Inclusion complexes of N-benzoyl-D-leucine and N-benzoyl-L-leucine with β-cyclodextrin by Raman spectroscopy

Article abstract of DOI:10.1366/0003702991946893

We report what is believed to be the first detection of chiral recognition in the inclusion complexes of cyclodextrins by Raman spectroscopy. The spectra of inclusion complexes of N-benzoyl-D-leucine or N-benzoyl-L-leucine with β-cyclodextrin were recorded in the 1500-1800 and 3000-3150 cm^-1 regions. These chiral molecules were chosen to provide group frequencies (NH₂₍₊₎ and benzoyl C=O) near the chiral center of the guest as well as COO^- and phenyl CH for probing the guest molecule without spectral interference from the vibrational bands of the cyclodextrin host. Frequency shifts and intensity decreases were observed for some Raman bands of N-benzoyl-leucines upon inclusion in the cyclodextrin cavity. In the Raman spectra of the inclusion complex, the frequency shifts were greater for N-benzoyl-D-leucine than for N-benzoyl-L-leucine.

Full text of DOI:10.1366/0003702991946893

D
Inclusion Complexes of N-Benzoyl- -leucine and
L
b
N-Benzoyl- -leucine with -Cyclodextrin
by Raman Spectroscopy
ROBIN SPIVEY and ROBERT L. SWOFFORD*
Department of Chemistry, Wake Forest University, Winston-Salem, North Carolina 27109
We report what is believed to be the ®rst detection of chiral rec-
bands. By selecting guest molecules with relevant vibra-
ognition in the inclusion complexes of cyclodextrins by Raman spec-
troscopy. The spectra of inclusion complexes of N-benzoyl-D-leucine
or N-benzoyl-L-leucine with b-cyclodextrin were recorded in the
tional bands in these spectral windows, one can study the
vibrations of the guest molecule without interference
from the cyclodextrin host.
In the present work, the Raman spectra of inclusion
complexes of N-benzoyl-D-leucine or N-benzoyl-L-leu-
1
1500±1800 and 3000±3150 cm2 regions. These chiral molecules
1
were chosen to provide group frequencies (NH2 and benzoyl C5O)
near the chiral center of the guest as well as COO 2 and phenyl CH
for probing the guest molecule without spectral interference from
the vibrational bands of the cyclodextrin host. Frequency shifts and
intensity decreases were observed for some Raman bands of N-
benzoyl-leucines upon inclusion in the cyclodextrin cavity. In the
Raman spectra of the inclusion complex, the frequency shifts were
greater for N-benzoyl-D-leucine than for N-benzoyl-L-leucine.
cine with -cyclodextrin were recorded. These guest mol-
b
ecules contain aromatic rings for favored interaction with
the cyclodextrin cavity and have vibrational absorption
in spectral regions with minimal interference from the
cyclodextrin host, namely, 1500±1800 and 3000±3150
cm21. Group frequencies for C O stretching, NH de-
1
5
2
formation, COO2 asymmetric stretching, and phenyl CC
stretching are observed in the lower energy region, and
phenyl C±H stretching is observed in the higher energy
region. Because the guest molecules are an enantiomeric
pair, they are expected to interact differently with the chi-
ral cyclodextrin host. To our knowledge, the present work
is the ®rst to study the effect of these differing interac-
tions on the Raman spectra of the guests.
Index Headings: Cyclodextrins; Inclusion complexes; Chiral; Raman
spectroscopy.
