at an accelerating voltage of 10 kV with a mass range of m/z
100±2400. The instrument was equipped with a standard
JEOL FAB source and an ion gun. Xenon was used as the
atom beam with an emission current of 10 mA and an
acceleration of 3 kV. The ion source pressure was typically
ca. 1±2£1025 Torr. Spectra were obtained with a magnet
scan rate of 10 s scan21 ,to m/z 2400) at room temperature
,258C) and the data were processed with a JEOL JMA-DA
6000 data processing system. Calibration was carried out
with CsI.
1
,0.460 mM mL21) and H-NMR spectra were measured in
order. To maintain the sample solution at a constant
temperature the NMR tube was maintained for 15 min on
the probe at 258C ,298 K) before measuring the NMR
spectra. The averages of the KS values estimated from the
concentration ratio [NH4SCN]/[b-MeCel2] and the
corresponding shifts of the following proton peaks were
adopted. It was assumed that a 1:1 complex is stoichio-
metrically formed in the solution because of the obser-
vation of the 1:1 complex ions in FAB mass spectra.
The K values were calculated using a general non-linear
method.22
A sample solution was prepared by mixing two solutions
and matrix ,3-nitrobenzyl alcohol, Aldrich). FAB mass
spectra were measured at room temperature with the deposit
of a 1 mL aliquot of the mixed solution, which stood over-
night to homogenize. The three solutions were as follows:
,1) 10 mL of a 1.33 M MeOH solution of a 1/1 mixture of
unlabeled ,R)- and labeled S-amino acid ester salts
,[AR1Cl2]ˆ0.67 M and [AS±dn1Cl2]ˆ0.67 M;,2) 5 mL of
a 0.20 M CHCl3 solution of a given permethylated oligo-
saccharide;and ,3) 15 mL of NBA matrix. The accuracy of
the 1/1 equivalent concentration of ,R)- and ,S)-enantiomers
of the amino acid ester salts was con®rmed from the relative
intensity ,IR/IS±dx) values with 18-crown-6 ,Tokyo Kasei),
which is an achiral host, and was experimentally obtained as
unity ,1.00^0.03).
The induced shift of 0.6 mL of b-MeCel2 in D2O ,Aldrich)
was measured adding 10 mg of ,S)-tryptophan 2-propyl
ester hydrochlorides. DSS was used as an internal standard.
4.12. Fluorescence spectrometry
FL spectra were measured while maintaining samples at
258C with a NESLAB endocal refrigerated circulating
bath. The exciting wavelength was 287 nm in the cases of
both ,S)-Trp-O-iPr1,Cl2) and ,S)-Trp-,S)-Trp-O-iPr1,Cl2).
The emission was scanned in the range 300±500 nm.
Disposable cells made from PMMA were used. The concen-
trations of the prepared solutions of ,S)-Trp-O-iPr1,Cl2)
and ,S)-Trp-,S)-Trp-O-iPr1,Cl2) were 3.53£1025 M and
2.32£1025 M, respectively. The ratio of the indole units in
these solutions was 1.0:1.1. The FL spectra of 3.5 mL
solutions were measeured before and after adding a
50-fold molar excess of permethylated oligosaccharides
,b-MeCel2, b-MeMal2, and b-MeCD).
Each relative intensity ,IR/IS±dx) value was the average of the
experimental values obtained during the 10th, 20th, 30th,
and 40th scans. The observed peak intensity of a perm-
ethylated oligosaccharide/S-amino acid ester complex ion
,IS±dx) inevitably contains contributions from the amounts
of ,M16) or ,M17) naturally abundant isotopes derived
from the peak intensity of a permethylated oligo-
saccharide/,R)-amino acid ester complex ion ,IR). However,
the theoretical distribution ,%) of the ,M16) or ,M17)
isotpes was too small ,,1%) to correct.
4.10. Molecular simulation
References
All molecular model simulations were calculated with a
software Cerius2 ,MSI, ver. 4.0) on a Silicon Graphic work-
station Unix system ,octane). The initial molecular
structures were constructed with a 3D-Sketcher software
module in Cerius2 and the molecular structures were
optimized with a Molecular Mechanics ,force ®eld:
PCFF).20 The molecular dynamics of the permethylated
oligosaccharides were simulated using a canonical ,NTV)
ensemble ,Tˆ298 K), and the temperature was controlled
by the Hoover dynamics method.21 One step of the
calculation is 1 fs. The calculation was repeated over
500,000 steps ,500 ps). The properties, which are an
averaged molecular length, a radius of gyration, and a
self-diffusion coef®cient were evaluated from the last
5,000,000 steps ,5000 ps).
Â
1. ,a) Dõhulst, A.;Berbeke, N. Chirality 1994, 6, 225±229.
,b) Yashima, E.;Okamoto, Y. Bull. Chem. Soc. Jpn. 1995,
68, 3289±3307. ,c) Yashima, E.;Yamamoto, C.;Okamoto,
Y. J. Am. Chem. Soc. 1996, 118, 4036±4048. ,d) Okamoto, Y.;
Yashima, E. Angew. Chem. Int. Ed. Engl. 1998, 37, 1020±
1043.