INTRODUCTION
Cyclodextrins are chiral, cyclic molecules consisting,
typically, of 6, 7, or 8 D-( )-glucopyranose units referred
1
, or -cyclodextrin, respectively. These slightly
to as
,
a b
g
water soluble molecules have the shape of a truncated
cone, with the 2- and 3-hydroxyl groups forming the wid-
er opening of the cone and the 6-hydroxyl units forming
the smaller opening. The interior of this cone is relatively
hydrophobic, while the exterior is hydrophilic. As a result
of these structural features, cyclodextrins can form inclu-
sion complexes with molecules having the appropriate
chemical af®nity, shape, and size to ®t inside the cyclo-
dextrin cavity.
Because cyclodextrins are chiral they will interact dif-
ferently with the enantiomers of the same compound.
This behavior is well known; cyclodextrins have been
used as the mobile and stationary phase in reversed-phase
separations,1,2 and derivatized cyclodextrins have been
used for chiral separations by gas chromatography.3 Cy-
clodextrins have also been used as chiral shift reagents
in NMR.4
EXPERIMENTAL
Reagents. -Cyclodextrin was kindly provided by
b
American Maize-Products Company. All other reagents
were obtained from Aldrich and used as received.
Synthesis of N-benzoyl Derivatives of D- and L-Leu-
cines. The synthesis of the N-benzoyl derivatives of D-
and L-leucine was carried out according to the Schotten±
Baumann procedure.8 Approximately 0.015 moles of the
amino acid was dissolved in an aqueous solution con-
taining 1.2 equivalents of sodium hydroxide, and approx-
imately 0.017 moles of benzoyl chloride was then added
slowly to the solution. The mixture was stirred for 15
min. The solution was acidi®ed and extracted with ether
and then ethyl acetate. The ethyl acetate was removed
under reduced pressure, and the product was dried under
high vacuum.
When a molecule is included in the cavity of a cyclo-
dextrin, it is held in place by hydrogen bonding and van
der Waal’s forces. Raman spectroscopy is an excellent
means of detecting these types of interactions, and inclu-
sion complex formation can be detected by monitoring
shifts in certain peaks in the Raman spectrum of the
guest.5,6 Recently, Raman spectroscopy has been used to
investigate these types of interactions on a chiral station-
ary phase.7 There are several regions in the Raman spec-
tra of cyclodextrins that are free from major vibrational
Synthesis of Inclusion Complexes. The inclusion
4
complexes were prepared by mixing 2 102 moles each
3
of -cyclodextrin and amino acid in a one-dram vial with
b
just enough deionized water to wet the mixture. A one-
inch magnetic stirring bar added to the vial stood nearly
on end. The vial was capped, and the mixture was
``kneaded’’ overnight by the rotating motion of the bot-
tom of the stirring bar. The product was freeze dried to
remove excess water.
Spectroscopy. The Raman spectrometer was con-
structed with the use of components from a Beckman
Model 700 double monochromator system ®tted with a
cooled Hamamatsu R943-02 photomultiplier tube. The
Received 6 August 1998; accepted 14 December 1998.
* Author to whom correspondence should be sent.
0003-7028 / 99 / 5304-0435$2.00 / 0
q 1999 Society for Applied Spectroscopy
Volume 53, Number 4, 1999
APPLIED SPECTROSCOPY
435