2. ,a) Soini, H.;Stefansson, M.;Riekkola, M.-L.;Novotny, M. V.
Anal. Chem. 1994, 66, 3477±3484. ,b) Kano, K.;Minami, K.;
Horiguchi, K.;Ishimura, T.;Kodera, M. J. Chromatogr. A
1995, 694, 307±313. ,c) Kano, K.;Negi, S.;Takaoka, R.;
Kamo, H.;Kitae, T.;Yamaguchi, M.;Okubo, H.;Hirama,
M. Chem. Lett. 1997, 715±716.
3. We have found few reports for the enantioselective formation
of 1:1 complexes of linear oligosaccharides with chiral
compounds until now. In the case of b-cyclodextrin, see:
Rekharsky, M.;Inoue, Y. J. Am. Chem. Soc. 2000, 122,
4418±4435.
4.11. NMR titration
1H-NMR spectra were measured using the 1H-NMR titration
technique reported before.9 Commercial acetone-d6
4. Sawada, M.;Shizuma, M.;Takai, Y.;Adachi, H.;Takeda, T.;
Uchiyama, T. Chem. Commun. 1998, 1453±1454.

4578
M. Shizuma et al. / Tetrahedron 57 72001) 4567±4578
5. ,a) Sawada, M.;Takai, Y.;Yamada, H.;Hirayama, S.;
13. Otsuki, J.;Kobayashi, K.;Toi, H.;Aoyama, Y. Tetrahedron
Lett. 1993, 34, 1945±1948.
Kaneda, T.;Tanaka, T.;Kamada, K.;Mizooku, T.;Takeuchi,
S.;Ueno, K.;Hirose, K.;Tobe, Y.;Naemura, K. J. Am. Chem.
Soc. 1995, 117, 7726±7736. ,b) Sawada, M. Mass Spectrom.
Rev. 1997, 16, 73±90. ,c) Sawada, M. J. Mass Spectrom. Soc.
Jpn. 1997, 45, 439±458. ,d) Shizuma, M. J. Mass. Spectrom.
Soc. Jpn. 1998, 46, 211±218.
14. Pirkle, W. H.;Pochapsky, T. C. Chem. Rev. 1989, 89, 347±
362.
15. Nishio, M.;Hirota, M. Tetrahedron 1989, 45, 7201.
16. ,a) Hakomori, S. J. Biochem. 7Tokyo) 1964, 55, 205±208.
,b) Conard, H. E. In Method in Carbohydrate Chemistry;
Whister, R. L., Bemiller, S. N., Eds.;Academic Press: New
York, 1972;6, pp 361. ,c) Melton, L. D.;McNeil, M.;Darvill,
A. G.;Albersheim, P.;Dell, A. Carbohydr. Res. 1986, 146,
279.
6. The selectivity of the permethylated inulo-oligosaccharides
toward the amino acid ester salts follows. IR/IS-dx.1.0,
R-selectivity; IR/IS-dxˆ1.0, non-selectivity; IR/IS-dx,1.0,
S-selectivity.
17. Amemura, A.;Moori, K.;Harada, T. Biochim. Biophys. Acta
1974, 334, 398±409.
7. Sawada, M.;Takai, Y.;Yamada, H.;Nishida, J.;Kaneda, T.;
Arakawa, R.;Okamoto, M.;Hirose, K.;Tanaka, T.;Naemura,
K. J. Chem. Soc., Perkin Trans. 2 1998, 701±710.
8. Lipkowitz, K. B.;Raghothama, S.;Yang, J.-a. J. Am. Chem.
Soc. 1992, 114, 1554±1562.
18. Kyba, E. P.;Timko, J. M.;Kaplan, L. J.;de Jong, F.;Gokel,
G. W.;Cram, D. J. J. Am. Chem. Soc. 1978, 100, 4555±4568.
19. Todd, J. S.;Proteau, P. J.;Gerwick, W. H. Tetrahedron Lett.
1993, 34, 7685±7688.
9. Takai, Y.;Okumura, Y.;Sawada, M.;Takahashi, S.;Siro, M.;
Kawamura, M.;Uchiyama, T. J. Org. Chem. 1994, 59, 2967±
2975.
20. Sun, H.;Ren, P.;Fried, J. P. Computational and Theoretical
Polymer Science 1998, 229±246.
21. Hoover, W. G. Computational Statistical Mechanics, Elsevier:
Amsterdam, 1990.
10. Live, D.;Chan, S. I. J. Am. Chem. Soc. 1976, 98, 3769.
11. ,a) Nishio, M. Tetrahedron 1995, 51, 8665. ,b) Nishio, M.
Bull. Chem. Soc. Jpn. 1998, 71, 1207.
22. ,a) Tone, K. In BASIC, Baifukan, 1981;pp 137±146 chap. 28.
,b) Tsukube, H.;Sohmiya, H. J. Org. Chem. 1991, 56, 875.
12. Suzuki, Y.;Morozumi, T.;Nakamura, H.;Shimomura, M.;
Hayashita, T.;Bartsh, R. A. J. Phys. Chem. B 1998, 102,
7910±7917.