FIG. 1. Raman spectrum of -cyclodextrin (A) and N-benzoyl-L-leu-
b
FIG. 2. Raman spectrum of -cyclodextrin (A) and N-benzoyl-L-leu-
b
1
cine (B) in the region 1500±1800 cm2
.
1
cine (B) in the region 2800±3150 cm2
.
due to a NH2 deformation mode.10 Finally, a benzoyl
1
double monochromator has a focal length of 400 mm and
a relative aperture of f /5.9. The coupled diffraction grat-
1
carbonyl stretch can be seen at 1644 cm2
.
Ê
1
In the 2800±3150 cm2 region of the spectra (Fig. 4)
the mechanical mixture contains a cluster of peaks in the
region 2850 to 3025 cm21. These peaks are assigned as
the aliphatic CH2 and CH3 vibrations from both the N-
benzoyl-D-leucine and the cyclodextrin. The major peaks
ings are blazed at 5000 A with 1180 grooves/mm. The
1
monochromator was calibrated by using the 17 268 cm2
line of a mercury lamp. The spectral slit width was 8
1
cm2 with an intermediate slit width of 12 cm21. The
solid samples were held in glass capillaries. Samples to
be closely compared were run sequentially and on the
same calibration to maximize precision. Under these con-
ditions, the reproducibility of the wavenumbers was
found to be within 1 cm21, in agreement with the man-
ufacturer’s speci®cations. Excitation was achieved by us-
ing a Coherent Innova 90 argon-ion laser at 514.5 nm
delivering approximately 300 mW to the sample com-
partment. The output from the photomultiplier tube
(PMT) was processed with the use of a Princeton Applied
Research Model 1121A discriminator control unit and
Model 1109 photon counter interfaced to a microcom-
puter. The spectrometer was controlled and data were col-
lected with a program written in QuickBasic. Peak fre-
quencies were assigned by inspection, and no data
smoothing was performed.
1
include one at 2912 cm2 and a group of at least four
peaks centered around 2950 cm21. The overlap of peaks
from the guest and host prevents de®nitive assignment of
these bands. A phenyl C±H stretching band occurs at
1
3076 cm2 with a shoulder at 3062 cm21, likely due to
another phenyl C±H stretching mode.
Raman Spectra of Inclusion Complexes N-Benzoyl-
leucines with b-Cyclodextrin. Signi®cant changes in the
Raman spectra are observed when guest±host complexes
are formed. More signi®cant, however, are the differences
between the enantiomeric guest±host complexes. The
peak maxima are summarized in Table I. The spectra are
presented in Figs. 3 and 4.
In the Raman spectrum of the -cyclodextrin complex
of N-benzoyl-D-leucine, the COO2 asymmetric stretch
b
RESULTS AND DISCUSSION
The Raman spectrum of pure N-benzoyl-L-leucine and
that of pure -cyclodextrin are presented in Figs. 1 and
b
2, respectively.
Mechanical Mixtures of the Guest and Host. As ex-
pected with all -amino acids, the Raman spectra of the
a
enantiomers of N-benzoyl-leucine were found to be iden-
tical.9 If no inclusion complexes were formed in a simple
``mechanical mixture’’ of the cyclodextrin host with ei-
ther enantiomeric guest, then the Raman spectra would
be identical for the two mixtures. This was found to be
the case for the enantiomers of N-benzoyl-leucine.
1
In the 1500±1800 cm2 region, the Raman spectrum
of the mechanical mixture of either N-benzoyl-D-leucine
or N-benzoyl-L-leucine with -cyclodextrin contains four
b
vibrational bands (Fig. 3). The lowest energy band occurs
1
at 1580 cm2 and is assigned as the COO 2 asymmetric
FIG. 3. Raman spectra of the inclusion complexes of N-benzoyl-D-
stretching motion.10 The most prominent peak occurs at
leucine with -cyclodextrin (A), N-benzoyl-L-leucine with -cyclodex-
b
b
1
1606 cm2 and is assigned as the phenyl CC quadrant
trin (B), and a mechanical mixture of N-benzoyl-D-leucine with -cy-
b
1
stretching mode.11 The shoulder at 1616 cm2 is likely
clodextrin (C) for the 1500±1800 cm2 region.
1
436
Volume 53, Number 4, 1999

TABLE II. Comparison of wavenumber shifts for inclusion com-
plexes of N-benzoyl-leucines.
Phenyl
CC
Benzoyl
C
Phenyl
CH
1
COO2
NH2
(cm2
O
5
1
1
1
1
1
(cm2
)
(cm2
)
)
(cm2
)
(cm2
)
D-Complex
L-Complex
Difference
2
0
2
0
0
0
28
22
6
28
24
4
´´´
´´´
´´´
4
0
4
2
2
1
inclusion on the NH2 and benzoyl carbonyl peaks as
compared with the COO2 peak. The disappearance of the
1
NH2 peaks, the considerable loss of intensity of the ben-
zoyl carbonyl peak, and the appearance of new peaks
suggest signi®cant interaction with the cyclodextrin host.
The effect on the COO2 peak was less. This result could
be explained by assuming that the guest molecule was
inserted in such a way that the benzoyl carbonyl and the
NH2 portions of the molecule were inside the cavity but
the COO2 portion of the molecule remained outside or
near the rim of the cavity.
FIG. 4. Raman spectra of -cyclodextrin inclusion complexes of N-
benzoyl-D-leucine (A) and N-benzoyl-L-leucine (B) and a mechanical
b
1
mixture of N-benzoyl-D-leucine with -cyclodextrin (C ) for the 2800±
b
1
3150 cm2 region.
Also of interest is the difference in the Raman spectra
of the enantiomers upon inclusion into the cyclodextrin
cavity (Table II). In the lower energy region of the spec-
tra, new peaks appear at different frequencies for the D
and L enantiomers. In the upper region of the spectrum
peak occurs at 1582 cm21, and its intensity is decreased
relative to the strong peak at 1606 cm21. The NH2 de-
1
1
formation, which appears as a shoulder at 1616 cm2
nearly disappears for the complex. The benzoyl carbonyl
,
1
peak at 1644 cm2 is diminished in intensity, and new
1
the peak at 2912 cm2 undergoes a larger shift for the L
peaks appear at 1668, 1698, and 1732 cm21. In the higher
enantiomer than for the D, and the peak shapes are dif-
1
1
energy region of the spectrum, the peak at 2912 cm2 is
ferent around 2950 cm2
.
Since -cyclodextrin is a chiral molecule, it was ex-
shifted to 2910 cm21, and the shape of the group of peaks
b
1
at 2950 cm2 is changed. The phenyl C±H peak is shifted
pected that the enantiomers of N-benzoyl-leucine would
interact differently with the cyclodextrin host. The fact
that these differences can be detected in the Raman spec-
tra suggests a new method for studying the complexes
formed by an enantiomeric pair with cyclodextrin.
to lower energy by 2 to 3074 cm21, and the shoulder at
1
3062 cm2 nearly disappears.
In the Raman spectrum of the -cyclodextrin complex
b
with N-benzoyl-L-leucine, the intensity of the COO2
asymmetric stretch peak is also decreased relative to the
strong peak at 1606 cm21. As with the D complex, the
CONCLUSION
1
NH2 deformation, which appears as a shoulder at 1618
cm21, nearly disappears. The benzoyl carbonyl peak at
We have investigated the changes in the Raman spectra
of the enantiomers of N-benzoyl-leucine upon inclusion
of the molecule into -cyclodextrin. These changes in-
b
dicate that the N-benzoyl-leucines did form inclusion
1
1644 cm2 is diminished in intensity, and two new peaks
also appear for the L complex at 1674 and 1732 cm21. In
1
the higher energy spectral region the peak at 2912 cm2
is shifted to 2908 cm21, and in the group of peaks at 2950
cm2 a peak at 2956 cm2 becomes most prominent. The
phenyl C±H peak is shifted to 3074 cm21, and the shoul-
complexes with -cyclodextrin. The guest molecules are
b
oriented so that the phenyl group is fully inserted into
1
1
1
the cyclodextrin cavity with the NH2 and benzoyl car-
1
der at 3062 cm2 disappears.
bonyl also inside the cavity. The small changes to the
COO2 stretch suggest that its vibrations are not in¯ u-
enced by the interaction of the molecule with the cyclo-
dextrin cavity.
The differences in the Raman spectra of the inclusion
complexes of the D and L forms of the guest molecule
suggest the formation of slightly different complexes
with the chiral cyclodextrin host and further suggest the
usefulness of Raman spectroscopy for studying these chi-
ral interactions.
On the basis of the number of changes in the Raman
spectra of these molecules, we conclude that an inclusion
complex was indeed formed with -cyclodextrin. Since
b
the hydrophobic cavity of -cyclodextrin is large enough
b
to accommodate molecules as large as naphthalene,12 it
is reasonable to expect that the aromatic portion of the
N-benzoyl-leucine will ®t inside the cavity along with
some portion of the amino acid moiety.
Of particular interest is the magnitude of the effect of
TABLE I. Summary of peak maxima for N-benzoyl-leucines inclusion complexes with b-cyclodextrin and a mechanical mixture of N-
benzoyl-D-leucine with b-cyclodextrin (cm21).
Benzoyl
C
1
COO2
Phenyl CC
NH2
O
Phenyl CH
5
D-Mechanical mix
D-Complex
L-Complex
1580
1582
1582
1606
1606
1606
1616
´´´
´´´
1644
1644
1644
´´´
1668
1674
´´´
1698
´´´
´´´
1732
1732
3062
´´´
´´´
3076
3074
3074
APPLIED SPECTROSCOPY
437

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